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THE AMERICAN JOURNAL

OF

ANATOMY

EDITORIAL BOARD

CHARLES R. BARDEEN, GEORGE 8. HUNTINGTON,
University of Wisconsin. Columbia University.
HENRY H. DONALDSON, FRANKLIN P. MALL,
The Wistar Institute. Johns Hopkins University.
THOMAS DWIGHT, J. PLAYFAIR McMURRICH,
Harvard University. University of Toronto.
SIMON H. GAGE, CHARLES 8. MINOT,
Cornell University. Harvard University.
G. CARL HUBER, GEORGE A. PIERSOL,
University of Michigan. University of Pennsylvania.

HENRY McE. KNOWER, Secretary,
University of Cincinnati.

VOLUME xX

PUBLISHED QUARTERLY BY

THE WISTAR INSTITUTE OF ANATOMY AND BIOLOGY
36th STREET AND WOODLAND AVENUE
PHILADELPHIA,-PA.

' BALTIMORE

PANY

PRINTED BY THE WILLIAMS & WILKINS CO

AT THE WAVERLY PR

CONTENTS. OF VOL. X
Emit Gortscn. The structure of the mammalian ocesoph-

Seventeen Figures.

J. B. Jounston. The limit between ectoderm and ento-

derm in the mouth, and the origin of taste buds.

Weg iniges ad yeh creer a2 Woao tease ik otep ee kee Oa ates ing erate
Twenty-one Text Figures.

S. W. Wituiston. The skull of Labidosaurus..........
Three Plates.

WatTeR E. Danny. A human embryo with seven pairs of
somites, measuring about 2 mm. in length........
Six Plates.

Victor E. Emmet. A study of the differentiation of
tissues in the regenerating crustacean limb.....
Hight Plates.

HELEN DEAN KinG. Some anomalies in the genital organs
of Bufo lentiginosus and their probable signifi-

Twenty-six Figures.

Gro. -S. HuntTINGToN anp C. F. W. McCuiure. The
anatomy and development of the jugular lymph
sacs in the domestic cat (Felis domestica).......

Sixty-six Figures.

JAcoB Parsons ScHAEFFER. The sinus maxillarus and
its relations in the embryo, child, and adult man. .
Thirty-one Figures.

Cuarues R. Strockarp. The influence of alcohol and
other anzsthetics on embryonic development... .
Twenty Figures.

(ili)

4]

69

109

159

177

313

369

iv Contents

CHARLES R. Srockarp. The independent origin and
development of the crystalline lens...............
Two Plates and Twenty-eight Text Figures.

J. F. McCienpon. The development of isolated blasto-
meres of thefroe’s-epei ee 0c ae cee et eee
Two Figures.

EK. C. MacDoweE.tu. Notes on the myology of Anthropopi-

Five Figures.

Daviy M. Davis. Studies on the chief veins in early
pig embryos and the origin of the vena cava in-
FETLON ss cick 2 baa ee ee eee ee

Three Text Figures and Six Plates.

Kpwin G. Kirk. On the histogenesis of gastric glands. .
Twenty-six Figures.

F. P. Jonnson. The development of the mucous mem-
brane of the cesophagus, stomach and small intes-
tine: in the human -emibryor% +... see. ae ee

Seven Plates.

393

431

461

473

521

THE STRUCTURE OF THE MAMMALIAN CSOPHAGUS.

I BY

EMIL GOETSCH.
From the Hull Laboratory of Anatomy, University of Chicago.

WiTH 17 FIGURES.

One of the most interesting features in the structure of the Mam-
malian Cfsophagus is the extreme variability in the degree of devel-
opment of the csophageal glands in different species. For example,
according to Ranvier (84) and others, the esophagus of the rabbit,
guinea-pig and rat is wholly devoid of glands, while in the dog, a
thick layer of mucous glands nearly filling the submucous coat is
found throughout the whole extent of the organ. Nor is this dis-
parity confined to species belonging to different orders, for the
cat and dog among carnivora and the sheep and pig among ungulates
present equally striking differences in this respect.

The reasons which have been advanced for this disparity are based
on the assumption that the function of the csophageal glands is to
furnish a secretion which will serve to lubricate the surface of the
cesophageal mucous membrane and so facilitate the passage of the
bolus of food in deglutition. Assuming that the secretion of the
cesophageal glands possesses this purely mechanical function, the
logical conclusion is that their development will be influenced by
several factors which determine the consistence of the food-bolus
which is to be swallowed, such for example as the character and bulk
of the food itself, the efficiency of the masticatory mechanism, and
the relative development of the salivary glands furnishing a secre-
tion by means of which the food is diluted and rendered of softer
consistence. Accordingly, various attempts have been made to
establish a correlation between the degree of development of the
salivary glands and the efficiency of the masticatory mechanism on

THH AMDERICAN JOURNAL Or ANATOMY.—VOL. 10, No. 1, JAN., 1910.

bo

Emil Goetsch.

the one hand, and the number and size of the csophageal glands
on the other hand. For example, Ranvier (84), speaking of the dif-
ference between the rodents and the dog in this respect, remarks
that this is easily comprehensible when one remembers that in the
rabbit and other rodents the bolus of food is liquid or semi-liquid,
and therefore mucous glands are not necessary, that on the other
hand the dog swallows greedily solid matters and untriturated bones
and hence the glands are indispensable. Renaut (97) similarly calls
attention to the difference in masticatory efficiency between the ox
and rodents on the one hand and the dog on the other and explains —
on this basis the differences in the number of cesophageal glands.

Many examples can be found in favor of this explanation. When,
however, one attempts to give it a general application, unexpected
difficulties arise. For example, there is not a sufficient difference
in the development of the salivary glands, of the masticatory mech-
anism, or in the consistence of the food, to explain adequately the
fact that glands are very numerous in the cesophagus of the dog and
wholly absent from that of the cat.

The possibility that the mechanical function of the cesophageal
glands may be a purely subsidiary one, and that their true function
may be something quite different from this, has received but little
attention at the hands of the investigators. Rubeli (90), it is true,
suggested that the secretion might be of use in digestion, but his
suggestion was based not on experimental data derived from mam-
mals but on the observations of Decker (87), Swiecicki (76), Lang-
ley (79), ete, on the formation of proteolytic enzymes in the
cesophagus of fishes and batrachians. As will appear later, these
observations deal with structures which are not homologous with
the cesophageal glands of mammals and can therefore not be used to
draw conclusions concerning the function of the mammalian cesoph-
ageal glands. In this connection a question of much importance which
has been variously answered by different observers is whether the
esophageal glands of mammals are pure mucous glands, or mixed
glands containing serous demilunes. If serous cells are present,
then one must at once think of a chemical function of the esophageal
secretion as well as a mechanical one. Klein (79) asserts that

The Structure of the Mammalian Csophagus. 3

demilunes occur in the cesophageal glands of the dog. Renaut (97)
describing the cesophageal glands of the dog and of man makes the
statement that they may be seen in preparations stained with his
elycerine-hematoxylin-eosin mixture, without specifying whether
he found them in one or both species. Schaffer (97) denies their
presence in the human. esophageal glands and Rubeli (90) failed to
find them in a number of domestic mammals. Stohr (87) appears
to have seen demilunes in his preparations, but to have interpreted
them in accord with his well-known phase-theory as inactive mucous
cells. More recently Helm (67) has described typical demilunes
in the dog and pig and has demonstrated in them, by means of the
iron-hematoxylin method, the intercellular secretion canaliculi. —

Assuming that the function of the secretion of these glands is in
part at least that of mechanically aiding deglutition, one would expect
that in those cases where the character of the food and the nature
of mastication suggest the need of these structures, but where, not-
withstanding, no cesophageal glands are present, a compensatory
development of other structures will be found, as, for example, a
thickening of the stratified epithelium or an increased development
of the muscularis mucose or of the external muscular coat. In
other words, it is to be expected that a correlation of some sort will
be found between the relative development of these structures and
that of the cesophageal glands.

Accordingly, it seemed desirable to extend the investigation of
the structure of the esophagus to a much larger series of animals
than has been considered hitherto by any single investigator, and to
determine, as accurately as may be, the specializations which have
arisen as a result of the response to the differences in the nature of
the food on which these animals subsist. Among these, it might
reasonably be expected that animals which live on coarse vegetable
food would develop either a thickened epithelium, or a more com-
pletely cornified epithelium, or a layer of glands furnishing a lubricant
secretion. It is, however, not by any means easy to estimate the
degree of cornification of an epithelium except in those cases where
a true stratum corneum composed of cells which have lost their
nuclei is present. The thickness of the epithelium too is variable

4. Emil Goetsch.

from animal to animal of the same species, and in a single animal
varies with the degree of extension of the membrane upon which it
rests. Only the more pronounced differences in these respects may
therefore be interpreted with caution from the standpoint of special-
ization.

It was apparent from the outset that very little help in interpret-
ing the cesophageal glands of mammals could be obtained from the
consideration of these structures in lower vertebrates, because glands
occur in reptiles only in the form of imperfect crypts in certain tur-
tles, and the so-called cesophageal glands of certain batrachia are, .
according to Bensley (00), in reality gastric glands.

Meruops or Strupy.

One difficulty that at once presents itself in studies on the cesoph-
agus is that of determining the point of transition of the pharynx
into the cesophagus. In animals like the dog, where there is a trans-
verse fold of the mucous membrane, corresponding in its posi-
tion to the lower border of the cricoid cartilage, and to an actual
change in structure of the mucous membrane, this is relatively easy,
but in the majority of cases no such superficial line of demarcation
exists, and the point of transition must be more or less arbitrarily
established. In the descriptions which follow, the lower border of
the cricoid cartilage has been taken as the point where the pharynx
passes into the cesophagus. In his recent article on the cesophageal
glands Haane (05) places the point of transition somewhat higher,
at the level of the corniculate cartilage, but designates the portion
of the tube included between this point and the lower level of the
ericoid cartilage in the dog as “Csophagus-vorraum.” In order to
include the doubtful region so designated by Haane, sections from
this region have been studied in each mammal examined, but struc-
tures occurring above the distal margin of the cricoid cartilage have
been referred to the pharynx, those below to the cesophagus.

In the case of the smaller mammals the entire cesophagus was
fixed, in the larger mammals, where this was out of the question,
a strip was taken including the whole length of the esophagus.

For fixation, Zenker’s fluid was employed, Bensley’s alcohol-

The Structure of the Mammalian (sophagus. 5

bichromate-sublimate mixture being used where a more detailed
study of the glandular epithelium was desired. The entire cesoph-
agus or a strip, after fixation, was cut into lengths of 1 cm. to
2 cm., and imbedded in paraffin. From each of these segments,
which were numbered consecutively from above downwards, sec-
tions were made at intervals of one millimeter, so that all portions
of the esophagus were examined. This method, however, did not
exclude the possibility in the case of those animals where the results
as regards the presence of glands were negative, that some glands
were missed in the short unsectioned portions. Accordingly the fol-
lowing method devised by Bensley was employed, where the material
was available, to make preparations in toto of the layer containing
the glands, staining the latter selectively so that every gland lobule
in the wsophagus was demonstrated clearly. The cesophagus was
pinned out on cork and placed in 70 per cent. aleohol for 24 hours.
Then, after a further stay of 24 hours in 95 per cent. alcohol, the
mucous membrane was dissected off, by dividing the tela submucosa
earefully with a scalpel close to the muscular tunic. In this way all
the glands come off with the layer of submucosa which remains
attached to the mucous membrane. The mucous membrane is placed
in water for one hour, then transferred to a mixture of one part of
strong muchzematein (see Bensley, 03) and five parts of distilled
water. In this staining solution the membrane remains for 48
hours, after which it is washed in distilled water and transferred to
95 per cent. alcohol containing two volumes per cent. of strong
hydrochloric acid. In this solution the preparation remains until
the glands stand out distinctly blue on a red background, when the
preparation is washed in several changes of alcohol, dehydrated in
absolute alcohol and cleared in benzole. Where the epithelium
is thick, as in man, dog, ete., it stains so intensely that it interferes
seriously with the transmission of light through the preparation. It
is easy, however, to remove the epithelium by stripping off with
forceps after clearing in benzole. By this means a preparation is
obtained in which every gland of the cesophagus is clearly visible
and their general relations to one another, the nature, course, and
branching, of their ducts may be seen. Furthermore, in such prep-

6 Emil Goetsch.

arations the branching of the tubule may be studied with ease, thus
avoiding the laborious method of reconstruction from sections.
Because of the lack of sufficient material I was unable to apply the
method in the case of the wild animals whose cesophagi were exam-
ined, but such preparations were made of all the domestic animals
and of man.

For staining sections hematoxylin and eosin, iron hematoxylin,
copper chrome hematoxylin, neutral gentian, muchematein, mucicar-
mine, Mallory’s connective tissue stain and acid violet-saffranin were
employed.

Opossum (Didelphys virginiana).

The mucous membrane of the cesophagus of the opossum exhibits
the usual transitory longitudinal folds observed in the empty
cesophagus. About 1 em. above the cardiac orifice of the stomach,
however, these disappear, and their place is taken by permanent
- transverse folds of the mucous membrane approximately 0.5 mm. in
width, and provided on their free surfaces with a network of second-
ary ridges. These folds are separated from one another by deep
sulci and, as will appear later, owe their occurrence in part to the
accumulation in the lamina propria mucose of masses of glands.

The epithelium of the cesophagus at its upper end is represented
in Fig. 1. It consists of a layer of somewhat irregular thickness,
owing to the projection into it, from below, of ridges longitudinal in
direction, belonging to the lamina propria. In full grown animal
weighing 3,000 grammes, the thickness of the epithelium at the
level of the cricoid cartilage was, in the spaces between the con-
nective-tissue ridges, 190-250 micra, on the summit of the ridges
72-110 micra. The irregularity in thickness presented by this
epithelium in transverse sections is due to high ridges of the lamina
propria, which are for the most part longitudinal in direction, but
are connected with one another by lower transverse and oblique ridges,
so that in sections parallel to the surface at the level of the ridges
a connective tissue network is seen surrounding islets of epithelium,
instead of the epithelial network seen at this level in sections of the
epidermis. As described by Oppel (97) in Phalangista, Phasco-
larctus and Aepyprymnus, true papille are wanting in the opossum.

The Structure of the Mammalian Csophagus. 7

On the surface of the epithelium a well-defined stratum corneum
is seen, of fairly uniform thickness, although it dips down some-
what in the intervals between the longitudinal ridges of the lamina
propria. This corneous layer presents two distinct strata, which
correspond in a general way in their appearance and staining reac-
tions to the stratum lucidum and stratum corneum of the epidermis.
The deeper layer, 85 micra in thickness, stains deeply in eosin, par-
ticularly at the deep and superficial margins, the intermediate por-

Stratum corneum

Stratum lucidum

: Stratum
e germinativum

— Lamina propria
Zt -mucose

Fic. 1. Transverse section of epithelium of cesophagus of Didelphys at
level of cricoid cartilage. 120.

tion exhibiting, as is often the case with the stratum lucidum, patchy
or irregular staining. ‘This layer is composed of flattened cells,
elongated, spindle-shaped in section, with a flattened nucleus rod-
shaped in section. ‘The superficial layer, 17 micra in thickness at
its thickest portion, stains but faintly in eosin. It consists of cells
of irregular polygonal shape similar to those of the stratum corneum
of the epidermis with the exception that remains of the nucleus are
to be found in them, in the form of shrunken remains of the nuclear
membrane and one or two chromatic particles.

8 Emil Goetsch.

The stratum germinativum requires no special comment except
that its superficial layers contain no granules of eleidin.

There is a gradual reduction in thickness of the epithelium going
down the esophagus. At the middle it is from 58-170 micra in
thickness with a corneous layer 35 micra in thickness. The super-
ficial layer of the corneous stratum here consists only of scattered
cells of the sort described above.

At the point where the permanent transverse folds make their
appearance about 1 em. above the cardia there is a change in the
character of the epithelium. On the surface and sides of these folds

aero / = — Lamina
SG Ee propria

SSS

Tic. 2. Epithelium from side of transverse folds at lower end of cesophagus
of Didelphys. > 750.

the epithelium varies from a double layer of cubical to polygonal
cells, 10 micra in thickness (Fig. 2), to several layers of cells, the
superficial layers flattened, 50 micra in thickness. The thin double
layer of cells is found here and there on the sides of the transverse
folds, the thicker epithelium on the summits. As shown in Fig. 2
there is frequently no sign at all of cornification of the superficial
layer.

The lamina muscularis mucosee is longitudinal in direction and is
found throughout the whole wsophagus. At the lower level of the
ericoid cartilage it makes its appearance as scattered bundles of

The Structure of the Mammalian Csophagus. 9

unstriated muscle. At the middle of the esophagus it forms a con-
tinuous layer of considerable thickness (205 micra in an animal of
3 kg.) and increasing to 420 micra at the cardiac orifice of the stom-
ach. In the lower portion, by reason of the fact that the glands are
practically confined to the mucosa, the lamina muscularis mucose
is separated by only a narrow band of collagenic connective tissue
representing the tela submucosa, from the tunica muscularis.
Mucous glands are present throughout the whole length of the
esophagus and in the pharynx. In the upper part of the esophagus

Demilune cells

Lumen

Mucous cells

Fic. 3. A. Section of a tubule of an csophageal gland of Didelphys show-
ing mucous cells and demilunes. B. Group of demilune cells with central
lumen and intercellular canaliculi. > 500.

they are located in the submucosa, but at the lower end where the
transverse folds occur they are found in the lamina propria of these
folds, superficial to the 1. muscularis mucose, although here a few
tubules may extend into the lamina muscularis mucosze and even
into the tunica muscularis.

The glands in all parts of the cesophagus of the opossum are mixed
glands, that is to say they consist of mucous cells and demilunes or
erescents. The latter are few in number at the upper end of the
cesophagus, but at the lower end where the glands are located in the
lamina propria mucose they are very abundant, as shown in Fig. 3.

10 Emil Goetsch.

The character of the mucous cells of these glands is well shown
in Fig. 8 and requires no special description. ‘Their secretory con-
tent stains selectively in muchematein.

The crescents (Figs. 8 and 4) are composed of cells which stain
intensely in eosin. In the glands of the upper portion of the cesoph-
agus they are to be found forming the characteristic crescent-
shaped groups at the ends or along the sides of tubules. In the
lower glands they form aggregates of considerable size, often sur-
rounding a lateral diverticulum of the lumen of the gland so as to
make a sort of sessile acinus on the side of a tubule. The character

Duct epithelium

Demilune

Denvilune

Iie. 4. Portion of duct of cesophageal gland from lower end of cesophagus
of Didelphys showing demilune cells alternating with duct epithelium. < 500.

of the cells is the same at all points in the esophagus. They are
cuboidal in shape and the aggregate presents a rather remarkable
resemblance to the parietal cells of the gastric glands. The cytoplasm
is finely granular, but the granules are less crowded than in the
parietal cells. Between the constituent cells of the complex, in iron
hematoxylin preparations, may be seen fine intercellular secretion
canaliculi, their outlines defined by fine cement lines. In Mal-
lory preparations the granules along the canaliculi and on the lumen-
border of the cell stain differently from the fine granules of the
cytoplasm and probably represent the secretion antecedent of these

The Structure of the Mammalian Césophagus. 11

cells. ‘There are no intracellular ductules. The crescents in the
glands of the lower portion of the esophagus are not confined to the
mucous portion of the gland, but occur also in groups along the ducts,
alternating with the non-secreting cubical epithelium of the ducts,
as shown in Fig. 4.

The ducts of the glands are lined at their point of origin from
the gland-tubule by a double layer of cuboidal cells. As they
approach the surface the number of layers increases, there being a
gradual transition to a stratified squamous epithelium. The majority
of the ducts pass into the gland about the level of the deep border
of the muscularis mucose. Frequently, however, they branch before
penetrating the muscularis mucosee, and occasionally they receive
a small group of mucous tubules in the lamina propria or in the
muscularis mucose. There are no aggregations of lymphoid tissue
around the ducts nor are the latter enlarged to form ampulle, as in
the pig.

The ducts of the superficial glands at the lower end of the cesoph-
agus are more numerous and the glandular masses are less com-
plex. The ducts here open into the depressions formed by the sec-
ondary network of ridges on the surface of the folds and also at

various points into the deep grooves between the principal folds.
_ The glandular tubules are supported by a thin basement membrane
of reticulum, between which and the bases of the cell fine fibres
resembling myofibrille may be seen. The latter stain red in Mal
lory’s reticulum stain, and probably belong to stellate cells (Korb-
zellen), although this could not be demonstrated by the technique
employed.

The tunica muscularis consists in the upper third of the cesophagus
of striated muscle. At the middle of the esophagus a few striated
fibres are still to be found, but below this point it is all unstriated.

RopeENTIA.

The literature of this subject contains so many references to the
structure of the cesophagus in Rodentia that it is unnecessary to
describe in detail the conditions found in the different rodents exam-
ined. It will suffice to discuss those structures which may be con-

12 Emil Goetsch.

sidered to be specialized in some degree to meet the mechanical
conditions imposed by the food.

The cesophagi of the following rodents were examined: Arctomys
monax, Sciurus hudsonicus, Tamias striatus, Cavia, Erethizon dor-
satus, Mus decumanus, Geomys bursarius, Lepus cuniculus, Lepus
nuttalli mallurus. In the ease of the porcupine (Erethizon) (Fig.
5 A) only the lower portion of the esophagus about three centimeters

Yj — Stratum corneum

if (
Z i,

-—— Stratum granulosum

“eum Lamina propria
S. mucose

“

Fie. 5. A
Transverse section of tunica mucosa of cesophagus of Erethizon.

in length was available. In addition to the above genera the follow-
ing rodents have been described by other authors: Mus musculus,
Arvicola amphibius and Arvicola arvalis, Hypudeeus arvalis, Sperm-
ophilus citillus.

With regard to the occurrence of glands, the results of all observa-
tions hitherto reported are negative except in the case of the rabbit,
where glands are reported by Graff (80) and by Vogt and Yung
(94), denied by Klein (71).

The Structure of the Mammalian Csophagus. 13

My observations are in accord with those of Ranvier (84), Briim-
mer (76) and others who deny the presence of cesophageal glands in
rodents. In none of the species mentioned above are cesophageal
glands to be found below the level of the cricoid cartilage. In the
rabbit, pocket gopher (Geomys) and chipmunk (Tamias) groups of
mucous glands were found above this layer in the submucosa of

Stratum
corneum

SS —— Stratum
granulosum

—— Stratum
germinativum

Lamina
propria mucose

——/. muscularis
MuUucOse

Tunica mucosa of cesophagus of guinea pig. Showing true stratum cor-
neum and stratum granulosum. > 120.

the pharynx, and it is probable that the occurrence of glands in this
portion of the pharynx is responsible for the statements of Graff
and of Vogt and of Yung that glands occur in the esophagus of the
rabbit.

The cesophageal epithelium in the rodents is a thick layer of strati-
fied squamous cells presenting in the different members of the order
two main types. One type (Fig. 5 A) is characterized by a very rapid

14 Emil Goetsch.

and complete cornification of the superficial layers of the epithelium
to constitute a true stratum corneum composed of flattened cells
wholly devoid of nuclei, and comprising over one-third of the total
thickness of the epithelium. Below the stratum corneum in this
type a true stratum granulosum is found consisting of two or more
layers of cells, of elongated fusiform shape in section, containing
numerous granules of eleidin staining blue in hemalum. The
stratum germinativum is relatively thin as compared with that of
the second type. This type of epithelium is found in the guinea pig
(Fig. 5 B), rat, pocket gopher and porcupine. ‘The second type of
epithelium (Fig. 6) also shows a considerable degree of cornification,
but a true stratum corneum is not present, the superficial layers of
flattened cells being nucleated and no stratum granulosum being
found. In this epithelium the process of cornification appears to go on
more gradually than in the first type and no sharp line can be drawn
between the stratum corneum and the stratum germinativum. ‘The
superficial half of the epithelium nevertheless consists of flattened
cells with elongated nuclei and stains more strongly in eosin than
the deeper layer. Comparing this epithelium with that of the dog
it is apparent that the degree of cornification is much greater in
the rodent and that the thickness of the epithelium considering the
relative sizes of the animals is much greater. True papille of the
lamina propria mucose are not present in the rodents examined. In
Cavia, Arctomys, Sciurus, Tamias and Lepus, however, as described
by Strahl (89), the lamina propria mucosz projects into the epithe-
lium in the form of irregular ridges for the most part longitudinal
in direction, and in Cavia and Tamias these have irregular summits
which in places approximate the formation of papille. In Mus,
Geomys, and Erethizon, there are neither ridges nor papille.

The 1. muscularis mucose is present throughout the whole length
of the cesophagus in all the rodents examined. It is particularly
well developed in the squirrels, where it forms a continuous layer
around the cesophagus.

The tunica muscularis in the rodents consists of striated fibres
throughout the greater extent of the esophagus. In Cavia, Geomys,
Mus, Tamias, and Lepus the striated fibres are found to the cardiac

The Structure of the Mammalian Csophagus. 15

opening of the stomach. In Arctomys striated fibers are found
right up to the cardia, but for a very short distance at the lower
end of the cesophagus are mixed with smooth muscle. In Sciurus
also there is a short distance at the lower end of the cesophagus in
which smooth muscle is found. In Erethizon the outer layer of

Surface
cornification

: i Stratum
ca \ germinativum

1 _L. muscularis
MUCOSe

Fic. 6. Transverse section of tunica mucosa of Lepus nutalli mallurus.
< 180.

muscle is striated to the cardia, but the inner layer has a thick lower
sphincter composed of smooth fibres. In Muscardinus the striated
fibres extend over onto the preventricular dilatation which, as is well
known, contains fundus glands and is therefore to be regarded as a
portion of the stomach.

16 Emil Goetsch.

INSECTIVORA.

The only observations on the structure of the esophagus in the
Insectivora which I have been able to find are the descriptions of the
cesophagus of Erinaceus given by Carlier (93) and Oppel (97).
Carlier’s description may be briefly summed up in the following
statements: The epithelium is thick and of the ordinary type; the
muscularis mucosee is greatly developed consisting of large bundles
of coarse non-striped fibers, longitudinally placed; the submucous
coat is reduced to a minimum, due to the total absence of all glan-
dular structures, mucous glands being entirely absent with the excep-
tion of a few scattered acini, situated near the cardiac end, internal
to the muscularis mucose, and therefore in the mucous membrane.
There are, however, according to Carlier, some serous glands in the
submucosa of the organ, arranged in a ring round the cardiac orifice
of the stomach, the long ducts of which pierce the muscle and epithe-
lium to open just above the border of the cwsophageal epithelium.
The muscular coat consists throughout its whole extent of striped
fibres. Oppel, on the other hand, found mucous glands in the upper
portion of the cesophagus only. These glands showed cells of two
types, as regards their affinity for hematoxylin and eosin respec-
tively, although Oppel did not decide whether this difference was
due to the physiological state of the cell or to a fundamental differ-
ence.

In Scalops aquaticus the conditions are very similar to those
described by Oppel in Erinaceus. That is to say the glands are con-
fined to the pharynx and to a very small portion of the upper extrem-
ity of the esophagus.

The epithelium in Scalops is thick and fairly uniform in thick-
ness throughout the length of the cesophagus, the increase in thick-
ness towards the lower end of the esophagus being but slight. At
the upper end it measured 127 micra, at the lower end 139 micra.
A true stratum corneum is not present, although a considerable
degree of cornification of the superficial layers is apparent. The
most superficial cells are nucleated and no stratum granulosum is to
be seen. The superficial layers, about 24 micra in thickness, stain
intensely in eosin. No true papille are present, although the deep

The Structure of the Mammalian (sophagus. 17

border of the epithelium presents a somewhat irregular outline in
section.

The lamina muscularis mucose is well developed, except at the
very beginning of the cesophagus. It forms a continuous layer com-
posed of two or three layers of longitudinally disposed bundles of
unstriated muscle.

The glands of the lower pharynx are mixed glands containing
mucous portions with demilunes and serous alveoli.

The external muscular coat is composed of striated muscle through-
out the whole length of the cesophagus.

CHIROPTERA.

This order is represented in my material by a single alcoholic
specimen of Vespertilio fuseus, the brown bat.

The epithelium in the csophagus of Vespertilio is thin (30-45
micra), and shows no stratum corneum, although the cells of the
superficial two-thirds of the epithelium are much flattened. No
papille are present.

The muscularis mucose is exceptionally well developed, reaching
a thickness of 50 micra in the lower third of the cesophagus. It
forms a continuous layer and consists of smooth muscle arranged
longitudinally.

No glands are found at any level in the cesophagus.

The external muscular coat consists of striated muscle in the upper
two-thirds and unstriated in the lower third.

CARNIVORA.

In the Carnivora the esophagi of the following species, hitherto
undescribed, have been examined: Proeyon lotor, Lutreola vison,
and Mephitis mephitica. One cesophagus of each species was avail-
able for this study including in the case of Procyon and Mephitis
the whole of the cesophagus and the adjacent portions of the pharynx
and stomach, in Lutreola the cesophagus and stomach only, so that in
the latter animal the pharynx and transition region was not obtained.
In addition the csophagi of the cat and dog were examined to con-
firm the work of earler observers. In these animals preparations

Emil Goetsch.

wer
CO

of the whole cesophagus were made according to the method outlined
in the introduction in order to determine positively the distribution
of glands in them. In the case of the cat three such preparations
were made, all of which demonstrated the complete absence of
glands below the level of the cricoid cartilage.

The cesophageal glands of the dog, like those of the opossum, are
composed of two kinds of cells, mucous cells and serous demilunes.
In this respect my observations confirm the statements of Klein (79),
Renaut (97) and Helm (07) and are opposed to those of Rubeli (90)

Serous
demilune

Mucous
cells

Fic. 7. Section of two tubules from csophageal gland of the dog showing
mucous cells and demilunes. >< 750.

and Haane (05). ‘The latter observer, it is true, does not specifi-
cally state that demilunes are absent, but implies it in the statement
that he was unable to find intercellular secretion-capillaries. The
demonstration of the demilunes in the cesophageal glands of the dog
is no difficult matter if thin sections of well-fixed tissue are examined,
and if an efficient differential staining method is employed, for
they occur in relatively large numbers in all the esophageal glands,
although less numerous than in the glands of the opossum and rac-
coon (Procyon). For staining neutral satireviolet-safranin is par-

The Structure of the Mammalian Csophagus. hg)

ticularly valuable, the mucous cells staining red in this method,
the demilunes violet. This method also shows well the intercellular
secretion capillaries, although the iron hematoxylin method is pref-
erable. The demilunes show the typical arangement as seen in
the submaxillary gland and are provided, as indicated above, with

Fic. 8. Drawing of an entire gland from cesophagus of dog, showing tubulo-
acinous character and mode of branching.

intercellular secretion capillaries resembling in every respect those
found elsewhere (Fig. 7).

The shape and mode of branching of the gland is well shown in
Fig. 8 drawn from a preparation stained in toto.

Procyon Loror.

As in the dog, there is, in the raccoon, in the lower part of the
pharynx, a circular fold of mucous membrane which projects into

20 Emil Goetsch.

the Jumen and forms a superficial demarcation between the cesopha-
eus and pharynx. This fold in the raccoon, however, is without
elands.

The epithelium (Fig. 9) varies in thickness according to the degree
of contraction of the surface upon which it rests from 158 micra to
220 micra. There is no marked thickening of the epithelium as
the stomach is approached. The lower border of the epithelium is
irregular in transverse sections owing to the projection into it of
high ridges of the lamina propria mucosee. These ridges are chiefly
longitudinal in direction, but are connected with one another by
numerous oblique ridges. On the summits of these ridges low
conical papille are found.

The muscularis mucose is well developed throughout the whole
cesophagus, consisting of two or more layers of longitudinal bundles
of unstriated muscle.

Glands are present in the tela submucosa throughout the entire
length of the esophagus, and are fully as numerous as in the dog.
The glandular lobules are ovoidal in shape, somewhat compressed
from side to side and so placed that their long diameter coincides
in direction with the long axis of the cesophagus. Many of the
lobules have independent duets, but in the majority of cases a duct
divides below the lamina muscularis mucosz into two ducts which
enter adjacent lobules. The glands are of the tubulo-acinous type,
the lobule being a system of highly branched tubules with small
acini along their course and at their terminations. There is no
difference between tubules and acini as to the character of the
lining cells. The epithelium changes from duct epithelium to secret-
ing epithelium at the point of entrance of the duet, so that all tubules
within the lobule are lined exclusively by glandular epithelium.

The character of the glandular epithelium is well shown in Fig.
10. It consists of mucous cells and serous demilunes. The latter
are very abundant and conspicuous. The mucous cells correspond
in character with those of other mucous glands and require no special
description. The demilunes in the glands of the upper part of the
esophagus have the characteristic crescentic shape in sections. In
the glands of the lower portion of the esophagus many demilunes

bo
rear

The Structure of the Mammalian Csophagus.

—— Epithelium

U, muscularis
MUCOSe

Duct of gland

Gland tubules
showing mucous
cells and demi-
lunes

Fic. 9. Transverse section of tunica mucosa and part of tela submucosa of
cwsophagus of Procyon. >< 120.

22 Emil Goetsch.

are so large that they form small sessile acini along the sides of
the tubules. In both cases intercellular capillaries are seen in iron
hematoxylin preparations, outlined by cement lines running between
the adjacent surfaces of the cells. In the acinus-like demilunes of
the glands at the lower end these intercellular ductules open into a
central lumen which is directly continuous with the lumen of the
main tubule. Although the material was not suitable for studying
the secretory content of the cells, because of the fact that the cesoph-
agus was fixed in Zenker’s fluid fully an hour after the animal was

Mucous cell

ab Ness
PU NL ete
em oaths aay a
¢

Serous
acinus
(demilune)

Fic. 10. Portion of tubule of cesophageal gland of Procyon showing
mucous cells and large demilunes with secretion granules. >< 500.

killed, yet in some of the more superficial glands the cells of the
demilunes contained well-fixed secretion granules (Fig. 10), which
stained strongly in eosin and in iron hematoxylin and which occu-
pied the portion of the cells bordering on the lumen. No basal
filaments, however, were demonstrated.

The ducts of the gland are lined for a very short distance at their
origin from the lobule, by a low simple cubical epithelium. A short
distance from its origin this changes to a two-layered epithelium,
the cells of the surface layer having their long axes perpendicular
to the basement membrane, those of the deep layer parallel to it.

The Structure of the Mammalian (Esophagus. 23

In the upper part of the duct a third layer is added and the surface
layer becomes flattened. The ducts enter the epithelium between the
ridges of the lamina propria.

The duets run perpendicular to the surface and have no ampulla-
like enlargements. No lymphatic nodules nor accumulations of
lymphocytes occur in relation to the ducts.

The tunica muscularis is composed of striated muscle throughout
nearly its whole extent. In the stratum circulare smooth muscles
make their appearance a short distance above the cardia and expand
at the cardia into a lower sphincter.

Merpuitis Mepurrica.

In Mephitis the epithelium is extremely uneven in thickness owing
to the fact that from the deep surface epithelial processes descend
into the lamina propria mucose. The connective tissue separating
these processes represents, somewhat more highly developed, the
longitudinal and transverse ridges of the esophagus of the raccoon.
The thickness varies from 122 micra to 294 micra, the difference
between these measurements indicating the height of the epithelial
processes. The epithelium shows very little evidence of cornifica-
tion, although the superficial layers of cells are much flattened and
stain more readily in eosin than the deeper ones. .

The lamina muscularis mucosz begins at the upper end of the
esophagus as scattered bundles, but rapidly becomes a complete
layer 47 to 118 micra in thickness, according to the degree of
extension of the mucous membrane. :

The only glands present in the esophagus are found just at the
cardiac orifice of the stomach. They are located in the submucosa
and are similar in structure to the glands of the esophagus of Pro-
‘eyon. The demilunes are particularly abundant and have the char-
acteristic form and structure. The ducts begin in the lobules with
a lining of cubical cells beneath which is an imperfect second layer
of cells. This second layer quickly becomes complete on going up
the duct and additional layers are added as the surface is approached,
as in esophageal glands elsewhere. These glands are therefore in

24. Emil Goetsch.

every respect typical esophageal glands and can not be confused with
the neighboring cardiac glands of the stomach.

The tunica muscularis consists of striated muscle throughout its
whole extent.

LurrEoLa VIsoN.

The epithelium in Lutreola is very similar to that in the cat.
No true papille are present and there is a very slight indication
of ridges of the lamina propria so that the deep border of the epi-
thelium is fairly regular. The cornification is imperfect but is
indicated by the different staining of the superficial layers of cells
which are much flattened, but which retain their nuclei. The thick-
ness is fairly uniform throughout except for such differences as
are the result of tension.

The lamina muscularis mucosee is well developed, forming a
layer 50-80 micra in thickness at the upper end of the oesophagus
and gradually increasing in thickness to 180 micra at the cardiac
orifice.

A few glands only are present, and these are confined to the upper
fourth of the wsophagus. They are similar in nature to the glands
of the esophagus of Procyon, that is they consist of mucous cells and
demilunes.

UNGULATA.

My observations on the structure of the esophagus of the ungu-
lates have been largely confirmatory of the work of Ellenberger (84),
Rubeli (90), Haane (05), and Helm (07).

To ascertain beyond doubt the presence or absence of glands in
the oesophagus of the sheep and ox, preparations were made of the
whole cesophagus according to the method already outlined, but with
negative ‘results. These osophagi contained no glands. In the
ease of the horse sections made at different levels were likewise
devoid of glands.

In the esophageal glands of the pig, I have found, in confirmation
of Helm (07), demilunes of serous cells, but in smaller numbers
than in any of the other animals whose glands were studied with
the exception of man. They are, however, by no means infrequent,

bo
Or

The Structure of the Mammalian (Césophagus.

although the complexes are smaller in size, more compressed, and
more easily overlooked than in other animals. These demilunes are
provided with intercellular secretion-canaliculhi.

In all the ungulates examined, as might be expected, the epithe-
lum shows a marked thickening, and an unusual degree of cornifica-
tion. The condition in the sheep as shown in Fig. 14 is typical
of that found in the sheep, ox and horse. The epithelium is thick
and presents on its deep surface an irregular outline owing to the
presence of longitudinal ridges of the lamina propria mucose. In
addition, both on the summits of these ridges and between them
there are extraordinarily long papille, for the most part simple,
which penetrate the epithelium as far as the stratum corneum and
even penetrate that layer for a short distance. A thick stratum
corneum forms the outer portion of the epithelium, forming from
one-third to one-fourth of the entire thickness of the epithelium,
the thicker portions being found in the lower regions of the cesoph-
agus. In this stratum corneum two secondary strata may be made
out which may be compared in general with the strata lucidum and
corneum of the plantar skin, although there are important differences
in structure. Both layers are very homogeneous and transparent in
the fresh condition, and in unstained alcohol-fixed sections. The deeper
layer is composed of fusiform flattened cells, but with flattened nuclei.
The superficial layer shows the wrinkled cell borders seen in section
of the stratum corneum of the skin. No true stratum granulosum
is present, but the superficial portion of the stratum germinatum
shows indications of a change preparatory to cornification, in that
the cell protoplasm of this layer stains more readily in fuchsin than
the deeper layers. Nuclei are present in the cells of the most
superficial layers of the stratum corneum, although they are much
degenerated. Thus while a deep stratum corneum is present the
changes do not involve the nuclei of the cells to the extent that they
do in the guinea pig.

In the pig, also, the epithelium is thick, but the degree of cornifica-
tion is much less than in the sheep, the contrast between the stratum
germinativum and the stratum corneum less striking, and the tran-
sition less abrupt from the one layer to the other. We do not see

26 Emil Goetsch.

in the esophagus of the pig the division of the stratum corneum
into two layers as in the sheep, and in the pig nuclei are more
numerous in the superficial layers and less degenerated. ‘There is
thus in these animals a very evident relation between the presence
of glands and the degree of cornification of the epithelium,

An interesting fact in connection with the structure of the pig’s
cesophagus is the arrangement of the muscularis mucose. In the
upper portion of the csophagus where the glands are abundant
there is no muscularis mucose. It makes its appearance a short
distance above the point at which the glands begin to thin out and
is well developed in the lower half of the esophagus where there are
few glands.

Man.

Much disagreement exists between observers as to the number and
distribution of the cesophageal glands in man. According to Toldt
(89) they are present in large numbers in the upper section of the
cesophagus and are wholly wanting in the lower segment. Accord-
ing to Klein (71) they are of rare occurrence. Kossowski (80)
on the other hand finds them most abundant at the lower end of the
cesophagus near the cardia. Dobrowolski (94) found that there were
considerable individual variations, but that the whole number did not
exceed two hundred, of which two-thirds were in the upper half of
the cesophagus. The latter observer’s results were based on prepara-
tions of the entire esophagus and his conclusions as to the indi-
vidual variability are supported by the preparations which I have
made by the method already outlined. The extent of the individual
variations is well indicated by figures 15 and 16 which show the
exact location of every glandular lobule in two human esophagi.
One of these contained 741 lobules pretty well distributed over the
whole cesophagus with the exception of a short area at the beginning
of the esophagus and another near the cardiac end where the glands
are relatively few in number. The other csophagus had but 62
glandular lobules, 58 of which were found in a segment 4 em. in.
length beginning 3 em. below the cricoid cartilage. In a third
cesophagus there were 140 lobules of which 43 were located in a

The Structure of the Mammalian Csophagus. 27

segment 3 cm. long at the lower end of the esophagus. Above this
a segment 3 em. long was free from glands. Above this 37 glands
were distributed over an area 10 em. in length. At the upper end
of the wsophagus 60 glands were distributed over a distance of 7
em., this group being separated from the middle group by a segment
2 em. in length devoid of glands. In addition to the glands there
were 54 cysts of various sizes. As the figure (15) indicates, there

Fic. 11. Photomicrograph of two glands from human esophagus stained
in toto. The long axis of the cesophagus coincides with that of the glandular
lobules and the ducts point obliquely toward the stomach.

is a marked tendency for the glands to be arranged in longitudinal
rows parallel to the long axis of the esophagus. These rows may
be as many as eight in number at one transverse level and are not
confined to the anterior surface and lateral surfaces as stated by
Dobrowolski, but are distributed indifferently on all surfaces of
the cesophagus.

The relation of the ducts to the lobules is well shown in Fig, 11.

-

bo
oe)

Emil Goetseh.

Jach duct is oblique in direction with reference to the axis of the
esophagus but has a general direction downward in the direction
of the stomach. The descending portion of the duct les in the
tela submucosa close underneath the lamina muscularis mucose, and
may reach a length of 4.6 mm., although the majority of them are
much shorter. Each duet branches at its termination into from two
to five rarely more secondary ducts, each of which enters a lobule.
Ampullary dilatations of the ducts are of very common occurrence
and affect most frequently the oblique portion of the main duct in
the submucosa. Almost as frequent, however, are the dilatations
into ampulle of the secondary ducts at the pomt of emergence from
the lobule or in the hilum of the lobule. The portion of the duet
contained in the lamina propria mucose is frequently surrounded,
as pointed out by Flesch (88) and Schaffer (97), by accumulations
of leucocytes and often by true lymphatic nodules with a germinal
center. At the point where the duct emerges from the lobule there is
also, frequently, surrounding the duet, an accumulation of reticular
tissue containing large numbers of leucocytes. ‘The nature of the
leucocytes found in these accumulations varies considerably, but in
some cases the predominating cell is Unna’s plasma cell. In other
eases the lymphocytes predominate. In all cases, however, there
are large numbers of plasma cells distributed along the duct and in
the reticular tissue forming the framework of the gland, and in those
cases where no nodule is present along the duct one finds occasionally
the whole duct surrounded by a narrow layer of reticular tissue
containing plasma cells and lymphocytes.

As regards the structure of the lobule and the character of the
individual cells composing the gland, little can be added to the
excellent description given of these structures by Schaffer (97) and
the description which follows is largely confirmatory of his obser-
vations.

The lobular duct gives off within the hilus of the gland a series
of intralobular duets which are arranged radially around it and are
of variable length, those which drain the portions of the lobule near
the hilus being short, those which go to the terminal portions of the
lobule longer. Each of these intralobular ducts drains a pyramidal

The Structure of the Mammalian Csophagus. 29

mass of tissue the apex of which is directed towards the duct, the
base towards the periphery of the lobule. These pyramidal masses
are the units of structure, being composed exclusively of tubules and
acini lined by secreting cells. As indicated above, the gland is
tubulo-acinous in type, the body of the unit composed of branching
tubules which terminate in bulb-like acini, tubules and acini being
occupied by the same type of epithelial cell. The acini show a
slight expansion of the lumen and the cells are frequently longer in
the acini than in the tubules, thus accounting for the difference
in size.

To the question of the presence of demilunes in the esophageal
elands of man I have given much attention. In view of the fact
that I have found these structures without difficulty in all cesophageal
elands of other mammals it was my expectation that they would
also be found in man. Accordingly I have studied carefully com-
plete series of sections, 5 micra thick, of glands from three different
wsophagi, using the methods of staining which I have found most
efficient in demonstrating these structures in other mammals. The
result of this study has, however, been wholly negative and I am
foreed to agree with Schaffer, who states that the human cesophageal
elands are pure mucous glands without demilunes. Referring to
the literature I find that neither Klein (79) nor Renaut (97), who
are quoted in this connection by Schaffer and Oppel, specifically
state that they found demilunes in the human cesophagus, although
the former found them in the dog and Renaut’s description applies in
general to the dog and man. Bohm and v. Davidoff (95) state that
there are but few mucous glands in the esophagus but that when
present they contain well marked demilunes, but it is diffieult to
determine whether these authors had before them the true cesophageal
elands occurring in the submucosa, or the superficial glands of
Riidinger which have been shown by Schaffer (97) and Hewlett
(00) to contain parietal cells and from which their illustration of
the cesophageal glands of man is clearly taken.

The character of the mucous cells forming the secreting tubules
is well shown in Fig. 12. Protoplasmic cells containing no mucin as
described by Schaffer occur also in my preparations, whole groups

30 Emil Goetsch.

of tubules being of this character. These I interpret, as did Schaffer,
as temporarily inactive mucous cells. These tubules are always sur-
rounded by a tissue containing large numbers of plasma cells.

The duets of the glands (Fig. 12) are lined by a stratified epithelium
except in the very beginning in the gland where for a short distance,
only 12 micra in some eases, a single layer of cylindrical cells is
found. The intralobular ducts have a double layer of cells and the
large ducts in the submucosa two to four layers of cells, the number
of layers increasing as the surface of the mucosa is approached. The
shape of the cells varies with the degree of tension of the surface,
produced either by distension of the duct with secretion or by the

Mucous
tubules

Fic. 12. Portion of a lobule of an cesophageal gland of man, showing
entrance of duct and transition of duct epithelium to glandular epithelium.
A. Lobular duct. B. Transitional duct lined by simple columnar epithelium.
<< 180.

formation of folds in the mucous membrane. In the relaxed duct
the superficial layers of cells are cylindrical, the deeper layers poly-
gonal except very near the epithelium, where the epithelium becomes
stratified squamous. In the distended or stretched duct both super-
ficial and deeper layers of cells become more or less fusiform in sec-
tion. In all cases the superficial layers of cylindrical or flattened
cells stain more strongly in eosin than the second layer, indicating
a probable change in a slight degree of the protoplasm in the direc-
tion of cornification.

As regards the superior cardiac glands of Schaffer and Riidinger
I have been able to make no study of them owing to the absence of

The Structure of the Mammalian (sophagus. 31

suitably fixed material. In one out of four wsophagi of which total
preparations were made to study the distribution of the glands, groups
of these glands were found in the upper part of the cesophagus
extending 4.5 em. below the lower border of the cricoid cartilage.
In this case twelve patches of these glands were found varying in
size from 1 mm. to 15 mm. in length and from 1 mm. to 3 mm. in
width.

The mucous tubules and acini are surrounded by a well marked
basement membrane composed of reticulum, and the tubules are bound
together by a reticular framework. No myoepithelial cells could be
demonstrated between the basement membrane and the epithelium
of the glandular tubules.

The epithelium in man is of considerable thickness, varying in
this respect with age, and with the degree of tension of the mucous
membrane. Another cause of variation in thickness in specimens
of the human csophagus is due to post-mortem loss of the super-
ficial layers of cells owing to maceration. In the newborn child it
has a thickness of 113 micra, in the adult from 260 to 440 micra.

The degree of cornification of the epithelium in man is about the
same as in the pig and considerably less than in the sheep and ox, that
is to to say that, although the superficial layers of cells show indica-
tions of chemical change in their greater transparency and in stain-
ing more readily in eosin, the degree of flattening of the cells is much
less than in the animals mentioned, and oval nuclei, which stain
well, are preserved even in the most superficial layers (Fig. 13).

The deep border of the epithelium is irregular owing to the pres-
ence of transitory folds of the lamina propria, described by Strahl
(89) and more particularly owing to the presence of large numbers
of high conical papille. The latter are arranged in linear rows
running parallel to the axis of the cesophagus and resembling in
their general arrangement those found in the skin of the palmar sur-
faces (Fig. 17). In some cases; as shown in the figure, the rows are
composed of several ranks of papille, the individual papille being
irregularly distributed.

The lamina muscularis mucose is well developed. It begins in
the lower part of the pharynx as scattered bundles of longitudinally

Slight cornification
of surface epithe-
lium

Str. germinativum

L. propria mucose

L. muscularis
mucose

Fic. 13. Transverse section of tunica mucosa of human cesophagus.
Ocular 3.

DO

The Structure of the Mammalian C¢sophagus.

.germinativum

oS

propria

i.

MUCOS

gus of sheep.

ucosa of cesopha

1a mM

if G8

ion oO

se sect

Transver

Wie. 14.

34. Emil Goetseh.

arranged unstriated muscle fibres, and rapidly becomes a complete
layer at the upper end of the cesophagus, attaining in the adult a
thickness of 0.4 mm.

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lig. 15. Fia. 16. Fie. 17.

Fics 15 and 16. Diagrams showing exact location and relative sizes of
every glandular lobule in two human @sophagi. Figures are reduced one-half.
The figures represent the oesophagus as seen when cut open along the poste-
rior border and spread out.

Iria. 17. Diagram showing exact distribution of papille on a small area
of human oesophagus. The rows are parallel with the cesophageal axis.

As regards the character of the tunica muscularis my observations
agree in the main with those of Welcker and Schweigger-Seidel

The Structure of the Mammalian Césophagus. 35

(61) except that I did not find the striated fibres extending so far
down. The following table shows the condition found on the pos-
terior wall of two human cesophagi.

No. LENGTH. | STRIATED. | MIXED. UNSTRIATED.
. | |
il. Longit. 21 cm. | 6.2 cm. 1.95 em. | 12.35¢em.
Cire. | Sol neGMme 4.63 cm. 12.35cm.
|
| Longit. 21.3 em. | 5.6 em 2.4 em. 13.3 em.
2. | Cire. | 3.6 em. | 4.75 em. | 12.95 cm.
| |

These measurements show that the unstriated fibers extend some-
what higher in the circular coat than in the longitudinal coat, but
that scattered striated fibers extend to about the same level in both
coats. Approximately the lower two-thirds of the cesophagus is pro-
vided with smooth fibres only. I did not find the striated fibres in
the lower end of the cesophagus which have been described by Gil-
lette (72) and Coakley (92). As Schaffer has shown, the unstri-
ated fibres may occasionally extend in the circular coat as far as the
lower border of the pharynx or even into the pharynx.

GENERAL CONCLUSIONS.

In addition to the specific data recorded in the preceding pages
certain general conclusions may be drawn from the observations and
from those of previous workers.

As regards the occurrence of cesophageal glands the species exam-
ined and those previously described may be divided into three groups:

A. Mammals in which the esophageal glands are wholly lacking
below the level of the cricoid cartilage. To this group belong all the
rodents examined, including the following species :—Arctomys monax,
Tamias striatus, Cavia, Erethizon dorsatus, Lepus cuniculus and
Lepus nuttalli, Mus decumanus, Sciurus hudsonicus, and Geomys
bursarius. In addition to these Sciurus vulgaris and Spermophilus
citillus have been reported by Oppel as lacking esophageal glands.
Here also belong the ungulate species Bos taurus, Ovis aries and
Equus caballus, the carnivor species Felis domestica, Vespertilio

36 Emil Goetsch.

fuscus belongiwg to the Chiroptera and according to Oppel the mar-
supial species, Trichosurus vulpecula, Aepyprymnus rufescens, and
Phascolarctus cinereus.

B. Mammals in which the glands are few in number. ‘To this
group belong Erinaceus, Scalops and Lutreola in which a few mucous
glands with demilunes occur in the upper end of the cesophagus, and
Mephitis in which a few glands of similar nature are to be found
at the. lower end of the cesophagus.

C. To the third class belong a small group of mammals belonging
to different orders in which the glands are present in considerable
numbers. These are Didelphys, Proeyon, Canis familiaris and C.
vulpes. Meles taxus and Nasua rufa, in which glands occur in
considerable numbers throughout the whole extent of the cesophagus,
Sus scrofa domestica in which they are abundant in the upper half
of the cesophagus and few in number in the lower half.

In structure the cesophageal glands are compound tubulo-alveolar
glands, consisting, except in those of man, of two kinds of cells,
mucous cells and serous cells. The latter elements, which have been
overlooked by previous observers with the exception of Klein (79),
Renaut (97) and Helm (07) are aggregated in the form of
demilunes such as we find in the submaxillary gland of the dog, and
like the demilunes of the salivary glands are provided with well
defined intercellular secretion canaliculi. In the glands of the lower
end of the csophagi of Procyon and Didelphys these serous cells
are so numerous that in many cases they form sessile alveoli on the
sides of the mucous tubules with an independent lumen from which
the secretion-canaliculi branch off. In man no demilunes could be —
found, thus confirming the observations of Schaffer.

A survey of the distribution of glands makes it apparent that the
occurrence of cesophageal glands is the exception rather than the
rule in mammals and that their presence bears no relation whatever
to the mechanical qualities of the food nor to the completeness of
mastication. Purely vegetable feeders are uniformly devoid of glands
in the esophagus and in many flesh feeders they are either few in
number or wholly absent. Ranvier it is true explains the absence of
glands in rodents on the basis of the thoroughness of mastication,

The Structure of the Mammalian (Ksophagus. 37

but if we examine the matter more closely we find that as a matter
of fact the csophagus of these animals has been specialized on
account of the coarse character of the food but in other ways than by
the production of mucous glands. On the other hand the theory that
the glands are developed in proportion to the needs of the animal
for lubrication of the food bolus does not adequately explain the fact
that in many Carnivora glands are few in number or wholly wanting
whle in others they are very abundant. If we examine the cases
of those animals which have a very large number of cesophageal
glands we find that there is no common quality as regards efficiency
of mastication relative to development of the salivary buccal and
cesophageal glands, or consistence of the food which would serve to
explain their presence. They are, however, without exception mixed
feeders, and this suggests that it is the composition of the food
rather than its consistence which has called forth the development
of mucous glands in the esophagus, or to express it differently, that
the secretion of these glands has a chemical value rather than a
mechanical one. This view is supported by the twofold cellular char-
acter of the glands, for it is difficult to understand why a serous
gland should have been developed where the need was simply the
mechanical need of lubrication.

In contrast to the glands the epithelium shows a very definite and
constant relation to the character of the food, and it is in the epithe-
lium that we find the esophagus undergoing specialization in those
animals which live on coarse vegetable food. The structure of the
epithelium is thus an accurate index of the character of the food
swallowed, inasmuch as we find a thickened and highly cornified
epithelium in those animals which live on coarse food, particularly
vegetable feeders, and a thin slightly cornified epithelium in animals
living on soft food, for example, the carnivors.

Correlated with this thickening of the epithelium we find, as
might be expected, an increase in the development of the muscularis
mucose, the probable function of this layer being to retract the
mucous membrane above the descending bolus of food. Conversely,
the muscularis mucose and the epithelium exhibit these specializa-
tions in a less degree in those animals in which large numbers of

Emil Goetsch.

co
ve)

mucous glands are present, indicating that, although the glands have
been developed primarily for another purpose, their secretion never-
theless serves the purpose of lubrication where it is present, and in
so doing modifies the degree of specialization of the epithelium and
muscularis mucose. For example in the pig, in the upper end of
the esophagus where glands are abundant, no muscularis mucose
is to be found, while in the lower end where few glands are present,
it is highly developed.

It will be noted that with regard to the processes of connective tis-
sue which project into the epithelium and which are usually described
as papille of the lamina propria mucose the observations here
recorded confirm in general the conclusions of Strahl (89), who
found that in many cases they were not true papille but elongated
ridges of the lamina propria running in a direction parallel to the
long axis of the cesophagus and connected with one another by oblique
ridges. This is a matter in which it is very easy to be mistaken
because ridges cut across are very similar in appearance to papillee.
It is only by examining earefully series of sections, or whole prepara-
tions of the epithelium, that this mistake can be avoided. Among
the mammals examined true papille were found in the pig, ox, horse,
sheep, and man and were associated in all these cases with longi-
tudinal ridges of the lamina propria. In Didelphys, Arctomys,
Sciurus, Tamias, Lepus, Canis, ridges only were present, although
in Cavia and Tamias the irregular summits of the ridges afford
suggestion of beginning papillae. In Procyon there are longitudinal
ridges also with suggestions of low papillze on their summits, while
in Mus, Geomys and Erethizon, there are neither ridges nor papille,
but an epithelium of fairly uniform thickness. In Mephitis, finally,
we have represented the exact opposite of connective tissue papille,
inasmuch as the epithelium sends processes into the subjacent con-
nective tissue. This is of course to be derived from a further devel-
opment of the system of longitudinal and transverse ridges which
have become so numerous and close that they surround papilla-like
processes from the deep surface of the epithelium.

With regard to the phylogeny of the cesophageal glands, the fact
that in whole orders no glands are to be found and that each of the

The Structure of the Mammalian Csophagus. 39

principal orders both of marsupials and placentals contain individuals
wholly lacking in cesophageal glands indicates that these structures
have been developed independently in the different orders in response
to needs that have arisen as result of food specialization. The alterna-
tive hypothesis that they were originally present in all mammals
and that they have disappeared from some is scarcely worth considera-
tion in view of the general absence of similar glands in lower verte-
brates.

In man alone is there any indication that the cesophageal glands
are disappearing. Here the great variability in number of the glands
and the constant presence of cyst formation and stasis of the secre-
tion and atrophy of the glandular cells might be taken as an indica-
tion of a disappearing structure.

Tn conclusion I desire to express my thanks to Prof. R. R. Bensley
for valuable suggestions during the course of this investigation. 1
desire also to thank Miss Katharine Hill for the accompanying

drawings.

BIBLIOGRAPHY.

BENSLEY, R. R., 00. The Gsophageal Glands of Urodela. Biological Bulle-
tin, Laneaster. Vol. 2, pp.
, 03. The Structure of the Glands of Brunner. The Decennial
Publications, University of Chicago. Vol. 10, pp. 279-326.

Brummer, J., 76. Anatomische und histologische Untersuchungen iiber den
zusammengesetzten Magen verschiedener Siugetiere. Deutsche
Ztschr. fiir Thiermed., Leipz., Bd. 2, pp. 158-186 and 299-319.

Caruier, EH. W. Contributions to the Histology of the Hedgehog (Hrina-
ceus europaeus). Part I. The Alimentary Canal. J. Anat. and
Physiol., London, Vol. 27, pp. 85-112.

Coaktey, C. S., 92. The Arrangement of the Muscular Fibres of the Csoph-
agus. Researches of the Loomis Laboratory of the Univ. of the
City of New York, Vol. 2, pp. 1138-4.
Decker, F., 92. Zur Physiologie des Fischdarmes, Festschr. Albert y. K6l-
liker, Leipzig, pp. 389-411. .
DosrowoLskI, Z., 94. Lymphknotchen (folliculi lymphatici) in der Schleim-
haut der Speiserdhre, des Magens, des Kehlkopfs, der Luftréhre und
der Scheide. Beitr. z. path. Anat. u. z. allg. Path., Jena, Bd. 16,
pp. 48-101.

EXLLENBERGER, W., 94. Handbuch der vergleichenden Histologie und Physio-
logie der Haussiiugetiere, bearbeitet von Bonnet, Csoker, Hichbaum,
ete, Bd. I, Histologie, Berlin, Parey.

40 Emil Goetsch.

Fiesu, M., ’83. Ueber Beziehungen zwischen Lymphfollikeln und secre-
tierenden Driisen im Gisophagus. Anat. Anz., Jena, pp. 283-6.
Gitterre, 72. Description et structure de la tunique musculaire de cesoph-
age chez homme et chez les animaux. J. de J’anat. et physiol.
(etc.), Paris, Vol. 8, pp. 617-44.

Grarr, K., 80. Lehrbuch der Gewebe und Organe der Haussiiugetiere. Jena.

HAAne, G., 05. Ueber die Driisen des Gisophagus und des Uebergangsge-
bietes zwischen Pharynx und Césophagus. Arch. f. wissensch. wu.
prakt. Thierh., Berlin, Vol. 31, pp. 466-83.

Heim, R., O07. Vergleichend anatomische und histologische Untersuchungen
iiber den Gsophagus der Haussiiugetiere. Inaug. Diss., Ziirich.

Hewtert, A. W., 01. The superficial glands of the Gsophagus. J. Exper.
Med., New York, Vol. 5, pp. 319-32.

Kier, E., ’°71. Handbuch der Lehre von den Geweben, herausgegeben yon
S. Stricker. Leipzig, Bd. 1, pp. 378-88.

"79. Observations on the Structure of Cells and Nuclei. II.

Epithelial and Gland-cells of Mammals. Quart. J. Micr. Sc., London,
n, s., Vol. 19, pp. 125-75.

LANGLEY, J. N., 79. On the Changes in Pepsin-forming Glands during Secre-
tion. Proc. Roy. Soc., London, Vol. 129, pp. 383-88.

Oppet, A., 97. Lehrbuch der vergleichenden Mikroskopischen Anatomie der
Wirbeltiere. Teil 2, Schlund und Darm. Jena, Fischer.

Ranvier, L., 84. Les membranes muqueuses et le systéme glandulaire. J.
des microg., Paris, Vol. 7, pp. 77-86.

RENAUT, J., 97. Traité dhistologie pratique. Paris, Vol. 2, pp. 590-600.

Ruse, C., 90. Ueber den CGsophagus des Menschen und der Hausthiere.
Arch. f. wissensch. u. prakt. Thierh., Berlin, Vol. 16, pp. 1-28 and
161-98.

Scuarrer, J., 97. Beitriige zur Histologie menschlicher Organe. IV. Zunge.
V. Mundboéhle-Schlundkopf. VI. Csophagus. VII. Cardia. Sitz-
ungsb. d. k. Akad. d. Wissensch. Math.-Naturw. Cl., Wien, Vol. 106,
Abth. 3, pp. 353-455.

Sréur, P., 87. Ueber Schleimdriisen. Festschr. Albert v. K6lliker, Leipzig,
pp. 421-44.

Sraut, H., 89. Beitriige zur Kenntniss des Baues des Gisophagus und der
Haut. Arch. f. Anat. u. Entweklngsgsch., Leipzig, pp. 177-195.

Swiecick!, H., v., 76. Untersuchungen iiber die Bildung und Ausscheidung
des Pepsins bei Batrachiern. Arch, f. d. ges. Physiol., Bonn, Vol. 13,
pp. 444-52.

Toutpt, C., ’88. Lehrbuch der Gewebelehre mit vorzugsweiser Berticksichti-
gung des menschlichen Ko6rpers. Stuttgart, Enke.

Voet, C., and Yune, E., 94. Lehrbuch der praktischen vergleichenden Ana-
tomie. Braunschweig.

THE LIMIT BETWEEN ECTODERM AND ENTODERM IN
THE MOUTH, AND THE ORIGIN OF TASTE BUDS.’

I. AMPHIBIANS.

BY
J. B. JOHNSTON.

WitH 21 TExtT FIGURES.

It has been generally recognized that in man it is not possible to
locate exactly the limit between ectoderm and entoderm after the
rupture and disappearance of the pharyngeal membrane. At the same
time it is clear that the tongue arises from the lower ends of the
visceral arches and that the floor of the mouth is in the greater part
lined by entoderm. In the roof the facts are not so clear, but it is
believed that the stomodzeum is much deeper above than below. Ob-
viously it can not include the eustachian tube nor the nasal cavity,
although the latter is of ectodermal origin. How much of the max-
illary process and the palatal shelf is covered by ectoderm, and how
much by entoderm, is not known.

The taste buds are all located in the pharyngeal portion or floor
of the mouth, with the exception of those in the soft palate. Whether
the soft palate is covered by ectoderm or entoderm remains to be
determined. In view of the fact that at least the great majority
of the taste buds are found in the area which all authors agree is
lined by entoderm in the embryo, the question may be asked what
reason, if any, is there for considering that the taste buds are not
of entodermal origin? The chief reason is the general doctrine that
nervous structures are of ectodermal origin. The history of that
doctrine can not be traced here, but it may be said that it does not
appear to be based upon an exhaustive study of the origin of all

‘Neurological Studies from the Institute of Anatomy, University of Minne-
sota, No. 7.

THH AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

42 J. DB. Johnston.

nervous structures. The origin of certain visceral plexuses in man
and of various peripheral nerve cells in other vertebrates has not
been determined, and the origin of taste buds is at present under
discussion. The statement that all nervous structures are derived
from the ectodermal layer of the embryo is not warranted by the
known facts. Another reason for thinking that taste buds are of
ectodermal origin is that in certain fishes they are found in the later
embryo and adult in ectodermal territory, namely, the outer skin.

In 1898 I made the statement that in the head of vertebrates ‘all
sensory structures of ectodermal origin are supplied by components
of the Vth (including spinal Vth components running in other
nerves), VIIIth and lateral line nerves, and that all fibres supply-
ing such structures have their central endings in the nucleus funiculi,
tuberculum acusticum or cerebellum. . . . On the other hand, all
sensory structures of entodermal origin are supphed by VIIth, I Xth
and Xth components, and all fibers supplying such structures have
their central endings in the lobus vagi.”

The inclusion of taste buds among structures of entodermal origin
was adversely criticised by Strong (1898), and it did not appear
that any neurologist was ready to entertain the idea that taste buds
were possibly of entodermal origin. I have since brought forward
evidence (1905) that in petromyzonts the taste buds arise in ento-
dermal area. In the ammoceetes stage I was able to find them only
in entodermal surfaces. In teleosts (Coregonus, Catostomus) the
buds first appear in the pharynx and cesophagus where there seems
to be no possibility of origin from any other source than entoderm.
The appearance of taste buds in entodermal area in teleosts has since
been confirmed by Landacre (1907), working on Ameiurus. He
recognized, however, the possibility of their being formed in ectoderm
also.

Marrriat and Mrrnops.

I have undertaken to study the limits of ectoderm and entoderm
in the mouth of several vertebrates, both for its general interest and
with especial reference to the origin of taste buds.

Amphibians offer especial advantages for the study of the relations

The Limit Between Ectoderm and Entoderm. 43

of ectoderm and entoderm owing to the persistence of yolk in the
entoderm cells after it has disappeared from the ectoderm. The
early development of the mouth has been studied fully in Amblys-
toma punctatum and the following description applies wholly to this
form. Necturus and frog embryos have been studied sufticiently
to control the chief facts.

While at West Virginia University I made a large collection of
Amblyostoma embryos and larve from the unsegmented egg to tad-
poles over an inch in length (about March 20 to July 1). Various
methods were tried both for fixation and staining. The fixing fluid
that gave the best results was the formol-sublimate-acetic mixture
devised by Worcester.? All the embryos used for this study were
fixed in that fluid. For staining, borax carmine, hemalum, heema-
toxylin with counter stains have been used. Sections prepared for
this particular purpose have all been stained by iron hematoxylin
and counter stained by acid fuchsin. It was found that iron heema-
toxylin stains yolk granules more intensely than any other element
in the tissues and remains in the yolk granules in the differentiating
bath until it is removed from all other structures, even the chromatin.
By differentiating to the last degree favorable for study of chromatin
and counter staining with acid fuchsin, clear transparent prepara-
tions are obtained in which the yolk granules stand out boldly on a
pink ground, while the nuclei and cell boundaries are lightly stained
but distinctly seen.

As is well known, the ectoderm and entoderm differ from the
earliest stages in the number and size of yolk granules contained
in the cells. This difference is readily made out in any sections
of well fixed material by careful study. In preparations stained as
above described the differences are sharply marked and become more
prominent and striking as development proceeds. The yolk granules
in the ectoderm grow smaller and less numerous and disappear at a
relatively early period. In the brain the yolk persists longer than in
the ectoderm, but in such small granules that there is no comparison
with the entoderm. The muscle-forming masses and some parts

*Formula: take 10 per cent formol, saturate with sublimate, add glacial
acetic acid to make 10 per cent of the whole.

44 J. B. Johnston.

of the mesenchyme retain large quantities of yolk, but these do not
enter in any way into the structures to be studied in this connection.
The ectoderm and entoderm are drawn as faithfully as possible in
the figures. When it is remembered that the general gray shading
of the cells represents fuchsin stain, while only the granules and
chromatin are black, it will be seen that the distinction between ecto-
derm and entoderm is many times more striking in the preparations
than it is in the figures. In late stages, when the yolk begins to
be used up in the entoderm, the sharp staining of the granules is of
the greatest advantage. Before that time the yolk has entirely dis-
appeared from the ectoderm and most of it from the nervous system.

In such a study as this good serial sections without tearing or
distortion are necessary, and the yolk itself is well known as a great
obstacle to obtaining such sections. I have tried some methods of
fixation in which the yolk is said to be made soft and easily cut. I
have not found, however, that such embryos show such perfect fixa-
tion as I desired for the study of the early stages in the formation
of taste buds. The fixing fluid used renders the yolk hard and brittle,
but fixes faithfully all elements of the tissues and without any dis-
tortion or shrinking. To obtain perfect serial sections in the three
planes I used the method of imbedding in a paraftin-rubber mixture
made as follows: to 100 ce. of paraffin which melts at 1 or 2 degrees
higher temperature than desired in the imbedding mass, add 1 to
9 grams of crude India rubber cut in as small bits or shreds as pos-
sible. Heat over boiling water 24 hours or let stand in the bath at
60° ©. for three days. Pour off the clear melted paraffin and keep
in the solid state. Use exactly as paraffin, clearing specimens for
imbedding in xylol or toluol. With this method I have obtained
many series of sections of all stages of Amblystoma as perfect as
it is usual to obtain from ordinary objects. Obviously I could not
use for this study preparations in which the yolk-bearing tissues
were torn, or in which the yolk granules were scraped out of their
position and driven along by the knife. In the figures given every yolk
granule is faithfully drawn under the camera as accurately as pos-
sible in its relations to the cell boundaries and nuclei. No yolk
granule is omitted because it seems to be displaced. No drawings

The Limit Between Ectoderm and Entoderm. 45

are taken from sections which show the slightest tearing or breaking

in the parts concerned.

1. Early Form and Relations of Entoderm.

After completion of gastrulation the archenteron has the well
known form characteristic of amphibian embryos: The enlarged
anterior part of the archenteron is bounded in the region where the
mouth will form by a relatively thin layer of entoderm and a thinner
layer of ectoderm (Fig. 1). In this region are two broad, very
shallow depressions in the ectoderm. The more anterior one repre-
sents the hypophysis, the posterior one the future mouth. Opposite
these the archenteron itself presents two prominent pits (or angles)
as seen in sagittal sections. The anterior one is a slender pointed

S|.

n eb

Fic. 1. Amblystoma punctatum, neural plate stage, sagittal section of mouth
region. Borax carmine stain.

cavity (Fig. 2), and the entoderm surrounding it forms a blunt
wedge projecting between the neural plate and the ectoderm. ‘The
posterior one is a broad depression which corresponds to the future
mouth opening. The entoderm surrounding the anterior pit is
the preoral entoderm and corresponds in every way to the preoral
entoderm of most selachians (cf. Johnston, 1909).

46 J. B. Johnston.

When the neural plate rolls up into a tube (Fig. 2) pressure is
exerted upon the archenteron by the brain portion of the neural
tube and the effect is seen in a compression of the archenteric space.
The two angles seen in sagittal sections are now more marked and
are separated by a thickening or fold of the intervening entoderm.
In the meantime the notochord and mesoderm are forming in the
same manner as in selachians. In the stages represented in Figs. 2
and 3 the notochord ends anteriorly in a median mass which is not
separated from the entoderm and is continuous laterally with the
mesoderm which is in process of splitting off from the entoderm.
This median mass remains in continuity with the preoral entoderm.

Ive, 2 lhe, 8%

Fic. 2. Amblystoma punctatum, neuropore stage. The foregut is already
compressed and the palestomal (p) and neostomal recesses are sharply
marked. Borax carmine.

Fic. 3. A. punctatum, after closure of neuropore. Borax carmine.

I have indicated in an earlier paper (1903) that the mesoderm
formation is somewhat delayed in the head of Amblystoma so that
the mesoderm of the hyoid and mandibular arches is split off from
the entoderm as separate rods of mesoderm. The median mass
mentioned is continuous laterally with the mesoderm which forms the
premandibular somite (Fig. 4, A and B). The splitting off of the
notochord stops at the posterior border of this median mass and in
later embryos the point at which the end of the notochord remained

The Limit Between Ectoderm and Entoderm. AT

longest in connection with the entoderm is marked by a sharp notch
(see Johnston, 1906, Fig. 36). In front of this the entoderm is not
thick as in selachians so that the term “median mass” is not so appli-
cable. However, from this part of the entoderm mesenchyme splits
off and leaves the entoderm very thin, often scarcely a continuous
membrane. This thin area is soon filled up by the shortening and

p cm.

IG. 45 A Jane, 2E 18

Fic. 4. A. punctatum, two parasagittal sections to show the relations of the
head mesoderm. Hzemalum.

shifting back of the preoral entoderm and by the general compression
which soon obliterates the anterior end of the archenteric cavity.

2. Development of Hypophysis.

The ectoderm which will form the hypophysis can be recognized
as soon as the neural plate is formed (Figs. 1, 2, 3). Below the

48 J. B. Johnston.

terminal ridge of the neural plate (see Johnston, 1909) the ectoderm
grows thinner. The wedge-shaped piece of ectoderm immediately
adjacent to the neural plate will form the hypophysis. Its position
is accurately indicated in Figs. 1 and 2. When the preoral entoderm
is more blunt or rounded the hypophysis presents in the earliest
stages a triangular form in sagittal sections (Fig. 3), one angle
being directed inward between the terminal ridge of the neural plate
and the preoral entoderm. When the neural plate rolls up the
terminal ridge participates in the rising up of the neural folds, and

en

Kia. 5. A. punctatum, stage when hypophysis begins active invagination.
Median sagittal section. Haemalum.

the distinction between the neural plate and hypophysis at once
becomes clearly marked (Figs. 2 and 3). From the earliest stage
the hypophysis rests against the tip of the preoral entoderm.

While the neural tube is rolling up, the hypophysial ectoderm be-
comes deeper, so that in sagittal section the depth of the triangle
becomes greater than the length of its base on the surface (Fig. 3).
Up to this time the cells within the triangle show no regularity of
arrangement. Immediately following this it is seen that the deeper
cells are arranged so as to suggest a short blind sac. From the start
(Fig. 5) this sac is thicker on the anterior or dorsal side. It has

The Limit Between Eetoderm and Entoderm. 49

no actual cavity, but the arrangement and pigmentation of the cells
suggest a cavity, and the wall next to the entoderm is thin and devoid
of nuclei. By this time the ectoderm of the mouth plate, caudal to
the hypophysis, consists of only a single layer of flat cells (see

Fie. 6. A. punctatum, in a stage slightly advanced over that in Fig. 5.
Medial sagittal section of hypophysis. Borax carmine.

Fic. 7. A. punctatum, at the height of the hypophysial invagination before
separation from ectoderm. Note the concavity of hypophysis and the absence
of ectoderm from the mouth plate. Heemalum.

below). As the hypophysis pushes in, its thin posterior or ventral
wall is devoid of cells, and soon the cavity of the hypophysial invagi-
nation is bounded on that side by entoderm (Figs. 6 and 7). The
hypophysis appears as a hook-shaped membrane (sagittal section)

50 J. B. Johnston.

extending in from the deeper layer of the ectoderm. A slight cavity
is contained within the recurved wall at the deeper end (Fig. 7),
the rest of the ventral wall being formed by entoderm.

While the hypophysis is pushing in, the preoral entoderm is pushed
back, flattened, and its cavity nearly obliterated. This cavity be-
comes less prominent before the ‘closing up of the entodermal mouth

Via. 8. A. punctatum, nearly median sagittal section to show the persistence
of the preoral or paleeostomal recess in the entoderm. The invagination of
the hypophysis is very tardy in this specimen. THeemalum.

pit. There is always to be seen, however, a shallow pit or an angular
prolongation of the archenteron which continues to indicate the
preoral cavity (Figs. 8, 9, 10). In some eases a cleft appears in
the entoderm leading toward the cavity of the hypophysis. By this
time the entoderm has been compressed and thickened, but the per-

The Limit Between Ectoderm and Entoderm. 51

sistence of the preoral cavity (in various degrees) serves to mark
the position of the preoral entoderm. It also explains the peculiar
form taken by the hypophysis. The cleft meeting the cavity of the
hypophysis suggests that the intact wall of the hypophysial invagina-
tion constitutes, with the entoderm dorsal to the cleft, the dorsal
wall or roof of the palzeostoma, while the absence of a ventral wall
to the hypophysial invagination allows the hypophysis to open into

Fic. 9. A. punctatum, after separation of hypophysis from ectoderm.
Medial sagittal section. The pointed recess of the archenteron directed toward
the letter m is the oral recess; the slight pit or concavity next to pr. en. is
the palzeostomal recess. Hzemalum.

the archenteron, the ventral wall or floor of the palseostoma being
formed by entoderm alone. These relations are indicated im a
diagram (Fig. 11).

After the stage at which the partial connection between hypophysis
and archenteron is seen, there is a further bending of the brain, the
flexure becoming excessive in amphibians. The bending, together
with the expansion of the forebrain, results in pushing the hypophysis

52 J. B. Johnston.

and preoral entoderm farther and farther back, until they “come
to lie close to the end of the notochord. Compare Figs. 3 and 10.
It is this movement of the preoral entoderm dorso-caudally that was
referred to above as the means of thickening up the wall of the archen-
teron which is left so thin by the splitting off of the head mesen-
chyme. The hypophysis finally comes to lie on the dorsal surface
of the preoral entoderm, and the notch which represents the original
preoral cavity is visible up to the stage when the velum transversum,
epiphysis, paraphysis and the inferior lobes of the brain are all
well formed (Fig. 10). The formation of the saccus vasculosus

Fic. 10. A. punctatum, stage after formation of epiphysis. Median sagittal
section. The palzostomal recess is a deep angular pit. The dental ridges:
are growing in. Heemalum.

and the differentiation of other structures in the region of the inferior
lobes takes place considerably later.

The shifting caudad of the preoral entoderm and hypophysis which
I have attributed to the brain flexure is accompanied by a very con-
siderable elongation of the mouth entoderm. While the shifting is
taking place the mouth entoderm is compressed into a solid mass,
and this becomes elongated into a comparatively slender cord, flat-
tened dorso-ventrally (Figs. 10, 12, 14, 16).

For some time after its separation from the ectoderm the hypophy-
sis continues to have the form of a shallow cup with the convexity

The Limit Between Ectoderm and Entoderm. 53

against the brain, the concavity toward the entoderm (Fig. 9). It
finally forms a somewhat ovoid mass and lies beneath the brain.
Its further history does not concern us in this study,

3. Coalescence of the Walls of the Foregut.

It has already been indicated that the formation of the neural tube
begins to compress the anterior part of the archenteron. This is

Vie. 11. A diagram of the relations of the palzeeostoma and neostoma in A.
punctatum. A camera drawing was made from the section from which Fig.
7 was drawn and the cleft connecting the archenteron with the hypophysis
was made wider, This is the only diagrammatic feature of the drawing. The
palzeostoma is represented by the cleft hy- p.

earried further by the formation of the mesodermic somites above
and at the sides and of the heart below, and the bending down of
the brain completes the influences which result in the complete oblit-
eration of the cavity of the anterior part of the future mouth (Figs.
12, 14). This condition is well known in amphibian embryos and
requires little comment. ‘Two things should be noted. (a) The
coalescence of the walls begins early and is followed by a relatively

4. J. B. Johnston.

Or

ereat elongation of the pharyngeal region. The necessity of this to
make room for the branchial apparatus, heart and brain, is clear
from Figs. 3, 9 and 10, and it is this chiefly which causes the for-
ward projection of the head from the yolk and the straightening
out of the embryo. (b) When the oral cavity appears it leads to the
place which has been indicated in the early embryo as the future
mouth opening. The history of the hypophysis and preoral ento-
derm given above and the later history of the preoral entoderm make

this clear.

‘SS.

Vic. 12. <A. punctatum, nearly median sagittal section of oropharyngeal
region after the coalescence of the walls of the primitive mouth cavity. The
yolk-laden entoderm is surrounded by the ingrowing dental ridges. Iron
hematoxylin, fuchsin.

4, Disappearance of Mouth Plate Hcetoderm.

In Amblystoma no stomodzeum is formed. Instead, the ectoderm
over the mouth plate area disappears, leaving the solid entoderm
exposed on the free surface. Later the mouth cavity appears as a
cleft in this entoderm.

The well known arrangement of the cells of the ectoderm in two
layers is clearly marked in the mouth plate area of Amblystoma in
the neural plate stage. As the neural tube rolls up the ectoderm

The Limit Between FEetoderm and Entoderm. 55

becomes somewhat thinner, but not as thin as that covering the
ventral surface of the yolk mass. Following this the inner cells of
ectoderm disappear from the mouth plate. In either transverse or
sagittal sections the entoderm is seen to be covered by the thin flat
cells of the outer layer of ectoderm, while the inner layer of more
cuboidal cells stops abruptly around the borders of the mouth plate
(Figs. 5, 6, 13). Anteriorly the mouth plate is bounded by the
hypophysis. I have not been able thus far to see in detail how this

Fic. 13. Detail drawing of the mouth plate and dental ridges in the section
drawn in Fig. 12. Here it is seen that the outer layer of ectoderm is partly
broken down but still covers the entoderm.

condition comes about. Whether the inner cells of ectoderm migrate
apart and draw away from the mouth area or whether some of these
cells disintegrate in situ is not wholly clear. I have seen no signs
of disintegrating cells, and there is to be seen a heaping up or thick-
ening of the inner layer of ectoderm around the mouth plate. I am
strongly inclined to think that the inner cells draw away from the
mouth plate in preparation for the formation of the dental ridges
to be described below. Certain it is that by the time the hypophysis
begins to be actively invaginated (Fig. 6) the ectoderm of the mouth

56 J. B. Johnston.

plate consists of the thin outer layer alone. As development goes
on this thin outer layer gradually breaks down and its cells seem to
scale off, leaving the yolk-laden entoderm cells freely exposed (Figs.
i 8; 0.10) 12, A@rotdonione

This disappearance of the mouth plate ectoderm I consider com-
parable to the rupturing of the pharyngeal membrane so far as its
ectodermal portion is concerned. The entodermal layer of the mem-
brane is not ruptured at this time beeause of the mechanical condi-

mand

Tic. 14. A. punctatum, stage of tooth formation. Median sagittal section
of the mouth region. The oral entoderm is drawn out into a slender flattened
rod and the formation of the mouth cleft will soon take place. A mandibular
tooth is seen in vertical section and dorsal to the entoderm, the ectodermal
in-growth in which the maxillary and vomerine teeth will appear is distin-
guished by the absence of yolk. Between d. max. and d. vom, is the projection
of entoderm which later meets the nasal sac. Iron hematoxylin, fuchsin.

tions which lead to the coalescence of the walls of the archenteron.
The appearance of the oro-pharyngeal cavity as a cleft in the entoderm
completes the process, which is comparable to the rupturing of the
pharyngeal membrane in other vertebrate embryos.

5. Formation of Dental Ridges and Teeth.

The thickening of the inner layer of ectoderm surrounding the
mouth-plate mentioned above soon becomes prominent, and in both

The Limit Between Ectoderm and Entoderm. 57

sagittal and transverse sections it is seen that this layer turns in
around the entoderm (Figs. 8, 9, 10, 12, 13). This in-growing
ectoderm pushes in farther on the dorsal surface of the entoderm than
on the ventral and soon forms a ring or collar around the solid oral
entoderm. At first the ring is incomplete in its mid-dorsal portion
where the inner layer of ectoderm is taken up in the formation of
the hypophysis. The ingrowing ectoderm spreads to fill up this

Fic. 15. A. punctatum, formation of dental ridges. A transverse section
through the mouth plate after disappearance of the ectoderm. Hzemalum.

gap, and pushes up on the dorsal surface of the oral entoderm to
about the point of contact of preoral entoderm and hypophysis.
This mid-dorsal portion of the in-growing ectoderm gives rise to the
vomerine teeth. The in-growing ectoderm forms the dental ridges
in which the teeth soon begin to make their appearance (Figs. 12-
18). For the purpose of this study the formation of the teeth need
not be followed in detail. One point only is of especial interest
here, namely, that when the teeth are formed they are separated from

58 J. B. Johnston.

the mouth eavity by a layer of entoderm. The eruption of the teeth
involves the piereing of this entoderm (Figs. 14, 16, 18, 20),

6. Appearance of Oro-Pharyngeal Cavity.

The splitting of the solid entoderm takes place progressively from
behind forward at a stage after the teeth are well formed in the
dental ridges (Figs. 14, 16, 17). By the time the splitting of the
oral entoderm takes place the mouth region has grown wider from side

Tia. 16. A. punctatum, appearance of mouth cleft. Median sagittal section
of the mouth. The ectoderm and dental ridges are more deeply shaded. The
yolk granules are all drawn and the outline between ectoderm and entoderm
is taken from the cell boundaries. Iron hematoxylin, fuchsin.

to side in anticipation of the future wide mouth. The whole anterior
part of the entoderm has taken part in this widening, so that the
oral entoderm is now a broad flat solid plate. As the pharyngeal
cavity progresses toward the mouth it advances most rapidly toward
the lateral angles of the mouth. When the tooth germs begin to
form there is a thin place left in the ectoderm between the maxillary
and vomerine teeth. At this place the entoderm forms an elbow
dorso-cephalad which is quite pronounced at the lateral border of
the tooth-forming areas (Figs. 14, 17, 18). In the meantime the

The Limit Between Ectoderm and Entoderm. 59

nasal sacs have been formed and the wall of the nasal sac comes into
contact with the elbow of entoderm lateral to the maxillary and
vomerine teeth. The pharyngeal cleft invades this part of the
entoderm first, and a connection is established between the cavity of
the nasal sac and the oro-pharyngeal cavity (Fig. 17). In this
way the alimentary canal has two openings to the exterior through
the nasal saes for one or two days before the mouth is open. When
the mouth cleft is finally completed the entoderm lines the oral

—_

~ nasal sae

Fic. 17. Parasagittal section from the same series as Fig. 16. The section
passes through the opening of the nasal sae into the oral cavity.

cavity to the extreme borders, and even extends out slightly on the
surface (Figs. 16, 17). Now the maxillary, mandibular and vome-
rine teeth pierce the entoderm and enter the mouth cavity.

In the series of embryos used for this study the limits of the
entoderm are clearly visible for a period of about ten days after the
mouth cleft was formed. (The rate of growth was slow as the
running water in the laboratory was colder than that in the ponds.)
The entoderm during this period is thicker than the ectoderm, con-
tains numerous yolk granules which are lacking in ectoderm and is
more opaque in appearance in sections. The entoderm lines the

60 J. B. Johnston.

whole oro-pharyngeal cavity, meets the ectoderm about midway on
the anterior and posterior surfaces of the branchial arches in the
gill slits and meets the ectoderm in front of the maxillary and
mandibular teeth at the lips.

7. Formation of Taste Buds.

The taste buds begin to be formed shortly before the mouth cleft
forms. They appear eventually, more or less numerous, on all parts
of the floor and roof of the oro-pharyngeal cavity forward to, but not
in front of, the maxillary and mandibular teeth. The buds appear

d.mand.

Fic. 18. <A. punctatum, stage when the teeth are forming. Median sagittal
section of mouth region to show forward projection of entoderm between
maxillary and vomerine teeth through which the connection of nasal and oral
cavities is formed. Iron hematoxylin, fuchsin. é

first in the pharynx and later farther forward, as I found to be the
case in teleosts (1905). In the larval stage of Petromyzon they are
found in the pharynx only. The buds in the greater part of the roof
of the mouth and pharynx and those in a large part of the floor are
formed in entoderm far removed from any ectoderm. There is, and
can be, no question as to their origin from entoderm. ‘The buds
which are formed on the gill arches are always on the inner surface,
and there is no reason to doubt that they are formed from entoderm
cells as are those in the roof of the pharynx far removed from the
ectoderm. The buds which are formed in the vicinity of the teeth
are the ones which one might think would be derived from the in-

The Limit Between Eetoderm and Entoderm. 61

growing ectoderm. ‘The fact that this ectoderm forms dental ridges
does not wholly exclude the supposition that some of the cells might
give rise to taste buds. Therefore, an account of the mode of forma-
tion of taste buds in the front part of the mouth will be sufficient to
determine whether any of the taste buds are derived from ectoderm.

Fig. 19 shows a taste bud forming in the floor of the mouth a
short distance behind the mandibular teeth at the time when the
mouth cleft is forming. The ectoderm is given a darker shade than

Fie. 19. A. punctatum, sagittal section near the median plane shortly before
the formation of the mouth opening. The section includes a part of the dental
ridge in the mandible and the tip of the hyoid arch. Between the two is seen
a taste bud forming in the entoderm. The asterisk marks the caudal limit of
the ectoderm of the vomerine tooth-germs. Iron hematoxylin, fuchsin.

the entoderm and all the yolk granules are drawn. For a short dis-
tance in the middle of the figure the nuclei are drawn. A small iso-
lated opening indicates the forming mouth cavity. Below it appear a
group of elongated nuclei placed vertical to the floor of the mouth.
This grouping of nuclei is the first indication of a taste bud. The
outlines of the cells are difficult to see on account of the great number
of yolk granules. This taste bud is forming in the entoderm between
the tongue and the teeth in the floor of the mouth. It is surrounded
on all sides by entoderm crowded with yolk granules and these gran-

62 J. B. Johnston.

ules are closely packed around and among the nuclei of the cells
which form the sense organ. It presents the same appearance as do
early taste buds in the roof of the pharynx or in any part of the
mouth, whether near the ectoderm or far from it.

Another taste bud in a shghtly later stage of development is shown
in Fig. 20. It is situated in the roof of the mouth just behind the
most lateral one of the vomerine teeth. The cells immediately around
the tooth together with the three or four deep nuclei at the right hand
end of the figure (asterisk) come from the ectoderm of the dental
ridge. All the rest of the cells are entoderm. At the left, several nuclei
vertically arranged indicate the beginning of a taste bud. It contains

Fic. 20. A. punctatum after the formation of the mouth cleft. Sagittal
section of the roof of the mouth in the vomerine region. At the left is a taste
bud forming. At the right the asterisk marks the border of the ectoderm.
Iron hematoxylin, fuchsin.

and is surrounded on all sides by large yolk granules such as are found
in this embryo only in entoderm and in parts of the mesoderm.
The ectoderm has no yolk at all in embryos of this stage and the
nervous system has very little.

In Fig. 21 are shown three taste buds and one neuromast from a
larva four or five days after the formation of the mouth opening.
The entoderm is still clearly distinguishable from the ectoderm and
extends beyond the teeth to the lips. At A in the figure is a taste bud
standing near the vomerine teeth. The deep nuclei which show active
nuclear changes belong to the ectoderm of the vomerine dental
ridge. The two cells with dark nuclei belong to the mesenchyme.
These cells and the definite outline of the entoderm show that there

The Limit Between Eectoderm and Entoderm. 63

is no relation between the taste bud and the ectoderm. At B is a
maxillary tooth and a taste bud just behind it. At C is a taste
bud which stands just behind the mandibular teeth. Note the yolk
granules in both of these, although the yolk is beginning to be
absorbed from the entoderm at this time. These are the most
anterior taste buds found in Amblystoma and none are found on the

Fic. 21. A. punctatum, buds and neuromast. A, taste bud near the yvomer-
ine teeth in a stage later than Fig. 20. B, taste bud just behind the maxillary
teeth. C, taste bud behind the mandibular teeth. D, a neuromast from the
snout. Iron hzmatoxylin, fuchsin.

outer surface of the head. At D in the figure is shown one of the
lateral line organs from the dorsal surface of the snout. Note the
pear-shaped sense cells reaching only half the depth of the epidermis.

By the stage represented by this figure taste buds have made their
appearance in all regions in which they are found in the adult. Up
to this time all the territory in which taste buds appear is lined with
entoderm only and the cells of the taste buds themselves are simply
re-arranged entoderm cells containing yolk granules. All these points

64 J. B. Johnston.

are demonstrated with the greatest ease. ‘The teeth are formed from
ectoderm which migrates in from the borders of the mouth. The
tooth germs and the teeth when well formed lie beneath the ento-
dermal lining of the mouth. The teeth pierce the entoderm at about
the time that the taste buds are becoming clearly recognizable. Up
to this time the entoderm forms a perfectly continuous lining of the
whole mouth. It is broken now only by the points of the teeth piere-
ing it. Up to the stage described no ectoderm cells have gained a
place in the lining membrane in any part of the mouth. The taste
buds are well formed by this stage, their formation from entoderm
cells is directly observed in Amblystoma, and they have no relation
to ectoderm.

Whether in later stages ectodermal cells of the dental ridges rise .
to the surface around the teeth and form the covering of the gums
or of a larger surface in the mouth I have not yet studied. It may be
difficult to determine this point, as the entoderm in the mouth loses
its yolk rapidly in the few days following the last stage described.
The innervation of the mouth suggests the probability that some part
of it is lined by ectoderm. The anterior part of the roof of the mouth,
in front of the vomers, receives cutaneous components from the
ophthalmicus profundus along with visceral sensory components from
the n. palatinus VII (Coghill, 1902). It seems probable that the
taste buds are innervated by the fibers of the palatine nerve, as they
are always innervated by branches of the communis VII. Whether
the trigeminal fibers invade purely entodermal territory or whether
ectodermal cells insinuate themselves into the lining of the mouth
in this region is unknown. This question, however, has no relation
to the question of the origin of taste buds since it is certain that no
ectoderm enters into the lining of the mouth until long after the taste
buds are formed, if at all. The fact that the trigeminus helps to
innervate the mouth, especially in mammals and man, has given
tacit support to the idea that the taste buds are of ectodermal origin.
The reasoning involved in this assumption is illogical, however, since
the taste buds are themselves innervated by the facial nerve which is
strictly a nerve related to entodermal surfaces in all vertebrates
except those fishes in which taste buds spread into the outer skin.

The Limit Between Ectoderm and Entoderm. 65

The presumption, from the standpoint of nerve distribution, is all in
favor of the origin of taste buds from entoderm.

Summary oF Resvutts.

1. In Amblystoma punctatum the relations of preoral entoderm,
premandibular somites, hypophysis and neural tube are essentially
the same as in selachians. There are strong indications that the
preoral entoderm and hypophysis together constitute the vestige of a
palzeostoma.

2. The entoderm surrounding the anterior part of the archenteron
is compressed and the mouth remains closed until a relatively late
period. The location of the palzeostoma and of the neostoma can be
traced continuously to the time when the mouth opening is formed.

3. The formation of the mouth opening includes three phases:
(1) the disappearance of the ectoderm over the mouth area; (2) the
relatively long period of solid entoderm; (3) the progressive forma-
tion of the oral cavity from behind forward as a cleft in the ento-
derm.

4. The deep layer of ectoderm gathers.into a thickening or collar
around the mouth plate and this grows in around the solid entoderm,
thus forming the dental ridges. From these are formed the maxillary,
mandibular and vomerine teeth. The ectoderm cells and teeth are
separated from the mouth cavity by a layer of entoderm. When the
teeth are fully formed they pierce this entoderm to enter the mouth.

5. The taste buds are all formed in entoderm. The entoderm lines
the whole mouth cavity to the very lips until some days after the
taste buds are well formed. The taste buds in all parts of the mouth
and pharynx are formed by re-arrangement of yolk-laden entoderm
cells. There is no evidence that any taste buds in Amblystoma
punctatum are of ectodermal origin. The relations of entoderm and
ectoderm are such that it is impossible that any of them should
arise from ectoderm in this animal.

66 J. B. Johnston.

ABBREVIATIONS USED IN ALL THE FIGURES.
arch., archenteron.
ch.op., chiasma opticum.
c.s., corpus striatum.
d., dental ridge.
d.max., portion of dental ridge forming maxillary teeth.
d.vom., portion of dental ridge forming vomerine teeth.
en., entoderm.
ep., epiphysis.
h., hyoid arch.
hy., hypophysis.
m., mouth plate.
mand., mandibular arch.
max., upper jaw.
n., neural plate or neural tube.
nch., notochord.
o.ph., oro-pharyngeal cavity.
p., preoral or palseostomal recess in the archenteron.
pr.en., preoral entoderm.
prm., premandibular somite.
r.m., recessus mammillaris.
r.p., recessus preeopticus.
r.po., recessus postopticus.
t., terminal ridge or chiasma ridge.
t.b., taste bud.
y.tr., velum transversum.

CoGHILL
1901.

1902.

JOHNSTON
1898.

1905.

1909.

LANDACRE
1907.

STRONG
1898.

The Limit Between Ectoderm and Entoderm. 67
LIST OF PAPERS CITED.

The Rami of the Fifth Nerve in Amphibia. Journal of Compara-
tive Neurology and Psychology, Vol. 11.

The Cranial Nerves of Amblystoma tigrinum. Journal of Com-
parative Neurology and Psychology, Vol. 12.

Hind Brain and Cranial Nerves of Acipenser. Anatomischer
Anzeiger, Vol. 14.

The Cranial Nerve Components of Petromyzon. Morphologisches
Jahrbuch, Vol. 34.

The Morphology of the Forebrain Vesicle in Vertebrates. Journal
of Comparative Neurology and Psychology, Vol. 19.

On the Place of Origin and Method of Distribution of Taste Buds
in Ameiurus melas. Journal of Comparative Neurology and
Psychology, Vol. 17.

Review of Johnston on the Cranial Nerves of the Sturgeon. Jouwr-
nal of Comparative Neurology and Psychology, Vol. 8.

THE SKULL OF LABIDOSAURUS.
BY

S. W. WILLISTON,
University of Chicago.

WITH 3 PLATES.

There are not a few problems in the morphology of the reptilian
skull which yet await satisfactory solutions. Both paleontology
and embryology have failed hitherto to determine indisputably the
homologies of the bones of the temporal region, scarcely any two
authors being in accord in the use of the terms applied to some of
them. Not only is there yet doubt as to the real structure of certain
parts of the skull, but the relationships of many groups, dependent
as they are so intimately upon cranial homologies, are in dispute.
It is evident that the final solution of these problems, problems
which resolve themselves at last into questions of relationships, must
come from paleontology, and especially from the study of the more
primitive and generalized land vertebrates.

Of the paleozoic reptiles the structure of the skull is perhaps best
known in the Pelycosauria, thanks chiefly to the studies of Case in
recent years. A recent review of the material in this group of rep-
tiles preserved in the University of Chicago museum, among the best
now known, that which served Case for the most part in his studies,
has convinced me of the general trustworthiness of his results, though,
as will be seen, I am not fully satisfied as to the legitimacy of some
of his conclusions. That the Pelycosauria are unrelated genetically
with the archosaurian reptiles | am now convinced; that they did
arise from that branch of reptiles usually called Cotylosauria in the
narrow sense, I am also convinced. The resemblances of the skeletal
characters between the Cotylosauria and the Pelycosauria have long
been evident; the modifications of the pectoral and pelvic girdles in

THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

70 S. W. Williston.

either group are of comparatively trivial importance; the remarkable
elongation of the vertebral spines in the more specialized pelycosaurs
is of scarcely greater moment than are similar modifications among
some existing lizards. ‘The presence of the cleithrum, the old-fash-
ioned character of the girdles in general, and the primitive struc-
ture ot the feet have been recognized in the pelycosaurs, but it has
been supposed that radical differences existed in the skull, differences
sufficient to separate the group not only ordinally but into a distinct
sub-class. My present studies of the skull of Labidosaurus, one of
the most primitive of the reptiles known from the Permian, prove,
satisfactorily to myself at least, not only that there are close morpho-
logical resemblances between the skeletons in these two groups of rep-
tiles, but that these resemblances extend to the skull as well; that we
have in Labidosaurus and its allies, the Pariotichide, a persistence
of those generalized characters which gave origin to the peculiar
specializations of the pelycosaurs; that, in other words, the pelyco-
saurs are simply an immediate branch from the primitive cotylosaurs,
having no direct affinities with the Rhynchocephalia; that their rela-
tionships are, probably, far closer with the ichthyosaurs and the
proganosaurs. I am aware that McGregor, in his latest studies of
the Proganosauria,’ still adheres to the belief that the group is closely
allied to the rhynchocephalian stem, a member not only of the “Diap-

sida” but also of the ‘‘Diaptosauria.”

His belief necessarily assumes
the presence of two temporal vacuities in Stereosternum and Meso-
saurus, Which has not been proven and of which I am very skeptical.
Moodie recently has ventured the suggestion that the reptiles have
arisen exclusively from the Microsauria and not from the Temno-
spondyli, but, until he finds better arguments to sustain his conten-
tion than he has so far furnished, I doubt whether his opinions will
obtain acceptance. To evolve five-toed reptiles from four-toed
amphibians; to develop a vestigial cleithrum in such forms as
Dimetrodon, Diadectes, Pareiasaurus, Dicynodon, ete., from forms
which did not possess it; to originate a vestigial intercentrum in rep-

1Commissao de Estudos das Minas de Carvao de Pedra do Brazil, II, p.
301, 1908.
“Geological Magazine, vol. vi, 1909, p. 216.

The Skull of Labidosaurus. 7a

tiles as an element distinct from the hypocentrum, would seem to dem-
onstrate the fallacy of any arguments in favor of the exclusive origin
of reptiles from the known microsaurs. The only legitimate conclu-
sion one may reach is that we have generally erred in the assumption
that reptiles arose in Permian times from known types of amphibians.
I believe that we have yet to discover the ancestral amphibians,
which were neither microsaurs nor temnospondyles in the present
sense, at least as far back as early Pennsylvanian times, and I also
believe that these ancestors had not only five toes in front, as have
the Eryopidee, with at least the modern reptilian phalangeal formula,
but that they also possessed cleithra, and rhachitomous vertebre.

Of the Cotylosauria the form hitherto best known is, perhaps,
Labidosaurus, a restoration of which has been published by Broili,?
with a partial description of the grosser characters of the skull. The
material of this genus in the collections of Chicago University, for
the most part obtained by the expedition of 1908,* is probably the
best yet secured. It consists of two excellent skulls, of typical size,
three nearly complete skulls of smaller size, and portions of four
of five others. Among these smaller skulls is the one described
by myself as Pariotichus (Labidosaurus) incisivus Cope.? That
the specific determination is wrong is very evident. Cope long ago
figured the duplicate rows of maxillary teeth in his Pariotichus
incisivus,’ whereas this specimen, like the typical labidosaurs, has
but a single row, the essential character of the genus, a character
indeed which induced Cope to separate the genus from the true
Cotylosauria and refer it to the Pareiasauria. That Labidosaurus is
closely related to Pariotichus is, I believe, assured, and its separation
by more than family rank therefore improper. That all of these
skulls belong in the same species as do the larger ones, L. hamatus,
is improbable, but their specific determination at the present time is

*Paleontographica, li, p. 65, 1904.
‘Several additional skulls and skeletons were obtained by the expedition
of 1909.

*"Case, Zoological Bulletin, ii, p. 231, 1899; Williston, Journal of Geology,
Xvi, 139, 1908.
*Trans. Amer. Phil. Soc., 1886, p. 290, pl. ii, ff. 4, 5.

~I
bo

_S. W. Williston.

of but little moment as having no bearing upon questions of morpho-
logical interest.

The two larger skulls, very clearly belonging to the typical species
L. hamatus, those which have furnished most of the information given
in the present paper, are somewhat obliquely compressed, as is often
the case with the Permian fossils of Texas; one of them toward the
right, the other toward the left. They are of precisely the same size,
and the correction of their distortions in the figures is a matter of
little difficulty (Plate I and Plate IL). The skull, it is seen, is remark-
able for its attentuated facial region, and for the beak-like extension
of the premaxillaries, terminating in the long, rake-like teeth. The
nares, situated nearly at the extremity of the rostrum, are semioval
in shape, directed outward. The face in front of the orbits is nar-
row, gently convex from side to side, with nearly vertical sides and
a gentle longitudinal convexity in the middle. The orbits are a
little longer than wide, their diameter a trifle greater than the inter-
orbital width. Posteriorly the skull is flattened in the middle above,
and greatly expanded in width, the expansion beginning near the
back part of the orbits, the lateral margins curving inward at the
extreme posterior part. The large pineal foramen is situated near
the front part of the parietal bone, about midway between a line
drawn through the hind margins of the orbits and the hind margin
of the skull in the middle line. There is a pronounced emargination
of the hind margin of the skull, extending the width of the parietal
bones. In well-preserved specimens the markings of the surface of
the skull are very distinct, consisting, for the most part, of round or
oval pits forming a reticulation, but not distinctly arranged in rows.
In other specimens these pits are less conspicuous, and the surface
in some appears almost smooth.

The relations of the bones of the upper surface of the skull have
been figured by Case and myself in the papers cited, but I am con-
vineed, from a careful study of the material at my command, that
we were both more or less in error in the supposed recognition of
elements which we took for granted must have been present. The
premaxillee are separated in several of the specimens in the museum.
They unite broadly above with the nasals, by a rounded border in

The Skull of Labidosaurus. Gas.

front of the middle of the nareal margins; and on the sides with the
maxille, below the nares. The two bones together present a strong
anterior convexity, with the alveolar border receding. Each has
three, elongated, pointed, slightly recurved teeth, of which the inner-
most 1s the largest, the outermost the smallest, less than half the length
of the longest. In the closed mouth these teeth, or the inner ones,
protrude quite a distance below the mandible, hook-like or rake-like, as
shown in Plate II, Fig. 2. This extraordinary development of these
teeth and their position, in association with the narrow, compressed
facial rostrum, remind one strongly of the phytosaurs and are sug-
gestive of like habits in the hving creatures, the exhumation of bur-
rowing invertebrates from the mud or sand of the shores or shallow
water. The maxille, free in one of our specimens, are rather slender
bones, with their greatest width a httle in advance of the orbits.
They extend back, decreasing in width, to nearly opposite the pos-
terior part of the orbits, uniting above in front with the elongated
lachrymals, behind with the anterior prolongation of the jugals. I
count in different specimens seventeen teeth, not very different in
size, the longest a little in front of the middle of the series, and the
series separated from the outermost of the premaxillary teeth by a
short diastema. The nasals form the upper side of the rostrum as
far as their union with the frontals, a little in advance of the orbits,
curving a little downward on the sides back of the nares, whose upper
borders only, do they form. The prefrontals are subtriangular in
shape and small; their inner sutures begin a little beyond the middle
of the upper orbital margin and are parallel with each other, extend-
ing a little beyond the end of the frontal bones. The lachrymals are
large bones, united broadly with the nasals anterior to the prefrontals,
and with more than half the length of the maxille below. They
form the posterior boundary of the nares and the larger part of the
anterior border of the orbits. The precise boundary between the
nasals and lachrymals may be somewhat indefinite; the sutural line
given is that in which four skulls seem to agree. The frontal bones
have nearly parallel sides, extending posteriorly a little beyond the
hind margins of the orbits, joining the parietals in a transverse ser-
rate suture, which appears on the under side somewhat in advance of

74 S. W. Williston.

the line above. The frontals form but a small part of the upper
orbital margin. The postfrontals are also small, forming the posterior
upper margin of the orbit, and leaving but a small space of frontal
margin between them and the prefrontals. The postorbitals are larger
than the postfrontals, and also extend a little further back of the
frontal suture. They form most of the hind border of the orbits,
articulating with the squamosal behind and the jugal below. The
jugals begin a httle in front of the middle of the orbit im an acute
point between the lachrymals and the maxilla. They are broader
just behind the orbit, where they articulate with the postorbitals
above and the squamosal behind. Below the latter they extend as a
rather narrow prolongation to or nearly to the hind angle of the
skull, and to the outer extremity of the ‘‘epiotic’” bones. In the
skull figured by Case and myself these posterior prolongations appear
to be suturally separated from the broader part of the jugals in
advance. <A careful examination of other specimens, however, reveals
no suture here and leads me to the belief that the supposed suture is
merely a fracture in the same place on each side, due doubtless to
the fact of the subangular narrowing of the jugal at this place.
If there be a distinct bone here I suppose that it must be the real
quadratojugal, notwithstanding it has no articulation with the quad-
rate. All the sutures I have so far described, save per-
haps that between the nasal and lachrymal, and that between
the postorbital and jugal, are decisively and clearly indicated in the
different specimens, some of them conspicuously so, and they, more-
over, agree in the different specimens, as long and patient examina-
tions and careful measurements testify. Cope’s determinations of
the cranial elements in Pariotichus and both Case’s and my own
in the small skull of Labidosaurus recognize another suture dividing
the so-called squamosal into two distinct elements, though we do not
agree in the position of this suture. In the Labidosaurus skull fig-
ured by myself there does appear to be a divisional line, indistinctly
shown and agreeing on the two sides pretty well. Unfortunately in
a half dozen other specimens showing this part of the cranial wall,
some of them in the most perfect condition both above and below,
I can find no trace of a divisional suture, even under the most careful

The Skull of Labidosaurus.

“I
or

examination with a lens. I am satisfied that there is none, that there
is but a single bone here and not two, and this conclusion was
reached before I perceived its significance in comparison with the
skull of Dimetrodon. ‘This large, flat and thin, or gently convex
bone unites on its inner side with the parietal, on the front side with
the postfrontal and postorbital, and on the lower or outer side by a
very squamous and loose suture with the posterior prolongation of
the jugal. This is precisely the arrangement of these bones in
Dimetrodon, and I am satisfied that the elements are morphologically
‘dentical. The chief difference between Labidosaurus and Dumet-
rodon consists in the rather large vacuity of the latter piercing what
otherwise would be the squamosal, jugal and postorbital bones. For
the present I accept Case’s determination of the squamosal element
as the prosquamosal, but I feel far less assured of its homology than
I did formerly, though I doubt not that it corresponds quite with
the element in the ichthyosaurs originally named prosquamosal by
Owen.

On the posterior or occipital side there are two cranial roof bones
on each side, clearly and positively shown in all our specimens, one
bordering the hind margin of the parietal, the other the squamosal,
and called by Cope respectively the supraoecipital and the tabulare,
that is the so-called epiotic of authors. They differ from the bones of
the upper surface of the skull in lacking the superficial markings
or pittings, and are suturally united with the superior bones at an
angle of nearly ninety degrees. The superior or inner of these
two pairs of bones, those bordering the parietals, the supraoccipitals
of Cope, are the narrower of the two. Their inner ends are curved
downward slightly, with an angular interval between them into which
fitted the small spine of the real supraoccipital described further on.
It has long been believed that the so-called supraoccipital of the
Stegocephala and of those reptiles in which a like bone is believed to
occur does not correspond to the true supraoccipital of the higher
reptiles and mammals. They are clearly membrane bones, and have
been called the postparietals by Broom. That they are not the real
supraoccipitals is very evident in this specimen in which a large
and well defined supraoccipital is found quite dissociated from the

76 S. W. Williston.

membrane bones of the cranial wall. These bones unite at their outer
end with the upper part of the so-called epiotic; the lower, thin and
somewhat concave border, is free. The epiotics are broader and
longer than this “postparietal,” with nearly parallel sides, the lower
margin thinned and free and concave in outline, the upper uniting
by suture with the squamosal at the angle of the skull. The inner
end, which is truncate, unites above with the “‘postparietal ;” below
it presents an oblique articular facet for union with the extremity of
the paroccipital. The outer extremity is rounded below, and extends
to the angle of the skull, articulating apparently with the posterior
end. of the jugal. On its inner surface near the roof it articulates
for a large part of its extent with the hind border of the quadrate.
Further observations on the homologies of this remarkable bone
will be given later.

The more complete of the two larger skulls has the palatal and
basicranial regions in excellent preservation, and but little distorted.
Just back of the transverse bones a recent fracture through the nar-
rowest parts of the free pterygoids has permitted the removal of the
posterior portion and its complete separation from the encrusting
matrix, both above and below, enabling one for almost the first time
to obtain a clear conception of the cranial bones and their relations to
each other. Very remarkable is the fact that all this portion has no
sutural connection with the cranial walls, the suture between the
quadrate and the epiotic being the only one, indirectly connecting
the vertebral elements with the superior membrane bones. This will
readily account for the fact, so often observed, of the loss of the
basioccipital and basisphenoid from the remainder of the skull, a
loss which, erroneously interpreted, induced Cope to give the name
Cotylosauria to the whole group. Above, in the middle, the “post-
parietals” merely touch the supraoecipital while the epipterygoids
further in advance touch the parietals in a mere rounded point.

The quadrate bone of the left side in this specimen lacks its
articular head, which had been, unfortunately, broken off with a part
of the articular and lost before the discovery of the specimen. The
remainder of the quadrate, however, is quite in position, overlapping
the pterygoids, and is complete. On the right side the quadrate,

The Skull of Labidosaurus. Me,

nearly complete, has been entirely separated from its articular rela-
tions. The vomers, anterior part of the pterygoids, the palatines and
the transverse bones are all in their normal positions. The nares,
situated far in front, probably directly below the external orifices,
are concealed by the mandibles, which are closed upon the maxillee ;
nor is the suture distinguishing the vomers from the posterior ele-
ments distinguishable. The narrow pterygopalatine shelf on each
side shows, on the upper side at least, a suture between the palatines
and pterygoids for a portion of the distance, though I can make out
no suture separating the transverse bones, though such doubtless
existed. The transverse bones are stout, forming a strong decliyity
from the plane of the palatines, and they abut massively against the
mandibles at least as far as their middle. In the middle, between
the pterygoids, opposite and in front of the transverse bones there
is a large ovate interpterygoidal space, in front of which the two
pterygoids approach each other closely, though not touching. Pos-
sibly in the living skull they actually met in the middle. In front of
the basisphenoid the pterygoids curve inward so that they meet in
the middle behind, leaving no space for a presphenoid or para-
sphenoid, which is certainly wanting in this specimen at least, though
distinctly present in a smaller skull, and recognized by Broili in
this species, The pterygoids unite firmly with the basisphenoid by
this inner sphenoid process. Along the margin of the interptery-
goidal opening for nearly its whole extent, there is a row, possibly
double in front, of small tubercular teeth; a patch of similar teeth
is also present in front of the transverse declivity of the palatines,
and yet another patch on the summit of each transverse bone.

The posterior prolongations or quadrate processes of the ptery-
goids, arising just back of the transverse bones from the base of the
stout sphenoid processes, are long, thin, divergent, oblique plates of
bone, extending back nearly to the hind margin of the skull, articulat-
ing broadly but loosely with the plate of the quadrate as shown by the
dotted lines in Plate III, Fig. 1. The inner border nearly touches
the sides of the basisphenoid; the lower, thin and nearly straight
border is continued to near the articular extremity of the quadrate.
The basisphenoid is narrow in front, gradually widened behind,

78 S. W. Williston.

erooved in the middle, shallowly in front, more deeply behind, where
it is bordered on each side by a descending process which terminates
in a free thin margin underhanging a fossa that opens backward. I
cannot distinguish with certainty the sutural division between the
basisphenoid and basioccipital, though it seems to be wholly back of
the lateral processes, since in another skull, in which the basioccipital
has been dislodged, the division has been made back of these processes.
On either side of the basiphenoid, or the conjoined basiphenoid and
basioccipital, an elongate, cylindrical or oval rod is given off, which
is directed downward, outward and backward, lying closely under the
posterior end of the pterygoid plate, and reaching nearly or quite to
the head of the quadrate. This process, clearly the stapes, seems to
be suturally united with the basiphenoid, as indicated in Fig. 1,
Plate II. The position of the bone in the specimen seems to be
quite normal and undistorted, and the bone is nearly complete,
though possibly the extreme end has been broken away; it seems to
be perforated proximally by a small foramen. The shape, form and
relations of the basisphenoid, stapes and pterygoids may be compared
with the author’s figure of the same parts in the remarkable rhachito-
mous amphibian recently described by myself.’ The basioccipital
bone, limited as I believe it to be in front, is small and is clearly
distinguishable from the exoccipitals. Its condyle is convex, oval
from side to side, somewhat pitted in its middle, and seems to be
wholly composed of the basioccipital. The exoccipitals are small,
apparently taking no part in the condyle. The suture limiting them
from the basioccipital is clearly seen at the sides below and joining
the margin of the foramen immediately at the side of the condylar
surface above. The suture separating it from the paroccipital passes
through the jugular foramen, thence directly upward and forward.
The exoccipitals join the supraoccipital by a transverse suture a
little below the summit of the foramen magnum. The foramen
magnum is heart-shaped, about eight millimeters in its greater diam-
eter. The paroccipitals or opisthotics are distinct elements, the dis-
tinguishing suture very clearly indicated, as already stated. They
are stout at their base, and are turned outward and backward to end

"Trematops milleri, Journal of Geology, vol. xvii, p. 686, 1909.

The Skull of Labidosaurus. 79

in a short clyindrical rod lying under the proximal posterior end of
the quadrate and articulating at the extremity with the facet already
described on the lower part of the inner end of the ‘‘epiotic.”
This articular arrangement is the normal one of the opisthotie with
the epiotic in the Stegocephala. Chiefly because of this fact I am
loath to identify the bone with the quadratojugal, to say nothing of
the anomalous position of the bone for a quadratojugal. Anteriorly
the suture separating the paroccipital from the supraoccipital passes
nearly directly forward to the outer side of the posterior lateral pro-
jections of the supraoccipitals where the dividing suture turns
inward. Of the suture separating the prodtics I am less certain,
though it seems to be quite_apparent in the position I have figured
it in the drawing. The supraoccipital is a large element, when seen
from above having a marvelous resemblance to the arch of a dorsal
vertebra. A small dorsal spine in the middle posteriorly is inter-
ealated in the angle between the inner ends of the postparietals, but
there is no sutural attachment. Anteriorly the two sides of the supra-
occipital diverge in the form of zygapophyses, with an emargination
between them exposing the cerebral cavity. From the median pos-
terior spine a ridge runs outwards to each ‘‘zygapophysis.” In
front of each lateral projection, the prodtic, distinguished suturally,
descends in a rounded margin to form the optic notch. In front
of these optic notches there is, on either side, a thin, vertical plate,
attached either to the protic or basisphenoid below the meeting in the
middle above, leaving an opening of rather small size between them.
The upper end of these plates is fractured, but it is very evident that,
in position, relations, and shape they agree quite with similar bones
bounding the cerebral cavity in most lizards, a small bone, usually
lost in the macerated skull, whose homology is not well understood.
Since their position is in front of the optic nerve it would seem to
preclude the possibility of their being alisphenoids. These elements
in the mosasaurs I have identified as orbitosphenoids (see University
of Kansas Geological Survey, Vol. IV, Pl. xxix, f. 5; Kans. Univ.
Quarterly, Vol. XT, p. 249), but neither identification is quite satis-
factory. The upper oblique surface of the pterygoid wings is
concealed posteriorly by the quadrates. In front of the quadrates,

80 S. W. Williston.

and occupying nearly the whole extent of their margin and the
upper half of their externo-superior surface, are the elongated and
oval epipterygoids. They continue the aclivity of the pterygoid
wings on the outer side a little more steeply, ending in an obtuse
point a little back of the orbitosphenoid plates, which touches, but
is not suturally united with, the parietals above. These epiptery-
goids are broader in front, where they come in contact with each
other over the pterygoids. The quadrates in the larger specimens,
and also in one of the smaller, are preserved nearly or quite intact, and
in their natural relations. They unite with four bones only, the ptery-
goids by a very broad and loose union, as shown in Fig. 1, Plate IIT;
with the outer ends of the paroccipitals, as also shown by the dotted
lines in the same figure and in Plate IJ, Fig. 1, and by their
posterior everted articular margin with the outer extremity of the
postparietals and, much more extensively, with the “epiotics” near
the cranial wall. The thin, expanded proximal plate of the quadrate,
as shown in Fig. 1, Plate III, narrows into a distinct neck, chiefly
by a groove which winds from the under side about the hind margin
a little above the articular extremity. The notch thus formed is
clearly the auditory notch, corresponding to the notch or foramen in
the quadrate of the mosasaurs and lizards; and possibly also it
corresponds with the so-called quadrate foramen of the Pelycosauria.
Doubtless the Cotylosauria had a small external ear situated nearly
as it is in the lizards, above the angle of the mandible. The artic-
ular surface of the quadrate for the mandible is oblique to the plane
of the bone, so as to look more nearly downward in the normal
position of the quadrate. Its outer side projects into a rather narrow
process, but does not touch, much less articulate with, the roof bones.

The mandibles, in comparison with the skull, are stout and heavy
bones, and amply attest the predaceous habits of the animals. They
are slightly expanded in front, where they meet in short symphysis,
heaviest and stoutest just back of the orbits and broadest also here.
They are nearly straight or gently incurved anteriorly, turned in-
wards in a broad curve behind. The splenial bones unite in a
median symphysis in front and extend back nearly to the articular,
leaving a broad, elongate open cavity on the inner side from imme-

The Skull of Labidosaurus. 81

diately back of the orbits. They also form a part of the lower
margin of the mandible, visible from below as far back as the middle
of the orbits, having between them and the hinder end of the dentary,
an elongate and acute projection of the angular. The suture between
the angular and the articular continues the curve of the inner border
of the mandible to the outer side of the extreme posterior end of the
angular process. The suture between the angular and the surangular
passes forward nearly midway of the mandible, and nearly parallel
with the upper border in the closed jaws, to the hind end of the
dentary, that is to nearly opposite the posterior end of the orbits.
The thin ascending plate of the surangular reaches at the summit
nearly as high as the lower margin of the orbits on the inner side
of the temporal roof. Over the summit the slender posterior end
of the coronoid is visible in the closed jaws, but its anterior part is
concealed by the transverse bones. The articular is a short bone,
turned inward, with a thin inner margin. It is apparently continued
forward as a slender prolongation above the margin of the splenial
or angular and forming the lower border of the mandibular cavity,
to a slender, acute point nearly as far as the hind end of the mandibu-
lar tooth series. Whether or not it is separated from the articular
as a distinct bone, the prearticular, or indeed of its precise relations
I will not be sure. Sixteen teeth I count in the mandibular series
in three different skulls. They resemble the maxillary teeth, but are
somewhat smaller. The first or second is distinctly larger than the
following ones. :

A minute comparison of the skull of Labidosaurus with that of
Procolophon or Pareiasaurus would be most interesting and instruc-
tive, but since this is impossible for me to make I trust that the
figures and descriptions herewith given may furnish the basis for
such a comparison by others. With the exception of the form
described by Broili (Seymouria), I believe that we have no positive
assurance of the simultaneous possession of all four roof-bones,
supraoccipital, epiotic, squamosal and prosquamosal in any coty-
losaurian, using the term in its broadest sense. That all these four
elements should occur concurrently in the primitive reptiles is of
course to be expected, and Broili’s genus seems to furnish an example.

82 S. W. Williston.

Two of these bones have been lost in all modern reptiles, and, with
the exception of the Squamata, three of them. We have generally
assumed that the supraoccipital and epiotic of the stegocephs are
these two, but I question very much, in the light of the evidence fur-
nished by Labidosaurus, whether this assumption is justified. I
have long believed that the small element intercalated between the
paroceipital and the so-called squamosal of the lizards corresponds
to the epioties of the stegocephs, and I am further confirmed in this
opinion by the position and relations of the bones in Labidosaurus.
However, my views as to the homologies of the cranial arches have
been not a little shaken by the structure of the labidosaurian and
pelycosaurian skulls, and I shall not venture here to offer any very
decided opinions, preferring to wait until further evidence is avail-
able, especially that to be yet furnished by the skull of the Diadec-
tide.

In the remarkable genus described by Cope and Case as Hdapho-
saurus a pair of bones almost certainly homologous with those here
provisionally called the “postparictals” are identified by Case as the
epiotics, but with a query.8 In the only known specimen of the
genus, the type, these bones and the parietals were crushed down
over the supraoccipital nearly to the foramen magnum, and this, I
believe, was their normal position. In the restoration given by Case,
however, they have been elevated nearly to the top of the supraoc-
cipital, depressing the paroccipitals so far that their outer extremity
ends near the articular end of the quadrate. In all probability it
articulated, as in Labidosawrus, with the outer end of the so-called
epiotics, or the inner end of the “quadrato-jugals,” as in Labido-
saurus. The relations of all these bones, the “epioties,” “quadrato-
jugals,” quadrate and paroccipitals appear to be quite the same in
Dimetrodon, Edaphosaurus, and Labidosaurus, and I doubt not that
they are all homologically identical. In Hdaphosaurus, however,
another bone is recognized between the parietal and prosquamosal,
clearly corresponding with the additional element supposed to occur
in Labidosaurus, but which I cannot find. Should it be a distinct
element in either or both these genera it does not in the least cast

®Pelycosauria, p. 151.

The Skull of Labidosaurus. S38

doubt upon the homologies of the two pairs of bones on the posterior
border, the squamosal and quadrato-jugal of Dimetrodon, the epiotics
and quadrato-jugal of Hdaphosaurus, the postparietals and epiotics
of Labidosaurus. EHdaphosaurus has a temporal vacuity situated
much higher than in Dimetrodon, by the sides of the parietal bones.
Case believes that there is also another vacuity below, separated by
a bar from the upper one. No certain evidence of such a vacuity
is yet forthcoming, and I shall doubt its presence until demonstrated.
I cannot believe that the presence of vacuities in the temporal region
of these early reptiles is of the great taxonomic importance that
has been ascribed to them.

Perhaps nothing is more noticeable in the skull of the present
reptile than the small comparative size of the brain cavity. While
the skull measures over seven inches in length and five in width, the
foramen magnum is of almost precisely the same size and shape as that
of a small Amblyrhynchus lizard whose skull measures but sixty
millimeters in length. Not only is the foramen of the same size,
but the brain cavity also is only a trifle larger in Labidosaurus.
Small brain capacity is of course to be expected in this old reptile,
and, moreover, the size of the brain cavity as compared with that
of the skull, may not be a fair criterion of the relative intelligence
of the two animals. Nevertheless, that their intelligence was rela-
tively much lower than that of the existing lizards cannot be doubted.

As to the habits of Labidosawrus and its allies, the pariotichids,
one can hardly doubt that the most of them at least were shore
dwellers. The form described by me as Pariotichus laticeps,° is
essentially identical in skeletal and skull structure with Labido-
saurus, so far as can be determined. Indeed it is not impossible
that the form may be a real Labidosaurus of a small species. Many
of the species of Pariotichus, like Labidosaurus hamatus, had the
peculiarly elongated premaxillary teeth, though perhaps never so
exaggerated as in the present species. The only use that I can con-
ceive for such teeth was the seizure of small creatures from burrows,
holes or crevices, or for detaching such as may have been closely
adherent to rocks. The maxillary and mandibular teeth were not
at all pointed, but were, rather more sectorial than prehensile.

"Biological Bulletin, vol. xvii, p. 241, 1909.

84 S. W. Williston.

It is generally assumed that the primitive vertebra from which
those of higher types have been developed was composed of several
distinct and separated bones, the neurocentra, pleurocentra, and
hypocentra, and perhaps others. It must also be assumed that the
differentiation of the three or four anterior vertebre to form the
brain capsule took place before the elimination of those parts now
absent in the higher animals. We should then expect to find in the
vertebrate skull the separate elements or some of them at least, more
or less differentiated in the embryonic condition. Furthermore we
know that, originally, the ribs were continued as vertebral append-
ages as far forward as the skull itself, as is evidenced by the atlantal
ribs still existent in the crocodiles, pelycosaurs, and other reptiles.
And why may we not also assume that the ribs originally continued
on the cranial vertebre 4? The brain capsule of Labidosaurus yet pre-
sents so much of its original vertebral condition that it has not
become attached to the membrane bones of the cranial walls, and its
occipital segment seems to present all the features of an undiffer-
entiated vertebra, the basioecipital representing the hypocentrum,
the exoccipitals the pleurocentra, the supraoccipital the neurocentra,
while the paroccipitals have all the characters of real ribs. To
carry the homologies to the next vertebra we would find the basi-
sphenoid homologous with the hypocentrum, the prodtics with the
pleurocentra, the alisphenoids or epipterygoids with the neurocentra,
while the stapes occupies the ideal position of the primitive rib. That
the stapes of the reptiles is not identical with that of the mammals
is, I believe, usually conceded; that it did primarily originate as
the rib seems not at all improbable. In this connection it is worth
while to observe that the stapes of the primitive reptiles seem always
of large size, a character also found in both the ichthyosaurs and
plesiosaurs.

a ane ¥ pare =

ae. va
‘

EXPLANATION OF PLATES.

PcAwr ele
Labidosaurus hamatus Cope. Skull, from above; two-thirds natural size.

PrArh ial:

Labidosaurus hamatus Cope. Fig. 1, skull, from below; two-thirds natural
size. A, articular; AN, angular; BS, basisphenoid; HP, epiotic; EX, exoc-
cipital; PP, postparietal; PT, pterygoid; Q, quadrate; ST, stapes.

Labidosaurus hamatus Cope. Fig. 2, skull, from the side; two-thirds nat-
ural size.

PLATE III.

Labidosaurus hamatus Cope. Fig. 1, right quadrate, from below; Fig.
2, the same, from above; Fig. 3, posterior basicranial bones from above;
Fig. 4, skull, from behind. All two-thirds natural size. EP, epiotic; EPT,
epipterygoid ; OC, occipital condyle; PO, paroccipital; PP, postparietal; PR,

proétic; PT, pterygoid; Q, quadrate; SO, supraoccipital.

THE SKULL OF LABIDOSAURUS. PLATE TI.

S. W. WILLISTON.

THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

THE SKULL OF LABIDOSAURUS, PLATE Il.

S. W. WILLISTON.

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THH AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAn., 1910.

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THE SKULL OF LABIDOSAURUS. PLATE III.
S. W. WILLISTON.

THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

A HUMAN EMBRYO WITH SEVEN PAIRS OF SOMITES
MEASURING ABOUT 2 MM, IN LENGTH.

BY

WALTER E. DANDY.
From the Anatomical Laboratory, Johns Hopkins University.

WITH 6 PLATES.

Among the youngest human embryos and one of the youngest in
Professor Mall’s collection, is Embryo No. 391, which he has very
kindly permitted me to reconstruct and describe. In general devel-
opment this embryo is almost identical with, probably a trifle older
than, the Kroemer-Pfannestiel embryo, Klb, which measures 1.8
mm. and has six pairs of somites. It is older than Eternod’s
embryo measuring 1.3 mm., Graf Spee’s Gle 1.54 mm., and Embryo
Frassi 1.17 mm., in the order named. It is younger than the
embryos of Unger and Bulle, of 9 and 14 somites respectively,
Eternod’s 2.1 mm. embryo, and embryos XLIV (Bff), LX VIII (Lg.
2.15 mm.), VI (B. R. 2.2 mm.) and VII (E 2.1 mm.) of His.

This very rare specimen came into Professor Mall’s possession
through the kindness of Dr. R. W. Pearce, of Albany, New York,
with the following history from the physician who handed the speci-
men to him. “The woman had passed her period about two weeks
when she performed an abortion with a stick about 8 inches long,
which she whittled out for the purpose. This she passed into the
uterus and 24 hours later this specimen was aborted. Her purpose
in calling me was to see if her object had been attained. I have
kept the specimen two years in a bottle of weak formaldehyde.”

Upon receipt the following measurements were made by Professor
Mall; ovum 16x14x12 mm., embryo about 2 mm. The specimen
was placed in fresh formalin, stained with alum-cochineal-eosin,
imbedded in paraffin and cut into serial sections 10 microns in
thickness.

From these sections (163 in number, excluding the bauchstiel),
the embryo was reconstructed upon a scale of 200 magnification,

THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

86 Walter E. Dandy.

giving the model a total length of 32.6 em. <A shrinkage space
between the layers of tissue aided materially in the reconstruction
without sacrificing tissue. The model is so constructed that every
detail may be exposed by cuts and windows.

Upon presentation of an embryo the question naturally arises,
is it normal? ‘The history of a mechanical abortion points very
strongly toward a normal embryo. It compares very closely with
the few other known embryos of this period. The tissues, including
the chorion, are excellently preserved and show no degenerative
changes.

AGE.

From the history that the menstrual period passed about two weeks,
the age of 13-14 days would fit very nicely into the conventional
Reichert-His theory of fertilization and age of embryos. Recent obser-
vations however by Mall in 1000 cases with menstrual history have
shown that we underestimate the age of young embryos, on an aver-
age about 10 days, that fertilization is not restricted to the period
immediately before the menstrual period, but may occur at any time
in the intermenstrual period. Bryce and Teacher also came to the
same conclusions and illustrate this in a well constructed diagram
of the menstrual cycle. This obviates the remarkable distortion
which has been necessary to harmonize age with size and progress
of development, and makes the human embryo much less precocious
in its development. In conformity to the above observations the
true age of this embryo is probably about 24° days, which would
make the time of fertilization occur about ten days before the time
for onset of lapsed menstrual period or about eighteen days after

beginning of the last menstrual period.

Tur ADNEXA.

Chorion. The chorionic membrane is about 0.1 mm. thick, and
is covered with many branching villi varying in size up to 1.25
mm. in length and 0.1 mm, in thickness. These villi are more
numerous at the point of attachment of the bauchstiel and gradually
fade away on all sides until finally a clear zone results from their
absence on the opposite pole.

(o'8)
a |

A Human Embryo with Seven Pairs of Somites.

The chorionic membrane and villi have the characteristic inner
loose mesenchymatous layer of beautiful branching spindle and. stel-
late cells with anastomosing processes and a somewhat jelly-lke inter-
cellular substance; an external double epithelial layer, consisting of
the inner Langhans layer of small cells with lightly staining nuclei
and cytoplasm, and the outer syncytial layer which stains more
deeply with eosin and has larger and more densely staining nuclei,
but no definite cell boundaries. From the epithelial layer of the
chorionic membrane and villi, numerous buds develop, some from
the syncytial layer alone, others from both layers of epithelium.
These represent proliferation and new formation of villi. The
mesenchymatous layer of the chorionic membrane contains many
newly forming capillaries, some of which extend into the villi. The
details of these will be considered later in the description of the
vascular system.

The Bauchstiel. The bauchstiel does not differ from that of
other young embryos. It consists of loose mesodermal tissue, lined
externally by a single layer of flattened mesodermal cells. It 1s con-
tinuous distally with the chorionic mesoderm and proximally with
the mesoderm of the umbilical vesicle, amnion and body of the
embryo. It contains the allantois, umbilical arteries and veins and
their resulting sinuses and branches.

Umbilical vesicle-—The yolk sae has no stalk but is attached to
the entire length and breadth of the embryo, with the exception of
small portions of the anterior and posterior ends, which are covered
by the short head and tail folds of amnion. At its attachment to
the embryo, the walls are very thin, consisting of two layers of flat-
tened cells—mesoderm and entoderm. These walls gradually grow
thicker distally, due to the degree of development of the blood
islands, which also cause a great distortion and knotty appearance
of the mesodermal surface. This vascular development extends
throughout the whole length of the umbilical vesicle, but is practi-
cally limited to the ventral or distal half, only a few islands being
seen in the dorsal half. The greatest development seems to be near
the center of the vesicle and is apparently developing to connect
with the future vitelline veins. This is in contradistinction to the
findings in Eternod’s 1.3 mm. embryo, in which the posterior region

88 Walter E. Dandy.

of the umbilical vesicle is drained by the sinus ensiforme. The ento-
dermal lining of the inner surface of the umbilical vesicle forms a dis-
tinet layer in places, whereas in other locations, especially in the
ventral half, no definite layer of cells can be made out, so intimately
is it fused with the knotty and thickened mesoderm.

Amnion.—The amnion is a completely closed cavity, with very thin
delicate walls of flattened cells, the imdividuality of ectoderm and
mesoderm being everywhere beautifully preserved. The cavity of the
amnion is very large, probably due, as suggested by Professor Mall,
to the osmosis of large quantities of dilute formalin in which it was
preserved for over two years. Anteriorly a rather deep pocket of
amnion dips ventral to the heart, evidently a sign of posterior exten-
sion of the head fold. The tail fold is very small, covering the
embryo for a distance of only 5 or 6 sections.

GENERAL APPEARANCE OF THE EMBRYO.

The embryo presents an anterior and posterior elevation with a
marked dorsal link. (Plate IV.) The anterior elevation is gradual,
the posterior very sharp, rising at an angle of about 80°. This kink
seems to be partly natural and partly an exaggerated post-mortem
condition. We should naturally expect a dorsal concavity due to
the greater development of the structures in both the anterior and
posterior regions of the embryo. ‘The marked accentuation how-
ever may be due, as suggested by Professor Mall, to the large
amnion filled with fluid, the weight of which would naturally act
upon the point of least resistance. Shrinkage incident to manipula-
tion and imbedding also plays an important role in its aggravation,
especially after location of the point of least resistance. This is
clearly shown by a comparison of the model with a sketch of the
embryo before imbedding, there being an accentuation of the kink
by almost 15°.

EcropERM.

Nervous system. The extent of development of the nervous sys-
tem is a medullary groove, open throughout its entire length. The
brain region is divided into three primary vesicles (Plate VI), the
anterior being equal in size to the other two combined. The first or

A Human Embryo with Seven Pairs of Somites. 89

anterior vesicle is long, wide and deep with edges everted and pro-
jecting outward over the anterior and lateral walls of the anterior
body elevation. The walls of the second and third vesicles have a
tendency toward inversion and the enclosed vesicles are more sharply
defined laterally than the first. The brain passes insensibly into the
spinal cord, which is much smaller in all diameters, but 1s nowhere
closed. Posteriorly the medullary groove forms a shallow dilatation
eradually fading into the flattened surface of the primitive streak.
(Plate IV.)

No neurenteric canal is visible, though present in the younger
embryos of Frassi, Graf Spee and Eternod and _ possibly the
Kroemer-Pfannenstiel Klb. No traces of spinal or cerebral ganglia
or nerves are visible. No suggestion of anlage of the lens, optic or
otic vesicles could be detected.

Primitive streak. Posteriorly and dorsally in the region of the
termination of the notochord and neural groove, ectoderm and meso-
derm gradually losing their individual morphological characteris-
ties, fuse to form a mass of a single variety of simple undifferenti-
ated cells, which extends to the posterior termination of the embryo.
This is the primitive streak (Fig. 7). The entoderm also seems to
be a part of this mass since its cells are directly continuous with it,
although the characteristic lining of the hindgut is still maintained
throughout. There is no groove except a very shallow flattened
dorsal depression which is gradually lost posteriorly and which 1s
continuous anteriorly with the neural groove. This primitive streak
region suggests a storehouse of simple undifferentiated cells supplying
mesoderm, ectoderm and entoderm in the earlier stages, later becom-
ing differentiated into the characteristic morphology and arrange-
ment of the different layers.

ENTODERM.

Entoderm lines the ventral surface of embryo, fore and hind
gut, allantois and the inner surface of the umbilical vesicle. In
the ventral entoderm of the embryo is a median longitudinal
groove corresponding to the location of the notochord and caused
by its adherence to the ectoderm of the medullary groove. On
either side of this groove and parallel to it is a ridge caused by

90 Walter E. Dandy.

ventral projection of the dorsal aorte. These are no doubt accentu-
ated by post-mortem shrinkage. The opening of the foregut is strad-
dled by two prominent ridges which unite above into a large ventral
bulging, represented by the overlying ccelom, and pericardial cavity
with the enclosed heart, respectively.

The foregut is present in 32 sections, representing a length of 320
microns. It ends blindly anteriorly and is separated by mesoderm from
the bueceal cavity which is forming by an invagination of ectoderm.
There is however no bucco-pharyngeal membrane. One small pharyn-
geal pouch is present on each side below the first and only complete
aortic arch. These pouches are continuous ventrally and unite to
form a ventral pouch. ‘There is no contact with the outer ectoderm,
although the mesoderm separating them is considerably thinner than
elsewhere.

The hindgut is a blind somewhat oval, dilated pouch, 120 microns
in length by sections, but on account of the dorsal kink of the embryo
and the consequent partially longitudinal plane of the sections, the
actual length is somewhat greater. The allantois arises from the
ventral surface of the hindgut. It passes from the embryo into the
bauchstiel in company with and between the umbilical arteries.
After the union of these arteries, it takes its position between the
arterial and venous sinuses (Plate VI) and bends at almost a right
angle to conform to the direction of these sinuses and the bauchstiel.
At this latter point it divides into a short stub measuring about 40
microns and a much longer branch. The terminus of each branch
may be seen in Figs. 8 and 9. The allantois is lined by a single
layer of cubical epithelium; it maintains a lumen to the Bera of
division, from which it consists of a solid core of cells.

No liver or thyroid anlage has made its appearance. No cloacal
or bueco-pharyngeal membranes can be distinguished.

Notochord. With the exception of the very posterior tip which
is entirely free for a distance of three sections, the notochord is
everywhere fused with and apparently an integral part of the ento-
derm. Posteriorly it is very conspicuous as a relatively large and
very compact knob of deeply staining cells on the dorsal wall of the
hindgut. This sharply differentiated mass invaginates. into the
ectoderm, with which is it not here united (Fig. 6). It gradually

A Human Embryo with Seven Pairs of Somites. 91

becomes smaller anteriorly, being represented by a very small thick-
ening of entoderm (Fig. 3) and finally is indistiguishable. Except
in the region of hindgut mentioned above, it, together with the ento-
derm, is everywhere in contact with the neural groove ectoderm, usu-
ally a line of contact being visible; in some places however a line of
demareation is exceedingly difficult to make out. No trace of a
notochordal eanal could be detected.

M&SoDERM.

Mesoderm.—Posterior to the somites is a large mass of very sim-
ple undifferentiated mesoderm which dorsally passes insensibly
into the primitive streak. It is largely responsible for the size and
shape of the posterior region of the embryo. Its only differentiation
is laterally, where it becomes somewhat more compactly arranged,
lining the spaces which are forming the peritoneal celom. Anterior
to the somites the paraxial mesoderm is also very simple and serves
as a wedge of tissue between entoderm and ectoderm. The highest
mesodermal differentiation is seen in the formation of the somites,
coelom and heart which will now be considered.

There are seven pairs of somites completely segmented. Each of
the first six pairs contains a cavity, unconnected laterally with the
eeelom as described by Keibel in the reconstruction of the Kroemer-
Pfannenstiel embryo, Klb. The first pair is apparently the most
highly differentiated, being the largest, having the largest cavity and
most compact peripheral arrangement of cells and is connected later-
ally by the most clearly defined and differentiated intermediate cell
mass. These characteristics are less pronounced in each successive
posterior somite until the last is the smallest, contains no cavity,
has no cellular arrangement, and shows no other signs of differentia-
tion. This would seem to indieate that the cavity is of secondary
origin; Graf Spee however finds a small mesodermal slit in embryo
Gle which he thinks is a myocele, although no somites are present.
In addition to the completely formed somites, another pair is just
beginning to show signs of constriction from the posterior paraxial
mesoderm. Anteriorly there is also another pair of somites about
half segmented with a very uniform compact mass of cells but con-
taining no cavity. This rudimentary pair probably represents the

92 Walter E. Dandy.

first pair of somites, which according to experiments in the chick
by Marion Hubbard and Patterson, is established first in time and
position, but develops slowly and maintains its connection with the
anterior paraxial mesoderm, thus causing the formation of somites
to take place only posteriorly.

Celom. In his article on the eclom for the new Keibel-Mall
Embryology (now in the press), Professor Mall used this embryo
as the second stage of human eccelomic development. Graf Spee’s
Gle 1.54 mm. is the first human embryo to show any sign of eccelom
formation, this being a very small bilateral slit in the mesoderm of
the anterior portion of the embryo—the beginning of the pericardial
colom. This slit is said to communicate externally by a very small
channel. The next stage, represented by this embryo, shows a very
large and well developed single, united pericardial cavity. (Figs.
1 and 11.)

The posterior continuation of the pericardial cavity on each side
(Figs. 11 and 15) probably represents the pleural ccelom, which
passes insensibly into the peritoneal eeelom, now forming from
multiple foci. A glance at Fig. 11 will show the extreme irregularity
and entire absence of any metameric arrangement in the formation
of the peritoneal eclom. Numerous irregular pockets of varying
size dip into the mesoderm from the extraceelom. This communica-
tion with the exterior may be either primary or secondary. The
diagram of the reconstructed ceelom, however, would seem to indi-
cate the former might be the case, because posteriorly a rather long
continuous slit is present in the mesoderm, the pockets of which are
shorter, much wider at the external opening, and diminish in size
internally. There are, however, several small independent cavities,
having no visible connection with each other or with the extraccelom,
which strongly suggest an independent formation by internal meso-
dermal cleavage, beginning in multiple small foci, which may later
connect with the other cavities and thus indirectly through them to the
extracelom. Judging from the structure of this specimen alone it
seems probable that the result may be a combination of both processes
which are in reality ultimately one and the same process differing
only in position. This irregular formation has been described by

A Human Embryo with Seven Pairs of Scmites. 93

Bonnet in young sheep embryos, both in pericardial and peritoneal
coelom, the latter only communicating with the extraccelom.

The pericardial cavity communicates freely with the pleural,
which in turn passes into the peritoneal ccelom and through it com-
munication is made on each side with the exocelom at the level
of somite IV (Fig. 11). ‘There is therefore a complete circuitous
eanal, by which it is possible to travel from the extraccelom on one
side through the embryonic ccelom to the extraccelom on the other
side.

Heart. The mesodermal walls of the heart are the result of
confluence, fusion and absorption of the walls of the pericardial
coelom. The heart is attached dorsally throughout its whole length
by a mesocardium; ventrally and laterally it is free in the peri
eardial cavity. The heart roughly fills about one-third of the
pericardial cavity, is bent upon itself almost at a right angle, with
convexity to right and coneavity to the left side of the embryo. The
endothelial heart is a simple tube of flattened cells lying within the
mesodermal walls but greatly shrunken and collapsed from fixation.

Nephric system. There is no evidence of the Wolffian bodies or
ducts. Three independent small cavities are seen in the lateral
masses of somites I and II on the right side and somite II on the
left side, none of which connect with celom. These cavities show
a partially differentiated wall of more or less distinct cubical epithe-
lium (Fig. 3) and are probably the earliest stages of the pronephric
tubules. Below somite II the lateral masses become very vague and
indistinct. Fig. 11 shows a number of cavities, some independent,
others connecting with the eelom, but agreeing in position with the
future lateral masses. They are merely small undifferentiated slits
in the mesoderm, have no definite lining and may or may not be
homologous with those mentioned above as pronephric tubules and in
earlier stages of development. They may, however, represent foci
or ccelomic development, as is suggested by several small spaces
similar in position and connecting with the ccelom.

Vascular system. The description of the vascular system of this
embryo might appear very unorthodox indeed, were it not for the
previous description by Eternod of a 1.3 mm. human embryo, which
for a decade has remained alone and unconfirmed, which shows, how-

94 Walter E. Dandy.

ever, all the signs of a normal embryo. The question of a primitive
volk circulation has always been regarded as an established and indis-
putable fact, based almost entirely upon comparative embryology.

Recent observations in human embryology, however, have shown
the human ovum to differ markedly from other comparative forms
in the very early stages of development. This embryo, together with
the early embryos which have been described, particularly that of
Eternod, seems to indicate that the primitive human vascular sys-
tem differs from the conventional comparative system in that the
umbilical circulation is the predecessor of the time-honored vitelline
circulation.

In studying the vascular system, every capillary was traced as far
as high magnification would permit. The recent work of H. M. Evans,
with injections of fresh specimens of young chick and pig embryos,
has however demonstrated the existence of large hitherto unrecogniz-
able capillary beds and has shown the comparative inefficiency of
studying fixed uninjected tissues‘for blood-vessels, even under the
most favorable conditions. We must therefore always bear in mind
the possibility of collapsed vessels, and that other things being equal,
too much emphasis cannot be placed upon negative evidence. Even
if there should be some small capillary beds, which are unrecogniz-
able by the microscope in the absence of injections, it would merely
be evidence in favor of vascular connection between the umbilical
vesicle and the embryonic circulation, and would not in any way
affect the question of priority, because of the relatively greater devel-
opment of the umbilical vessels. A glance at Fig. 15 and Fig. 12
will conclusively show that the functioning system is umbilical or
chorionic and that if a connection could be traced through the large
gap between the umbilical vesicle and the small sprout of the possible
vitelline vein, it would be comparatively insignificant. Since the
embryonic vessels are full of blood and there is no apparent connec-
tion with the blood-forming area in the umbilical vesicle, the ques-
tion naturally arises, how are we to account for the presence of the
blood corpuscles? This is explained by finding many beautiful exam-
ples of endothelial proliferation of blood corpuscles from the capil-
laries in the chorionic membrane (Fig. 10), probably supplying the
embryo with blood until the time of connection with the yolk sac.

A Human Embryo with Seven Pairs of Somites. 95

A survey of the literature gives very limited information on the
primitive vascular development. ‘The first suggestion of a vascular
anlage is a very indistinct knotty appearance in Peters’ embryo;
Keibel, however, who made a reconstruction of this ovum, says this
appearance is merely suggestive and that no positive conclusions
can be drawn. None of the several other young ova show any vas-
cular formation. Embryo Frassi 1.17 mm. shows the first blood
vessels, which are distributed in the chorion, bauchstiel and ventral
pole of the yolk sac. No vessels were observed in the body of the
embryo. Graf Spee’s Gle 1.54 mm. is probably next in point of age,
followed very closely by Eternod’s 1.3 mm. embryo. Graf Spee
describes the first anlage of the heart, otherwise there are no vessels
in the body of the embryo; blood islands are present in the ventral
pole of the yolk sac, but no mention is made of vessels in the bauch-
stiel or chorion. Eternod’s 1.3 mm. embryo is the first specimen to
present a complete circulation. It has a very well developed umbil-
ical circulation and the villi are partly vascularized, but no vitel-
line vessels could be detected. The Kroemer-Pfannenstiel embryo
Klb. of approximately the same age as the embryo under considera-
tion is said to have a very large umbilical artery, unaccompanied
however by a corresponding vein, also omphalomesenteric veins are
present but no corresponding arteries. ‘The vessels in the yolk sac are
full of corpuscles, but no mention is made of any chorionic vessels.

This summary of the earliest vascular development is by no means
evidence in favor of a primitive vitelline circulation. Although the
facts are very meager on account of the scarcity of material and
nothing positive can be deduced, nevertheless there seems to be some
evidence in support of a primitive umbilical circulation. The first
embryo having a complete circulation is Eternod’s (1.3 mm.), with
a well developed umbilical circulation. From the other remaining
specimen (Kroemer-Pfannestiel Klb.) between Eternod’s and our
embryo it would be hard to draw definite conclusions on account of
the presence of the artery of one system and the vein of another,
forming no complete circulation.—

The presence of capillaries in the chorion, bauchstiel and yolk sac
in Embryo Frassi would seem to indicate that the mesoderm which
forms all of the above structures, is endowed with a high power of

96 Walter E. Dandy.

vascularity in its early history, irrespective of position, and that from
this vascular formation, whether from single or multiple points of
origin, two main primitive vascular areas originate practically syn-
chronously and develop independently toward the embryo. Eternod’s
embryo together with the one under consideration, which are the
two earliest embryos with a complete circulation, would seem to indi-
cate that the umbilical circulation was the first to attain this fune-
tion.

As mentioned before, this embryo agrees in general very closely
with Eternod’s 1.5 mm. embryo, differing only in a few details. The
yolk vascular system is developing rapidly and consists of numerous
large blood islands, forming irregular, disconnected channels and
masses of blood-forming tissue, situated almost entirely on the ventral
half of the umbilical vesicle (Fig. 12), extending from the anterior
to the posterior poles and more marked in the region of the future
vitelline veins. A few very small blood islands extend along the
dorsal half of the yolk sac, but never is there any visible vascular
connection within the embryo and always a considerable distance
exists between the blood islands and the vessels of the body of the
embryo.

The umbilical circulation consists of a simple vascular cycle
comprising the umbilical veins, heart, dorsal aortee and umbilical
arteries, with capillary connection in the chorion and chorionie villi.
A number of villi show the early stages of vascularization, which are
not limited to the region around the attachment of the bauchstiel, but
extend anteriorly beyond a point corresponding with the anterior limit
of the embryo. There appear to be several seemingly independent
small systems of capillaries in the chorionic membrane, into which
drain the capillaries of the villi. Although these systems seem to have
no connection with each other, the difficulty of following capillaries
in uninjected specimens would make a positive statement impos-
sible. As mentioned above, many of these chorionic vessels show
beautiful examples of what appears to be blood formation from
endothelial proliferations (Fig. 10) and is probably the source of
the blood for the present circulation.

Two large independent blood reservoirs (Figs. 8, 9 and 15) in
the bauchstiel connect distally with the chorionic vessels; proximally

A Human Embryo with Seven Pairs of Somites. of

the smaller gives off the paired umbilical veins; the larger is formed
by the confluence of the umbilical arteries.

The umbilical veins run in the dorsolateral portion of the body
of the embryo, at the point of origin of the amnion. The left vein
gives one, the right vein two, very short branches to the amnion at
the level of the posterior region of the embryo. Nothing comparable
to the sinus ensiforme of Eternod’s 1.3 mm, embryo was observed.
While still in the dorsal position and just before its course ventral-
ward, each umbilical vein gives off a short branch which runs ven-
trally through the body of the embryo toward the umbilical vesicle.
On one side it terminates before reaching the yolk sac, on the other
side it runs a short distance in the wall of the yolk sac and seems
to end blindly. On each side a couple of spaces, presumably capil-
laries, although they do not contain any blood, are in the immediate
vicinity of these short branches, but no connection can be detected.
These short branches and possible capillaries may be the beginnings
of the vitelline veins, although the dorsal point of origin from the
umbilical veins is rather against this view, unless there is a subse-
quent ventral wandering, as has been observed in many other vessels.
This is the only visible evidence of the possibility of the vitelline
veins. (Plate IV.)

The umbilical veins now run ventrally around the celom and
unite to form the heart just anterior to the origin of the foregut.
It is a simple tube of endothelium throughout. From its anterior
extremity one complete aortic arch is given off on each side (Fig.
- 1) and in addition, four short stubs on the right and two on the
left sides, which represent rudiments of future aortic arches. No
corresponding buds are given off by the cephalic portion of the
dorsal aortee to connect with these. The heart is a step in advance
of that of the Kroemer-Pfannenstiel embryo, which has one aortic
arch on each side, but a paired heart is still present. Eternod’s
1.3 mm. has a slightly higher development, in that three and _ pos-
sibly four complete arches are present on each side.

The dorsal aorte, paired throughout, begin anteriorly in a small
dilatation and terminate caudally at the posterior border of the fore-
gut in a much larger dilatation, from which the umbilical arteries are
the direct continuation (Plate VI). Each aorta gives off a series of

98 Walter E. Dandy.

eleven lateral branches, beginning as very minute and rather indefinite
branches posterior to the first somite; these become progressively
larger and reach a climax at the posterior dilatation of the aorta.
In the somite region they are solid buds (Fig. 5), the last five aris-
ing from the dilatation have a lumen, but end blindly in the meso-
derm of the splanchnopleure (Fig. 5). On the right side the last
three branches seem to end in a common blood island, giving the
impression of a small capillary plexus; this was, however, not
ebserved on the other side. These branches probably represent the
segmental arteries of Mall destined for the umbilical vesicle. In
the somite region there are six of these branches for the seven
somites; on one side all six arise between the somites; on the other
side three arise intersegmentally, the other three come off nearer
the center of the somites. From the dorsal and caudal end of the
posterior aortic dilatation there is a very short stub on each side,
which may represent the beginning of the caudal aorta. Otherwise
no dorsal branches of the aortee were observed.

The umbilical arteries, somewhat larger than the aorte, are
direct posterior continuations of the posterior aortic dilatation. In
the bauchstiel they unite to form a large sinus (Fig. 9) which lies
posterior to the smaller venous sinus. (Fig. 8.) It sends off numer-
ous large branches to the chorion, some of which anastomose and
redistribute branches to the chorionic membrane and villi.

It is a pleasure to extend my heartiest thanks to Professor Mall
for his kindness in allowing me the privilege of describing this very
rare specimen, for his valuable advice and suggestions and for the
use of two manuscripts which are not yet in print.

A Human Embryo with Seven Pairs of Somites. 99

BIBLIOGRAPHY.

BENEKE, R. Ein sehr junges menschliches Ei. Deutsche med. Wochenschr.,
Jg. 30, p. 1804.

Broman, Ivar. Ueber die Entwicklung, “Wanderung” und Variation der
Bauchaortenzweige bei den Wirbeltieren. Ergebn. d. Anat. u. Ent-
wegesch., Bd. 16, 1906.

Bryce, J. H., and Teacuer, J. H. An Early Human Ovum. Glasgow, 1908.

Erernop, A. C. F. Il y a un lecithophore dans l’embryon humain. Biblo-
graphie anatomique, T. 15, 1906.

. Communication sur un wuf humain avec embryon excessivement
jeune. Comptes Rend. du XI. Congrés int. Méd., Rome, 1894.

Sur un ceuf humain de 16.3 mm. avec embryon de 2.1 mm. Actes
de la Soe. helv. des Se. nat., 1896.

Premiers stades de la circulation sanguine dans l’ceuf et l’embryon
humain. Anat. Anz., Bd. XV, 1898.

Evans, H. M. On the Earliest Blood Vessels in the Anterior Limb Buds of
Birds. Amer. Jour. Anat., April, 1909.

On the Development of the Aorta, Cardinal and Umbilical Veins
and other Blood Vessels in the Vertebrate Embryo from Capillaries.
Anat. Record, Vol. III, 1909.

Frassi, L. Ueber ein junges menschliches Hi in situ. Archiv f. microsk.
Anat., Bd. 70; 1907.

Weitere Ergebnisse des Studiums eines jungen menschlichen Eies
in situ. Archiv f. mikr. Anat., Bd. 71, 1908.

Gacr, SusANA Puetps. Three Weeks’ Human Embryo. American Jour.
Anat., Vol. IV, 1905.

His, WitHetm. Anatomie menschlicher Embryonen. Leipzig, 1880-1885.

Incatts, N. W. Beschreibung eines menschlichen Embryo von 4.9 em. Archiv
f. mikr. Anatomie, Bd. 70, 1907.

Keiser, Franz. Ueber den Schwanz des menschlichen Embryo. Archiv f.
Anat. u. Physiol., Anat, Abt., Jg. 1891, pp. 356-398.

Ein sehr junges menschliches Ei. Archiv f. Anat. u. Physiol.
Anat. Abt., Jg. 1890, pp. 250-267.

Normentafel zur Entwicklungsgeschichte der Wirbeltiere. Jena,
1908.

KoLt~MAN, Jutius. Handatlas der Entwicklungsgeschichte des Menschen.
Jena, 1907.

KroeMer. Wachsmodell eines jungen mensch]. Embryo. Verhand. d. Ges.
f. Gyniikologie, 1903.

LENHOSSEKS, M. V. Die Entwicklung der Ganglien-Anlagen bei dem mensch-
lichen Embryo. Archiv f. Anat. u. Physiol., Anat. Abt., Vol. —, 1891.

Linuiz, FRanK R. The Development of the Chick. New York, 1908. (Ref.
to Marion Hubbard and J. T. Patterson.) (Pp. 111-114.)

MacCatium, JouHn Bruce. Notes on the Wolffian Body of Higher Mammals.
Amer. Journ. Anat., Vol. I, 1902.

Matt, F. P. Age of Embryos and Feetuses. Keibel-Mall Embryology (in
press) (manuscript by permission of author).

100 Walter E. Dandy.

Marr, F. P. On the Development of the Human Diaphragm. Bull. Johns
Hopkins Hosp., Vol. XII, 1901.

———. A Human Embryo of the Second Week. Anat. Anz., Jg. 8, 1893,
pp. 680-638.

———. Human Embryo, twenty-six days old. Journal of Morphology,
Vol. V, 1891.

—————, Development of the Human Celom. Journal of Morphology, Vol.
XII, 1897.

———. Development of the Human Ce@lom. Keibel-Mall Embryology (in
press), Leipzig and Philadelphia, 1910.

Minor, C. S. Human Hmbryology. New York, 1892.

Quarn’s Anatomy, Vol. I, London, 1908.

SPEE, FERDINAND, GRAF y. Beobachtungen an einer menschlichen Keim-

scheibe mit offener Medullarrinne und Canalis neurentericus. Archiy .-

f. Anat. u. Physiol., Anat. Abt., 1889, pp. 159-176.
Neue Beobachtungen tiber sehr friihe Entwicklungsstufen des

menschlichen Eies. Archiv f. Anat. u. Physiol., Anat. Abt., 1896,
DD =A

TANDLER, JULIUS. Ueber Vornierenrudimente beim menschlichen Embryo.
Anat, Hefte, Bd. 28, 1905.

THOMPSON, PETER. Description of a Human Hmbryo of Twenty-three Paired
Somites. Journ. of Anat. and Physiol., Vol. XULI, 1907.

WILLIAMS, J. Wurrripce.: Textbook of Obstetrics, New York, 1907.

102 Walter E. Dandy.

EXPLANATION OF PLATES.
PLATE ] (Fics. 1-7).

Fie. 1. Section 22, x 50. Bi, First primary brain vesicle; Oo, Aorta; A1,
Complete view of longitudinal section of first aortic arch; A2, Cross section
of stub of second aortic arch; En, Endothelial heart; Ht, Mesodermal wall of
heart; P.C., Pericardial cavity; Fg, foregut; Am, Amnion; Mes, Mesoderm:
Ent, Entoderm.

Fic. 2. Section 538, «x 50. Ch, Chorda; Ao, Dorsal aorta; V, Umbilical
vein; Fg, Foregut; P.C., Pericardial cavity; B2, Second primary vesicle of
the brain.

Fic. 3. Section 76, x 50. Pr, Pronephros; Coe, Celom; Pl, Pleural Ceelom ;
V, Umbilical vein; Br V, Branch of umbilical vein, only suggestion of a vitel-
line vein, it unites with umbilical vein two sections below; Ch, Chorda; M1,
First Somite in each side.

Fic. 4. Section 93, x 50. Br, Cross section of lateral branch of dorsal
aorta; V, Umbilical vein; M4, Somite IV, with cavity (anterior tip of somite
IV on opposite side) ; Coe, Celom; B.C., Communication of ceelom with ex-
terior; Ch, Chorda.

Fie. 5. Section 101, x 50. Br. Lateral branch of Aorta; Ao, Aorta;
V, Umbilical vein; M4, Somite IV with cavity; M5, Tip of Somite V; Coe,
Ceelom.

Fig. 6. Section 187, & 50. Ao, Dorsal aorta; Br, Lateral branch of aorta
(branch on left side terminates in a mass of mesodermal cells which may be
a small blood island); Ch, Chorda at point of greatest development; Coe,
Celom; S.M, Slit in mesoderm. (See also Fig. 11.) Pr. S, Beginning of
primitive streak. U. V, Umbilical vesicle.

Fig. 7. . Section 152, « 50. Shows primitive streak region (Pr. S.) from
fusion of ectoderm, mesoderm and entoderm. Hg, Hindgut; All, Allantois,
near point of origin from hindgut; U, Umbilical artery; V, Umbilical vein;
S. M, Slit in mesoderm. Ect, Hctoderm; Hnt, Entoderm; Mes, Mesoderm.

A HUMAN EMBRYO WITH SEVEN PAIRS OF SOMITES

WALTHER E. DANDY.

PLATE

Coe M4

M4 S

BIG os

Fig. 4.

*
3
E
2

Ws,

THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

104 Walter E. Dandy.

EXPLANATION OF PLATES.
PLATE II (F ies. 8-11).

Tic. 8. Section 177, « 25. Outline of section to show size of the sinus
formed by union of umbilical veins; V, Umbilical venous sinus (from union
of the umbilical veins); All, Allantois, shown in two places, the lower is
before division, the upper is terminus of long branch; Ch, Chorion; Vi,
Chorionic villus, showing syncytial bud; U, Umbilical artery, shown in two
places below, before union to form umbilical arterial sinus, and above, tip
of sinus and large branches.

Fic. 9. Section 188, % 25. Outline of section to show size of sinus formed
from union of umbilical arteries. U, Umbilical arterial sinus; All, The
terminus of the short branch of the allantois; Ca, Capillaries and small
branches of sinus in chorion; Ch, Chorionic Membrane; Vi, Chorionic villus,
showing capillary entering.

Fic. 10. > 525. Capillaries from chorion, showing apparent formation
of blood corpuscles from endothelium of capillaries.

Fig. 11. Reconstruction of ccolom, showing relation to somites; Arabic
numbers on side represent the numbers of sections; Roman numerals in
center represent the paired somites; P.C., Pericardial celom; Pl, Pleural
ceelom; Coe, Peritoneal celom; E.C., External Communication of the ccelom ;
Pr, Pronephros; Ht, Projection of the heart in the pericardial cavity; X,
Outer limit of body wall; Mes, Paraxial mesoblast ; Br.M., Bridge of meso-
dermal tissue extending across the mesodermal slit; Pr.S., Primitive streak.

A HUMAN EMBRYO WITH SEVEN PAIRS OF SOMITES.

Jaireis

Tete'g tte)

THH AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

PLATE II.

WALTHER BH. DANDY.

j
1
WL

| | HH |

sos

Ere. il.

106 Walter E. Dandy.

PLATE III.

Ic. 12. Diagram of a sagittal view of embryo to show the vascular system.
Ht, Heart; A I-II-III, First, second and third aortic arches; Ao, Dorsal aorta ;
C.Ao, Caudal aorta; U, Umbilical artery (unite in the bauchstiel) ; V, Um-
bilical vein (unite in the Bauchstiel) ; Vit, Branch of umbilical vein, is only
suggestion of a possible vitelline vein; V. am, Venous branch to amnion; Ca,
Capillaries; do not contain blood and do not connect with veins; B.I., Blood
Islands; Ect, Ectoderm; Mes, Mesoderm; Ent, Entoderm; Fg, Foregut; Hg,
Hindgut; All, Allantois; Ch, Chorionic membrane; V1, Chorionic villi, showing
vascularization; B.S, Bauchstiel; M.C., Mouth Cavity.

A HUMAN EMBRYO WITH SEVEN PAIRS OF SOMITES. PLATE III.
WALTER B. DANDY.

Amnion

Unmbilical Vesicle

Jee, 112)

THE AMDRICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, Jan., 1910.

108 Walter E. Dandy.

EXPLANATION OF PLATES.
PLATEVIVE

Fie. 13. Dorsol-lateral view of model of the embryo, with the amnion
removed. Umbilical vein with possible vitelline branch (?) seen on right side.
(The vein is projected on exterior for purpose of clearness.) Large um-
bilical venous and arterial sinuses posteriorly, with allantois lying between.
Arterial sinus is partially cut away, venous is intact. Neural canal is open
throughout, and shows the three primary vesicles of the brain.

AWE Vis
I'ic. 14. Same view as Plate IV: window of ectoderm removed, exposing
the somites and mesoderm.

PLATE VI.

Itc. 15. Same view as in Plates IV and V. Mesoderm is removed.
I, II, I11-Three primary brain vesicles; I-First aortic arch; studs of arches
2, 3, 4, 5 are seen just posterior; U-Umbilical arterial sinus; V-Umbilical
veins (umbilical veins and umbilical venous sinus have been cut away to
show the arterial system). Vi, Branch of umbilical vein, only suggestion of
possible vitelline vein; Fg-Foregut; Hg-Hindgut; All-Allantois; Ch-Chorda ;
Ht-Heart; Coe-Ceelom; P.C.-Pericardial coelom; Ca-Capillary, but no connec-
tion; Pl-Plexus of lateral aortic branches (shown only in this place). Mes-
Mesoderm in primitive streak region separated from the ectoderm by a
shrinkage space, which is only present laterally, in this region. (See also
LS ee)

A HUMAN EMBRYO WITH SEVEN PAIRS OF SOMITES. PLATE IV.

WALTER B. DANDY.

THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

A HUMAN EMBRYO WITH SEVEN PAIRS OF SOMITES. } PLATE V.

WALTDR BH. DANDY.

THE AMERICAN JOURNAL OF ANATOMY.—-VOL. 10, No. 1, JAN., 1910.

A HUMAN EMBRYO WITH SEVEN PAIRS OF SOMITES. . PLATE VI.

WALTER BH. DANDY.

Ch.

Mes.

All.

THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

A STUDY. OF THE DIFFERENTIATION OF TISSUES IN
THE REGENERATING CRUSTACEAN LIMB.!

BY
VICTOR HE. EMMEL.
From the Department of Comparative Anatomy, Harvard Medical School,
Boston, Mass.

WitH 8 PLATES.

CONTENTS.

PAGE

PeR TUG CO CLUE OL OM pet per ey eens, Sete eaten: cheer eke Se mes Ctae aueec te eeane ete wieceae 110
Hie Trend lean Gh MeKMOUS =n sce, a.cnsteps «iss ous artes theleneehersisia iontatere tase crs ape eee alti
ean aLOmIGHEsiruiGenunEexor-tne Chelar si. .s. eoeisiene so ele ele © 6) eleisseur er 112
ee SCLIMENESMIISCLESs ANG eT OUMESY 2)... aerate ce access eseke exe. siecuoke 112

PS CBIHSM gate TORE, ey eeaPueeacic ean oer cee ae RCo eIeEc ood One enc Bene Pern 1s}

(CGB) MONO STIRCVET ON zp sare a aganl oT ahcler a reeset ne tomeben oT ive foretel sveseleire 1138

(GD) Phloodevesselseamd =Mervesy dari. sercuetess ost eke becsiwis cess ei 115

(c) an embryonic or transitory IVUISCLE S255 Siees «seeks Goatees 117

(OE) SCMUILyONIVOTR ART a etene caplet nrc ciao etree oc SIGHS Dio COI EC Dt oeer aia 118

Mee harlvestacessor the resenerating Limb) . os cc.celeer sc stew se + eles ars 119
ESR enagiustmentofanjimred TISSUES 245.6 sce ae lam cle wrs aaron ss = 119

mV ATCA Uel Og lessee aanre ancien chen seek pees cae ot aor one’ aacbareys, Oe) oaats, afecePehene teas 120

See haneeceam th Orr Gl tien vacraya sew ele sratees eee oka. & sratione ves elses wag nes 120

APe SIT OM CYLON ASINT Be ot sete eves aoe ace os hele rote alote oak e eel sel et ehlecore, ce: cvarete 125

pee Cilia Vat SLO MNes <gevans fave cnc atone oteasc kot oroNtutie’ ol caohoat osu euc telah OOS ie eeve ees 126
VeLORTT Aone Ol JOULES ANC SCLMEMIS 2.0... teeiloctavsins ostece sys, s 0-8 6/6 3 127
em OTOH MOTs Wi COI Sata teyafe. + ele, echoes evel arancictie! orster=. 8 otensiche co sila «auc, Siete fs ovatoy seus 130
MPM Te TeTiAtTOM V Ol TISSUES! aye). ore’ acts ch clcite ante ole oeteyer eres, «) eile elit im "el alae ener aie 132
PMR CH Tel Cee S ClO one che - ayatersne eseserts a7oele erste ec. c0et leas als siscene dienes es Abe

(CA MISCOSCMESIS@ Face sere euencies se elclesevetecta cue ee iene) orale al etait 132

(oO) attachment torexoskeletiomy 2535 52.5 .sse0% see oss s 136

C@)MGISGUSSTON ANG <COMEMISLOMS eye crete s eaiiete fay overs sc) oes, 137

PMID S Teich mR MP an, Ag op fee en hurd IE alta. a ea ay crlateh opie dscteRen uote e seheseoe ene 142
UOMO CLV EISSUTOs et .coisiz/shciers)<, sais whois op =) apaie elelsne 5 c:N ies. os (op acelelfe = 16s 144

‘This paper is one of a series of experimental and anatomical studies of
regeneration (’05, ’06', ’06?, ’06*, ’07', ’07, ’08).

THE AMPRICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910,

110 Victor E. Emmel.

PAGE.

VIII. Directionvof differentiation” 22-45...-7520e eee ee eee 146
1. Chitinogenous plates or muscle tendons ..................- 147

2. Myoibrillse: +20..cire aooitasiteatier eta notre Hee eee 150

3. striation of muscesibrilsye-ee ere eestor eee eee 151

4. ‘Conclusions. .iscc4 025, foe enien tee ee ee eee 151

PX.” -SUMMALY’ esis crectials SEs Slee eyotere Sse ree eee ern aero eee 152
RR. Diteratures sa vic Halos v ors ererever the eanteee nie cess ial ore eee 155
<I. ~Description of plates i522. cascicetern tec ie «sie he clot onic one ee 157

INTRODUCTION.

The recent work by Reed ((04) on regeneration in the crayfish,
Ost ((06) on Oniscus, and Steele (’08) on the regeneration of the.
compound crustacean eye, draws attention to certain fundamental
questions. Their results are in marked contrast with the observa-
tions made on vertebrates where it appears that a regenerating tissue
almost invariably arises from similar pre-existing tissue. The im-
portant relations of this subject to the problem of both the normal
and pathological origin and cytomorphosis? of new tissue cells, render
it desirable to investigate further the histogenesis of regenerating
tissues. The present paper contains the results of a cytological
study of the cells and tissues in the regenerating chela of the lobster,
a crustacean which has been especially favorable for this study, both
on account of its highly developed capacity for regeneration and
because of the unsurpassed opportunity for obtaining suitable mate-
rial at the lobster hatcheries.

The results to be described deal more especially with (1) the eyto-
logical changes observed in the regenerating ectodermal cells; (2)
the genesis of striated muscle, nerve, and connective tissue; and (3)
the morphology of certain structures at the breaking joint of the limb,
which as far as the writer is aware, has not previously been con-
sidered.

I desire to express my thanks to Professor C. S. Minot, of the
Harvard Medical School, for interest and encouragement during the

*This term is used “to designate comprehensively all the structural modifica-
tions which cells or successive generations of cells may undergo, from the
earliest undifferentiated stage to their final destruction.” See Minot (’01,
p. 494.)

Tissues in the Crustacean Limb. alg bl

progress of the present work. I am also indebted to Professor
A. D. Mead, of Brown University, for generously permitting me
the use of materials and apparatus at the Experiment Station of
the Rhode Island Commission of Inland Fisheries; and to Professor
R. J. Terry, of Washington University, for valuable suggestions
during the completion of the present paper.

Il. Materirat anp Mrtuops.

A serious obstacle encountered in determining the origin and dif-
ferentiation of a regenerating tissue is the difficulty of securing a
complete series of successive developmental stages. If a number
of animals are operated upon at the same time and material fixed
at successive intervals, it is usually found necessary in subsequent
study to make an extensive rearrangement of the material on account
of variations in size and rate of growth. These technical difficulties
were considerably lessened in the present work because of the excel-
lent material obtained at the lobster hatchery of the State Experiment
Station at Wickford, Rhode Island. From a hatchery pool contain-
ing several thousand lobster fry, it was possible to select a large
number of animals which were practically equal in age and size,
and which had all moulted to the same stage within less than twenty-
four hours. This last point is of considerable importance, since
it had been ascertained in previous experiments (’06) that the age
of the animal and the time, in relation to moulting, at which the
operation is made are important factors influencing the rate of regen-
eration. These young lobsters were also further favorable for study
because the exoskeleton had not as yet attained such an amount of
chitin and lime salts as to necessitate decalcification before sectioning.

The operation consisted in the autotomous removal of both chelze.
The lobsters, about 300 in number and all practically equal in age
and size, were placed in floating cars and kept in as nearly normal
condition as possible. Specimens were then preserved at two, four,
and eight hour intervals throughout a period of fourteen days, in
which time the chele had undergone complete regeneration. From
this succession of stages about 54 were selected, sections of the chele
were cut 5 micra in thickness and mounted in series. In order to

112 Victor E. Emmel.

facilitate a comparative study, the specimens were all cut in a nearly
constant plane. In addition to this series, sections of the regener-
ating limbs of older lobsters were also studied.

The material used was fixed in a saturated 35 per cent alcoholic
solution of corrosive sublimate. Several other fixing agents were
tried, but all solutions containing acids proved unsatisfactory on
account of their injurious reaction with the chitin and salts of the
exoskeleton. The nuclear and cytoplasmic stains employed were
alum heemotoxylin, Heidenhain’s iron-alum-hematoxylin, Congo-red,
eosin, and Mallory’s connective tissue stain.

III. AnaTomMiIcaL STRUCTURE OF THE CHELA.

1. Segments, Muscles, and Joints.—The following account is con-
fined to those structural characteristics of the limb which are essential
for the subsequent description of its regeneration.

The chela of the lobster is composed of six segments (Fig. 37).
Proceeding in a disto-proximal direction these segments are known
successively as a dactylopodite or dactyl (d), propodite (p), carpopo-
dite (c), meropodite or meros (m), ischipododite (7), and basipodite
(bs). A seventh basal segment not involved in the present study is
the coxopodite. The index is the distal part of the propodite which
opposes the dactyl and forms one jaw of the large claw. All of these
seven segments are united to each other by flexible joints with the
exception of the last two,—the ischiopodite and basipodite. These
are fused together into an immovable joint or “breaking-joint,” a
structure especially adapted to the process of autotomy. ach joint
is like a hinge, articulating at two points and permitting only a
simple flexion and extension of the segment. The musculature of
the chela consists of a pair of opposing muscles for each of the
segments with the exception of ischiopodite (Fig. 37). The pair
of muscles in any given segment is attached proximally to the chitin
of the proximal region of the segment. Distally the converging
muscle bundles are attached to a chitinous plate or ingrowth of the
exoskeleton of the next segment distad. The origin and insertion
of the extensor and flexor are on opposite sides of the joint. In
each of the first three pairs of muscles, the flexor is larger than the
opposing extensor.

Tissues in the Crustacean Limb. 133

The ischiopodite is an exception to the above description, for the
muscles are not arranged to act in opposition. There are two
muscles in this segment, but they both have their origin on the same
side of the limb, 7. e., on the ventral side of the fourth joint (Fig.
37, e*). One muscle is slightly larger than the other, but both are
smaller than any other muscles of the chela. The larger muscle is
inserted partly on the morphologically anterior side and partly on
the dorsal surface of the exoskeleton in the proximal part of the
ischiopodite. The smaller muscle is inserted directly on the morpho-
logically anterior wall of the segment. Both muscles, therefore,
appear to function as extensors. A similar arrangement of muscles
has been described by Fredericq (’92) in the ischiopodite of the crab.
That a similar condition exists in the crayfish is not so clear, how-
ever, for Reed (’04) states that the ischiopodite of the cheliped in the
crayfish possesses ‘‘both an extensor and a flexor” (p. 310).

2. The “Breaking Joint.”—a. Exoskeleton—By the breaking
joint (Fig. 37, bk) is meant the region in which a fusion has taken
place between the ischiopodite and the basipodite. Since it is at
this region that the chela is amputated in autotomy and the new limb
begins to regenerate, a more detailed description of the breaking
plane or joint is essential for our present purpose.

The external appearance of the breaking joint in the adult has
been described by Herrick (’95) as a “fine hair line leading from
the small spur next to the articular facet on the under side, round
the anterior border to the upper side of the joint. It then bends
forward and abruptly backward, crossing the small proximal end of
the joint, to near its point of departure” (p. 10). This hair line
represents the plane of union of the basipodite and ischiopodite into
a solid compound segment. In the first three larval stages there is a
movable articulation between these two segments and as described
by Herrick, “there is no true fusion of the segments until after the
fifth stage” (p. 102). It may be questioned, however, whether there
is any longer a functional articulation of these segments even during
the fourth and fifth stages of development. Although externally
the joint may present the appearance of a free articulation as indi-
eated in Herrick’s figures, the examination of longitudinal sections

114 Victor E. Emmel.

of the fourth stage limb shows a different condition. As may be
seen in Figs. 1 and 2, there is strictly no free articulation evident,
especially on the morphologically ventral side (see also Fig. 3, bk),
where there is an almost perfect continuity of the exoskeleton from
one segment to the other. On the dorsal side the union is less com-
plete. A difference between the two sides of the breaking joint
is evident even in the adult where the chitin is still much thicker
on the ventral than on the dorsal surface. In section the region of
the breaking joint can easily be identified by the characteristic color-
ation which the chitin takes with Mallory’s connective tissue stain.
With this reagent the exoskeleton, with the exception of the outer
lamellee of chitin, takes on a bright blue color, whereas the line of
fusion between the two segments is sharply differentiated by becom-
ing dark red.

The internal structure of the breaking plane of the limb may be
described as follows. The epithelium (epidermis or “hypodermis’’)
of the exoskeleton is continuous across the breaking plane, from one
segment to the other. The connective tissue occupies the interior of
the breaking joint and is complex in its arrangements. Certain
structural relations of this tissue were discovered, which differ con-
siderably from those previously recorded. They will presently be
deseribed.

Tt can be readily appreciated that the mechanism for preventing
an excessive loss of blood is an important element in the perfection
of a breaking joint adapted to the autotomous remoyal of the limb.
Such a mechanism is supplied by the connective tissue structures.
Frederieq (’92) was among the first to describe the separation of
the blood eavities of the basipodite and ischiopodite by a “cloison
membraneuse” or circular diaphragm, stretched across the distal part
of the basipodite (p. 177). Similar descriptions have been written by
Andrews (790) for Libina, and by Herrick (’98) and Reed (709) for
the lobster and the crayfish. This transverse membrane was at once
interpreted as a structure for preventing hemorrhage after autotomy.
Andrews describes the membranous fold in the breaking joint of
the spider crab as extending “from the epidermis to the central nerve
and blood vessels,’ and states that after autotomy the membrane covers

Tissues in the Crustacean Limb. 115

the entire surface of the stump except at the centre, ‘where there
is a rounded hole with a little torn tissue and blood exposed”. Reed
states that in the lobster and the hermit crab “the opening through
the membrane for the nerve and the blood vessel is in about the centre
of the exposed surface when the leg is thrown off” (p. 309). In an
earlier description of the lobster, the writer (705) also described this
membrane as “extending almost entirely over the basipodite” and
“perforated only at the centre by an artery, the blood sinus, and a
large nerve” (p. 89). It was with considerable surprise, therefore,
that it was found in the present study of serial sections of the chela
that the above descriptions cannot, for the lobster at least, be regarded
as strictly accurate, particularly in regard to the relation of the blood
sinuses.

In order to obtain a more thorough conception of these structural
relations, graphic reconstructions of the breaking joint have been
made from serial sections of the right chela of a fourth stage lobster.
The tissue structures of the breaking joint at this stage are perhaps
somewhat simpler than in the chela of an older animal, but their
essential relations are, I believe, practically the same. It should
also be stated that these reconstructions are in one detail not complete,
since the connective tissue network in which the arteries and nerves
are partly embedded as they pass along the walls of the large venous
blood sinuses, is not represented in its entirety. Figs. 1 and 2, re-
spectively, show the outer and inner, or morphologically posterior and
anterior, parts of the breaking joint and adjacent segments. Tig. 3
represents a median section between the two parts shown in Figs.
1 and 2. The latter figures do not show quite half of each side.

b. Blood Vessels and Nerves.—It may be observed in these figures
that there are two nerves, two arteries, and a large venous sinus
(divided into two separate channels) which pass through the region
of the breaking joint. The two nerves n', n?, lie on the inner or
medial side of the joint, the larger nerve (n') giving off a small
branch as it leaves the basipodite. The main artery (a') passes
along the outer wall. In the basipodite it gives off a small branch
(a?) which takes an oblique course across toward the inner wall of
the ischiopodite. The venous blood of the chela is carried toward

116 Victor E. Emmel.

the body through an immense blood sinus (vs). As this sinus passes
through the ischiopodite it is temporarily separated into two channels
by a connective tissue partition or septum (S). One of these chan-
nels passes ventrally and the other dorsally, through the breaking
joint. The two nerves and arteries lie within the septum and
adjacent connective tissue separating the two venous channels.

In the course of these two blood channels just as they enter the
basipodite after having passed through the breaking joint, there
occur two very interesting valve-like folds of connective tissue. These
two structures arise in the following manner from the median septum
separating the blood channels. As the septum extends proximally
into the basipodite it becomes divided into two plates or folds
(v', v2) much like the arms of an inverted “Y,” one of the arms
passing toward the dorsal wall and the other toward the ventral
wall of the segment. The fold extending toward the dorsal wall is
somewhat shorter and thicker than the one on the ventral side.
These two folds do not completely unite with the walls of the basi-
podite. A part of the border of each fold retains a free edge, which,
together with the opposing wall of the segment, completely encloses
the lumen of each blood channel. Beyond (7. e., proximally) these
folds the two blood channels reunite to form again a common venous
sinus (vs”).

While endeavoring to understand the significance of these folds
as they were first seen in the serial sections, it was suggested that
they might possibly function as valves for the venous blood channels,
the valves closing when the limb had been autotomously removed.
Following up this suggestion, serial sections of specimens which had
been fixed immediately after the autotomy of the chela, were then
examined. It was at once evident that the folds in question had
assumed in these specimens a position different from that observed
in sections of a chela before the limb had been removed. Each fold
had now become distended in a distal direction as if caught by an
outward blood pressure, the result being the complete closure of the
ruptured end of the two blood channels (Fig. 4, v?). Desiring to
make a still further test, a number of live lobsters were then obtained
and the chele autotomously amputated. At first there was a short

Tissues in the Crustacean Limb. sia Eg

jet of blood from the exposed stump, and then a slight bulging of the
tissues over the surface of the stump with no further escape of blood.
The location of the two venous channels could be distinguished quite
readily by the transparency of the now turgid valves closing each
lumen. The two valves were then gently pressed inwards with the
tip of a pencil and in each case, a slight opening of the valve being
effected, there at once followed a jet of blood. The valvular function
of these folds seems, therefore, to be clearly demonstrated.

This description may be summarized with the statement that a
septum of connective tissue extends across the cavity of the limb
in such a way as to divide the venous sinus into two channels in
its passage through the breaking joint. Through this septum and the
adjacent connective tissue pass the two arteries and nerves. Proxi-
mally the septum forms two folds which function as valves for the
venous channels.

It can readily be appreciated how in previous descriptions some
of the relations of these structures may have been overlooked. For
when the valves have closed after autotomy, the remaining stump
presents every appearance of a continuous membrane extending over
its exposed surface, ‘‘perforated only by a nerve and a blood vessel,”
so that in the following statement Fredericq (’92) even indicates
a doubt regarding the passage of the vein, “Seul le nerf mixte de la
patte, V'artére (et un sinus veineux?) franchissant cette limite et
passent du basipodite dans Vischiopodite, 4 travers un orifice étroit,
creusé dans la membrane obturatrice” (p. 177).

e. An Embryonic or Transitory Muscle-—Both Fredericq (792)
and Morgan (’00) have recorded the fact that in the chela of the
adult crayfish or lobster “no muscle passes from the ischiopodite to
the basipodite”’. In other words, in the adult lobster no muscle
extends across the breaking joint. It was with considerable surprise,
therefore, that it was found that this is not true for the larval lobster.
In the study of sections of the chela of larval animals, a well devel-
oped muscle was discovered extending from the ischiopodite to the
basipodite, and consequently passing through the breaking joint.
This muscle is easily seen in a longitudinal section of the limb (Fig.

5, m). It is attached to the inner side of the ischiopodite, passes

118 Victor EK. Emmel.

through the ventro-anterior region of the breaking joint, and is
inserted upon the inner wall of the basipodite. It functions as a
flexor. This muscle persists throughout the first four larval stages
of the lobster’s development. During the fifth stage, however, it
begins to degenerate, and at the sixth stage the former muscle is
represented by a mere bundle of connective tissue (Fig. 6, ct.), con-
taining a few remnants of the degenerating muscle fibers (md).

It is interesting to note that the stage of the lobster’s development
at which the degeneration of this muscle occurs is not only the one
just preceding the stage in which the asymmetrical differentiation
of the chela first becomes evident, but, as the writer (’08) has previ-
ously shown, it is also the stage after which the asymmetrical differ-
entiation of the chelze can no longer be experimentally controlled.

The discovery of this muscle introduces a new fact to be taken into
account in the question of the morphological significance of the
breaking joint. Andrews (’90) maintains that the origin of the
breaking joint may be ‘“‘explained as a modification of a former free
joint” (p. 142). On the other hand Reed (’04) concludes, from a
comparative study of the crustacean limb, that it ‘“‘seems, therefore,
erroneous to state that the breaking joint corresponds to the lost joint”
(p. 311). Without entering into a discussion of this question, it
may be observed that the existence of a muscle in the chela of the
larval lobster which crosses the breaking joint and is attached to its
two adjacent segments, seems a strong point in favor of Andrew’s
conclusion.

d. Autotomy.—The process of autotomy or defensive mutilation
among crustaceans has been fully described by Frederieq and Mor-
gan. When the nerve within the chela is experimentally stimulated
a vigorous contraction occurs in the muscles of the basal segment,
followed by a sudden snapping off of the limb at the breaking joint,
leaving the exposed surface of the stump as smooth.as if eut with a
keen-edged knife. Regarding autotomy in the larval lobsters, I ean
confirm Herrick’s observation that the “casting of the claw” does
not occur before the fourth larval stage. In experiments with
several hundreds of first, second, and third stage lobsters in which
one or both chelee were removed, I have never observed the limb to

Tissues in the Crustacean Limb. 119

be thrown off autotomously, although under a slight tension, such
as may be produced by a gentle pull with a small forceps, the chela
will, however, usually part in the region between the basipodite and
ischiopodite, 7: e., at the joint which is destined to develop later into
the breaking joint of the adult animal.

TV. Earzty Sraces oF THE REGENERATING Lime.

1. The Readjustment of Injured Tissues.—Fig. 4 represents a
longitudinal section of the stump of the right chela just after
autotomy. ‘The valve (v?) has been carried outwards as the result
of the amputation and now closes the venous channel ve. The ex-
posed edge of the layer of epithelial cells of the epidermis (@) pro-
jects slightly above the level of the broken chitin (ch) of the stump.
Shortly after autotomy the vascular cavities at the distal end of the
stump become filled with a dense mass of nucleated blood corpuscles
(Fig. 7, bc). The muscle (Figs. 2 and 5, m), which was torn by
the operation, degenerates and is not restored again in the later
regeneration of the limb, a result which might be expected, for it
will be recalled that normally this muscle degenerates during the
fifth stage and is absent in the adult lobster.

A protecting crust of coagulated blood plasma and blood corpus-
cles soon becomes evident over the exposed surface of the stump.
This blood clot (Fig. 7, bcl) forms above, or in other words distal to,
the connective tissue membrane extending across the limb. The
relation of this blood clot to the connective tissue membrane differs
somewhat from that described by Reed (’04) as occurring in the
erayfish, where the “blood cells form a plug at the opening through
the membrane, and collect in a thick layer just beneath the ectoderm
cells, which form the proximal layer of the membrane of the breaking
joint. . . . After a time these blood cells arrange themselves
in horizontal lines crowded close under the ectoderm cells of the
membrane.” Reed further states that “in a few hours from the time
that the leg is thrown off the membrane takes on the appearance of
dead tissue. . . . The blood cells soon degenerate, and most
of them are thrown away with the membrane, when this is later cast
off” (p. 312). In contrast to this, as may be observed in Figs. 4

120 Victor E. Emmel.

and 7, the corresponding membrane of the chela of the lobster is
composed only of connective tissue and does not appear to degenerate
to any appreciable degree. On the contrary, in the lobster the origi-
nal connective tissue membrane and the valves for the venous sinuses
persist throughout the regenerative activities, and retain their original
function in the new limb. In Fig. 7 it may be seen that as the
regenerating epidermal cells begin to migrate across the stump they
pass between the outer blood clot (bel), and the connective tissue
membrane and the venous valves. ‘The venous valves no longer ex-
erting a resistance to the blood stream, now begin to assume their
original and normal position (Figs. 7 and 8). There is little, if
any, degeneration of the connective tissue. In the blood clot, how-
ever, the corpuscles soon deteriorate, the nuclei become flattened in
the disto-proximal direction of the limb, and both the nuclei and the
cytoplasm of the clot assume a deeper stain with hematoxylin and
Congo red.

9. Migration of Epidermal Cells.—As the blood clot forms, the
epithelium of the exoskeleton retracts slightly within the stump.
There is, however, no other noticeable change in the position of the
epidermal cells until after the fourteenth hour following the opera-
tion. The first regenerative activity which is then observed is a
migration of ectodermal cells from all sides of the epidermal wall.
This movement of the epidermal cells appears at first to be more
pronounced in the region of the limb nerves on the inner side of the
stump ; in a section taken through the outer side of the stump twenty-
four hours after autotomy (Fig. 7) the migrating epidermal cells (e*,
e!) from the opposite sides are still quite widely separated, but in a
section from the inner side of the stump the migrating cells are found
much nearer each other. These cells continue advancing centripetally
until they unite to form a complete layer of epidermal cells over the
surface of the stump. During the migration, the congested mass of
blood corpuscles within the distal end of the stump begins to disinte-
erate, but the old connective tissue remains intact as already de-
scribed.

3. Nuclear Changes in the Epidermal Cells.—The epidermal
epithelium consists of low columnar or cuboidal cells, apparently

Tissues in the Crustacean Limb. 191

fused into a true syncytium. Indeed, Huxley’s description of the
epidermal epithelium of the crayfish apples equally well to the lob-
ster,—‘‘It is found to consist of a protoplasmic substance in which
close set nuclei are embedded . . . there can be no doubt that
it is really an aggregate of nucleated cells, though the mits between
the individual cells are rarely visible in the fresh state” (p. 178).
It may be added that in the lobster at least, cell boundaries are no
more clearly defined even after fixation and staining. In view of
these facts it becomes evident that in making observations upon cell
changes in such a tissue, attention is necessarily more largely directed
toward the nuclei of the tissue cells.

The characteristic structure of normal epithelial nuclei as they
appear immediately after the autotomy of the limb is shown in Fig.
11. They may be described as more or less elongated or oval in
form with a clearly defined nuclear membrane. A structural charac-
teristic is the segregation of the chromatin into relatively large angu-
lar masses or karyosomes. ‘These karyosomes take on a heavy stain
with hematoxylin, and are rather evenly distributed throughout the
periphery of the nucleus. ‘The ground substance stains a uniformly
light color. The four nuclei in Fig. 11 were from the epidermal
epithelium just below the breaking plane and from a region at the
inner side of the basipodite near the limb nerves.

Fig. 12 represents epidermal nuclei fourteen hours after autotomy.
To facilitate a more direct comparison of structural characteristics,
these nuclei were taken from practically the same region as the nuclei
shown in Fig. 11, 2. e., from the inner side of the basipodite,—a
region of the epidermis where regenerative activities are first ob-
served. Of these four nuclei, a and b are proximal of ¢ and d and
farthest from the region of the first regenerative changes. In con-
trasting c and d with a and b, and also comparing with Fig. 11, it
may be noticed that the nuclei are becoming larger in size and the
chromatin is undergoing certain changes. In a and b the karyosomes
are still large and are similar to those in Fig. 11. But in ¢ and d
the chromatin is more finely subdivided and arranged in a reticulum
of chromatin strands and knots. In nuclei a and ¢ a round, centrally
located body is seen, which takes a relatively lighter stain and has the

122 Victor E. Emmel.

characteristics of a nucleolus. Whether the presence of nucleoli is
typical of this stage is, however, questionable, for with the same stain
they can also be found, although less frequently, in the normal or
resting nuclei.

The nuclei shown in Fig. 13 are from a limb fixed twenty-four
hours after autotomy. At this time the migrating epidermal cells
have progressed considerably in their advance over the surface of the
stump. ‘The nuclei drawn are from the same specimen as are those
of Fig. 12, but they have been selected from a region where the
advancing epithelium has already partly closed over the inner side
of the stump. As the epidermal nuclei migrate, their long axes
rotate through an angle of about 90 degrees from their former posi-
tion, and in such a way that at the center of the stump the long axis
of the nucleus assumes a position practically parallel to the long axis
of the limb (Fig. 8). In Fig. 13 the nuclei ¢ and d are nearer the
center of the stump. As the nuclei approach the central region, there
is an increase in their shortest diameter, giving each nucleus a broader
and less elongated form. They now also become more widely sepa-
rated from one another. In the earlier stages of migration the lower
or proximal ends of the nuclei tend to assume a tapering or pointed
form.

A striking structural characteristic is the unequal distribution of
the chromatic material (giving the nuclei a “loaded” appearance).
The proximal end of each nucleus takes a relatively deep stain, while
the upper or distal end of the nucleus is so pale that frequently little
or no stain can be detected except in the few scattered granules of
chromatin. In nuclei a, 6, and d, one can see that the boundary be-
tween the light and dark areas, or zones, lies at such an angle across
the nucleus that the darker zone occupies the lower and left region,
and vice versa for the lighter zone. The nuclear contents have much
the appearance of having been centrifuged. In nucleus ¢ the plane
separating the two zones is more nearly equatorial in position. It will
also be reealled that these nuclei were taken from the left side of the
section of the limb (¢ being nearest the center of the limb), In nuclei
taken from the opposite or right side of the same section, it is inter-
esting to find the localization of the two zones just reversed, 2. ¢., the

Tissues in the Crustacean Limb. 123

more lightly stained area is now at the upper and left side, whereas
the darker zone at the proximal end is directed toward the right side
of the nucleus. In studying the sections, a considerable enlargement
or expansion of the nucleus is frequently found at the distal end, or
region of the light zone, in some cases being even more marked than
is shown in nucleus d (Fig. 13). At this stage of migration the
former karyosomes have now almost disappeared. The chroinatin
has disintegrated into fine granules, which appear to be either per-
ipherally distributed or axially segregated into a dense mass. It is
also to be observed that the denser central mass is always located at
the darker pole of the nucleus.

The significance of this polarization of the nuclear contents (if it
may be thus designated) is not readily apparent. It can hardly be
considered an artifact resulting from faulty fixation or staining.
On the contrary, its regular occurrence, its reversal at opposite sides
of the limb, the frequent expansion of the lighter pole, the associa-
tion of the axial chromatin mass with the darker pole, together with
the fact that similar conditions are even more conspicuous in later
stages of the regenerating limb as will be preseatly described, is
evidence which supports the conclusion that this polarization of
nuclear contents is a characteristic structural feature of certain phases
in the regenerative activity of the epidermal cells.?

Rand’s (’04) discussion of the regenerating epidermis of the
earthworm applies equally well to the migrating epidermal cells in
the regenerating lobster’s limb. He says: “The movement of the
cells at the margin of the layer is not a passive one, resulting from
external pressure. Nor is there the slightest ground for supposing
the existence of a force acting from a position anterior to the cut
epidermal edges and serving to pull the epidermal cells over the cica-
trix. We are compelled, therefore, to look in the individual epi-
dermal cell itself for the immediate source of the activity. This
activity, by whatever mechanism effected, must be occasioned by an
agency external to the cell, viz., by some factor of the conditions
resulting from the injury.” Beyond this we must admit that as yet we

“The resemblance of the “polarized” nuclei in appearance to certain stages
of synapsis of the nuclei in spermatogonia may be noted, although it has
suggested no further explanation of the “polarization.”

124. Victor E. Emmel.

have not succeeded even in determining whether the stimulus thus
controlling the cell activity is some “chemical peculiarity” of the
injured and exposed tissue, or whether it is some “‘inter-action” of
the epidermal cells, questions which are merely preliminary to under-
standing the factors which may account for the apparent polarization
of nuclear contents just described. These nuclear changes might
possibly be classed among the phenomena designated as ‘“‘cytotaxis”
(Roux, 796), although in doing so it is not evident that we advance
materially in a solution of the problem. At the end of the next
fourteen hours, 7. e., thirty-eight hours after autotomy, the epidermal
cells have spread completely over the stump and the first mitotic cell
division appears. During this period the nuclei have assumed a
more spherical form and appear somewhat larger. Both the nuclear
sap and chromatin have become more equally distributed. The
chromatin is now more coarsely granular, and as the nucleus ap-
proaches the prophase of mitosis, the chromatin collects in con-
spicuous rod and knot-like masses or chromosomes (Fig. 14, a). The
nuclear membrane then disappears and the cell passes through the vari-
ous mitotic phases, of which a later metaphase is shown in Fig. 14, D.

The volumetric increase of nuclear material during the initial regen-
erative activities presents another fact for consideration. In many
respects the problem arising from the phenomena of cytomorphosis
and regulation in the reproduction of a part of the organism are
comparable with similar problems in the development of the original
embryo. In both processes the differentiation of anatomical structure
is preceded by the formation of a more or less undifferentiated mass
of cells, characterized, in the case of embryonic development, by
the segmentation of the ovum, and in the case of regeneration, by
the proliferation of a mass of cells from the tissues in the region
of injury. While the present data hardly justify an extended dis-
cussion, attention may, however, be called to the fact that in both
regenerative and ontogenetic processes, the initial cytological changes
are attended by a characteristic increase in the amount of nuclear
material.‘

‘It should be observed that in the case of regeneration, at least in the earlier
stages, the increase of nuclear material apparently is not associated with cell
division.

Tissues in the Crustacean Limb. 125

In regard to the segmentation of the ovum, Professor Minot (’08)
emphasizes this increase in nuclear material as a characteristic
feature of “the process of rejuvenation,’ and as a result of his
studies of the structural relations of cytoplasmic and nuclear mate-
rial during the development of the organism, concludes “that as we
define senescence as an increase and differentiation of the proto-
plasm, so we must define rejuvenation as an increase of the nuclear
material,” p. 167. Whether we shall be justified in attaching a
similar significance to the corresponding nuclear changes in regen-
elation, or whether in the latter case the nuclear phenomena are
fundamentally different in nature, are questions for further inves-
tigation.

4. Cytoplasm.—In describing the cytoplasm of the epidermal cells,
it is necessary to consider the cells collectively, on account of the ill-
defined or even non-existent cell boundaries. In the normal or rest-
ing cell the cytoplasm has a reticular structure of rather even appear-
ance and is traversed by larger delicate supportive fibrils lying more
or less parallel to the long axis of the cell. As the epidermal cells
migrate over the surface of the stump, their cytoplasm ceases to be
characterized by the presence of fibrillar structures, but appears to
be composed of fine granules rather uniformly distributed (Figs. &
and 9). A definite reticular structure is no longer evident. The
inner surface of this sheet of migrating cells, instead of being smooth,
presents an uneven contour due to numerous rounded inward projec-
tions of cytoplasm. These projecting masses of cytoplasm never
show definite limiting membranes, but their boundaries can, however,
be traced upward into the general layer of cytoplasm as far as the
level of the epidermal nuclei, the relations of each mass being such
as to indicate that it represents the unit of cytoplasm coming under
the influence of each nucleus. The volume of cytoplasm appears
to have increased, as is indicated by the wider separation of the
nuclei and the greater depth of the epithelium after its migration
has been completed.

One result of these observations on the epidermal cells is to empha-
size the fact that in the migration of these cells as the first step in
regeneration, there occurs a parallel series of definite structural

126 Victor E. Emmel.

changes in cytoplasm and nucleus. It remains for further observa-
tion to answer the interesting question whether the parallel occur-
rence of these structural changes is due to the existence of a signifi-
cant correlation between the nucleus and the cytoplasm in the regen-
erative activities of the cell. Such a correlation is supported, for
example, by Eyeleshymer’s (04) thorough work on the muscle cell of
Necturus, in which that investigator finds strong evidence that the
nuclear material plays a most important role in cytoplasmic syn-
thesis. He suggests that “cellular degeneration and regeneration are
accompanied by volumetric, structural and chemical changes in
chromatin” (p. 307).

5. Cell Division.—Reference has already been made to the fact
that mitotie cell division was not observed until the second day follow-
ing the operation, and then only in the epidermal cells. Thirty-
eight hours after the amputation, two mitotic figures were found in
serial sections of the right chela and four figures in the left chela of
the same specimen. Twelve hours later, eighteen mitotic figures
were counted. Preceding mitosis the nucleus migrates toward the
outer surface of the epidermis so that the mitotic figures are always
found near the periphery of the epithelium. Regarding the ques-
tion of amitosis in the early stages of regeneration, it may be said
that there was no evidence found for direct or amitotie division of
the epidermal cells during the thirty-eight hours preceding the first
mitosis. Each migrating nucleus seems to maintain its original
unity up to the time at which mitosis first occurs.

Without entering further into a discussion of this interesting ques-
tion of amitosis, attention may be called to the contrast of the present
results as compared with ohservations on the compound crustacean eye,
where it appears that in “all cases of the regeneration of the eye the
nuclei are increased by amitotic division” (Steele, ’08, p. 183).
Steele, referring to Reed’s (704) conclusion that mitotic figures do not
occur in the early stages of the regenerating limb of the crayfish, sug-
gests (p. 184) that amitotie division may possibly have taken place
“during the preparatory stages at least.” In regard to this question,
the evidence, in the lobster at least, is of a negative character. It ap-
pears rather that an increase in both the quantity of cytoplasm and

Tissues in the Crustacean Limb. 127

the size of the nuclei, together with their wider separation during
migration are sufficient to account for the formation of the first
epithelium over the injured surface of the limb.

Similar preliminary regenerative conditions have also been observed
in other animal forms. Among the vertebrates, Barfurth (790),
from a study of regeneration in the amphibian tail, concludes that
the new cells which first cover the wound ‘“‘stammen her vom persis-
tierenden Epithel der Wundrander, sind nicht etwa durch Theilung
aus diesen Epidermiszellen hervorgegangen, sondern haben sich aus
dem Epithelverbande losgelést, sind embryonal beweglich (amdéboid)
geworden und schieben sich langsam iiber die Wundflache vor, bis sie
mit den Zellen der anderen Seite Fiithlung gewonnen haben”. The
process continues until “eine mehrfache Schicht die Wunde bedeckt”’
(p. 417). Among invertebrates, Rand (704, p. 39), in his study of
regeneration in the earthworm, states that “the wound surface becomes
completely covered by an epidermal layer derived from the existing
epidermis, without the occurrence of cell proliferation in that layer”.

V. Tur ForMATION OF SEGMENTS AND JOINTS.

In this section will be described the earlier stages in the regenera-
tion of the limb. The description of the further differentiation of
the tissues and the sequence in which the segments develop is deferred
to a later section.

At the time when the first mitotic cell division appears there is
already present a thin lamella of chitinogenous cuticle formed on the
outer surface of the epidermal cells (Fig. 9, ch’). The fact that a
cuticle could be detected as early as the twenty-fourth hour after
operation indicates that cuticle differentiation may begin before cell
division has occurred.

With the appearance of mitosis, there soon follows a rapid accu-
mulation of cells in the central region of the epidermal disc of cells
which has formed over the injured surface of the stump. This disc
of ectodermal cells consequently becomes thicker, its cuticle increases
in amount, and at the end of fifty hours the regenerating cells (Fig.
9) are seen pushing outward and breaking through the blood clot
at the. exterior, to form a papillalike mass near the center of the
stump.

128 Victor E. Emmel.

Examined in section, the papilla is seen to consist of an evagina-
tion of regenerating epidermal cells. Toward the end of the third
day after the operation, the regenerating bud has increased consider-
ably in size (Fig. 10). A characteristic developmental phenomenon
is that the lateral or outer wall of the bud grows faster than the inner
or more mesial wall, so that, of the two sides, the outer wall becomes
both thicker and larger in extent of surface, and contains a greater
number of mitotic figures and nuclei. It is also on the outer or ven-
tral sides that the first appearance of joint formation becomes appar-
ent (Fig. 31). The result of this asymmetrical growth is that the
regenerating bud bends upward or inward toward the body of the
animal; a position which evidently reduces the chances for injury
through friction with external objects.

It is near the end of the third day, too, that the first differentiation
of limb segments becomes evident. <A slight groove appears near the
apex of the bud, and thus marks the first step in the development of
the two jaws of the claw. It is characteristic that this groove is
not located directly at the apex of the bud, but is somewhat ventral
to the tip, with the result that during the earlier stages of develop-
ment the dactyl is relatively larger than the opposing segment -or
index of the claw. The fact that this relation in the size of the
dactyl and the index becomes reversed in the fully regenerated claw, is
especially interesting since it is a process parallel with the series of
changes which occur in the normal ontogeny of the claw (Emmel
06). During the next six days the outlines of the dactylopodite
and propodite become more definite; the anlagen of all the seg-
ments appear, and eventually the four joints of the regenerating
limb become clearly defined (Fig. 31-36).

As each point develops, an ectodermal invagination arises in rela-
tion with it. This invagination, which more or less completely sur-
rounds the joint, soon develops, at two opposite sides of the joint,
into two proximally directed processes, which are destined to form
attachments for the flexor and extensor muscles. In the interior
of each invagination, there is secreted a lamella of chitin which is
readily demonstrated by its differential stain with Congo red.

During the development of the joints the epidermal cells present

Tissues in the Crustacean Limb. 129

characteristic structural changes. The mitotic figures occur at the
periphery of the bud, the spindle being almost invariably parallel
with the surface. A dividing nucleus is rarely found at a deeper
level. The cytoplasm is granular in appearance. No cell walls were
detected. In the vicinity of the mitotic figure, however, the mass of
cytoplasm immediately surrounding the dividing nucleus is less
granular and takes on a lighter stain.

Fig. 15 (five days and ten hours after operation) represents a
group of epidermal cells in the region of the invagination for the
flexor muscle of the fourth segment. The four nuclei at the right
in this figure are migrating inward in the process of invagination.
The three nuclei at the left are in the epidermal wall of the segment,
while the remaining nucleus, in the prophase (?) of mitosis, occupies
the typical superficial position. In these nuclei there may again be
seen a polarization of the nuclear material similar to that described
in the earlier stages of the regenerating cells. The dark staining
chromatic elements are found at the inner or more mesial pole of the
nucleus (with the exception of the nucleus undergoing mitosis). At
the same time the opposite pole of the nucleus takes only a lght
stain and evidently contains a much smaller amount of chromatin.
Here again the position of the plane of division between these light
and dark zones varies with the position of the nucleus. (In Fig. 15
the left and right parts are respectively distal and proximal with
reference to the limb.) The conditions odnerally found may be
stated as follows: In nuclei lying approximately transverse to the
long axis of the regenerating bud, the darker staining material is at
the inner end or pole of the nucleus. In nuclei not lying in a trans-
verse plane, as occurs in the region of joint formation, the chromatic
material, although still found in the inner region of the nucleus,
now becomes shifted either to the proximal or to the distal side of
the nucleus according as the inner pole of the nucleus is directed in
a distal or a proximal direction. The darker pole also frequently
contains a darker body or nucleolus, while at the lighter pole of the
nucleus there was usually found a reticular arrangement of fine
granular elements. The inner or darker pole of the nucleus was
generally found smaller and more tapering or pointed in form as

130 Victor E. Emmel.

compared with the more expanded outer peripheral region. Here
again the significance of this apparent polarization of the nuclear
contents is not clear, but, as was mentioned in the ease of migrating
nuclei, the general occurrence of these structural characters in dif-
ferent specimens and with different stains, and the variation of the
polarization of the elements in conformity with variations in the
position of the nuclei, hardly permits of the interpretation of this
phenomenon as a mere artifact.

As the invagination of the ectodermal cells at the joints advances,
the involved nuclei undergo further typical changes. Fig. 17 (seven
days, six hours) represents several nuclei taken at a later stage in
the development of the same invagination shown in Fig. 15 (five days,
ten hours). A marked difference is at once apparent in these two
groups of nuclei. In Fig. 17 the chromatin is evenly distributed,
and the nuclei have become elongated to such a degree that frequently
they are more than two and one-half times longer than at an earlier
stage. See Fig. 16 (six days, six hours) for intermediate stages.
It will also be observed in this latter figure that at the center of the
invaginating mass of cells, a thin lamella of chitin (mp) is now be-
coming evident, which later serves for muscle attachment.

VI. Origin or New CBrEtts.

We have already learned that the regenerative process is initiated
by epidermal cells, which migrate across the wound, form a layer
over the injured surface, then multiply by mitotic division and evag-
inate to form the new bud. It is evident, therefore, that the outer
layer of cells covering the regenerating limb bud is entirely of ecto-
dermal origin.

The evagination of the ectodermal cells encloses a cavity at the
center of the bud, which becomes filled with a core of cells. The
origin of this core of cells becomes at once a vital question in the
further study of the histogenesis. Since the interior of the fully
developed limb consists of such tissues as striated muscle and con-
nective tissue, it might be expected that this internal core of cells
would be derived from similar tissues in the old stump, and conse-
quently would be mesodermal in origin.

Tissues in the Crustacean Limb. Bai

A careful study of the successive stages of the regenerating bud,
however, does not warrant such a conclusion. Fig. 9 is typical of
the conditions found at early stages of the regenerative process. The
section is taken through the large limb nerve (n”) which is here seen
passing through a mass of connective tissue (ct). The regenerating
layer of ectodermal cells (ec) is increasing in thickness. Between it
and the connective tissue membrane enclosing the nerve trunk, a
small space has arisen in which may be observed several nuclei
lying in what seems to be a syncytial mass of cytoplasm. In their
form, structure, and reaction to stains, these nuclei resemble the
regenerating ectodermal nuclei. At certain points they are evidently
migrating directly from the ectodermal layer of cells; and ocea-
sionally nuclei are seen which seem to be migrating inward from the
edges of the old epidermis. On the other hand no evidence was
obtained indicating any proliferation of the old connective tissue
cells. Mitotic figures were not found, nor did there seem to be any
migration of the connective tissue cells into the space beneath the
evaginating ectodermal plate. Occasional blood cells, however, were
observed within this space.

Fig. 10 (two days, twenty-two hours) is a section of a regenerating
bud in which the invagination for the two jaws of the claw has begun.
The central core of cells is now considerably larger. A few blood
corpuscles are present in the proximal region of the core, but the
nuclei of the central mass of cells still resemble the ectodermal
nuclei. A slight migration of ectodermal cells may be observed
from the sides of the limb bud, but it is in the region of the invagina-
tion for the first joint that the migration is most extensive.
From this advancing invagination the ectodermal cells migrate in
large numbers, and thus fill the central cavity of the regenerat:
ing bud. In this central mass of cells mitotic figures were very
rarely observed. As other limb joints are formed, a similar migra-
tion of ectodermal cells was found to occur at the invagination for
each limb segment. No evidence was found indicating the possible
derivation of these internal cells from the underlying connective
tissue of the old stump; if it does occur, it seems evident that it
cannot be to any large degree. This conclusion is based upon (1) the

132 Victor E. Emmel. |

lack of evidence of cell division in the connective tissue, and (2)
the fact that at no stage of development did there appear to be a
migration of connective tissue cells into the regenerating bud.

In a word, the results of these observations lead to the unexpected
conclusion that certainly the greater part, and probably the entire
mass, of cells composing the interior as well as the wall of the regen-
erating limb bud, are ectodermal in origin.

VIL. Tur DirFERENTIATION OF TISSUES.

1. Striated Muscle—a. Histogenesis.—The first differentiation ~
of striated muscle, as indicated by the formation of myofibrillee,
became apparent as early as five days and twenty-two hours after
amputation. On the sixth day the fibrillee were readily distinguished.
At the center of a longitudinal section through the second segment or
propodite (Fig. 16, six days, six hours) may be seen the tongue of
ectodermal cells invaginated for the flexor muscle of the dactyl. The

space between the epidermal cells (e) of the regenerating exoskele-
ton, seen at the lower side of the figure, and the invagination, 1s filled
by a mass of cells, the ectodermal origin of which has already been
considered. It is within this central mass of cells that the myofibrillee
(mf) first appear. These fibrillee, instead of extending as straight
fibers between their origin in the invagination and their insertion in
the epidermal wall, are considerably curved, being convex in a proxi-
mal direction. This curvature of the myofibrils does not entirely
disappear until after the moult of the lobster has occurred and the
regenerated limb has become functional. It may be justly questioned
whether there is any correlation between the appearance of these fibrils
and functional activity, such, for example, as Eycleshymer (’05)
describes in the Necturus embryo, for there is no evidence of func-
tional activity in the regenerated limb until it has been liberated
from its enveloping membrane during moulting.

The cytoplasm in which the myofibrils are first seen to differentiate
(Fig. 22) appears finely granular in structure and is traversed by a
delicate cytoplasmic reticulum (7t). As soon as the fibrillee can be
distinguished from the cyto-reticulum they appear as heavier fibers

Tissues in the Crustacean Limb. 133

(mf) taking a darker stain with Congo red. The very earliest fibrils
are, however, not so readily identified on account of their similarity
to the cytoplasmic network and their close relations with it; for at
the time of their first differeniation the fibrille stain but slightly with
Congo red, are irregular or wavy in structure, and appear to be in
continuity with the cyto-reticulum. These characteristics indicate
that the contractile elements in the regenerating limb of the lobster
may be derived trom the cyto-reticulum, and consequently favor the
“network” rather than the “fibrille” theory for the origin of striated
muscle fibers. It should be added, however, that no such constancy
in relation was observed between the cytoplasmic network and the
fibrille, as would seem to be necessitated by MacCallum’s (’98)
theory for striated muscle of vertebrates.

As late as the seventh day of regeneration each myofibril (Fig. 23)
still retains its individuality as a single structure. During the eighth
day, the fibrils began to appear double or in pairs. In Fig. 24 one
of the fibrils is still a single structure, but the remaining eight fibrils
are in pairs, the members of each pair appearing in cross section as
half cylinders. At later stages of differentiation each pair of
fibrillee becomes represented by a group of four fibrils; the number
in each group then increases until as many as twenty fibrille could
be counted in each bundle, which in transverse section may now be
recognized as a “Cohnheim’s area”. As to how the fibrillee multiply,
the evidence from appearances in both cross and longitudinal sections
of the regenerating muscle in this invertebrate supports the conclu-
sions of Heidenhain, Eycleshymer, and other investigators for the
striated muscle of vertebrates,—that an increase in the number of
myofibrillz arises through longitudinal division of the fibril.

Preceding the first division or splitting there is a marked increase
in the diameter of each fibril (compare Figs. 22 and 23). At the
same time the cytoplasm immediately surrounding the fibril becomes
less granular and is distinguished from the neighboring cytoplasm
by its lighter stain (Figs. 23 and 24). The developing fibril is
consequently enclosed by a sheath of modified cytoplasm, which is
evidently correctly interpreted as representing the beginning of a
Cohnheim area of the mature muscle fiber.

134 Victor E. Emmel.

Reference has just been made to the fact that each myofibril
retains its individuality as a single structure until the seventh day of
regeneration. At this time the light and dark bands of the myofi-
brill: are already present, but the ‘‘Z” line, or membrane of Krause,
could not be observed. By the eighth day the ‘‘Z” line had become
differentiated, the first indication of these lines being found during the
early part of the seventh day. It appears, therefore, that the “Z”
lines differentiate later than the light and dark bands. In this
respect the differentiation of the regenerating crustacean muscle
resembles the histogenesis of the striated muscle as found in the
21 mm. pig embryo, where, as described by Bardeen (700, p. 392),
“The fibril bundles are composed of longitudinal fibrils made up
of longer deeply staining segments alternating with shorter lighter
segments which do not stain. It is only in older embryos that lines
through the light areas, corresponding to Krause’s membrane, may
be clearly distinguished.”

The problem of the origin of the multinucleated muscle cell and its
sarcolemma in the regenerating tissue of the lobster differs from the
same problem in the vertebrates, because in the former there appar-
ently are no myoblasts concerned in muscle formation.

The structure in the adult muscle of the lobster, which is known
as the sarcolemma, is a membrane surrounding the muscle fiber and
containing elongated nuclei. This membrane is usually regarded
as developed ontogenetically from connective tissue elements. No
evidence, however, was obtained for a similar origin during regenera-
tion. When the myofibrille first appear in regeneration, definite
cell membranes cannot be identified, but the sarcolemma first appears
in a later stage of differentiation (Fig. 25, sa), and seems to arise
by a modification of the cytoplasm of the ectodermal cells.

During the differentiation of the myofibrille (which are at first
placed centrally and later become located eccentrically in the muscle
fiber), the nuclei undergo certain characteristic changes in form and
structure. The elongation of the ectodermal nuclei, as they pro-
liferate and migrate inward at the joint, has already been described.
As the migration advances, the nuclei in the region where the myo-
fibrils differentiate, assume a peripheral position in the developing

Tissues in the Crustacean Limb. 135

muscle fiber and become more spherical in form (Figs. 16, 22, 24,
and 25). These nuclei now seem somewhat larger than the earler
ectodermal nuclei, an appearance which may be partly due to the
change from an elongated to a rounded shape. The chromatin ap-
pears coarsely granular and has a fairly even distribution, the nucleus
as a whole taking a lighter stain. In later differentiation the nuclei
again become flattened and considerably elongated in the direction
of the long axis of the muscle fiber. The present observation indi-
cates that the more peripheral of these nuclei eventually become the
nuclei of the sarcolemma, but more evidence is necessary to establish
this point.

In the formation of muscle fibers from a syncytium of ectodermal
cells, each fiber is from the very first multinucleated. Instead of
“each muscle bundle developing from a single cell” (Claus, ’86, p.
33), as seems to be the case in the normal development in Branchipus,
for example, each bundle is multicellular in origin. Furthermore,
the nuclei increase in number, with the growth of the fiber, by mitotic
division, the spindle frequently occurring at right angles to the long
axis of the fiber. Consequently in regard to the discussed question
of direct and indirect division of muscle nuélei, it is evident that in
the regenerating muscle of the lobster at least, mitotic division per-
sists even after the differentiation of myofibrille has advanced to a
considerable degree.

The conclusion that regenerating striated muscle is ectodermal in
origin, draws attention to the problem of the genetic relationship
of tissues and the primary germ layers, the question at issue being
whether in regeneration a given tissue arises from the same germ
layer as in normal development. An adequate discussion of this
question in the case of the crustacean muscle, involves, of course,
an accurate knowledge of the histogenesis of the tissue under con-
sideration, in both the regenerative and ontogenetic processes.

With regard to regeneration, as a result of the work of Reed
(704) on the oerayfish, Ost (’06) on Oniscus, and the present study
of the lobster, we have at hand a fairly exact mass of data concern-
ing the genesis of the regenerating crustacean muscle, but as for its
origin during normal ontogeny, the evidence does not appear suffi-

136 Victor E. Emmel.

ciently conclusive. In the ontogeny of vertebrates, the striated mus-
culature is evidently mesodermal in origin, but whether the same
is true in crustacean development, especially in the case of the limb
musculature, is not, so far as the writer is aware, clearly established.
Consequently it seems obvious that we are not as yet justified in
concluding that the ectodermal origin of the regenerating crustacean
muscle is a divergence from ontogenetic development. Indeed, as
Ost (06, p. 312) points out, it is not impossible ‘‘dass die embryo-
nalen Muskeln von den ectodermalen Sehneneinstiilpungen aus-
sprossten und dann Regeneration und Embryonalentwicklung sogar
iibereinstimmten,” a conclusion which is certainly not rendered less
improbable when one considers the degree to which the generally
accepted conclusions regarding genetic relationships between ecto-
derm, mesoderm, and certain forms of connective and muscle tissue
have been called into question by the work of such investigators as
Katschenko (88), Kolliker (’84), and Platt (798).

b. Attachment to the Exoskeleton.—Among the first investigations
on the attachment of the crustacean muscle is Claus’s (?86) work
on Branchipus and Artemia. Claus concludes that in many eases
the muscle fibers are attached directly to the exoskeleton. (pp. 22 and
29), a conclusion which involves the assumption that among some
invertebrates at least the muscle may be attached to the skeleton
without the intervention of a connective tissue tendon, such as are
typical of muscle-skeletal attachments among vertebrates. . Since
1886 the subject has received considerable attention both for erusta-
cea and insects, with the result that several divergent opinions have
arisen. As far as the writer is aware, however, the problem has not
been studied from the standpoint of regeneration; consequently data
derived from the present investigation of the lobster may not be
valueless.

Attention has already been directed to the fact that in the early
stages of the regenerating bud and up to the time when the muscle
fibrils have become well differentiated, there is no definite boundary
between the outer epithelial cells related to the chitin and the internal
cell mass in which muscle develops. Cell walls are not evident, and
the cytoplasm of the two regions appears syneytially related, the

Tissues in the Crustacean Limb. 137

only noticeable difference being that the epidermal cytoplasm next to
the chitin takes a slightly darker stain (Fig. 16). The question
now arises, what is the relation of the developing myofibrille to the
external epidermal cells and how is their attachment to the chitin
established ¢

In studying the regenerating tissue at the stage when the myo-
fibrille can just be identified (Fig. 16, six days, six hours), an im-
portant fact becomes evident, viz., that fibrils develop in the epider-
mal layer of cells, simultaneously with the differentiation of the
myofibrille in the internal cells, apparently without any discon-
tinuity between them. In other words in early differentiation the
myofibrilla can be traced into the layer of epidermal cells, and in
some cases at even this early stage (six days, six hours) myofibrils
were found continuous even through to the chitin.

The differentiating muscle fibrils early assume a characteristic
striation (Fig. 18, mf). At the same time more clearly defined
differences are becoming evident between the epidermal and muscle
cells. The muscle nuclei are more nearly spherical in form, while
the epidermal cells take a distinctly heavier stain. Small vacuoles can
also be observed at the boundary between the two groups of cells.
The striation of the muscle fibrils gradually becomes more distinct.
These striations could be traced beneath the inner surface of the
epidermis, frequently extending to the level of the proximal ends of
the nuclei (Fig. 18). No evidence was obtained, however, that the
fibrillee are ever striated entirely through the epidermis to the chitin.

A definite boundary (basement membrane) now becomes apparent
between the epidermal and muscle cells (Fig. 19, em). There is an
interesting variation in the general level of the boundary or inner
surface of the epidermis in the region of muscle attachment. The
epidermis between the attachment of two or more muscles, frequently
projects inward in rounded elevations or broad papilla. The result
is that in the immediate vicinity of the muscle attachment the inner
surface of the epidermis appears to follow the fiber for some distance
toward the chitin, thus forming a sort of pit, through the bottom of
which passes the muscle fiber (Figs. 19 and 20). Since this eleva-
tion of the epidermis increases in the later stages of the regenerating

138 Victor E. Emmel.

limb, it is probably due to the rapid growth of the epidermal cells,
which are consequently forced inward between the muscle fibers.
After the moult, the regenerating limb expands and the elevations
then disappear as the epidermis spreads out to cover the now larger
surface of the limb.

The muscle fibers appear to be continuous throughout the epi-
dermis to the chitin (Figs. 18, 19, and 20). Frequently just before
the fibers reach the chitin they spread out in a brush-like manner
and fuse with the chitin (Fig. 20). In this fusion the fibrils fre-
quently terminate on knob-like thickenings or inward projections of
the chitin (Fig. 19). These chitinogenous processes were found
only in connection with the attachment of the regenerating muscle
fibers.

In later stages of differentiation, the epidermal nuclei adjacent
to the muscle attachment frequently become considerably lengthened
in the direction of the long axis of the muscle fiber. In addition
to the fibrils concerned with muscle attachment, there are other fibrils
which later differentiate in the cytoplasm of the epidermal cells (Fig.
19, ef). These latter fibrils do not appear to be associated with
muscle attachment. In structure they are much finer and are dis-
tinguished from muscle fibrils by their stain reaction. With Mal-
lory’s connective tissue stain they appear light blue, whereas the
fibrils concerned with muscle attachment take a dark red or purple
stain similar to the striated part of the myofibril.

On the inner surface of the fully developed epidermis of the
crustacean there occurs a very thin layer or lamella of tissue which
has been termed the ‘‘Grenzlamelle” (Schneider). The fact that a
flat nucleus is occasionally found in this “border lamella” has
favored the interpretation that it is a connective tissue formation
and consequently mesodermal in origin. Other investigators (Claus,
for example) have regarded this layer or “basement membrane,”
as merely a differentiation of the inner surface of the epidermal
cells. In regard to this question it is clear that in the early regenera-
tion of the lobster’s limb there is no definite hmiting membrane be-
tween the epidermal and muscle cells. The first limiting membrane
to appear during the differentiation of the epidermal cells is evidently

Tissues in the Crustacean Limb. 139

epidermal in origin and to that extent is ectodermal. This earlier
membrane seems similar to the basement membrane which MeMur-
rich (796) describes on the epithelium of the mid gut of isopods, and
which he coneludes ‘‘was formed from the epithelial cells and not
by the mesoderm” (p. 89). It is to be observed, however, that at
a later stage of development (Fig. 21) a thin nucleated layer does
appear on the inner surface of the epidermis, which is perhaps to
be regarded as the true “Grenzlamelle.” The origin of this latter
layer has not been satisfactorily determined. It was noticed that
it is first apparent in the region of the developing blood sinuses,
where it appears to serve as a vascular epithelium.

c. Discussion and Conelusions.—The study of the attachment of
the arthropod muscle is attended with considerable technical diffi-
eulty. That the question is still an open one is indicated by the
diverse opinions existing at the present time. The chief points
at issue are involved in the solution of the questions whether the
muscle fibers are attached (1) directly to the chitin of the exoskeleton,
or (2) indirectly by means of intervening epidermal cells; and in the
latter case, whether (3) in addition to the epidermal cells there is also
an intervention of connective tissue between the muscle fibers and the
epidermis, in a manner somewhat analogous to the muscle attachment
typical among vertebrates.

The literature upon this subject is becoming very extensive, espe-
cially for insects, where the question of muscle attachment has been
studied by Henneguy (’06), Holmgren (’02), Lecaillon (’07), Riley
(08), Snethlage (’05),° and others. Without reviewing this grow-
ing literature, we may advantageously confine our discussion to the
present observations on the lobster and their bearing on the problem
among crustacea.

The third of the different modes of muscle attachment just con-
sidered involves the union of three sets of fibrils,—the fibrils of the
muscle, of connective tissue, and of the epidermal cells. Schneider
(702) is inclined to regard this method of attachment as typical for
crustacea, and his description of the jaw muscle of the crayfish states

*Unfortunately, I have been able to have access only to abstracts of Sneth-
lage’s work, “Ueber die Frage vom Muskelansatze, ete., bei den Arthropoden.”

140 Victor E. Emmel.

that “Das Bindegewebe ist an den Muskelenden als typisches Faser-
gewebe entwickelt, das sich vom faserigen Zellengewebe durch reich-
liche Entwicklung extracellulirer fibrillirer Bindesubstanz, im Um-
kreis spindeliger Bindezellen scharf unterscheidet. Die Myofibrillen
einerseits und die Deckzellenfibrillen andererseits senken sich in eine
dicke Lamelle ein, in der Bindefibrillen in dichter Anordung, von
sparlicher Grundsubstanz verkittet, entsprechend den Myo- und Stiitz-
fibrillen verlaufen” (p. 494). On the other hand, Claus (’86) in
his study of Branchipus finds that ‘‘An vielen Stellen heften sich
aber die Muskelsehnen nicht direct mittelst Connectivfaden am Inte-
gument an, sondern gehen in einer durch solche suspendirte Lamelle.
iiber, welche sich der Integumentflache parallel als Basalplatte unter-
halb jener ausbreitet” (p. 29).

The critical question here is in regard to the continuity of the
muscle fibers within the epidermal cells. If the muscle fibers can
be traced below the level of the inner surface of the epidermis, inde-
pendent of any connective tissue elements, evidently the third mode
of attachment may be eliminated from further consideration. That
such is certainly the case in the regenerating musculature of the
lobster’s limb has already been indicated, and the crucial fact remains
that this relation of the myofibrille was already evident at a stage
of development before there was any differentiation of connective
tissue elements or of a ‘““Grenzlamelle.” Consequently, at this time
at least, the muscle fibrils are found passing into the epidermis unac-
companied by any connective tissue elements. The apparently close
relation, which is later established between the muscle fibers and the
connective tissue, may perhaps be not incorrectly regarded as a sec-
ondary development resulting from a later differentiation of connec-
tive tissue around the already formed muscle fibrils, rather than as
a primary functional relation. It is of interest to observe that
even in the fully developed lobster, as admitted by Dahlgren and
Kepner (’08), the connective tissue elements are in some places “so
small as to be apparently absent, and it would seem possible that in
some attachments they were absent altogether and the muscle joined
directly with the epithelium” (p. 66).

Jn considering the twe remaining modes of attachment, certain

Tissues in the Crustacean Limb. 141

aspects of the problem, as they present themselves in other animal
forms, disappear in approaching the subject from the standpoint of
regeneration. For example, the question whether the fibrils by which
muscle’ attachment is accomplished are inter- or intra-cellular with
reference to the epidermal cells, loses much of its significance here
on account of the syncytial structure of the cytoplasm. Genetically,
there is perhaps no distinction to be drawn between the purely epider-
mal fibrils and the myofibrille, for if the conclusions regarding the
origin of the muscle are correct, both groups of fibrils are derivatives
of the cytoplasm of epidermal cells, and are consequently also both
ectodermal. Structurally, however, they have diverged in their
differentiation, in conformity with their differences in function. The
striation of the muscle fibril can frequently be traced for some dis-
tance into the epidermal cytoplasm, but it was not observed that the
striation of the fibrils ever continues through to the chitin; it appears
that the peripheral ends of the original myofibrille diffentiate as
tensile rather than as contractile structures. But even the non-
contractile elements thus directly concerned with the attachment of
the muscle to the chitin can be distinguished from the purely sup-
portive fibrils of the epidermal cells by (a) their staining reaction
with Congo red and Mallory’s stain; (b) their frequently larger size ;
(c) their evident continuity with the contractile part of the muscle
fiber; and (d) their relations with the exoskeleton where they ter-
minate in characteristic end-plates or chitinogenous processes.

The conclusions which these observations support may, therefore,
be summarized as follows:

1. The myofibrillz differentiate in the cytoplasm of a syncytial
mass of cells, derived from the epidermis and consequently ecto-
dermal in origin.

2. The original fibrille ultimately differentiate throughout their
whole length into true striated muscle elements, except in the region
of skeletal attachment, where the peripheral ends of the fibrils remain
unstriated, and serve as tensile structures directly uniting the con-
tractile elements with the chitin of the exoskeleton.

3. In later development connective tissue elements may form over
the inner surface of the epidermis and around the muscle fibers, and

142 Victor E. Emmel.

purely supportive fibrils differentiate within the epidermal cells,
but these structures are secondary in their relation to muscle
attachment.

2. Nerve Fibers.—In studying the regeneration of nerve fibers
we are at once involved in the intricate questions of the origin of
the neurilemma, and of the axis cylinder with its neurofibrille.
Complete agreement regarding ectodermal or mesodermal origin of
these structures has not yet been attained. Regarding the neuri-
lemma, for example, the conclusions of one group of investigators
may be summarized in the statement that sheath cells “are true con-
nective tissue cells from the locality through which the nerve fiber
has passed in its development” (Dahlgren and Kepner, ’08, p. 188).
On the other hand, the ectodermal origin of the sheath cells appears
to be strongly supported by experiments in which it has been shown
that in the tadpole at least ‘the source of the sheath cells, both of the
motor and sensory nerves is in the ganglion crest’? (Harrison, 08,
p- 393). Although the present observations on the lobster are, on
account of a lack of material especially prepared for neurological
study, not sufficiently extensive to render them very important in
relation to these fundamental problems, still they may be of some
value.

In early stages of the regenerating limb bud there is found a cord
or column of cells extending from the tip of the bud to the end of
the old nerve trunk. As the limb segments are formed, this cord of
cells is joined by similar strands from the various segments. Within
these cords the axis cylinders of the nerve fibers develop, and from the
cells composing the cord the sheath cells are differentiated. The
important question is as to the origin of these cords of cells; have
they migrated outward from the sheath cells of the old nerve trunk, or
are they derived from the ectodermal cells of the regenerating bud ?

The present observations support the latter conclusion. Among
the old sheath cells no evidence of cell division was obtained,
nor was there observed any extensive proliferation or outward migra-
tion of the sheath cells from the old nerve trunk. On the other
hand, cells from the first-formed ectodermal plate migrate inward
toward the injured nerve, as has already been described in the early

Tissues in the Crustacean Limb. 143

stages of the regenerating bud (Fig. 9). At later stages this inward
migration of ectodermal cells is quite marked in certain regions
(Fig. 28, n), where they evidently contribute to the formation of
the cords of cells just described. The distal end of each cord
always contains a larger number of cells than its more proximal end,
giving it a club shape, with the smaller end joined to the old nerve.
Occasionally mitotic figures were found, but they always occurred
among the more distal eclls of the cords. The nuclei of these distal
cells were generally large and spherical in form, but in passing proxi-
mally toward the old nerve trunk, there was a gradual transition
from the spherical nuclei to the long flat nuclei of the sheath cells,
indicating a direct differentiation of the neurilemma from these ecto-
dermal cells.

These observations on the lobster, together with the results of
Reed’s (’04) on the crayfish, and Steele’s (’07) on the nerve endings
in the ommatidia of the eye, furnish collective evidence for the par-
ticipation of ectodermal cells in the regeneration of the nerve fibers
in crustacea. It remains to be determined, however, whether this
conclusion can be extended to arthropods in general. For it must
not be overlooked that different results have been obtained by Ost
(706) in his study of the regenerating antenna of Oniscus, in which
form he finds that the neurilemma regenerates ‘‘durch Nachschieben
vom proximalen Ende her . . . Die Regeneration des Antennen-
nerven von Oniscus geht also nach meinen Beobachtungen durch
direktes Auswachsen junger Nervenfasern aus dem alten Stumpf vor
sich” (p. 313). It is noteworthy that Ost was unable to obtain
any evidence of cell division among the sheath cells, for he states,
“Mitosen oder sonstige Teilungsvorginge konnte ich an diesen Ker-
nen freilich nie beobachten,”’—a result corresponding with the con-
ditions found in the regenerating lobster’s limb, 7. e., in the region of
the old nerve trunk.

By the fifth day of regeneration (five days, ten hours) the differ-
entiation of an axis cylinder within the cord of cells just described,
had advanced sufficiently to present delicate neurofibrille.® These

‘The nerve fiber thus appears to develop earlier than the muscle, for in
all cases regenerating nerve fibers were found before myofibrille could be
detected.

144. Victor E. Emmel.

fibrillee were not of equal size and were distributed in an interesting
manner. In some of the preparations, the larger or heavier fibrils
were mostly at the periphery of the axis cylinder, whereas the finer
ones were nearer the center (Fig. 29). Consequently at certain
stages of regeneration the nerve fiber could be structurally analyzed
into four parts: (a) a central core of very delicate fibrille; (b) a
peripheral layer, not sharply marked off from (a) but containing
heavier fibrils; (c) the external sheath of nucleated cells or future
neurilemmma, and (d) a sheath of cytoplasm between (b) and (ce)
relatively free from neurofibrille. In later stages of differentiation
the heavier fibrils predominate (Fig. 30). At the time when the
neurofibrille are beginning to appear, the cytoplasm of fhe nucleated
sheath, as compared with that of the axis cylinder, is more coarsely
granular. In some regions there also appears to be a membrane
present between the nucleated sheath and axis cylinder.

No conclusive evidence was obtained as to whether the axis cylin-
der is formed in situ from the sheath cells, or whether it is an out-
growth from the axis cylinder of the old nerve cell. In either case,
however, if the present observation is correct, that the nerve fiber
develops within a cord of cells proliferated from the ectoderm of the
various new limb segments, it points to the sheath cells as one of the
factors involved in determining the final distribution of nerve fibers
within the regenerated limb. It was not determined whether the
neurofibrillee differentiate distally or proximally, or whether they
appear simultaneously throughout the regenerating nerve fiber. The
fact that in at least certain stages of differentiation the finer fibrille
occur at he center, and the heavier fibrils at the periphery of the fiber,
apparently lends support to the conclusion that the neurofibrille are
being derived from the cytoplasm of the axis cylinder.

3. Connective Tissuc.—The connective or supporting tissue of the
crustacean limb consists mostly of broad loose bands or sheets ex-
tending between opposite walls of the exoskeleton, around the ex-
tremities of each segment, and in certain regions filling almost the
entire segment. In addition to these sheets and bands of connective
tissue there is also a thin layer of simple flat epithelium found cover-
ing the inner surface of the epidermal cells (here known as “Grenz-
lamelle”) and surrounding the muscle bundles.

Tissues in the Crustacean Limb. 145

Let us consider first the broad sheets of tissue extending from one
wall to the other within the limb segment. As the segments of the
regenerating limb become well differentiated, there occurs an exten-
sive inward migration of ectodermal cells in those regions where there
later develop the bands of connective tissue characteristic of the adult
limb. This migration is especially well defined in the regenerating
meropodite. The epidermal cells in this region migrate inward from
the epidermal wall and form a broad sheet extending across the central
cavity of the segment (Fig. 26, 7c). The nuclei become elongated,
and distinct fibrils arise in the cytoplasm. At later stages (Fig. 27,
eleven days, ten hours) the cytoplasm, with the exception of the
epidermal cells next the exoskeleton, becomes vacuolated. These inter-
cellular spaces then become filled with blood plasm (Fig. 27, 5),
and eventually there is thus formed the vascular and connective
tissue characteristic of the fully developed limb.

It seems evident, therefore, that in at least certain regions of
the regenerating limb a supportive and apparently true connective
tissue may be derived from ectodermal cells. Nor is this divergent
from conditions found in normal development among crustacea, for
Claus (’86) in his work on the embryology of Branchipus, concludes
that the broad connective tissue bands which unite the opposite sur-
faces of the integument, “sind Erzeugnisse der Chitinogenzellen der
Hypodermis”’, and quotes Braun to the effect that in the crayfish
it is “nicht méglich, eine scharfe Grenze zwischen den Erzeugnissen
von Chitinogenzellen und den mesodermalen Bindegewebebildungen
festzustellen” (pp. 22-23).

We may now briefly consider the tissue characteristics of the so-
called “Grenzlamelle.”” In the description of the regenerating mus-
cle, reference has been made to the fact that a “Grenzlamelle” is
not present under the epidermis at the time when myofibrille begin
to differentiate. It is only after the muscle fibers and the epidermis
have become well developed that an epithelial layer is formed over
the inner surface of the epidermal cells. In various parts of the
limb this “Grenzlamelle” is apparently the only tissue between the
blood sinuses and the epidermis, and consequently in at least certain
regions it seems to serve as a vascular epithelium (cf. Williams, ’07,
p. 155).

146 Victor E. Emmel.

As to the origin of the regenerating ‘‘Grenzlamelle,” the present
observations fail to furnish conclusive evidence. The fact, how-
ever, that the regenerating bud becomes filled with ectodermal cells
involved in the regeneration of muscles and nerves, and the lack of
clear evidence of cell proliferation from the old connective tissue,
suggests an ectodermal origin. Of interest in this connection are
Claus’s observations on the normal development of Branchipus :—
“Was man als solche (connective tissue) beim ersten Blick in An-
spruch zu nehmen geneigt ist, erweist sich wenigstens an vielen
Stellen nach genauer Betrachtung als eine Art innere Cuticula, die
aus der Basis des Epithels durch Erhartung des Protoplasmus ent-
standen ist” (p. 21). On the other hand, Schneider (’02) from a
comparative point of view is unable to support this conclusion: “Nach
Claus stammen die Grenzlamellen und die Muskelsehnen mindestens
zum Teil vom Epiderm, dessen Deckzellen die basalen ausscheiden
sollen. Dieser Ansicht kann in Riicksicht auf die Verhialtnisse bei
anderen Arthropoden, wo die Sehnen und Lamellen unzweifelhaft
Bindegewebsbildungen sind, nicht beigestimmt werden, wenn auch

die Bildung von Seiten des Bindegewebes nicht bekannt ist” (p.
466).

VIII. Tue Direction or DIFFERENTIATION.

The question of the direction in which the process of differentia-
tion proceeds in both regeneration and normal ontogeny is a subject
which has given rise to diverse opinions. In elaborating certain
theories of form regulation, Holmes (’04) assumed that in a regen-
erating appendage, differentiation proceeds from the base toward
the tip, whereas Child, at least in an earlier paper (’06), was in-
clined to regard the reverse as generally true.’?. Zeleny (’07) found
in the regenerating antennule of Mancasellus that there are two
distinct periods of development, in the first of which the process of
differentiation “begins at the base and travels irregularly outward”,
while in the second period, the process is in the reverse order, 2. ¢.,
beginning with the terminal segment and differentiating inwards (p.

"Holmes (’07) and Child (’08) have later restated their theories as not
being inconsistent with differentiation in either direction.

Tissues in the Crustacean Limb. 147

325). Haseman (’07) concluded that in the regenerating cheliped
of the crayfish, differentiation proceeded in a disto-proximal direc-
tion. Haseman, appreciating the importance of determining whether
changes in external form could be safely taken as an index of internal
changes, gave some attention to the internal differentiation of tissues.
But as a rule conclusions upon this subject have been largely based
upon evidence derived from modifications in external structure,
rather than from internal cytological and histological changes in the
segment tissues. In view of this fact, the following study has been
made with the purpose of obtaining further data upon the direction of
differentiation among the internal regenerating tissues.

The results to be described were derived from the study of serial
sections of somewhat over a hundred regenerating chele at various
stages of development. In tracing the progress of differentiation
it becomes necessary to rely on such criteria as are most clearly
defined. In the present case the following structural characters
have been chosen on account of their ready identification by means
of selective stains: (1) the formation of chitinogenous muscle plates
or tendons; (2) the differentiation of myofibrille; and (8) the
striation of the muscle fibrils.

In the description of the differentiation of the regenerating limb,
it will be recalled that distal to the breaking joint there are five
segments. Kach segment contains a pair of muscles controlling the
extensor and flexor movements of the next distal segment (see Fig.
37). It is important to note that in each pair of muscles the flexor is
the larger, and that of the four pairs of muscles, it is the first and third
pairs (counting from the distal end) which contain the largest muscles,
7. €., the muscles which lie in the propodite and meropodite. Each
muscle takes its origin from the exoskeletal wall of the segment in
which it lies, and is inserted upon the chitinogenous plate extending
proximally from the next distal segment. The differentiation of these
chitinogenous plates will first be considered.

1. The Chitinogenous Plates or Muscle Tendons.—The differen-
tiation of chitin within the joint invagination is indicated by a char-
acteristic brick-red stain reaction with Congo red, or bright blue with
Mallory’s connective tissue stain. ‘The developing chitin is thus

148 Victor E. Emmel.

sharply contrasted with any surrounding tissue. Using this stain
reaction as an index of differentiation, serial sections were studied
and graphic reconstructions made of different stages in the regenera-
tion of the limb (Figs. 31-36).

a. Two Days and Twenty-two Hours after Operation (fig. 31).—
Although externally at this time there was little, if any, indication
of segmentation, internally there was found a well defined invagina-
tion of epidermal cells, within which a differentiation of chitin (7)
was already evident. This invagination marks the beginning of the
first and second distal segments. It is here that the chitin plate is
formed for the flexor muscle of the dactyl, which, it is to be observed,
is normally not only much larger than the opposing extensor, but
is also the largest muscle of the limb.*

b. Four Days and Sia Hours (ig. 32).—The differentiation of
additional chitin plates is not evident until on the fourth day of
regeneration. At this time two more invaginations were beginning
to develop chitin. Of these two invaginations the one representing
the extensor for the dactyl (e') showed as yet but a slight differen-
tiation of this tissue. The other invagination (7°) represents the
flexor for the meropodite, and shows a well-defined plate of chitin.

In regard to the latter invagination (7?) there are two important
points to be emphasized. - First, the invagination is not in the next
or third proximal segment, but in the fourth segment or meropodite ;
and second, of the two muscles in the meropodite, the invagination
represents the flexor.®

‘This invagination occurs at one side of the apex of the bud, so that at
first the dactyl is relatively Jarger than the index,—a relation, it is interesting
to note, which corresponds with the condition found in the larval stages of
development, but is reversed in the adult. (Hmmel, ’06*).

°In the external segmentation of the limb at this stage, the groove between
the second and third segments is quite as well marked, if not more so, than
the groove between the third and fourth segments. This agrees with Haseman’s
(07) observations on the cheliped, and with my own earlier description (’06)
for the lobster, that the external segmentation appears to proceed in a disto-
proximal direction; but the crucial fact that in the present case it is the
tendon in the fourth segment instead of in the third segment which differ-
entiates next after the second segment, demonstrates that the external seg-
mentation cannot, therefore, be safely relied upon alone as a correct index
of internal differentiation.

Tissues in the Crustacean Limb. 149

The first point becomes especially interesting when it is considered
that according to the requirements of the theory that differentiation
proceeds in a disto-proximal direction, the next chitin muscle plate
to develop after those of the propodite should have been in the third
segment or carpopodite instead of the fourth segment or meropodite.
Serial sections of the carpopodite were carefully examined, but there
was no evidence that any chitinization had yet taken place for the
tendon plates in this segment.

c. Hour Days and Twenty-two Hours (Fig. 33).—At this stage a
fourth chitinous tendon (77) has become evident. This tendon is
in the third segment and is the one involved in the development of
the tlexor muscle. The extensor (e') and flexor (r!) in the propo-
dite, and the flexor (7*) in the meropodite, show an advance in differ-
entiation as compared with the preceding stage, but otherwise there
is no evidence of chitin differentiation \in any of the remaining
muscles. Consequently at this stage only four chitinogenous plates
or tendons are present; two in the propodite and one in each of the
next two proximal segments. The important point is that in each
case it is the tendon for the flexor muscle which differentiates first
in each segment.

d. Five Days and Six Hours (Fig. 34).—The tendon for a second
extensor muscle (¢*) is now beginning to differentiate. Here again
the next tendon to develop is not for the extensor in the carpopodite,
but for the one in the meropodite. At this time two other chitinous
tendons (e*) are also just becoming evident in the fifth segment or
ischiopodite. The muscles correlated with these last two tendons, it
will be recalled, are both extensors.

e. Live Days and Nine Hours (Fig. 35).—The chitinous tendon
(e*) for the extensor of the carpopodite is now present. This intro-
duces, therefore, the last of the eight muscle tendons to differentiate.

f. Len Days and Two Hours (ig. 36).—By this time the eight
chitinogenous tendons and muscles of the regenerating limb have
become fully developed. The animal is now about to undergo the
process of moulting, in which the regenerated limb will be liberated
from its membranous sac and become a functional appendage.

In summarizing these observations it may be stated that the chitin-

150 Victor E. Emmel.

ogenous plates or tendons for each of the eight muscles distal of
the breaking joint were found to be differentiated in the following
order: (1) for the flexor muscle in the second limb segment; (2) the
flexor in the third segment, together with the extensor in the second
segment; (3) the flexor in the third segment, and at the same time
the two extensors in the fifth segment; (4) and last, the extensor in
the third segment. From these results it seems evident that the
direction of differentiation cannot be described as being any more a
disto-proximal one than the reverse. The important point disclosed
is that in each segment the tendon for the flexor, or larger muscle
of the pair, is differentiated first; and further, of these flexors the
larger ones are the first to develop.

2. Differentiation of Myofibrilla.—The results of the microscop-
ical study of the earliest differentiation of the myofibrille in each
pair of regenerating muscles are presented in the following table. In
this table the first column gives the time of regeneration. The signs
++ and 0 indicate, respectively, the presence or absence of myofibrils.

TABLE SHOWING THE SEQUENCE OF DIFFERENTIATION OF MYOFIBRILLAE IN HACH
OF THE EHicHTt LIMB MUSCLES DISTAL TO THE BREAKING JOINT.

Series Second Segment.| Third Segment. Fourth Segment.) Fifth Segment.
Regeneration. |
No. Flex. Exten. Flex. Exten. Flex. Exten. Exten. Exten
29 5 da. 9 hrs ar 0 0 0 0 0 0
30 | 5 da. 10 hrs + 0 0 +* 0 0
31 5 da. 22 hrs. + + + +* + +%* 0 0
3a sG.das Guhirs: +oo+ + + + + + +

*Differentiation just beginning.

In this table it will be seen that the first muscle fibrils to be differ-
entiated were those of the flexor in the second segment; the next to
appear were those of the flexor in the fourth segment. Up to this
time no muscle fibrillse were found in either the third segment or
in the extensors in the second and fourth segments. The flexors in
the fifth segment were the last to develop. Here again, differentia-

Tissues in the Crustacean Limb. 151

tion does not proceed definitely in either a proximal or a distal
direction, but appears rather to be correlated with the size and fune-
tion of the muscles concerned.

3. Differentiation of Striw in the Myofibrille.—The series were
also studied to ascertain the time at which myofibrils in each muscle
showed striations. The following table represents the results ob-
tained. The signs + and 0 indicate, respectively, the presence and
absence of striation.

TABLE SHOWING THE SEQUENCE IN THE DIFFERENTIATION OF STRIAE IN THE
REGENERATING Limp MUSCLES.

Series Second Segment. Third Segment. |Fourth Segment. Fifth Segment.
Regeneration.
No Flex. Exten. | Flex. Exten. | Flex. Exten. | Exten. Exten.
34 6 da. 22 hrs. +%* + 0 0
35 |7da. 2 hrs. | oL ea eee + + Se Sap + if

*Striations were not found until after the myofibrillz had begun to be
differentiated in all the segments.
{Differentiation of strise just becoming evident.

The time intervening between the appearance of the first stris
and the striation of all the muscles is apparently much shorter than
the time between the first differentiation of myofibrils and their
complete development for all the muscles. N evertheless, striation
does not occur simultaneously throughout the limb. The strie differ-
entiate first in the flexor museles in the second and fourth segments,
and later they appear in the remaining limb muscles.

4. Conclusions.—The results of the present study of the differen-
tiation of the chitinogenous muscle tendons, the myofibrille and their
striation justify two conclusions:

First, that the differentiation of the regenerating musculature
proceeds in neither a definite disto-proximal direction, nor the
reverse.

Second, that whatever sequence there may be in this differentia-
tion is correlated rather with the size and functional relations of the
various muscles concerned, the larger muscles differentiating first.

152 Victor E. Emmel.

Reference has already been made to the recent discussions of the
direction of differentiation by Child (06, ’08), Holmes (’04, ’07),
Zeleny (07), and Haseman (’07). Among these investigators Hase-
man has studied the form most closely related to the lobster. From
his observations upon the regenerating appendages of the crayfish,
he concludes that differentiation in the chela proceeds from the tip
toward the base, while the reverse is true for the walking legs.

In view of the results of these investigators Zeleny has discussed
three possibilities regarding the direction of differentiation in the
development of an appendage.

1. That “all the parts may appear at once”; (2) that “the progres-
sion may be outwardly directed’, or (3) “it may be inwardly di-
rected”. Zeleny points out that the first lacks evidence, that the
second has been more especially favored by Pfliiger and Holmes in
connection with certain general theories of development, and the third
by Morgan, Driesch, and Child. From his own study Zeleny con-
cludes that the second method may predominate in some animals,
the third in others, and further that both modes may predominate
at different times in the same animal, as appears to be the case in
the antennule of Maneasellus macrourus. It is evident that the
present data from the lobster fail to conform with any of the three
modes of differentiation. But the evidence does support a fourth
proposition, viz., that sequence of differentiation may be influenced
or determined by the relative size and functional role of the regenerat-
ing structures. Consequently, differentiation in the regenerating
appendage is not necessarily either “centrifugal” nor “centripetal,”
the new structures, on the contrary, differentiating rather according
to size and functional relations, in a manner perhaps similar to the

formation of organs in embryonic development,—a most striking
illustration of which is furnished by Sutton’s (’83) rule for the
differentiation of the epiphyses of the long bones in human osteology :
“The centers of ossification appear earliest for those epiphyses which
bear the largest relative proportion to the shafts of the bones to

which they belong” (p. 480).

IX. Summary.

1. Two new structures are recorded in connection with the break-
ing joint of the chela of lobsters.

Tissues in the Crustacean Limb. 153

a. A muscle crossing the joint, but degenerating during the fifth
larval stage.

b. Valves in the venous blood sinus which prevent excessive hem-
orrhage after the autotomy of the limb. The valves persist through-
out life.

The following conclusions concern the regeneration of the limb
after autotomy.

2. The first layer of cells,
wound is formed by the migration of epidermal cells.

aside from the blood clot, to cover the

3. The nuclei of these migrating cells enlarge and become some-
what wedge-shaped. The karyosomes disintegrate into fine granules
which tend to collect at the proximal end of the nucleus. Thus the
nuclear contents appear polarized into an inner zone of darkly
stained chromatin, and an outer zone relatively free from chromatin
granules.

4. The significance of this polarization is not clear, but its regular
occurrence, and is reversed on opposite sides of the limb, together
with the frequent expansion of the higher pole, and the association
of axial chromation masses with the darker pole, indicate that it is a
phenomenon characteristic of certain stages in the regenerative activ-
ity of the epidermal cell.

5. During these nuclear activities the cytoplasm loses its suppor-
tive fibrille and reticular structure, and becomes finely granular.
Although definite cell membranes are never evident, there are more
or less clearly defined cytoplasmic units around each nucleus.>

6. Mitotic cell division begins after the formation of the first
epidermal plate over the wound. The formation of this plate seems
sufficiently accounted for by the volumetric increase of the cytoplasm,
together with the enlargement and wider separation of the nuclei
of the migrating cells.

7. The wall of the regenerating bud, and a large part, if not the
entire core of cells filling its interior, are derived from epidermal
cells. Since there is no conclusive evidence of either cell division
or migration among the old muscle and connective tissue cells, these
tissues appear to contribute little, if anything, to the formation of the
new limb bud.

154 Victor E. Emmel.

8. a. Regenerating striated muscle is ectodermal in origin.

b. The myofibrille appear genetically related to the eyto-reticulum.
Each fibril is early surrounded by a sheath of modified cytoplasm,
multiplies by longitudinal splitting, and in its differentiation the “z”
line or membrane of Krause, becomes evident after the formation
of the light and dark bands.

e. The muscle fiber or cell is multinuclear when first formed. Later
the nuclei come to be peripheral in position while the fibrille he
to the side of the fiber nearest the center of the muscle bundle. The
nuclei multiply by mitotic division, and with the development of the
sarcolemma they become flattened and elongated in the long axis of
the fibers.

d. The myofibrille differentiate throughout their whole length into
true striated muscle elements, except in the region of skeletal attach-
ment. There the peripheral ends of the fibrils remain unstriated,
and serve as tensile elements.

e. The connective tissue over the inner surface of the epidermis,
and the purely supporting fibrille within the epidermal cells develop
later, but these structures appear to be secondary in their relation to
muscle attachment.

9. a. The present observations indicate that the neurilemma is
derived from the regenerating epidermal cells.

b. During certain stages in the differentiation of the axis cylinder
the finer neurofibrille are central and the coarser fibrils more per-
ipheral in position.

10. In at least certain regions of the regenerating limb, supporting
and apparently true connective tissue differentiates from epidermal
cells. ;

11. a. In the development of the chitinogenous muscle plates and
the myofibrille and their striz, differentiation appears to be neither
directly “centrifugal” nor “centripetal” in direction.

b. On the contrary, the facts warrant the statement that whatever
sequence there may be in differentiation, it is correlated with the size
and functional relations of the muscles concerned, rather than with
any distal or proximal relation of these structures to the organism.

Cr

Tissues in the Crustacean Limb. 15

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Herrick, F. H., ’95. The American Lobster. Bull. U. S. Fish Commission.
252 pp. Fifty-four plates.

HouMEs, S., 04. The Problem of Form Regulation. Arch. f. Ent.-Mech., Bd. 17.

. 07. The Behavior of Loxyphyllum and Its Relation to Regenera-
tion. Journ. Exp. Zo6l., Vol. IV, pp. 3899-480.

HouMGREN, N., ’02. Ueber das Verhalten des Chitins und Epithels zu den
unterliegenden Gewebarten bei Insecten. Anat. Anz., Bd. 21, pp. 480-488.

Huxtry, T. H., ’88. The crayfish. New York.

KASTSCHENKO, N., ’88. Zur Entwicklungsgeschichte des Selachierembryos.
Anat. Anz., Bd. 3, pp. 445-467.

KG6.LiriKer, A., von, ’84. Die embryonalen Keimblitter und die Gewebe. Zeit-
schrift fiir wissenschaftliche Zoologie, Bd. 40.

LécarLton, A., ’07. Recherches sur la structure de la cuticle tegumentaire
des Insectes. Bibliog. Anat., t. 16, pp. 245-261.

MacCatium, J. B., 98. On the Histogenesis of the Striated Muscle Fiber and
the Growth of the Human Sartorius Muscle. Johns Hopkins Hosp.
Bull., Balt., Vol. IX, pp. 208-215.

McMourricu, J. P., 96. The Epithelium of the So-called Mid-gut of Terres-
trial Isopods. Journ. of Morph., Vol. 14, pp. 88-103.

Minor, C. S., 94. Text-book of Human Hmbryology. Philadelphia.

, O01. The Embryological Basis of Pathology. Science, Vol. XIIT,
pp. 481-498.
. 08. The Problem of Age, Growth and Death. New York.

Morean, T. H., 00. Further Experiments on Regeneration of the Appendages

of the Hermit Crab. Anat. Anz.,-Vol. 17.
. 01. Regeneration. New York.

Ost, J., 706. Zur Kenntnis der Regeneration der Extremitiiten bei den Arth-
ropoden. Arch. f. Ent.-Mech., Bd. 22, pp. 289-324.

Puattr, JULIA B., 98. The Development of the Cartilaginous Skull and of the

- Branchial and Hypoglossal Musculature in Necturus. Morphologisches
Jahrbuch, Bd. 25, pp. 877-464.

Ranp, H. W., ’04. The Behavior of the Hpidermis of the Earthworm in
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Reep, M. A., ’04. The Regeneration of the First Leg of the Crayfish. Arch.
f, Ent.-Mech., Bd. 18, pp. 307-316. Two plates.

Tissues in the Crustacean Limb. 157

Ritzy, W. A., 08. Muscle Attachment in Insects. Annals of the Entomolog.
Soc. of Am., Vol. I, pp. 265-269. One plate.

Roux, W., 96. Ueber die Selbstordnung (cytotaxis) sich “beriihrender” Fur-
chungszellen des Froscheies durch Zellenzusammenftigung, Zellentren-
nung und Zellengleiten. Arch. f. Ent.-Mech., Bd. 3, pp. 381-468. Two
plates.

SCHNEIDER, K. C., 02. Lehrbuch der vergleichenden Histologie. Jena.

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STEELE, M. I., ’07. . Regeneration in Compound Eyes of Crustacea. Journ.
Exp. Zool., Vol. V, pp. 163-248. Sixteen plates.

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and Physiol., Vol. XVII, p. 479-4838.

WiiiAMsS, L. W., ’07. The Stomach of the Lobster. 37th Report of Inland
Fisheries of Rhode Island, pp. 153-180. Ten plates.

ZELENY, C., 07. The Direction of Differentiation in Development. Arch. f.
Ent.-Mech., Bd. 23, pp. 324-348. Seven plates.

XI. DESCRIPTION OF PLATES.

Unless otherwise indicated, all figures are from camera drawings made
from longitudinal sections of the chelze of fourth stage lobsters.

List OF ABBREVIATIONS.

a, artery.

at, a’, arteries passing through the breaking joint.

b, blood plasm.

be, blood corpuscles.

bel, blood clot.

bk, breaking joint.

bs, basipodite.

c, carpopodite.

ch, chitin.

ch‘, new chitin.

cm, connective tissue membrane,

ct, connective tissue.

d, dactylopodite.

e, epidermis.

e*, migrating epidermal cells.

e*, e, e*, e*, extensor mucles in the propodite, carpopodite, meropodite, and
ischiopodite, respectively.

ec, regenerating and proliferating epidermal cells.

ef, Supportive fibrille.

em, epidermal membrane.

en, regenerating epidermal nuclei.

i, ischiopodite.

158 Victor E. Emmel.

ic, inward proliferation of epidermal cells to form connective tissue.

in, invagination of epidermal cells at the joints

m, muscle crossing the breaking joint.

ma, muscle attachment.

mb, muscle in the basipodite.

me, meropodite.

mf, myofibrille.

mn, muscle nuclei. E

mp, chitinogenous muscle plate or tendon.

mt, cells undergoing mitotic division.

n, nerve.

nt, n2, nerves passing through the breaking joint.

nf, neurofibrille.

ng, nuclei of the “Grenzlamelle.”

nn, nuclei of the neurilemma.

p, propodite.

m, 7, 73, flexor muscles in the propodite, carpopodite, and meropodite, re-
spectively.

rt, cyto-reticulum.

s, septum of connective tissue dividing venous blood sinus into two channels.

sa, sarcolemma.

v, valves in venous blood channels.

v, valves in venous blood channels; v', v*, inner and outer channels, respect-
ively.

ve, venous blood channel.

vs, yenous sinus; vs', vs?, respectively distant and proximal of the breaking
joint.

PLATE I.
STRUCTURE OF THE CHELA IN THE REGION OF AMPUTATION.

Graphic reconstructions of the breaking joint and related segments of the
left chela, to show the anatomical relations of the blood vessels, muscles.
nerves, epidermis, and connective tissue. The drawings are incomplete in one
particular because the finer meshwork of connective tissue in which the
arteries and nerves are suspended is not represented in its entirety.

Fic. 1 represents the outer third, and

Fic. 2 the inner third of the breaking joint and segments. The sides of the
figures nearest each other are ventral. > 263.

PLATE I,

TISSUES IN THE CRUSTACEAN LIMB.

EMMEL.

VICTOR E.

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Vou. 10, No. 1, JAN., 1910.

THE AMERICAN JOURNAL OF ANATOMY.

PLATE II.
STRUCTURE OF THE CHELA IN THE REGION OF AMPUTATION.

Fic. 8. Section of the breaking joint from the region midway between the
parts represented in Figs. 1 and 2, to show the relations of the epidermis,
nerve trunk, and connective tissue membranes preceding autotomy. < 368.

Fic. 4. Showing relation of tissues after the autotomy of the limb, and the
function of the valve (v?) in closing the venous blood channel (ve). >< 368.

Fie. 5. Section showing the relations of the muscle (mm) discovered at the
breaking joint (bk). < 204.

Fic. 6. In the fifth stage lobster this muscle has almost entirely degen-
erated only remnants (md) of the former muscle being present. < 210.

?

PLATE II.

TISSUES IN THE CRUSTACEAN LIMB.

EMMEL.

VICTOR E.

"9g “DIA

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THE AMERICAN JOURNAL OF ANATOMY.—-VOL. 10, No. 1, JaNn., 1910.

PLATE III.
SUCCESSIVE STAGES IN THE REGENERATION OF THE LIMB,

Fic. 7. Regeneration, 1 day. Epidermal cells (e') beginning to migrate
across the wound beneath the blood clot (bel). >< 240.

Fic. 8. Regeneration, 1 day, 14 hours. Migrating epidermal cells have
formed a complete plate or disc over the wound. Valve (v) of the venous
blood channel now assuming its normal position. < 240.

Fic. 9. Regeneration, 2 days, 2 hours. The beginning of cell multiplication
(mt) in the first formed epidermal plate. >< 240.

Ira. 10. Regeneration, 2 days, 22 hours. Epidermal cells (ec’) at the apex
of the bud are migrating inward to form the flexor muscle of the claw. x 240.

PLATE III.

TISSUES IN THE CRUSTACEAN LIMB.

VICTOR BE. EMMEL.

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THE AMERICAN JOURNAL OF ANATOMY.

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PLATE IV.
NUCLEAR CHANGES.

Changes in epidermal nuclei during regeneration. The groups of nuclei in
Figs. 11-13 taken from the same region of the limb, 7. e., on the same side of
the limb just below the breaking joint; in each case (c) and (d) are nearer
the center of the wound than (a) and (0).

Fic. 11. Normal nuclei before the amputation of the limb. >< 2000.

Wie. 12. Regeneration, 14 hours. < 2000.

Fig. 18. Regeneration, 24 hours. x 2000.

Fic. 14. Regeneration, 2 days, 2 hours. Nuclei undergoing mitosis; @
(apparently), prophase; b, anaphase. > 2000.

Fig. 15. Regeneration, 5 days, 10 hours. Nuclei in the epidermal wall of
the regenerating limb bud, in the region of the invagination (in) for the
flexor muscle in the meropodite. >< 1250.

TISSUES IN THE CRUSTACEAN LIMP. PLATE Iv.

VICTOR E. EMMEL.

Hie, 15,

THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

PLATE VY.
REGENERATION OF STRIATED MUSCLE,

Fie. 16. Regeneration, 6 days, 6 hours. Myofibrillee (af) just beginning to
differentiate for the flexor of the dactylopodite. >< 334.

Fic. 17. Regeneration, 7 days, 6 hours. Showing the great elongation of
epidermal nuclei during their invagination for the formation of the muscles,—
in this case the flexor is the meropodite. >< 1250. (Cf. with Fig. 15.)

Fic. 18. Regeneration, 12 days, 10 hours. Shows striation of muscle fibrils
(mf) and their attachment to the chitin of exoskeleton. > 334.

Ilia. 19. Showing skeletal attachment of regenerating muscle fibrils. (From
the regenerating chela of a two-year-old lobster.) The myofibrillee appear to
be attached directly to the chitin (ch’). x 292.

PLATE V.

TISSUES IN THE CRUSTACEAN LIMB.

VICTOR E. EMMEL.

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Fic. 16.

THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

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PLATE VI.
REGENERATION OF STRIATED MUSCLE.

Iie. 20. Regeneration, 10 days, 6 hours. Showing the muscle fibrils form-
ing brush-like end pieces in their attachment to the chitin. Epidermal mem-
brane (em) just beginning to differentiate. > 1250.

Fic. 21. Completely regenerated epidermis in the chela of a sixth stage
lobster just after moulting, showing a nucleus (ng) of the “grenzlamelle,”
and the relation of the “grenzlamelle” to the epidermis and muscle fibers.
< 750.

Vig. 22. Regeneration, 6 days, 2 hours. Showing the close relation of the
myofibrillae (mf) to the cytoplasmic reticulum (rt) during early differentia-
tion. > 1250.

Fic. 28. Regeneration, 7 days, 6 hours. Myofibrillae (mf) surrounded by
a sheath of modified cytoplasm (transverse section). 1250.

Via. 24. Regeneration, 8 days, 10 hours. Transverse section of myofi-
brillae (mf), Showing the first longitudinal splitting of the fibrils. >< 1250.

ia. 25. Regeneration, 11 days, 10 hours. Transverse section of a fully
formed muscle fiber showing membrane or sarcolemma (sq@), nuclei, and Cohn-
heim’s areas. >< 1250.

TISSUES IN THE CRUSTACEAN LIMB.
VICTOR E. EMMEL.

---- em

wae — ~~~ — 6

Fie. 20.
mn rt
1 i
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Fic. 22.

=

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PLATE VI.

ng I

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Fig. 23.

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THE AMERICAN JOURNAL oF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

PLATE VII.
REGENERATION OF CONNECTIVE TISSUE AND NERVES.

Fic. 26. Regeneration, 10 days, 2 hours. Shows inward proliferation of
epidermal cells to form a sheet of connective tissue (ic) across the segment
(ineropodite). > 417.

Fig. 27. Regeneration, 11 days, 10 hours. A later stage in the differentia-
tion of the connective tissue (in the meropodite) showing vacuolation and
infiltration of blood plasm (6). < 834.

Fic. 28. Regeneration, 5 days, 10 hours. Proliferation of epidermal cells
(e) in the formation of a nerve fiber (nm). 582.

Fie. 29. A stage in the differentiation of the nerve fiber showing a nucle-
ated sheath and axial core of delicate neurofibrille (nf) in the axis cylinder.
At the periphery of this core of fibrille may be observed two heavier fibrillz.
< 750. :

Ilia. 30. A stage of the regenerating nerve fiber in which the heavier neu-
rofibrillae predominate. >< 750. (Figures 29 and 30 were made from sections
of the regenerating chela of a two-year-old lobster in which the neurofibrillz
were more sharply defined than in preparations from younger specimens. )

TISSUES IN THE CRUSTACEAN LIMB. PLATE VII.

VICTOR E. EMMBEL.

Hic. 26:

Y

- Wie. 30. ' Fie. 29.

3

THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

%

PLATE VIII.
THE DIRECTION OF DIFFERENTIATION,

The direction of differentiation of the eight chitinogenous muscle plates or
tendons of the limb. Figs. 31-36 are reconstructions from serial sections.

Fic. 31. Regeneration, 2 days, 22 hours. x 60.

lig. 32. Regeneration, 4 days, 6 hours. x 60.

Fic. 33. Regeneration, 4 days, 22 hours. x 60.

ic. 34. Regeneration, 5 days, 6 hours. » 60.

Fic. 35. Regeneration, 5 days, 9 hours. » 60.
Fic. 36. Regeneration, 10 days, 2 hours. 54.
Fig. 37. Diagrammatic drawing of a fully developed chela showing the

relative size and position of the eight limb muscles distal of the breaking joint
(bk). 4-9.

TISSUES IN THE CRUSTACEAN LIMB, PLATE VIII.
VICTOR E. EMMEL.

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THE AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

SOME ANOMALIES IN THE GENITAL ORGANS OF
BUFO LENTIGINOSUS AND THEIR
PROBABLE SIGNIFICANCE.

BY

HELEN DEAN KING,

Assistant in Anatomy, Wistar Institute.
WITH 26 FIGURES.

During the course of a series of investigations on the germ-cells
of the common American toad, Bufo lentiginosus, I have had occa-
sion to examine a large number of these amphibians at various
stages of their development, and I have found many individuals in
which the genital organs showed marked deviations from the normal
type. The most striking of these anomalies are described in the
present paper. Many cases of this kind have a direct bearing on the
question of the existence of hermaphroditism in the primitive verte-
brates, and but few of them have as yet been recorded for any
amphibian other than Rana.

I. ANOMALIES IN THE GenrTAL Orcans or Youne Toaps.

Anomalies occur much more commonly in the sex-glands of young
toads than in those of adults, and at least two per cent of the young
individuals that I have examined showed more or less marked irregu-
larities of this kind. As most of the toads in which abnormalities
were found were reared in the laboratory, one might infer that the
anomalies were the result of pathological changes produced in the
genital organs by abnormal environmental conditions. That such
is not the case, however, is shown by the fact that eleven individuals
in which the sex-glands were anomalous in some respect were found
in a lot of 500 young toads that had completed their metamorphosis
under natural conditions.

Although in Bufo sex is probably determined at or before the
time that the egg is fertilized, the gonads are often in an apparently

THH AMDPRICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, JAN., 1910.

160 Helen Dean King.

indifferent state even at the time of metamorphosis, and in many
instances the genital glands must be examined histologically before
the sex of an individual can be ascertained. The toad grows very
rapidly after completing its metamorphosis, and then, except in very
rare cases, the sexes are readily distinguished by an examination of
the sex-glands under a dissecting lens. At this period of development
the testes are long, cylindrical bodies with a smooth contour (Figs. 3
and 6, Left) ; the ovaries, although they are of about the same length
as the testes, are relatively broader and they have an irregular, jagged
outline (Figs. 5 and 7, Left). At the anterior end of each genital
gland is a large rounded body, Bidder’s organ (Fig. 1, B. O.), to
which the corpora adiposa are attached (Fig. 1, C. A.). In young
toads Bidder’s organ is of the same size and structure in both sexes:
it persists throughout the lifetime of the male, but it disappears in
the female towards the end of the second year. <A study of the struc-
ture and development of Bidder’s organ (King, ’08 a) has shown that
this body is undoubtedly a rudimentary ovary in which the cells have
marked characteristics which readily distinguish them from the eggs
developing in the ovaries.

As a rule there is but one Bidder’s organ at the anterior end of a
sex-gland, and it usually measures about 0.4 mm. in diameter. In
many young toads Bidder’s organ shows considerable deviation from
the normal size and shape. In some instances this structure is very
large, measuring 0.6 mm. in diameter (Figs. 1, 2, 7); in rarer cases
it is less than one-half of its usual volume (Fig. 13). Examined
histologically such Bidder’s organs are found to differ from those of
normal size only in the number of ova that they contain.

Not infrequently a Bidder’s organ is found that is greatly elon-
gated (Figs. 7,9, 12). In such eases this body is usually indented in
the middle region (Figs. 7, 9), although sometimes it has a perfectly
smooth contour (Fig. 12). Anomalies of this kind indicate clearly
the steps by which the one Bidder’s organ becomes divided into two;
and the presence of two of these structures at the anterior end of an
ovary or of a testis is probably the most common abnormality occur-
ring in the genital organs of young toads. Usually, in cases like this,
one Bidder’s organ lies directly behind the other (Figs. 1, 10, 11,

Anomalies in the Genital Organs of Toads. 161

14); more rarely these bodies are separated as shown in Fig. 3.
I have found one instance only in which there were three Bidder’s
organs at the anterior end of a sex-gland (Fig. 16). Abnormalities
of this type can doubtless be ascribed to the mechanical effect of
pressure exerted by some structure on Bidder’s organ at a very early
period in its development.

In many individuals one or more rounded bodies, similar in
appearance to the normal Bidder’s organ although usually much
smaller, appear along the course of the sex-gland (Figs. 1, 2, 5, 8,
ete.). Structures of this kind may occur on any part of the gland,
and they are found in both sexes, although they are, perhaps, more
common in males than in females. The probable origin of these
bodies will be considered later.

Fig. 12 shows in outline the most interesting of the anomalies
found in the genital organs of young toads. On the left side Bidder’s
organ appears much enlarged and greatly elongated; on the right
there are four rounded Bidder’s organs lying one behind the other.
Sections of these bodies show that each has the structure typical of
a normal Bidder’s organ. As in this individual the ovaries were not
more than one-half of their normal length, it is evident that some
of the germ-cells in the anterior part of each ovary became incor-
porated with Bidder’s organ and assumed the characteristics of
rudimentary ova. A somewhat similar case in which the anterior
part of a testis has been changed into a Bidder’s organ is shown in
Fig. 16. Abnormalities of this kind can hardly be due to any
mechanical cause, although possibly a lessening of the normal blood
supply to the anterior part of a sex-gland might induce such changes
which undoubtedly must be considered as degenerative in their
character.

In all of the cases so far described, and in many others of the
same general character, sections were made of the genital glands
and the anomalies appearing in them carefully studied. All enlarge-
ments of the sex-glands, no matter what their position or their size,
were found to contain large cells having all of the characteristics of
the rudimentary ova of which the normal Bidder’s organ is composed.
If there are two Bidder’s organs at the anterior end of a sex-gland,

162 Helen Dean King.

each has the structure typical of the normal Bidder’s organ. Most
of the large bodies found along the course of the sex-glands (Figs.
6, 13, 16) also appear similar in structure to Bidder’s organ, and
they are directly connected above and below with germ-cell tissue
that appears normal in every respect.

The structure of many of the smaller bodies found on the sex-
glands does not conform strictly to that of Bidder’s organ. Fig.
17 shows a longitudinal section of the enlargement on the lower
part of the left ovary, which is outlined in Fig. 10. The ovarian
wall has a perfectly normal structure here as in other parts of the
sex-gland, being composed of cysts of secondary oégonia and of
odeytes in the synizesis stage of development (Fig. 17, S8.). In
the cavity of the ovary is a mass of large cells inclosed by follicle
cells and a thin membrane. These large cells have all of the char-
acteristics of the large rudimentary ova normally found in Bidder’s
organ; they do not bear the slightest resemblance to the odgonia
and odeytes of which the ovarian wall is composed. A somewhat
different arrangement of tissues is shown in Fig. 18 which is a
drawing of a transverse section through the smaller of the two
bodies on the right testis outlined in Fig. 2. Here the spermato-
gonial tissue does not inclose the group of cells which appear like
rudimentary ova, but is collected together at one side of it and the
cells are surrounded in great part only by a thin covering of mesen-
tery. A similar condition of the tissues was found in the enlarge-
ments of the testes shown in outline in Figs. 8 and 15.

As a rule only the enlarged portions of the sex-glands contain
any of the large cells which appear like rudimentary ova, in all
other parts the glands have a normal structure. In but one instance
have I found cells of this character among germ-cells when their
presence was not shown by an examination of the sex-gland under
a dissecting lens. In this case the cavity of one of the ovaries in a
young female contained three of these large cells which were sepa-
rated a considerable distance and thus gave no external evidence of
their presence.

During the course of my investigations on the toad (King,
07, 708, ’08a) I have made sections of the gonads of a large

Anomalies in the Genital Organs of Toads. 163

\

number of tadpoles in various stages of development from the time
of hatching up to metamorphosis. In no case have I found any
abnormalities in the gonads proper, although in several instances
Bidder’s organ on one or both sides had been divided as shown in
Figs. 11 and 14. The large cells resembling rudimentary ova which
are found singly or in groups in the genital organs of so many
young toads that have recently completed their metamorphosis must,
therefore, develop very quickly, presumably just before or during
the period of metamorphosis. It is highly improbable that these
cells originate in Bidder’s organ and subsequently migrate into the
sex-gland, as there is never any opening between these structures
except in the female toward the end of the second year when the
entire Bidder’s organ is degenerating. Such cells, moreover, never
show the slightest evidence of amceboid movement, and in many
instances the membrane surrounding them is continuous with that
inclosing the primordial germ-cells themselves. Judging from my
previous investigations I am strongly inclined to the opinion that
these cells are primordial germ-cells in which, for some unknown
reason, the course of development has been changed so that the cells
increase in size with unusual rapidity and assume the character-
istics of rudimentary ova. Cells of this kind must, therefore, be
considered as degenerating cells. ‘Their presence in the sex-gland
is apparently not harmful to the individual, since none of the
young toads in which they are found seem to differ in any other
way from the normal type. In Bufo, with rare exceptions, all cells
of this character must become absorbed early in the life history of
the individual. I have never found any of them in the sex-glands
of toads that were more than three or four months old.

According to Pfliiger, ’82, there are three kinds of individuals
to be found among young frogs killed soon after completing their
metamorphosis: males, females, and hermaphrodites. In the course of
a few months the hermaphroditic forms become either definite males
or females, and in few cases only does the hermaphroditie condition
persist until the individual becomes an adult. During the course
of a series of investigations on the determination of sex in frogs,
Hertwig, 06, ’07, has found a number of individuals in which sex

164 Helen Dean King.

could not be ascertained even after metamorphosis was completed.
These individuals, Hertwig believes, correspond to the ‘hermaphro-
ditic’”’ forms described by Pfluger. Schmitt-Marcel, ’08, has recently
studied the structure of the sex-glands in young Rana temporaria
that appeared to be hermaphrodites. He states that in all cases the
sex-glands of such individuals, which he calls “intermediate forms,”
appear much more like ovaries than like testes, yet they probably all
ultimately develop into testes. In accounting for the origin of such
forms Schmitt-Marcel assumes that they are derived from young
females, and he concludes: “Die Veranderung also, die von einer
normalen weiblichen Driise zu dieser Bildung fiihrt, besteht im
wesentlichen wohl darin dass bei einem Wachstum des Organes nicht
in der ganzen Keimdriise ein Heranwachsen von Urkeimzellen zu
jungen Eizellen stattfindet, sondern dass ganze Strecken auf dem
Stadium der Urkeimzellen stehen bleiben und sich als solche weiter
vermehren, wihrend gleichzeitig eine ausserordentliche in der nor-
malen weiblichen Keimdriise fehlende Vermehrung der Stroma
Platz greift.” The young ova grow to a considerable size and then
degenerate ; meanwhile indifferent germ-cell tissue spreads through-
out the whole sex-gland and later develops into spermatogonia so
that the individual eventually becomes a male. Schmitt-Marcel
states that he finds an increasing number of such forms among young
frogs killed from the second to the tenth month after metamorphosis,
and that the number of these forms gradually decreases after this
time. Among adult frogs such “intermediate forms” are not known,
males and females being found in about equal proportions.

There is a great similarity between the sex-glands of the “inter-
mediate forms” found among young Rana temporaria and those of
young toads having the anomalies described above. In both cases
large cells, having the general appearance of young ova, are found
among primordial germ-cells which are still in an apparently indif-
ferent state. These cells develop to a certain size and then undergo
processes of degeneration and absorption, leaving the gland male
or female as the case may be. The chief differences between such
anomalies in Rana and those in Bufo consist in the fact that the
large cells which appear in the sex-glands of young toads are, as a

Anomalies in the Genital Organs of Toads. 165

rule, segregated into one or several masses which can be seen under
a low magnification before the gland is sectioned, and they have
very distinctive characteristics that distinguish them from normal
ova; in Rana, according to Schmitt-Marcel, the large cells are scat-
tered throughout the sex-gland, and they appear in every respect like
normal ova.

Schmitt-Marcel offers no suggestion as to how or why, in an
individual in which the female sex is already determined, the pri-
mordial germ-cells can change the course of their development and
subsequently alter the sex of the individual. If this phenomenon
is of common occurrence in young Rana temporaria, then it would
seem as if some clue might be’ obtained as to the causes which deter-
mine sex in this species if an extensive series of experiments was to be
made with young individuals that had recently completed their meta-
morphosis.

Except in rare cases, the sex of young toads in which anoma-
lies appear in the genital organs can readily be ascertained, since,
although the primordial germ-cells may appear alike in both sexes
at this time, the ovary has a central cavity which is lacking in the
testes. There are, therefore, no young toads that can properly be
ealled “intermediate forms,” as each individual is at this time defi-
nitely male or female, no matter how many cells appearing like
rudimentary ova may be present in the sex-glands.

The development of the genital organs must be considerably
slower in Rana temporaria than in Bufo lentiginosus, since in the
former species it is impossible to ascertain the sex of many indi-
viduals until they are nearly a year old. As according to Schmitt-
Marcel the germ-cell tissue which spreads throughout the sex-glands
in these intermediate forms is still in an indifferent state I can see
no valid reason for the assumption that such forms were originally
females. In Bufo, as far as I can judge from the number of cases
(about 75) that have come under my observation, large cells with
the characteristics of rudimentary ova appear somewhat more com-
monly in the genital glands of young males than in those of young
females. Assuming that a similar condition exists in young Rana
temporaria, one readily accounts for the fact that the great majority

166 Helen Dean King.

of “intermediate forms” ultimately become males, and one also
avoids the necessity of presuming that sex can be altered after an
individual has completed its metamorphosis.

In young toads, and possibly also in young frogs, primordial
germ-cells that fail to undergo normal processes of development
assume the characteristics of rudimentary ova, regardless of the sex
of the individual in which they occur. An explanation of this phe-
nomenon seems to me possible if one accepts for the amphibians
Haeckel’s, ’74, view ‘“‘dass das dlteste und urspriinglichste Ge-
schlechtsverhiltniss die Zwitterbildung war und dass aus dieser erst
secundir (durch Arbeitstheilung) die Geschlechtstrennung hervor-
ging.” In the male amphibian at the present time many of the
primordial germ-cells still have the power, under certain conditions,
of developing into ova which, since they cannot leave the sex-gland
or come to maturity, are destined to degeneration and absorption.
Except in very rare cases the primordial germ-cells in the female
are no longer able to develop into spermatogonia. When, there-
fore, these cells fail to develop along normal lines they become rudi-
mentary ova which are similar in structure to the rudimentary ova
derived from the primordial germ-cells in the male and they have
a similar fate. —

In adult amphibians, as a rule, germ-cells which fail to undergo
normal processes at any stage of their development disintegrate at
once and become absorbed. Some few cases have been found, how-
ever, in which germ-cells in adult males have changed the course
of their development and become rudimenary ova. Cole, 795, Fried-
mann, 98, Latter, ’90, and Punnett, ’00, have noted the presence
of rudimentary ova in the testes of various species of frogs, and I
also have a preparation of the testis of an adult Rana pipiens which
shows this anomaly. Rudimentary ova have also been found in the
testes of adult Bufo vulgaris by Spengel, ’76, Moffmann, ’86, Knappe,
’86, and Friedmann, 798, but as yet no cells of this kind have been
found in the testes of the American species, Bufo lentiginosus.

Fig. 4 shows a type of abnormality which is found occasionally
in the sex-glands of young toads. The median genital ridge, which
is usually divided into two ridges when a tadpole is from twelve to

Anomalies in the Genital Organs of Toads. 167

fourteen days old, has separated only in the anterior region and it
still remains undivided posteriorly even after the toad has com-
pleted its metamorphosis. Sections of the sex-glands show no other
apparent abnormality.

Il. ANOMALIES IN THE GENITAL OrGaANS or Aputt Toaps.

Among the large number of adult toads which I have examined
during the past ten years there was but one individual in which the
sex-glands appeared in any way abnormal. This toad, which proved
to be a rudimentary hermaphrodite, was brought into the vivarium
of the University of Pennsylvania on the morning of March 30, 1907,
having been captured in a pond near West Philadelphia. The abnor-
mal character of the sex-glands was noticed as soon as the body
cavity was opened, and these organs were removed at once and fixed
in Flemming’s fluid; the body of the toad being placed in a 4 per
cent formalin solution for future examination.

Measurements that were made showed that this toad was
somewhat larger than the average male as it had a body length of
8.5 em. The thumbs were deeply pigmented, and they bore pads
similar to those found on the thumbs of all male toads during the
breeding season, although these structures were somewhat smaller
than normal.

Photographs of the genital organs of this individual are shown in
the text-figure. On the left side (B) there is a well developed testis
(T), measuring 8 mm. in length and 3 mm. in width. Above the testis
is a large, pigmented, irregularly shaped body (O), unquestionably
a rudimentary ovary, which is 15 mm. long and 7 mm. across at its
widest part. The genital organs on the right side (A) are noticeably
smaller than those on the left, although they have a similar structure.
The testis (T) is but 7 mm. long; the rudimentary ovary above it
(O), which has a much more regular outline than that on the left
side, measures 10 mm. in length and 7 mm. in width. There is no
Bidder’s organ on either side, the fat bodies (C. A.) being attached
directly to the anterior surface of the rudimentary ovaries.

Transverse sections through the testes show that these bodies are
composed of cysts filled either with ripe spermatozoa or with sper-

168 Helen Dean King.

matocytes and spermatids in various stages of development. A careful
study of many cysts failed to show a single instance in which the
development of the germ-cells appeared to be abnormal. The cysts
in the centre of the testes are not crowded quite as closely together
as are the cysts in the testes of the normal male, and the stroma
separating them is unusually thick; in all other respects the testes
appear perfectly normal. Each testis was connected with the kidney
by vasa efferentia, the genital ducts appearing normal. In this
individual the Miillerian ducts were still present; but, although
they were quite large and much convoluted, they showed no dilatations
either in the anterior or in the posterior region.

Interest in this individual centers in the structure of the rudimen-
tary ovaries lying above and attached to the testis on either side.

A B

Photographs of the right (A) and of the left (B) ovo-testis found in an
adult Bufo lentiginosus. O, ovary; T, testis; C. A, corpora adiposa.
The position of these bodies and the fact that no Bidder’s organs
are to be found indicate that they have been produced, in part at
least, from the cells which ordinarily form Bidder’s organ. Since
in this individual the testes are somewhat shorter than those usually
found in adult males, it seems probable that primordial germ-cells
in the anterior part of the germinal ridge, which normally would
have developed into spermatozoa, joined with the cells of Bidder’s
organ to form these rudimentary ovaries. The force, whatever its
nature, which modified the development of the cells and brought
about the formation of these rudimentary ovaries must have acted

Anomalies in the Genital Organs of Toads. 169

at a very early period in the life history of the individual, as in a
normal toad the cells which develop into Bidder’s organ begin to
show characteristics which distinguish them from the other germ-
cells when a tadpole is about two weeks old.

Sections show that each ovary contains a well defined central
cavity which is lined by epithelial cells, and that the structure
of the ovarian wall is the same in each case. In both ovaries the
great majority of the ova are practically of the same size and
in about the same stage of development. It is not possible, there-
fore, to trace the various stages in the growth of the cells or to dis-
cover any apparent reason for their unusual mode of development.
With but a few exceptions the cells in both ovaries have developed
uniformly along the same lines, and those in the upper part of each
ovary bear no more resemblance to the cells which are typical of
Bidder’s crgan than do the ones lying more posteriorly which were
derived, presumably, from primordial germ-cells belonging to the
sex-gland proper.

The smaller ova, which usually le at the periphery of the ovaries,
are rounded in outline, and they have an average diameter of 0.14
mm. As a rule, the cytoplasm of these cells appears uniformly
granular, although sometimes it contains yolk nuclei, as shown in
Fig. 19, Y. N. The nuclei are round, or slightly oval, and they
measure about 0.08 mm. in diameter; with but few exceptions, all
of them are in the early post-synizesis stage of development (Fig.
22). I can detect nothing in the structure of the great majority of
these small cells that would in any way serve to distinguish them
from normal young odcytes of the same size.

In some few cases the smaller cells of these rudimentary ovaries
exhibit all of the characteristics of the young ova normally found in
Bidder’s organ. A section of a cell of this type is shown in Fig. 19.
The nucleus contains a number of nucleoli of various sizes and
chromatin threads which are composed of a series of deeply stain-
ing, rounded granules. Two of the larger nucleoli show degenerative
changes that are similar to those taking place in the large nucleoli
of the cells of Bidder’s organ which are beginning to disintegrate.
The body of the cell contains a number of finely granular masses,

170 Helen Dean King.

sharply distinct from the cytoplasm, which stain very intensely with
iron hematoxylin (Fig. 19, Y.N.). These are the so-called “yolk-
nuclei” which are always present in the normal ova at certain stages
in their development, and which sometimes appear in the young
cells of Bidder’s organ. The arrangement of the yolk-nuclei in
the cell shown in Fig. 19 is very similar to that of the yolk-nuclei
in the cells of Bidder’s organ. I have not been able to find any
cells in these rudimentary ovaries that might be considered as inter-
mediate in structure between normal and rudimentary ova.

The larger cells in the rudimentary ovaries border the central
cavity, and, owing to pressure, they are usually greatly distorted in
shape. The average diameter of these cells is 0.6 to 0.7 mm.; while
the diameter of their nuclei ranges from 0.17 mm. to 0.25 mm. As
a rule the nuclei of these cells resemble to a remarkable degree those
of young ovarian ova of the same size: they are rounded in outline and
contain a large number of scattered nucleoli; the karyoplasm appears
uniformly finely granular; and the chromatin threads have the
feathery structure characteristic of the chromosomes in the young
ovarian odcytes (Fig. 25).

The structure of the cell body in many of these larger cells differs
considerably from that normally found in the ovarian odcytes of
the same size. As a rule the cytoplasm is much vacuolated, as if
degenerative processes had already begun in it. In some cases the
vacuoles extend radially from the periphery of the cell to the nucleus,
giving the eytoplasm a striated appearance when examined under a
low magnification; in other cases the vacuolated area extends only
around the periphery of the cell and the cytoplasm surrounding the
nucleus appears uniformly reticular (Fig. 23). There is a possi-
bility that the vacuolization of the cytoplasm in these cells is due,
in part at least, to the way in which the material was preserved.
Flemming’s fluid, although an excellent fixing agent for the testes
at, all stages of their development, is but a very indifferent fixative
for amphibian ova after they have passed the synizesis stage. The
cytoplasm of these large cells would doubtless appear much less
vacuolated had some other fluid with greater powers of penetration
been used in fixing the ovo-testes.

Anomalies in the Genital Organs of Toads. 7a
ro)

Many of the ova contain large numbers of yolk spherules which
have formed, as in normal ova, at the periphery of the cell. In
some few cases I have found two distinct layers of yolk spherules ;
one lying at the periphery, the other half-way between the surface
of the cell and the nucleus (Fig. 24). In eases of this kind it is
probable that the inner layer of yolk spherules was derived from
yolk-nuclei which appeared in a ring midway between the nucleus
and the periphery of the cell and were there transformed directly
into yolk spherules. I have found one or two cases somewhat similar
to this occurring in the ovarian ova (King, 708).

This hermaphroditic toad was killed at the breeding season when
the germ-cells have become mature and the large ova in the normal
Bidder’s organ have reached the highest stage in development of which
they are capable. It seems probable, therefore, that the large cells
in the rudimentary ovaries have also attained their maximum devel-
opment, and that they would have undergone rapid degeneration
and absorption to give place to another generation of similar cells,
had the individual not been killed. This assumption seems the more
plausible since a number of the largest ova in the rudimentary
ovaries already show marked degenerative changes.

The processes of degeneration occurring in the nuclei of the large
cell found in these rudimentary ovaries are somewhat different from
those taking place in the large ova of Bidder’s organ, and they differ
also from the disintegration processes occurring in mature eggs that
for some reason have failed to leave the ovary. The first evidence of
degeneration is the migration of the nucleoli to the centre of the
nucleus where they lie massed together as shown in Fig. 20. At this
time ali of the nucleoli are rounded in outline and, with but few excep-
tions, they stain uniformly black with iron hematoxylin. The chro-
matin threads can sometimes be seen in the nucleus at this stage,
although they cannot be found at a later period owing, possibly, to
the fact that Flemming’s fluid does not fix the chromosomes well in
cells of this size. The next step in the degeneration of the nucleus
is shown in Fig. 20. Many of the nucleoli appear vacuolated, others
stain faintly and are evidently being dissolved. Only that portion
of the karyoplasm in which the nucleoli lie is uniformly finely

ie Helen Dean King.

granular at this time, elsewhere it contains numerous rounded
granules, staiming very intensely, that are much smaller than the
nucleoli and many times larger than the minute granules which
normally form the karyoplasm. In a later stage (Fig. 21) the
nucleus is filled with numerous short fibres composed of deeply stain-
ing, rounded granules which appear similar to those scattered through
the nucleus at the stage of Fig. 20. There is no finely granular
karyoplasm anywhere in the nucleus at this time, and the fibres,
as well as the remaining nucleoli, lie in an apparently fluid space.
The immense number of the large granules in the nucleus at this
stage of degeneration seems to preclude the possibility that these
bodies have been derived from chromatin or from the substance of
the relatively few nucleoli that have been dissolved. It seems prob-
able, therefore, that these granules have originated from the granular
karyoplasm, since their number increases in proportion as the minute
karyoplasmic granules disappear. At the stage of Fig. 21 follicle
cells and blood capillaries are beginning to enter the cytoplasm of the
cells to complete the processes of disintegration and absorption. The
nuclear membrane breaks down at or soon after the stage shown in
Fig. 21, and the nuclear contents come in direct contact with the
cytoplasm. Unfortunately the rudimentary ovaries contain no later
stages in the degeneration of the large ova.

The granular fibres which fill the greater part of the nucleus at
the stage of degeneration shown in Fig. 21, bear a very striking
resemblance to the “oxychromatin” fibres found in connection with
the nucleoli during the early post-synizesis stages in the development
of the young odcytes. It is possible, therefore, that the latter struc- |
tures are not composed of chromatin but of fused karyoplasmic
granules which have great affinity for the chromatin stains. If this
interpretation is correct, then the chromatin in the amphibian egg
is probably not concerned in any way with the formation of the
nucleoli which are doubtless waste products of nuclear metabolism.

Cerruti, ’07, has recently given a brief description of two cases
of hermaphroditism which he has found in Bufo vulgaris. In one
individual the anterior part of each testis had developed into a small
rudimentary ovary lying between Bidder’s organ and the testis proper:

Anomalies in the Genital Organs of Toads. 173

this case is similar to that found by Spengel, ’76, in Bufo cinereus.
In the other individual Cerruti found that each Bidder’s organ had
developed into a rudimentary ovary which contained many large ova
in various stages of degeneration and also ova that appeared like
normal odcytes except that they did not have any yolk spherules.

In the second cases noted by Cerruti the condition of the sex-
glands is much like that in the hermaphroditie Bufo lentiginosus
described above. In both of these individuals it is evident that the
rudimentary ovaries were derived from cells which normally would
form a Bidder’s organ. In young toad tadpoles, as shown in a
previous paper (King, 08a), the cells which are destined to form
Bidder’s organ are directly continuous with the primordial germ-
cells that later form the sex-gland; they are similar to the germ-cells
in structure, and they develop like the germ-cells up to the synizesis
stage. These facts seem to me to prove conclusively that the cells
which form Bidder’s organ are degenerate germ-cells. It is not sur-
prising, therefore, that occasionally these cells develop into ova which
apparently have a normal structure.

Many instances of hermaphroditism have been recorded for various
species of frogs. Cases similar to those found in Bufo, where a
rudimentary ovary has developed at the anterior end of a testis,
have been described by Marshall, ’84, Kent, ’85, and Ridge-
wood, °88; the reverse condition, with the ovary below the testis,
has been found by Bourne, ’84, in Rana temporaria. Marshall,
and also Smith, *90, have reported cases in which one of the sex-
glands in an individual was an ovary and the other a testis. This
latter form of hermaphroditism is extremely rare among amphibians,
and it has not yet been found in Bufo. Evidently hermaphroditism
occurs much less frequently among the Urodela than among the
Anura, as only two cases have as yet been reported for this group
of amphibians. La Valette St. George, 93, has given a_ brief
description of a case of hermaphroditism in Triton teeniatus, and
Knappe, ’86, has noted the presence of a Bidder’s organ in a young
salamander; neither investigator gives any details regarding the
structure of the ovo-testes in these forms. ;

Abnormalities of the kind described above, whether they occur

174 Tlelen Dean King.

in the sex-glands of young or of adult amphibians, seem explicable
only on the assumption that the primitive amphibians were herma-
phroditic, and that in the course of their evolution this primitive
hermaphroditic condition has given place to the bi-sexual condition
found at the present time. Although a rudimentary sex-gland (Bid-
der’s organ) is found only among the Bufonide at present, traces
of the primitive hermaphroditie condition still exist in other forms,
as is shown by the fact that in some individuals the genital glands
contain both ova and spermatozoa. Such individuals, however, are
only rudimentary hermaphrodites, since apparently only one kind
of germ-cell ever becomes functional. The spermatozoa are prob-
ably a much more specialized type of cell than the ova, and it might
be expected, therefore, that the male germ-cells would assume the
characteristics of ova more frequently than that the female germ-
cells would be able to develop into spermatozoa. ‘This is doubtless
the reason that, with but very few exceptions, all of the anomalies
found in the sex-glands of adult amphibians occur in individuals
which must be considered as males since only the sperm-cells are able
to reach maturity.

LITERATURE.
Bourne, A. G.
1884. On Certain Abnormalities in the Common Frog (Rana temporaria).
Quart. Journ. Micr. Sci., Vol. XXIV.
CERRUTI, A.
1907. Sopra due casi di anomalia dell’ apparato reproduttore nel Bufo
; vulgaris Laur. Anat. Anz., Bd. XXX.
Cone, F. J.
1895. A Case of Hermaphroditism in Rana temporaria. Anat. Ang., Bd.
DiGi :
FRIEDMANN, IF.
1898. Rudimentiire Wier im Hoden von Rana viridis. Arch. mikr. Anat.,
Bd. LII.
HAECKEL, E.
1874. Anthropogene oder Entwickelungsgeschichte des Menschen. Leip-
21g.
Hertwia, R.
1906. Weitere Untersuchungen itiber das Sexualitiitsproblem. Verhandl.
deutsch. zool. Gesellsch.
1907. Untersuchungen iiber das Sexualitiitsproblem. III. Verhandl.
deutsch. zool. Gesellsch.

a

Anomalies in the Genital Organs of Toads. Wied

HOFFMANN, C. K.
1886. Entwicklungsgeschichte der Urogenitalorgane bei den Anamisia.
Zeit. wiss. Zool., Bd. XLIV.
KeEnT, A. F. S.
1885. A Case of Abnormal Development of the Reproductive Organs in
the Frog. Journ. Anat. and Phys., Vol. XIX.
KXInc, HELEN DEAN.

1907. The Spermatogenesis of Bufo lentiginosus. Amer. Journ. Anut.,
Vol. VII.

1908. The Odgenesis of Bufo lentiginous. Journ. Morph., Vol. XIX.
1908a. The Structure and Development of Bidder’s Organ in Bufo lenti-
ginosus. Journ. Morph., Vol. XIX.
ISNAPPE, EP.
1886. Das Bidder’sche Organ. JMJorph. Jahrb., Bd. XI.
Arr, OF EH:
1890. Abnormal Reproductive Organs in Rana temporaria. Journ. Anat.
and Phys., Vol. XXIV.
MarsHatt, A. M.
1884. On Certain Abnormal Conditions of the Reproductive Organs in
the Frog. Journ. Anat. and Phys., Vol. XVIII.
PFLUGER, E.
1882. Ueber die das Geschlecht bestimmenden Ursachen und die Ge-
schlechtsverhiltnisse der Frésche. Arch. gesammte Phys., Bd.
XXITX.
PUNNETT, R. C.
1900. Notes on a Hermaphroditic Frog. Annals and Mag. Nat. Hist.,
Vol. VI.
Ripcewoop, W. G.
1888. On an Abnormal Genital System in a Male of the Common Frog.
Anat. Anz., Bd. III.
Sr. GrorGE, LA VALETTE.
1895. Zwitterbildung beim kleinen Wassermolch (Triton taeniatus).
Arch. mikr, Anat., Bd. XLV.

ScHMITT-MARcEL, W.
1908. Ueber Pseudo-Hermaphroditismus bei Rana temporaria. Arch.
mikr. Anat., Bd. LX XII.
SmitH, W. R.
1890. A Case of Hermaphroditism in a Common Frog. Journ. Anat. and
Phys., Vol. XXIV.
SPENGEL, J. W.
1876. Das Urogenitalsystem der Amphibien. Arbeit. zoolog.-zootom.
Inst. Wiireburg, Bd. III.

EXPLANATION OF FIGURES.

All figures were drawn with the aid of a camera lucida. They have been
reduced two-thirds.

Fies. 1-16.—Outline drawings showing the various types of anomalies found
in the genital organs of young toads killed soon after completing their meta-
morphosis. C.A, corpora adiposa; B.O, Bidder’s organ; T, testis; O, ovary ;
N, kidney. 26.

Kia. 17.—Longitudinal section through the enlargement in the lower part of
the left ovary which is outlined in Fig. 10. > 250.

Fic. 18.—Transverse section through the enlargement in the lower part of
the right ovary which is outlined in Fig. 2. 250.

Fic. 19.—Section of an ovum found in the rudimentary ovary of an her-
maphroditic toad. This cell has all of the characteristics of the young ova
normally present in Bidder’s organ. Y.N, yollk-nuclei. >< 1000.

Fie. 20.—Section of the nucleus of a rudimentary ovum which is just
beginning to degenerate. < 333.

Fic. 21.—A later stage in the degeneration of the nucleus of a rudimentary
ovum. 333.

Fic. 22.—-Section of the nucleus of a young odcyte which appears normal.
>< 1000.

Fie. 23.—Part of a section of a large ovum showing vacuolization of the
cytoplasm. 3383. P

Fig. 24.—Part of a section of a large ovum having two distinct layers of
yolk spherules in the cytoplasm. > 338.

Fic. 25.—Section of the nucleus of a large ovum which appears normal.
< 333.

i it

ANOMALIES IN THE GENITAL ORGANS OF TOADS.
JIELEN D. KING.

THE AMERICAN JOURNAL OF ANATOMY.—Vo.L. 10,

No. 1, JAn., 1910.

st.

ANOMALIES IN THE GENITAL ORGANS OF TOADS.
HELEN D. KING.

H AMERICAN JOURNAL OF ANATOMY.—VOL. 10, No. 1, Jan., 1910.

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VoL. 10, No. 1, JAn., 1910.

BH AMERICAN JOURNAL OF ANATOMY.

THE ANATOMY AND DEVELOPMENT OF THE JUGU-
AR oy MPH SACS. INTHE “DOMESTIC, CAT
(FELIS DOMESTICA).

BY

GEORGE S. HUNTINGTON AND CHARLES F. W. McCLURE.

From the Anatomical Laboratories of Columbia and Princeton Universities.

Wirr 66 FricureEs.

CONTENTS.
TEER OMUNCEION RMP Gh rae ti eke thee Nicaee TO LS Renata se Petras Skee werk ae oe Ay 178
Nr ipo lMrmease tu alltex a MTEC cy. reese sae St cose ce oon ate each ees sea BLE eo uct ae ees 181
A summary of former investigations bearing upon the development of the
iceman alien, Toyo ee Naioayo) MEO sen one enon oe too nkeae ds adnan geeetod 5 o- 182

The retention of the embryonic jugular lymph sac as an adult structure and
the general character of the lymphatico-venous connections in the adult.. 188
On the establishment of the lymphatico-venous connections of the adult.. 190
Development of the veno-lymphatic plexuses and sacs of the early embryonic
stages and the establishment of the structural basis for the subsequent

development or the jucular lymph sacs. 2.200. oc -.s sree ss de ees 196

I General ground-plan of the embryonic venous area involved in the sub-
sequent development of the jugular lymph sacs........:............... 197

II Analysis of the developmental stages in the formation of the jugular
bymph sacs... .. nee eer een 6 2d Rp Sse PERN Noten aie oor ye 202
WetMMNOMOn Me ermMstenestratlon’. sees ces edias asi gee ace ens = levee ors 203

First period- of development, from the early venous stages up to the
establishment of the three primary veno-lymphatic plexuses .......... 208

Second period of development, from the establishment of the three primary
veno-lymphatiec plexuses to the attainment of the adult condition of the
OD erm ANAT) NSS Cine Testi ote sauce he ome Dee tees Gas ak PE eam eh ote 218
These two periods of development are illustrated in the “‘Analysis”’ by
the series of diagrams, figs. 8 to 21, inclusive.
III Detailed description of the individual stages based upon astudy of the

RECOM ICH MGaeay ta sigs ns eee AN oie he ager Ca. MDE IES Raenhelag als 222
MEET VaTenI OMS Sham CSu airs Ha secant: bic sResiels oto salt a ie Wace ocho ta ere coer 226
2. Intermediate early stages of pre-and posteardinal veno-lymphatic

GB ENyCellOyOMAKEATLE BS Sel a eee ee CREME Rime RS Or eg sce ns Ee a ne Aa 233
3. Subsequent early stages illustrating veno-lymphatic development... 254

4. Two embryos illustrating the lateral displacement of certain veno-
lymphaties which are secondarily separated by fenestration from
the caudal division of the ventral veno-lymphatic plexus ....... 270

_ THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 2.

178 George 8S. Huntington and Charles F. W. McClure.

ContTents—Continued
5. Fully developed veno-lymphaticstage...................:tee- veces 277
6:. Pre-lymphatie staize ic: a ite wee Sater tern ooh cites ced hat Oe eee 288
7. Lymphatic stage or stage of definitely organized jugular lymph sac. . 297
LV General considerations and conclusions... 05.222. a ene ree 307
Explanation of -firuresw sss. sek de nee ee ae ee ee ee 308

INTRODUCTION.

In January, 1908, the writers read a paper before the Associa-
tion of American Anatomists upon the ‘‘Anatomy and Develop-
ment of the Jugular Lymph Saes in the Domestic Cat.’’ This
paper, which includes merely an outline of the development
of the sacs was subsequently published in the Proceedings of the
Association.’

In dealing with this subject in the present paper 1% 1s the inten-
tion of the writers to give a minute and detailed account of the
development of these sacs in the domestic cat so that every phase
and condition of their development may be represented and illus-
trated by a series of reconstructions, which are as accurately repro-
duced as their complicated character will permit. The object of
presenting so detailed an account is, to place beyond the range of
inference all doubt that has hitherto existed regarding the venous
origin of the jugular lymph saes, as well as to establish a basis
for the study of the development of this portion of the lymphatic
system in other mammalian embryos.’

In a more recent article on the “Cervical Veins and Lymphatics
in Four Human Embryos” (Am. Jour. of Anat., vol. 9, page 38),
Dr. Lewis states as follows concerning the first appearance of
lymphatics in human embryos:

No lymphatics could be found in a 9.2 mm. embryo, so that the jugu-

lar lymphatics probably arise in human embryos of about 10 mm. This ,

‘Huntington and McClure: The Anatomy and Development of the Jugular
Lymph Saes in the Domestic Cat. The Anatomical Record, vol. 2, nos. 1 and 2,
1908 (pages 1-18, 17 figs.).

*In a paper by N. W. Ingalls (The Anatomical Record, vol. 2, no. 8, 1908),
dealing with the vascular system of a 4.5 mm. human embryo, certain vessels are
figured which open into the post and precardinal veins near their confluence to
form the duct of Cuvier. These are undoubtedly, as Ingalls suggests, veno-
lymphatic anlages of the jugular lymph sae, if judged by the corresponding
structures observed in embryos of the cat.

——

Development of the Jugular Lymph Sacs. 179

accords with the observation that they first appear in rabbits of 9.5 to
10 mm., but does not agree with Ingalls’ opinion that in a 4.9 mm.
human embryo certain vessels represent the first anlage, or earliest fore-
runners, perhaps, of the lymphatic system in man. The vessels in cues-
tion are clearly veins.

Unless the conditions in man differ widely from those in the cat,
it is evident to the present writers that Dr. Lewis has not as yet
identified the earliest venous anlages or forerunners of the jugular
lymph sac in human embryos, but has observed only the deriva-
tives of these anlages after they have become distinctly lymphatic
in character and when, by themselves, at this stage of development,
they present no direct evidence that they have been derived
from the veins.

In the course of our work on this subject, which has extended
over a period of four years, we have been constantly surprised by
the extraordinary variability and complexity which characterize
the detailed development of the jugular lymph sacs, and it has
only been possible to interpret these conditions after the study of
avery complete series of embryos, including a number of em-
bryos of approximately the same ages. It has also been neces-
sary to consider, in connection with the embryonic history of the
jugular lymph sacs, these structures and their variations in the
adult.

The primary principles underlying the development of the
jugular lymph sacs are (1) the development of a secondary chan-
nel parallel to the embryonic precardinal and the Cuvierian end of
the posteardinal; (2) the association with this secondary channel
of a certain number of dorsal precardinal tributaries, and (3) the
separation of these two sets of venous elements, which we have
termed “‘Veno-lymphaties,”’ from the main venous channels and
their subsequent conversion into the definite jugular lymph sacs
by a process of growth and fusion.

Although a general principle of development has not been
difficult to establish, it has proved, in some cases, a matter of the
greatest difficulty to determine the actual mode of origin of cer-
tain of the venous anlages (veno-lymphaties) of the jugular lymph
sacs owing to the variable manner in which these veno-lymphatics

180 George 8S. Huntington and Charles F. W. McClure.

develop in conjunction with the main venous channels and their
tributaries, as well as the variable manner in which they fuse
together. On account of the large number of reconstructions (71),
which we have made of the region in which the jugular lymph
sac is developed, we feel, however, that we are able to present a
fairly accurate account of all stages of their development.

Our observations have been based wholly upon the study of
wax reconstructions made after a slight modification of the method
of Born. The method of plastic reconstruction was alone used
by us for the reason that we found it the only one by means of
which it was possible to get every vascular element in the recon-
struction and to establish thereby a graded series of stages,
complete in all details, in which the vascular elements could
be studied and compared in their proper relations. Moreover, in
view of recent publications on the vascular system, it is necessary
to point out the self-evident fact that the only other available
extra-vitam method of studying the development of vascular ele-
ments, viz., by injection, will, if successful, only demonstrate
channels or spaces actually continuous with each other at the time
of the injection, but will completely fail in revealing vascular
spaces as yet independent of those injected, although subse-
quently a connection between the two may be established.’

We concluded at the beginning of our investigation that the
method of injection would prove inadequate as a means of deter-
mining the actual mode of origin of the anlages of the jugular lymph
sacs. That our conclusion was correct or not, the reader may best
judge for himself.

Most of the embryos studied by the writers were fixed in
Zenker’s fluid and the sections then stained on the slide with De-
lafield’s haematoxylin and Orange G. For fixation of tissues and
differentiation of vascular structures, this method has proved
most satisfactory.

Opposite is a list of the cat embryos studied and in part re-
constructed by the writers in connection with this investigation.

“See figs. 1 and 2in Dr. Sabin’s paper on The Lymphatic System in Human
Embryos. The American Journal of Anatomy, vol. 9, 1909, page 64.

Development of the Jugular Lymph Sacs. 181

LIST OF MATERIAL EXAMINED.

SOMITES SERIES | MILLIMETERS SERIES | MILLIMETERS SERIES MILLIMETERS SERIES
I eg wer 3 12 ie kG 12
13 86 8 Sox 12 97° .| 16 15
eeS.5 102 — be ao 100 | 16 95
Millimeters Scries| 8.5 T5A| 12 Zl 4 16 96
45 Baile 5 12.5 Boh. AG 222
5 134 9 elena ean Dt 16 224
5 ie 9 106 | 13 35 | 16 230
5 ATS 10) 136s" I> ES 92 | 16.5 197
5 + 30-40. -9:5 D Spale? 107 | 16.5 240
Bo ieee O s5 239 13.5 MG6-7|. 31% 23
5.18 (SBP) 10 Ee alia fe 189°") 17 36
5.6 110 | 10 79 2185 223 17 94
5.7 103° - |) 416 101 14 ese 142
6 CA t6 111 14 SAE 17 190
6 85. | <10 142-0}, 14 GALLI 17
6 115 10 ASI 14: 122 | 18 41520
6 116 | 10 TAN 14 1272 ule 19 87
6 i alltel 140 | 14 210 18 88
6 128 | 10 237 14 Fi Liltne Wels 10
6 rey 1083 81 14 212) 218.5 21
6.2 109 |- 10.5 GE D140 2A 199
6.5 130 | 10.5 120 | 14.5 eae a9 80
6.5 131 | 10.7 474(Har-| 14.5 38 | 20 83
6.5 129 vardEmbryological 15 53 21 242
6.5 186 (Collection). 15 53 | 22 16
6.8 LOS) 1d 6 | 15 "fs eae 22
7 iBSecbee th rey | a Oi» |i 25 44
if mater tl Fefh 15 216 25 147
7 adie bt Final abs FAS | 31 144
7 135 | 11 98 |-15 F19 -|- +34 168
7 11S ype ee 313. | 15 243 | 35 90
ier 108) 4) 125 29. | 15 244 | 37.5 99
2 119112 1 15.5 141 | 45 14
7.2 Pipi 24 | 15.5 143 | 51 104
7.25 Clee ees 55) sag 215
7.50 Sos ie 26 245 |

The measurement of each embryo was made after fixation. All
measurements of embryos referred to in this paper represent the
distance between the crown of the head and the baseof the tail
(Crown-rump measurement).

Series 1 to 74 belong to the Princeton Embryological Collection,

182 George 8. Huntington and Charles F. W. McClure.

and from series 75 on, with the exception of series 474, to the
Embryological Collection of Columbia University (College of
Physicians and Surgeons).

A SUMMARY OF FORMER INVESTIGATIONS BEARING UPON THE
DEVELOPMENT OF THE MAMMALIAN JUGULAR LYMPH SACS.

It is not the purpose of the present paper to give a complete his-
torical review, nor a statement concerning the differences of opin-
ion which exist at the present time in regard to the development
of the lymphatic system in general. Both of these subjects have
been fully considered by Huntington‘ and Sabin®, and can be
referred to by anyone interested in the matter.

It may be stated here, however, that all American anatomists
who have thus far investigated the development of the mamma-
lian jugular lymph sac, as distinguished from the general system
of lymphatic vessels, now agree that it is derived directly from the
venous system. It may, therefore, be of interest to examine the
evidence upon which this conclusion was originally based. With
the exception of the present writers, Sabin and Lewis, as far as we
are aware, are the only investigators who have studied the develop-
ment of the mammalian jugular lymph sacs.

Sabin’s® original statement concerning the development of the
lymphatic system, including the jugular lymph sacs, was as
follows:

The lymphatic system of the embryo pig begins as two blind ducts
which bud off from the veins in the neck. At the very start the openings
of these ducts into the veins are guarded by valves formed by the direction
which the endothelial bud takes as it grows from the vein. In the ducts
themselves there are no valves at first. From these two buds, and later
from two similiar buds in the inguinal region, ducts grow toward the skin
and widen out to form four sacs or lymph hearts and from these saes the
lymphatics grow to the skin and cover its surface (page 387, 1902).

‘The Genetic Interpretation of the Development of the Mammalian Lymphatic
System. The Anatomical Record, nos. 1 and 2, vol. 2, 1908, pages 19 to 45.

5Sabin, Florence R. Further Evidence on the Origin of the Lymphatic
Endothelium from the Endothelium of the Blood-vascular System. The Anatomi-
cal Record, vol. 2, 1908. .

6 The American Journal of Anatomy, vol. 1, 1902; vol. 3, 1904 and vol. 4, 1904.

Development of the Jugular Lymph Saes. 183

Sabin’s?7 recent statement on this subject is a retraction of her
former view and a complete acceptance of the work of Lewis,*
concerning which she writes as follows:

He (Lewis) showed that there is first a plexus of veins in the region of
the sac; these veins are in free connection with the jugular vein. Then
the plexus of veins is cut off from the jugular and appears full of blood,
but without venous connections. Later the plexus forms a sac and rejoins
the main vein. After it has joined the vein it becomes empty of blood.
This discovery of Dr. Lewis I can entirely confirm.

In view of her present position, that the anlages of the lymph
sacs are first cut off from and then later rejoin the veins, it appears
to the writers that Sabin’s original observations upon the develop-
ment of the jugular lymph sacs must have been made upon
embryos in which the lymph sacs had already been established, and
in which the secondary permanent connections with the veins
had taken place, since in all of the embryos injected she found the
lymph sacs to be in open communication with the veins. If a
connection between lymph sac and vein served as the only eri-
terion for inferring that the lymph sac is derived from the veins, it
appears to the writers, in view of her more recent statement on the
subject, that a similar and equally valid inference might have been
drawn by observing the conditions which prevail in the adult.
Considering our present knowledge of their development, it is evi-
dent, from the very nature of the case, that the injection method,
as used by Dr. Sabin, has not as yet given us the slightest clue as to
the actual manner in which the anlages of the lymph sacs are
derived from the veins, nor, in fact, any evidence at all that they
even possess & Venous origin.

The chief service thus far rendered by the injection method is
the determination of the extent to which the lymphatic develop-
ment has progressed in the body after the lymphatics themselves
have been established.

7 Loc. cit. p. 49. The Lymphatic System in Human Embryos. With a Con-
sideration of the Morphology of the System, in The American Journal of Anatomy,
vol. 9, 1909. ;

8 Lewis. F. T. The Development of the Lymphatic System in Rabbits. The
American Journal of Anatomy, vol. 5, 1905.

184 George 8. Huntington and Charles F. W. MeClure.

We now pass to a consideration of Lewis’ investigations upon the
development of the mammalian jugular lymph sacs.

Lewis is unquestionably the first investigator to furnish the
clue for the proper interpretation of the development of the
mammalian jugular lymph sae. In the summary of -his paper
(p. 110) he states that the lymphatic system begins along the in-
ternal jugular vein as a detached sac formed by the coalescence
of several venous outgrowths. This sac, after uniting with other
independently derived lymphatics to form a continuous system,
acquires a new and permanent opening into the vein near the
subclavian termination.

Lewis’ paper deals chiefly with a description of stages in which
the jugular lymph sacs have already been formed. Only two em-
bryos are figured and described by him in which there appears to
be any evidence whatever that the lymph sac is derived from the
veins, and, on the basis of these two embryos his above-mentioned
conclusions concerning the development of the jugular sac were
apparently formed. These are two rabbit embryos measuring
9.5 and 10 mm., respectively, in length and concerning which he
writes as follows:

In a rabbit of 13 days,9.5 mm., no lymphatics could be found. The
reconstruction, fig. 1, shows the veins along which the first lymphatics
are soon to appear. The internal jugular vein receives a great many small
branches. One of these, nearly parallel with the dorsal border of the
vein and wider than the others, opens into the vein at either end. It is
in relation with the third cervical nerve. From its position and appear-
ance it is believed that this branch of the vein becomes a lymphatic
vessel (p. 98).

The second reconstruction is a 10 mm. embryo of 14 days. In this
specimen a chain of lymphatic spaces has appeared along the internal
jugular and the dorsal root of the primitive ulnar veins. The most an-
terior segment of the chain extends back to the third cervical nerve. It
sends out short blind sprouts like a vein and contains many blood cor-
puscles. The partition between it and the jugular vein is very thin, and
at one point there is a suggestion of communication between the two, as
shown in the figure. No opening into the vein can be demonstrated,

9 Loe. cit.

Development of the Jugular Lymph Saes. 185

however. The second segment of the chain, proceeding posteriorly,
extends to the fifth nerve. It equals the internal jugular vein in diameter,
and is closely applied to its wall. Behind the third nerve it sends a blind
diverticulum around the ventral end of the dorsal body muscles, into
the deep subcutaneous tissue of the back. This diverticulum, not matched
on the opposite side of the embryo, contains blood which apparently
entered it from rough treatment in preserving the specimen. The third
segment of the chain, between the fifth and sixth nerves, seems to connect
with the root of the ulnar vein. This connection, however, lies in the
plane of section, and a thin intervening wall may have been carried away
in the process of cutting. A detached lymph space follows the dorsal
root of the ulnar vein. A small and somewhat questionable one, not
matched on the opposite side, rests against the superior vena cava between
the roots of the ulnar vein. The most significant structure found in
this embryo is a space filled with blood, which opens into the external
jugular vein near its junction with the internal jugular. This space lies
quite near the third segment of the lymphatic chain. On the opposite
side of this embryo, and in the following one, this blood-filled sac con-
necting with the vein appears to be replaced by a lymphatic space,
detached from the vein, but connecting with the chain (p. 99).

Concerning the next oldest stage he states that

Fig. 3, from an embryo of 14 days, 11 mm., shows the fusion of all the
lymphatics of the previous stage into one large sac which encircles the
external jugular vein. On neither side could this sac be seen to communi-
eate with the vein (p. 99).

Upon stages older than those mentioned above he made the
following observations concerning the relations of the jugular
lymph sacs to the veins:

In a 14.5 mm. rabbit (14 days, 18 hours) he found that

on the right side of the embryo, in one section (No. 476), a minute
orifice connected the sac and the vein. It was not in the position of
the adult opening between these structures, and was not matched on
the opposite side (p. 101).

In a 20-(21)mm. pig embryo he states that “no connection be-
tween the jugular sac and the veins could be detected”’ (p. 103).

186 George 5. Huntington and Charles F. W. McClure.

In a 15mm. cat embryo he observed that

in one section (266) a branch of the jugular sac may enter the innominate
vein a little anterior to the subclavian, but it is not clear that an actual
opening exists and none can be found on the opposite side (p. 103).

Finally, referring to a 21mm. rabbit of 17 days, he states that

the jugular lymph sae on the left side, except for an extensive rupture
does not connect with the vein. On the right, a pore is found leading
from the sac to the internal jugular vein near its union with the external,
but this also may be artificial. Thus in all the series of rabbits no bilateral
communication of the lymphatics and veins, in the position of the adult
openings, could be found. The pores, sometimes detected in various
positions, are not adequate to empty the large sacs, and may indeed be
artifacts. Communication with the veins in these must be by osmosis,
therefore, and the permanent outlets of the lymphatic system must
develop later (p. 109).

The above quotations include all of the data given by Lewis
which bear in any way upon the actual development of the jugu-
lar lymph sacs. On examining these data it can be seen that his
general statement, given in the Summary (p. 110), that the
lymphatic system of rabbits (jugular lymph sac) begins along the
internal jugular vein as a detached sac formed by the coalescence
of several venous outgrowths, covers the general ground of lymph
sac formation, but that his conclusions, while perfectly correct
in their bearing on veno-lymphatic development, are based on
insufficient observations. We therefore feel justified in making
the statement at the beginning of our paper that our object in
presenting so detailed an account is to place beyond the range of
inference all doubt that has hitherto existed regarding the venous
origin of the jugular lymph saes. Although Lewis has been justi-
fied in drawing this inference, had he examined a large number
of embryos of each stage, ranging between 6 and 14 mm. in length,
he would have, unquestionably, been able to establish a definite
venous origin for each of the segments of the lymphatic chain in
the 10 mm. rabbit, without expressing any doubts whatever.

Considering the fact that we now know the jugular lymph sac
to be derived from the veins, the question arises as to the develop-

Development of the Jugular Lymph Saes. 187

mental period in which we may regard these venous derivatives as
having assumed a lymphatic significance. Is it at a time when
the adult condition has been reached and the lymphatic system
begins to perform its destined function? Is it at a time, as in
Lewis’ 11 mm. rabbit (14 days) in which the venous anlages have
fused into two large sacs, containing no blood corpuscles, which,
in virtue of their complete separation from the main venous
channels, cannot, in the generally accepted sense, function in their
present condition in the lymphatic organization of the body? Is
it at a time, as in Lewis’ 10 mm. rabbit (14 days), in which his
chain of lymphatics, constituting the jugular lymph sac, contains
many blood corpuscles, and possibly communicates with the veins?
Or, finally, is it at a time, as in the 8.5 mm. cat and in stages
which precede it, in which the anlages of the jugular sacs are in
the process of being separated from the venous channels but are
still in communication with them?

The development of the jugular lymph sacs is not a haphazard
process, and their anlages, although derived from the embryonic
veins, are as definite in their character and mode of development
as those of other organs. If we are able to draw a distinction be-
tween thymus and pharyngeal tissue at any time before the thy-
mus anlages have separated from the pharynx and become trans-
formed into a functional body, we are likewise justified in draw-
ing a similar distinction between the lymphatic and venous
structures. It is not the purpose of this paper to enter further
into a discussion of this question of specificity of tissues, but
merely to point out that the claim, made by Dr. Sabin and others,
that no lymphatics make their appearance in the body before the
jugular sacs are formed, can best be answered by asking for the
definition of a lymphatic.

In a former contribution to this subject, the writers applied the
term “Veno-lymphatics” to all the venous anlages of the jugular
sacs, at a time when these anlages were filled with blood and in
free communication with the venous channels. In early stages
the only structural distinction that can be made between the
venous anlages (veno-lymphatics) of the jugular sacs and the
fully formed sacs themselves, is that the former, being in com-

188 George 8. Huntington and Charles F. W. McClure.

munication with thé veins, are filled with blood and appear to
function as veins, while the latter apparently do not.

THE RETENTION OF THE EMBRYONIC JUGULAR LYMPH SAC AS AN
ADULT STRUCTURE AND THE GENERAL CHARACTER OF THE
LYMPH ATICO-VENOUS CONNECTIONS IN THE ADULT CAT.

A detailed description of the variable conditions manifested by
the lymphatics of the neck region in the adult cat is a subject upon
which one of the writers (Huntington) is at present engaged.

The question concerning the fundamental character of the com-
munications which exist between the lymphatics and the veins
on each side of the body in the adult cat, as well as that concerning
the persistence of the embryonic jugular lymph sac in the adult,
have, however, claimed our attention. '

An examination of a large number of adult cats!® has shown
that a cemmunication between the lymphatics and the systemic
veins may normally occur, on each side of the body, at either one
of two or at two typical districts which correspond, approximately,
to the angles of confluence formed by the union of the internal
and external jugular" (common jugular angle) and of the com-
mon jugular and the subclavian veins (jugulo-subclavian angle),
respectively. We have found that neither one of these two angles
of venous confluence predominates as the place of communication,
but that either one of the two or both may serve in this capacity,
and for this reason both districts must be regarded as constituting
the normal points at which in the adult cat the lymphatics
communicate with the systemic veins.

The three forms of communication normally met with in the
adult cat are shown in fig. 1? (left side) in which a communication

10 Figs. 1, 2 and 3 are selected from a series of 180 lymphatic injections of the
adult cat (Nos. 13, 26 and 40), which form the basis of a forthcoming publication
on lymphatie variation in this species. They are reproduced here to define the
fundamental plan of the adult connections of the jugular lymph sacs with the veins.

11 'This vein, strictly speaking, is acommon jugular vein in the cat but on account
of its large size, as compared with the internal jugular, is usually spoken of as the
external jugular vein, of which the internal jugular is a tributary.

” All of the figures referred to in this paper are arranged in sequence on
plates and a description of the same, preceding the plates, may be found on
page 308,

Development of the Jugular Lymph Saes. 189

is alone present at the common jugular angle (Common Jugular
Tap); in fig. 1 (right side) and fig. 2. (left side), in which a com-
munication is alone present at the jugulo-subclavian angle (Jugulo-
Subclavian Tap) and in fig. 2 (right side) and fig. 3 (both sides)
in which a communication is present at both of these angles.

As will be fully deseribed in the following pages, the duplex
character of the communication in the adult finds its explanation
in the circumstance that the embryonic jugular lymph sac on each
side of the body develops two caudally directed processes (Jugular
and Subclavian Approaches), which are potentially capable of
establishing a communication with the veins at the common jugu-
lar and jugulo-subeclavian angles, respectively, and through which
either one of the two or both of the typical points of communica-
tion in the adult cat are invariably established.

The jugular lymph sac of the embryo, which is, relatively, a
very large structure, is in the adult much reduced in size and
extremely variable in form. Figs. 2 (right side) and 3 (both sides)
clearly illustrate the fact that the relations of the thyro-cervical
artery to the jugular lymph sac are the same in the adult as in
the embryo. This artery, a branch of the subclavian, lies ven-
tral to the caudal end of the jugular lymph sac, both in the
embryo and in the adult, and passes between the two typical
points at which the jugular lymph sac communicates with the
veins. As far as we are aware the persistence of the jugular lymph
sac asa distinct anatomical structure in an adult mammal has not
been hitherto recognized.

We must also protest against the assumption that the jugular
lymph sacs as a whole undergo subsequent transformation into
lymph-nodes. Since they receive on the one hand the main
systematic lymphatic trunks of the entire body, and on the other
empty into the venous system, they necessarily maintain their
lumen uninterrupted. In our series of 180 adult cats they
uniformly appear as distinct, more or less well-defined sacs.
Their walls are definite, firmer than those of the lymphatic
vessels which empty into them, somewhat thinner than the walls
of the veins, and often, in fully injected preparations, multilocular
or diverticular. Exactly the same conditions are presented in

190 George 8. Huntington and Charles F. W. McClure.

the adult human subject, and the investigations of MeClure and
Silvester show that this arrangement is found in a very large
series of mammalian forms. As far as our own observations on
the cat and in man are concerned, we have drawn the following
conclusions:

(1) The Saccus lymphaticus jugularis is a distinct adult
mammalian structure, connected on the one hand with the syste-
mic lymphatic trunks of the entire body, and on the other with
the systemic veins, and transmitting the flow of lymph from the
lymphatic into the venous system.

(2) Serving in this manner as the lymphatico-venous connect-
ing channel, the jugular sac maintains throughout adult life its
character as a receptacle of distinct and relatively large caliber,
and is not extensively involved in lymph node formation.

(3) The current anatomical descriptions of the termination
of the ‘‘thoracic duct’ in the ‘“‘jugulo-subclavian angle,’’ or,
according to variations, in the individual veins of this conflu-
ence, should be modified to correspond to the actual conditions
above described.

(4) It is scarcely necessary to call attention to the surgical
importance of the jugular sac in man. In cases of large and
multilocular sacs the structure might be easily wounded in opera-
tions involving the jugulo-subelavian region. On the other
hand, under these circumstances, it should be possible to close
the wound successfully by suture of the walls of the sac, without
interfering with lymphatic return through the thoracic duct and
the other tributaries of the sac.

ON THE ESTABLISHMENT OF THE LYMPHATICO-VENOUS CON-
NECTIONS OF THE ADULT.

One of the greatest difficulties encountered in our investigation
was the deterinination of the exact manner in which the perma-
nent lymphatico-venous connections are established in the adult.

Do the veno-lymphatic anlages of the jugular lymph saes under-
go a complete separation from the main veins and then, after
fusing to form the jugular sacs, secondarily rejoin the veins: at
either one of the two or at both of the points at which a communi-

Development of the Jugular Lymph Sacs. 191

cation may be found in the adult (jugulo—subclavian angle and
common jugular angle), or, do the two points of communication
found in the adult represent primary connections between the
main venous channels and the veno-lymphatic anlages of the
jugular sacs which have persisted throughout development?

There appear to be only two methods of determining these
questions, namely, by the study and reconstruction of serial sec-
tions and by the injection of the venous, veno-lymphatic and
lymphatic vessels in embryos of the appropriate stages. As far
as our observations on embryos of Felis domestica are concerned,
we are in the position to make the following definite statement
which is based on an exhaustive study of numerous series of
embryos.

In embryos between 10.7 and 14 mm., inclusive, (crown-rump
measurement) a very constant and definite picture is offered, in
strong contrast both with the free veno-lymphatic connections of
earlier stages, and with the definite and typical taps at the two
above-defined points which are encountered in the later stages and
which are present, one or both, in the adult. In this intermediate
condition, the caudal portion of the emptied sac appears to end
blindly in two processes (the Jugular and Subclavian Approaches).
These blind terminals extend to the immediate neighborhood of
the common jugular and jugulo-subclavian angles, in other words,
to those points at which in the earlier stages the last of the primi-
tive connections between the venous system and the lymphatic
sac persist for a time, and at which in later stages, after the estab-
lishment of the adult type, the wedge-shaped lymphatico-venous
entrance with valve formation is found. But, in the intermediate
stages now under consideration, the most careful scrutiny under
high power, of excellently fixed and stained series in large number,
fails to reveal any definite continuity whatever between the lymph
sac and the vein. We have here, therefore, a positive and appar-
ently uniform observation which, in judging the general question
at issue, cannot be disregarded, since one well-established fact
is of more value than any number of conjectures. This fact we
regard as definitely determined, within the natural limits which
are imposed by errors in observation and interpretation of a

192 George 8. Huntington and Charles F. W. McClure.

microscopical field. We have had no personal experience with the
injection method, as practised by Professor Sabin and others,
but have had the opportunity of seeing at a meeting of the Anatom-
ical Association a number of her series sectioned after injection.
The chance of extravasations of the injection fluid, coupled with
the further chance of injury in section-cutting, seems to us to
preclude the possibility of drawing definite conclusions by this
method as to the communication or non-communication of
venous and lymphatic spaces. The close approach in many in-
stances of the blind ends of the lymphatic sac to the angles of
venous confluence and the delicacy of the intervening tissue,
raises the possibility of a rupture occurring, to a probability, if an
injection were made in either direction. However valuable the
injection method may prove in determining the extent and fur-
ther growth of the lymphatic system when once established, we
have felt that it would not yield trustworthy results in determining
the problem now under consideration. On the other hand, in
view of the uncertainty of injections, it is evident that even a
negative result would not be conclusive and could not be accepted
as valid evidence that the lymphatic sae no longer communicated
with the venous system.

We are obliged, therefore, in the present state of our knowledge,
to confine ourselves to the above statement, viz., that, as far as
observation can determine, an intermediate stage exists in
which temporarily the lymph sac is completely separated from the
venous system, with which at a later period it makes secondary
connections which constitute the definite and permanent adult
lymphatico-venous taps.

As bearing somewhat on this question, the conditions noted
above in the adult cat (ef. p. 188) deserve further consideration.
While in many individuals both the common jugular and jugulo-
subclavian taps are present (fig. 3), others possess only a single
adult lymphatico-venous connection on each side (fig. 1), which
may be located at either one of the two typical points. We con-
sequently deal in these latter cases with instances in which one or
the other of the primitive taps (always present in the early veno-
lymphatie stages) is given up during the process of development

Development of the Jugular Lymph Saes. 193

and is not re-established. If oneof the original connections is thus
abandoned, it is at least evidence that both are not inviolable
lymphatico-venous junctions which persist throughout develop-
ment. It therefore appears quite as logical to suppose that, after
temporary complete separation of the lymph sac, in the aberrant
adults referred to, a secondary venous connection is established
at only one of the two points of election, as to argue that one of the
primitive taps persisted throughout in an unrecognizable form,
while the other was entirely lost.

The view that the lymphatico-venous communications of the
adult are secondarily established in the embryo was first advanced
by Lewis who arrived at his conclusion on the grounds that he was
unable to determine the presence of a communication on opposite
sides of the embryo (rabbit and pig) in the position of the adult
openings, and that in many instances no communication could be
detected at all.

McClure and Silvester!® have recently shown that the plan of
communication in the adult rabbit and pig is exactly the same as
that in the adult cat, in that it may occur on each side of the body,
at either one of two, or at two typical districts (common jugular
and jugulo-subclavian angles). Therefore, while the absence of
a symmetrical arrangement of communications on both sides of
the embryo is no longer proof that the communications are second-
arily established, the fact that no communications have been
detected at all in certain stages cannot be disregarded in favoring
this view.

Following is a description of the relations observed by the
writers which exist in the cat between the jugular lymph sae and
the veins in embryos measuring over 10.5 mm. in length:

In a 10.7 mm. embryo (Harvard Embryological Collection,
Series 474) no communication could be observed between lymph
sac and vein on either side except possibly at a point situated
slightly cranial to the jugulo-subclavian junction (right side, slide
K, section 194, and left side, slide E, sections 197-198). These
points of communication are extremely doubtful in character but

18 McClure and Silvester. A Comparative Study of the Lymphatico-venous
Connections in Adult Mammals. The Anatomical Record, vol. 3, no. 10, 1909.

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 2.

194 George S. Huntington and Charles F. W. MeClure.

are of interest on account of their bilateral arrangement as well
as for the reason that they correspond approximately in their
position to that of the primary connection which exists at an
earlier stage between the veno-lymphatic plexus and the base of
the primitive ulnar vein. If it is true that the sac separates
temporarily from the venous system, then it is quite natural that
pictures should be observed during the process of separation
and before the same is quite completed, which would lead to the
description of these “‘extremely doubtful’’ connections.

In a 11 mm. embryo (series 27) in which the jugular lymph sae
was established, no communication between lymph sac and vein
could be detected. Two processes of the lymph sac, however, ap-
proach and almost reach the two points at which the lymph sae
was formerly connected with the veins. In a former paper we
have termed these two processes the Jugular and Subclavian
Approaches, respectively.

In a 12 mm. embryo (series 78) no communication could be
detected on the right side. On the left side of the same embryo
however, the anterior end of the lymph sac was found to be in
wide-open communication with the internal jugular vein (anterior
tap of evacuation) but was completely separated from the veins
at all other points. This condition in an embryo of this age will
be further considered in connection with another topic.

In a 18 mm. embryo no communication between lymph sac
and veins could be detected on either side.

In a 14 mm. embryo examined by the writers a doubtful
communication was observed on the left side, situated slightly
cranial to the jugulo-subclavian junction (series 34, slide 21, sec-
tion 24). No communication could be detected at the junction
of the external and internal jugular veins.

In another 14 mm. embryo (series 37) a-tap was established at
the common jugular angle, but no connection was found at the
jugulo-subclavian confluence.

In the above sense, if the final connections are secondary
then, in the beginning of their establishment (14 mm.), stages
might be found which give incomplete and doubtful pictures of
lymphatico-venous connection when compared with the definite
and typical arrangement once fully established.

-

Development of the Jugular Lymph Saes. 195

Ina 15mm. embryo (series 75) the caudal end of the left jugular
lymph sae (subclavian approach) was filled with blood corpuscles
and communicated with the venous system in an unmistakable
manner at the jugulo-subclavian junction, by means of a wide
opening (slide 8, section 22, left side). A doubtful communication
was also apparent at a corresponding point on the right side, but
in neither case could one be detected at the junction of the external
and internal jugular veins. In another 15 mm. embryo (series 53,
slide 11, sections 23 and 24) the caudal end of the lymph sac was
in wide-open communication with the vein on both sides of the
embryo, at a point slightly in front of the jugulo-subclavian June-
tion. These connections in the 15 mm. embryo may be secondary
taps, fully established, but not yet invaginated into the typical
wedge-shaped entrance, and prior to the valve formation.

In a 15.5 mm. embryo (series 141) no indication of a communi-
eation could be observed on either side. In this case either the
measurement of the embryo may have been modified by varia-
tions in curvature, or the secondary connections belated.

From the 16 mm. embryo on, in which the thoracic duct. usually
forms a continuous vessel and communicates with the left jugular
lymph sac, we have found no difficulty in determining the pres-
ence of a lymphatico-venous connection or tap at either one of the
two, or at both of the points at which the lymphatic system com-
municates with the veins in the adult. The lymphatico-venous
connection or tap met with in these more advanced embryos
differs widely in character from that found in the 15 mm. embryos
as well as from that found in the preceding veno-lymphatic stages,
in which a wide-open communication may be present. The lym-
phatico-venous connection in these advanced embryos is formed
at each of the two typical adult points of entry by a wedge-shaped
process of the jugular lymph sae which is deeply invaginated into
the angle of confluence of two veins, or into one of the veins econ-
tiguous to the point of confluence, and which opens into the vascular
lumen by a narrow slit-like aperture bounded by a two-lipped
valve. This type of embryonic lymphatico-venous connection
was first demonstrated by the writers at the meeting of the

196 George 5. Huntington and Charles F. W. MeClure.

Association of American Anatomists in 1906" and is well illus-
trated by figs. 4, 5, 6, and 7 which are photomicrographs of
transverse sections of a 16 mm. cat embryo taken at the level at
which the external jugular joins the jugulo-cephalic trunk. Figs.
4, 5, 6 and 7 represent, respectively, sections 233, 237, 239 and
241 of series 15.

DEVELOPMENT OF THE VENO-LYMPHATIC PLEXUSES AND SACS OF
THE EARLY EMBRYONIC STAGES AND THE ESTABLISHMENT
OF THE STRUCTURAL BASIS FOR THE SUBSEQUENT DEVELOP-
MENT OF THE JUGULAR LYMPH SACS.

The general development of the jugular lymph sacs in the eat
presents four successive periods:

1. THE PRIMARY VENOUS STAGE, in which the ground-plan of
the primitive embryonic venous system is laid down.

These early embryonic venous channels appear to crystallize
out of an antecedent capillary network along definite hydrostatic
lines.

2. VENO-LYMPHATIC STAGE, In which, with reduction and further
definition of the venous channels, a portion of the same is sur-
rounded in certain areas by a secondary capillary network which,
together with a larger or smaller portion of the main embryonic
vein, is condensed into a uniform structure and separates from the
permanent venous channel as the anlage of the future jugular
lymph sae.

In view of the double relation which this structure sustains, on
the one hand to the embryonic venous system from which it is
directly derived, and on the other to the general lymphatic
system with which it establishes secondary connections, we have
designated this period as the Veno-Lymphatic Stage in the develop-
ment of the definite jugular lymph sac, and have described the
portions separated from the early venous channels as the Veno-
Lymphatic Plexuses or Sinuses, which form the anlage of the
final definite jugular lymph sac.

3. PRE-LYMPHATIC STAGE, in which the early veno-lymphatic
plexus becomes sac-like and evacuates its blood-contents, and

' f’untington and McClure. Figs. 14 to 17 inclusive, in vol. 2 of The Anatomi-
cal Record.

Development of the Jugular Lymph Sacs. 197

then appears to become temporarily detached from the venous
channels from which it arose.

4. SECONDARY OR DEFINITE LYMPHATIC STAGE, In which the evac-
uated sac becomes assigned to the lymphatic system, establishing,
on the one hand, connections with the independently developed
general systemic lymphatics, and, on the other, reéntering the
permanent venous channels by apparently secondary connections
made at typical points, thus forming the link which in the adult
unites the definitely organized venous and lymphatic systems.

It is desirable, for reasons which will appear in the presentation
of the subject, to deal with the individual problems involved on the
following basis and in the order indicated:

I. General Ground-Plan of the Embryonic Venous Area
Involved in the Subsequent Development of the Jugular
Lymph Sacs.

II. Analysis of the Developmental Stages in the Formation
of the Jugular Lymph Sacs.

III. Detailed Description of the Individual Stages.

IV. Summary and Conclusions.

I. GENERAL GROUND-PLAN OF THE EMBRYONIC VENOUS AREA
INVOLVED IN THE SUBSEQUENT DEVELOPMENT OF THE JUG-
ULAR LYMPH SACS.

It is necessary to consider in the first place the general struc-
tural ground-plan of the principal embryonic venous channels
concerned in this process, and to establish certain main divisions
of the same to which subsequent reference can be made in dealing
with the individual series.

The portion of the embryonic venous system involved in the
development of the jugular lymph sacs includes a large part of
the precardinal vein, the district of its confluence with the post-
cardinal vein to form the duct of Cuvier, and the proximal seg-
ment of the postcardinal vein caudal to this point.

This venous area can be mapped out into the districts shown in
the composite average schematic fig. 8 containing the following
subdivisions:

198 George 8. Huntington and Charles Ff. W. McClure.

Precardinal area

1. Arched Portion or Cephalic Arch of the Precardinal
Vein
2. Caudal or Straight Segment of the Precardinal Vein

Area of Pre- and Postcardinal Confluence and of Postcardinal Veins

3. Jugular Promontory and Duct of Cuvier
4. Primitive Ulnar Segment of Posteardinal

The definite connections between the fully developed lymphatic
and venous systems in later embryonic stages and in the adult fall
within these districts and occupy normally two typical and, within
certain limits, constant points:

1. Common jugular tap: At the angle of confluence of
internal and external-jugular veins.

2. Jugulo-subclavian tap: At the jugulo-subclavian angle
of confluence.

PRECARDINAL AREA.

1. The Cephalic Arch of the Precardinal Vein.

The precardinal vein begins as a strongly curved intracephalic
arch receiving a number of dorsal tributaries along its convexity
and a number of branches along its concavity.

A. Dorsal Tributaries of the Cephalic Arch.

The dorsal tributaries entering the cephalic arch along its
convexity vary in number and in mutual relations in individual
embryos 22.4 in embryos of closely related stages.

One large tributary of this group (A—B in the schematic figs.
8 to 16) is uniformly present and in later stages forms the direct
continuation of the caudal or straight segment of the precardinal
vein. .

In addition to this large tributary a number of smaller secondary
dorsal tributaries of the cephalic arch may be encountered both
cephalad and caudad of this main vessel (fig. 8, a’, b’, ete.).

Development of the Jugular Lymph Saes. 199

These additional tributaries (b’, b”, b’’’) caudad of the main
trunk of A-B are serially more or less in direct line with the set
of primary dorsal precardinal tributaries (1, 2, 3 and 4 in fig. 8)
entering into the subsequent formation of the veno-lymphatic
plexuses. As far as our observations extend, tributaries b’, b’’
and b’” do not generally enter into the composition of these
plexuses, although it is possible that in the later stages, in cer-
tain circumstances, they may be retained as secondary anterior
channels of communication between the fully established veno-
lymphatic sac and the precardinal vein.

The possible involvement of these tributaries (b’, ete.) of the
cephalic arch in the late veno-lymphatie stages, in connection
with the establishment of an anterior tap of evacuation is discussed
in dealing with the series concerned (ef. p. 292, series 78, fig. 57),

B. Ventral Tributaries of the Cephalic Arch.

In the early stages the concavity of the cephalic arch receives
a number of tributaries which temporarily drain a territory sub-
sequently drained by the permanent external jugular vein. The
vagus and spinal accessory nerves lie in close relation to these
branches, and in later stages these vessels anastomose around the
spinal accessory so that the nerve then penetrates the precardinal
arch from the medial to the lateral aspect through an oblique
foramen.

2. Caudal or Straight Segment of the Precardinal Vein.

The cephalic arch is continued caudad into a second nearly
straight segment of fairly uniform caliber. This portion of the
vein is the more important of the two, since both the dorsal cir-
cumference of the vein itself and its dorsal tributaries (1, 2, 3 and
4, in fig. 8) are involvéd in the subsequent development of the
veno-lymphatiec plexuses.

A. Dorsal Tributaries of the Caudal or Straight Segment of the
Precardinal Vein.

This segment may receive four distinct and separate tributaries
(1, 2, 3 and 4, fig. 8) or four distinct groups of tributaries, depend-

200 George 8S. Huntington and Charles F. W. McClure.

ing upon the developmental stage and individual variation. Of
these tributaries the one labeled 4 in fig. 8 is the most important,
not only on account of the extensive drainage area which it con-
trols, both caudad and cephalad of its precardinal terminal, by
means of its branches a and b, but also on account of the relations
which it subsequently maintains with respect to the development
of the anlages of the jugular lymph sac (caudal division of the
ventral veno-lymphatic plexus).

B. Ventral Tributaries of the Caudal or Straight Segment of the
Precardinal Vein.

This portion of the precardinal usually receives two main
branches, frequently joined at their terminal to include a fenestra.

AREA OF PRE- AND POSTCARDINAL CONFLUENCE.

3. Jugular Promontory and Duct of Cuvier.

At the confluence of the pre- and posteardinal veins the main
venous channel enlarges into a capacious swelling, which gives
rise ventrally to the duct of Cuvier, while it projects dorsally as a
rounded protuberance which we have designated as the Jugular
Promontory. This promontory constitutes an important region
in the future history of the veno-lymphatic and the definite lym-
phatic structures. Primarily it develops as the result of the con-
fluence of large venous trunks. It receives on its ventro-lateral
aspect the cephalic vein, and ventrally the anlage of the external
jugular which, after confluence with the cephalic vein, forms a
trunk of large size entering the ventro-lateral aspect of the promon-
tory.. The jugular promontory usually receives in later stages
the three or four well-defined dorsal tributaries (5S, 6S, 7S, and 8S),
in fig. 8, which enter its convexity along the dorso-medial aspect.
In the later stages the jugular promontory also receives dor-
sally the drainage from the body-wall and anterior limb-bud by
the primitive ulnar vein. The development of this vein will be
considered under the following topic The Primitive Ulnar Seg-
ment of the Postcardinal Vein.

—

Development of the Jugular Lymph Sacs. 201

In the early stages, the cephalic arch of the precardinal and the
drainage area of its caudal or straight segment is situated lateral
to the otocyst. Subsequently a network of anastomosing venous
channels develops on the medial aspect of this structure and fin-
ally forms the main intracranial channel. Tributaries draining
this area in part empty directly into the newly formed channel and
in part pass caudad to terminate in the dorso-medial aspect of the
jugular promonotory caudal to the entrance of the primitive
dorsal somatic branch 4 in fig 8. This change enables the caudal
or straight portion of the precardinal to give up, in the majority
of cases, the dorsal tributaries 1, 2, 3, and part or the whole of 4,
and to contribute them toward the development of the veno-
lymphatic plexuses. Consequently in certain stages the straight
sepn ei t of the precardinal vein may appear partially denuded of
dorsal somatic tributaries. Thesomatic vessels from 5S lo 7S or 8S,
inclusive, descending caudad, enter the dorso-medial aspect of the
promo.tory, while the dorso-lateral part of the promontory and
precardinal, cephalad of the promontory, as well as precardinal
tributaries 1, 2, 3 and 4, in whole or in part, are involved in the
veno-lymphatic development.

It is thus seen that the large size of the jugular promontory is
due to the number and increasing caliber of the trunks, which
progressively become confluent at this point, and that it represents
so to speak, the slack of the early embryonic jugular system
which permits, in the subsequent cardiac descent, the drawing
out of the redundant promontory into the elongated vessel of
later stages.

4. Primitive Ulnar Segment of the Postcardinal Vein.

Caudal to the entrance of dorsal somatic tributary 6S or 7S,
(6S, 7S, fig. 8) into the jugular promontory, and to the origin
ventrally of the duct of Cuvier, the postcardinal vein of the early
stages, in the area receiving the seventh or the eighth, to the
twelfth dorsal somatic branch (7S, 88S, fig.8), gives rise to a sec-
ondary parallel channel, the primitive ulnar vein. This vessel
develops along the dorsal circumference of the postcardinal by

202 George S. Huntington and Charles F. W. McClure.

condensation of a peri-venous plexiform capillary network. The
impetus to its formation is apparently given by the return circu-
lation from the anterior limb-bud through the marginal vein.
This vessel, in its rudiments, at first drains into the somatic
branches of the umbilical vein. With the diversion of the um-
bilical to the hepatic circulation and with the resulting transferal
of the somatic return to the posteardinal line, the primitive ulnar
appears as a secondary derivative from the latter, serving, before
the establishment of the subclavian vein, as the main channel
returning the blood from the anterior extremity. As such, it
develops from the capillary network surrounding the postcardinal
vein by condensation of the plexiform reticulum along the dorsal
circumference of the main vessel, and opens into the promontory
at the level of the sixth dorsal somatic tributary (6S in series
106, 9 mm. embryo, fig. 40).

The series of fenestre in the postcardinal vein (fig. 8), leading
up to the jugular promontory, indicate the line along which sub-
sequently the primitive ulnar vein, by further extension and con-
fluence of the fenestrae, continues to separate from the postcardinal
vein proper, until only its cephalic termination into the dorso-
lateral aspect of the jugular promontory is retained as its definite
point of connection with the embryonic venous system.

The part played by this vessel in the early drainage of the ante-
rior limb-bud, in the subsequent development of the dorsal
veno-lymphatic plexus, and its final relation to the adult type of
lymphatic organization, makes it one of the most important fac-_
tors in the evolution of the definite lymphatic system. The de-
tailed history of this vessel may therefore be properly considered
under the separate headings, dealing with the individual stages.

Il. ANALYSIS OF DEVELOPMENTAL STAGES IN THE FORMATION
OF THE JUGULAR LYMPH SACS.

As already stated, the jugular lymph sac, or anterior lymph
heart, develops in the embryo of the Domestic Cat as a direct
derivative of the early embryonic venous system. It arises from
certain of the dorsal tributaries of the precardinal and from the

Development of the Jugular Lymph Sacs. 203

main channel of the pre- and posteardinal veins, adjacent to
and including their point of confluence to form the duct of Cuvier.
Two distinct ontogenetic types are involved in the production of
this structure:

1. A certain number of dorsal somatic tributaries of the early
precardinal vein become dilated and fuse together, retaining at
first their primary connections with the main vein. This process
in the mammalian embryo, preliminary to definite lymphatic
formation, is strictly homologous to the development of the lymph
hearts in the lower vertebrates. Sala’s'! researches on the
development of the posterior lymph heart in the embryo of the
common fowl prove that this organ is developed entirely by dilata-
tion and fusion of the lateral branches of the first five coccygeal
veins.

In the mammalian embryo, and specifically in embryos of Felis
domestica, the first four dorsal somatic branches of the straight
or caudal segment of the precardinal vein are, to a greater or less
extent, involved in the formation of the jugular lymph sac, the
homologue of one of the lymph hearts of lower vertebrates.

2. Thus, while in mammals this phylogenetic type of lymph
heart formation is preserved, it plays but a secondary part in the
production of the jugular lymph sac. The greater part of this
structure, which in certain embryonic stages reaches a relatively
enormous development; as compared with its condition in the
adult, is directly derived from the main systemic veins by a proc-
ess, which we have called ‘“‘fenestration.”” By this designation we
do not mean to define a method of histogenetic formation of the
early embryonic blood vessels, but to accentuate, by the use of a
brief and serviceable term, the fact that in certain stages in the
development of the venous system, stages of the greatest impor-
tance in preparation for the appearance of the veno-lymphatic or
lymph heart division of the lymphatic system, the early embry-

16 Sala, Luigi, Ricerche fatte nel laboratorio de Anatomia Normale della R.
Universita di Roma: vol. 7, pp. 263-269, April, 1900.

204 George S. Huntington and Charles F. W. McClure.

onic pathways exhibit a typical and characteristic picture, best
defined by the term ‘‘fenestration.”’ The main lines of venous
drainage have already become fairly defined out of the antecedent
primary capillary network from which they arise. In the further
growth of the embryonic tissues secondary plexiform capillaries
surround the main vessel and become connected with the same.
By condensation of this plexus and enlargement of the interreticu-
lar spaces the principal embryonic veins appear in this develop-
mental phase as channels perforated by larger spaces or “fenestrae”
within the general confines of the main trunk. In a subsequent
stage confluence of these ‘‘fenestree’’ results in the more or less
complete separation of the primary vein into two parallel second-
ary channels, which appears to be a uniform principle in the devel-
opment of parallel venous trunks. Or, as in the case of the jugular
lymph sae, further extension of this same process may result in
separating from the main venous channel elements which unite to
form a closed sae entirely distinet from the vein from which it
arose.

* means, therefore, in the sense in
which it is employed in this paper, one of the last stages in the
definite crystallization of the venous system out of an indefinite
antecedent plexiform condition, and the determination of an
important element in lymphatic organization, closely associated
with the embryonic venous system. It does not mean the forma-
tion of lacuns or spaces in veins already fully established, but is
used by us as a convenient and short term to define the conditions
actually and frequently encountered in reconstructions of the
veno-lymphatie period. Thus the description of a portion of the
arly venous area ‘‘separating by fenestration’? from the remainder
to form a discrete and separate element is to be interpreted as
indicating in a few words the more complicated processes just
outlined. Such ‘‘fenestral separation’? may result in the estab-
lishment of a secondary vein paralleling the course of the main
channel, as in case of the posteardinal and primitive ulnar veins,
or, specifically, in the development of the jugular lymph saes_ the
areas involved ‘“‘separate by fenestration”’ from the vein proper to
constitute the anlages of that portion of the lymphatic system

The term ‘fenestration’

Development of the Jugular Lymph Saes. 205

which is derived, as a veno-lymphatic plexus or sac, or modified
mammalian lymph heart, directly from the veins. Some of the
general problems involved in the question of the early embryonic
venous organization have been recently considered by Clark,"
Evans,'? and by Schulte and Tilney,!® while the mechanical basis
of early embryonic vascular organization forms the subject of a
classical monograph by Thoma.!®

To return to the analysis of the development of the jugular
lymph sae we therefore find that along the dorso-lateral circum-
ference of the early pre- and posteardinal veins, in the appropriate
stages, the redundant embryonic venous channels appear perfor-
ated by a linear series of openings or fenestrae. At first these
fenestre are small, but subsequently they enlarge, elongate and
then adjacent fenestrae become confluent.

In this manner a segment of the originally uniform pre- and
posteardinal veins gradually becomes separated from the dorsal
circumference of the parent vein, and forms a secondary parallel
“‘para-precardinal” or ‘‘para-posteardinal’’ channel which, to-
gether with the above mentioned precardinal tributaries, consti-
tutes the anlage of the jugular lymph sac. To these venous deriva-
tives of the precardinal tributaries and of the main channels we
have, as above mentioned, given the name ‘‘veno-lymphatics.”’

One important condition develops as the result of this method
of formation, namely, the progressive increase, up to a certain
point, in the number of the communications which may obtain
between the veno-lymphatic anlages of the jugular sac and the
systemic veins. At first these communications in the precardinal
region are represented by the terminals of the three or four dorsal

16 Clark, Elliott R: Observations on Living Growing Lymphatics in the Tail of
the Frog Larva. The Anatomical Record, vol. 3, 1909.

17 vans, H. M. On the Earliest Bloodvessels in the Anterior Limb Buds of
Birds and their Relation to the Primary Subclavian Artery. The American
Journal of Anatomy, vol. 9, no. 2, pp. 281-319, May, 1909.

18H. von W. Schulte and Frederick Tilney: A Note on the Organization of the
Venous Return, with Special Reference to the Iliac Veins. The Anatomical
Record, vol. 3, no. 11, 1909.

19Thoma, R. Untersuchungen iiber die Histogenese und Histomechanik des
Gefiisssystems, Stuttgart, 1893.

206 George 8S. Huntington and Charles F. W. McClure.

precardinal tributaries. The process of fenestration not only
involves the portions of the precardinal vein intermediate between
the terminals of its dorsal somatic branches (1, 2, 3 and 4, in fig. 8.),
but also invades the terminals themselves (fig. 10) which usually
appear enlarged and dilated in a funnel-shaped fashion at their
points of union with the main trunk.

With the more marked separation of the veno-lymphatic plex-
uses from the main venous channels the interfenestral spaces
become drawn out into elongated channels which function as num-
erous secondary avenues of communication between the anlages
of the jugular sac and the systemic veins.

The process of secondary capillary perivenous formation lead-
ing to ultimate fenestration may first involve the main precardinal
vein and then extend into the dorsal tributaries of the vein, or it
may begin in the funnel shaped terminals of the tributaries them-
selves, and subsequently progress in them to such an extent that
the original single dorsal tributary becomes split up, or separated
into two or more components, depending upon the type of the
plexiform and fenestral development (fig. 11).

This separation of the dorsal tributaries of the precardinal vein
into a number of components is a fundamental process of veno-
lymphatic development, since certain of the elements, thus second-
arily derived, constitute, in part, the veno-lymphatic anlages of
the jugular lymph sac, while others retain the original character
of the early dorsal somatic tributaries, prior to veno-lymphatic
organization, and form the series of dorsal precardinal branches,
for the most part temporary, which lie in line with those opening
further caudad into the promontory and the postcardinal vein.

Prior to the detailed consideration of the development of the
jugular lymph saes it is advisable to determine definitely the
status of the dorsal precardinal tributaries and their share in the
development of the veno-lymphatic anlages of the jugular lymph
sacs.
Although the general principles underlying the development
of the jugular lymph sacs are the same for all embryos, the range
of individual variation is considerable. No two embryos of
approximately the same measurement, in the series examined by

Development of the Jugular Lymph Sacs. 207

us, agree in every detail, and even the same embryo often shows
variation and different degrees of development on opposite sides.

In the description of the individual series these special differ-
ences will be taken up in detail, but prior to their consideration it
is desirable to establish the general conditions and the genetic
principles involved in veno-lymphatic development.

A great part of the observed variability depends upon the fact
that the jugular lymph sac of the later stages (fig. 17) is preceded
by the formation of three primary veno-lymphatic plexuses
(fig. 12). Subsequently these unite to form the complete jugular
lymph sac (figs. 12 to 17).

In this process two of these plexuses first unite to form a common
ventral division of the sac, which then.fuses with the third plexus
placed dorsally, to constitute the common jugular lymph sac of
the later embryonic stages and of the adult.

We have consequently adopted the following descriptive ter-
minology:

I. Ventral Veno-Lymphatic Plexus.
A. Cephalic or Anterior Division.
B. Caudal or Posterior Division.
II. Dorsal Veno-Lymphatic Plexus.

These three veno-lymphatic plexuses are of primary importance
and represent a developmental stage intermediate between the
purely venous and early veno-lymphatic conditions, and the sub-
sequent period, in which the veno-lymphaties have fused to form
a common sac. (cf. figs. 12 and 46).

For clearness of description we will divide the account of the
development of the jugular lymph sac into two periods:

I. From the Development of the Early Venous Stage up
to the Establishment of the Three Primary Veno-
Lymphatic Plexuses.

A. Development of the Ventral Veno-Lymphatic Plexus.
1. Cephalic or Anterior Division.
2. Caudal or Posterior Division.

B. Development of the Dorsal Veno-lymphatic Plexus.

208 George 8. Huntington and Charles F. W. McClure.

II. From the Establishment of the Three Primary Veno-
Lymphatic Plexuses or Sinuses to the Attainment of the
Adult Condition of the Jugular Lymph Saes.

FIRST PERIOD.

2. FROM THE HARLY VENOUS STAGES TO THE ESTABLISHMENT
OF THE THREE PRIMARY VENO-LYMPHATIC PLEXUSES.

A. Development of the Ventral Veno-Lymphatic Plexuses.

In some instances the ventral veno-lymphatic plexus is largely
derived from the primary dorsal somatic branches 1, 2, 3 and 4,
while in others the main portion of the plexus differentiates by
plexiform condensation from the precardinal vein, carrying the
tributaries with it, in which case these latter play a secondary
part in the development of the plexus. Finally in a third group
both somatic and main precardinal components share nearly
equally in the formation of the plexus. Again one portion of the
plexus may be mainly precardinal in derivation, another largely
composed of elements contributed by the somatic tributaries.

Hence the ventral veno-lymphatic plexus, in both of its sub-
sidiary parts, exhibits a considerable range in variation, as will
appear in the detailed description of the individual series, and it
becomes necessary to analyze the two developmental processes
in detail in order to prepare the way for the consideration of the
conditions presented by any given case.

Moreover, in this general plan of development of the ventral
plexus, dorsal somatic precardinal tributary 4 and the associated
segment of the main precardinal vein take a share approximately
equalling that contributed by the remaining three (anterior)
tributaries (1,2 and 3) and by the portion of the precardinal vein
included within their area. This results in the establishment of
the two primary divisions of the ventral veno-lymphatic plexus,
viz:

1. <A cephalic or anterior division.
2. A caudal or posterior division.

eo

Development of the Jugular Lymph Sacs. 209

Thus, while the genetic principles involved in the formation of
the two divisions of the ventral veno-lymphatic plexus are iden-
tical, it will prove more serviceable to distinguish between the two
in the preliminary analysis, because certain special features in the
relation of dorsal somatic tributary 4 to the precardinal vein and
the promontory modify the picture of the development of the
eaudal portion of the ventral veno-lymphatic plexus and serve to
distinguish it in early stages from the cephalic division of the same.

We will therefore first consider the development of the cephalic
or anterior division of the ventral veno-lymphatic plexus, then
that of the caudal division, thirdly that of the dorsal veno-lym-
phatie plexus, and finally take up the mutual relations of these
three divisions prior to their fusion into the common veno-
lymphatic sac.

This can best be accomplished by following the veno-lymphatic
development on the hand of the series of diagrams shown in
figs. 8 to 13 which, with the exception of fig. 11A, represent in
schema lateral views of the reconstructions of the left side in
profile, but which answer equally well for the conditions found on
the right side. Fig. 11A represents a dorsal view of fig. 11 in the
region of the jugular promontory.

1. Development of the Cephalic or Anterior Division of the
Ventral Veno-Lymphatic Plexus.

The formation of this portion of the ventral veno-lymphatic
plexus can be analyzed by considering the two main types of
development which it exhibits, with the understanding that in
individual instances one or the other of these types may predomi-
nate, or that they may share equally inthe production of this
portion of the ventral plexus.

First Mode of Development. In this type the cephalic part of
the ventral veno-lymphatic plexus is largely developed from the
three anterior dorsal somatic branches of the precardinal vein.

The early precardinal tributaries 1, 2, and 3 in fig. 8, enlarge
at their terminals in a funnel-shaped manner, and these expanded
bases present a more or less distinct plexiform or fenestrated ap-
pearance (1, 2, and 3 in fig. 10).

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 2.

210 George 8. Huntington and Charles F. W. McClure.

By enlargement of these fenestrae or by confluence of adjacent
inter-reticular spaces, the terminals of the primary. tributaries
divide into two components, a lateral veno-lymphatic (VL) and
a medial somatic (S) element (fig. 11).

The lateral elements, (1VL, 2VL, 3VL, in fig. 11) shift to the
dorso-lateral circumference of the precardinal vein, the medial
elements (18, 2S, 38, in fig. 11), conversely enter the dorso-
medial aspect of the main channel.

At first these two sets of components are still connected by
transverse channels (figs. 11 and 18).

The best example of this stage encountered in our collection is
represented by Series 138 (ef. figs. 33, 34, 35 and 36 and the
diagrams, figs. 18 and 37).

Subsequently these tranverse connections are given up and then
the dorso-lateral elements enlarge and constitute the cephalic
division of the ventral veno-lymphatic plexus (1VL, 2VL, 3VL,
in figs. 12 and 19), while the dorso-medial branches (18, 2S, 38S
in figs. 11, 12 and 19) continue to function for a time as dorsal som-
atic tributaries of the precardinal, and are not further involved in
veno-lymphatic development.

After the dorsal somatic branches (1S, 2S, and 3S) have separ-
ated completely from the veno-lymphatic components (1VL,
2VL, 3VL in figs. 12 and 19), the latter may or may not fuse with
each other in the early veno-lymphatic stages. Usually, however,
the veno-lymphatic components of tributaries land 2 (1VL, 2VL,
in fig. 13), or of 2and 3 (2VL, 3VL,.in fig. 12) or of 1.2>angus
(1VL, 2VL, 3VL, in fig. 20) unite with each other so that their
individuality is lost and they then present a more sac-like ap-
pearance than in the preceding stages.

As a result of this fusion one or two of the originally separate
veno-lymphatic components give up their primary connection
with the precardinal vein (fig. 13 and 21). All of these variations
are not shown in the diagrams but can easily be followed in the
figures of the individual reconstructions.

Typical instances of the separation of the dorsal tributaries
into their veno-lymphatic and dorsal somatic components are
afforded by series 138, (figs. 33-37), while the three primary divi-

Development of the Jugular Lymph Saes. 211

sions of the veno-lymphatic plexus are well shown in series 102,
fig. 46.

Second Mode of Development. In this form the dorso-lateral
circumference of the straight segment of the precardinal, in the
area included between the levels of the first and third dorsal
somatic tributaries (1, 2 and 3, in fig. 8), retains the plexiform con-
dition and by continued growth and enlargement of the inter-
reticular spaces forms a multi-fenestrated appendage to the main
venous channel (cephalic division of ventral veno-lymphatic
plexus in fig. 9). By confluence of these fenestral spaces this
appendage is separated as a veno-lymphatic sac from the definite
vein except at its cephalic end, where one or two of the primary
connections between the veno-lymphatic structure thus formed
and the precardinal vein proper persist until a later stage, serving
as channels for the evacuation of the blood from the veno-lym-
phatic sac into the definite venous system, after the three primary
veno-lymphatic plexuses have fused to form a common sac. The
tributaries, 1, 2 and 3 (1, 2 and 3, in fig. 9) appear to be taken up
as a whole in this veno-lymphatic development and to lose their
individual character.

2. Development of the Caudal or Posterior Division of the Ven-
tral Veno-Lymphatic Plexus.

This portion of the ventral veno-lymphatic plexus is also formed
in one of two ways:

1. From part of primary dorsal tributary 4 (4, fig. 8), combined
with a portion of the precardinal vein which, between the early
terminal of 4 and the promontory, crystallizes out of the plexi-
form network of the primary vein along its dorso-lateral cireum-
ference as a “‘para-precardinal channel”’ (figs. 8, 9 and 10).

2. From the para-precardinal channel alone. In this case
tributary 4 is not involved directly in the veno-lymphatic process
and remains in its entirety as a dorsal somatic branch secondarily
entering the promontory.

In the early venous stages (fig. 8), as mentioned above, the
fourth dorsal tributary of the precardinal (4) is by far the largest

212 George 8. Huntington and Charles F. W. McClure

and the most variable in form and composition of all the dorsal
branches of this vein. It is formed by the confluence of two trunks
(a and b) which, cephalad and caudad, follow the dorso-lateral
border of the neural tube, and it opens into the precardinal by an
enlarged and expanded base, frequently fenestrated, which re-
ceives a large single cephalic tributary (4S in fig. 8).

Preeardinal tributary 4 is on the border line dividing the series
of the preceding branches 1, 2, and 3, uniformly involved to a
greater or less extent in the development of the cephalic division
of the ventral veno-lymphatic plexus, from the series of per-
manent dorsal somatic tributaries of the promontory (5S, 6S, and
7S, in fig. 8). Consequently, while 4 shares with the anterior
branches 1, 2, and 3, the tendency to divide by enlargement of the
plexiform terminal into a medial somatic and lateral veno-lym-
phatic component, the tributary, as a whole, inclines in individual
cases in one or the other of these directions, i. e., either toward
the more complete veno-lymphatic or the dorsal somatic type of
development. The genetic characters of 4 are more marked, com-
pared with the preceding branches 1, 2, and 3, by reason of its
large size and extensive drainage area. Certain instances, or
rather certain developmental stages (cf. series 188, figs. 35, 37, 18
and 114A), offer striking pictures of the separation into a dorso-
lateral veno-lymphatic (4VL) and a dorso-medial somatic compo-
nent (4S), connected by a number of transverse anastomoses.

Further, in the same sense, the entire caudal division of the
ventral veno-lymphatic plexus is interposed between the ceph-
alic division of the same and the dorsal veno-lymphatic plexus,
which is developed entirely from the promontory by condensation
of the plexiform dorso-lateral cireumferemce of this segment of
the main venous channel, without involving the promontorial
dorsal somatic branches (5S, 6S, etc.).

Hence the caudal division of the ventral veno-lymphatic plexus
of the early stages inclines in some instances toward the develop-
mental type of its cephalic associate, i. e., it involves dorsal
somatic tributary 4 to a greater or less extent. In other cases
it follows more closely in its development the succeeding dorsal
veno-lymphatic plexus, i. e., it is derived largely or entirely from

1s)

Development of the Jugular Lymph Sacs. 21

the circumference of the main vein, by further extension of the
para-precardinal channel, while in a third group both the pre-
cardinal vein and tributary 4 share more or less equally in fur- .
nishing the elements which enter into the composition of the
caudal division of the ventral veno-lymphatie plexus.

Consequently an analysis of primary somatic tributary 4 and
of the related precardinal segment, in reference to veno-lymphatic
development, will give the clue to the interpretation of the various
conditions encountered in the detailed study of the individual
series.

In the early stages the blood current of tributary 4 (4 in fig. 8),
empties by a dilated terminal into the straight segment of the
precardinal.

Subsequently, a secondary channel (para-precardinal channel)
is separated by condensation of plexiform areas of the main vein
from the latter between the terminal of 4 and the promontory.
This secondary “para-precardinal channel”’ (figs. 9 and 10) now
drains the blood current of tributary 4, originally emptying into
the precardinal, for the greater part directly into the promontory.
The dilated and frequently fenestrated terminal of 4 and the para-
precardinal channel, secondarily segregated from the main _pre-
cardinal line, undergo veno-lymphatic development and form the
structural basis for the further growth of the caudal division of the
ventral veno-lymphatic plexus.

In this process as stated above, tributary 4 may be largely
taken up in the veno-lymphatic organization, or the latter may
chiefly involve the main precardinal channel, leaving 4 to main-
tain its somatic character.

Hence in general the caudal division of the ventral veno-lym-
phatic plexus presents the two main genetic types:

1. Derivation from the ¢.ra-precardinal channel and the
veno-lymphatic component of 4, leaving the dorsal somatic
element of 4 as the first of the promontorial dorsal somatic
branches.

2. Derivation entirely from the para-precardinal channel,
without involving any portion of tributary 4, which

214 George 8. Huntington and Charles lk’. W. McClure.

remains intact as the first somatic promontorial
tributary and does not participate in veno-lymphatic
development.

The two main genetic types observed by us in the development
of the caudal division of the ventral veno-lymphatiec plexus, in
embryos between 7 and 10 mm., may be presented schematically
in the series of diagrams figs. 8 to 14 (lateral views, left side) and
in figs. 11A, 18 and 19 (dorsal views).

1. Deriviation from para-precardinal channel and veno-lym-
phatic component of precardinal tributary 4.

Early Venous Stage (fig. 8).

Tributary 4 of the precardinal series is marked by its large size
and extensive area of drainage, and by its expanded plexiform
terminal.

Intermediate Stage.

The beginning of veno-lymphatic organization in general is indi-
cated along the line of the primary dorsal precardinal tributaries
by expansion and reticular fenestration of their terminals. In
the area controlled by precardinal tributary 4, the precardinal
begins to differentiate along its dorsal circumference a sec-
ondary para-precardinal channel (figs. 9 and 10).

Early Veno-lymphatic Stage.

The precardinal tributaries, in general, have begun to separate
into dorso-medial somatic (1S, 2S, 3S, and 4S), and dorso-lateral
veno-lymphatic components (1VL, 2VL, 3VL, and 4VL, in figs.
11, 11A and 18).

The para-precardinal channel, having, for the most part,
assumed the function of draining the territory formerly controlled
by the primary precardinal terminal of tributary 4 (fig. 8), has
become associated with the veno-lymphatic component of 4 (4VL),
which in turn is in process of multiple fenestral separation from
the somatic element of 4 (4S). Cf. fig. 114 which is a dorsal view
of fig. 11 in the region of the promontorio-precardinal angle and
fig. 18, alsoa dorsal view, which shows this process of separation ata
more advanced stage of development. —

Development of the Jugular Lymph Sacs. 215

Later Veno-lymphatic Stage (figs. 12, 18, 14 and 19).

The separation of the veno-lymphatie (lateral) and somatic
(medial) components of tributary 4 has been completed. Each
component now obtains a separate and independent entrance into
the main venous channel.

The para-precardinal channel has further separated from the
main vein and has fused with the veno-lymphatic element of tribu-
tary 4(4VL) into asingle sac (figs. 13, 14and 19), to form the caudal
division of the ventral veno-lymphatic plexus which is now no
longer connected by transverse anastomoses with the somatic
portion (48) of tributary 4.

The above outlined plan of development for the caudal division
of the ventral veno-lymphatic plexus, appears to be more or less
uniform in character for cat embryos, in general, in which the
para-precardinal channel and tributary 4 combined, enter into
the formation of this plexus.

2. Derivation entirely from the para-precardinal channel,
without involving any portion of tributary 4.

This condition is illustrated by series 2, fig. 32,and by the dia-
gram fig. 19.

In this case the caudal division of the ventral veno-lymphatic
plexus appears to be developed entirely by further growth and
enlargement of the para-precardinal channel, without involving
dorsal somatic tributary 4 which secondarily enters the promon-
tory independently of the para-precardinal channel, as the first
of the series of dorso-medial branches draining into this segment
of the venous system.

Here the caudal division of the ventral veno-lymphatiec plexus
follows absolutely the developmental path of the dorsal veno-
lymphatic plexus.

In connection with these two main genetic types of development
two conditions have been observed by us which are worthy of
special mention.

(a.) In one ease (fig. 20, diagram) the veno-lymphatic (4VL)
and somatic (4S) elements of tributary 4 separate completely by

216 George S. Huntington and Charles F. W. McClure.

fenestration in the usual manner. The latter (4S) then enters the
promontory as its first dorsal somatic branch. The former (4VL)
joins the veno-lymphatic para-precardinal channel, enlarging the
‘audal division of the ventral veno-lymphatic plexus and very
probably forming in part or in whole the rounded appendage of the
same noted in some of the later stages (fig. 14 and series 77, figs.
Sl and (52):

(b.) In another case (fig. 21, diagram), the caudal division of
the ventral veno-lymphatic plexus joins the cephalic division and
retains its early connections with the precardinal vein. The para-
precardinal channel separates from the promontory at its caudal
extremity, leaving the original point of connection with the main
vein as a prominent conical teat or projection, lateral and close
to the entrance of 4S into the promontory.

It is of course evident that in an analysis of this kind it is
practically impossible to distinguish always between clearly de-
fined types of development and stages fixed in individual embryos
which may or may not represent partial approximations to a single
common genetic ground-plan. In other words, it is impossible, in
studying any given series, to foretell the exact details of further
development which this particular embryo would have exhibited
if allowed to continue its course.

On the other hand, we feel justified in establishing the above
indicated gradations of the veno-lymphatic development by rea-
son of the very large number of embryos which we have critically
examined. We believe that this exhaustive study has enabled us
to present a concise composite account of veno-lymphatic develop-
ment to which each series subsequently described in detail can be
correctly referred, so that the unity and consistency of the funda-
mental plan underlying veno-lymphatic development in the cat
becomes apparent.

B. The Development of the Dorsal Veno-Lymphatic Plexus

This third component of the general veno-lymphatic plexus
is developed entirely from the main venous channels of the prom-
ontory and posteardinal, without involving the dorsal somatic
tributaries (5S, 6S, ete.), of this area.

Development of the Jugular Lymph Saes. 2i7

The dorso-lateral circumference of the promontory and_post-
cardinal carries a plexiform net work which, along the lines above
indicated (page 205), condenses by fusion and enlargement of the
interreticular spaces into a dorso-lateral component of the main
channel, representing the anlage of the dorsal veno-lymphatic
plexus. The result of this process is shown in figs.8, 9and 10, in
which the dorsal veno-lymphatic plexus appears as a dilated
sac, extending cephalo-laterad from the promontory over the
junction with the precardinal and separated from the main sys-
temic channel by a linear series of fenestree. This plexus subse-
quently develops into the dorsal division of the general veno-
lymphatic plexus, and forms, in later stages, after amalgamation of
the veno-lymphatic components into the single jugular lymph sae,
the portion of the latter to which we have applied the term ‘‘ sub-
clavian approach. ”’

Figs. 8, 9, 10, 11 and 12 illustrate the general character and pro-
gressive growth of the dorsal plexus in the early veno-lymphatic
stages, and fig. 12 the drawing out of the interfenestral areas into
elongated channels, preparatory to the separation of the dorsal
plexus from the promontory.

The secondary capillary network in which the dorsal plexus takes
its origin from the main venous channel, extends caudad along
the promontory and postcardinal vein beyond the confines of the
dorsal veno-lymphatic plexus proper.

Thus the linear series of fenestrae shown in figs. 9 and 10, by
enlargement and confluence, finally separate a secondary channel
from the main posteardinal vein, which retains its cephalic con-
nection with the promontory near the entrance of the sixth or
seventh dorsal somatic tributaries (6S, and 7S, in figs. 11, 12 and
13). This secondary parallel vessel is the primitive ulnar vein,
serving as a temporary drainage channel for the anterior limb-
bud. Subsequently, in repetition of its own genesis from the post-
cardinal vein, a plexiform capillary network, extending along the
dorsal circumference of the primitive ulnar vein, condenses into
a parallel vessel which terminates in the caudal part of the dorsal
veno-lymphatic plexus. Hence we have in many of our diagrams
designated the area involved as the ‘‘anlage common to the primi-
tive ulnar vein and the primitive ulnar veno-lymphatice. ”’

218 George 8S. Huntington and Charles F. W. McClure.

In later stages this primitive ulnar veno-lymphatie becomes
secondarily connected with the systemic lymphatic vessels of the
anterior extremity, in the same way in which on a larger scale,
the thoracic duct secondarily taps the process of the jugular
lymph sae which forms its portal of entry into the same. The
partial separation of the common anlage of these two structures,
viz., primitive ulnar vein and primitive ulnar veno-lymphatie,
is shown in fig. 12. Their subsequent development can be followed
in figs. 13 to 17 inel.

SECOND PERIOD.

II. From THE ESTABLISHMENT OF THE THREE PRIMARY VENO-
LYMPHATIC PLEXUSES OR SINUSES TO THE ATTAINMENT OF
THE ADULT CONDITION OF THE JUGULAR LymPpH Sacs.

This period is characterized by the following well-marked char-
acters:

1. Further reduction of the multiple early connections between
the veno-lymphatic plexuses and the permanent veins, and the
consequent more complete separation of the former from the latter,
the plexuses or sac-like structures assuming a greater degree of
independence. ‘This general process of separation and loss of
early communicating channels appears to proceed from both
extremities of the area toward the jugular promontory where the
embryonie connections are longest retained and where the two
permanent adult communications between the lymphatics and
the veins are established. It is to be noted, however, that in
many observed cases the termination of one or more of the anterior
dorsal somatic tributaries persists as an open communication
between the cephalic division of the ventral veno-lymphatic plexus
and the anterior part of the precardinal, and in later stages even
enlarges considerably, forming the main veno-lymphatic-venous
connection and functioning as the anterior tap of evacuation
(cf. figs. 12 to 15, and series 78, fig. 57).

2. The three primary veno-lymphatic plexuses begin to fuse
with each other to form a common sac.

Development of the Jugular Lymph Sacs. 219

Figs. 12 and 13 indicate diagrammatically the origin of the veno-
lymphatic components of the jugular lymph sac above described
in detail, and their partial fusion to form the single jugular sac of
the later stages.

In the succeeding stage (figs. 14 and 15) the veno-lymphatic
plexus further condenses by amalgamation and fusion of the
primary dorsal and ventral divisions, and the communications
with the main venous channels are reduced typically to three:

(1) Tap A (figs. 14 and 15). The Anterior Tap of Evacuation.

(2) Tap B (figs. 14and15). At the Jugulo-promontorial Angle,
the future common jugular confluence, i. e., junction of internal
jugular vein (precardinal) with common trunk formed by union
of external jugular and cephalic veins.

(3) Tap C (figs 14 and 15). At the site of the connection of
the primitive Ulnar Vein with the promontory, corresponding,
approximately, to the future jugulo-subclavian junction.

The veno-lymphatic plexus as a whole is filled with blood and
the plexiform character of the capillary network from which it
arose is still manifested by the multiplicity of the fenestral spaces.
On transverse section the walls of the original vessels entering into
its composition still appear as partitions or septa which divide the
interior of the sac into a complicated system of interecommunicat-
ing channels. Later, these septa become greatly reduced in num-
ber and extent so that there results a capacious sac lined smoothly
by an endothelial layer, continuous with the intima of the veins.

This sac subsequently becomes emptied, the contents being
evacuated through the connections still persisting with the general
venous system (fig. 15, Taps A, Band C). This process of evacu-
ation occurs with great rapidity, so that in embryos of the appro-
priate stages (10.5 to 12 mm.), it frequently happens that the sac
on one side is completely emptied of its blood-contents, while on
the opposite side it is still well filled with blood. The evacuation
occurs normally at the above mentioned points, where the veno-
lymphatic sac retains longest its connection with the venous sys-
tem. On account of the small openings by means of which the
veno-lymphatie sac usually communicates with the veins at Taps
B and C, we have reason for believing that, in the majority of

220 George 8. Huntington and Charles F. W. McClure.

cases, the sac empties itself chiefly, if not entirely, through one of
the anterior connections (one or two of the primary dorsal somatic
tributaries) into the cephalic end of the straight segment of the
precardinal, or even that a secondary connection is established
between the anterior end of the veno-lymphatic sac and the poste-
rior extremity of the cephalic arch (through b,! ete., fig. 8), for the
purpose of serving as a portal through which the blood-contents
of the sac are poured into the permanent venous channels. (Com-
pare p. 290, description of series 78, 12m).

We have designated the connections thus longest retained be-
tween the veno-lymphatic sae and the veins, as the “‘ taps of evacu-
ation,’ because, after the sac is fully formed, they serve the pur-
pose of draining the blood contained in the sac, during the earlier
periods, into the permanent venous channels, so that after comple-
tion of this process the sac appears entirely empty of blood, lined
by endothelium, continuous through the taps of evacuation,
with the intimal lining of the large veins.

As the process of evacuation takes place with great rapidity
there is some difficulty in ascertaining, in individual cases, the
exact point at which it occurs. The taps of evacuation apparently
enlarge very much at this time, and, when the process is completed,
as rapidly close. It is therefore necessary to examine numerous
stages during this very short and evanescent period in order to
establish the actual conditions.

After the evacuation is completed the now fully organized
lymph sae separates, in the cat embryo, apparently completely
for a short time from the adjacent veins, by breaking away at the
evacuating point or points (fig.-16). This closed sac subse-
quently establishes two sets of secondary connections (fig. 17):

(a) With the independently formed systemic lymphatics.
(b) With the venous system.

These latter connections are normally formed at the two points
at which the primitive promontorial connections are, inthe major-
ity of instances, longest retained, viz., at the common jugular
confluence (common jugular permanent tap) and the jugulo-
subclavian angle (jugulo-subelavian permanent tap, fig. 17, and

Development of the Jugular Lymph Saes. pa

figs. 1,2, and 3). The latter may be double, with a larger dorsal
and a smaller ventral connection. The embryonic arrangement
tallies with the conditions observed in the adult, in which either
the common jugular, or the jugulo-subclavian tap, or both, func-
tion in individual cases as the chief portal of adult lymphatico-
venous connection.

After the secondary connections with the venous system have
become established, the jugular lymphsac decreases relatively
in size, while the systemic lymphatic channels increase propor-
tionately in extent and complexity. In the adult, therefore, the
jugular sac appears as a greatly reduced remnant of the extensive
embryonic sac and serves merely as the connecting link between
the systemic lymphatics and the venous system. In a few in-
stances portions of the early structure become invaded by ade-
noid growth and are thus partially transformed into lymph nodes.
The lumen of the jugular sac is, however, even in these cases
maintained, since it forms the channel of adult lymphatico-
venous entry.

The significance of the jugular lymph sae of the mammalian
embryo and the homology existing between it and the lymph
heart formation in lower vertebrates has been considered in a
separate communication?® by one of the writers.

Finally, a word concerning the relations of the thyro-cervical
artery and the first six spinal nerves to the veno-lymphatic
plexuses and the fully established jugular lymph sac.

The thyro-cervical artery at first passes forward from _ its
subclavian origin along the dorsal surface of the main venous
channel to a point in front of the venous arch formed by the
primitive ulnar vein as it enters the dorso-lateral circumference
of the promontory. Here the artery turns abruptly laterad on
to the lateral surface of the promontory where it divides into
branches (figs. 13). In the later veno-lymphatic and lymphatic
stages (figs. 14 to 17), the thyro-cervical artery still maintains

°° G. 8. Huntington, ‘‘The Genetic Interpretation of the Development of the
Mammalian Lymphatic System,’’ The Anatomical Record. Vol. 2, nos. 1 and
2, pp.» 19-45, 1908.

222 ~=George 8. Huntington and Charles F. W. McClure.

similar relations to the main venous channel and the jugular
lymph sac, lying ventral to that portion of the latter which is
derived from the dorsal veno-lymphatic plexus.

In the course of its development, a large foramen is formed in
the jugular lymph sac through which the first four spinal nerves
pass (SP. N.I-IV, figs. 15 and 16). The stages which lead up to
the establishment of this foramen are shown in figs. 13, 14 and 15.
At a later stage, the dorsal and ventral portions of the jugular
sac bounding the foramen, separate anteriorly and then connect
secondarily with the systemic lymphatics which are formed in-
dependently of the jugular sac (fig. 17) and not, as claimed by
Sabin, as the result of a centrifugal growth of the same.

The fifth spinal nerve (SP.N.V, in figs 13. to 17) also penetrates
the jugular sac, but through a separate foramen. This condition
is retained until a late stage of development.

The sixth spinal nerve (SP.N.VJ) does not penetrate the veno-
lymphatic plexus nor the jugular sac at any time. From its
origin in the spinal cord it at first passes ventro-laterad between
the primitive ulnar and posteardinal veins and contiguous to the
point where the primitive ulnar vein arches ventrad to open into
the promontory (figs. 18, 14 and 15). After the primitive ulnar
vein has given rise to the primitive ulnar lymphatic and has lost
its connection with the promontory, the sixth spinal nerve then
passes ventro-laterad and ventral to the primitive ulnar lymphatic
(figs. 16 and 17).

III. DETAILED DESCRIPTION OF THE INDIVIDUAL STAGES.
GENERAL CONSIDERATIONS.

The jugular lymph sacs of the cat present a considerable range ,
of variation both in the adult structure and in the details of
their development. This variability appears not only in different
embryos from the same litter, and possessing approximately the
same measurements, but even upon opposite sides of the same
embryo, just as in the adult the right and left sides frequently
show marked differences in composition and relations in the same
individual. Moreover, the individual embryos differ greatly in

Development of the Jugular Lymph Saces. 228

a chronological sense. In some, one portion of the veno-lym-
phatic system is well developed, while the remainder is retarded.
Ultimately, within certain limits, all embryos develop congruent
conditions, but owing to this variability as to the coincidence
of all the factors at a given period, it has been found necessary to
examine a number of embryos of about the same age, and to group
the vessels thus obtained in certain cases into a composite picture
which will serve as a guide for the proper interpretation of each
individual embryo falling within a given period, although all of
the developmental possibilities of this period may not be pre-
sented by each and every individual embryo in the series. Sucha
series of composite pictures has already been discussed in con-
nection with the preceding topic (figs. 8 to 21 inclusive.)

Of course, in addition to this general source of variability,
individual embryos present a certain range of personal and acci-
dental variation in development. Not only will individual
embryos show advanced development in certain areas, and retarded
conditions in others, but they also differ in the minor details,
such as in the number and arrangement of secondary tributaries
and their anastomotic conditions.

In order to facilitate the interpretation of the individual reconstruc-
tions figured in this paper, the reader is advised to refer each one to
the series of diagrams already described in the preceding pages
(figs. 8 to 21, inel.).

In the figures of the individual reconstructions the approxi-
mate derivation of the components of the jugular lymph sac
is indicated by the same color scheme as that adopted in the
introductory analysis for the diagrams, figs. 8 to 21, inclusive, so
that comparison of any given series with the corresponding
generalized developmental stage described in the analysis can
readily be made.

We have adopted this plan in order to emphasize the uniform
genetic type of lymph sac development in an extensive series of
embryos presenting the inevitable individual variations. Of
course the value assigned by us in this presentation to the indi-
vidual components in a given series is a question of personal inter-
pretation of the actual conditions offered by the embryo. This

224 George 8. Huntington and Charles F. W. McClure.

interpretation is, however, based on a vastly larger number of
embryos than those figured in this paper. Each of the series
quoted in our list of material has been carefully and completely
examined by one or both of the authors, and many reconstruc-
tions have been made in addition to those here published. We
consequently feel that the tentative value given by us to the indi-
vidual developmental components of the jugular lymph sacin the
embryos of the cat is based on sufficiently extensive observation
and comparison, and that we are warranted in offering an onto-
genetic history of the jugular lymph sae in which definite embry-
onic stages follow each other with remarkable uniformity in the
entire series and Jead uninterruptedly to the conditions established
in the adult.

On account of the very narrow limits of the venous system
within which the development of the jugular lymph sacs is con-
fined, it was found advantageous, in reconstructing certain stages,
to elongate the reconstruction by using thicker wax plates than
the #-measurement and magnification of the sections actually
called for. This was done with the purpose of separating, and
thus more easily determining, the arrangement and position of the
complex of veno-lymphatic components of the jugular sacs.

With these few exceptions, all of the reconstructions shown in
this paper have been reconstructed and drawn to scale. The few
that have not, however, illustrate equally well the main princi-
ples of veno-lymphatic development, but appear slightly more
elongated than the actual dimensions call for.

In the early stages the precardinal vein is, in conformity with
the more elongated form of the embryo, less curved in its anterior
or arched segment than in the later stages, and less distinctly
divided into the typical districts shown in the composite scheme
(fig. 8). The caudal or straight portion of the precardinal is
somewhat elongated, and presents, at more or less regular inter-
vals, oval or spindle-shaped enlargements (series 30 and 31, figs. 22
to 24), which correspond to the confluence of ventral and dorsal
tributaries with the main channel. These enlargements of the
precardinal are evanescent and are evidently produced by the
increased amount of blood entering the main vein at these points

Development of the Jugular Lymph Sacs. 225

through the tributary branches, repeating, on a small and serial
scale, the phenomena observed in the development of the jugular
promontory at the site of the principal venous confluence of this
region, the promontory representing in its inception a correspond-
ing enlargement of the main vein, as yet indistinctly indicated, at
the junction of pre- and posteardinal veins to form the duct of
Cuvier.

These early stages offer a characteristic irregular and redundant
appearance of the embryonic vein channels, which in part are
but poorly differentiated against the surrounding tissue, a con-
dition likewise characteristic of the ventral and dorsal tributaries
of the main vein. In place of the more clearly defined series
‘of collateral branches of the succeeding stages, the earlier embryos
offer a number of short and more or less irregalar branches, which,
in their aggregate, represent the single more fully formed tribu-
tary of the later stages.

This appearance is due to the numerous secondary capillary
vessels which develop around both the main vein and its tribu-
taries, and which subsequently give rise, as above stated, to the
fenestrated character of the channels, and to the appearance of the
veno-lymphatic anlages.

In the following description of the dorsal tributdries of the
caudal or straight segment of the precardinal, the assignment of
a complex of several branches to the valuation of one of the single
tributaries of later stages is based primarily upon the latter’s rela-
tion tothe localized enlargements of the precardinal into which
the tributaries of earlier stages open. The increased vascularity
of the tissues, and the resulting augmentation of the number of
capillary vessels found in the area of each of the principal tribu-
taries, is one of the phenomena preceding and directly leading up
to the fenestral process above described as active in producing the
veno-lymphatic condition. This secondary capillary reticulum
to a certain extent masks the individuality of the primary pre-
cardinal tributaries. We have evidence that a localized group
or complex of tributaries which opens dorsally into the precardinal
vein in early stages may be represented by a single well-defined
branch in later stages. We also have positive evidence that this

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 2.

226 George 8. Huntington and Charles F. W. McClure.

single tributary may become secondarily surrounded by, and
involved in, a capillary network. We are therefore justified in
assuming that the formation of a reticular complex in connection
with the individual precardinal tributaries is a normal develop-
mental character, leading up to the typical fenestral condition of
the veno-lymphatic stage, and that its establishment may occur
very early or be somewhat retarded.

This view certainly coincides with the multiple arrangement
of the first three dorsal precardinal tributaries on the left side of
series 30 (1, 2 and 3, fig. 22), and series 31 (1, 2 and 3, fig. 24)
which are represented on the right side of series 30 by three dis-
tinct and single tributaries (1, 2 and 3, fig. 23).

EARLY VENOUS STAGES.

Series 80, 5+-""" Embryo
Reconstruction of left side,
Lateral aspect, fig. 22

The cephalic or arched portion of the precardinal is moderately
curved. The branches A—B are large and in line with the straight
precardinal segment, entering the arch at its confluence with the
latter. Double fenestration occurs at the junction of A—B with
the main vein, and cephalad of the arch are several detached
vascular islands. The proximal precardinal spindle I (I, fig. 22),
at the posterior extremity of the cephalic arch, is only moderately
developed.

The first three dorsal branches of the straight segment of the
precardinal are represented on the left side of the embryo partly
by single vessels and partly by plexiform capillaries. Tribu-
tary 1 (1, fig. 22) is a single vessel, which opens into the precar-
dinal close to the terminal of tributary A—B. Tributary 2 is
formed by three components (2, 2’ and 2”, fig. 22).

The spindle-shaped enlargements JJ and J/J in the course of
the straight precardinal trunk, receiving dorsally the terminals
of 1 and 2, are, like these, closely approximated to each other.

Dorsal tributary 3 is represented on the left side of this embryo
by three small branches, entering the dorsal aspect of precardinal

Development of the Jugular Lymph Sacs. pag

spindle JV, and by a detached vascular island not distinctly
joined to the main vein, or to the tributary complex to which it
belongs.

Dorsal tributary 4 (4, fig. 22) is large and typically developed.
It is formed by the confluence of two extended paraneural channels
(a and b) converging from the cephalic and caudal directions.
Their confluence already exhibits the irregular dilatations charac-
teristic of veno-lymphatic formation. The main right-angled
trunk receives in addition a cephalic branch (4S), which represents
the element of precardinal tributary 4 so often found in subse-
quent stages entering the promontorio-precardinal junction on
its medial aspect as the first of the series of promontorial dorsal
somatic tributaries (4S, figs. 11 and 12).

The precardinal spindle V is only moderately indicated. On the
other hand, the distal enlargement VI is well marked, representing
the anlage of the future jugular promontory. It marks the con-
fluence of the pre- and posteardinal veins, which are still in a
direct line, and gives origin ventrally to the duct of Cuvier, receiv-
ing just cephalad of this point a short branch of considerable
size which probably represents the rudiment of the definite
external jugular vein.

The dorsal region of this area is most interesting and suggestive
of the later developmental stages. By the formation of a large
fenestral space in the dorsal portion of the postcardinal, opposite
to the Cuvierian confluence, an irregular dilated venous plexus
_(H) has been produced which projects from the vein in the area of
the future promontory. This plexus represents the anlage of
the dorsal veno-lvmphatic plexus.

Later, with the further development of the promontory and the
resulting dorso-ventral arching of the postcardinal, the dorsal
veno-lymphatic fenestrated area gains the level of the promontory
and rides on its cephalo-dorsal aspect.

Cephalad of this main fenestral arch H a second smaller arch
(K) is developed in a similar manner from the dorsal circumference
of the precardinal. This arch (K) probably represents, in part,
the capillary plexus involved in the formation of the para-pre-
cardinal channel of later stages (fig. 9).

228 George 8. Huntington and Charles F. W. McClure.

Series 30, 5-+"""" Embryo
Reconstruction of right side,
Lateral aspect, fig. 23

A—B, as the main dorsal branches of the cephalic arch, occupy
relatively the same position as on the left side. They are
reversed in size, A representing the main drainage channel, while
on the left side B is the larger of the two vessels.

The anterior precardinal spindle J (fig. 23, I) is well marked.
A small fenestra in the dorsal portion of spindle J may represent
the beginning separation of the element B', which on the left
side forms a dilated appendage to B.

Dorsal tributaries 1, 2 and 3 are single and distinctly defined.
1 and 2,as on the left side, are closely approximated, so that pre-
cardinal spindles J and J/ are practically confluent. ~ Precardinal
spindle JV is well developed. It receives dorsally tributary 3.

Spindle V is not pronounced, the dilatation at this point being
taken up by the marked enlargement of the terminal of dorsal
tributary 4. The broad quadrilateral channel formed by the
confluence of its cephalic and caudal tributaries is fenestrated at
the junction with the precardinal so that the tributary enters the ~
main vein by an anterior and posterior trunk. This fenestra
represents the space framed by the venous arch which 4 in the
later veno-lymphatic stages so frequently forms at its junction
with the main vein, and its presence denotes the beginning for-
mation of a reticular and fenestrated complex by means of which,
as above defined, this tributary may become connected with the
promontory in the later stages. From our reconstruction of these
later stages it is evident that its transferal to the promontory
may take place in one of two ways: |

(a) The anterior of the two trunks forming the terminal of .
4 may retain its connection with the precardinal vein, and the
posterior trunk may travel back, along the line of the fenestrated
reticulum, to the promontory, in which case the arch frames
dorsally a vertical parajugular foramen (figs. 27, 30 and 31) or, (6)
both trunks may become thus secondarily drained into the pro-
montory, in which case a closed venous arch projects forward
from the anterior face of the promontory (fig. 38).

Development of the Jugular Lymph Saes. 229
oa )

There is no indication of the presence of the dorsal somatic
element 4S, which is present on the left side of this embryo
(fig. 22). The reconstruction suggests that in this instance,
as frequently happens, the primitive dorsal tributary 4 becomes
involved in its entire extent in, the development of the caudal
division of the ventral veno-lymphatic plexus.

Caudad of tributary 4 the elongated distal segment of the pre-
cardinal receives dorsally three small branches (K) which may
represent the element K of the opposite side, (fig. 22).

Spindle shaped dilatation V/is bent sharply ventrad and receives
two large ventral branches, which represent the anlage of the per-
manent external jugular vein. As a matter of fact these vessels
now open into the cardinal end of the duct of Cuvier, but with the
caudal extension of the promontory, which is not yet fully estab-
lished in this embryo, they will be included in later stages within
the promontorial area. (See fig. 8, which illustrates diagramma-
tically the progressive caudal extension of the promontory and
the gradual inclusion within the same of the terminals of the exter-
nal jugular vein).

Dorsally, the postcardinal, at its confluence with the precardinal
channel is dilated, and perforated by irregular fenestrae of varying
size.

Comparison with the left side (fig. 22) suggests that confluence
of these separate openings would establish the single large fenes-
tra there encountered, in which case the channel H! (fig. 23)
would form the main postcardinal path, while the remaining
dorsal fenestrated arch would yield a para-cardinal venous arch H
which constitutes the anlage of the dorsal veno-lymphatic plexus.

The two sides of this embryo control and supplement each
other extremely well and present a typical picture of the stage
concerned.

Series 31, 5+" Embryo
Reconstruction of left side,
Lateral aspect, fig. 24

The cephalic arch is somewhat more decidedly ante-flexed, and
better differentiated against the succeeding straight segment of

230 George 8. Huntington and Charles F. W. McClure.

the precardinal (fig, 24), and tributaries A-B are more dis-
tinctly assigned to its caudal portion than in series 30. The ven-
tral tributaries of spindle J are relatively small, but more numer-
ous.

Spindles J and J/ are practically confluent. Spindle JJ
receives dorsally the well developed trunk of dorsal somatic
tributary 1 (1, fig. 24), formed by the confluence of two branches,
and, further caudad a secondary element, 1', belonging evidently
to the same drainage area, and representing the branch interme-
diate between 1 and 2 in series 30 (fig. 22).

There is a well marked constricted interval of the precardinal
caudad of spindle /7, between it and spindle J/J. The latter
receives dorsally four branches, representing the drainage of the
area assigned to dorsal tributary 2 (2 in fig. 24).

Another elongated constricted precardinal segment intervenes
between spindles //7 and JV. Spindle JV receives along its dorsal
circumference six individual branches representing the drainage
of dorsal tributary 3, in the condition of multiple small elements
entering the precardinal separately (3 in fig. 24).

Spindle V is occupied dorsally by the broad and complex union
with dorsal tributary 4. This tributary is typically dilated near
its Junction with the main precardinal vein, and contains two
fenestral openings, which by confluence would establish the single
space seen on the right side in embryo 30 (fig. 23) at the entrance of
the expanded terminal of tributary 4 into the precardinal.

Owing to imperfections in the series, the embryo was not recon-
structed caudal to this point. In general, compared with series
30, the embryo represents a slightly earlier developmental phase,
marked by the larger number of individual elements composing the
complex of dorsal tributaries 1, 2, and 3, and by the presence of
small intermediate tributaries between the main districts.

The general ground-plan of the embryonic venous system
shown in the two series just considered (series 30 and 31) affords
the most convenient approach to the detailed study of the sub-
sequent stages in venous and veno-lymphatic development.
Before proceeding to this study, it may be of advantage to con-
sider briefly the steps by means of which the venous conditions in

Development of the Jugular Lymph Sacs. 231

series 30 and 31 have become established. They are apparently
derived from the antecedent type shown in series 134 and 47.
These are the earliest stages which we have completely recon-
structed, and they show the building up of the primitive main
systematic venous system by the confluence of at first three and
then of four principal channels which unite to form the duct of
Cuvier and open through the same into the sinus venosus.

Series 134, 5°” Embryo
Reconstruction of left side,
Lateral aspect, fig. 25

In series 134 the duct of Cuvier is formed through the con-
fluence of the precardinal, the posteardinal, the omphalo-mesen-
teric and the umbilical veins.

The omphalo-mesenteric vein, receiving a number of small trib-
utaries from the pre-aortic region, possibly subcardinal radicles,
arches dorso-ventrad and joins the precardinal to form the duct of
Cuvier. The proximal dilated end of the omphalo-mesenteric
receives on its lateral aspect the terminal of the umbilical vein.
The latter is a vessel of large size, receiving by numerous tribu-
taries the drainage from the body walls and the anterior limb
bud. Along its dorsal circumference it is irregularly dilated and
fenestrated. ;

The precardinal forms a short, strongly curved arch, receiving
anteriorly several short tributaries. Near its posterior extremity
it receives dorsally two very large irregularly dilated branches
(4 in fig. 25). Comparison with the succeeding stages (series 30
and 31, figs. 22 to 24) suggests that they represent the elements
of precardinal tributary 4.

The postcardinal vein begins anteriorly in a dilated and fenest-
rated extremity (H) which closely approaches the large dorsal
tributaries of the precardinal but does not communicate with them.
Near its anterior extremity the posteardinal, by a ventrally
directed branch (LZ), establishes its connection at the cardinal-
Cuvierian junction. Along the rest of its course it follows closely

232 ~=George 8. Huntington and Charles F. W. McClure.

the dorsolateral aspect of the umbilical vein and receives a
number of serially arranged branches, which open into it dorsally.
Caudal to the last of these tributaries indicated in the figure the
posteardinal appears dilated and fenestrated and then rapidly
diminishes in caliber.

Series 47, 5°” Embryo
. Reconstruction of right side,
Lateral aspect, fig. 26

The junction of the pre- and postcardinal veins is, relatively,
of narrow caliber and perforated by a fenestra, agreeing with the
peculiar slender and irregular formation exhibited by the Cuvier-
ian confluence in series 134 and 30, figs. 25, 22 and 23. The om-
phalo-mesenteric and umbilical veins have shifted their terminals
caudad and now obtain an opening into the sinus venosus inde-
pendently of the duct of Cuvier proper. A branch from the
body wall entering the umbilical at the sinus venosus seems to
correspond to the primitive drainage of the external jugular terri-
tory. The umbilical vein also receives further caudad a number
of somatic tributaries and branches from the anterior limb bud.
This drainage area, compared with series 134 (fig. 25) appears
reduced and in the process of transferal to the path of the postcar-
dinal whose somatic branches are more fully developed.

The greatest interest attaches to the neighborhood of the pre-
and posteardinal confluence. In both embryos (134 and 47) con-
ditions are presented in this region which directly lead up to those
found in the succeeding stages (embryos 30 and 31). Two points
are here involved:

(a) Precardinal

The dorsal precardinal tributary 4 (4 in fig. 26), which is
destined to play so important a réle in the subsequent venous and
veno-lymphatic development, appears to be laid down at a very
early stage. In series 134 (4, fig. 25), it is represented by the two
large dilated and tortuous branches which enter the dorsal cir-

Development of the Jugular Lympn Saces. 233

cumference of the caudal part of the precardinal. In series 47
(4, fig. 26), the precardinal end of this element appears as the large
single trunk entering the precardinal in the same situation. The
formation of tributary 4, with its cephalic and caudal tributary
a and b, in series 30 (figs. 22 and 23), has already been considered
in detail, and corresponds evidently to a further development in
these later embryos of the conditions presented by the earlier
series.

(b) Postcardinal

The redundant plexiform area of the postcardinal (H) corre-
sponding to the dorsal aspect of the Cuvierian junction in series
30 and 31, and representing the early anlage of the dorsal veno-
lymphatic plexus is already well indicated in both of the earlier
stages. In series 134 (fig. 25) it is represented by the dilated and
fenestrated anterior blind end of the posteardinal (H). In series
47 (fig. 26), it appears as the fenestrated and plexiform append-
age of the dorsal circumference of the postcardinal (7) just caudal
to the latter’s entrance into the duct of Cuvier.

INTERMEDIATE EARLY STAGES OF PRE- AND POSTCARDINAL
VENO-LYMPHATIC DEVELOPMENT

Series 109, 6.2" Embryo
Series 2, 7” Embryo
Series 188, 7"”" Embryo
Series 18, 7.25"™" Embryo
Figs. 27 to 39

A. GENERAL CONSIDERATIONS

The main venous trunks have now begun to assume the arrange-
ment typical of the later stages.

The cephalic arched segment, conforming to the increased
cranial flexure of the embryo, has turned ventro-caudad, present-
ing a well-defined convexity, which receives the most anterior
dorsal tributaries (A-B).

The straight precardinal segment, shortened anteriorly by the

234 George S. Huntington and Charles F.. W. McClure.

development of the cranial bend, is further condensed and appears
relatively shortened by the further development of the jugular
promontory.

The dorsal tributaries of the precardinal are condensed, and
approximated to each other. The more numerous individual
branches of the earlier stages are replaced by larger single trunks
representing the three typical anterior sets, tributaries 1, 2, and 3.

Dorsal tributary 4 of the earlier stages now terminates, for the
most part,inthe promontory at the promontorio-precardinal junc-
tion. It (4) may be in part or entirely taken up in the subsequent
veno-lymphatic formation, or it may be permanently retained as a
somatic branch, in which case it forms the first of a series of
somatic tributaries entering into the dorso-medial aspect of the
promontory, or, finally, it may exhibit distinctly both veno-
lymphatic and dorsal somatic characters, with numerous trans-
verse anastomoses between the two elements.

In fact, all of the fundamental processes involved in veno-

lymphatic development, as described above under Analysis of
Developmental Stages in the Formation of the Jugular Lymph
Saes, are represented in one or the other of the above mentioned
series, and may be summed up as follows:
- (1.) Progressive enlargement and caudal extension of the
jugular promontory. (2) Increase in secondary capillary forma-
tion and resulting increase in the reticulated and fenestrated dorso-
lateral circumference of the promontory and the postcardinal vein,
and consequently an enlargement of the dorsal veno-lymphatic
plexus. (3) The establishment of the anlage common to
the primitive ulnar vein and the primitive ulnar veno-lymphatie.
(4) The transferal of the main blood current in tributary 4 from
the precardinal vein to the promontory through the secondarily
formed para-precardinal channel. (5) Separation of precardinal
tributary 4 and its associated para-precardinal channel into a
veno-lymphatie and a dorsal somatic component, (6) Sepa-
ration of the precardinal tributaries 1, 2 and 3 into their
veno-lymphatie and dorsal somatic components. (7) Estab-
lishment of the cephalic and caudal divisions of the ventral
veno-lymphatic plexus.

Development of the Jugular Lymph Sacs. 235

B. DETAILED STUDY OF THE INDIVIDUAL SERIES.

Series 109, 6.2" Embryo
Reconstruction of left side,
Lateral aspect, fig. 27 and
Medial aspect, fig. 28

The cephalic arch is bent ina wide curve ventro-caudad. The
tributary A-B of the convexity of the arch is of large size, and has,
relatively, moved forward with the better differentiation of the
arch from the straight segment of the precardinal. The area of the
precardinal which receives the dorsal tributaries 1, 2 and 3 (in figs.
27 and 28), is somewhat expanded and dilated along its
dorsal circumference, a feature which characterizes this region
of the precardinal preparatory to the separation therefrom of a
secondary channel by crystallization of the perivenous capillary
network. Asa result of this expansion of the precardinal vein, the
first three dorsal tributaries of the latter (1, 2 and 3, figs. 27 and 28),
assume a funnel-shaped form and frequently become confluent
at their precardinal termination. This funnel-shaped enlargement
of the terminals of dorsal tributaries 1, 2 and 3, is well shown in
fig. 27, where tributary 1 opens into the precardinal as an inde-
pendent vessel, while tributaries 2 and 3 are confluent at their
bases, although a small vascular island, apparently belonging to
the latter, appears detached. <A similar detached venous element
lies between tributaries 1 and 2. These detached vascular areas
are of common occurrence in later stages, as portions of the sec-
ondary capillary plexus entering into the formation of the jugu-
lar lymph sacs. Caudal to tributary 3 the presence of a small
fenestra in the precardinal vein indicates the condensation of the
perivenous plexus to form the beginning of a small secondary
channel. Dorsal tributary 4, as in series 30 and 31 (figs. 22, 23 and
24) is still the largest and most prominent of the series of pre-
cardinal dorsal branches. As a whole it presents a condition pre-
liminary to its differentiation into a medial dorsal somatic and a
lateral veno-lymphatic component. In addition to its primary
point of communication with the precardinal, at X in fig. 27, it
now also opens caudad into the promontory. This condition, as

236 «George S. Huntington and Charles F. W. MeClure.

compared with the preceding series 30 and 31, has been developed
as the result of the formation of the elongated fenestra Y (figs. 27
and 28), by means of which a dorsal para-precardinal channel has
become separated from the main precardinal vein in the area
between the promontory and the primary precardinal connection
(X) of tributary 4 (fig. 27, X). This para-precardinal channel may
have been formed as the result of the caudal extension along the
precardinal of a fenestral process in the terminal of tributary 4,
similar to that present in series 30 (fig. 28).

It is not possible to state definitely what the subsequent fate of
tributary 4 and its associated para-precardinal channel would
have been in this specific instance, if the embryo had continued to
develop. Tributary 4 would, however, be potentially capable of
separating into a dorsal somatic (48 in fig. 11A) and a veno-
lymphatic component (4VL in fig. 11A), the former opening into
the promontory as the first of the series of promontorial somatic
tributaries, and the latter entering into the formation of the caudal
or posterior division of the ventral veno-lymphatic plexus, as well
as contributing to the forward extension of the dorsal veno-
lymphatic plexus (fig. 13).

The promontory is a capacious sac, formed largely by the di-
lated proximal end of the postcardinal vein and is continued
ventrally into the duct of Cuvier. The dorso-medial aspect of
the promontory and of the postcardinal vein receives at regular
intervals the dorsal somatic tributaries 5S, 6S, 7S, and 8S (fig. 28,
medial aspect). J

The dorso-lateral aspect of the promontory (figs. 27 and 28)
presents a most typical picture of the redundant venous develop-
ment of this stage. It is occupied by an elongated capacious and
fenestrated sac (dorsal veno-lymphatic plexus) in wide open com-
munication with the main vein, continued caudad into the anlage
common to the primitive ulnar vein and the primitive ulnar veno-
lymphatic. A series of five fenestrae run along the junction of the
dorsal veno-lymphatie plexus and the promontory at irregular
intervals, and are continuous with those which separate the anlage
common to the primitive ulnar vein and veno-lymphatie from the
posteardinal.

Development of the Jugular Lymph Sacs. Zeit

The dorsal veno-lymphatic plexus opens into the promontory
at four points, separated by fenestrae of varying width. The
anlage common to the primitive ulnar vein and its associated
veno-lymphatic has separated from the postcardinal to a point
about midway between the dorsal somatic tributaries 7S and 8S
(7S and 8S in fig. 28), where it arches over the sixth segmental
nerve to open into the dorso-lateral circumference of the promon-
tory, and to become continuous in front with the dorsal veno-lym-
phatic plexus. The external jugular anlage (figs. 27 and 28) still
opens into the cardinal end of the duct of Cuvier, which has not,
as yet, been incorporated in the promontory.

The chief features of the left side of this embryo, as compared
with the preceding earlier stages, may be enumerated as follows:

1. More marked differentiation of cephalic arch from succeed-
ing segment.

2. Confluence and dilatation (with beginning fenestration) of
terminals of dorsal precardinal tributaries 1, 2, 3.

3. The establishment of a secondary promontorial connection
for precardinal tributary 4.

4. Clear differentiation of the jugular promontory from the
straight portion of the precardinal and dilatation of the dorsal
veno-lymphatic plexus. More marked separation from the post-
-eardinal, by condensation of the capillary plexus, of the anlage
of the primitive ulnar vein and veno-lymphatie.

5. Assignment of three dorsal somatic tributaries (5S, 6S, 7S)
to dorso-medial aspect of promontory (fig. 28).

6. Differentiation of tributary 8S as first dorsal somatic branch
of the postcardinal, caudad of primitive ulnar entrance.

It is necessary to compare the lateral and medial aspects of the
reconstruction in order to see the special features presented in
this stage. They are shown in figs. 27 and 28.

238 George 8S. Huntington and Charles F. W. McClure.

Series 109, 6.2" Embryo
Reconstruction of right side,
Lateral aspect, fig. 29 and
Medial aspect, fig. 30

The reconstruction of the right side of this embryo corresponds
in general with that of the left. The veno-lymphatic development
and the incident plexiform formation in the area of tributary 4
shows, however, a greater degree of development.

The first three dorsal tributaries of the precardinal (1, 2 and 3,
figs. 29 and 30) are dilated and funnel-shaped, the second and
third having fused at their bases. This funnel-shaped form pre-
cedes their separation into veno-lymphatic and dorsal somatic
components, the former of which will constitute the anlages of the
cephalic division of the ventral veno-lymphatic plexus. As on the
left side of this embryo (fig. 27), dorsal tributary 4 (4 in figs.
29 and 30), has obtained a promontorial connection through the
formation of a secondary channel (para-precardinal channel),
separated from the precardinal vein by the elongated fenestra Y
(fig. 30). In addition, a second fenestra (fig. 30, Z) has developed
at the promontorial end of the para-precardinal channel, so
that the latter is now composed in this region of two parallel
vessels opening separately into the promontory. The plexus at
this stage presents the condition of tributary 4 just prior to
its separation into definite veno-lymphatic and dorsal somatic
components, the former associated with the para-precardinal
channel to form with it the caudal division of the ventral veno-
lymphatic plexus.

The dorsal veno-lymphatie plexus, as on the left side of this
embryo, is an extensive plexiform sac occupying the dorso-lateral
aspect of the promontory (figs. 29 and 30). The sac projects for-
ward over the precardinal vein and is separated from the promon-
tory by a row of six fenestrae between which it communicates at
seven points with the main venous system. The anlage common to
the primitive ulnar vein and primitive ulnar veno-lymphatic has

Development of the Jugular Lymph Saes. 239

not separated from the postcardinal as far forward on the right
side (fig. 29) as on the left (fig. 27). In other respects its relations
to the dorsal sac and the main systemic veins are the same. Figs.
28 and 30, which represent the medial aspects of the left and right
sides, respectively, clearly show the relations of the dorsal veno-
lymphatic plexus and the dorsal somatic tributaries (5S, 6S, 7S
and 8S) to the promontory and posteardinal vein. In this stage,
the first of the promontorial dorsal somatic tributaries is the fifth
of the entire precardino-postcardinal series (5S).

In a subsequent stage the fourth of the entire series (4$) is the
first promontorial branch (compare figs. 28 and 30 with fig. 45).

In the embryo at present under consideration tributary 4, as a
whole, is still included, together with the para-precardinal channel,
in a common plexus, which will subsequently differentiate into a
veno-lymphatic portion (para-precardinal channel plus veno-
lymphatic component of 4) and into a somatic element. The
former will constitute the caudal division of the ventral veno-
lymphatic plexus. The latter, as the somatic component of tri-
butary 4, will enter the promontory as the first of the promontor-
ial dorsal somatic tributaries (4S) in line with the rest of the series
(5S, 6S, 7S,) and independently of the caudal division of the ven-
tral veno-lymphatic plexus.

This ontogenetic transferal of the fourth dorsal somatic tribu-
tary to the promontory is illustrated diagrammatically in figs. 8, 9,
10, 11 and 114A, and is further explained in detail in dealing with
series 138 presently to be considered. As stated above the dorsal
somatic tributaries (5S, 6S, etc.), of the promontory do not enter
into the formation of the veno-lymphatic plexus.

The embryo also exhibits well the early disposition of the veno-
lymphatic plexus to shift distinctly to the dorso-lateral circum-
ference of the precardinal and promontory, while the somatic
tributaries are grouping themselves into a regular series entering
the dorso-medial aspect of the promontory.

240 George 8. Huntington and Charles F. W. McClure.
Series 2, 7" Embryo
Reconstruction of right side,
Lateral aspect, fig. 31 and
Reconstruction of left side,
Lateral aspect, fig. 3%

~

t

This embryo, although of the same crown-rump measurement
as the following series 138 (figs. 33 to 36), is less developed than
the latter in so far as its veno-lymphatic elements are concerned.
There is also considerable variation present upon opposite sides
of the embryo, the right side having progressed to a lesser extent
than the left.

On the right side the dorsal precardinal tributaries 1 and 2
(fig. 31), are expanded and plexiform at their terminals, and repre-
sent a stage preparatory to their final separation into dorsal
somatic and veno-lymphatic components, as seen in series 138
(figs. 38 and 36). (Also compare with schematic diagram, fig. 10,
which corresponds in this respect to the condition presented by
this embryo).

Precardinal tributary 3 is not represented on the right side
by any well-defined vessel. It may have become incorporated in
the funnel-shaped expanded terminal of 2, as in series 109 (fig. 29),
or subsequently detached from the same, as in fig. 28, in which
case it may be represented on the right side in series 2 by the
detached element labelled 3VL(fiz. 31).

Precardinal tributary 4 presents essentially the same condi-
tions as on the left side of the 6.2 mm. embryo (series 109, fig. 27).
It opens cephalad into the precardinal vein at X and caudad into
the promontory. The former connection (fig. 31, X) represents
its original junction with the precardinal vein (compare series
30 and 31, figs. 22, 283 and 24). Its connection with the promon-
tory has been secondarily acquired by the condensation of the
precardinal plexus into a para-precardinal channel, which, separ-
ated from the main vein by the fenestra Y (fig. 31, Y), spans
the interval! between the promontory and the primary terminal
of 4 (fig. 31, X), and now drains the greater part of the blood cur-
rent of 4 directly caudad into the promontory.

Development of the Jugular Lymph Sacs. 241

As will be seen by comparison with the next series 138 (fig. 35),
tributary 4 on the right side of the embryo now under considera-
tion represents a stage in which the capillary plexus surrounding
its terminal has not yet been clearly defined into somatic and
veno-lymphatic components. A comparison of this reconstruc-
tion with the one shown in fig. 35 may, however, justify the
inference that the irregular, bifurcated dorsal portion of tributary
4 (4Vl, fig. 31) may represent a veno-lymphatic anlage, while
the dorsal somatic component (4S in fig. 31,) in this case would
separate subsequently from the veno-lymphatic component of
tributary 4 and from the para-precardinal channel, and gain an
independent entrance into the dorso-medial aspect of the promon-
tory. This is one of the instances, so frequently encountered in
the course of this investigation, in which it is evident that the
embryo in question has been fixed during a period of transition
from one well-defined stage to the succeeding condition. If we
were basing our conclusions on a small number of sectioned and
reconstructed embryos, it would be necessary to record the
observed conditions without venturing a positive interpretation.
Our material, however, has been so extensive and conclusive, and
so representative of each important ontogenetic phase, that we
feel justified in foretelling the significance of conditions which, in
a given embryo, unmistakably point toward the stage, which,
without interruption, would have been attained.

We may be pardoned if, in connection with the series under dis-
cussion, we venture to emphasize the importance of embryological
investigations based on the careful analysis of a large number of em-
bryos of approximately the same age. Our previous experience in
the study of the embryonic venous variations of the cat has proved
conclusively to us that it is necessary to determine normal onto-
genetic conditions for each main developmental period, and that
the question of embryonic departures from this norm equals
in importance and significance the value which should be assigned
to variations of the adult vascular system.

On the left side of this embryo (fig. 832) the ventral curvature of
the cephalic arch is well established.

A-B enters as a large single vessel at the junction of the cephalic

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 2.

242 George 8. Huntington and Charles F. W. McClure.

arch and straight portion of the preeardinal. The precardinal
receives dorsal somatic tributaries 1, 2, 3 and 4.

Several detached vascular islands, notably between A-B and
tributary 2 are present. The para-precardinal channel has been
separated from the main precardinal vein independently of tribu-
tary 4, or, if at one time connected with it, this connection has
been lost. In other words, the embryo under discussion is one of
the instances illustrating the type of development described in
the analysis (cf. p. 215, fig. 19), in which the caudal division of the
ventral veno-lymphatic plexus is apparently derived entirely
from the para-precardinal channel, without involving tributary 4.
In the present instance, the elongated fenestra Y in fig. 31,
between the precardinal and promontorial terminals of the para-
precardinal channel, is replaced on the left side of the embryo
by two rectangular foramina (fig. 32, Y’ and Z); consequently the
para-precardinal channel here drains through the resulting inter-
fenestral pathways, by three separate and distinct openings into
the main channel.

The fenestral character of the terminal of tributary 4 (4, fig. 32),
indicates that a secondary channel is being formed along the pre-
cardinal vein, independently of the para-precardinal channel, by
means of which tributary 4 will subsequently drain into the
promontory.

The dorsal veno-lymphatic plexus is more advanced in develop-
ment on the left side than on the right. It is continuous cephalad
with the caudal division of the ventral veno-lymphatic plexus,
and caudad with the anlage common to the primitive ulnar vein
and the primitive ulnar veno-lymphatic. This anlage has differ-
entiated completely from the postcardinal vein, caudal to the
termination in the latter of dorsal somatic tributary 7 (fig. 32, 7S).
The anlage of the permanent external jugular system is represented
on the left side by a single trunk (fig. 32), corresponding to the
double vessel found on the right side (fig. 31) in the same situation.

Compared with the preceding series (109, 6.2mm., figs. 27 and
29), the right side of this embryo (fig. 31) shows retarded develop-
ment of the dorsal veno-lymphatic plexus. A single channel,
closely following the dorso-lateral circumference of the promon-

Development of the Jugular Lymph Saces. 243

tory, and only differentiated from the same by a linear series of
three fenestrae, represents the extensive plexiform area which in
series 109 (fig. 27) occupies the dorso-lateral circumference of the
promontory. The same relative reduction of the dorsal veno-
lymphatic plexus is seen on the left side of this embryo (fig. 32)
when compared with the same side of the 6. 2 embryo (series 109,
fig. 27.)
Series 138, 7°” Embryo

Reconstruction of left side,

Lateral aspect, fig. 33 and

Medial aspect, fig. 34

This embryo offers a remarkably clear picture of veno-lym-
phatic development and illustrates well the relation of the veno-
lymphatic plexuses to the system of the primitive dorsal somatic
tributaries.

The plexiform and dilated terminals of precardinal tributaries
1, 2 and 3 (as seen in figs. 27, 29 and 31), and the terminal of
dorsal precardinal tributary 4 and its associated para-precardinal
channel, reveal here very strikingly their separation into dorsal
somatic and veno-lymphatic components. Another feature of
fundamental importance to the interpretation of succeeding
stages is the tendency displayed by these veno-lymphatic deriva-
tives to shift their connection with the precardinal from the dorsal
to the dorso-lateral circumference of the main channel.

After these veno-lymphatic derivatives have assumed their
typical dorso-lateral position with respect to the main systemic
venous channel, they are then in line to fuse with one another and
with the dorsal veno-lymphatic plexus which, from the beginning,
projects cephalad from the dorso-lateral circumference of the
promontory.

This embryo likewise throws considerable light on the genesis
of the variations presented by the terminal of dorsal somatic
tributary 4 of the early series, in reference to the double character
assumed by the same in both the somatic and the veno-lymphatic
direction. For these reasons the embryo requires special and
detailed consideration.

244 George 8. Huntington and Charles F. W. McClure.

Tributary 1 (fig. 33, lateral aspect) enters the cephalic end of
the straight precardinal segment by two branches, inclosing a
wide fenestral space. The caudal of these two branches (fig. 33,
1VL) opens into the dorso-lateral circumference of the precardinal,
a circumstance which, in addition to its plexiform and dilated
character, designates it as a veno-lymphatic structure. Here the
crystallization of the plexiform reticulum into definite somatic
and veno-lymphatie lines is still evident within the basal area of
the tributary.

Tributary 2 is represented by a veno-lymphatic component
(2VL) which opens into the dorso-lateral, and by a dorsal somatic
component (28, fig. 34), which opens into the dorso-medial cir-
cumference of the precardinal.

The original anastomosis between these components of 2 has
in this instance been lost, and they now open into the precardinal
as separate and independent tributaries.

This interpretation is supported by the disposition of the suc-
ceeding branches in this embryo.

Dorsal somatic tributary 3 has separated into a lateral veno-
lymphatic (3VL) and amedial dorsal somatic component (38).
The veno-lymphatic component (3VL) consists of alargeexpanded
double branch (3VL) and of asmaller caudal division (3VL’). Both
of these veno-lymphatic elements open into the dorso-lateral
circumference of the precardinal.

The dorsal somatic component of tributary 3 (3S) is extremely
small, and its relation to the veno-lymphatie element of this tribu-
tary clearly indicates its original connection with the same.

The veno-lymphatic components of the first three dorsal precardinal
tributaries constitute the anlage of the Cephalic or Anterior Division
of the Ventral Veno-Lymphatic Plexus. (Cf. figs. 10, 11, 12 and 13).

The caudal or posterior division of the ventral veno-lymphatic
plexus is already fully established. It is formed by the para-
precardinal channel and by the veno-lymphatic components of
4 (4VL in figs. 33 and 34); while the dorsal somatic element of 4
(4S in figs. 33 and 34) now opens into the dorso-medial aspect of
the promontory.

The veno-lymphatic element of 4 represents the fenestrated

Development of the Jugular Lymph Saces. 245

part of the terminal of the early branch 4 ina more advanced state
of development. It has retained a cephalic connection with the
precardinal (X, fig. 33), but the para-precardinal channel has
broken away at the usual distal tap (at B) into the angle between
the precardinal and promontory, apparently by secondary fusion
at this point with the dorsal veno-lymphatic plexus.

This embryo may possibly offer the ontogenetic explanation of
the adult cases in which the lymphatic system, through the Jugu-
lar lymph sac, communicates with the venous system only at
the distal of the two customary taps, viz., at the jugulo-subclavian
angle. By early union of the elements of the vencral veno-
lymphatic plexus with the dorsal plexus, the terminal of the
latter, at the jugulo-subclavian angle, may lead secondarily to the
formation of the only definite adult lymphatico-venous portal.
As previously stated (p. 192) the results of this investigation
oblige us to assume that the early embryonic connections of the
veno-lymphatic plexuses with the permanent veins are given up
and that the adult lymphatico-venous taps are formed secondarily.

Compared with series 109 (figs. 27 and 30) this side of the
embryo clearly shows the double character now assumed by pre-
cardinal tributary 4 and the associated para-precardinal channel
(figs. 833 and 34), and the lines along which the veno-lymphatie
and dorsal somatic components separated. On the left side of this
embryo this separation is complete, except at one point (figs. 33
and 34, A) where the original connection between the two still

‘persists.

The veno-lymphatic component of 4 (4VL in fig. 33) and its asso-
ciated para-precardinal channel is expanded and dilated. It has,
as in series 199 (figs. 27 and 30), retained its original point of
connection with the precardinal vein (X in fig.33). The promon-
torial opening of the para-precardinal channel at the promontorio-
precardinal angle has been much reduced, owing perhaps to the
dorsal extension of the fenestra Y. (fig. 33), resembling somewhat
the condition found in series 109 (fig. 30). The small curved
tributary B in fig. 33 at the promontorio-precardinal angle, prob-
ably represents a portion of the para-precardinal channel separated
from the main vessel by the extension of fenestra Y. The veno-

246 George S. Huntington and Charles F. W. McClure.

lymphatic complex of 4 and of the para-precardinal channel has
become confluent with the dorsal veno-lymphatic plexus.

An elongated diverticular process near the promontory (A P, fig.
33) opens on the left side into the para-precardinal channel. The
same element is present on the right side (AP, fig. 35) but, in
this case, does not terminate in the para-precardinal channel, but
in the dorsal somatic component of precardinal tributary 4 (4S)
which is still but incompletely differentiated from the para-pre-
cardinal channel.

The dorsal somatic component of 4 (4S, fig. 34) is composed
of two parallel channels, united anteriorly by a short transverse
branch, which open into the dorso-medial circumference of the
promontory by a common trunk. The ventral and longer of
these two channels follows closely the dorso-medial circumference
of the precardinal vein and terminates cephalad in a blind ex-
tremity, near which it communicates (figs. 33 and 34, at A)
with a veno-lymphatic component (4VL) of tributary 4. About
midway in its course the ventral channel also communicates with
the precardinal vein (fig. 34, X’), thus retaining its primary con-
nection with the main vein as in figs. 22, 23 and 24, as does also
the veno-lymphatic derivative of tributary 4 (fig. 33, X). The
dorsal smaller subdivision of the somatic element of tributary 4
terminates cephalad in a blind end.

The dorsal division of the veno-lymphatic plexus is an extensive
multi-fenestrated product of the dorso-lateral portion of the prom-
ontory, the plexiform network extending caudad along the line
of the future separation of the primitive ulnar from the main
posteardinal vein.

The promontory in series 138 (figs. 33 to 36) is a well-defined
area. The external jugular anlage opens into the promontory
near the latter’s point of confluence with the duct of Cuvier.

Series 188, 7" Embryo
Reconstruction of right side,
Lateral aspect, fig. 35 and
Medial aspect, fig. 36

The right side of the embryo presents the same clear differen-
tiation into dorso-lateral veno-lymphatiec and dorso-media]

Development of the Jugular Lymph Sacs. 247

somatic precardinal elements, but with a number of communications
still persisting between the corresponding elements of the two sets.

Precardinal dorsal tributary 1 is fenestrated at its junction
with the main vein, so as to open into the same by two inter-
fenestral channels (1VL, fig. 35). It occupies the dorso-lateral
aspect of the vein aad may be fairly regarded as the most ante-
rior veno-lymphatic element. There is no corresponding dorso-
medial or somatic branch.

The dorsal precardinal tributary 2 is represented by a dorso-
lateral, dilated veno-lymphatic (2VL), which opens dorso-laterally
into the precardinal vein (fig. 35), by two small detached veno-
lymphatics (2VL’, figs. 35 and 36), and by a small dorsal
somatic branch (28, fig. 36) which opens dorso-medially into the
precardinal.

The precardinal dorsal tributary 3 is still in the plexiform con-
dition, and although it has not as yet divided into separate and
independent veno-lymphatic (3VL) and dorsal somatic (3S) com-
ponents, as in the case of tributary 2, these components can be
clearly differentiated from each other on account of the relations
which they bear to the dorso-lateral (3VL) and dorso-media! (3S)
circumference of the precardinal vein (figs. 35 and 36).

The two detached vascular elements which lie dorsal to the area
of tributary 2, on account of their dorso-lateral position, are veno-
lymphatics and have probably seen separated from the veno-
lymphatic component of tributary 3.

All of these veno-lymphatic components of tributaries 1, 2 and 3
constitute the anlage of the cephalic division of the ventral veno-
lymphatic plexus (cf. figs. 11 and 12).

In the area of the complex formed by the precardinal tributary
4 and its associated para-precardinal channel (caudal division of
the ventral veno-lymphatic plexus) we encounter an excellent
example of the incomplete separation between the veno-lymphatic
and dorsal somatic components into which the early complex has
subdivided, as the two sets of components are still connected with
each other by means of a number of transverse vessels.

The dorso-lateral veno-lymphatic component of tributary 4 (4VL,
fig. 35) appears as an extensive sac, which ends anteriorily in a

248 George S. Huntington and Charles F. W. McClure.

blind diverticular extremity. When compared with the recon-
struction of the left side (fig. 33) it appears possible that this
diverticular extremity may have been derived from tributary 3.
Such a condition might easily be brought about as the result of a
fusion between the veno-lymyhatic components of tributaries 3
and 4. Measurements of the two reconstructions seem to bear
out this view. (See also 3VL, in fig. 47).

The dorso-laterally situated veno-lymphatic complex of tribu-
tary 4 (4VL) and its associated para-precardinal channel (para-pre-
cardinal channel, VL) communicate at three points on _ the
right side of this embryo with the main precardinal vein (at X, X’
and X”’, figs. 35 and 36). The most anterior point of communica-
tion at X” (fig. 35) is not present on the left side of the embryo
(fig. 33) unless, as intimated above, it may be represented by the
precardino-veno-lymphatie confluence of tributary 3. The com-
munications with the precardinal at X and_X’, however, are repre-
sented on both sides of the embryo, as is evidenced by compara-
tive measurements of the two sides. (Compare X and X’ in figs.
33, 34 and 35). }

On the right side of the embryo the communication with the
precardinal at. X occurs dorso-medially and in common with a
secondary somatic component of tributary 4 (49’, fig. 36), while on
the left side of the embryo the communication with the precardi-
nal at X occurs dorso-laterally and independently of a somatic
branch (X, fig. 33).

The same reverse conditions prevail regarding the point of
communication with the precardinal at X’, since it occurs dorso-
laterally on the right side (X’, fig. 835) and dorso-medially on the
left (X’, fig. 34).

The communication at X on both sides of the embryo probably
represents the persistence of the primary union of tributary 4 with
the precardinal (see series 109, X, fig. 28), while the communi-
cation at X’ probably represents an instance in which the para-
precardinal channel has not been completely separated along its
entire extent from the precardinal vein.

Caudally, the veno-lymphatic complex of tributary 4 and its
associated para-precardinal channel enlarges and presents a multi-

EE ——————— a |

Development of the Jugular Lymph Saes. 249

ple plexiform formation, in line with the fenestrae along the prom-
ontorial and primitive ulnar areas, and is directly continuous with
the dorsal veno-lymphatic plexus (fig. 35). The position occu-
pied by the fenestre in the complex suggest that the latter may
subsequently be separated into a number of parallel channels
(see fig. 45).

Medially, the veno-lymphatic complex of tributary 4 and the
para-precardinal channel communicates by means of six trans-
verse branches (a, b, c,d, e andf, figs. 35 and 36) with a large dorso-
medial somatic trunk (4S) which, as the chief representative of the
dorsal somatic element of precardinal tributary 4, now enters the
promontory at the promontorio-precardinal angle and forms the
first of a series of segmentally arranged promontorial dorsal
somatic tributaries (4S, 5S, and 6S, figs. 36). The promontorial
dorsal somatic tributary 7S is not shown in the figure.

The presence of the above mentioned transverse communica-
tions at a,b, c,d, e, and f (figs. 35 and 36) represents a stage of
development in which the separation of the early complex of pre-
cardinal tributary 4 and its associated para-precardinal channel
into its veno-lymphatic and dorsal somatic components is still
incomplete, and more so than on the left side of the embryo, where
only one such connection exists (A, figs. 33 and 34).

Fig. 37 is a scheme of the right side of series 138 (fig. 35) viewed
from its dorsal aspect, in which the dorso-lateral (veno-lymphatic)
and dorso-medial (somatic) components of tributaries 1, 2,3 and 4
have been slightly divaricated in order to expose the dorsal sur-
face of the precardinal and promontory and to emphasize, thereby,
the double character of the precardinal tributaries, as well as the
relation subsequently assumed by their veno-lymphatie and
somatic components with respect to the main venous channels.

The legends in fig. 37 correspond to those in figs. 35 and 36.
The following description of the diagram (fig. 37) will make clear
the complicated conditions observed in series 138.

In the area of primitive dorsal tributary 4 we encounter, in its
fullest degree, the development of the dorso-lateral veno-lym-
phatic component, of the dorso-medial somatic branch, and of the
transverse communications between the two. The dorso-lateral

250 George 8. Huntington and Charles F. W. McClure.

veno-lymphatic element, (4VL, fig.37) appears asan extensive and
fenestrated sac. It ends anteriorly in a blind diverticular ex-
tremity. Two channels of communication through X”” and X’
lead into the main precardinal channel, between which a trans-
verse branch connects a medial somatic tributary (48’) with the
veno-lymphatie component of tributary 4 (4LV), Proceeding
eaudad, the veno-lymphatic sac of 4 enlarges and presents
multiple fenestral formation in line with the fenestre of the pro-
montorial and primitive ulnar areas, and unites with the redun-
dant veno-lymphatie dilatation of this region (dorsal veno-lym-
phatic plexus). Medially it connects by six transverse branches
(a,b,c, d,e,f) with a well developed dorso-medial trunk (4S) which,
as the main representative of the somatic element of 4, enters the
dorso-medial angle of the promontory in line with the succeeding
dorsal somatic promontorial branches 5S and 68, ete.

It will be noted that in the general plan of organization the two
sides of this embryo agree closely and present a consistent picture,
characterized by three main features.

1. Distinct development of a dorso-lateral series of dilated
veno-lymphatic elements derived from the precardinal tributaries
1, 2and 3 and from precardinal tributary 4 and its associated para-
precardinal channel. Of these the derivatives of tributaries 1, 2
and 3 represent the anlage of the cephalic division of the ventral .
veno-lymphatic plexus, while the element derived from the tribu-
tary 4 and the para-precardinal channel forms the anlage of the
caudal division of the ventral veno-lymphatic plexus. In early
stages the latter is often continuous with the dorsal veno-lym-
phatic plexus, which is derived from the dilated, plexiform dorso-
lateral aspect of the promontory and the primitive ulnar segment
of the postcardinal.

2. Coincident with this development is the appearance of
a series of dorso-medial branches which represent the somatic
elements of the primitive precardinal tributaries. These are very
variable in their character, as may be seen by comparing the oppo-
site sides of the embryo (figs. 34 and 36).

3. The most remarkable feature is the persistence between the
dorso-lateral veno-lymphatic and dorso-medial somatic elements
of a series of transverse communications.

Development of the Jugular Lymph Sacs. LGM

The embryo illustrates exceedingly well the fact that, in cer-
tain stages, or in certain individuals, while the early dorsal tribu-
taries 1, 2 and 3 are largely taken up in the veno-lymphatic
organization and shifted to the dorso-lateral circumference of the
precardinal, portions of the early elements may remain as small
and temporary channels in line with the succeeding dorsal somatic
series, and function to a limited extent as dorsal somatic tribu-
taries of the precardinal.

Moreover, this embryo clearly establishes the compound
character of the element 4, and its associated para-precardinal
channel, and accounts for the many variations encountered in its
connections with the main channels and its relation to the veno-
lymphatie sac. In fact this entire question can best be discussed
on the hand of the conditions shown by the reconstruction Just
deseribed, compared with other typical series.

Series 18, 7.25" Embryo
Reconstruction of right side,
Lateral aspect, fig. 38 and
Reconstruction of left side,
Lateral aspect, fig. 39

This embryo is especially interesting for the reason that it
illustrates very clearly the range of variation, as regards veno-
lymphatic development, which may be presented on opposite
sides of the same embryo.

On the right side of the embryo (fig. 38) the straight segment
of the precardinal receives at regular intervals three well-defined
dorsal tributaries (1, 2 and 3 in fig. 38). In front of its confluence
with tributary 2 the precardinal is much dilated.

Precardinal tributary 4 of the right side is represented by two
veno-lymphatic structures (4VL and 4VUL’, fig. 38). One is a
triangular-shaped element(4VL’) which rests upon the dorsal
surface of the precardinal at the promontorio-precardinal angle
and lies in line with the precardinal tributaries 1, 2 and 3. As
far as we are able to determine this structure does not commu-
nicate with the venous system and may, in this case, constitute
the anlage of the dilated, rounded appendage of the ventral divi-

252 George 8. Huntington and Charles F. W. McClure.

sion of the veno-lymphatic plexus, which occupies a similar posi-
tion and is so often met with in later stages (see series 77, fig. 51).
The other veno-lymphatic component (4VL) of precardinal tribu-
utary 4 is the larger of the two. It occupies a dorso-lateral posi-
tion, has given up its connection with the precardinal and opens
into the promontory by a right-angled line which is directly
continuous with that of the dorsal veno- lymphatic plexus.

This embryo shows a condition which can be derived from that
obtaining in series 30 and 31 (figs. 23 and 24) by development of
a para-precardinal channel involving the fenestrated terminal
of tributary 4 and the subsequent detachment of the complex
thus formed from the precardinal and promontory back to the
point where if becomes continuous with the dorsal veno-lymphatic
plexus.

The right-angled limb of this veno-lymphatic structure shows
clearly the course along which the process of fenestration pro-
ceeded. We have here an instance, in all probability, in which the
precardinal tributary 4 has been transformed in its entirety into
veno-lymphatic components. If such is actually the case, dorsal
somatic tributary 5S, as in the present case (fig. 38), will form the
first of the promontorial tributaries.

Veno-lymphatic formation along the dorso-lateral cireum-
ference of the promontory and postcardinal vein is well advanced
on the right side of this embryo except for an area which inter-
venes between the dorsal veno-lymphatic plexus and the anlage
common to the primitive ulnar vein and primitive ulnar veno-
lymphatic. This anlage opens into the main channel at three
points in the neighborhood of the termination of the promon-
torial dorsal somatic tributary 7S (fig. 38). Caudad of these
connections the primitive ulnar vein has completely separated
from the posteardinal.

The promontory is not as yet a capacious sac and is formed on
the right as well as upon the left side (fig. 39) by the enlarged
Cuvierian end of the posteardinal. The anlage of the external
jugular vein opens on each side into the proximal end of the duct
of Cuvier, a circumstance which denotes the still incomplete
development of the promontory.

Development of the Jugular Lymph Saes. Dae
=| .
Series 18, 7.25" Embryo
Reconstruction of left side,
Lateral aspect, fig. 39

The two salient features of this embryo which distinguish
the left side (fig. 39) from the right (fig. 38) are that on the left
side the dorsal precardinal tributaries, with the exception of
tributary 1, are either rudimentary or are practically wanting as
such, and that the promontorial end of the straight segment of the
precardinal consists of two parallel channels, subequal in caliber,
which are separated from each other by an elongated fenestra
Ce tie 39).

The dorsal precardinal tributary 1 (fig. 39) is a vessel of fairly
large size. It opens dorso-medially into the precardinal ard as
yet shows no sign of fenestration.

Preeardinal tributary 2 is represented by a veno-lymphatic
element, which opens dorso-laterally into the precardinal (2VL,
fig. 39) and probably by the detached element which lies contig-
uous to the latter.

Precardinal tributaries 3 and 4, as distinct branches of the pre-
cardinal, are wanting, as no trace of any tributaries opening into
the precardinal could be found in the territory usually occupied
by them. In their place, however, and occupying a dorso-lateral
position with respect to the para-precardinal channel and in line
with the dorsal veno-lymphatic plexus, are situated a number of
independent veno-lymphatics (3, 4VL, fig. 39) which probably
represent these tributaries, and which in further development
would probably contribute to the formation of the continuous
veno-lymphatic arch of later stages, which lies dorso-lateral to the
straight segment of the precardinal.

The separation of the promontorial end of the straight segment
of the precardinal into two parallel channels is a feature of develop-
ment which has already been described in connection with series
109 (fig. 28) and series 2 (fig. 31). Usually, however, the more
dorsal of the two channels, which we have termed the para-pre-
cardinal channel, is the smaller of the two and serves, at a cer-
tain period of development, as the main pathway through which
the precardinal tributary 4 drains into the promontory. In the

254 George F. Huntington and Charles F. W. McClure.

present instance, having lost its connection with the precardinal
tributary 4, it functions in a manner similar to that of the more
ventrally situated main precardinal channel and_ represents
an advanced stage in the separation from the latter of what con-
stitutes the anlage of the caudal division of the ventral veno-
lymphatic plexus.

The veno-lymphatic development along the dorso-lateral cir-
cumference of the promontory and posteardinal vein has pro-
gressed to a slightly greater extent than on the right side. The
broken line along the promontory (fig. 39) shows the extent to
which the veno-lymphatiec ridge projects from the promontory.
The anterior end of the dorsal veno-lymphatic plexus extends for-
ward over the para-precardinal ¢ghannel, where it ends blindly and is
continuous caudad with the anlage common to the primitive
ulnar vein and primitive ulnar veno-lymphatic. This vessel opens
into the posteardinal by means of a single channel, and caudad of
this point, as on the right side, it has completely separated from
the postcardinal.

We have here an instance, as on the right side of this embryo,
in which the first promontorial dorsal somatic tributary (5 S, fig.
39) vs represented by the fifth of the series, and in which, in all proba-
bility, precardinal tributary 4 has been transformed in its entirety
into veno-lymphatic components which will aid in the forward exten-
sion of the dorsal veno-lymphatic plexus. Also, on the left side,
fig. 89, a case in which the caudal division of the ventral veno-lym-
phatic plexus is derived solely from the para-precardinal channel.
Compare figs. 39 and 40.

SUBSEQUENT EARLY STAGES ILLUSTRATING VENO-LYMPHATIC
DEVELOPMENT

Series 102, 8.5" Embryo
Series 106, 9°" Embryo
Series 19, 9°" Embryo

This group of embryos presents the following features of veno-
lymphatic development in advance of those described in connec-
tion with the preceding group:

Development of the Jugular Lymph Saes. Zo

1. As the result of its caudal extension, the promontory has
become a capacious sac, involving in its caudal growth the proxi-
mal end of the duct of Cuvier and of the posteardinal vein (see
fig. 8). The permanent external jugular anlage now opens into
the promontory.

2. Precardinal dorsal tributaries 1, 2, 3 and 4 are completely
differentiated into definite veno-lymphatic and dorsal somatic
components.

3. The veno-lymphatic components of precardinal tributaries
1, 2 and 3 have become much dilated, and secondary plexus for-
mation has now invaded the anterior portion of the straight
segment of the precardinal.

4. In addition to the two veno-lymphatic plexuses met with

in the preceding group (dorsal plexus and caudal division of
ventral plexus) the cephalic division of the ventral plexus makes
its first definite appearance in the group under consideration.
- Embryo 102, in spite of its smaller measurement (8.5 mm.),
possibly due to greater curvature, offers more advanced develop-
mental conditions than the remaining members of the group.
It is therefore desirable to consider the last two series first.

Series 196, 9°” Embryo
Reconstruction of left side,
Lateral aspect, fig. 40

This embryo illustrates very well a condition frequently observed
in the early stages, of comparatively advanced development of the
veno-lymphatic area in one section, with retarded development in
another.

Tributary A-B forms a prominent trunk with smaller accessory
branches (b, b’) entering the convexity of the cephalic arch caudal
to its termination.

Only the veno-lymphatic components of the precardinal dorsal
tributaries 1, 2 and 3, are shown in the figure. Two prominent
dorsal somatic tributaries open into the dorso-medial circumfer-
ence of the precardinal in the area occupied by tributaries 1 and
2. These lie in a line with the series of promontorial and post-
cardinal dorsal somatic vessels and have been derived from the

256 George S. Huntington and Charles F. W. McClure.

plexus of the dorsal precardinal tributaries of earlier stages as the
result of the condensation into dorsal somatic and veno-lymphatic
components (see description of series 138, and figs. 33 and 35).

The veno-lymphatiec component of tributary 1 (1VL, fig. 40)
is dilated dorsally into an elongated sac which opens into the dorso-
lateral circumference of the precardinal by a slender channel.
From its general appearance and relations to the area of the pre-
cardinal occupied by precardinal tributary 2, it is possible that this
dilated sac may be a compound structure formed through a fusion
of veno-lymphatics derived from tributaries 1 and 2. After
fusing with the more caudally situated veno-lymphatics, however,

‘it will form the most anterior portion of the continous dorsal
veno-lymphatic sac of later stages and its primary connection
with the precardinal vein may serve as the channel of exit into the
systemic veins of the blood contained in the veno-lymphatic
sac (see figs. 12, 13, 14 and 15).

The area of the precardinal occupied by tributary 2 (2 VL) is
redundant and fenestrated, a condition which foreshadows the
formation of an anterior para-precardinal channel which may
involve in its separation from the precardinal the bases of trib-
utaries 1, 2 and 3, At a later stage this channel frequently
fuses along the lateral surface of the precardinal with an anterior
prolongation of the caudal division of the ventral veno-lymphatic
plexus (see figs. 12, 18, 14 and 15).

The veno-lymphatic component of precardinal tributary 3
(3VL) is, relatively, a large and somewhat dilated, single vessel
which opens dorso-laterally into the precardinal, caudal to the
territory occupied by tributary 2 (2VL).

Precardinal tributary 4 presents an interesting condition in
this embryo and one which, in some respects, resembles that
found on the right side of the 7.25 mm. embryo (fig. 39), since it
has severed its connection with the para-precardinal channel
and the precardinal vein. The primary point of connection which
formerly existed between tributary 4 and the precardinal vein
(X, fig. 40) is indicated in the figure by broken lines.

In the area of precardinal tributary 4 and its associated para-
precardinal channel the process of fenestration is completed. A

Development of the Jugular Lymph Sacs. 257

dorsal somatic branch (4S) has been separated from tributary 4
and the para-precardinal channel, which now opens into the dorso-
medial surface of the promontory, where it forms the first of a
series of promontorial somatic tributaries (4S, fig. 40). The
veno-lymphatic component of tributary 4 (4VL, fig. 40) which
has severed its connections with the precardinal and the para-pre-
cardinal channel, now occupies a dorso-lateral position with respect
to the precardinal, and lies in line with the dorsal veno-lymphatic
plexus and the veno-lvmphatics derived from tributaries 1, 2 and
3, with which it will subsequently fuse to form the dorsal veno-
lymphatic arch in fig. 14. The para-precardinal channel still
communicates with the precardinal and the promontory, being
separated from the former by the fenestra Y. The para-pre-
cardinal channel on the left side of this embryo constitutes the sole
anlage of the caudal division of the ventral veno-lymphatic plexus,
precardinal tributary 4 per se not contributing to its formation.

As the result of a caudal extension which involved the proximal
ends of the postcardinal and the duct of Cuvier, the promontory
has increased in size in an antero-posterior direction. In con-
sequence of this caudal extension, the duct of Cuvier joins the
posteardinal further caudad than in the earlier stages and no
longer receives the anlage of the permanent external jugular vein,
which now opens into the ventro-lateral circumference of the
promontory. The promontory has not yet assumed the rounded
and sac-like appearance in this embryo which is so characteristic
of this area in certain other members of this group.

The anlage common to the primitive ulnar vein and primitive
ulnar veno-lymphatic has completely separated from the post-
cardinal. After arching over the sixth spinal nerve (SF.N.V1Z,
fig. 40) it opens into the dorso-lateral circumference of the prom-
ontory, contiguous to the latter’s point of confluence with the
sixth dorsal somatic tributary (6S). Anterior to its union with
the promontory it becomes continuous with the dorsal veno-
lymphatic plexus.

The most marked advance is shown in this embryo in the dors9-
lateral circumference of the promontory and of the arch of the
primitive ulnar vein. This segment of the primitive venous

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 2.

258 George 8. Huntington and Charles F. W. McClure.

pathway has developed a large dilated veno-lymphatie sac
representing the dorsal veno-lymphatie plexus. It forms an
irregularly quadrilateral dilated and diverticular appendage,
projecting from the dorso-lateral aspect of the promontory and
primitive ulnar arch. The persistent pathways between the
fenestral spaces furnish four channels of communication between
this veno-lymphatic sac and the permanent vein. Only three
of these channels are shown in the figure. This represents an
advance over the conditions shown in the earlier stages in this
region. The fenestrae have become elongated, thus drawing out
the persistent patent intervals between them into definite and
well-marked channels of communication. At a later stage of
development, and in connection with the formation of the primi-
tive ulnar veno-lymphatic, the dorsal veno-lymphatic plexus,
through confluence of its fenestrae, becomes separated from the
primitive ulnar vein, but retains a single point of communication
with the promontory near the latter’s junction with the primitive
ulnar vein (see Tap C, figs. 40 and 44).
As previously stated, this embryo offers a marked example of
the fact that veno-lymphatic development does not proceed uni-
-formly in all parts in the primary venous area involved, and that
in individual portions of this area one segment may be develop-
mentally much in advance of the rest, in a given stage. Thus
in the present instance the dorsal veno-lymphatic plexus is
considerably further developed than the ventral division (para-pre-
vardinal channel), while in other examples the reverse may obtain
(series 2, fig. 32). The reconstruction of this embryo also fur-
nishes a marked instance of the multiple openings, characteristic
of the intermediate stages, between the anlages of the future defin-
ite veno-lymphatic plexus and the permanent veins. With
the more complete separation of the veno-lymphatic sac from
the main venous channels the intervals between the individual
fenestral spaces elongate and become drawn out into narrower and
longer branches of communication between the two vascular areas.
Thus, in the present instance, we find, caudal to the junction of
tributary A—B with the cephalic arch, ten distinct openings lead-
ing from the anlages of the veno-lymphatic plexuses into the dorsal
circumference of the precardinal vein and promontory (fig. 40).

Development of the Jugular Lymph Sacs. 259

Subsequently, as stated in the preliminary analysis of veno-
lymphatic development, and as will be shown in the course of the
detailed consideration of the more advanced stages, these multiple
early connections undergo great numerical reduction.

This embryo, as well as series 13 (fig. 39), is also of great
importance in illustrating a feature of veno-lymphatic develop-
ment which, as yet, has not been carefully considered and one
which is even more marked in certain of the succeeding stages.
We refer to the presence of the detached and isolated veno-lym-
phatic saes, or spaces which appear as outlying islands to the main
divisions of the veno-lymphatic plexus and in the area in which the
jugular lymph sacs are subsequently found. These detached sacs
may or may not contain blood corpuscles and, in the latter case,
they present the appearance of typicalisolated lymphatic spaces.

The gradual separation and detachment of the main veno-lym-
phatic plexus from the systemic veins constitutes a uniform and typt-
cal phase in the ontogeny of the jugular lymph sacs. It, therefore,
appears to us that the detachment of these small veno-lymphatic
elements, in advance of the main veno-lymphatic system, may be
explained on the ground that their detachment merely anticipates the
main sac in attaining this stage.

As to the subsequent fate of these detached spaces there can
be no doubt. They enlarge, fuse with one another and with the
main veno-lymphatic plexuses in forming the anlage of the jugu-
lar lymph sae.

Series 19, 9°" Embryo
Reconstruction of right side,
Lateral aspect, fig. 41 and
Reconstruction of left side,
Lateral aspect, fig. 42

This embryo is markedly curved and shows a distinct demarca-
tion into cephalic arch, straight precardinal segment and promon-
tory. | It affords an instance of the developmental type described
in the analysis (p. 211) and illustrated by fig. 9, in which the
cephalic division of the ventral veno-lymphatic plexus is formed
primarily from the peri-precardinal capillary plexus, involving

260 George 8S. Huntington and Charles F. W. McClure.

secondarily the terminals of dorsal precardinal tributaries 1, 2
and 3, which are taken up in the veno-lvmphatic organization.
The two sides of this embryo present two distinct stages in the
condensation of the peri-precardinal plexus and the subsequent
separation of the resulting anlage of the cephalic division of
the ventral plexus from the main channel.

Reconstruction of Right Side, Lateral Aspect, Fig. 41.

On the right side, the area receiving the terminals of dorsal
tributaries 1, 2 and 3 in fig. 41, is plexiform and redundant.
A portion of this plexus is condensed into a well-defined veno-
lymphatic channel partially separated from the main vein by a
fenestra through which the arrow passes in the figure. This
channel, the principal anlage of the cephalic division of the
ventral veno-lymphatic plexus, is a much more prominent struc-
ture than its appearance would denote in the figure.

It is also significant in this embryo that dorsal somatic tribu-
taries 1, 2 and 3 are practically not involved in the development
of the cephalic division of the ventral veno-lymphatic plexus,
except in so far as their expanded terminals are included in the
redundant precardinal area which is responsible for the formation
of this portion of the plexus.

The complex formed by the precardinal tributary 4 and its
associated para-precardinal channel has separated into its dorsal
somatic (4S) and veno-lymphatic (4VL) components.

The dorsal somatic component of tributary 4 (4S, fig. 41)
opens into the dorso-inedial circumference of the promontory and
now forms the first of the promontorial series of somatic tribu-
taries (4S, 5S and 6S, fig. 41).

The veno-lymphatic derivative of the primary dorsal tributary
4 (4VL, fig. 41) consists of an expanded, dilated and fenestrated
sac which communicates with the precardinal at its primary point
of connection with the latter (X in fig, 41; see also figs. 22 and
23) and which also drains into the promontory through a para-
precardinal channel that has developed from the peri-precardinal
plexus in front of and contiguous to the promontory. Strictly

Development of the Jugular Lymph Sacs. 261

speaking, this para-precardinal channel, in this instance, as well
as in all others, in which it serves as a portal through which tribu-
tary 4 drains into the promontory, must be regarded as constitut-
ing a portion of tributary 4. In addition to its points of communi-
eation with the precardinal and promontory, just mentioned, the
veno-lymphatic component of tributary 4 (4VL, fig. 41) also
communicates with its companion dorsal somatic component (4S,
fig. 41) near the point where the latter opens into the promontory,
a circumstance which betrays the common origin of these two com-
ponents from the primary dorsal plexus of tributary 4 of earlier
stages (see figs. 1/A, 35 and 37). The small diverticular-shaped
process which projects ventrad from the para-precardinal channel
(AP, fig. 41) corresponds to a similarly situated process on the left
side of series 138 (fig. 33).

In front of the primary point of connection of tributary 4 with
the precardinal (X in fig. 41) the latter receives a small inverted
L-shaped tributary (Z in fig. 41) which opens into its dorso-lateral
circumference. This small vessel is probably related to the plexus
of the primary terminal of tributary 4 (X) from which it has been
separated in the condensation of the channels. Its position with
respect to the main channel establishes its veno-lymphatic character
and indicates that it will enter into the formation of the ventral
division of the veno-lymphatic plexus as in series 102, fig. 44.

The dorsal veno-lymphatic plexus was only partially recon-
structed on the right side of this embryo and, therefore, will not be
considered.

Reconstruction of Left Side, Lateral Aspect, Fig. 42.

A comparison of the right and left sides of this embryo affords
an interesting illustration of the unequal chronological development
of the three main divisions of the veno-lymphatic plexus which may
occur upon opposite sides of the same embryo. On the left side
(fig. 42) the cephalic division is much advanced in development
and the caudal division of the ventral plexus extremely rudimentary,
while on the right side (fig. 41) the reverse conditions obtain.

The anlage of the external jugular vein is well developed and
opens into the promontory.

262 George 8S. Huntington and Charles F. W. McClure.

The anterior part of the straight precardinal segment, in the area
corresponding to the dorsal tributaries 1, 2 and 3, carries on its
dorso-lateral aspect a large diverticular sac, which extends caudad
in the direction of the promontory. This sac, produced by conden-
sation of the peri-precardinal capillary network. forms the anterior
or cephalic division of the ventral veno-lymphatic plexus (cj. fig. 9).
On the right side of this embryo (fig. 41) this structure is in the pro-
cess of differentiating from the main vein. In the later stages it
will fuse with the dorsal plexus derived from the promontory, and
with the caudal division of the ventral plexus, and will then form
the anterior portion of the continuous veno-lymphatic sac (fig. 42,
area of 1,2and3). Atits anterior end it is still in wide-open communi-
cation with the precardinal and it terminates caudally in a blind
and deeply bifurcated extremity which overlaps the dorso-lateral
aspect of the straight precardinal. When the cephalic division of
the ventral plexus is formed in this manner the precardinal tribu-
taries 1, 2 and 3 appear to be taken up as a whole in the veno-lym-
phatie organization and do not divide into veno-lymphatic and
somatic components as far as we can determine.

The left side of this embryo apparently represents an instance in
which primary dorsal tributary 4 of the precardinal has been trans-
formed in its entirety into a dorsal somatic promontorial branch (48) and
has not contributed a component to the ventral veno-lymphatic
plexus.

It may be possible that the more dorsal of the two processes
(4VL’) which, form the bifurcated extremity of the anterior veno-
lymphatic sac has been derived from tributary 4 and has been sec-
ondarily joined to the’sac. If such is the case, we are not in a
position to prove it, and must, therefore, abide by our original
contention that tributary 4 per se, in this particular instance,
does not furnish a veno-lymphatic component.

The dorsal somatic tributary 4 (4S, fig. 42) is formed by two
confluent branches which enter the dorso-medial aspect of the pro-
montory at the promontorio-precardinal angle. The vessel is
expanded and fenestrated near its promontorial connection and les
in line with a series of promontorial and postcardinal dorsal somatic
tributaries (5S, 6S, 7S and 8S, fig. 42), of which it forms the first
or most anterior member of the series.

~~ ss

Development of the Jugular Lymph Sacs. 263

The para-precardinal channel, so prominently developed on the
right side in connection with the veno-lymphatic component. of
tributary 4 (fig.41),is represented on the left side by a small diver-
ticular process of the promontory which is connected with the dorso-
lateral circumference of the latter (para-precardinal channel, fig.
42). An examination of the actual reconstruction of the left side
shows more clearly than is indicated in the figure that this short
stump-like para-precardinal element was probably continuous at
one time with the more ventrally situated of the two processes which
form the posterior extremity of the cephalic division of the veno-
lymphatic plexus or sac, and that it has separated from the latter,
during the course of development. Such as it is,however, it repre-
sents in its entirety, on the left side of this embryo, all that is present
of the caudal division of the ventral veno-lymphatic plexus, which is
well developed on the right side (fig. 41), as well as in certain embryos
of the earlier stages (fig. 82). (Cf. also fig. 21).

The dorsal! veno-lymphatic plexus of the left side (fig. 42) is
extensively developed. It has given up its connection with the
anterior end of the primitive ulnar anlage and lies further from the
promontory than in earlier stages (fig. 31), as the result of an en-
largement and elongation of the fenestree which lie between its
promontorial connections. The dorsal veno-lymphatic plexus, as
already described, has been separated by fenestration from the dorso-
lateral circumference of the promontory and primitive ulnar segment
of the posteardinal, and its general phases of development can now
be easily followed by comparing the preceding figures.

The anlage common to the primitive ulnar vein and the primitive
ulnar veno-lymphatic is fully established and opens into the dorso-
lateral circumference of the promontory about opposite the latter’s
point of confluence with the seventh dorsal somatic tributary (7S, fig.
42). The anlage of the primitive ulnar veno-lymphatic has not
yet begun to separate from the anlage common to it and the primitive
ulnar vein.

The anlage of the cephalic vein has made its appearance on the
left side of this embryo as a small vessel which opens laterally into
the promontory, dorsal to the anlage of the permanent external
jugular vein.

264 George 8. Huntington and Charles F. W. McClure.

This embryo shows most clearly that the veno-lymphatic area
of the promontory has differentiated along its dorso-lateral cir-
cumference, leaving its dorso-medial aspect free for the entrance of
the dorsal somatic tributaries (4S, 5S, 6S and 7S, fig. 42). A
similar dorso-lateral position with respect to the precardinal has
already been described and emphasized as being characteristic of
the veno-lymphatic derivatives of its dorsal tributaries (see figs.
33, 34, etc.).

In general, the relative position of the dorso-lateral veno-lymphatie
(dorsal veno-lymphatie plexus and caudal division of ventral veno-
lymphatic plexus) and the dorso-medial somatic (48, 5S, 6S) ele-
ments with respect to each other and to the precardinal vein, is
clearly shown in fig. 43, which is a photomicrograph of a transverse
section of al0 mm. cat embryo taken at a level just anterior to
the jugular promontory (series 114, slide 4, section 27).

Series 102, 8.5" Embryo
Reconstruction of the left side
Lateral aspect, fig. 44 and
Medial aspect, fig. 45
Reconstruction of right side,
Lateral aspect, fig. 46

Although this embryo measures less than other members of
this group, due probably to a greater degree of body and head
curvature, it represents a more advanced stage of development,
which bridges the interval between the intermediate and defi-
nitely established veno-lymphatic periods.

Reconstruction of Left Side, Lateral Aspect, Fig. 44.

The three primary divisions of the veno-lymphatic plexus are
fully established.

With the exception of the element contributed by dorsal tribu-
tary 1 (1VL), which forms a dilated and fenestrated sac, connected
with the precardinal, all of the components of the ventral division
of the veno-lymphatic plexus have fused with one another to
form a large and multi-fenestrated sac.

EE ——

Development of the Jugular Lymph Sacs. 265

The cephalic division of the ventral plexus has been formed by
tributary 1 (1VL) and from the area of the precardinal which
receives dorsal tributaries 2 and 3 (2VL and 3VL), the latter
two having retained their original precardinal terminations.

The caudal division of the ventral veno-lymphatic plexus is a
capacious and multi-fenestrated sae which lies dorso-lateral to the
precardinal and for the most part in front of the promontory. It
is continuous in front with the anterior division of the ventral
plexus and communicates with the promontory by two well-de-
fined apertures: one, the smaller of the two, being situated near
the promontorio-precardinal angle on the lateral surface (Tap B,
fig. 44), and the other (Tap D, fig. 44) on the cephalic surface of
the promontory in close association with the promontorial end of
the dorsal somatic tributary 4 ( 4S, fig. 45).

This capacious sac which constitutes the caudal division of the
ventral veno-lymphatic plexus has been formed as the result of a
further development of a condition similar to that found in series
109 (fig. 30), series 2 (fig. 32) or series 19 (fig. 41), and has been
derived either from a para-precardinal channel or from a para-pre-
cardinal channel.in connection with a veno-lymphatic component
derived from precardinal tributary 4.

Although we know, in general, the elements which may enter
into the formation of the ventral plexus, on account of the vari-
ability of the process, it is extremely difficult to determine defi-
nitely in all cases of advanced stages exactly how its formation
has been brought about. In view of the intimate relations which
exist between this plexus and the first of the promontorial dorsal
somatic tributaries (4S) which was derived from precardinal trib-
utary 4, we are inclined to believe that in this case the veno-
lymphatic component of the latter as well as the para-precardinal
channel have entered into its formation.

The dorsal veno-lymphatic plexus is represented by a pouch-
like fenestrated sac which was derived from the dorso-lateral
aspect of the promontory. It projects forward, dorsal to the
caudal division of the ventral veno-lymphatic plexus, and now
communicates with the promontory by only a single narrow open-
ing (Tap C, fig. 44) near the former’s point of confluence with the

266 George 8S. Huntington and Charles F. W. McClure.

primitive ulnar vein. As mentioned above, in connection with
the description of series 106 (fig. 40), this condition has been
reached as the result of a separation of the sac from the anterior
end of the anlage common to the primitive ulnar vein and prim-
itive ulnar veno-lymphatic, and by the retention of only one of
its primary promontorial points of connection. This single tap
(Tap C, fig. 44) corresponds, approximately, in its position to that
of the lymphatico-venous connection of later stages which is found
at the jugulo-subelavian junction, while the more ventrally situ-
ated tap (Tap 2) corresponds, approximately, to that met with at
the junction of the external and internal jugular veins.

The thyro-cervical artery passes laterad close to the surface
of the promontory, between the dorsal plexus and the caudal divi-
sion of the ventral veno-lymphatic plexus.

This artery, from this stage on, is constantly encountered in
this position and offers a uniform relationship to the components
of the veno-lymphatie plexus. It thus constitutes an important
landmark in differentiating in later stages and in the adult the
parts of the united jugular lymph sac derived, respectively,
from the dorsal plexus and caudal division of the ventral veno-
lymphatic plexus.

The promontorial end of the dorsal veno-lymphatic plexus
constitutes the subclavian approach of later stages and can al-
ways be identified by the constant relation which it holds to the
ventrally situated thyro-cervical artery.

The anlage common to the primitive ulnar vein and primitive
ulnar veno-lymphatic has separated into these two components.
The former arches over the sixth spinal nerve (SP.N.VI) and
opens into the dorso-lateral circumference of the promontory be-
tween the latter’s point of confluence with the sixth and*seventh
dorsal somatic tributaries (68 and 7S, figs. 44 and 45). The prim-
itive ulnar veno-lymphatie is a small vessel which runs parallel
to the primitive ulnar vein. It has been completely differentiated
from this vein except at its anterior end where indications of its
former connection with the same are still evident. At a later
stage it will completely separate from the primitive ulnar vein and,
after joining the dorsal veno-lymphatie plexus, form a caudal
extension of the latter.

Development of the Jugular Lymph Sacs. 267

Series 102, 8.5" Embryo
Reconstruction of left side
b
Medial aspect, fig. 45

This view of the reconstruction shows three tiers of elongated
fenestrae and deep depressions on the medial surface of the caudal
division of the ventral veno-lymphatic plexus which appear to
divide it into four parallel channels. Taken by itself this view
of the reconstruction suggests that this portion of the plexus has
been formed as the result of a fusion between four parallel vessels.
Such is not the case, however, since the ontogenetic history of
this plexus proves conclusively that these parallel vessels are
secondarily formed as the intervals between the fenestral spans
developed in the plexiform anlage of this structure.

The medial view of the reconstruction also shows the extensive —
and dilated character of the promontory and the relations which
it holds to its dorsal somatic (4S, 5S, 6S, and 7S, fig. 45) and
veno-lymphatic structures (Cf. also fig. 43).

Series 102, 8.5"”" Embryo
Reconstruction of right side,
Lateral aspect, fig. 46

As on the left side of this embryo the three primary divisions
of the veno-lymphatic plexus are fully established on the right
side.

I. VENTRAL VENO-LYMPHATIC PLEXUS
1. Cephalic Division

The precardinal dorsal tributary 1 (1 VL) is represented by a
dilated, elongated and fenestrated sac which has retained its pri-
mary connection with the precardinal. It has not yet fused at
its base with the adjacent veno-lymphatic components (2 VL,
3 (VL, and in this respect both sides of this embryo agree. In
addition to its contribution, at its base, to the ventral plexus, this
veno-lymphatie component of tributary 1 (1 VL) will, by its
further dilation and caudal extension, fuse with the dorsal veno-

268 George 8. Huntington and Charles F. W. McClure.

lymphatic plexus and thus contribute to the forward extension of
the latter. If, in the meantime, it still retains its primary con-
nection with the precardinal, this connection may serve*as a por-
tal through which the blood is evacuated from the plexus into
the systemic veins (Tap of evacuation).

The distinctive element of the cephalic division of the ventral
plexus in this region is represented by a capacious sac which has
shifted to the lateral aspect of the precardinal and which com-
municates with the same by a single opening (Tap A’). This sac
occupies an area of the precardinal which formerly received the
dorsal tributaries 2 and 3. The primary connection of one of
these two tributaries (probably 2) with the precardinal has un-
doubtedly persisted as the opening through which the sac now
communicates with the vein, while the sac itself has probably
been formed through a fusion of the veno-lymphatic components
of these two tributaries, possibly in conjunction with an element
derived from the plexiform dorso-lateral circumference of the main
channel of the precardinal vein.

The cephalic portion of the ventral division of the veno-lym-
phatic plexus is not always developed to the extent shown in the
present embryo but, when present, its origin can always be traced
back to the veno-lymphatic components of the precardinal trib-
utaries 1, 2 and 3, to the area of the precardinal which receives
these tributaries, or, to both of these sources.

2. Caudal Division.

The caudal division of the ventral veno-lymphatic plexus,
which has undoubtedly been derived from the veno-lymphatic
component of precardinal dorsal tributary 4 and its related para-
precardinal channel, forms an extensive sac with three elongated
cephalic processes. The sac has shifted laterad and caudad on to
the lateral aspect of the precardinal and promontory, a position
which becomes more pronounced in later stages. It communicates
laterally (fig. 46) with the precardinal near the promontorio-
precardinal angle (Tap 6’) and with the promontory slightly cau-
dal to this point (Tap B).

7

Development of the Jugular Lymph Sacs. 269

The dorsal somatic component of tributary 4 (4S) is entirely
detached from the veno-lymphatic plexus, and enters, as the first
of the somatie series, the dorso-medial aspect of the promontory.

II. Dorsat VeENo-LYMPHATIC PLEXUS.

This is a much dilated and fenestrated sac which lies dorso-
lateral to the promontory, and which has been derived from the
same. It extends forward for some distance in front of the prom-
ontory and communicates with the latter by a narrow channel
which opens into the promontorial end of the primitive ulnar
vein (Tap C). The presence of a single point of communication
between the dorsal plexus and the promontory at the primitive
ulnar entrance corresponds to the conditions already described
for the left side of this embryo (Tap C, fig.44). The relations of the
the dorsal plexus to the thyo-cervical artery are also the same and
need, therefore, no further mention.

The primitive ulnar vein makes a distinct arch in its terminal
portion and receives a long cephalic tributary which follows the
dorsal border of the dorsal veno-lymphatic plexus. The primitive
ulnar veno-lymphatic was not fully reconstructed on this side of
the embryo but presents about the same conditions as on the left
side.

In addition to the three primary veno-lymphatic plexuses already
described, a number of detached veno-lymphatic elements are
met with on the right side of this embryo. These are four in num-
ber and are labelled VL’ in fig. 46. The significance and subse-
quent fate of these detached veno-lymphatics has already been
discussed on a preceding page (page 259) and therefore need not
be further considered here.

The medial aspect of this reconstruction, which is not figured,
like that of the left side (fig. 45), offers a typical instance of the
serial arrangment of dorsal somatic tributaries, beginning with
4S, which enter the dorso-medial aspect of the promontory.

As this embryo serves as a connecting link between the early
and later veno-lymphatic stages, it may be well to summarize
the principal features which it illustrates.

270 ~=69©George 8. Huntington and Charles F. W. MeClure.

1. Gradual reduction of the multiple early veno-lymphatico-
venous connections, but retention, in some cases, of the primary
precardinal connections of tributaries 1, 2, and 3.

2. The gradual establishment of two definite points of com-
munication between the promontory and the dorsal (Tap C)
and ventral (Tap B) veno-lymphatic plexuses, respectively, which
are usually the last to be retained during the separation process and
which correspond, approximately, in their position to the points
where the jugular lymph sae taps the venous system in the adult.

3. Clear differentiation of the veno-lymphatic plexus into
three primary divisions and the beginning of a fusion between the
two divisions of the ventral plexus.

4. Complete separation of the dorsal and ventral veno-lym-
phatic plexuses by a clear interval which serves for the transit of
the thyro-cervical artery.

5. Separation of the dorsal plexus from the anterior end of the
anlage common to the primitive ulnar vein and primitive ulnar
veno-lymphatie.

6. Separation from this common anlage of a small parallel

channel which forms the primitive ulnar veno-lymphatic. This
veno-lymphatic joins the dorsal plexus and remains connected with
the caudal end of the same after the primitive ulnar vein has been
replaced in the drainage of the anterior limb by the definite sub-
clavian vein.
7. The presence of a number of detached veno-lymphatics
which subsequently fuse with one another and with the three
primary divisions of the plexus in establishing the single veno-
lymphatic sac of later*stages.

8. The definite establishment of theanlage of the cephalic vein.

TWO EMBRYOS ILLUSTRATING THE LATERAL DISPLACEMENT
OF CERTAIN VENO-LYMPHATICS WHICH ARE SECONDARILY
SEPARATED BY FENESTRATION FROM THE CAUDAL DIVISION
OF THE VENTRAL VENO-LYMPHATIC PLEXUS,

Series 112, 10" Embryo
Series 118, 10™™ Embryo

As the result of an excessive retiform development in the caudal
division of the ventral veno-lymphatic plexus it frequently,

Development of the Jugular Lymph Sacs. 211

though not invariably, happens that a portion of the same becomes
partially or wholly detached and is displaced to the lateral sur-
face of the precardinal vein and the promontory.

This feature of development appears to be of sufficient importance
to warrant a careful consideration, as it further emphasizes the
~variable character of the process of fenestration, as well as the
variable form which may be assumed by this division of the ven-
tral plexus as the result of this process.

The two embryos in question illustrate exceedingly well the
partial separation of a portion of the ventral plexus and its dis-
placement to the lateral surface of the promontory and precardi-
nal vein. They also give us the clue as to the origin of the inde-
pendent and completely isolated veno-lymphatic sacs or spaces
which are met with in certain cases on the lateral surface of the
promontory and precardinal vein (fig. 50).

Series 112, 10°” Embryo
‘Reconstruction of left side,
Lateral aspect, fig. 47

The Caudal Division of the Ventral Veno-Lymphatic Plexus.

The caudal division of the ventral vcno-lymphatic plexus in
this embryo (fig. 47) is a less condensec and compact structure
than in the 8.5 mm. embryo (figs. 44 and 46), Just considered.
This does not necessarily mean that it is less advanced in develop-
ment than in the 8.5 mm. embryo (series 102), but rather that
the plexiform character has been longer retained in it than in the
case of the latter, at a stage which precedes the general amalgama-
tion of all the veno-lymphatics to form a single common sac.
In this embryo (fig. 47) the caudal division of the ventral veno-
lymphatic plexus consist of three distinct and parallel channels.

A large dorsal channel, continuous with the dorsal veno-lvm-
phatic plexus, which joins anteriorly the ventral of the three
channels by two transverse vessels; an intermediate smaller
channel which opens into the lateral surface of the promontory
at Tap B’ and into the dorsal surface of the ventral channel, and
of a large ventral channel lying upon the lateral surface of the pre-

272 George S. Huntington and Charles F. W. McClure.

‘ardinal and promontory, which communicates with the latter at
Tap B and at its cephalic extremity with the precardinal vein
(Tap X).

On comparing the caudal division of the ventral plexus in the
10 mm. (fig. 47) and 8.5 mm. (figs. 44 and 45) embryos, it is evi-
dent that a further confluence of the fenestra in the caudal divi-
sion of the ventral plexus of the 8.5 mm. embryo (fig. 45) would
produce a series of parallel and partially separated channels which
hold the same relations to the precardinal and promontory, as do
the three parallel channels in the 10 mm. embryo (fig. 47). It is
also clear, considering our knowledge of the ventral plexus, in
general, that the ventral and intermediate of the three channels
in the 10 mm. embryo (fig. 47) must have been separated from that
portion of the veno-lymphatic complex of tributary 4 and its
associated para-precardinal channel, which has been contributed
by the para-precardinal channel. ,

A comparison of the 10 mm. (fig. 47) and 7 mm. (series 138,
figs. 33 and 35) embryos also gives us a clue, at this late stage of
development, concerning the origin of the most dorsal of the three
channels which compose the caudal division of the ventral veno-
lymphatic plexus in the 10 mm. embryo (fig. 47).

The Dorsal Division of the Veno-Lymphatic Plexus.

This embryo is characterized by extensive development in the
area of the dorsal division of the veno-lymphatic plexus, and by
the complete separation of the same from the systemic veins
except by means of its connection with the latter through the cau-
dal division of the ventral plexus. It therefore prepares the way
for the consideration of subsequent stages in which the entire
veno-lymphatic plexus, as a closed and empty sac, appears to
separate temporarily from the systemic veins.

The possibilities of forward extension of the dorsal veno-lym-
phatic plexus are represented here by a number of detached and
closed sacs (VL’ in fig. 47), lying in the course of its secondary for-
ward growth, which subsequently by fusion with it and with the
veno-lymphatics of the cephalic division of the ventral plexus, will

Development of the Jugular Lymph Sacs. 213

complete the secondary dorsal veno-lymphatie arch of later stages
(fig. 51, series 77).

The primitive ulnar veno-lymphatic has become detached from
the vein from which it arose by condensation of the surrounding
capillary network, and now enters the caudal extremity of the
dorsal veno-lymphatiec plexus.

The relations of the thyro-cervical artery to the dorsal veno-
lymphatic plexus and to its ventro-caudal extension, the sub-
clavian approach, are the same as in the preceding embryo (series
102, fig.44). The jugular approach is represented by that portion
of the ventral plexus which opens into the promontory at the two
points labelled Tap B and Tap B’ in fig. 47.

The Cephalic Division of the Ventral Veno-Lymphatic Plexus.

The veno-lymphatice components of tributaries 1, 2 and 3 which
constitute the anlages of this division of the ventral plexus (1, 2
and 3VL, fig. 47), although modified by retiform fenestration, still
retain their individuality in this embryo, a condition which may
or may not obtain up to a late stage of development.

The embryo offers a good concrete example of the following
general phases of veno-lymphatic development:

1. Unequal development of the two main veno-lymphatic
plexuses at any given stage.

2. Beginning separation of the veno-lymphatic sac, formed by
further condensation of the plexuses from the definite venous
channels.

3. The formation of isolated and completely closed sacs or
spaces, detached both from the veins and from the main veno-
lymphatic plexuses, which are subsequently incorporated in the
jugular lymph sae.

4. The genesis of the secondary dorsal veno-lymphatic bridge
or arch, which in certain subsequent stages connects the cephalic
end of the dorsal plexus with the anterior veno-lymphatic channels
of the ventral division. This structure which, e. g., appears in
its characteristic development in series 77 and 101 (figs. 51 and 50),
is apparently developed in different individuals to an unequal
degree and in one of the following ways:

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 2.

274 George 8S. Huntington and Charles F. W. McClure.

(a) Cephalic extension of the dorsal veno-lymphatic channel
to union with the cephalic division of the ventral plexus.

(b) Caudal growth of part of the cephalic division of the
ventral plexus until union with the dorsal plexus has been effected.

(c) Development of intermediate detached areas of veno-lym-
phatie character, which, isolated at first, subsequently unite
with both the dorsal veno-lymphatic plexus and with the cephalic
division of the ventral plexus, and thus effect the junction of the
secondary arch with the primary veno-lymphatic elements.

Series 118, 10°" Embryo
Reconstruction of left side
)
Lateral aspect, fig. 48

This embryo is characterized by multiple discrete veno-lym-
phatie development along the entire dorso-lateral circumference
of the preeardinal and promontory, resulting in the formation of
numerous early communication between the veno-lymphatic
plexuses and the main venous channel. It also shows marked
displacement of a portion of the caudal division of the ventral
veno-lymphatic plexus from the dorsal to the lateral aspect of the
- promontory.

The Caudal Division of the Ventral Veno-lymphatic Plexus.

This division of the ventral plexus is composed of a number of
elements, with separate entrances into the precardinal and
promontory. :

These elements group themselves into two main areas, anterior
and posterior, each of which contains subordinate components.

1. Anterior Area of the caudal division of the ventral plexus.

The anterior portion of the caudal division of the ventral plexus
is separated, as in the preceding series 112, into two parallel ele-
ments, a ventral and a dorsal (fig. 48, V and D), which are joined
by a narrow curved piece. ‘The former element (V), representing

Development of the Jugular Lymph Saes. 275

the para-precardinal channel, is displaced ventro-caudad from the
dorsal to the lateral surface of the precardinal and promontory,
forming a dilated sac which communicates with the promontory
at three points (area of Tap B, fig. 48), while in series 112 (fig. 47)
the promontorial end of this correspondingly displaced portion
of the ventral plexus consists of two parallel channels which open
independently of each other into the lateral surface of the prom-
ontory (Tap B, and B’, fig. 47). The difference in form, as well as
the difference in the number of promontorial communications
encountered in the two embryos, is due to varying phases of capil-
lary condensation and amalgamation in the development of these
partially detached portions of the ventral plexus. A comparison
of figs. 44, 45, 47, and 48 clearly shows the transitional stages
leading up to the conditions presented by series 113 (fig. 48).

The second element (D) composing this anterior portion of the
caudal division of the ventral plexus consists of a channel, lying
cephalad of the promontory and parallel to the precardinal vein.
It communicates with the latter at two points, and in addition,
as above stated, is connected by a narrow curved channel with the
first expanded sac-like element (V) on the lateral surface of the
promontory. It, including its dorsal branches (4VL in fig. 48), has
been derived from the original complex formed by precardinal
tributary 4, and the para-precardinal channel.

The original connection of element D with the promontory is,
however, still represented by the most anterior of a series (1’, 2”,
and E’’’) of small veno-lymphatic channels (fig. 48), which form
together the posterior or caudal portion of the caudal division of
the ventral plexus. This small veno-lymphatic (fig. 48, #”) opens
into the promontory in line with the elements D of the anterior
portion of the caudal division, and probably has become second-
arily detached from the same in the course of development. Com-
pare fig. 48 with fig. 33, in which a similar separation of the ventral
plexus from the promontory at B has taken place, and with fig.
42 in which the caudal division of the ventral plexus is alone
represented by a small promontorial veno-lymphatic, the remain-
der of the complex of tributary 4 having developed into dorsal
somatic tributary 4S.

276 ~=George 8S. Huntington and Charles F. W. McClure.

2. Posterior Area of the caudal division of the ventral plexus.

In this embryo, as clearly indicated by the course of the thyro-
cervical artery, the caudal division of the ventral plexus receives
an addition of plexiform veno-lymphatiec anlages (fig. 48, #’, H”,
E’"’) which communicate with the lateral surface of the promon-
tory by five openings. This veno-lymphatic plexus intervenes
between the caudal termination of the two elements (V and D)
forming the anterior portion of the caudal division of the ventral
plexus, and the dorsal veno-lymphatic plexus. They are undoubt-
edly destined to fuse with each other and with the remaining
components of the caudal division. After this fusion has occurred
the thyro-cervical artery will then occupy its typical and invari-
able position, arching over the shoulder of the promontory be-
tween the dorsal veno-lymphatie plexus and the united caudal
division of the ventral plexus.

The Dorsal Veno-Lymphatic Plexus.

The dorsal veno-lymphatie plexus of this embryo (fig. 48) is
still incompletely separated from the promontory. It forms a sac-
like fenestrated structure which opens into the distal portion of
the promontory (fig. 48, Tap C), as well as into the primitive ulnar
arch.

The Cephalic Division of the Ventral Veno-Lymphatic Plexus.

The veno-lymphatic components of precardinal tributaries 1,
2 and 3 (fig. 48, 1VL, 2VL 3VL), which will unite to form the
cephalic division of the ventral plexus, are still individually
distinct; IVL and 2VL are united by a narrow channel, 3VL is
still separate, partially hidden in the figure by the second spinal
nerve (fig. 48, SP.N.IT).

In addition to the separation of the caudal division of the ven-
tral plexus into separate components and the displacement of
one of these components from the dorsal to the lateral surface of
the promontory, this embryo is characterized by the large number
of discrete connections which still persist between the veno-lym-
phatics and the systemic veins. Beginning with precardinal tribu-

(= Yor d

Development of the Jugular Lymph Sacs. Pir

tary 1 (fig. 48, 1VL) as many as sixteen of these connections are
met with along the precardinal vein and the promontory.

FULLY DEVELOPED VENO-LYMPHATIC STAGE.

Series 101, 10” Embryo
Series 77, 11°” Embryo

Series 101 is characterized by enormous cephalic extension of
the dorsal veno-lvmphatice plexus, which, as a minutely fenes-
trated multilocular sac, extends forward to the level of the ceph-
alic arch of the precardinal.

The two main divisions of the ventral veno-lymphatic plexus
are well differentiated, but still entirely separate from each other
and from the dorsal plexus.

Series 101, 10°” Embryo
Reconstruction of left side,
Lateral aspect, fig. 49

The group of veno-lymphatics which constitutes the cephalic
division of the ventral veno-lymphatic plexus, derived from the
precardinal tributaries 1, 2 and 3, is represented by two indepen-
dent dilated sacs both of which still retain a connection with the
precardinal. The united trunk of the first and second spinal
nerves (SP.N.J and JJ) passes ventro-laterad between the ceph-
alic of these two sacs and the precardinal, while the third nerve
(SP.N.IIT) passes between the caudal sac and the dorsal veno-
lymphatic arch.

The caudal division of the ventral veno-lymphatic plexus is an
irregular-shaped sac which lies upon the lateral surface of the pre-
cardinal and promontory. It communicates with the venous sys-
tem at only one point (Tap B), by a wide orifice situated on the
lateral surface of the promontory near the promontorio-precard-
inal angle. This sac must be regarded as representing a further
development of the para-precardinal channel and the associated
veno-lymphatic component of precardinal tributary 4.

In conformity with a greater degree of veno-lymphatic develop-
ment, the caudal division of the ventral plexus has given up its

278 George 8. Huntington and Charles F. W. McClure.

multiple connections with the precardinal and promontory and
retained only one which corresponds, approximately, to the pri-
mary point of connection of the para-precardinal channel with
the promontory. This point of connection, as mentioned above.
is one of the last to be given up, and also corresponds, approxi-
mately, in its position to that of the common jugular tap in the
adult (cf. fig. 1, left side).

Just dorsal to the entrance of the cephalic vein into the lateral
surface of the promontory, the thyro-cervical artery passes ventro-
laterad in the interval between the ventral and dorsal divisions of
the veno-lymphatic plexus.

The dorsal veno-lymphatic plexus is now a very extensive
structure which extends from the region of the promontory, as
far forward as the level of the cephalic arch of the precardinal
where it ends blindly. In the region of the promontory the dorsal
plexus is still broadly confluent with the anlage common to the
primitive ulnar vein and primitive ulnar veno-lymphatic which
has not as yet separated into its two components. The line of
cleavage along which the separation will take place, however, is
clearly indicated by a row of fenestrae which will subsequently
become confluent. (Compare with fig. 47).

Just in front of its connection with the primitive ulnar arch
the dorsal plexus gives off a ventrally directed tongue-shaped
process (subclavian approach) which communicates with the prom-
ontory by a single opening (Tap CY and which les near the en-
trance into the promontory of the primitive ulnar vein. This
point of communication with the promontory corresponds in all
respects to that previously described for this plexus in connection
with series 102 (Tap C, figs. 44 and 46).

This extensive development of the dorsal veno-lymphatic
plexus is the result of a forward growth of the redundant plexi-
form area of the promontory and convexity of the primitive ulnar
arch and inclusion within the same of detached veno-lymphatics
which have been derived from the dorsal tributaries of the pre-
cardinal.

The fifth spinal nerve (SP.N.V) penetrates the dorsal plexus,
while the sixth (SP.N.VJ), and following spinal nerves in this

a

Development of the Jugular Lymph Sacs. 279

region pass between the primitive ulnar anlage and the main
systemic venous channel.

Series 101, 10°" Embryo
Reconstruction of right side,
Lateral aspect, fig. 50

5)

The right side of this embryo (fig. 50) offers an interesting com-
parison with the veno-lymphatic development found on the left
side (fig. 49).

A. Dorsau Veno-LympHatic Plexus.

The dorsal sac presents the same forward extension and multi-
fenestrated appearance as that on the left side, but its relations
to the promontory and to the straight segment of the precardinal
differ in a number of important features from those of the oppo-
site side.

1. On the right side (fig. 50) the dorsal plexus is plainly separ-
ated into two portions, one derived from the promontory proper,
the other from the arch of the primitive ulnar vein.

The posterior and smaller part of the general dorsal veno-lym-
phatic plexus is not connected, as it is on the left side, with the
larger anterior part.

2. The anterior end of the dorsal plexus has established a con-
nection, near the level of the cephalic arch, with the straight seg-
ment of the precardinal (Anterior Tap of Evacuation). This —
connection results from the fusion between the anterior end of
the dorsal plexus and the veno-lymphatic component of a dorsal
tributary of the straight segment of the precardinal which has
retained its primary connection with the latter, or, which, in case
of early multiple precardinal terminals, has continued to develop
one of these, thus accounting for the difference in the level of the
first precardinal veno-lymphatiec tap observed on the two sides of
this embryo in reference to the level of the spinal nerves.

On the left side the abandonment of the early precardinal termi-
nal of tributary 1, together with the retention of the terminals of
tributaries 2 and 3, would produce the relations to the segmental
nerves found on this side of the embryo, while the relations of the

280 George 8. Huntington and Charles F. W. McClure.

nerves to the veins and veno-lymphatics of the right side suggest
that here the terminal of tributary 1 has been retained, while
tributaries 2 and 3 have lost their precardinal connections. Thus
on the left side the common trunk of the first and second cervical
nerves comes to lie cephalad of all veno-lymphatic terminals into
the precardinal, while on the right side this trunk is placed caudad
of the first veno-lymphatie precardinal tap.

3. On the left side (fig. 49) the extension cephalad of the dorsal
plexus involves both the portion derived from the promontory
and that furnished by the primitive ulnar arch, both components
uniting in the plexiform network of the dorsal veno-lymphatic
arch. On the right side (fig. 50) the primitive ulnar arch yields a
relatively small plexiform veno-lymphatic cephalic prolongation,
which, while undoubtedly destined to unite eventually with the
other components of the dorsal veno-lymphatic arch to form its
caudal extension is, in this stage, still separate and discrete. The
arch is in this instance evidently produced by cephalic extension
of that portion of the dorsal plexus, which was originally derived
from the promontory, but which, in establishing a secondary ante-
rior connection with one of the veno-lymphatic compounds of the
cephalic division of the ventral plexus, has given up its original
promontorial connection in the area of Tap C (cf. fig. 44).

4. On the right side (fig. 50) the promontorial portion of the
dorsal plexus has given up its original connection with the prom-
ontory, still present on the left side, and now ends in this region
in a caudal blind prolongation which lies dorsal to the thyro-
cervical artery, and constitutes that portion of the future lymph
sac which we have termed the subclavian approach.

5. This right side of this embryo (fig. 50) illustrates an impor-
tant phase in veno-lymphatic development, which, in a more
advanced condition, is found in series 78 (fig. 57). In embryos
ranging between 10 and 12.5 mm. the dorsal veno-lymphatic
plexus makes a secondary, although temporary, connection with
the anterior part of the straight segment of the precardinal, and
in doing so, often gives up its original promontorial connection
(series 78, fig. 57). The secondarily acquired anterior connection
with the preecardinal vein apparently serves for the evacuation of

_——

Development of the Jugular Lymph Saes. 281

the blood-contents from the plexiform sac into the systemic veins.
After the process of evacuation is completed this precardinal con-
nection is abandoned. It is not possible to state definitely that this
anterior tap of evacuation invariably develops in all cat embryos,
but we can state that it occurs with frequency in embryos rang-
ing between 10.5 and 12.5. mm. in length. (Cf. p. 291.)

B. VENTRAL VENO-LYMPHATIC PLEXUS.
1. Cephalic division

This portion of the ventral plexus (fig. 50) is represented by two
veno-lymphaties, one of which has fused with the anterior pro-
longation of the dorsal veno-lymphatic plexus while the other, a
small dilated sac, fenestrated at its base and still connected with
the precardinal, lies directly caudal to the anterior tap of evacu-
ation.

2. Caudal division.

A considerable difference is met with on opposite sides of this
embryo concerning the make-up of this portion of the ventral
plexus. On the left side (fig. 49) it consists of a single, irregular
sac which communicates with the promontory by a single opening
(Tap B, fig. 49). On the right side (fig. 50) it is composed of two
entirely detached and independent sacs and of a portion whieh still
communicates with the systemic veins at three points.

The two detached and independent sacs are completely closed
and, as far as we can determine under a high magnification, do
not communicate with the venous svstem. The anterior and
Jarger of the two sacs is irregular in form and lies on the lateral
surface of the preeardinal and promontory near the promontorio-
precardinal angle. In its form and position it closely resembles
the caudal division of the ventral plexus on the left side of the
embryo (fig. 49). It contains no blood corpuscles and from a
histological standpoint cannot be distinguished from a typical
lymphatic structure. The other closed sae lies upon the lateral
surface of the promontory, slightly dorsal and caudal to the first
and between the latter and the anlage of the cephalic vein. This
sac still contains a large number of blood corpuscles.

282 George S. Huntington and Charles F. W. McClure.

The remaining portion of this plexus presents an appearance
which has been shown to be typical for this structure in a number
of the preceding stages (e. g. fig. 48). It consists of a vessel running
parallel to the precardinal, somewhat arched anteriorily, which
opens into the precardinal at two points (Tap X and Tap X’,
fig. 50) and into the lateral surface of the promontory by a single
orifice (Tap B). Opposite its point of communication with the
precardinal at Tap X’, it is connected by means of a very narrow
stalk with a dilated oval blind sae whose caudal extremity ap-
proaches the promontory just in front of the thyro-cervical artery.
This expanded appendage to the caudal portion of the ventral veno-
lymphatic plexus is an important feature of development and
appears to represent a condition frequently obtained by this plexus
previous to its subsequent amalgamation with the dorsal plexus
and the cephalic division of the ventral veno-lymphatic plexus to
form a common sac (cf. fig. 14).

Although exactly the same conditions seldom prevail in any
two embryos, it is not difficult to interpret the origin of the caudal
division of the ventral plexus in the present instance on the basis
of a further development of the conditions shown in series 19
(fig. 41, 9 mm. embryo).

The channel of the caudal division of the ventral veno-lymphatic
plexus which, in the 10 mm. embryo (fig. 50, series 101) runs
parallel to the precardinal and taps the systemic veins at three
points (Tap X, X’ and B), has its homologue in the 9 mm. embryo
(fig. 41, series 19) in the para-precardinal channel and the element
L which has been separated from the latter, while the oval blind
sac in series 101 (fig. 50) which is connected with this parallel
channel by a narrow stalk has been derived from the veno-lym-
phatiec component of precardinal tributary 4 in series 19 (4VL,
fig. 41). On the other hand, the two detached and closed sacs in
series 101 (fig. 50) are the secondary products of the original para-
precardinal plexus (fig. 41, see also fig. 31) which have become
separated from the same by an increase in the process of conden-
sation, as in figs. 47 and 48.

The transformation of the complex of tributary 4 and the para-
precardinal channel into two or more parallel channels is a feature

ee eee

Developmert of the Jugular Lymph Sacs. 283

of common occurrence, and is well illustrated by series 102 (fig.
45) and by series 112 (fig. 47) in which two such channels are
met with opening into the lateral surface of the promontory at
Tap B. In the latter series, if these two channels were to separ-
rate completely from each other and from the remaining portion
of the ventral plexus, they would oecupy the same relative
position with respect to each other and to the precardinal vein
and promontory as do the two closed sacs in series 101 (fig. 50).

Series 77 mbryo
S ie EB b y
Reconstruction of left side,
Lateral aspect, fig. 51 and
) aD
Reconstruction of right side,
Lateral aspect, fig. 52

This 11 mm. embryo offers an exceedingly interesting and sug-
gestive comparison with the 10 mm. embryo just considered (figs.
49 and 50), since it further emphasizes two principal features of
development first noted in the latter:—(1) the forward extension of
the dorsal veno-lymphatic plexus and the establishment of an
anterior tap of evacuation and (2) the beginning of the amalgama-
tion of the three primary divisions of the veno-lymphatic plexus
to form a common single veno-lymphatic sac.

On both sides of the embryo, (figs. 51 and 52) an anterior tap
of evacuation has been established at the level of the cephalic arch,
between the anterior end of the dorsal veno-lymphatic arch and
the straight segment of the precardinal. As mentioned above, we
have every reason to believe that this communication, secondarily
established, between the dorsal arch and the precardinal, repre-
sents a normal phase of development and serves as the main, if
not the only exit, in some cases, by which the blood is evacuated
from the veno-lymphatic sac into the systemic veins.

Further, on both sides of thisembryo (figs. 51 and 52), the cephalic
division of the ventral veno-lymphatic plexus has become incorpor-
ated by fusion with the anterior end of the dorsal plexus so that
this portion of the ventral plexus no longer appears as a separate
structure.

On the left side (fig. 51) the dorsal veno-lymphatic plexus com-

284 George 8S. Huntington and Charles F. W. McClure.

municates with the lateral surface of the promontory, through the
subelavian approach, by a single opening (Tap C). This point of
communication lies between the openings into the promontory of
the cephalic and primitive ulnar veins and corresponds in its
position and relations to a promontorial communication of the
dorsal plexus observed in series 102 and 101 (Tap C, figs. 44, 46,
and 49). In addition to this promontorial communication the
dorsal plexus still retains a connection with the primitive ulnar
vein and also communicates with the caudal division of the ven-
tral veno-lymphatic plexus by a narrow transverse channel. Cau-
dally the dorsal plexus of the left side (fig. 51) receives the primi-
tive ulnar veno-lymphatic which is now completely separated
from the primitive ulnar vein. The latter, however, has retained
its original connection with the dorsal plexus, but has differentiated
from the posteardinal and promontory except at its anterior end
where it arches over the sixth spinal nerve (SP. N.V/J, fig. 51).

On the left side (fig. 51) the caudal division of the ventral veno-
lymphatic plexus consists of a single sac which, as mentioned above,
communicates by a narrow channel with the dorsal plexus and by
a single opening (Tap B) with the lateral surface of the promon-
tory near the union of the latter with the cephalic vein. ‘This
poitt of communication with the promontory (Tap B) corresponds,
approximately, in its position and relations to the promontorial
communication of the ventral plexus observed in series 102, 112
and 101 (Tap B, figs. 44, 46, 47, 49 and 50), and the portion of the
ventral plexus directly involved in the communication consti-
tutes that portion of the future jugular lymph sae which we have
termed the “jugular approach.” By tracing back its relations to the
promontory in the above mentioned series, it will be seen that the so-
called ‘jugular approach’ has its origin in the para-precardinal
channel of earlier stages and that the communication at Tap B repre-
sents one of the persistent openings by which the para-precardinal
channel and its associated veno-lymphatic component of precardinal
tributary 4 communicates with the promontory.

The caudal division of the ventral veno-lymphatic plexus, as
mentioned above, has assumed a characteristic form which appears
to precede its fusion with the dorsal plexus. It is composed of a

Development of the Jugular Lymph Saces. 285

ventrally situated elongated portion which lies along the lateral
surface of the precardinal and promontory and of an expanded,
oval dorsal portion which is flexed mesad so that it lies directly
upon the dorsal surface of the precardinal, just in front of the
promontory (figs. 14 and 20). The promontory, however, does
not form a prominent swelling of the main venous channel on the
left side of this embryo.

On the right side of this embryo (fig. 52), in addition to the an-
terior tap of evacuation, the dorsal veno-lymphatie plexus com-
municates with the systemic veins at three points, all of which are
situated on the dorso-lateral surface of the promontory. The
most anterior of these three points of communication (Tap C),
made through the subclavian approach, lies dorsal to the cephalic
vein and caudal to the thyro-cervical artery and in this respect
corresponds to the Tap C observed on the opposite side of the
embryo (Tap C, fig. 51). The two points of communication caudal
to Tap C on the right side (fig. 52) represent a condition in which
the process of separation of the dorsal plexus from the promon-
tory is less complete, owing to the establishment of secondary
connections representing the anlage of the subclavian vein ter-
minal. The primitive ulnar veno-lymphatic is fully established
on the right side but, as it was not reconstructed, has not been
indicated in the figure.

In addition to its connection with the systemic veins, the dorsal
veno-lymphatic plexus also communicates, by two narrow trans-
verse channels, with the caudal division of the ventral plexus.
These two connections, as well as the single one of the opposite side,
must be regarded as secondary communications which have been
established between the dorsal and ventral plexus preliminary to
a complete fusion between the two. The composition of the cau-
dal division of the ventral plexus is essentially the same as on the
left side. It consists of an elongated portion, which lies along the
lateral surface of the precardinal and promontory, and communi-
cates with the latter (Tap B) near the entrance into the promon-
tory of the cephalic vein,and of a dorsal oval, expanded portion
which is flexed mesad so that it lies upon the dorsal surface of the
precardinal, just in front of the promontory. The promontory is

286 George 8. Huntington and Charles F. W. McClure.

a prominent structure on the right side, its outline being indicated
in the figure by a broken line.

The relation of the first five spinal nerves to the veno-lymphatic
plexusesis indicated in the reconstruction of the right side (fig. 52).
The first and second (SP.N.J and JJ), third (SP.N.JII) and
fourth (SP. N. IV) nerves pass ventro-laterad between the cephalic
and eaudal divisions of the ventral veno-lymphatic plexus, and
the fifth (SP.N.V) nerve passes directly through the dorsal
plexus. The sixth spinal nerve (SP.N.V/J, fig. 51) lies behind
the dorsal plexus and arches ventro-laterad under the primitive
ulnar vein.

The thyro-cervical artery, as in the preceding stage, lies ventral
to the subclavian approach and between the dorsal and ventral
divisions of the veno-lymphatic plexus.

These relations of the spinal nerves are retained in later stages
after the primary divisions of the veno-lymphatic plexus have
fused to form a common sac. Subsequently, after the comple-
tion of the veno-lymphatic process, the resulting jugular lymph
sac divides at its anterior extremity (fig. 17), thus freeing the
nerves, which then come to pass, first between the dorsal portion
of the sac and the internal jugular vein, and, finally, entirely dor-
sal to the sac. The relation of the thyro-cervical artery to the
lymph sac is, however, retained throughout and persists in the
adult.

In view of the importance of the two embryos, which consti-
tute this group (series 101 and 77), since they represent a stage of
development which immediately precedes the establishment of
the jugular lymph sacs, it may be well to tabulate the principal
features which they illustrate, as follows:

Series 191, 10°" Embryo
Figs. 49 and 50

1. This embryo furnishes an excellent example of the three
main divisions of the veno-lymphatic plexus, fully developed, but
still quite distinct from each other.

2. The extensive forward growth of the dorsal veno-lymphatic
plexus and the establishment, on the right side, of an anterior

a ee ae Po

Development of the Jugular Lymph Sacs. 287

tap of evacuation through its union with a dorsal tributary of the
precardinal.

3. The gradual reduction in the number of early multiple
connections which exist between the veno-lymphatic plexus and
the veins.

4. The complete separation of the dorsal plexus from the
promontory on the right side and the persistence of a single point
of connection on the left through the subclavian approach (Tap
C).

5. The persistence of a single point of connection between the
caudal division of the ventral veno-lymphatic plexus and the
promontory on both sides of the embryo (Tap 6), this represent-
ing the primary promontorial connection of the para-precardinal
channel and its associated veno-lymphatic component of precar-
dinal tributary 4.

6. The presence of two closed sacs on the right side, one with-
out blood corpuscles, which have become detached from the para-
precardinal channel and its associated veno-lymphatic component
of precardinal tributary 4.

Series 77, 11”” Embryo
Figs. 51 and 52

1. Forward extension of the dorsal veno-lymphatic plexus
and the establishment on both sides of the embryo of an anterior
tap of evacuation.

2. Fusion of cephalic division of ventral veno-lymphatic
plexus with anterior end of dorsal plexus.

3. With the exception of the anterior tap of evacuation no
communication exists between the straight segment of the pre-
cardinal and the veno-lymphatic plexus.

4. Two points of communication between the general veno-
lymphatic plexus and the promontory are present which, in
the entire series of embryos examined, appear the last to be
given up, as well as the most constant in character; one is with the
dorsal veno-ly mphatie plexus through the subclavian approach (Tap
C), the other with the caudal division of the ventral plexus through
the jugular approach (Tap B). These two points of communica-

288 George 8. Huntington and Charles F. W. McClure.

tion between the promontory and the veno-lymphatic plexus
correspond, approximately, to the two points at which in later
stages the jugular lymph sac taps the venous system (see fig. 3 of
adult).

5. The embryo represents a stage just preceding the complete
fusion of the dorsal and ventral plexus to form a common sac.

6. The caudal portion of the ventral plexus has assumed a
characteristic form which appears to precede this fusion, its
dorsal, dilated oval portion being flexed mesad so that it les upon
the dorsal surface of the precardinal just in front of the promon-
tory, while its ventral, elongated segment follows the lateral sur-
face of the promontory and precardinal, preparatory to its fusion
with the anterior end of the dorsal plexus, which, in this case, also
includes the cephalic division of the ventral plexus.

7. Relation of spinal nerves to the veno-lymphatic plexus
to be compared with relation of these nerves to the jugular lymph
sacs of later stages and of the adult.

8. Constant relations of thyro-cervical artery to the sub-
clavian approach, to the veno-lymphatico-venous connection
of the dorsal plexus at Tap C, and to the interval between the
dorsal veno-lymphatic plexus and the caudal division of the ven-
tral plexus.

PRE-LYMPHATIC STAGE.

Sracr OF EvacuaTION OF BLOOD CONTENTS OF EARLIER VENO-LYMPH-
atic PLexus, AND APPARENT CoMPLETE TEMPORARY SEPARATION OF
tHE Empry LympHatic Sac FROM THE PERMANENT VENOUS BLoop
CHANNELS.

Series 78, 12” Embryo
Series 474, 10.7"™ Embryo, Harvard Embryological Collection

We have already stated previously that no hard and fast lines can
be drawn between the veno-lymphatic and definite lymphatic stages
as far as the development of the jugular lymph sac is conncerned.
It is convenient, however, to define certain derivatives of the
venous system, constituting the anlages of the jugular lymph sacs,
as ‘“‘veno-lymphaties,”’ as long as these derivations are still filled

—

Development of the Jugular Lymph Saes. 289

with blood and remain in free communication with the veins. Be-
tween this veno-lymphatic stage and the stage of final organiza-
tion of the lymph saes as definite portions of the entire lymphatic
system, intervenes a very short period which we have designated
as the pre-lymphatic stage. In this stage the venous derivatives
appear fused together to form a single sac which then undergoes
the following successive changes;

1 The blood contents of the sac are emptied in many cases
into the systemic vein through a large connection, secondarily
established, between the anterior end of the common sac and the
precardinal vein (Anterior tap of evacuation). Except for this
anterior connection between the systemic veins and the sac, the
latter is distinctly lymphatic in character, and, from a histological
standpoint, cannot be distinguished from the lymph sac of much
later stages. It is possible that in certain embryos the promon-
torial connections serve as posterior taps of evacuation, in which
ease the anterior tap is not established.

2 After the sac has become emptied of blood contents the an-
terior tap of evacuation closes rapidly, and the sac, according to
our observation, separates for a short time entirely from the ven-
ous system. <A few doubtful points of apparent communication
observed in some instances are very likely artifacts. In any case
they differ entirely in character and appearance both from the
earlier connections of the veno-lymphatic plexus with the veins,
and from the later typical lymphatico-venous taps found after
the adult conditions have been established.

Although in this pre-lymphatic stage no definite promontorial
connection between the sac and the veins can be determined,
the sac sends out two processes which approach and almost reach
the two points which in the adult constitute the area of lymphatico-
venous connections. These two points correspond, respectively,
to those at which in the earlier stages the dorsal and ventral veno-
lymphatic plexuses opened into the jugular promontory. (Com-
pare fig. 16 with figs. 14, 15 and 17). It is difficult to account on
physiological grounds for this apparent temporary separation of
the sac from the veins. Since the permanent connections of the
lymphatic and venous system take place apparently at the iden-

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 2.

290 George 8S. Huntington and Charles F. W. McClure.

tical points occupied in the earlier stages by the terminals of the
ventral and dorsal veno-lymphatic plexuses, it appears much more
reasonable to assume a continuity of the channels throughout
development. The picture afforded by the proper stages is,
however, so definite and conclusive, that it becomes necessary to
accept this separation as a normal feature of the pre-lymphatic
stage, and as differentiating it sharply both from the earlier veno-
lymphatic condition and from the later stage in which the definite
secondary lymphatico-venous taps are established.

Series 78, 12°" Embryo
Reconstruction of left side,
Lateral aspect, fig. 57

The left side of this embryo illustrates an extremely important
and evidently very evanescent stage in the transferal of the veno-
lymphatic sac to the definite lymphatic system as the jugular
lymph sac. It has already been referred to in connection with
the significance of the conditions shown by series 101 and 77 (figs.
50, 51 and 52) in which an anterior tap of evacuation was observed
for the first time.

The main features furnished by the left side of this embryo are
as follows:

1 The three primary divisions of the veno-lymphatic plexus of
earlier stages (figs. 12 and 49) have fused with one another to form
a common sac (fig. 57). As compared with series 77 (fig. 52), the
relations of the spinal nerves and thyro-cervical artery to the
resulting capacious sac elearly indicate the role played by the three
primary divisions of the veno-lymphatic plexus in forming the sac.

2 The primary promontorial connections of earlier stages,
which exist between the venous system and the plexus (Taps Bb
and C, figs. 51 and 52) at the jugulo-subclavian and ecmmon
jugular angles, have been completely lost as far as can be deter-
mined by the most careful observations. The jugular and sub-
clavian approaches, however, form prominent processes of the sac,
and terminate blindly near the point where they formerly com-
municated with the lateral surface of the promontory (fig. 51, 11

Development of the Jugular Lymph Saes. 291

mm. embryo). In other words, the jugular and subclavian ap-
proaches retain exactly the same relations to the promontory, the
cephalic vein and the thyro-cervical artery in the present case
as they do in the 11 mm. embryo (figs. 51 and 52) where they com-
municate with the systemic veins.

The significance of the jugular and subclavian approaches, as
well as their relation to the early veno-lymphatic plexus, should be
constantly kept in mind in reading the following pages, since it is
through them that the permanent connections between the lym-
phatic and venous systems are subsequently established.

3 Anteriorly, the veno-lymphatie sac opens by a broad ori-
fice into the straight segment of the precardinal near the level of
the cephalic arch, and is also connected with the same by asmaller
channel slightly caudal to this point. This wide-open communica-
tion between the veno-lymphatic sac and the precardinal vein consti-
tutes the anterior tap of evacuation which on the left side of this
embryo is much more extensive in character than in the 10 and 11
mm. embryos (figs. 50, 51 and 52. This extensive entrance into
the vein, which now serves as an outlet for the blood contents of
the sac, is connected with the area of the precardinal which for-
merly received the dorsal tributaries 1, 2 and 3, and its mode of
origin in this case was probably the same as that of a similarly
situated element which was described in connection with series
19 (9 mm., fig. 42).

The genesis of this anterior tap of evacuation can be further
traced from the preceding 10 and 11 mm. stages, by comparing
figs. 47, 48, 49, 50, 51 and 52 with fig. 57. <A comparison of
figs. 47, 48 and 49 with fig. 57 will show that the area of the tap
of evacuation in the latter corresponds to the area of the veno-
lymphatic derivatives of tributaries 1, 2 and 3, as seen in the for-
mer, before confluence of the individual elements to form the con-
plete sac has occured.

In fig 50 (series 101, 10 mm.,right side) the anterior tap of
evacuation is small but distinct,the dorsal plexus arching to its
precardinal junction over the combined trunks of the first and
second spinal nerves. Cephalad of this point the junction of the
arched and straight segment of the precardinal carries a dilated

292 George 8. Huntington and Charles F. W. McClure.

and fenestrated appendage (fig. 50, b) which is also seen in series
77 (fig. 51,b). From the position of this element b,it seems prob-
able that its junction with the small anterior tap of evacuation
will establish the wide portal of entry, which in series 78 (fig 57)
forms the main anterior connection of the sac with the veins. In
this case it appears likely that the small caudal connection in ser-
ies 78 (fig. 57) represents the original anterior tap as seen on the
right side of series 101 (fig. 50), and on both sides of series 77 (figs.
50, 51, and 52). Finally, on the left side of series 101 (fig. 49), a
stage is shown, just prior to the establishment of the anterior tap
of evacuation, in which the dorsal veno-lymphatic arch has not
yet: united with the anterior precardinal veno-lymphatic elements
to form the anterior tap of evacuation.

4. The blood contents of the earlier veno-lymphatic plexus
have for the most part, been completely evacuated on the left
side of this 12 mm. embryo (fig. 57), evidently through the large
anterior venous connection just described. Here and there in the
caudal portion of the sae small outlying pockets still contain a
few blood cells, but as a whole the former veno-lymphatic plexus
represents on this side an enormously expanded empty sac, de-
detached at all points from the venous system except at the very
large and the small anterior channels described. A comparison of

the left (fig. 57) with the right side of this embryo (not figured) is

most instructive. On the right side the sac is completely closed, con-
tains no blood corpuscles and, as far as we can determine, does not
communicate with the veins at a single point. Since the promontor-
ial connections of the sac in both of the embryos which immediate-
ly precede this stage appear to be too insignificant to permit of
the passage of a large amount of blood, we can only infer that on
the right side, as on the left, an anterior tap of evacuation was at
one time present, and that it served as the main exit for the blood
contents of the veno-lymphatic sac. Figs. 53 and 54 (series 78,
slide 3, sections 34 and 40) show the appearance of the anterior tap
of evacuation and of the connected portion of the sac in transverse
section on the left side, and, on the right side, the complete sep-
aration of the jugular sac from the veins. Figs. 55 and 56 show the
caudal portion of the lymph sac (slide 4, sections 33 and 35).

Development of the Jugular Lymph Sacs. 293

The dorsal and ventral portions of the sac are clearly differen-
tiated on the left side in Fig. 55. The multilocular character of
the empty sac and the incomplete septa and partitions in the
interior are well shown in both figures.

Secondary features of significance illustrated by the left side
of this embryo are as follows:

1. The thyro-cervical artery on reaching the level of the pro-
montory bends laterad on to the lateral surface of the promontory
in the interval which separates the jugular from the subclavian
approach. On reaching the lateral surface of the promontory the
thyro-cervical artery gives off a branch which runs dorsally along
the lateral surface of the veno-lymphatic sac. This latter branch
is accompanied by a branch of the cephalic vein and in later
stages both this vein and the artery which accompanies it may
become embedded in the jugular lymph sae.

2. The primitive ulnar vein has given up its anterior connec-
tion with the promontory and the drainage of the anterior limb
is now assumed by the definite subclavian vein. The subclavian
vein opens into the promontory slightly caudal to the point where
the latter formerly received the primitive ulnar vein. (Compare
fig. 51, in which the subclavian connection is established). Before
the primitive ulnar vein gives up its anterior connection with the
promontory, the sixth spinal nerve (SP.N.VJ) arches under or
ventral to the primitive ulnar vein and primitive ulnar veno-
lymphatic (figs. 45,47 and 48). With the loss of its promontorial
connection, however, this relation is alone maintained to the
primitive ulnar veno-lymphatie (fig. 57). Cf. figs, 13, 14, 15 and 16.

3. The primitive ulnar veno-lymphatic forms a well-defined
caudal prolongation of the veno-lymphatic sac and can be traced
to the base of the anterior limb.

4. ‘The marked convexity of the cephalic arch has been reduced
with the descent of the heart and the resulting elongation of the
straight segment of the precardinal. The promontory no longer
forms a prominent swelling at the cardinal-Cuvierian junction,
and has been reduced by the caudal descent of the heart.

5. The relations of the spinal nerves to the veno-lymphatic
sac have already been referred to and need no further explanation,

294 George S. Huntington and Charles F. W. McClure.

6. A new element has been added to the veno-lymphatie sac
which Lewis has described in connection with the lymph sae of
the rabbit under the name of the subcutaneous duct. It consists
of an appendage of the dorsal arch of the veno-lymphatie sac,
which is connected with the lateral surface of the same at two
points. This appendage increases in size from now on, and, as it
is constant in its appearance, is worthy of consideration. As we
have not followed its origin closely, however, we are unable to
speak authoritatively concerning it.

Series 474, Harvard Embryological Collection, slides C, D, EL, and F.
10.7" Embryo
Reconstruction of left side,
Lateral aspect fig. 58 and
Reconstruction of right side,
Lateral aspect, fig. 59

We owe the opportunity of studying and reconstructing this
very important and interesting embryo to the courtesy of Prof. C.
S. Minot of Harvard University. The length of the embryois given
as 10.7 mm., but in the development of its jugular lymph sac, it
corresponds to specimens of our collections, which measure con-
siderably more-(from 11 to 12.5 mm.). This discrepancy may,
however, be due to different technique in taking the total length
measure, as to unusual curvature of the embryo.

Serially, from the developmental aspect, it follows the stage
previously described (series 78, 12 mm.), and is an excellent
example of the apparent complete bilateral temporary separation
of the voided veno-lymphatic sac, now the closed and empty jugu-
lar lymph sac, from the permanent veins. The extremely doubt-
ful connection of the sae with the venous system found by us
near the jugulo-subclavian angle is cqnfined to a single section on
the right side (slide E, section 194), and to two sections on the
left side (slide EK, sections 197-198). Even if this connection
exists, it differs entirely from both the earlier veno-lymphatic and
the later definite lymphatico-venous taps. As previously stated
(p. 194), if the empty sae separates temporarily from the ven-
ous system, and subsequently rejoins the same by establishing

Development of the Jugular Lymph Saes. 295
secondary connections, it is to be expected that certain embryos
will be fixed at periods in which, on the one hand, the temporary
detachment has been nearly, but not quite completely attained,
and, on the other, in which the secondary connections are just be-
ginning to be established. In either case the resulting histolog-
ical picture will yield these so-called ‘‘doubtful’’ connections.
The evidence that the jugular lymph sac becomes temporarily
separated from the veins is based upon the following observations:

1. We have been unable to find any connection between sac
and vein on the right side of embryo 78 (12 mm.).

2. There is no connection on the left side of this embryo
series 78 at either of the typical points of final lymphatico-venous
entry, viz., the common jugular confluence and the jugulo-sub-
clavian angle.

3. No connection between sac and veins exists at any point
in all mm. embryo (series 27), in which the jugular lymph sac
is fully established.

4. No connection is found in the 13 mm. embryos.

5. Absence of the typical communications in the Harvard
series 474 (10.7 mm), although the minute doubtful connections
above described may be present.

The definite and permanent connections of the jugular lymph
sac and the veins do not appear prior to the stage between 14 and
15 mm., from which stage on they are usually constant.

Reconstruction of left side, lateral aspect (slides C, D, E, and F) fig. 58.

The jugular lymph sae contains no blood and is composed of
clearly defined confluent elements, (fig. 58), as follows:

1. The part derived from the cephalic division of the ventral
veno-lymphatic plexus (J).

2. The part derived from the caudal division of the ventral
veno-lymphatie plexus (//) terminating in a blind prolongation
(jugular approach), which is directed toward, and nearly reaches,
the angle of confluence of the internal jugular and the combined
trunk of the external jugular and cephalic veins (common jugular
angle).

296 George 8. Huntington and Charles F. W. McClure.

3. The entire dorsal arch is derived from the dorsal veno-
lymphatic plexus and the veno-lymphatic components of the
precardinal tributaries, and terminates caudally in a pointed
extremity (subclavian approach), which is directed toward the
jugulo-subclavian angle, near which it forms a doubtful connection
with the venous system (slide E, sections 197-198).

4. The primitive ulnar lymphatic, derived from the primi-
tive ulnar veno-lymphatic. This structure now contains no blood
cells, is distinctly lymphatic in structure, and forms a blind ap-
pendage to the caudal end of the jugular sac, arching dorso-
ventrad over the brachial plexus, the sixth spinal nerve of which
is represented in the figure SP. N. VJ.

These components of the jugular lymph sac can readily be
homologized with their equivalents in series 102, (fig. 46), series
101 (fig. 49), series 77(fig. 51) and series 78 (fig. 57).

The relations of the spinal nerves to the jugular lymph sac in
the 10.7 mm. embryo remain fundamentally the same as in the
later veno-lymphatice and pre-lymphatic stages and may be com-
pared with those in figs. 49, 50, 52 and 57.

The external jugular terminal which opens singly into the pro-
montory in series 102, (fig. 46), has united with the base of the ceph-
lic vein, forming the common jugulo-cephalic trunk so character-
istic of later stages.

In this, as well as in all subsequent stages, the promontory is no
longer recognizable as an expanded and dilated portion of the
main venous channel, but its former position is easily determined
by the relations of the thyro-cervical artery and by the presence of
a series of dorsal somatic tributaries which open into the jugular
vein near the caudal end of the jugular lymph sac. These tribu-
taries have been fully described in connection with the preceding
stages (see 4S, 5S, 6S, 7S and 88, fig. 45).

Reconstruction of Right side, Lateral A spects (slides C, D, andE), Fig. 59.

The right side of this embryo presents identically the same con-
ditions as the left, including the presence of a doubtful lymphatico-
venous connection near the jugulo-subelavian angle (slide #
section 194), with the exception that the cephalic and caudal divis-

Development of the Jugular Lymph Sacs. 297

ions of the ventral arch of the lymph sac have not united to form
the ventral boundary of the large foramen through which the first
and second (SP.N.J and JI) and third and fourth (SP.N.III
and JV) spinal nerves pass on the left side (fig. 58). The fifth
spinal nerve (SP. N.V), as on the left side, and as in all of the pre-
ceding stages, penetrates the lymph sac.

LYMPHATIC STAGE OR STAGE OF THE DEFINITELY ORGANIZED
JUGULAR LYMPH SAC.

Series 87, 14" Embryo
Series 15, 16"”" Embryo
Series 86, 17°" Embryo
Series 88, 18" Embryo
Series 22, 25"”" Embryo

As mentioned above the jugular lymph sac consists of a single
sac, containing no blood or only isolated corpuscles, which, after
its apparent temporary separation from the veins in the preceding
pre-lymphatic stage,now secondarily rejoins the venous system
through its jugular and subclavian approaches at the common
jugular confluence, or at the jugulo-subclavian angle, or at both
points, depending upon the definite type of adult lymphatico-
venous connection to be established in the individual instances.
(See p. 188). With the assumption of this definite lymphatic
stage the organization of the jugular sac is completed. The further
development involves merely some topographical changes in rela-
tion to the spinal nerves and the establishment of secondary con-
nections with the independently formed systemic lymphatics.

Series 87, 14°" Embryo
Reconstruction of left side,
Dorsal aspect, fig. 60 and
Ventral aspect, fig. 61

Only the caudal end of the lymph sac is shown in this recon-
struction.

The jugular lymph sac is much enlarged in this embryo, and the
subcutaneous duct is still a prominent tributary of the sac.

298 George 8. Huntington and Charles F. W. McClure.

The internal jugular vein has relatively decreased in size, while
the external jugular and cephalic veins have correspondingly
enlarged. The two divisions of the lymph sac (dorsal and ventral)
have become closely amalgamated in the caudal portion and the
united sac is still perforated some distance from its dorsal margin
by the fifth cervical nerve (S.P.N.V.). The sac terminates
-audally in four prolongations:

1. The jugular approach (fig. 61) terminates in a pointed
extremity, which enters, as previously described, in a wedged-
shaped invagination, with valve formation, the dorsal aspect of
the angle of confluence of the internal jugular (precardinal) and
the combined trunk of the external jugular and cephalic veins.
This forms the typical secondary and final lymphatico-venous
tap at the common jugular confluence. (Compare with figs. 1,
2 and 3 of adult).

2. The subclavian approach (fig. 61), is a much elongated pro-
cess of the ventro-caudal end of the jugular sac, which extends
caudad, ventral to and somewhat beyond the jugulo-subclavian
junction, where it ends in a pointed blind extremity. A doubt-
ful point of connection exists with the vein near the jugulo-
subelavian junction. The subclavian approach is potentially
in position to establish a typical secondary entrance into the
dorsal aspect of the jugulo-subelavian angle. We have, therefore,
probably here an instance in which, while the common jugular
secondary lymphatico-venous tap is already established, the sub-
clavian connection has not yet been made. It is, of course, on the
other hand, possible that we are dealing with an individual which
in the adult state would. have presented only a single (common
jugular) lymphatico-venous connection, and in which the secon-
dary jugulo-subeclavian tap was never formed. (Fig. 1, left side).

3. The third prolongation is formed by the primitive ulnar
lymphatic as a blind process of the sac, arching caudad over the
sixth spinal nerve (fig. 61, S.P.N.VJ.), along the dorsal aspect
of the subclavian vein.

4. The point at which in later stages the proximal end of the
thoracic duct will connect with the jugular sac, is represented
(fig. 60), by a short process which lies upon the dorsal surface

Development of the Jugular Lymph Sacs. 299

of the internal jugular vein, between the sympathetic nerve
and the thyro-cervical artery. It extends caudad as far as
the base of the first of the (promontorial) dorsal somatic tributar-
ies (4S in figs. 34, 45 and 60), where it ends blindly. The connec-
tion of this process with the jugular sac lies dorsal to that of the
jugular approach, the position of the latter, on the ventral surface
of the vein, being represented by a plus sign (+) in fig. 60.

The thyro-cervical artery, a branch of the subclavian, runs
cephalad along the dorsal-lateral surface of the jugular vein and
on reaching the lymph sac turns abruptly laterad, ventral to the
subclavian approach, and then divides into three branches one of
which runs dorsad along the lateral surface of the jugular sac.
The relations of the thyro-cervical artery to the jugular sac are
represented in fig. 60. The branch which follows the lateral sur-
face of the lymph sac and its accompanying vein are not repre-
sented, however. It will be observed that the relations of the
thyro-cervical artery to the jugular lymph sac are exactly the
same as in all of the preceding stages (figs. 49, 51, 52 and 57).

The external jugular vein is a vein of large size in this embryo
(fig. 61), having practically assumed its adult condition. It usually
functions as the main drainage canal of the head and neck in
the adult although cases are sometimes met with in which the
external and internal (precardinal) jugulars are subequal in
size.

Series 15, 16” Embryo
Reconstruction of left side,
Dorso-lateral aspect, fig. 62

The jugular lymph sac now appears relatively reduced and
shortened in its cephalo-caudal dimension. It endsblindly in front
in a bluntly pointed extremity, and receives the subcutaneous
duct on its lateral surface.

A number of features characterize this embryo, which show
an advance in lymphatic development over that of the preceding
stage (14 mm.).

1. The jugular approach, not shown in the dorso-lateral view,
communicates with the venous system at the angle of confluence
of the internal jugular and the combined trunk of the external

300 George 8S. Huntington and Charles F. W. McClure.

jugular and cephalic veins (common jugular tap). This lymphati-
co-venous connection in no way resembles those of the early veno-
lymphatic, nor the doubtful ones in the lymphatic stages, but is
established by a wedge shaped process of the lymph sae which is
deeply invaginated into the angle of confluence of two veins and
opens into the vascular lumen by a narrow slit-like aperture
bounded by a two-lipped valve. This type of valve, also char-
acteristic of the adult, is illustrated by figs. 4, 5, 6 and 7, which are
photomicrographs of transverse sections through the region of the
valve in the embryo under consideration.

2. The subclavian approach also communicates with the ven-
ous system at the jugulo—subelavian junction by a narrow slit-
like aperture (jugulo-subclavian tap).

In view of the conditions observed in the preceding stages we
are bound to infer that these two taps have been secondarily
formed.

By comparing fig. 62 with fig. 3, it will be seen that the adult
condition has been reached.

3. The separate anlages of the thoracic duct, formed inde-
pendently of the jugular lymph sac, have now united with each
other, and with the dorsal process of the lymph sac above described.
The thoracic duct, therefore, for the first time,now forms a con-
tinuous vessel between the posterior thoracic region and the
jugular lymph sac into which it opens dorsal to, and slightly in
front of the tap at the common jugular confluence. In the por-
tion of the embryo figured the thoracic duct crosses the dorsal
surface of the subclavian artery, passes ventral to the sympa-
thetic nerve, and from this point on to its entrance into the jugu-
lar sac it lies between the sympathetic nerve and the thyro-cervical
artery. Along this course it follows the dorsal surface of the left
innominate and common jugular veins.

4. The primitive ulnar lymphatic has lost its connection
in this embryo with the jugular sac, as may be seen by the rela-
tions of the sixth spinal nerve (SP.N.VIJ, fig. 62). It may be
stated in general that the primitive ulnar lymphatic retains its
connection with the jugular sac through the 14 mm. stage, after
which this connection appears to be lost. This change, therefore,

—_

Development of the Jugular Lymph Sacs. 301

approximately coincides with, or follows very shortly upon, the
establishment of the subclavian vein as the chief channel of venous
return from the anterior limb and the resulting abolishment of the
primitive ulnar vein.

The further fate of the primitive ulnar lymphatic is involved
in the establishment of the definite systemic lymphatic channels of
the anterior extremity, and is hence not considered at this time.

5. The relations of the thyro-cervical artery to the subclavian
approach and to the common trunk formed by the external jugu-
lar and cephalic veins, is well illustrated by the series of photomic-
rographs taken through this region, and should be compared with
the reconstruction (fig. 62). Beginning caudad, figs. 7, 6, and 5
represent the thyro-cervical artery situated dorsal to the com-
mon trunk of the external jugular and cephalic veins and medial
to the subclavian approach. Fig. 4, on the other hand, represents
a transverse section taken through the thyro-cervical artery as it
bends laterad, ventral to the subclavian approach and dorsal to
the jugulo-cephalic trunk. This last section lies shghtly caudal
to the point where the thyro-cervical artery divides into three
branches; one ventrally directed branch; one which follows the
cephalic vein, and another which runs dorsally along the lateral
surface of the jugular sac and is accompanied by a branch of the
cephalic vein (fig. 62). The latter branch as well as the vein, as
mentioned above, are at times embedded in the lymph sac (see fig.
64, series 88).

6. The first, second, third and fourth (fig. 62, SP.N. DIG EE
and JV) spinal nerves no longer pass through a foramen in the
lymph sac as in fig. 57, but now pass between the sac and the in-
ternal jugular (precardinal) vein. The fifth (SP.N .V) spinal
nerve still penetrates the jugular sac while the sixth (SP.N. VI)
lies upon the lateral surface of the subclavian approach, and, as
stated above, is now in no way related to the primitive ulnar
lymphatic. .

The present relations of the first four spinal nerves to the jugu-
lar sac are explained by comparing fig. 17 with the preceding
fig. 16. In course of further development, as illustrated in the
present instance, the large lymphatic foramen through which

302. George S. Huntington and Charles F. W. McClure.

the nerves pass, is broken by reduction of its anterior extrem-
ity. The lymph sae now is continued cephalad into two
processes, a smaller ventral and larger dorsal. The former, which
accompanies the internal jugular vein, becomes further reduced
and recedes caudad, thus freeing the nerves and allowing them to
pass between the internal jugular vein and the larger dorsal pro-
cess which remains as the main component of the lymph sae in
this stage.

Series 36, 17°" Embryo
Reconstruction of left side,

Dorso-lateral aspect, fig. 63

The general composition of the caudal end of the jugular lymph
sac, which is alone represented in the figure, corresponds quite
closely to the 16 mm. stage.

A lymphatico-venous connection, as in the 16 mm. embryo, has
been established on the dorsal aspect of the common jugular con-
fluence (common jugular tap) by the jugular approach. This tap
cannot be seen from the dorsal view of the reconstruction but its
relative position is indicated by a plus sign (+) in the figure.

The jugular sac is prolonged caudad into a subclavian approach
which now bifureates near the jugulo-subclavian junction into a
dorsal and ventral division. The ventral division constitutes the
caudal extension of the original subclavian approach of the pre-
ceding stages and in the present embryo communicates ventrally
with the veins at the jugulo-subclavian junction (jugulo-subclay-
ian tap). The dorsal division, not present in the 16 mm. embryo,
extends caudad for a short distance on the dorsal aspect of the
jugulo-subclavian angle where it ends blindly. It is now in a
position, however, to establish the secondary dorsal lymphatico-
venous connection at this point which is frequently met with in
the adult.

The thoracic duct is represented by a more complex system of
vessels in this embryo than in the 16 mm. stage. A number of
vessels from the thoracic region converge to form a single trunk
which unites with the jugular sac dorsal to and slightly in front
of the common jugular tap. The relations of the thoracic duct

Development of the Jugular Lymph Saes. 303

to the subclavian, thyro-cervical arteries and to the sympathetic
nerve remain the same as described above for the 16 mm. embryo.

Two small lymphatic vessels lying along the internal jugular
vein open into the medial aspect of the jugular sae just in front of
the entrance into the latter of the thoracic duct. These vessels
represent the approach of the sac to union with the lymphatic
vessels which have begun to form along the course of the internal
jugular vein.

The fifth spinal nerve (SP.N.V.) still penetrates the jugular
sae and the relations of the thyro-cervical artery and its branches
to the jugular lymph sac and systemic veins are the same as in
the 16 mm. embryo (fig. 62).

Series 88, 18" Embryo
Reconstruction of the jugular lymph sac of the right and left sides
Ventral View, fig. 64 and
Lateral view of the left side, fig. 66

This reconstruction is especially valuable in showing the sym-
metrical degree of development obtained by the jugular lymph
sac on opposite sides of the same embryo, as well as the fact that
up to acertain stage of development both sacs are ontogenetically
of equal importance (fig. 64).

In the 18 mm. embryo the arteries end main systemic veins
have practically reached the adult concition. The external and
internal jugulars are more elongated than in the preceding stage
as the result of the continued descent of the heart. The external
jugular is a vein of large size, although its caliber does not yet exceed
that of the internal jugular. The cephalic vein is also of large
size and,after joining the external jugular to form the common
jugulo-cephalic trunk, the latter, which has become somewhat
elongated, joins the internal jugular at the common jugular angle.
The resulting common trunk formed by the confluence of the
internal. jugular and jugulo-cephalic trunk, as far caudad as its
confluence with the subclavian, now constitutes the common
jugular vein (see fig. 1 of adult).

Beginning in the 17 mm. embryo, slightly caudal to the level
of the subclavian vein and dorsal to the thymus glands, the two

304. George S. Huntington and Charles F. W. McClure.

precaval veins (precardinal) anastomose with each other across
the median line and thereby establish the anlage of the definite
right and left innominate (brachio-cephalic) veins. In the 18 mm.
embryo this anastomosus is completed and the blood from the
left jugular and left subclavian districts is now, for the most part,
directed into the right precava. As the result of this transfer the
right precava in the 18 mm. embryo has become much enlarged and
the left correspondingly reduced.

In addition to the innominate anastomosis, mentioned above,
another anastomosis occurs in this region ventral to the thymus
gland. This makes its appearance in the 15 and 16 mm. embryos,
and before the definite innominate anastomosis has been estab-
lished; but with an increase in size of the latter, this anas-
tomosis soon disappears. In the 18 mm. embryo, however,
the thymus is still crossed ventrally by a net-work of vessels
which represents the remains of this sub-thymic anastomosis.

The transferal of the blood current from the left jugular and
left subclavian veins to the right precava is illustrated by figs. 13,
14, 15, 16 and 17 to which the reader is referred. An examination
of these figures will make plain the conditions of the venous sys-
tem in this (figs. 64 and 65), and in the embryo next to be con-
sidered (25 mm., fig. 66).

Lymphatic development has reached an advanced stage in the
18mm. embryo. The capacious jugular sac of each side has estab-
lished in this instance the typical taps at the common jugular and
jugulo-subclavian angles, respectively. (Cf. fig. 3 of adult). The
left jugular sac also receives the large thoracic duct on its dorsal
aspect. The lateral view of the left side of the reconstruction
(fig. 65), shows this connection.

The jugular sacs present further evidence of the reduction of
the ventral portion, the main sac being formed by the dorsal
part of the earlier lymphatic ring surrounding the first four spinal
nerves. Asa result of this change, with the exception of the fifth
nerve (fig. 64,SP.N.V), which still penetrates through the sac, the
remaining four anterior nerves pass between the sac and the internal
jugular vein and the lymphatics which accompany that vessel.

The branch of the thyro-cervical artery which extends dorsad

Development of the Jugular Lymph Saces. 305

along the lateral surface of the jugular sac in earlier stages (fig. 62)
is embedded in the lymph sac in this 18 mm. embryo (fig. 64)
with an accompanying branch of the cephalic vein.

Lymphatic formation, independent of the jugular lymph sae,
is in full swing,and some of these independent lymphatics have
made secondary connections with the jugular sacs. Closed lym-
phatic channels or sacs stud the course of the jugular, cephalic
and subclavian veins, and an especially prominent lymphatic
follows the course of the diminishing left precava (fig. 65).

Series 22, 25°" Embryo
Reconstruction of left side,
Ventral aspect, fig. 66

Only the caudal end of the lymph sac and its related veins are
shown in the figure.

As far as the jugular lymph sac is concerned, the final approach
to the adult condition is achieved in the 25 mm. embryo. There
is a further reduction in the size of the internal and a correspond-
ing enlargement of the external jugular vein. The common vessel
(jugulo-cephalic trunk) formed by the confluence of the external
jugular and cephalic veins has, with further cardiac descent, be-
come much elongated and the common jugular angle lies rela-
tively further caudad than in the 18 mm. embryo. This descent
has drawn out the lymph sac of the preceding stage into an elon-
gated channel which lies along the veins and taps them in this
case also at the two typical points.

The dorsal and ventral divisions of the subclavian approach,
described above in connection with the 17 mm. embryo (fig. 63),
now anastomose with each other just caudal to the jugulo-sub-
clavian confluence, where they become continuous with a lymph
channel which follows the left precava. This latter lymphatic
was described in connection with the 18 mm. embryo (left pre-
vacal lymphatic, fig. 65), where a connection between it and the
jugular sac does not yet exist.

Numerous independent lymphatic structures are found along
the course of the large veins. Among these may be mentioned a
large complex lymphatic sac which envelops the subclavian vein,

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, No. 2.

306 George 8. Huntington and Charles Ff. W. McClure.

and which does not yet join the jugular sac, as far as we can de-
termine. This lymphatic complex increases in its dimensions
from before backward.

An elongated and fenestrated lymphatic vessel, not yet joined
to the subclavian approach, lies ventral to the left precava and
the anlage of the left innominate vein, while another isolated lym-
phatic is found at the jugulo-cephalic junction. None of these
lymphatic structures communicate with the veins.

The lymphatic organization of this embryo foreshadows the
conditions in the adult.

The innominate cross anastomosis, the establishment of the

right and left innominate veins and the reduced size of the left
precava, are well shown in the 25 mm. embryo (fig 66).

Development of the Jugular Lymph Sacs. 307

IV. GENERAL CONSIDERATIONS AND CONCLUSIONS

In the preceding pages the writers have presented a detailed
account of the development of the jugular lymph sacs in the
domestic cat (Felis domestica).

The primary principles underlying the development of the jugu-
lar lymph sac are: (1) the development of a secondary channel
parallel to the embryonic precardinal and the Cuvierian end of
the postcardinal; (2) the association with this secondary channel
of a certain number of dorsal precardinal tributaries, and (3) the
separation of these two sets of venous elements, which we have
termed ‘Veno-lymphatics’, from the main venous channels, and
their subsequent conversion into the definite jugular lymph sacs
by a process of growth and fusion.

For a complete and short résumé of the development of the
jugular lymph sac, illustrated by diagrams (figs. 8 to 21), the
reader is referred to the topic in the preceding pages, entitled
‘Analysis of Developmental Stages in the Formation of the
Jugular Lymph Saes’ (page 202). <A short résumé, although
less complete than the one referred to, was published in the Ana-
tomical Record, Vol. II, 1908, to which the reader is also referred.

The writers wish to make it perfectly clear that the present
communication deals solely with the anatomy and development of
the JuGuLAR LympH Sacs of the cat, and does not touch on the
question of the origin of the independently formed systemic lym-
phatic vessels which unite with the jugular sacs.

Accepted by the Wistar Institute of Anatomy and Biology, December 2Ist, 1909. Printed May
25th, 1910.

308 George 8. Huntington and Charles F. W. McClure.

We take pleasure in expressing our thanks and gratitude to our
friend Mr. M. Petersen for his untiring efforts in our behalf, as well
as our appreciation of the manner in which he has reproduced our
complicated reconstructions for publication.

EXPLANATION OF FIGURES

Figs. 1, 2, and 3. Dissections of the lymphatics in the neck region of the adult
cat (Felis domestica), showing the three normal forms of lymphatico-venous com-
munication which may occur on each side of the body, namely, at the common
jugular angle, at the jugulo-subclavian angle or at both of these points.

Figs. 4, 5,6 and 7. Photomicrographs of transverse sections of a 16 mm. cat
embryo (series 15), sections 233, 237, 239 and 241, respectively, taken at the level
of the junction of the left Jjugulo-subclavian trunk and left internal jugular vein
(common jugular confluence) illustrating the ty pe of lymphatico-venous connection
met with in advanced stages at the two typical points of lymphatico-venous entry.

Figs. 8 to 11. Composite diagrams illustrating the development of the jugular
lymph sacs in the cat. Lateral views of left side.

Fig. 11 A. Dorsal view of fig. 11 in the region of the jugular promontory.

Figs. 12 to 17. Composite diagrams, continued, illustrating the development
of the jugular lymph sacs in the cat. Lateral views of left side.

Fig. 18. Composite diagram (dorsal view) of the precardinal vein and promon-
tory of the left side, illustrating the separation of the precardinal tributaries
1, 2, 3 and 4 into their dorso-lateral veno-lymphatice and dorso-medial somatic
components; also the relation of the dorsal veno-lymphatic plexus to the promon-
tory and to the promontorial somatic tributaries, 5S, 68, etc.

Fig. 19. Diagram of precardinal vein and promontory (dorsal view), illustrat-
ing a case in which the para-precardinal channel is alone involved in the formation
of the caudal division of the ventral veno-lymphatic plexus.

Fig. 20. Diagram of left precardinal vein and promontory (dorsal view),
illustrating the role played by the veno-lymphatic component of the precardinal
tributary 4 (4VL) in forming the rounded appendage of the caudal division of the
ventral veno-lymphatic plexus, sometimes met within the later stages.

Fig. 21. Diagram of left precardinal vein and promontory (dorsal view) illus-
trating a case in which the para-precardinal channel has given up its connection
with the jugular promontory, leaving thereon a small conical teat- like process
near the future common jugular junction.

Development of the Jugular Lymph Sacs. 309

ALL OF THE RECONSTRUCTIONS MENTIONED BELOW INCLUDE THE REGION OF
THE PRECARDINAL VEIN AND ITS CONFLUENCE WITH THE POSTCARDINAL VEIN AND
THE DUCT OF CUVIER.”!

Fig. 22. Reconstruction of a5 + mm. cat embryo (series 30), left side, lateral
aspect.

Fig. 23. Reconstruction of a5 + mm. cat embryo (series 30), right side,
lateral aspect.

Fig. 24. Reconstruction of a5 + mm. cat embryo (series 31), left side, lateral
aspect. The postcardinal vein was not reconstructed in this embryo.

Fig. 25. Reconstruction of a 5mm. cat embryo (series 134), left side, lateral
aspect.

Fig. 26. Reconstruction of a5mm. cat embryo (series 47), right side, lateral
aspect.

Fig. 27. Reconstruction of a 6.2 mm. cat embryo (series 109), left side, lateral
aspect.

Fig. 28. Reconstruction of 6.2 mm. cat embryo (series 109), left side, medial
aspect.

Fig. 29. Reconstruction of a 6.2 mm. cat embryo (series 109), right side,
lateral aspect.

Fig. 30. Reconstruction of 6.2 mm. embryo (series 109), right side, medial
aspect.

Fig. 31. Reconstruction of a 7 mm. cat embryo (series 2), right side, lateral
aspect.

Fig. 32. Reconstruction of a 7 mm. cat embryo (series 2), left side, lateral
aspect.

Fig. 33. Reconstruction of a 7 mm. cat embryo (series 138), left side, lateral
aspect.

Fig. 34. Reconstruction of a7 mm. embryo(series 138),left side, medial aspect.

Fig. 35. Reconstruction of a7 mm. cat embryo (series 138), right side, lateral
aspect.
Fig. 36. Reconstruction of 7mm. embryo(series 138),right side, medial aspect.

Fig. 37. Diagram (dorsal view) of fig. 35.

Fig. 38. Reconstruction of 27.25 mm. cat embryo (series 13), right side, lateral
aspect.

Fig. 39. Reconstruction of a 7.25 mm. cat embryo (series 13), left side, lateral
aspect.

Fig. 40. Reconstruction of a 9 mm. cat embryo (series 106), left side, lateral
aspect.

Fig. 41. Reconstruction of a 9 mm. cat embryo (series 19), right side, lateral
aspect.

*1 In a few cases in which the first four spinal nerves (SP.N.J-IV) were omitted
in the reconstructions they have been added to the drawings of the same in order
to show the relations which they bear to the developing jugular lymph sac. In
all other respects the drawings of the reconstructions are accurate reproductions
of the models which they represent.

310 George 8. Huntington and Charles F. W. McClure.

Fig. 42. Reconstruction of a 9 mm. cat embryo (series 19), left side. lateral
aspect.

Fig. 48. Photomicrograph of a transverse section of a 10 mm. cat embryo
(series 114, slide 4, section 27), taken at a level just anterior to the jugular prom-
ontory and showing the relative position of the dorso-lateral veno-lymphatic
and dorso-medial somatic elements with respect to each other and to the precar-
dinal vein.

Fig. 44. Reconstruction of an 8.5 mm. cat embryo (series 102), left side,
lateral aspect. ;

Fig. 45. Reconstruction of an8.5mm. cat embryo (series 102), left side, medial
aspect.

Fig. 46. Reconstruction of an 8.5 mm. cat embryo (series 102), right side,
lateral aspect.

Fig. 47. Reconstruction of a 10 mm. cat embryo (series 112), left side, lateral
aspect.

Fig. 48. Reconstruction of a 10 mm. cat embryo (series 113), left side, lateral
aspect.

Fig. 49. Reconstruction of a10inm. cat embryo (series 101), left side, lateral
aspect. (The dotted lines represent the actual cephalic and caudal limits of the
area reconstructed).

Fig. 50. Reconstruction of a 10 mm. cat embryo (series 101), right side, lateral
aspect.

Fig. 51. Reconstruction of a 11 mm. cat embryo (series 77), left side, lateral
aspect.

Fig. 52. Reconstruction of a 11 mm. cat embryo (series 77), right side, lateral
aspect. ;

Figs. 53 and 54. Photomicrographsof transversesections of al2mm. eat embyro
(series 78) passing through the anterior tap of evacuation on the left side and show-
ing the absence of this tap on the right side of the embryo (sections 34 and 40,
respectively, of slide 3). :

Figs. 55 and 56. Photomicrographs of transverse sections of a 12 mm. cat
embryo (series 78, slide 4, sections 33 and 35), passing through the caudal end of
the jugular lymph sac, anterior to the jugular promontory. The multilocular
character of the jugular sac is clearly shown in fig. 56.

Fig. 57. Reconstruction of a 12 mm. cat embryo (series 78), lateral aspect,
left side.

Fig. 58. Reconstruction of a 10.7 mm. cat embryo (Harvard Embryological
Collection, series 474), left side, lateral aspect.

Fig. 59. Reconstruction of a 10.7 mm. cat embryo (Harvard Embryological
Collection, series 474), right side, lateral aspect.

Fig. 60. Reconstruction of a 14 mm. cat embryo (series 37), left side, dorsal
aspect. :

Fig. 61. Reconstruction of a 14 mm. cat embryo (series 37), left side, ventral
aspect.

Fig. 62. Reconstruction of a 16 mm. cat embryo (series 15), left side, dorso-
lateral aspect.

Fig. 63. Reconstruction of a 17 mm. cat embryo (series 36), left side, dorso-
lateral aspect.

ad te

Development of the Jugular Lymph Sacs. 311

Fig. 64. Reconstruction of an 18 mm. cat embryo (series 88), both sides, ventral
aspect.

Fig. 65. Reconstruction of an 18 mm. cat embryo (series 88), left side, lateral
aspect.

Fig. 66. Reconstruction of a 25 mm. cat embryo (series 22), left side, ventral
aspect.

JUGULAR LYMPH SACS IN THE DOMESTIC ved
GeEorRGE 8S. HUNTINGTON aND CHARLES F. W. McCLuRE

INT JUGULAR VEIN

~

RIGHT EXT. JUG. V. Yi IS LEFT EXT. JUG. v.

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THE AMERICAN JOURNAL OF ANATOMY—VoL. 10, No

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE S. HUNTINGTON AND CHARLES F. W. McCLuRE

INT. JUGULAR VEIN

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JUGULAR APPROACH |

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THE AMERICAN JOURNAL OF ANATOMY--VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE 8. HUNTINGTON AND CHARLES F. W. McCuiure

\

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THe AMERICAN JOURNAL or ANATOMY—VOL. 10, No. 2

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GeEorRGDP S. HUNTINGTON AND CHARLES F. W. McClure

THYRO-CERVICAL A. THORACIC

SUBCLAVIAN

APPROACH
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CONFLUENCE
{TAP}

THE AMERICAN JOURNAL OF ANATOMY--VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
Georeep S. HuNTINGTON AND CHARLES F. W. McCiure

ARCHED PORTION
OF PRECARDINAL

CAUDAL OR STRAIGHT

"SEGMENT OF PRECARDINAL

5S

————— DORSAL VENO-LYMPHATIC
° PLEXUS

EXT. JUG. V.

JUGULAR PROMONTORY AND,
iW DUCT OF CUVIER

6S
7S

8S

Iv PRIMITIVE ULNAR SEGMENT
OF POSTCARDINAL

DUCT OF CUVIER
POSTCARDINAL

AREA OF CONFLUENCE BETWEEN
DUCT OF CUVIER AND POSTCARDINAL

8

TuE AMBRICAN JOURNAL OF ANATOMY--VoL. 10, No. 2.

ANLAGE OF PRIMITIVE ULNAR
VEIN AND VENO-LYMPHATIC

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE 8S. HunTINGTON AND CHARLES F. McCLuRE

cEPHALIC ARCH

CEPHALIC DIVISION VENTRAL
FRECAR DT VENO-LYMPHATIC PLEXUS

(STRAIGHT SEGMENT)

PARA-PRECARDINAL
CHANNEL

DORSAL VENO-

AREA OF TAP B
LYMPHATIC PLEXUS

AREA OF TAP C

DUCT OF CUVIER
7 ANLAGE COMMON TO
PROMONTOR 7 PRIMITIVE ULNAR VEIN

AND LYMPHATIC

POSTCARDINAL

THr AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
Georep S. HUNTINGTON AND CHARLES F. W. McCLurE

PRECARDINAL
STRAIGHT SEGMENT

PARA-PRECARDINAL ie
CHANNEL
EXT. JUG. V. ( 6S

ae)
om

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AREA OF TAP B

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DUCT OF CUVIER PROMONTORY

POSTCARDINAL

10

Tue AMERICAN JOURNAL OF ANATOMY—VoOt. 10, No. 2.

DORSAL VENO-
LYMPHATIC PLEXUS

a ae AREA OF TAP C
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LYMPH sac sh

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GeoRGE S. HunTINGYON AND CHARLES F. McCLuRE

1VL

2vL

PRECARDINAL
(STRAIGHT SEGMENT)

PARA-PRECARDINAL

58
CHANNEL (VL)

DORSAL VENO- LYMPHATIC
PLEXUS

AREA OF TAP B 6S

oh
: AREA OF TAP C
EXT. JUG. V.

7S

ANLAGE COMMON TO
PRIMITIVE ULNAR VEIN
AND LYMPHATIC

DUCT OF CUVIER
PROMONTORY

POSTCARDINAL

11

THe American JouRNAL or ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN ‘THE DOMESTIC CAT
GEORGE S. HUNTINGTON AND CHARLES F. McCLuRE

PRECARDINAL

4S

PARA-PRECARDINAL
CHANNEL

6S

DORSAL VENO-
LYMPHATIC
PLEXUS

PROMONTORY

11a

WALIc ARCH A-B

2s

CEPHALIC DIVISION VENTRAL
/ VENO-LYMPHATIC PLEXUS

PRECARDINAL
(STRAIGHT SEGMENT)

4VL

PARA-PRECARDINAL
CHANNEL

CAUDAL DIVISION VENTRAL
VENO-LYMPHATIC PLEXUS

>

EXT. JUG. v. >

TAP. B

5S DORSAL VENO- LYMPHATIC

PLEXUS
6S

AREA OF TAP C

7s
CEPHALIC VEIN

THYRO-CERVICAL A.

PRIMITIVE ULNAR

DUCT OF CUVIER VENO-LYMPHATIC

PRIMITIVE ULNAR VEIN

POSTCARDINAL

PROMONTORY
12

THe AMERICAN JOURNAL Or ANATOMy—VoL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GeorG®P S. HunTINGTON AND CHARLES F. W. McCLurE

PRECARDINAL
(STRAIGHT SEGMENT)

CAUDAL DIVISION VENTRAL

VENO-LYMPHATIC PLEXUS ——— |

PARA-PRECARDINAL CHANNEL
JUGULAR APPROACH

TAP B

EXT. JUG. V. 4)

CEPHALIC VEIN
TAP C

PRIMITIVE ULNAR VEIN

DUCT OF CUVIER PROMONTORY

POSTCARDINAL

13

THE AMERICAN JOURNAL OF ANATOMY--VOL, 10, No. 2.

CEPHALIC DIVISION
— VENTRAL VENO-

LYMPHATIC PLEXUS

SP. N. IV
4VL
ave

SS SP. N. v

DORSAL VENO.- LYMPHATIC
6s PLEXUS

7s

SUBCLAVIAN APPROACH
“—~SP..N. VI

8S
i RO=CERVIGAILSAG
PRIMITIVE ULNAR

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9S

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SUBCLAVIAN VEIN
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aA ;

if

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JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GrEorRGE S. HunTINGTON AND CHARLES F. MCCLURE

TAP A

ANTERIOR TAP
OF EVACUATION

SP. N. I

sp. N. Il

SP. N.. Ill

SP. N. IV

PRECARDINAL

(STRAIGHT SEGMENT) 3VL

DORSAL VENO-LYMPHATIC
ARCH
4VL

“Z sP. N. V

VENTRAL VENO-LYMPHATIC SAC 4s

CAUDAL DIVISION
DORSAL VENO-LYMPHATIC

SAC

JUGULAR APPROACH
5S
TAP B
CEPHALIC VEIN SP. N. VI
EXT. JUG. V.
6S

SUBCLAVIAN APPROACH THYRO-CERVICAL A.

PRIMITIVE ULNAR VEIN
7s

TAP C
PRIMITIVE ULNAR

VENO-LYMPHATIC

SUBCLAVIAN V.

SUBCLAVIAN A.

DUCT OF CUVIER

POSTCARDINAL

14

THE AMERICAN JOURNAL OF ANATOMY—VoOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
Grorap S. Huntincron AND CHARLES F. W. McCLuRE

eommm SP..N. I!

ANTERIOR TAP
OF EVACUATION

SUBCUTANEOUS
DUCT

PRECARDINAL
(STRAIGHT SEGMENT)

INT. JUG. V.
COMMON JUGULAR
LYMPHATIC SAC
SP. N. V
CEPHALIC V.
JUGULAR APPROACH
EXT. JUG. V.

4s

5S

SP. N. VI

TAP B

SUBCLAVIAN APPROACH
THYRO-CERVICAL A.

7S

PRIMITIVE ULNAR

TAP C
VENO-LYMPHATIC

PROMONTORY
SUBCLAVIAN V.

DUCT OF CUVIER
SUBCLAVIAN A.

POSTCARDINAL

15

Tun AMERICAN JOURNAL Or ANATOMY—VoOt. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE S. HUNTINGTON AND CHARLES F. W. McCiure

SP.N. 1

SUBCUTANEOUS DUCT

PRECARDINAL (INT. JUG. V.)

JUGULAR LYMPH SAC
JUGULAR LYMPH SAC

Cc
: SP. N. V

EXT. JUG. V.

THORACIC DUCT CEPHALIC vy.

ENTRANCE

JUGULAR APPROACH SUBCLAVIAN APPROACH

COMMON JUGULAR V.

LEFT INNOMINATE V.

(PROMONTORY) THYRO-CERVICAL A.

—— SUBCLAVIAN v.

SUBCLAVIAN A,

LEFT DUCT OF CUVIER

LEFT POSTCARDINAL

16

Tuer AMERICAN JOURNAL OF ANATOMY--VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE S. HUNTINGTON AND CHARLES F.. MCCLURE

*

‘SP. N. 1

SPLN.
INT. JUG. V.

(STRAIGHT SEGMENT
OF PRECARDINAL)

ge SUBCUTANEOUS DUCT

JUGULAR LYMPH SAC

JUGULAR LYMPH SAG

_S SP. N. V

THORACIC DUCT ENTRANCE

EXT. JUG. V.

JUGULAR LYMPH
SAC

CEPHALIC VEIN

SP. N. VI
JUGULAR APPROACH

JUGULO-CEPHALIC TRUNK SUBCLAVIAN APPROACH

THYRO-CERVICAL A.

COMMON JUGULAR TAP PRIMITIVE ULNAR

LYMPHATIC

SUBCLAVIAN V,
COMMON JUGULAR V.

JUGULO-SUBCLAVIAN TAP SUBCLAVIAN A,

LEFT INNOMINATE V.-—~
(PROMONTORY)

LEFT PRECAVA

LEFT POSTCARDINAL

7

THE AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
Gerorce S. HuntTINGTON AND Cuar.es F. W. McCLure

1VL SF Qs

VENTRAL
VENO-LYMPHATIC
PLEXUS

CEPHALIC DIVISION 2VL

2S

3S

3VL

PRECARDINAL
S48

f=)

4VL AND

VENTRAL (PARA-
VENO-LYMPHATIC< PRECARDINAL
PLEXUS CHANNEL

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19

Tur Amprican JouRNAL or ANaTomY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE 8. HUNTINGTON AND CHARLES F. W. McCLure

VENTRAL
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THE AMERICAN JOURNAL OF ANATONY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE S. HuwrINGTON AND CHARLES F. W. McCLurRE

CEPHALIC ARCH

STRAIGHT SEGME
OF PRECARDINAL

EXT. JUG. V.
CARDINAL-CUVIERIAN
JUNCTION
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POSTCARDINAL

22

Tue AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GeEorRGDP S. HUNTINGTON AND CHARLES F. W. McCiureE

CEPHALIC ARCH

EXT. JUG. V.

= CARDINAL-CUVIERIAN
js JUNCTION

POSTCARDINAL DUCT OF CUVIER

23

Tue AMERICAN JOURNAL OF ANATOMY—Vot. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GerorcE S. HunTINGTON AND CHARLES F. W. McCLuRE
CEPHALIC ARCH

STRAIGHT SEGMENT
OF PRECARDINAL-
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24

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Tue AMERICAN JOURNAL OF ANATOMY—VoL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
Gerorep S. Huntineton anp Cuartes F. W. McCiure

STRAIGHT SEGMENT
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THE AMERICAN JOURNAL OF ANATOMY--VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE S. HountTINGTON AND CHARLES F. W. McCLureE

CEPHALIC ARCH

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29

THE AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE 8S. HUNTINGTON AND CHARLES F. W. McCLureE
A-B

DORSAL
VENO-LYMPHATIC
PLEXUS

PARA-PRECARD
CHANNEL

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DUCT OF CUVIER

POSTCARDINAL

30

CEPHALIC AnCH

STRAIGHT SEGMENT
OF PRECARDINAL

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Tue American Journat or ANAToMy—Vot. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GrorGe S. Huntincton AND CHartzes F. W. McCriure

EXT. JUG, V.

DORSAL
VENO-LYMPHATIC
PLEXUS

DUCT OF CUVIER

: .
POSTCARDINAL . : ANLA MON us

32

THE AMERICAN JOURNAL OF ANATOMY--VOL. 10, No. 2.

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JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE S. HUNTINGTON aND CHar_Les F. W. McCiure

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33

THE AMERICAN JOURNAL OF ANatomy—Vot. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE S. HUNTINGTON anpD CHartEs F. W. McCiure

SOR PREC AHDINAL |

PARA-PRECARDINA
(SOMATIC

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DUCT OF CUVIER

34

THE AMERICAN JOURNAL or ANATOMy—VoL. 10, No. 2.

5 CHANNEL

aj EXT. JUG. V,

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GrorGp S. HuNTINGTON AND CHARLES F. W. McClure

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39

THE AMERICAN JOURNAL OF ANATOMY--VoOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GrorGE S. HunTINGTON AND CHARLES F. W. McCLurE

CEPHALIC ARCH

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THe AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
Georee 8. Huntineron AND CHARLES F. W. McCiure

PARA-PRECARDINAL
CHANNEL (VL)

DORSAL
VENO-LYMPHATIC
PLEXUS

PROMONTORY

6S

POSTCARDINAL

37

~ 40

THE AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

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JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE S. HuNTINGION AND CHARLES F. W. McCLure

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39

THE AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GeorGE S. HunTINGTON AND CHARLES F. W. McCLureE

CEPHALIC ARCH fo.

EiVL

CEPHALIC DIVISION
VENTRAL VENO-
LYMPHATIC PLEXUS

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PROMONTORY
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PLEXUS

CEPHALIC VEIN

ANLAGE COMMON TO
PRIMIT!VE ULNAR VEIN
AND VENO-LYMPHATIC
DUCT OF CUVIER

POSTCARDINAL

Tue AMERICAN JOURNAL OF ANATOMY—VoOkL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE S. HUNTINGTON AND CHARLES F. W. McCLureE

AREA OF 1, 2 AND 3,

PARA-PRECARDINAL CHANNEL (VL)

AP

EXT. JUG. V.

PROMONTORY

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CEPHALIC DIVISION VENTRAL
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EXT. JUG. v. PLEXUS

DUCT OF CUVIER

ANLAGE COMM
BRIMITIVE ULNAR VGN
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POSTCARDINAL
PROMONTORY

42

Tue AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE 8. HUNTINGTON AND CHARLES F. W. McCLure

LEFT
DORSAL

p—_V ENO-LYMPHATIC
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43

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VEIN a

44

THE AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEORGE S. HUNTINGTON AND CHARLES F. W. McCLureE

CEPHALIC ARCH

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46

THe AMERICAN JouRNAL OF ANATOMY—VoL. 10, No. 2.

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PRIMITIVE ULNAR VEIN

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VENO-LYMPHATIC

THE AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GporGE S. HunTINGTON AND CHARLES F. W. McCiur®

‘CEPHALIC ARCH
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DUCT OF CUVIER &

A COMMON TO
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ANLAGE COMMON TO

PRIMITIVE ULNAR VEIN
AND VENO-LYMPHATIC

CEPHALIC VEIN

50

Tur AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GeEorGD 8. HunTINGTON AND CHARLES F. W. McCLurE

CEPHALiC ARCH

SPINAL ACCESSORY
NERVE
CAUDAL DIVISION DORSAL
VENO-LYMPHATIC PLEX AE Di SEN ARGHIeOn

EXT. JUG. V.— Qa

SUBCLAVIAN
APPROACH
JUGULAR APPROACH
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THYRO-CERVICAL A.~

DUCT OF CUVIER

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ANTERIOR TAP
=e OF EVACUATION
SPINAL ACCESSORY:
NERVE CEPHALIC ARCH

THYRO-CERVICAL A.

PRIMITIVE ULNAR VEIN

52

Tue AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GrEorGE S. HuNTINGTON AND CuHarves F. W. McCiure

yes ae ts LEFT
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el ae “af 4 /
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JUGULAR LYMPH—&
SAC

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INT UG INTERNAL JUGULAR
(PRECARDINAL)

54

THe AMERICAN JoURNAL or ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GrorGe 8. HunTINGION AND CHARLES F. W. McCLurRE

DORSAL
ARCH . OF
DORSAL goa A REYNE
UGULARLIVMPH
: SAC VENTRAL
ARCH OF
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VENTRAL. SAC
ARCH OF
oe eA LYMPH
AC LEFT
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INTERNAL JUGULAR
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LEFT
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RIGHT

JUGULAR LYMPH SAC<\ INTERNAL JUGULAR

(PRECARDINAL)

INTERNAL JUGULAR
(PRECARDINAL)

Tue AMERICAN JOURNAL Or ANATOMY—VOL. 10, No. 2

ae

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GEorGE S. HuntTINGION AND CHARLEs F. W. McCLurRE

ANTERIOR TAP

_OF EVACUATION

AREA OF 1, 2 AND+3

PRECARDINAL

VENO-LYMPHATIC SAC——
VENTRAL ARCH

-VENO-LYMPHATIC sac

JUGULAR APPROACH DORSAL ARCH

THYRO-CERVICAL A.

JUGULO-CEPHALIC VEIN

PROMONTORY— |

\
SUBCLAVIAN V.

57

THE AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GeorcE S. Hunrineron anp CHArRtes F. W. McCiure

CEPHALIC ARCH

JUGULAR LYMPH SAG

SPINAL ACCESSORY NERVE

SUBCUTANEOUS DUCT
SP. N. 1, Il

TERNAL JUGULAR
IN PRECARDINAL)

VENTRAL ARCH

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CEPHALIC VEIN

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SUBCLAVIAN APPROACH

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CEPHALIC ARCH

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SPINAL ACCESSORY NERVE

SP..N. |
INTERNAL JUGULAR
SP. N (PRECARDINAL)
THYRO-CERVICAL A, ;
SP.N. Ill IV—

JUGULAR LYMPH SAC CEPHALIC VEIN

-EXT. JUG. V.

SP. N. V
JUGULAR APPROACH
SP. N. VI
SUBCLAVIAN APPROACH
IV
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PRIMITIVE ULNAR SUBCLAVIAN Vv,
LYMPHATIC

59

Tue AMERICAN JoURNAL OF ANATOMY—VOL. 10, No. 2.

:
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JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GuorGe 8. HuntTmInGTON AND CHARLES F. W. McCiure

JUGULAR LYMPH SAC

sp. N. V

SUBCUTANEOUS
DUCT CAROTID A.

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CEPHALIC VEIN

SUBCLAVIAN APPROACH

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LYMPHATIC

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AORTIC ARCH

INT. MAMMARY A.

60

Tae AMBRICAN JOURNAL OF ANATOMY—VoL. 10 No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT

GEORGE S. HUNTINGTON AND CHARLES F. W. McCiure

EXT. JUG. V. JUGULAR LYMPH SAC

INT. JUGULAR VEIN

VAGUS N._
SUBCUTANEOUS DUCT

CAROTID A. ——
JUGULAR APPROACH SP. N. V

CEPHALIC VEIN

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SUBCLAVIAN APPROACH

INNOMINATE A.

PRIMITIVE ULNAR
LYMPHATIC

INT. MAMMARY A.
AORTIC ARCH
INT. MAMMARY V.

SUBCLAVIAN V.

61

JUGULAR LYMPH SAC

SUBCUTANEOUS DUCT

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62

Tae AMERICAN JOURNAL oF ANATOMY—VoOL. 10 No. 2.

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
GeEorGE S. Huntinci1on anp CuHaruss F. W. McCiure

INT. JUGULAR VEIN

JUGULAR LYMPH SAC
SP. N. V

CAROTID A.

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CEPHALIC VEIN

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EXT. JUG. V.
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SUBCLAVIAN APPROACH
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—— SUBCLAVIAN A.

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INT. MAMMARY A.

LEFT POSTCARDINAL
(CUVIERIAN JUNCTION)

AORTIC ARCH

LEFT POSTCARDINAL

63

Tae AMERICAN JOURNAL OF ANATOMY—VOoOL. 10, No 2

JUGULAR LYMPH SACS IN THE DOMESTIC CAT
Georce S. HuntiIncton anv Cuartes F. W. McCiure

LEFT INT. JUG. Vv.

—CAROTID A, CAROTID A -

INTERNAL JUGULAR —
LYMPHATICS

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RIGHT PRECAVA

64

Tue AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2

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Tue AMERICAN JOURNAL OF ANATOMY—VoOL. 10, No. 2.

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66

THE AMERICAN JOURNAL OF ANATOMY—VOL. 10, No. 2.

THE SINUS MAXILLARIS AND ITS RELATIONS IN THE
KHMBRYO, CHILD, AND ADULT MAN,
Ae
JACOB PARSONS SCHAEFFER,
Instructor in Anatomy, Cornell University Medical College, Ithaca, N. Y.

WiruH 31 Ficurss.

This paper is based upon a study of the sinus maxillaris in the
embryo in successive stages up to the fetus at term, as well as in
the child and adult man. Most of the adult specimens ranged in
age from 18 to 80 years.

The lateral nasal wall of the embryo, at different stages of
growth, was modeled; thus showing the relations of the sinus
maxillaris and its progress in development. The blotting-paper
method was used in all the reconstructions. Both solid structures
and cavities were modeled, thus securing positive and negative
views.

The work also covers a general consideration of the sinus in
the child and the adult; including a study of the sinus relations,
its ostium or aperture, and the ostium accessorium. Special
attention was also given to the cause and effect of recesses occur-
ring on the walls of the adult sinus. Dissections were made to
cover all phases of the problem.

In determining the size of the sinus the following measurements
were taken: (1) dorsosuperior diagonal; (2) ventrosuperior diag-
onal; (3) superoinferior; (4) ventrodorsal; (5) mediolateral.

The capacity of the sinus was determined by filling the cavity
with a portion of a previously measured liquid, or by measuring
the amount of water the hardened mucous membrane (represent-
ing the exact shape and size of the sinus) would displace.

I wish to take this opportunity for expressing grateful acknowl-
edgment to the heads of the Departments of Anatomy, and
Embryology and Histology for valuable criticisms and helpful

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 2.

314 Jacob Parsons Schaeffer.

suggestions. I also wish to express my grateful appreciation
of the abundant material and other facilities placed at my dis-
posal by the aforementioned departments. To Professor and Mrs.
Gage, for the loan of embryos from the research collection, I
wish to express thanks.

THE EMBRYOLOGY AND EARLY RELATIONS OF THE SINUS
MAXILLARIS.

About the tenth week of fetal life the mucous membrane
in the primitive middle meatus of the nose begins to pouch
laterally. This pouch represents the Anlage of the sinus maxil-
laris, which pushes from the originally simple furrow separating
the maxillo-turbinal (later concha nasalis inferior) and the first
ethmo-turbinal (later concha nasalis media).

In order to gain a clearer conception of the location and rela-
tions of this primary maxillary pouch, and to better interpret
adult conditions, a brief consideration of the lateral nasal wall,
of the embryo, is necessary.

_ During the second month of intrauterine life, before the cavum
nasi and the cavum oris have become separate cavities, we
find three swellings on the lateral wall of the nasal fossa
(maxillo-turbinal, appearing first; ethmo-turbinal, appearing
next; naso-turbinal—extremely rudimentary in man, appearing
later) (fig. 1). The maxillo-turbinal corresponds to the adult
concha nasalis inferior. The naso-turbinal, which is termed by
Peter, ‘‘der Agger nasi,”’ and by Killian, in conjunction with the
primitive processus uncinatus, ‘‘der erste Hauptmuschel,”’ per-
sists in the adult as the agger nasi. The ethmo-turbinal under-
goes subdivision, and by this division, according to Killian, five
ethmo-turbinal plates, defined by six grooves, are usually formed.
EK. Kallius says, ‘‘ Dass alle diese 6 Furchen ausgebildet sind, ist
selten.’’ Zuckerkandl’s investigations show that three ethmo-
turbinal plates are the typical number. He says, ‘‘ Drei Siebbein-
muscheln repriisentieren demnach die typische Faltungsweise
des Siebbeines.”’ According to the embryos studied for this
paper, I find that the number of furrows and ethmoidal conch
varies, but in the specimens examined, four plates are rather

The Sinus Maxillaris in Man. B05

common. The ethmo-turbinal plates and the resultant furrows
become reduced in number as development goes on, and finally
represent the conchz nasales, media and superior, and the
meatus nasi, medius and superior, respectively. The reduction
in number may not be carried so far, and this accounts for the
supernumerary ethmoidal conche and meatus in many adults.

Enc’lon.

Fic. 1 (X 10). Drawing of a frontal section through the head of an embryo
aged about 45 days, in the region immediately dorsal to the organon vomerona-
sale (Jacobsoni), note the development of the early turbinal processes, and that
there is no cartilage laid down in them at this early period. This is contradic-
tory to the theory that the turbinal processes are primarily the result of an
inpushing of the lateral nasal wall by cartilaginous strands which later become
the nasal conche. ;

Enc’lon, = encephalon; E. T., = ethmo-turbinalia; M. T., maxillo-turbinale;
P. Pal., = processus palatinus; Sep. Nasi, = septum nasi.

Just how the primitive nasal processes and furrows are formed
is interpreted differently. Some claim that the projections are
due to an inpushing of the lateral nasal wall by the cartilaginous
strands which become the nasal conche. This latter claim I
have been unable to verify, because I find that the folds or
projections are invariably present before cartilage is found in them

316 Jacob Parsons Schaeffer.

(fig. 1). The elevations at first consist of a duplication of the
ectoderm filled with indifferent mesenchyma, which, in part, later
changes into cartilage. Thenasal conchal cartilages are, therefore,
a result and not a cause of the early condition. Schoenemann
claims that they are elevations left by excavations of furrows
on the lateral wall of the nasal fossa. Killian and Mihalkovies
hold that the projections are free ingrowing folds on the lateral
wall of the nasal fossa. Glas concludes a discussion on the nasal
conche in rats thus:

Die Bildungsmodus der Muscheln, ist die Resultierende zweier
Komponenten; (1) des Auswachsens die Wandpartien einwachsender
Epithelleisten (Fissuren). (2) des Vorwachsens bestimmter Wand-
partien.

After a study of these early conditions I am led to believe that
the primitive furrows are primarily the result of an outpushing
or outgrowing of the mucous membrane on the lateral wall of the
nasal fossa. The projections, by duplication of the mucous
membrane (especially true in the ethmo-turbinal region) (fig. 1),
and the deepening of the furrows, become rapidly prominent.
At times the two processes, an outgrowth or outpushing, and a
duplication of the intervening mucous membrane and mesen-
chyme, seem to be at work simultaneously in forming the early
projections and furrows. The theory that the furrows are
primarily started as an outpushing or outgrowing of mucous mem-
brane is entirely in accord with, apparently, similar processes
taking place in the early embryo nose; namely, the pouching or
outgrowing of the mucous membrane as the Anlagen of some
of the sinus paranasales. That a similar process should cause
the formation of primarily similar outgrowths seems plausible.

It is, however, not the province of this paper to speak in detail
of the development of the early projections and furrows; suffice
it to say that it is from the furrow separating the primitive
conche nasales, inferior and media, that the maxillary pouch
evaginates. It is, therefore, the primitive meatus nasi medius
with its contained structures, and the naso- and first ethmo-

The Sinus Maxillaris in Man. S17

turbinals that especially concern us in the development and
relations of the primitive sinus maxillaris.

Killian terms the naso-turbinal and the subdivisions of the
ethmo-turbinal, ‘‘Hauptmuscheln;”’ and the smaller projections
appearing in the furrows between these ‘‘Hauptmuscheln,”’ as
‘“Nebenmuscheln.’’ What he terms ‘‘die zweite Hauptmuschel”’
will be spoken of in this paper as the concha nasalis media (first
ethmoidal concha).

The naso-turbinal plus the processus uncinatus and the concha
nasalis media have marked bends, thus presenting ascending
and descending crura. Correspondingly the furrow between
these conche has a bend, and presents ascending and descend-
ing limbs. For the sake of description we will consider the
processus uncinatus as the descending crus of the naso-
turbinal. The primitive meatus nasi medius has, therefore, as
inferior boundaries the cura of the naso-turbinal and the space
existing between the conch nasales, media and inferior. The
superior boundaries of the space are the crura of the concha
nasalis media (zweite Hauptmuschel of Killian) (fig. 4). To
say then, as has been done earlier in this paper, that the maxillary
pouch evaginates from the space separating the primitive conch
nasales, inferior and media, is not giving the pouch its definite
location. The actual point of this primary pouching is from
the primitive infundibulum ethmoidale, or the ‘‘unterer Re-
cessus des absteigenden Astes der ersten Hauptfurche”’ of
Killian.

This pouch is a minute epithelial sac, and forms the Anlage
of the sinus maxillaris. Its earliest establishment precedes the
appearance of the cartilage which later surrounds it. This is
in accord with the statement of E. Kallius, that

: die Nebenhdhlen der Nase schon angelegt sind,
ehe dee Knorpel entsteht, und dass also das Skelett sich erst sekundir
um jene herumlegt.

"The relation of the space existing between the conche nasales, inferior and
media, to the descending ramus or limb of the first furrow is spoken of thus by
Killian, ‘“‘Der Raum zwischen zweiter Hauptmuschel und unterer Muschel ist
demnach nur eine Art Vorhof zum absteigenden Theil der ersten Haupfurche.’’

318 Jacob Parsons Schaeffer.

According to my observations the earliest evidences of maxil-
lary pouching are found about the seventieth day of fetal life. Kal-
lius places the time of the primary evagination. during the
middle of the third month, ‘‘Die Oberkieferhohle erscheint in
der Mitte des 3. Monats.”’ J. Kollman places the time of pouch-
ing later, ‘‘Seine Anlage beginnt erst bei Foeten Von 8 cm.
Linge.’’ Gegenbaur quotes Dursy as authority for the following:

Schon bei 8 em. lange Embryonen buchtet sich der Raum der Nasen-
hdhle zwischen mittlerer und unterer Muschel gegen den hier verdickten
Knorpel der Seitenwand der Nasenhohle aus und bildet die Anlage des
Sinus Maxillaris.

I have found the primitive maxillary pouch duplicated, 1. e.,
two pouches growing laterally side by side. (This may explain
some of the duplications of the adult ostium maxillare—the two
primary pouches fusing distally, leaving the two points of evagi-
nation as the ostia maxillaria of the adult sinus. Other duplica-
tions of the adult ostium may be caused in a manner similar
to the formation of the accessory ostium).

This embryonal condition probably explains some of the cases
in which the sinus maxillaris is divided into two partially or
wholly separate compartments by a vertical partition, i. e., each
pouch developing into an adult cavity independent of its mate
(see subsequent paragraph).

The primary ostium maxillare varies greatly in its dimensions
in different embryos (figs. 2, 3). This is entirely in accord with
adult conditions, since the ostium of the adult sinus has a great
range of dimensions (table D). The great differences in the
dimensions of the ostium may be due to early fusion of two or
more primary maxillary pouches; or the primitive pouching
may have been single but extensive, as is frequently the case.
These two latter would give rise to long slit-like ostia, while the
single and less extensive pouching would give us the typically
shaped and average sized adult ostium. :

Sometime prior to the establishment of the maxillary pouch?

7 It is indeed difficult, in some cases, to say which structure is the primary one
in establishing an Anlage. It most cases the processus uncinatus is the first to

The Sinus Maxillaris in Man. 319

a ridge appears immediately inferior to the point of maxillary
evagination. This ridge is the Anlage of the processus uncinatus
and, as said before, will be considered, merely for the sake of
description, as the descending crus of the naso-turbinal. It will
be recalled that Killian terms the latter two structures, ‘‘die

M.N.M C.N
4s AS:
Le. M. N.M
O. M

S. M.

M. N. 1 Ps

Fie. 2 (X10). Drawing of a reconstruction of portion of the right nasal cavity;
including the meatus, infundibulum ethmoidale, and the sinus maxillaris. Only
that portion of the nasal cavity necessary to include the sinus maxillaris and its
relations is shown. :

Compare the size of the ostium maxillare with the corresponding aperture
in fig. 3.

The model was reconstructed from the nose of an embryo aged 105 days. It
must be remembered that the drawing represents cavity and is, therefore, a
negative.

M. N. M., M. N. L, =meatus nasi, medius et inferior; I. E., = infundibulum
ethmoidale; C.N., = cavum nasi; H. S., = hiatus semilunaris; O. M., = ostium
maxillare; S. M., =sinus maxillaris.

appear, and in some instances it is impossible to say whether the pouching of the
mucous membrane, or the formation of the ridge is first. It may, however, be said
that both structures are more or less dependent upon each other in establishing
Anlagen.

320 Jacob Parsons Schaeffer.

erste Hauptmuschel.’’ This ridge has its free border directed
superiorly, and it extends in a ventrosuperior direction. It early
tends to form a shallow groove immediately superior to it, which
is the primitive infundibulum ethmoidale (Recessus inferior des
absteigenden Astes der ersten Hauptfurche of Killian). To be

Fic. 3 (X 10). Drawing of a reconstruction of portion of the right nasal
cavity of an embryo aged 120 days. The drawing includes the meatus, infundi-
bulum ethmoidale, and the sinus maxillaris. Only that portion of the cavity,
was modeled so as to include the sinus maxillaris and its relations. Since it
represents cavity it is a negative.

Note the very extensive ostium maxillare in comparison to that in fig. 2.

M. NS., = meatus nasi superior; M.N.M.,M.N.1, = meatus nasi, medius
he inferior; I. E., = infundibulum ethmoidale; C.N., = cavum nasi; H. S., =
ttaius semilunaris; O. M., = ostium maxillare; S. M., = sinus maxillaris.

accurate, then, we must say that the maxillary pouch evagi-
nates from the primitive infundibulum ethmoidale —a part of the
meatus nasi medius.

Some time after the appearance of these structures there is
a more or less uneven bulging on the lateral wall of the primitive

The Sinus Maxillaris in Man. Soi

meatus nasi medius, immediately superior and lateral to the free
border of the processus uncinatus (fig. 4). This is the Anlage of
the bulla ethmoidalis. The slit-like space existing between the
free border of the processus uncinatus and the bulla ethmoidalis
is the primary hiatus semilunaris. Through this slit the infun-
dibulum ethmoidale communicates directly, and the primitive
sinus maxillaris indirectly, with the meatus nasi medius.

Palatum

Fic. 4 (X6.6). Drawing of a frontal section of the nose of an embryo aged
120 days. The section is 7.25 mm. from the tip of the nose. Note that on one
side the section is in the region of the ostium maxillare, on the other it is dorsal
to it; also the fusion between the processus uncinatus aad a frontal concha.

Inf. Eth-, = infundibulum ethmoidale; 0. M., = ostium maxillare; S. Max., =
sinus maxillaris; Os., = developing bone; F. Cr. Ant., = fossa cranii anterior;
B. Eth.,=bulla ethmoidalis; H. Sem., = hiatus semilunaris; Proc. Unc., = pro-
cessus uncinatus.

Killian subdivides the uneven thickening on the lateral wall
of his “Ramus decendens der ersten Hauptfurche” into small pro-
jections with very shallow intervening furrows. The projections
are his “‘absteigende Nebenmuscheln”’, and the furrows the ‘‘ obere
und untere Zwischenfurchen”’ of the first furrow, or the primitive
meatus nasi medius. He concludes that the bulla is formed by

322 Jacob Parsons Schaeffer.

the early fusion of some of the processes (Nebenmuscheln).
The space immediately inferior to his ‘‘untere Nebenmuschel”’
or the space he designates as the ‘‘ Recessus inferior”’ of his first
chief furrow, is the infundibulum ethmoidale—the exact place
of the primary maxillary pouching.

At this juncture mention must be made of the primary pouch-
ing of the sinus frontalis in order to interpret later conditions in
connection with adult fronto-maxillarv relations. It will be
remembered that the furrow from which the maxillary pouch
evaginates has ascending and descending rami, and that from
the descending ramus the maxillary evagination takes place.
The ascending ramus of this furrow widens and pushes ventrally
and superiorly. Turner says:

It is generally held that the frontal sinus commences to develop at
the end of the first or the beginning of the second year of life, as an up-
ward expansion of the ethmoid cell labyrinth.

Hartman quotes Steiner for the following:

Der erste Anlage der Stirnhéhle ist in der Anlage des Knorpeligen
Siebbein-labyrinthes gegeben mit der Entwickelung der zelligen Raiume
des vorderen Siebbeinlabyrinthes beginnt auch die der Stirnhdhle, denn
letztere stellt eben nur die Ausdehnung der vorderen Siebbeinzellen
nach oben dar.

Hartman makes the following statement:

Aus dem aufsteigenden Ast der ersten Hauptfurche bildet sich durch
oberflachliche Verwachsung eine sackartige Bucht, der Recessus ascen-
dens od. R. frontalis, die Stirnbucht. Aus der Stirnbucht entwickelt
sich die Stirnhohle.

The embryos studied showed evidence of a slight pouching at
the superior and ventral end of the primitive meatus nasi medius.
This doubtless corresponds to the ‘‘Recessus frontalis” of the
first chief furrow of Hartman and Killian.

According to Killian’s commendable work there are three
processes and four furrows on the lateral wall of the recessus
frontalis, which are designated by him as ‘‘Stirnmuscheln und
Stirnfurchen,”’ respectively. The processes, according to Killian,

The Sinus Maxillaris in Man. 323

merge, and form the cellule ethmoidales anterior, or cellule
frontales as some call them. From this he concludes that the sinus
frontalis may continue its development in one of the following
ways: (1) by extension of the frontal recess (direct method),
(2) by extension of a frontal cell (indirect method), (8) by ex-
tension of the frontal recess and a frontal cell, (4) by extension
of two frontal cells. With these facts kept in mind—allowing
for further differentiation during development, it is easier to
understand why the sinus frontalis, in the adult, connects either
with the infundibulum ethmoidale, with the meatus nasi medius,
or with both. These embryological facts are of importance in
connection with adult fronto-maxillary relations. Doubtless
more work should be done on the development of the nasofrontal
duct in order to clear up some points in connection with the rela-
tions existing between the sinus frontalis and maxillaris. J am
now working along this line and hope to report my findings at
some future time. |

Although the pouching to form the recessus frontalis, or what
may be termed the Anlage of the sinus frontalis, begins during
the third month of fetal life, as does that of the sinus maxillaris,
the extension of the sinus frontalis is for a time so small that it is
usually regarded as wanting at birth. This is in part due to
the fact that the sinus frontalis is as a rule looked for in the frontal
or vertical portion of the frontal bone, while the first evidences of
it are to be sought elsewhere. In fact, according to Lothrop’s
investigations, the sinus frontalis of the adult does not reach
the vertical or frontal portion of the frontal bone in about three
per cent of cases—the only evidences of the sinus appearing in
the horizontal or orbital portion of the frontal bone. It must,
however, be said that the sinus frontalis is tardy in its develop-
ment until after birth; while, on the other hand, the pouch form-
ing the Anlage of the sinus maxillaris develops more rapidly and
occupies a definite space in the lateral wall of the nasal fossa by
the end of the third fetal month (figs. 2, 3).

By the simultaneous processes of resorption of surrounding
tissue and the growth of the maxillary pouch, the primitive
cavity gains more amd more capacity. The pouch soon acquires

324 Jacob Parsons Schaeffer.

Fig. 5 (x 4). Drawings of the mucous membrane, representing the exact
shapes of the sinus maxillaris at different stages of its development in the fetus
and child.

A. From a fetus aged 4 months. B. From a fetus at term. C. From a child
aged 18 to 20 months. D. From a child aged 20 to 23 months.

ee en

The Sinus Maxillaris in Man. 325

a slit-like shape at the side of the nose (fig. 4). It has its greatest
measurement in the ventrodorsal direction, while mediolaterally
the cavity occupies comparatively little space. In embryos aged
from 100 to 105 days the ventrodorsal measurement is about 2
mm. (fig.2). In a120-day embryo the distance is about 2.5 mm.
(figs. 3 and5 A). Ina 100-day embryo the most ventral spur of
the sinus is about 6.5 mm., and the most dorsal spur 8.5 mm.
from the tip of the nose.

It will be remembered that, in the embryo, the processus
alveolaris of the maxilla is in proximity to the orbit, and when
we recall the fact that the unerupted teeth are contained in this
situation, it at once becomes evident that the sinus maxillaris
must be correspondingly small at this time. Because of these facts
the sinus of a 7-month fetus measures only 5 mm. in the ventro-
dorsal plane, while that of a fetus at term has increased this
distance to approximately 7 mm. (fig. 5 B). During the latter
month of intrauterine life the sinus gains in the mediolateral
plane, so that at term this distance measures from 3 to 4 mm.

It is generally stated that the deciduous teeth hold the sinus
maxillaris in check, and that the cavity rapidly assumes larger
dimensions as the first dentition progresses. I, however, find
that the growth of the sinus is rather uniform, and that the first
dentition has little to do with any rapid increase in the size of the
eavity. The age of the child and the size of the sinus, apparently,
progress pari passu (fig. 6).

The ventrodorsal measurement of the sinus in a child aged
6 months is 10 mm., but the cavity has not developed sufficiently
in the mediolateral plane to reach beneath the orbit. In a child of
9 months the ventrodorsal distance is 14 mm., with a superoin-
ferior measurement of 5mm. At the end of the first year the sinus
has reached a ventrodorsal measurement of 16 mm., a superoin-
ferior of 6 mm.; and has now reached a mediolateral point suffh-
cient to pass beneath the orbit. As the maxilla grows, the sinus
remains for some time on the medial side of the infraorbital
canal (fig. 10). By the twentieth month the sinus measures
ventrodorsally 20 mm. (fig. 5) and has, as a rule, extended above
the rudimentary first permanent molar tooth. 3

326

VENTRODOR

MEASUREMENT

20 mm.

mm.

mm,

7 mm.

mm.

mim.

mm.

mm.

mm.

mm.

10

mim.

mm,

mm,

mm.

mm.

mm.

mm.

3 mm.

2 mm,

1 mm.

Fia. 6.

dorsal plane.

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Jacob Parsons Schaeffer.
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The Sinus Mazxillaris in Man. ae

Before and during dentition the sinus maxillaris is separated
from the front of the maxilla by the unerupted teeth (fig. 17).
After the eruption of the deciduous teeth the cavity continues
to have a more or léss rounded and elongated shape (fig. 5 D).
It is really never spherical as is often stated, but has an irregular
elongated form from the beginning.

After the eruption of the permanent teeth the sinus begins to
lose its rounded and elongated shape and to assume the adult
pyramidal form (figs. 7, 8, and 9). By the twelfth or fifteenth
year of age, when the second molar has appeared, the sinus
approaches, though it has not yet attained its definite shape.
The sinus reaches its full size between the fourteenth and eigh-
teenth year. .

THE ADULT SINUS MAXILLARIS.

The adult sinus maxillaris was known to Galenus (130-201),
‘but apparently Dr. Nathaniel Highmore was the first to give any
detailed description of it. In his work (1651), ‘‘Corporis Humani
Disquisitio Anatomica,” he describes the cavity in the maxilla,
to which his attention was drawn by a lady patient, in whom
an abscess of this cavity, since known as the antrum of Highmore
(sinus maxillaris), was drained by the extraction of the canine
tooth (left). The following are the exact words of Doctor
Highmore in describing the cavity located in the body of the
maxilla. His report of the case also follows.

; ‘ANTRUM MAXILLA! SUPERIORIS.”’

Antrum hoc utrinque unum, sub oculi sede inferiore ubi os ad ocul
tutelam quodammodo protuberat, ad latera inferiora nasi situm est.
Insigniter cavum sphaericum, aliquantulum vero oblongum, et ita
amplum ut articulus pollicis majoris pedis ultimus in illo delitescat,
a oe Osse attenuata seu squamma osseA obtegitur: Os enim
quod illud includit, et quod a dentium alveolis extremis distinguit,
crassitie chartam Emporeticam non multum excedit.

In basi hujus protuberantes quaedam eminentiae cernuntur, :
Quibus dentium apices tenniores includuntur. . . ._. Dentium
alveoli margini hujus ossis inferiori insculpuntur, quibus dentes in-

328 Jacob Parsons Schaeffer.

figuntur. . . . . Antrum hoc frequentius vacuum, aliquando
muco repletum reperitur, in quod humores a capite per meatum quendam
a cavitate illa in osse fronts et ab osse ethmoeide destillare poterunt.

Atque hic silentio praeterire non possumus, quod generosae cuidam
foeminae sub nostra cura laboranti accedit. Cum sub ferina eaque
continua falsi humoris distillatione, per multos retro annos laborasset,
omnesque pené dentes corrosos ac cariosos evulserat; nec tamen a
dolore liberata, tandem dente canino sinistri lateris effosso
Simu! squammosa illa distinctio inter cavitatem hance et dentis fanpeane
eriptur, adeo ut humorum, per alveolum dicti dentis, ab antro illo
perrenis successerit destillatio; Qua multum perterrita, stylo argenteo
in alveolum immissa originem fontis hujus exploratura, usque ad
oculum, per uncias pené duas sursum adegit; magis adhuc metuens,
pennam minorem plumis decerptis totam pené ad_ longitudinem
palmae unius immisit. Iam maximae consternata, ad Cerebrum usque
decurrere existimans, me inter alias consulit; ubi autem singulas exam-
inavimus circumstantias, pennae reduplicationes, illamque per cavi-
tatem hane circumgyrare invenimus. Atque sic, ubi in figura seguenti
cavitatem hane designavimus, illam de usu ac necessitate hujus satis
instructam, perennisque illius fontis patentissimam habuimus, a timore
et medicina simul desistit.

Antrum hoe levitatis ossium causa, quae hic oculorum situs gratia
crassa esse debuere, factum esse arbitramur.

The antrum (sinus maxillaris) described by Highmore must
have been an exceptionally large one, because the canine tooth
does not as a rule come in relation with the sinus. This same
tooth is mentioned by some writers even today as the tooth to
extract in draining the sinus. It is a very bad tooth to select
for this purpose in the great majority of cases. This fact will be
referred to in a subsequent paragraph when considering teeth
relations. It will also be noticed that Highmore had a somewhat
faulty idea of the shape of the adult cavity. He, however,
mentions some very essential conditions in his descriptions of
the adult sinus. His consideration of the cavity is very brief
and many important factors are omitted. His report of the case
through which his attention was called to this cavity, is unique and
interesting.

The Sinus Maxillaris in Man. 329

The adult sinus maxillaris, as we know it, is the large cavity
within the body of the maxilla. It is the largest of the sinus
paranasales, save in exceptional cases, when it is comparatively
small and may be exceeded in size by the sinus frontalis and the

Sinus Frontalis

Cc. Ethmoidales

Sinus Maxillaris

Fic. 7 (0.9). Photograph of a dissection of the sinus paranasales of the
left side. The mucous membrane is shown, the bony walls have been dissected
away after first hardening the subject in formalin.

C. N. M., C.N. Inf., = conche nasales, media et inferior; Sep. Nasi, = septum nasi;
Cc. Ethmoidales, = cellulae ethmoidales; S. Sphenoidalis, = sinus sphenoidalis ;
M. N.S.,M.N. M., M.N.I., = meatus nasi, superior, medius, et inferior, respect-
ively.

sinus sphenoidalis. It lies lateral to the cavum nasi and resembles
in shape a three sided pyramid (fig. 7). It follows in the main
the shape of the body of the maxilla; and may be described as
having a roof, a floor, and three walls. The walls of the sinus
vary in thickness, usually from 5 to 8 mm.; but they may be

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 2.

330 Jacob Parsons Schaeffer.

reduced to a papery delicacy. The base or median wall is directed
towards the cavum nasi, and the apex of the sinus extends into
the root of the processus zygomaticus of the maxilla. It may even
extend into the maxillary border of the zygomatic bone; thus
extending the recessus zygomaticus of the sinus maxillaris.

The ventral wall of the cavity corresponds to the anterior or
facial surface of the maxilla, and looks ventrolaterally. Part of
this wall is at times greatly approximated to the dorsal wall
and base of the sinus, due to a very prominent fossa canina.
Occasionally the whole ventral wall bulges markedly into the
savity of the sinus.

The dorsal wall of the sinus corresponds to the infratemporal
surface of the maxilla. It is a thin plate of bone, also forming
the ventral boundary of the infratemporal and the pterygo-
palatine fossee. This wall is usually the thickest of the sinus
walls—it is, however, occasionally extremely thin (the processus
alveolaris being recognized as the floor of the cavity and not as
a wall).

The base or median wall is directed towards the cavum nasi.
It presents a very irregular orifice—hiatus maxillaris, in the dis-
articulated bone. In the articulated skull this opening is partly
filled in by the pars perpendicularis of the palate bone, the
processus uncinatus of the ethmoid bone, the processus max-
illaris of the inferior nasal concha, and a portion of the lacrimal
bone. In the undissected state this irregular aperture tormed
by these bones is rounded by mucous membrane, which is con-
tinued into the sinus maxillaris from the cavum nasi. This
rounded opening—ostium maxillare, may be duplicated; and such
duplication must not be confused with the ostium maxillare
accessorium, which is a direct passageway between the sinus
and the cavum nasi. The ostium or ostia maxillaria establish a
communication between the sinus and the infundibulum eth-
moidale. The medial wall immediately inferior to the attach-
ment of the concha nasalis inferior is very thin and is easily
punctured in this region. This wall also forms the lateral bound-
ary of the cavum nasi and often markedly encroaches upon the
cavity of the sinus maxillaris, which greatly influences its size.

The Sinus Maxillaris in Man. 331

The roof of the sinus maxillaris is a very thin plate of bone,
at times of a papery delicacy. It also forms the floor of the orbit
and the orbital surface of the maxilla. It is often modeled by a
ridge formed by the infraorbital canal. In some cases the ridge is
replaced by a groove which is covered over with the mucous
membrane of the cavity. At times the roof of the sinus is par-
tially divided into two plates separated by air cells. Occasionally
the palate bone aids in forming the roof.

The floor of the sinus is formed by the processus alveolaris
of the maxilla. It is by far the thickest of the osseous bound-
aries of the cavity—the thickness of the floor depending upon
the degree of hollowing out of the process. In cases where the
hollowing out has been earried far, the floor of the sinus will bear
an important relation to some of the teeth and their sockets. The
floor of the sinus may be thrown into irregular elevations by the
fangs of the teeth—this depending upon the thickness of the
layer of spongy bone. This layer varies in thickness in different
skulls and there may be considerable asymmetry on the twe sides
of the same skull. The relation of the teeth will be considered
in a subsequent paragraph.

THE RELATION OF THE SINUS FLOOR TO THE NASAL FLOOR.

The relation of the floor of the sinus maxillaris to the floor of
the nose depends largely upon the degree of hollowing out of the
processus alveolaris of the maxilla. The degree of arching of the
pvalatum durum—-thereby affecting the floor of the nose, has also
some bearing on this relation. When the layer of spongy bone
is thin, 1. e., the processus alveolaris of the maxilla markedly
hollowed out, the floor of the sinus is at a level inferior to the
nasal floor. On the other hand, when the processus alveolaris
is comparatively thick, the floor of the nose is inferior to the
sinus floor. Occasionally both floors are in the same plane
(figs. 8,9 and 11). When the anterior surface of the maxilla and
the lateral wall of the nose are markedly bulging towards the sinus
maxillaris, the floor of the nose is, as a rule, inferior to the floor
of the sinus. It, however, remains that the majority of sinuses

332 Jacob Parsons Schaeffer.

have their floors, at varying distances, inferior to the floor of the
nose. Sixty adult specimens were examined to ascertain this
relation, with results as appended:

NUMBER SINUS FLOOR NOSE FLOOR SAME
EXAMINED INFERIOR INFERIOR LEVEL
60 39 12 9

The difference in levels of these two floors, when not in the
same plane, varies from one-half to 10 mm. C. Resschreiter
says that it is a male characteristic to find the sinus floor at an

Fie. 8 (X. 385). Photograph of a dissection of the sinus maxillaris and
frontalis from a ventral view.
S. Fron., S. Max., = sinus frontalis et maxillaris, respectively.

inferior level to the nasal floor. I have, however, been unable
to verify this statement, and give the following as typical of my
findings:

NUMBER SINUS FLOOR NOSE FLOOR SAME
EXAMINED INFERIOR INFERIOR LEVEL

12 (female) 9 2 1

ea a ee

The Sinus Maxillaris in Man. aaD
RELATION OF THE SINUS MAXILLARIS TO THE TEETH.

Since the sinus maxillaris varies greatly in size in different
skulls, and on the two sides of the same skull, it at once becomes
apparent that the relations of the teeth to the sinus cannot be
constant. As stated before, the layer of spongy bone between

S. Fron.
S. Fron.
Ge. Eth.
Suoph, O.Sph,
S: Sph.
ean S. Max.
Palatum M, C.N. Inf.
Sep. Nasi
Palatum D.

Fic. 9 (X .64). Photograph of a dissection of the sinus paranasales from a
dorsal view. The mucous membrane of the cavities is shown, the bony walls
have been dissected away. The sinus frontalis and sphenoidalis, and the cellu-
le ethmoidales of the right side have been opened.

S. Fron., S. Sph., S. Max., = sinus frontalis, sphenoidalis, and maxillaris,
respectively; Palatum M., Palatum D., = palatum molle and palatum durum,
respectively; Cc. Eth., = cellule ethmoidales; O. Sph., = ostium sphenoidale;
C. N. Inf., = concha nasalis inferior; Sep. Nasi., = septum nasi.

the roots of the teeth and the floor of the sinus varies in thickness
in different skulls, and the asymmetry on the two sides of the
same skull is at times marked. When this layer of spongy bone
is comparatively thin the projecting tooth fangs form elevations,
of a greater or less degree, on the floor of the sinus. These eleva-
tions at times aid in recess formation (fig. 16). Direet ecommuni-

334 Jacob Parsons Schaeffer.

cation between the fangs of the teeth and the mucous membrane
of the sinus, due to extreme hollowing out of the processus
alveolaris of the maxilla, occurs most frequently in the aged
(fig. 15). This latter condition does, however, occasionally pre-
vail in the young adult (fig. 16). That very intimate relations
frequently exist between the teeth and the sinus maxillaris is a
fact that we should be cognizant of, but I find that these intimate
relations have been somewhat exaggerated by some writers.

Fic. 10 (X .8). Photograph of a frontal section of a child’s face aged from
16 to 18 months. Note the infraorbital canal and nerve, and the relation of the
sinus maxillaris to the developing teeth. It will be noticed that the sinus max-
illaris has developed sufficiently to reach beneath the orbit but that it is medial
to the infraorbital canal.

C. Info., = canalis infraorbitalis; S. Max., = sinus maxillaris.

The number of teeth that bear a direct relation to the sinus
is necessarily inconstant, as stated before. In exceptional
cases, when the cavity is very large—especially in the line of the
ventrosuperior diagonal, all of the teeth of the trwe maxilla may
be in relation with the sinus (fig. 15). It is, however, only an
occasional occurrence to have the canine in direct relation with
the sinus. In a certain number of cases the first premolar tooth

The Sinus Maxillaris in Man. 335

bears a direct relation to the cavity, and in a slightly larger per-
centage of cases the second premolar bears a similar relation.
The three most constant teeth, however, in direct relation to the sinus
are the three molars. When the sinus maxillaris is small the first
molar must be omitted from the direct relation (figs. 12 to 16).

Orbita

S. Max.

H.Sem.

Proc. Unc.

Fic. 11 (X .8). Photograph of a frontal section of an adult’s face in the
region of the sinus mavillaris. Note the location of the ostium maxillare and th
infundibulum ethmoidale. Although the section is not exactly a frontal one
there is nevertheless a marked asymmetry in the size of the two sinus maxillares.
There is also quite a difference in the relation of the sinus floor to the nasal floor
on the two sides.

S. Max., = sinus maxillaris; B. Eth., = bulla ethmoidalis; H. Sem., = hiatus
semilunaris; Proc. Unc., = processus uncinatus; C. Info., = canalis infraorbi-
talis; Inf. Eth., = infundibulum ethmoidale; O. M., = ostium maxillare.

It is a fairly safe rule to follow that, when the canine fossa
and the lateral nasal wall are simultaneously approximated, the
canine and premolar teeth do not bear a direct relation to the
sinus maxillaris. In such cases a perforator pushed through a pre-
molar-tooth socket might readily enter the lateral nasal wall—even
pass through it, passing entirely free of the sinus cavity. Again, if
the perforator were pushed through the lateral nasal wall, inferior

336 Jacob Parsons Schaeffer.

Frias. 12 to17. Photographs of dissections showing variations in the teeth
relations of the sinus maxillaris.

Fias. 12, 13 (x .48). Note the short ventrosuperior diagonal of the sinus
maxillaris, due to the simultaneous approximation of the ventral wall and the
base of the sinus. As a result, the only teeth in direct relation to the cavity are
the second and third molars. Maes

Fia. 14 (x .456). Note that the premolar teeth are not in direct relation, and
that the canine tooth, as in the preceding figures, would certainly be a bad tooth
to use in attempting to drain the cavity through the canine socket.

Fic. 15 (x .48). This figure shows to what extreme the body of the maxilla
may be hollowed out by the sinus maxillaris. Note the very delicate walls of the
cavity, especially the shell-like alveolar process. Due to the extensive recessus
alveolaris the ‘“‘remaining tooth’’ projects into the lumen of the cavity and is
merely covered with the mucuous membrane of the sinus.

Fig. 16 (Xx .552). Although the processus alveolaris is comparatively thick,
the second-molar tooth fangs just reach the mucous membrane of the cavity.
Note the large ostium maxillare accessorium.

Fia. 17 (X 1.28). Showing the relations of the developing teeth to the sinus
maxillaris, of a child aged from 18 to 20 months. Note the position of the Anlagen
of the permanent teeth.

S. Max., = sinus maxillaris; C. P., = crescentic projection; D. p. r., = dentes
permanentes rudimentii; Sep. Nasi., = septum nasi; F. Info., = foramen infra-
orbitale; D. d., = dentes decidui.

ee ee

The Sinus Maxillaris in Man. BBY

Orbita
Sep. Nasi

F. Info.

338 Jacob Parsons Schaeffer.

to the concha nasalis inferior, the instrument could readily be
pushed through the soft structures of the cheek, unless the point
were directed well superodorsally.

RIDGES, CRESCENTIC PROJECTIONS, AND SEPTA ON THE SINUS
‘ WALLS.

It is important to note how frequently the walls of the sinus
are found uneven. ‘These irregularities may consist of mere
ridges or of different sized crescentic projections. The crescentic
projections have been reported occasionally replaced by septa
which completely divide the sinus into two cavities, each hav-
ing its independent opening into the nasal fossa, but not com-
municating with each other (fig. 22). The smaller ridges are of
little consequence and may be omitted from further consideration.
The larger ridges and crescentie projections, on the other hand,
tend to form pockets and recesses, of varying depths, within the
cavity. The septa, when they exist, may be placed either supero-
inferiorly or ventrodorsally; thus forming either ventral and dor-
sal, or inferior and superior compartments, respectively. Accord-
ing to Zucherkandl’s findings, the superior and dorsal cavities
communicate with the meatus nasi superior, and the ventral and
inferior cavities with the infundibulum ethmoidale. Bruhl found
the inferior compartment communicating with the meatus nasi
inferior. Gruber found a complete division of the sinus maxil--
laris in 2.5 per cent of cases.

In the material used for this paper no sinus was found show-
ing division into two distinctly separate compartments, but the
specimens repeatedly showed crescentic projections and ridges
which formed pockets of a greater or less depth (figs. 18 to 23).

Sixty sinuses were examined to cover this phrase of the work,
with results as follows:

NUMBER RIDGES OR CRESCENTIC SINUS WALLS
EXAMINED PROJECTIONS EBYEN
60 29 31

It must be borne in mind that, in the 29 positive sinuses,
quite a number of them showed mere ridges—the latter will be
omitted from further study. The remaining number of the

The Sinus Maxillaris in Man. 339

positive group fall, however, into a very important class of speci-
mens. That these crescentic projections offer, at times, almost
insuperable obstruction in attempting to drain fluid from the
sinus through an opening either in the processus alveolaris, or in
the meatus nasi inferior, is a fact that we should be cognizant
of. This was repeatedly demonstrated by first filling the sinus
with a liquid, then making an opening at some point on the
processus alveolaris; thus draining out what would come away.
If some of the fluid was retained—allowing for adherence to
mucous membrane—the facial or anterior surface of the maxilla
was removed to find where the remaining fluid was lodged. As
a rule the portion of fluid was retained by a recess or recesses on
one or more of the sinus walls. At other times a second and
even a third opening was made, either through the alveolar
border or -through the meatus nasi inferior, before the remain-
ing fluid would come away. If after repeated attempts the fluid
could not be located, the ventral wall of the cavity was removed
to ascertain the reason for its retention, and the fact was thus
demonstrated that repeated punctures, in some cases, would not
reach all of the recesses.

Just what these recesses mean in all cases is difficult to say.
Some of them are of course formed by elevations caused by tooth
fangs, but these as a rule are of minor importance and only
occasionally form deep recesses. Others are formed by projec-
tions of mucous membrane, which may or may not be caused
by ecrescentic bone projections. Where complete septa exist, the
sinus maxillaris very likely developed from two primary pouches.
In some cases the intervening wall may have disappeared in
part, thus leaving the larger crescentic projections which occa-
sionally are found in the adult sinus. A double pouching of the
primitive sinus maxillaris was mentioned in a previous para-
graph on the development of the cavity. Unequal resorption
of the bone during the growth of the sinus is doubtless a cause
for some projections occurring on the walls of the cavity.

Recess

CUP:

Fig. 22 Fig. 23

Fics. 18 to 23. Drawings of specimens showing septa and crescentie pro-
jections on the walls of the sinus maxillaris. Note how these projections form
recesses within the cavity. (Fig. 22 is modified from E. Zuckerkandl, Normaie
und pathologische Anatomie der Nasenhdéhle und ihrer pneumatischen Anhiinge).

C. P., = crescentic projection; O. M., = ostium maxillare; 0. M. A., = ostium
maxillare accessorium; S. Max., =sinus maxillaris.

The Sinus Maxillaris in Man. 341
THE SIZE OF THE SINUS MAXILLARIS.

The sinus maxillaris varies greatly in size in different individ-
uals. There may also exist considerable asymmetry on the
two sides of the same individual. The statement that all old
people have large sinuses is very fallacious, as is also the state-
ment that all females have smaller sinuses than males (tables A,
B,C.)

The investigations of Zuckerkandl have shown that enlarge-
ment of the sinus maxillaris may be produced by:

a. Hollowing out of the processus alveolaris of the maxilla
(recessus alveolaris) ;

b. Exeavation of the floor of the nasal fossa by a pushing of
the recessus alveolaris between the plates of the palatum durum
(recessus palatinus) ;

c. Extension of the sinus maxillaris into the processus fron-
talis of the maxilla (recessus infraorbitalis) ;

d. Hollowing out of the processus zygomaticus of the maxilla
(recessus zygomaticus) ;

e. Extension to, and appropriation of an air cell within’ the
processus orbitalis of the palate bone;

To these should be added, according to my findings:

f. Extreme hollowing out of the body of the maxilla in all
directions, thus causing the sinus walls to be thin and the recesses
all markedly developed;

g. The rarer condition when the lateral nasal wall is bulging
towards the cavum nasi;

h. The extension of the recessus zygomaticus of the sinus
maxillaris into the maxillary border of the zygomatic bone. |

Zuckerkandl! has found that the sinus may be made smaller,
on the other hand, by:

a. Deficient absorption of the cancellated bone on the floor of
the sinus;

b. Encroachment of the ventral wall of the cavity;

c. A deep fossa canina;

d. Thick sinus walls;

e. Excessive lateral bulging of the nasal wall;

342 Jacob Parsons Schaeffer.

f. A combination of the above conditions;

qg. Imperfect dentition.

The thickness of the sinus walls varies from 5 to 8 mm. and
down to that of a papery delicacy. The statement that all large
cavities have thin walls and small cavities invariably thick walls
does not hold in all cases. The smallest sinus measured in this
series had the thinnest walls—of a papery delicacy. The small-
ness of this cavity was in part due to the marked simultaneous
approximation of the ventral and medial walls.

The size of the sinus maxillaris is best determined by a series
of measurements, V1z:

1. Dorsosuperior diagonal (D.5. D.)
2. Ventrosuperior diagonal (V.5. D.)
3. Superoinferior (5. 1.)
4. WVentrodorsal (V. D.)
5. Mediolateral (M. L.)

These several measurements are determined thus (fig. 24):

Fic. 24. Schematic drawing of the right maxilla. The ventrolateral wall of
the sinus maxillaris has been removed, thus exposing the base or median wall
of the cavity. The lines drawn on the base indicate the position of the several
measurements.

1, Dorsosuperior diagonal; 2, Ventrosuperior diagonal; 3, Superoinferior;
4, Ventrodorsal; 5, Mediolateral.

The Sinus Maxillaris in Man. 343

1. The dorsosuperior diagonal, from the most dorsal and
lateral part of the sinus floor diagonally across the base or median
wall of the sinus, to the most medial and superior part of the re-
cessus infraorbitalis;

2. The ventrosuperior diagonal, from the most ventral and
medial part of the recessus alveolaris diagonally across the base
of the sinus, to the most lateral and superior point of the cavity;

3. The superoinferior, from the roof or infraorbital wall of
the sinus, to the sinus floor (always using uniform points) ;

4. The ventrodorsal, from the most ventral point of the
cavity midway between the roof and the floor, to the dorsal wall;

5. The mediolateral, from the base midway between its
most ventral and dorsal points, to the precessus zygomaticus of the
maxilla (in some cases this extends into the maxillary border of
the zygomatic bone due to the extension of the recessus zygomat-
icus of the sinus maxillaris into this bone).

The ventrodorsal distance is especially affected by the degree
of approximation of the ventral wall of the sinus; the superoin-
ferior by the degree of hollowing out of the processus alveolaris
of the maxilla; the mediolateral by the degree of encroachment of
the lateral nasa! wall; the ventrosuperior diagonal by the extent
of the recessus alveolaris; and the dors >superior diagonal by the
extent of the recessus infraorbitalis. Of cours? there are other
contributing factors to shorten or leng hen these distances, but
these are the primary factors especially affecting the several
measurements.

In order that the measurements of the sinus maxillaris may be
of most value, it is necessary to compare the two sinuses of the
same individual, to compare them with the respective sinuses of
another individual; also to consider the age and the sex.

A careful examination of the following tables (A, B, C) will
show conclusively that the sinus maxillaris has a rather wide
range of variation. These tables also show that in the adult,
age does not have much bearing on the size of the cavity. A
reference to table C will show that the smallest cavity is that
of an old man, aged 70 years; while the largest cavity is also that
of an old man, aged 77 years. This same table shows that the
cavity of a, young adult, aged 21 years is a close second to the

344 Jacob Parsons Schaeffer.
largest sinus found in the whole series. Although the cavity in
the male averages slightly larger than that of the female, a
reference to table C will show that sex affects the size of the sinus
but shghtly.

The following may be given as average measurements of the
adult sinus maxillaris, based on the measurements of 90 speci-

mens:
num.
! _Dorsosuperior diagonal. sa, oe 2. ae Reb Po en Nh 38
2 Vienbrosupenlor dra conmals, meena se cots eae ae ee eats ene 38.5
Bd SUperolmher On Mets senses eee eae TIS cen rae tg setae eae ene peenn 33
Aim (Viembrod Onsale eis cei cssra ese ie eee Ce teed meee ease cae ete 34
Br IMediolaberall.. cus d vx heey acca pat Ses mie ene ye ita Nene ae neue a 23

Due to the great differences in the several measurements, the
capacity of the sinus, in different individuals, must also differ.
The range in capacity, of the sinuses studied to ascertain this
fact, was from 9.5 ce. to 20 ec., with an average of 14.75 ce.

The tables A, B and C show the range of measurements.

The conditions which produce these varied differences in the
dimensions of the sinus maxillaris may be readily ascertained.
Take for example the following two conditions which showa marked
difference in the mediolateral plane and yet the other measure-
ments are inversed:

No. ; VoD; M. L. Soule IDS TS; 1D) Vien)
mm. mm. mm. mm. mm.

Le Aer AE 30 18 40 41 41

2 30 30 35 40 30

In case number 1 the- lateral nasal wall was markedly bulging
towards the sinus. In consequence of this encroachment, the
mediolateral distance was greatly lessened. In case 2 the recessus
alveolaris was poorly developed, hence the short ventrosuperior
diagonal in comparison with the respective measurement in case
number 1. These cases show that even though a sinus may greatly
exceed another in one of its measurements, it may be exceeded in
size in its other planes.

Again there may be a great difference in the ventrodorsal dis-
tance. This means a marked inpushing of the ventral wall of the

The Sinus Maxillaris in Man. 345

NUMBER |

10.

| M

M

| M

M

| M

M

36

TABLE A.
Bb a eeraiseeqaneeas|oamerioa: b, Suremon | bsurrecx
right 26 15 20 30 26
left 30 16 22, 32 26
right 40 22 50 50 50
left 30 24 39 45 50
right 32 32 40 40 38
left | 30 18 40 41 Al
| |
right etn 15 Bier ei S38 30
left 25 15 25 35 36
right 40 25 40 45 A5
left 40 22 38 36 45
right 40 21 32 | 50 38
left 32 25 30 | 32 43
right 30 22 45 | 45 40
left | 40 18 35 40 45
right 40 22 33 45 45
left 40 35 40 50 45
right 30 30 35 41 40
left | 43 20 30 41 37
right 31 24 30 | 30 38
left 32 20 35 40 40

346 Jacob Parsons Schaeffer.

TABLE B.
OSes fins a ees +e OS ee
Bs | fae. DORSO- VENTRO-
S| sex | AGE SIDE pi pecan aes pA deste SUPERIOR SUPERIOR
p | } | ei } ‘5 DIAGONAL DIAGONAL
Zz |
Be; | | | mm. : | mm. ; mm. mm. mm.
right 35 35 35 40 30
1| F | 684|
left 40° a2 | 16 30 43 36
[| right | 35 21 30 40 40
2 | TRH 52 f
| | left 35 24 28 38 45
| right 40 25 30 60 42
Bie Ba labs
left 33 30 45 45 - 46
right 36 296 25 37 PY aia
PN Uae: eg
left 37 28 35 35 37
right 33 24 31 38 40
5| F | 73
left 37 24 37 30 42
right 33 17 30 35 40
6| F | 50
left 33 22 33 38 34
right 30 18 32 30 35
7B 38
left 30 21 30 BOrS i oe a
right 34 25 33 32. | = 85
8) W389 | |
Sebi a4 33 22 33 32) | see
| |
Pielaeible gee 25 38 Bi ine a
97) awe : )
deft 35 23 38 33 35
right 35 21 30 40 36
10| F | 52 ae
left 35 21 32 38 35

eS < “ns
@ - a —
aye e on
- >./ tee .
» aly a ry +
ah _+ he >> al 2
- be em
< q~ » a
ei nae
- be y ree t

The Sinus Maxillaris in Man. 347

TABLE C.

ow)
g DOSRO- VENTRO-
| sex | ace | sIDE WAGiNTO BBE AGNES SUPERIOR SUPERIOR
A DORSAL LATERAL INFERIOR DIAGONAL DIAGONAL:
a

mm. mm. mm. mm. ‘ mm.
1|M | 70 | right 15 1st 21 ire 18
2}|M | 70 | left 16 12 21 21 20
3|/M | 35 | left 22 20 30 31 25
4|M | 54 left 25 15 22 32 27
o} EF | 54 | right 26 15 20 30 26
6;|M | 60 left 30 20 22 38 25
AR Sa rere site 35 25 30 37 38
8 | M_-| 59 left 40 22 32 45 45
Ole Nie 21h right 46 33 26 50 50
10} M | 77 left 47 40 50 57 60

sinus, on the one hand, and a shallow fossa canina with a lessened
encroachment on the other hand. Thus:

No. V. D. M. L. Sip Jf Diss D: Wietsn 10%
mm. mm. mm. mm. mm.

IL crepes cones ae 25 15 25 39 36

Dea tee Nek 43 20 30 41 37

If the body of the maxilla is hollowed out to a marked degree
in all directions the measurements will be correspondingly
lengthened. When this hollowing out has not been carried far,
and when associated with some of the above mentioned condi-
tions, the measurements will be markedly lessened. Thus:

No. Verb: M.L. Seale DiS: Db: V.S. D.
mm. mm. mm. mm. mm.

lee sree ah See tas 47 40 50 57 60

eget Oe ica iea ace 16 12 21 21 20

These few examples show how anatomical conditions will
affect the measurements of the sinus maxillaris. It, therefore,
appears reasonable that, by examination of the anterior surface
of the maxilla and the lateral nasal wall, the size of the sinus may
be approximately determined and the teeth relations judged. It
does, however, not necessarily follow, because the ventral and

348 Jacob Parsons Schaeffer.

median walls of the sinus are closely approximated, that the
sinus capacity is markedly lessened, These sinuses may have
marked infraorbital recesses and the processus alveolaris may be
hollowed out towards its dorsal termination. In this manner com-
pensation may be made for the marked bulging toward the cavity
of the ventral and median walls of the sinus. It, however,
remains that in the vast majority of cases, where these walls are
simultaneously bulging into the cavity, the sinus 1s correspond-
ingly reduced in size and the canine and premolar teeth not in
direct relation to the sinus.

Fic. 25 (* 1). Composite chart showing how anatomical variations in the
extent of the recesses and the approximation of the walls of the sinus maxillaris
affect the shape and size of its median wall or base.

1 to 6, = outlines of different bases; V. D., S. I., V.S. D.,D.S. D., = ventrodorsal,
superoinferior, ventrosuperior diagonal, and dorsosuperior diagonal, respectively.

These variations in the approximation of the sinus walls, and
the great difference in the extent of the various recesses, have
a marked effect on the shape of the base of the cavity. <A refer-
ence to figure 25 will show various shapes and sizes. Note espec-
ially case 4 in which the ventrosuperior diagonal is very short,
and the dorsosuperior, because of a marked infraorbital recess,
comparatively long. The great difference in the two diagonals
produces a peculiarly shaped base.

The Sinus Maxillaris in Man. 349
THE OSTIUM MAXILLARE.

When considering the embryology of the lateral nasal wall it
will be remembered that the primitive maxiliary pouch had cer-
tain relations of importance. These structures were the processus
uncinatus, the infundibulum ethmbdidale, the hiatus semilunaris,
and the bulla ethmoidalis. The location of the ostium maxillare,
of the adult, corresponds to the place of the primitive maxillary
pouch. This pouch gradually develops into the pyramidal cavity
of the adult, leaving the place of communication with the infun-
dibulum ethmoidale at the point of primary evagination. It is,
therefore, quite evident that these structures which in the embryo
bore so close a relation to the Anlage of the sinus maxillaris, must
now bear even more important relations to the ostium maxillare.

On raising or removing the middle nasal concha, in the adult,
a rounded elevation—the bulla ethmoidalis, is seen. This
structure is directed inferiorly and ventrally. Immediately
beneath it is the well defined curved margin of the processus
uncinatus of the ethmoid bone. Between these structures there
is a narrow slit or semilunar cleft—the hiatus semilunaris, which
is from 15 to 20 mm. long. This is an important opening, for it
serves as the communication between the meatus nasi medius
and the gutter-like groove (infundibulum ethmoidale) formed by
these structures. The bulla ethmoidalis varies considerably in
size. At times it is feebly developed and again it may assume
comparatively large proportions. The size of the bulla greatly
influences the width of the semilunar cleft or hiatus semilunaris.
The bulla may be so large that its convexity comes in direct con-
tact with the free margin of the processus uncinatus of the eth-
moid bone. In other cases the hiatus semilunaris may be of con-
siderable width.

It is easy to conclude what effect these conditions will have on
the ostium maxillare directly, and on the sinus maxillaris indirectly.
In one case the cleft of communication between the ostium maxil-
lare and the meatus nasi medius is practically shut off, while
in the other case a freer communication exists. It must be
remembered that, even though the bulla touches the free margin

350 Jacob Parsons Schaeffer.

of the processus uncinatus—thus greatly narrowing the hiatus
semilunaris, the infundibulum ethmoidale may be of average
dimensions. This is an important fact, and must always be borne
in mind when considering the fronto-maxillary relations.

The processus uncinatus with its covering of mucous membrane
projects inferiorly and dorsally. By its free superior border it
forms the inferior boundary of the hiatus semilunaris. This pro-
cess frequently terminates dorsally in what may be termed two
roots; the inferior one passes towards the superior edge of the
concha nasalis inferior, while the superior root curves superiorly
behind the dorsal termination of the bulla ethmoidalis (figs. 28 and
29). Such a condition, as the latter, causes the infundibulum
ethmoidale to end dorsally in a pocket. This fact is of extreme
importance because the pocket is so situated that it will direct
any fluid coming to the dorsal end of the infundibulum eth-
moidale into the sinus maxillaris, via the ostium maxillare which
is in the immediate location.

The infundibulum ethmoidale is a groove or gutter situated
upon the lateral nasal wall. It is bounded superiorly by the
inferior surface of the bulla ethmoidalis throughout the greater
part of its extent, save ventrally and superiorly where the bulla
is replaced by some anterior ethmoidal cells. The inferior and
medial boundary of the groove is formed by the lateral surface
of the processus uncinatus. This groove communicates with the
meatus nasi medius through the hiatus semilunaris. The in-
fundibulum may end, as stated above, in a pocket; or may loose
its depth gradually and be lost in the meatus nasi medius (figs. 28,
29, 30). The superior and ventral end of the infundibulum may
terminate blindly without dilatation, or in an air cell; or may be
continuous with the nasofrontal duct. The lateral wall of the
infundibulum is formed partly by mucous membrane. The depth
of this gutter-like channel, or the distance from the superior
border of the processus uncinatus to the floor of the groove,
varies from 1 to 12 mm., with approximately an average of 5 mm.

The sinus maxillaris communicates ¢ndirectly with the meatus
nasi medius by means of an opening—the ostium maxillare—
which pierces the superior and ventral part of the base of the

The Sinus Mawxillaris in Man. 351

cavity to open into the infundibulum ethmoidale, thence via the
hiatus semilunaris into the meatus nasi medius. It must be
clearly kept in mind that the ostium is located in the superior
part of the sinus, and that it opens into infundibulum ethmoi-
dale and not into the hiatus semilunaris as many writers say.
The ostium maxillare may be either in the most dependent part
of the infundibulum or in the lateral wall of this channel. This
opening varies in distance from the hiatus semilunaris from
1 to 12 mm. This distance is dependent upon the width of the
processus uncinatus and the resultant depth of the infundibulum
ethmoidale at this point.

Fic. 26 (X .66). Drawing of a specimen showing a direct communication
between the sinus frontalis and maxillaris (indicated by the arrow).

Note the very large slit-like ostium maxillare. The lateral wall of the infun-
dibulum ethmoidale is entirely wanting.

S. Fron., = sinus frontalis; S. Max., = sinus maxillaris; O. M., = ostium max-
illare.

The ostium may be round, but as a rule is either oval or ellip-
tical. In my series of 90 cases it has a great range of dimensions;
varying from 1 to 20 mm. in length, and from 1 to 6 mm. in width.
In some cases where the ostium has reached considerable size
it may almost entirely replace the lateral wall of the infundibu-
lum ethmoidale, thus forming a long slit-like communication

aa)

352 Jacob Parsons Schaeffer.

between the sinus maxillaris and the infundibulum ethmoidale
(fig. 26) (table D, nos. 7 to 12).

The following table gives an idea of the range of dimensions
of the ostium maxillee as found in the series of specimens studied:

TABLE D.

NUMBER LENGTH WIDTH
mm. mm.
Ee eee) Pe Pee ES on okie Wn VR ee lh Dae ea ate 1 1
i a BY RE UT ig Adam ns yan en te Pe; 2 3 3
Se Le Ee EE tec eo a RS ashi atte eo OS 3 2
A ete a aE ache PEE Eero ens Ae es SER RIEL CI Ee 5 3
EE tk fhas hci MARS en Raa cb oe a at ke ce i 4
OAS SES cae SoC AIR et teh re gta cea eee Sree n 8 3
1 SRS Oe pI A MRE a, oe MA A dnt OST AN gh WON TG 10 6
Oitodiee Saree cant ake! auth tae ath as shes AA, Reais iva ene Stee Mn hs ete Wil +
Qt gtahnaaihs Sts SOUR Sg) Dk Aetna mS CAL olaeh URN SNH i iil 6
1U URS ch cea ayy Ae a ER at ae Oo ts Oe oe els a eedonee = 14 3
10 ay ee ee Ae etre Pee on MOR DAL NT 10s Ue ie nar am SER De Seance 19 3
I ae RTE Ge bret MAT MEATS Ss oo Eco nny Attn MM eA Ab = 20 3

THE OSTIUM MAXILLARE ACCESSORIUM.

In many cases the sinus maxillaris has an accessory ostium
communicating directly with the meatus nasi medius—the ostium
maxillare accessorium: This opening is, as a rule, situated in the
membranous portion of the lateral wall of the meatus nasi medius
a short distance above the superior border of the concha nasalis
inferior, at about the junction of its middle and _ posterior,
thirds. In some instances the accessory ostium is placed imme-
diately behind the dorsal termination of the infundibulum eth-
moidale (fig. 27). This accessory ostium must not be confused
with the duplication of the ostium maxillare, which communi-
cates with the infundibulum ethmoidale.

According to Chiari and Hajek an accessory opening is found
in every fifth case in the meatus nasi medius, posterior and inferior
to the normal aperture. Giraldés says it is found in 10 per cent
of cases, and represents a pathological condition. Zuckerkandl
and Kallius report it present in 10 per cent of cases. Turner
found it four times in nine dissections.

That this accessory opening occurs more frequently than is
generally supposed seems proven by the study of 80 adult speci-

The Sinus Maxillaris in Man. 353

Fic. 27. Diagrams of the lateral nasal wall. The conche nasales medize
have been partially cut away so as to bring to view the underlying structures.
Note the positions and varying sizes of the ostium maxillare accessorium in the
different diagrams. The accessory ostia are designated by the deep black circles.
The upper and lower right hand diagrams show two accessory ostia and the others
but one.

354 Jacob Parsons Schaeffer.

mens. Out of SO specimens examined 35 showed accessory ostia,
or a percentage of 43; while three cases had two accessory ostia, or a
percentage of 3.75. From this it seems that the former figures
were much too low. Whether this series had a special run for
accessory ostia or whether too few specimens were used in the
former reports is of course not known. It may, however, be
said that the opening occurs very frequently and that the ear-
lier reports, apparently, placed the percentage of occurrence far
too low.

Just what the ostium maxillare accessorium means in all cases
is indeed difficult to say. It seems almost incredible that so
large a percentage of specimens should have pathological open-
ings. Giraldés bases his claim of a pathological origin on the
facts that the accessory ostium is absent in the young individ-
ual, and that the mucous membrane becomes thinned out in this
locality—even though the opening fails to establish itself. Zuck-
erkandl corroborates the thinning of the mucous membrane in
this locality at times, but claims that we have no evidence that it is
always a pathological process causing this condition. He says
that occasionally the accessory opening is caused by neighboring
structures.

Seltenenfalls entsteht ein Ostium maxillare accessorium durch Druck
von Seite nachbarlicher Organe; ich habe gesehen, das ein abnorm breiter
zugespitzer Hakenfortsatz der Nasenscheidewand an der hinteren
Nasenfontanelle eine Durchlocherung veranlasst hatte.

I have not found the accessory ostium present in the fetus and
infant. Unfortunately I have been unable to secure a sufficient
number of specimens between the ages of 6 and 15 years to draw
any conclusions of value on the occurrence of the ostium maxillare
accessorium during this period of time. I found the accessory
opening occasionally present in the young adult—17 to 20 years.
The specimens (adult) studied, ranged in age from 17 to 80
years, with the majority from subjects over 50 years old. That
some cases of accessory ostia are of pathological origin is doubt-
less true, but many cases certainly do not give any evidence
of a pathological process. The thinning of the-mucous membrane.

The Sinus Maxillaris in Man. 355

of which Giraldés and Zuckerkandl speak, is very evident in
many specimens. I, however, believe with Zuckerkand|! that we
must, in the majority of cases, look elsewhere than to a patho-
logical process for the determining factor in this condition.

In this connection it is important to note that out of the 34
stnus maxillares having accessory ostia, 27 of them had positive
relations with the sinus frontalis; i. e., the infundibulum eth-
moidale continuous with the nasofrontal duct. This would
indicate that 77 per cent of sinus maxillares having positive
fronto-maxillary relations have accessory ostia communicating
directly with the meatus nasi medius.

Another explanation for this accessory ostium may be found
in the fact that since the sinus maxillaris develops by the growth
of the sae and resorption of surrounding bone, its walls have a
tendency to become thinned out most at points of least resist-
ance. Such a point is found in the membranous portion of
the base of the sinus, where bone is entirely wanting—the usual
seat of the accessory opening. The mucous membrane in this
position may become thinned out to such an extent, by the
growth of the sinus, that an opening is formed; thus establishing
the ostium maxillare accessorium.

Since the ostium maxillare opens into the infundibulum eth-
moidale, and secondarily by way of the hiatus semilunaris
into the meatus nasi medius, it is apparent that the ostium max-
illare accessorium, with its more dependent location and direct
communication with the meatus nasi medius, is more advanta-
geously placed as a drainage opening for the sinus maxillaris. In
some cases the ostium maxillare certainly seems inadequate—
due to its position, relations, and size—to properly drain the sinus.
Why then may we not say that this accessory ostium, in some
cases, of necessity comes to be formed as a means by which the
sinus maxillaris can more readily dispose of accumulated fluid?
The process by which this is brought about need not necessarily
be termed pathological. Doubtless more information is neces-
sary on this point before we dare draw conclusions.

Of course some specimens present accessory ostia which look
decidedly pathological; and as Zuckerkandl points out some are

356 Jacob Parsons Schaeffer.

due to pressure caused by neighboring structures. I hope to
study the subject more extensively in the embryo and child to
see whether the opening, after all, at times, does not have an
embryological significance. Thus far I must agree with Giraldes
that the ostium maxillare accessorium does not appear in the
embryo and young child.

The accessory ostium varies much in size. In the series I
studied the range of measurements was from 1 to 10 mm. long,
and from one-half to 10 mm. wide. The opening may be round
or elliptical.

The appended table selected from a series of 80 specimens
gives the range in size:

TABLE E.

LONG WIDE
mm. mim.
1 i
2 it
4 4
6 4
7 5
10 10

THE FRONTO-MAXILLARY RELATIONS.

It is interesting to note that Nathaniel Highmore (1651)
recognized the fact that the sinus maxillaris at times receives
fluid from other sources. In his description of the cavity (see
previous paragraph) he makes brief mention of this important
condition.

Antrum hoc frequentius vacuum, aliquando muco repletum reperitur,
in quod humores a capite per meatum quendam a cavitate illa in osse
frontis, et ab osse ethmoeide distillare poterunt.

Although mentioning that fluid from the cavities in the frontal
and ethmoid bones occasionally reaches the sinus maxillaris by
way of the ‘‘meatum,’’ he does not attempt to explain how this
is brought about.

Tillaux (40) found when injecting fluid into the sinus fron-
talis that some of it passed into the sinus maxillaris, instead of

The Sinus Maxillaris in Man. Bie

the whole amount passing into the meatus nasi medius. Cryer
(94, ’01, 07), Fillibrown (96, 97), reported on fronto-maxillary
relations. Lothrop’s investigations (98) show that in 47 per
cent of cases the infundibulum ethmoidale is continuous with the
nasofrontal duct, while 53 per cent show that the infundibulum
ethmoidale has no connection with the sinus frontalis. ‘Turner
(01) speaks briefly about the relation, and Wilson (’08) in his
paper on the “‘ Variations of the Ostium Frontale’’ alludes to this
important relation. Some clinicians have reported isolated
cases where they believed the maxillary trouble secondary to
preéxisting frontal trouble; without, however, attempting to
explain any anatomical conditions which would justify the
clinical conclusions.

In order to secure the fronto-maxillary relations in the speci-
mens at hand, I undertook a series of investigations; including
special dissections, filling the sinus frontalis with a fluid to deter-
mine the direction of drainage, and the determination of the effi-
ciency of the infundibulum ethmoidale.

It will be remembered that the infundibulum ethmoidale at
its superior and ventral termination is either continuous with
the nasofrontal duct; or ends blindly without dilation, or in an air
cell. The cases where it is continuous with the nasofrontal duct
or with the sinus frontalis directly, represent what will be here
spoken of as the positive fronto-maazillary relations. Where the
infundibulum ends blindly or in an air cell, the conditions will
be spoken of as negative fronto-maxilary relations.

According to the specimens I examined, the sinus frontalis
may discharge fluid put into it in one of the following ways:

a. By the nasofrontal duct or the sinus frontalis being
continuous with the infundibulum ethmoidale (in some cases
there is no nasofrontal duct and the sinus frontalis is directly
continuous with the infundibulum ethmoidale) (positive relation)
(fig. 28).

b. By the nasofrontal duct communicating directly with the
meatus nasi medius (negative relation) (fig. 29).

c. By a combination of the above conditions—in which case
the sinus frontalis had two nasofrontal ducts; one continuous

Fra. 28. A semidiagrammatic drawing of the lateral nasal wall showing positive
fronto-maxillary relations. Note that the infundibulum ethmoidale is directly
continuous with the naso-frontal duct. Note also the superior and lateral curv-
ing of the processus uncinatus at its dorsal termination, thus forming a pocket at
the dorsal end of the infundibulum ethmoidale. This pocket is so situated that
it will direct fluid coming to the dorsal end of the infundibulum ethmoidale to
the ostium maxillare and into the sinus maxillaris.

The concha nasalis media is in part cut away so as to expose the underlying
structures.

Fic. 29. Asemidiagrammatic drawing of the lateral nasal wall with the concha
nasalis media partially removed. Note that the infundibulum ethmoidale termi-
nates blindly at its superior and ventral end. The nasofrontal duct communi-

cates directly with the meatus nasi medius and not with the infundibulum eth-

moidale as in the preceding figure (28). This represents negative fronto-maxillary
relations.

Fic. 30. A semidiagrammatic
drawing of the lateral nasal wall
showing that the sinus frontalis,
in this ease, has two nasofrontal
ducts, one communicating with
the infundibulum ethmoidale and
the other with the meatus nasi
medius.

Note that the infundibulum eth-
moidale terminates at its dorsal
extremity in the meatus nasi me-
dius without a pocket formation
(compare this condition with figs.
28, 29).

C. N.Sup.,C. N.Med., C. N. Inf., =
conche nasales, superior, media
and inferior; S. Sph.,=sinus sphen-
oidalis; S. F., = sinus frontalis; B.
Eth.,=bulla ethmoidalis; Inf. Eth.,
= infundibulum ethmoidale; Proc.
Unc., = processus uncinatus.

The Sinus Maxillaris in Man. 359

with the infundibulum ethmoidale, and the other communicat-
ing directly with the meatus nasi medius (positive and negative
relations) (fig. 30).

d. By the nasofrontal duct being continued down to. the
infundibulum ethmoidale; and in conjunction there being a
passage-way between the ventral attachment of the concha
nasalis media and the processus uncinatus of the ethmoid bone,
to the meatus nasi medius (considered as positive relations).

e. By a direct communication between the sinus maxillaris
and the sinus frontalis, by what may be termed the maxillo-
frontal duct (direct relation) (fig. 26).

Of the SO specimens studied to ascertain the fronto-maxillary
relations; 45 showed a positive relation, or a percentage of 56.25:
32 a negative relation, or a percentage of 40: 2.a combination of post-
tive and negative, or a percentage of 2.5 ; 1 a direct communication
between the two sinuses, or a percentage of 1.25.

The importance of the above conditions was in each case
tested by putting fluid unto the sinus frontalis to determine
the course of drainage. It at once became apparent that the
specimens falling under classes (a) and (d) should be classed
together as representing positive fronto-maxillary relations.
The only difference in the above two conditions is that in class
(a) all of the fluid put into the sinus frontalis will reach the
supertor and ventral part of the infundibulum ethmoidale;
while in class (d) some of it. will pass directly into the meatus
nasi medius, and the remaining portion to the infundibulum eth-
moidale.

Class (6) will drain fluid from the sinus frontalis directly into
the meatus nasi medius. It is, however, important to know
that even in these cases some fluid may reach the infundibulum
ethmoidale, because of the intimate relations existing between
the nasofrontal duct and the superior and ventral end of the
infundibulum ethmoidale (fig. 29).

Class (c), where the sinus frontalis has two nasofrontal ducts,
the drainage is of course partly into the meatus nasi medius and
partly into the infundibulum ethmoidale. This class leads to
similar results as mentioned above, the only difference being that

360 Jacob Parsons Schaeffer.

the infundibulum does not receive as much fluid in a given
time.

Class (e) fortunately represents a rare condition. Here the
sinus frontalis draii.s Cirectly into the sinus maxillaris. In the
specimen I found with this direct relation there was also a com-
munication between the infundibulum ethmoidale and the sinus
frontalis. Cryer, Bryan, and Brophy have reported direct rela-
tions between the two sinuses, which I have been able to verify
in this one specimen. In cases where the lateral wall of the in-
fundibulum ethmoidale is largely wanting, fluid from the frontal
sinus (providing the infundibulum ethmoidale is continuous with
the nasofrontal duct) will pass almost directly into the sinus
maxillaris, and will, therefore, very closely simulate a direct
communication between the two sinuses. A probe passed from
the sinus frontalis will, in such cases, also pass into the sinus
maxillaris. Doubtless some of these cases have been considered
by some clinicians as direct communications, whereas a further
dissection would have proved them otherwise (fig. 26).

The question now arises—what happens to the fluid that has
reached the superior and ventral end of the infundibulum eth-
moidale? In the first place it may be said that the efficiency of the
infundibulum ethmoidale, as a carrier of fluid, is in direct ratio
to its depth and to the degree of overhanging of the mucous
membrane from the free border of the processus uncinatus of the
ethmoid bone. In some cases the processus uncinatus is so nar-
row that the infundibulum ethmoidale has no appreciable depth
at its superior end, and in these cases the fluid which has reached
it from the frontal region will soon leave the shallow groove
after entering it—at least a goodly portion of it. In other cases
the processus uncinatus is broad, and the resultant infundibulum
ethmoidale deep and channel-like. It must also be recalled that
in a previous paragraph mention was made of the fact that fre-
quently the infundibulum ethmoidale ends dorsally in a pocket, so
situated that it will direct the flow of fluid coming to the dorsal
end of the infundibulum ethmoidale into the ostium maxillare—
thence into the sinus maxillaris (the ostium maxillare being
patent) (figs. 28, 29).

The Sinus Mawyillaris in Man. 361

We have, therefore, a gutter-like channel, of varying depth and
efficiency, communicating between the frontal region and the sinus
maxillaris; including the sinus frontalis in 56 per cent of cases and
some of the cellule ethmoidales anterior in nearly all cases.

In the cases where the infundibulum ethmoidale does not end
in a pocket dorsally (fig. 30), much of the fluid that would other-
wise be directed into the sinus maxillaris by this pocket, passes
from the dorsal termination of the infundibulum ethmoidale into
the meatus nasi medius. This, however, makes little difference—
the very fact that some of the fluid gets into the sinus maxillaris
makes the condition similar to the above. It requires merely
more time to accomplish the same end result—a filled sinus
makxillaris.

In case the ostium maxillare is not patent, the fluid after
reaching the dorsal end of the infundibulum ethmoidale rises
in the channel and finally passes through the hiatus semilunaris
into the meatus nasi medius.

That the sinus maxillaris, because of its position and relations,
is a reservoir for some or all of the fluid coming to the DORSAL END
of the infundibulum ethmoidale, is a fact that admits of no debate
(the ostium maxillare being patent).

IMPORTANT NERVE RELATIONS OF THE SINUS MAXILLARIS.

The roof or orbital wall of the sinus maxillaris is traversed
by the infraorbital suleus and the infraorbital canal. These
passageways transmit the infraorbital vessels and nerve (con-
sidering the maxillary nerve as the infraorbital nerve from the
proximal end of the infraorbital sulcus on). As a rule the canal
has comparatively thick walls, but in many cases the inferior
wall of the canal is of a papery delicacy and is easily compressed
against the contained nerve and vessels. Frequently the canal
is replaced by a groove, with the opening of the groove directed
towards the sinus mazxillaris. The structures—infrarobital
nerve and vessels—contained in the groove are merely covered
with the mucous membrane of the sinus.

The posterior superior alveolar (dental) nerves, branches of

THE AMERICAN JOURNAL OF ANATOMY, VoL. 10, NO. 2.

362 Jacob Parsons Schaeffer.

the maxillary nerve, in most of my cases were found to pass
inferiorly and ventrally upon the infratemporal surface of the
maxilla, through the alveolar foramina into the alveolar canals.
They thus aided in the formation of the superior dental plexus of
nerves. Occasionally some of the branches of these nerves
instead of taking the above course, passed entirely through the
infratemporal surface of the maxilla into the sinus maxillaris.
They then passed under cover of the mucous membrane of the
sinus inferiorly and ventrally to the sinus floor; thence to the
superior dental plexus.

Fia. 31 (X .633). Drawing from a disS8ection showing the anterior superior
alveolar nerve (N.A.S.A.) passing diagonally from the roof or orbital wall of the
sinus to the ventral or facial wall. The nerve in this position is suspended freely
in the cavity of the sinus maxillaris merely covered with mucous membrane.

N.A.S.A. = nervus alveolaris superior anterior; N. Info., = nervus infraor-
bitalis; S. Max., = sinus maxillaris.

The middle superior alveolar (dental) nerve, a branch of the
infraorbital nerve, was as a rule given off in the proximal part of
the infraorbital canal. It passed inferiorly and ventrally in a
canal in the lateral wall of the sinus maxillaris and aided in
establishing the superior dental plexus of nerves. The nerve I
found in one case to arise from one of the anterior superior
alveolar nerves. It also rarely passed under cover of the mucous
membrane of the sinus to the superior dental plexus.

The Sinus Maxillaris in Man. 363

The anterior superior alveolar (dental) nerve was given off
from the infraorbital nerve, proximal to the infraorbital foramen.
It passed inferiorly in the alveolar canal of the anterior surface
of the maxilla and took part in forming the superior dental plexus
of nerves. From this plexus arose the superior dental nerves
which supply the fangs of the teeth, the gums, and give numerous
branches to the maxilla and the mucous membrane of the sinus
maxillaris.

I also observed—a very important condition—that in one case
the anterior superior alveolar nerve came off’ from the infra-
orbital nerve quite a distance proximal to the infraorbital fora-
men. The nerve then passed through the inferior wall of the
infraorbital canal and took a course diagonally across the sinus
from the roof to its ventral wall. The nerve thus suspended
freely in the cavity of the sinus maxillaris was surrounded merely
with mucous membrane (fig. 31).

CONCLUSIONS.

1. The Anlage of the sinus maxillaris appears during the third
month of fetal life as a minute epithelial sac evaginating and
growing at first inferiorly, later more laterally, from the dorsal
end of the primitive infundibulum ethmoidale.

2. The primitive maxillary pouch may be duplicated. In
some cases this may account for the duplication of the ostium
maxillare of the adult sinus, i. e., the two pouches fusing distally,
leaving the two points of evagination as the adult ostia. Other
duplications of the ostium may develop in a way similar to that
of the accessory ostium.

3. The primitive ostium maxillare varies very much in its
dimensions in different embryos. This is entirely in accord with
adult conditions, since the ostium of the adult sinus has a great
range of dimensions.

4. Dentition seems to influence the size of the cavity but
little. The age of the child and the size of the sinus apparently
progress pari passu.

5. The cavity enlarges by the simultaneous growth of the

364 Jacob Parsons Schaeffer.

sac and the resorption of surrounding tissue. These two pro-
cesses taking place pari passu with the growth of the face.

6. In a fetus at term the ventrodorsal measurement of the
sinus is about 7 mm., and in a child aged 20 months it is about
20 mm. The cavity reaches its full, size from the fourteenth to
the eighteenth year.

7. The following may be given as average measurements of
the adult sinus maxillaris, based on the measurements of 90
adult specimens:

MM
i ~ Dorsosuperiordingonal.:. \ ese.) cee See oe eine 38
2; Ventrosuperior-diagonal. 17 Ae Gee ee 38.5
@ - PUPSTOIMLEDION . 5 1.55 ore eee eal eens kN ae eae 33
4: N emibrodorsale 5 5 50 5 50h it ee ee ES ee, eee 34
5° Mediolatenalr sc 2.35). cee San ee are ee ee ae ee 23

8. The range in capacity of the sinuses studied to ascertain
this fact was from 9.5 ce. to 20 ee.; with an average of 14.75 ee.

9. In the majority of cases the sinus floor is-at an inferior
level to the nasal floor. This distance varies from one-half to
10mm. Sex has little influence on this relation.

10. The number of teeth that bear a direct relation to the
sinus is inconstant, due to the great difference in the size of the
cavity in different individuals. The three most constant teeth
in direct relation are the three molars.

11. The tooth fangs may cause the formation of elevations
on the sinus floor. Occasionally the fangs of some teeth are in
direct communication with the mucous membrane of the cavity.

12, Frequently the walls of the sinus are uneven, due to
ridges, or crescentic projections. These prominences form pockets
and recesses within the cavity. Occasionally the cavity is divided
by a septum into two distinctly separate compartments, each
having an independent opening into the nasal fossa, but not
communicating with each other.

13. The adult sinus varies much in size in different individuals,
and the asymmetry on the two sides of the same individual is
often marked.

14. Age, sex, and side influence the size of the adult sinus
but little.

The Sinus Maxillaris in Man. 365

15. The adult ostium maxillare varies much in size. It is
located in the superior and ventral part of the base of the cavity,
and serves as a means of communication between the sinus
maxilaris and the infundibulum ethmoidale. Occasionally it
replaces the greater portion of the lateral wall of the infundibu-
lum ethmoidale, and represents a slit-like aperture. The ostium
may be duplicated.

16. The ostium maxillare accessorium is of very frequent
occurrence. It serves as a means of direct communication
between the sinus maxillaris and the meatus nasi medius. In my
series of specimens it was present in 48 per cent of cases. The
aperture was not found in the fetus and infant,

17. Most of the accessory ostia do not look pathological, and
the writer believes that we must, in many cases, look elsewhere
than to a pathological process for the determining factor in this
condition.

18. Of the specimens studied to ascertain the fronto-maxillary
relations, 56 per cent showed that the infundibulum ethmoidale
was intimately related with the nasofrontal duct or with the sinus
frontalis directly,—in case the nasofrontal duct was wanting; 40
per cent showed that the nasofrontal duct communicated directly
with the meatus nasi medius—the infundibulum ethmoidale
ending blindly or in an air cell; 2.5 per cent showed two naso-
frontal ducts, one continuous with the infundibulum ethmoidale,
and the other communicating with the meatus nasi medius;
1.25 per cent showed a direct communication between the
sinus frontalis and maxillaris.

19. Since the infundibulum ethmoidale receives the ostium
maxillare at its dorsal and inferior end in all cases, and the naso-
frontal duct, or the sinus frontalis directly, at its ventral and
superior end in over one-half the cases, it very frequently
serves as a gutter-like channel, of varying depth and efficiency,
communicating between the frontal region and the sinus max-
laris.

20. The sinus maxillaris, therefore, acts as a reservoir for
fluids coming to the dorsal end of the infundibulum ethmoidale
(the ostium maxillare being patent).

366 Jacob Parsons Schaeffer.

21. Frequently the processus uncinatus by a superior curv-
ing at its dorsal end causes the infundibulum ethmoidale to end
in a pocket. This pocket is so situated that it directs
fluids coming to the dorsal end of the infundibulum ethmoidale
into the sinus maxillaris,—via the ostium maxillare which is in
the immediate vicinity.

_ 22. Occasionally branches of the superior alveolar nerves in
passing to the superior dental plexus pass entirely through the
walls of the sinus, thence under cover of the mucous membrane
of the cavity to their destination. Rarely the anterior superior
alveolar ramus, instead of taking its usual course, passes diagon-
ally from the roof of the sinus to its ventral wall,—the nerve thus
suspended freely in the cavity is merely covered with mucous
membrane.

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THE INFLUENCE OF ALCOHOL AND OTHER ANAS-
THETICS ON EMBRYONIC DEVELOPMENT

CHARLES R. STOCKARD

Anatomical Laboratory, Cornell University Medical School, New York City

WITH TWENTY TEXT FIGURES

The adult nervous system is peculiarly sensitive in its responses
to the influence of alcohol and other anesthetics. The writer has
found it to be equally true that alcohol and anesthetics exert a
most striking influence over the development of the central ner-
vous system and the organs of special sense. There is consider-
able variation in the way in which the several aneesthetics act on
the developing animal; some of them, such as ether and chlore-
tone, producing effects of a general nature, while alcohol and mag-
nesium are more localized or specific in their action. A similar
statement is true for the actions of different anzesthetics on the
adult body. :

In attempting to explain the occurrence of asymmetrically
monophthalmic, cyclopean and blind individuals among’ fish
that had been developed in solutions containing magnesium, the
writer advanced the hypothesis that the anesthetic property of
Meg was the causal factor. Many reasons for such a view were put
forward in a paper on the artificial production of these monsters
(1909). To experimentally test this hypothesis various other
anesthetic agents have been used and all of them to a higher or
lower degree inhibit the development of the optic vesicles in fish
embryos, and thus give rise to various ophthalmic defects. Alco-
hol is most decided in its action, causing in some experiments as
high as 90 to 98 per cent of abnormal eyes, generally cyclopean,
which far surpasses the highest results obtained with Mg.

The effect of alcohol on the general development of the nervous
system is more pronounced than that of Mg, and only a few of the

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10 No. 3.

370 Charles R. Stockard.

alcoholic specimens ever develop sufficiently to hatch and swim
about as do the Mg embryos. An explanation for this may
be that Mg exerts an influence to inhibit dynamic processes, such
as the out-pushing of the optie vesicles, while alcohol acts more
especially on the nervous tissues. Mayer (1908) has shown that
Mg inhibits muscular contractility without affecting in any way
the nervous impulse or nervous rhythm.

The eye defects, it must be remembered, have only been ob-
tained in solutions of one or another anesthetic; the many other
salt and sugar solutions which have been experimented with dur-
ing four years (06 and ’07) have failed entirely to produce similar
results.

The most important outcome of these experiments has been to
prove conclusively that many monsters which occur in nature
may be artificially produced by changing the environment of the
normally developing eggs. The present experiments will demon-
strate that this may be done even after development has pro-
ceeded for some time. ‘These anomalous structures being the
results of external influences and not germinal variations are to
some extent within scientific control. A promising field is thus
opened in the devising of means to control or regulate the devel-
opment of the embryo and possibly to obviate certain monstrous
conditions at least, Such possibilities were of course beyond our
reach if defective germ cells were actually the cause of these mon-
sters.

Mall (08) has brought forward evidence to show that im-
proper placentation or unfavorable developmental environment
is responsible for most human monstrosities, many of which are
aborted before reaching term. There is evidently much need of
investigation aiming toward the control and regulation of the
developmental environment of mammals.

METHOD AND MATERIAL

The eggs of the fish, Fundulus heteroclitus, were used in all of
the experiments. The method of treatment varies somewhat
for the different solutions employed, so that it is best to describe
each separately.

Effeet of Aleohol and Anzsthetics. Syl

Alcohol solutions were prepared in sea- ee on the percentage
basis. The strength used being 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 16,
18 and 20 per cent, 60 ec. of each solition was eae into finger .
bowls and from 60 to 100 eggs, in the early cleavage stages, four
and eight cells, were placed in each bowl. The stronger solu-
tions killed all of the eggs, and those from 3 to 9 per cent gave the
best results. In the 3 per cent alcohol solutions at times as many
as 90 in every 100 embryos showed abnormal conditions of the
eyes, being either eyeless, asymetrically monophthalmic or cyclo-
pean, while in one experiment in a 5 per cent solution of alcohol
in sea-water, there were 146 ophthalmic monsters against only 3
individuals with two separate eyes.

Chloretone of 0.1 per cent and 0.066 per cent in sea-water
caused abnormalities similar to those produced by other anzs-
thetics. This substance is more general in its anesthetic action
than either alcohol or Mg, as will be seen in the discussion to fol-
low.

Ether and chloroform also produce rather general effects on the
developing embryo, yet a small percentage of cyclopean monsters
occur among the embryos which are treated with 60 ec. of sea-
water to which 2,2.5, and 3 ec. of ether has been added. In solu-
tions of chloroform of about the same proportions and slightly
weaker, a few monsters occurred of the type common to the other
anesthetics. Chloroform is rather toxic in its action on these
eggs, large numbers dying in the weaker solutions, while others
are so inhibited in their development, that various abnormal con-
ditions follow.

Eggs were not exposed to any of the above anesthetics for
more than twenty-four or thirty-six hours. They were then
placed in pure sea-water and continued development showing
the abnormal conditions of the eyes and central nervous system
that had been ‘induced by their sojourn in the unusual environ-
ment.

Similar Mg solutions to those formerly employed were again
used. A gram-molecular solution of MgCl, in distilled water was
titrated and kept, to be diluted with sea-water to the proper
strength just before the eggs were placed in it. The most

372 Charles R. Stockard.

favorable results were obtained in solutions of 16, 17, 18, 19, 20,
21 and 22 ce. of molecular MgCl, made up to 60 ce. by the addition
of a sufficient amount of pure sea-water; e.g., 44 cc. of sea-water
was added to the 16 ec. of molecular MgCl, and 43 cc. of sea-water
to the 17 cc. of MgCl.. The solutions are, therefore, ¢0, tb Go»
ete., parts MgCl. to sea-water. In the ¢o solution 66 per cent
of the embryos were cyclopean in one of the experiments. Eggs
were exposed to the action of Mg shortly after fertilization and at
various other times until they reached an early periblast stage, or
were fourteen hours old, all with similar results. Although the
most favorable time for introducing the eggs into Mg solutions
is during the eight-celled stage. The developing embryos were
returned to pure sea-water after the third day. “The Mg is so
slightly toxic that eggs may be kept in it and will continue to
develop; the embryos actually hatch and swim about in the solu-
tion, being, however, slightly slower in their developmental rate
and not so hardy as the specimens which are returned to the sea-
water.

Tue AcTION OF ALCOHOL ON DEVELOPMENT

Weak solutions of alcohol exert a most decided effect on the
_developing fish embryos, causing deformities of the central nervous
system, the eyes, and ears in a very large percentage of the speci-
mens.

a. Defects of the eyes

Typical cases of cyclopia showing in the different specimens
all gradations, from merely closely approximated eyes, hour-glass
eyes with two pupils and two lenses, oval eyes having the two
component intimately associated, typical median cyclopean eyes
with scarcely an indication of their double nature and extremely
small ill-formed cyclopean eyes, were present in the weak alcohol
solutions. All of these have been fully described in a former
‘paper (’09) on the artificial production of cyclopea as a result of
the action of Mg. The alcohol monsters in some cases also
‘present various degrees of the monophalmicum asymmetricum

Effect of Aleohol and Anesthetics on Development. 373

defect which was common in the Meg experiments. Individuals
may have one normal eye and the other eye in different conditions
of arrested, development from slightly small and defective to
entirely absent, see figs. | and 2. An important point that was
brought out by the alcohol monsters which was not noticed in
the magnesium specimens is the fact. that some of the embryos
have both eyes equally small and defective, figs. 3, 8, 9, 10, and 11.
The two eyes are symmetrically defective and the head appears
to have small eye-spots instead of normal eyes; compare figs. 3,
and 7, 9,10 and 11 and 12. Finally, as in Mg solutions so also
in the aleohol, many eyeless individuals are present .

The eye in some of the alcohol monsters is rather different from
that found in the Mg embryos, and may possibly serve to indicate
something of the condition of the eye anlagen in the brain. Many
embryos possess optic cups with their concave surfaces facing
almost directly toward the median sagittal plane of the head. In
life the eye presents a heavily pigmented solid convex surface to
the side of the head instead of the usual open pupil through which
the lens may be seen within the cavity of the optic cup. Fig.
5 and 6 show front and lateral views of such an embryo; the side
view indicates the peculiarly solid convex object seen when looking
towards the lateral eye. Fig. 18 represents a section through
this eye. The choroid coat of the eye-ball is pressed close against
the body wall on the sides of the head and the concave retinal
surfaces which should face outward are turned directly towards
one another; a lens lies between the two eye components which are
really separate except along their dorsal borders. Fig. 4 shows
a somewhat similar specimen in which the eyes are entirely sepa-
rate yet they have an arrangement almost identical to that
just deseribed. The eyes face the mid-plane of the head and
turn their convex choroid coats out against the body wall. The
only place at which the inner face of the eyes touches the ecto-
derm is the ventral body wall and from this a lens has arisen and
lies between the two eyes. Fig. 14 illustrates a section through
these eyes, in which a most peculiar arrangement exists. Optic
fibers probably arising from the ganglionic layer of the retina,
(although this can not be positively demonstrated in the sections),

374 Charles R. Stockard.

CaMERA DrRawineGs orf Livinc FunpuLUS EMBRYOS

Fig. 1. An embryo twenty days old which was treated with5 per cent alcohol.
Only one eye is formed and it faces in a ventro-median direction, instead of to-
wards the lateral wall of the head.

Fie. 2. An asymmetricum nonophthalmicum monster with one almost typical
eye while the other eye is small and poorly formed, from a 4 per cent solution of
alcohol in sea-water.

Fia. 3. An embryo with both eyes small and closely approximated; the eyes
face ventrally. This type is common in all of the anesthetic solutions.

Fic. 4. A nineteen day embryo from 5 per cent alcohol. The two eyes are not
connected yet their convex surfaces are turned out against the lateral walls of the
head and the pupils face the median plane. A single lens lies between the two
eyes. Fig. 14 shows a section of these eyes.

Fic. 5. A nineteen day embryo from 5 per cent alcohol. This front view shows
the two eyes joined dorsally and facing one another with the lens between them.

Fig. 6. shows a lateral view of the same head with the convex choroid surface
of the eye-ball close against the side of the head. Fig. 13 illustrates a section
through these eyes.

Fig. 7. A normal Fundulus embr you when eight Baus old drawn to the same scale
as the monsters.

Fic. 8. A twenty day embryo from 5 per cent alcohol; the eyes are small and
defective as shown in the section fig. 10.

‘

Effect of Alcohol and Anesthetics.

376 Charles R. Stockard.

are collected into optic nerves which pursue extraordinary courses;
instead of passing through the outer retinal layers and the choroid
coat they take an almost directly opposite course and run across
what should be the optic cup cavity (the humor cavity in these
specimens is filled with loose cellular tissue) and out through the
wide-open ‘‘pupil,”’ forming a perfect cross and then passing into
the base of the brain to end in the optic lobes.

The position of the lens must not be supposed to determine the
actual pupil region of these eyes, as the lens clearly lies between
them; see fig. 4 of the living embryo. The wide open pupils of
the two eyes face or lead directly into one another. The position
might be taken that the entire arrangement represents one
large eye; this is not true however since the eyes are entirely
separate in all of the sections and the optic cross could searcely be
expected to exist within the base of the eye itself as would be the
case if this were one huge eye. The eyes really hang down from
the brain as two large retinal disks. Fig. 11 represents a similar
case with the eyes rather more flattened out laterally, and the
existence of all gradations between the two conditions substan-
tiates the above statement.

The retinal layers of these eyes which are nineteen days old,
are poorly differentiated, the inner layer consisting. merely of
indefinitely arranged cells; a better differentiation is usually
attained in normal specimens by the sixth or seventh day. A
comparison of Figs. 13 and 14 with Fig. 12 illustrates in a way the
more definite orientation of the inner retinal cells in the normal
individual when compared with the defective eyes. It will also
be noticed that the lenses in. the two-eyed specimen are surrounded
by clear humor spaces while the lens in the defective eyes lies
buried in loosely arranged cellular tissue.

The right retina of the monster faces in the same direction as
the left retina of the ordinary individual yet there is no indication
of a reversal of the layers which might possibly beimagined in such
a case.

The optic stalk was scarcely formed in these eyes, which is
not infrequently true in cyclopia. The path of the optic
nerve is, therefore, evidently not that usually taken along the

Effect of Aleohol and Anesthetics on Development. 377

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eestsoe

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3 Q 0. 4: 6.08
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Fig.9. A-section through the small defective eyes of an eight day embryo, after
treatment with a4 per cent solution of etherinsea-water. The brainis narrow and
poorly developed. °

Fig. 10. A section through the head of the embryo shown in fig. 8. Both of the
eyes are small and defective with ill-fitting lenses and face in a ventral direction.

Fig. 11. A section through the eyes of a monster commonly found in the alco-
hol solutions. The eyes are joined beneath the bilateral brain and face ventrally.
' Fie. 12. A section through the eyes of a normal thirteen day embryo.

a4
378 Charles R. Stockard.

optic stalk, but the optic nerve fibers grow directly into the brain
from the region of the eye cavity itself.

It is interesting to find in this connection that Lewis (’70)
describes in his experiments on tadpoles a strikingly similar
course pursued by the nerves arising from some of the optic cups
he had transplanted to various positions along the hind brain
region of the embryos. He states that

In a few of these somewhat irregular transplanted eyes the optic nerve
takes a very curious course, passing across the cup cavity from the gang-
lionic layer, through the pupil and then into the mesenchyme, ending
there. In both of these experiments a small bundle of optic nerve fibers
pierces the retina as far as the pigment layer. In transplanting these
eyes the ganglionic layer was probably injured in such a way as to inter-
fere with the normal path of the nerve fibers, and so they have prob-
ably followed the path of least resistance through the pupil and out into
the mesenchyme.

Via. 138. A section of the embryo shown in figs. 5 and 6. The optic cups are
joined dorsally and face the median plane of the head; a lens lies between them
and is surrounded by loose cellular tissue instead of by the humor.

Fic. 14. Section of the eyes in the embryo shown in fig. 4. The optic cups are
not joined yet they face towards one another with their convex choroid surfaces
pressed close against the head wall. The optic nerves run across what should be
the humor chamber of the eyes and out through the wide pupils to form a perfect
cross; the optic fibers then enter the brain floor. A lens lies between the eyes.

Effect of Aleohol and Ansestheties. 379

Lewis’ illustrations are similar to Fig. 14 in so far as the course
of the optic nerve is concerned. These experiments would seem
to indicate that the direction taken by the optic nerve fibers is
not firmly fixed but that they may pursue an almost reverse
direction from that generally followed. Many of the eyes in the
writer’s specimens show various conditions of this kind and he
must agree with Lewis in the conclusion that

It would seem to me impossible to explain these various conditions
of the optic nerve on any other basis than that they are outgrowths of
nerve cells of the ganglionic layer of the retina.

Direct evidence is thus furnished for the outgrowth theory of the
nerve fiber which has been so ably supported in the last few years
by Harrison’s (’08) experiments.

The present experiments warrant the following explanation
for incomplete cyclopean eyes, or double eyes, when compared with
the usual condition.

In normal development the eye anlagen push out from the ven-
tro-lateral borders of the brain and turn dorsally as indicated in
the diagram, fig. 15 A. The abnormal individuals with two
eyes facing the median plane also have them more ventrally
situated in relation to the brain, and it may be supposed that when
the eyes arose from the brain their formation was directed ven-
trally instead of dorsally, fig. 15 B. This causes the eyes to hang
below the brain and face one another as already shown in figs.
4, 5, 6, 13, and 14, instead of turning dorsally and facing outward
as in figs, 11 and 12.

Similar conditions are also found in the development of a single
eye. Fig. 1 shows an embryo with an eye on the right side only,
yet this eye faces the median plane and is unusually ventral in
position; it probably arose as indicated in the diagram fig. 15 D,
where as other single-eyed individuals, the commoner type, have
an eye looking out from the usual lateral position, fig. 15 C.

From these conditions we may determine whether cyclopia is
brought about by a failure of certain central tissues of the brain
to develop, thus allowing the eye anlagen to come together as
Lewis (’09) has suggested, or whether through a lack of develop-
mental energy necessary for the optic cups to grow dorsally and

380 Charles R. Stockard.

outward to meet the ectoderm as the writer (09) has supposed.
Considering the case of the single eye it might be held that the fail-
ure of certain central tissues of the brain to develop would cause
the eye to arise too near the median line, but this lack of central
tissue does not explain why the eye faces the median plane instead
of the lateral head wall, and it is much less able to account for the
absence of the eye on the opposite side of the head. On the other
hand, if the conditions are due to a lack of the necessary develop-
mental energy or an anesthesia produced by the experiment, then
itis evident that although one eye does succeed in pushing out from
the brain it might not have sufficient developmental energy to grow
dorsally and outward to the lateral body wall, but droops, as
it were, into a more ventro-median position and faces in toward
the median plane. Thus one-half of an incomplete eyclopean
eye is formed. The other eye was entirely suppressed, lacking
the energy- necessary to push itself out from the brain. This
inequality in the developmental powers of the two eyes is indi-
cated by their frequent asymmetrical condition.

The two eye components do not always face the median plane
and in such cases the eyes merely fail to grow out laterally. They
come off ventrally from the brain and either face in a ventral
direction or grow so as to face outward.

The experiments fail to give any definite clue as to where the
optic anlagen are located in the brain before they become visible,
although Lewis’ operations on the embryonic shields of older
embryos would seem to indicate that at that time the anlagen
occupied somewhat lateral positions.

It is clear from the foregoing consideration that aleohol has
the power to induce the same typical ophthalmic defects that were
formerly described in the embryos from the Mg solutions. The
property common to both Mg and alcohol is their anesthetic
effect on animals. The writer concludes that cyclopia, mon-
ophthalmicum asymmetricum and entire absence of eyes, all of
which are more or less arrested or inhibited condition of develop-
ment, result from anesthesia during certain embryonic stages.
Of course this may not be the sole cause of such defects; on the
contrary the fact that they are produced in this way would indicate

Effect of Alcohol and Anesthetics on Development. 381

: 15

Fig. 15. A diagram illustrating several positions taken by the optic. cups in
development. A, the usual case in which both cups push out trom the brain and
turn dorsally so as to face the lateral head wall with their convex choroid surfaces
towards the brain; see fig. 12. B, the optic cups push out from the brain but in-
stead of growing Morealie: they fang ventrally and face in towards the median
plane, - with their convex choroid surfaces against the outer head wall; see figs, 13
and 14. Cand D, the same conditions are often realized when only one optic ves-
icle arises from the brain.

382 Charles R. Stockard.

that any factor which might come in during early development
to lower the developmental energy could possibly induce similar
defects. In mammals such monsters probably arise as the result
of some weakening or debilitating influence of the environment
during early developmental stages, which need only have acted
for a short space of time.

b. Defects of the auditory organs

A very pronounced suppression in the development of the audi-
tory apparatus is often noticed in the embryos which have been
treated with weak solutions of alcohol. In many individuals only
one ear exists. When this condition is found in an embryo with
only one eye, two unequally developed eyes or a cyclopean eye
with asymmetrical components, it is of interest to find that in all
cases observed, the ear is on the same side of the head as the better
formed eye. In rare cases both ears are absent, and again it
often happens that the ears are apparently normal while the
eyes are deformed. Fig. 16, a horizontal section through the
head, shows two small abnormal eyes with a lens between them
and two perfect ears with cartilaginous capsules, near the hind
brain. Fig. 18, which is a section through the ear region of the
embryo shown in fig. 4, illustrates two poorly formed ears; on the
right side the ear issmall and two semicircular canals are represented
only by their ampulla, the epithelial lining of which forms papillz
of cells with long hair-like processes growing from them as is
indicated in the drawing. The left ear is almost entirely absent,
its median section showing only the small cavity and ampullary
papilla seen in the figure. . Both ears, however, are surrounded by
well formed cartilaginous capsules.

A remarkably abnormal ear is seen in fig. 17. The auditory
vesicles have united so as to occupy a dorsal position above the
posterior end of the brain. Only two semicircular canals are devel-
oped on each side. The cartilaginous capsules in this case seem
unable to meet the situation and extend for only a portion of the
way around the huge auditory cavity. This union of the lateral
auditory vesicles, although formed by an entirely different princi-
ple, suggests the large double cyclopean eye.

383 Effect of Aleohol and Anesthetics.

The final persistence of the ampulla-like cavities seems to be
the rule, these structures being present even when all other por-
tions of the internal ear are absent. The ampulle of the canals
are perhaps particularly useful as organs of equilibration in these
animals, and their stuborn persistence may be indicative of an
ancient origin and suggests the primary function of the ear as an
organ of equilibrium.

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Fic. 16. A horizontal section through the head of a thirteen day embryo from
a 5.per cent alcohol solution. Two very small and defective eyes have alens be-
tween them, while more posteriorly two perfectly formed auditory vesicles are seen
with cartilaginous capsules surrounding them.

Fic. 17. A section through the auditory region of an eight day embryo treated
with a4 per cent ether solution. The two auditory vesicles unite dorsally to form
a huge cavity above the medulla and spinal cord. This embryo also shows an in-
complete cyclopean eye and spina-befida. ‘sc, semicircular canal; a, auditory
vesicle, ; ph, pharynx.

Fig. 18. A section through the auditory vesicle of the embryo shown in fig. 4
and section fig. 14. The entire auditory vesicle is suppressed except the ampulle
of the semicircular canals. The right ampulla is larger and shows a papilla with
hair-like fibers, while the left-ampulla is almost completely closed, yet, it too, shows
the papilla with projecting hairs. The cartilaginous capsules are small and thus
adjusted to the tiny ear parts.

384. Charles R. Stockard.

Although all parts of the ear are absent the cartilaginous cap-
sules are present. The shape and size of the capsules, however,
seem to be adjusted to that of the auditory vesicle when any part
of it exists, as is indicated on the two sides of fig. 18.

c. Defects of the central nervous system

The abnormalities of the brain shown by the specimens treated
with alcohol might easily form the subject of an extensive mcno-
graph so various and numerous are they. Only a few of them
will be briefly mentioned.

In rare cases the brain is almost normal; the fore brain, however,
is usually very narrow ard gives to the head a characteristically
pointed appearance. Dcrsal hernie at times occur in the region
of the optic lobes and the hind brain. The histological structure
of the brain is often peculiarly abnormal in both the arrangement
and the appearance of the cells. The cells may be hyaline and in
the region of the central cavity fail to take the stains. They may
even be diffusely scattered in peculiarly defective specimens.

The spinal cord in some individuals also shows the hyaline
appearance about its central canal, and spina-bifida in not uncom-
mon. The latter condition no doubt results from the general
inhibition in rate of development which is constantly true for
the specimens in the alcohol solutions. The germ ring is slow in
surrounding the yolk and consequently the trunk region of the
early embryo is abbreviated. This condition interferes with the.
median cell proliferation forming the spinal cord so that a split
or divided cord results and may extend for various distances in
the trunk region. Fig. 19 shows a section through a trunk region
with a double cord, ch, the notocord, nch, is also divided. Fig.
20 is a more posterior section of the same embryo and shows the
cord and notocord again single as they are in more anterior region
than that shown by fig. 19.

Many of the defects of the central nervous system are of a
general nature and almost any substance that inhibits or inter-
feres with the normal developmental rate may cause them. The
writer does not intend to convey the idea that these are characteris-

4 jee

Effect cf Aleohol and Anesthetics on Development. 385

Fic. 19. A section through the trunk region of an eight day embryo from a 4
per cent ethersolution. Thespinal cord, ch, isdouble (spina-bifida) and the noto-
chord, nch, is divided into three parts.

Fie. 20. A more posterior section of the same embryo, showing the spinal cord
and notocord again single as they are in regions more anterior than fig. 19.

THE AMERICAN JOURNAL OF ANATOMY, VOL 10, NO. 3.

386 Charles R. Stockard.

tic anssthetic effects, but they are strikingly common in the
alcohol solutions. On the other hand, similar abnormalities of
the nervous system are really infrequent in the weaker Mg solu-
tions.

The slight effects of Mg on the development of the central
nervous system are interesting when compared with the marked
effects of other anzesthetics on these tissues. In this regard it is
important to remember that in physiological experiments on nerve-
muscle preparations Mayer (’08) has found that Mg acts directly
as an inhibitor of muscular activity, exerting little if any effect on
the activity of the nervous parts. The action of Mg in these
experiments is not particularly on the nervous system but more
largely on the dynamic processes concerned in the outpushing of
the eyes.

The occurrence of the several ophthalmic anomalies is common
to all of the anesthetic solutions and similar conditions have not
been found in embryos treated with various salts and sugars
(06, ’07) which may inhibit general development and induce
many other abnormalities.

THE Errects oF CHLORETONE, ETHER AND CHLOROFORM ON
DEVELOPMENT

Chloretone, ether and chloroform when employed in weak
solutions influence developing eggs in a way somewhat similar
to that described for alcohol. The action of these substances is
not so pronounced and may be described more as an inhibition of
the general developmental processes. The embryos are usually
small and recover very slowly from the inhibiting effects after
being returned to sea-water. A few of the individuals in all
these solutions exhibit the cyclopean defect in its various degrees
just as it has been described in the specimens treated with alcohol
and Mg.

The ophthalmic defects, cyclopia, monophthalmicum asymme-
tricum and entire absence of optic vesicles, are all conditions of
arrested or inhibited development and are prevalent among
embryos treated with solutions having anesthetic properties.

Effect of Aleohol and Anzsthetics. 387

The writer is led to conclude, therefore, that his former hypo-
thetical explanation of why cyclopia occurred in embryos treated
with Mg solutions was correct. The evidence strongly indicates
that the ophthalmic abnormalities produced in these experiments
are the result of an anzsthetic action during the early develop-
mental stages.

THE PERIOD OF DEVELOPMENT AT WHICH CYCLOPIA MAY BE IN-
DUCED BY CHEMICAL AGENTS

The Mg experiments were repeated mainly to ascertain at
how late a period in development eggs could be subjected to the
solutions and subsequently develop into cyclopean monsters.
The original experiments seemed to demonstrate the fact that
eyclopia was due to external influences acting on the development
of the optic vesicles, and not in any sense to a germinal variation.
Nevertheless, H. H. Wilder (08) in the face of these results,
advanced a germinal theory to account for the origin of cyclopia
and attempted to explain away the obstacle offered in the experi-
ments referred to by claiming that the eggs were subjected to the so-
lutions at so early a stage in development that germinal variations
might still have been induced. This is impossible, asthe writer has
pointed out elsewhere (’09b), since germinal variations may only
be induced before embryonic development has begun. After
the two-or four-cell stage (the time at which the eggs were sub-
jected to Mg) is reached any thing done to the egg has its effect
on the developing embryo to cause this or that abnormal condition.
The only germinal variation possible at such a period would be in
the primordial germ cells of the developing individual, a variation
which would not manifest itself until the next generation of
individuals.

The following experiments prove beyond doubt that cyclopia
may be produced by the action of environmental influences.

When eggs in the eight-cell stage, or three and one-half hours
after fertilization, are subjected to the action of Mg solutions
many of the resulting embryos will show the cyclopean defect.

388 Charles R. Stockard.

If eggs be placed in Mg solutions five hours after fertilization,
when in the sixteen or thirty-two-cell stage, an almost equally
large number of cyclopean individuals will occur. The same is
true when they are subjected to the solutions after having devel-
oped in pure sea-water for seven and one quarter hours and reached
the sixty-four or higher cell stages.

Eggs that have developed eleven hours in pure sea-water and
are in the early periblast stage with a somewhat flattened blast-
odermic cap, may be put into Mg solutions and caused to form
cyclopean monsters. The percentage of cyclopean embryos
arising from eggs treated at this late period is small, yet even after
developing for fifteen hours in pure sea-water some eggs may be
induced to form cyclopean monsters by treatment with MgCl, in
sea-water solutions.

Eggs that were older than this before being introduced into the
solutions failed to respond, all developing into ordinary two-
eyed individuals. This is due to the fact that a considerable
amount of time is necessary for the Mg to act-upon the body sub-
stances of the early embryo and prevent normal eye developnfent.
The optic vesicles begin to push out from the brain: before the
thirtieth hour in development. Thus, after the first six or seven
hours, the longer the eggs have been allowed to develop naturally
the smaller the proportion of cyclopean individuals that may be
artificially induced. After fifteen hours no embryo will be so
affected, since an insufficient amount of time exists for the Mg to
act on the eye anlagen.

The solutions are effective up to a stage in development pre-
ceding the formation of the germ ring and embryonic shield, and
the action of the Mg on the eye anlagen probably takes place
while the embryonic shield and outline of the embryo are forming.

There can be no further doubt that cyclopean monsters are
caused by the action of a strange environment on the developing
fish embryos. With such evidence at hand it is also highly
probable that mammalian cyclops are due to the action of external
influences on the embryo and not to an abnormal germinal tend-
ency.

Effect of Aleohol and Anesthetics on Development. 389

OTHER CASES OF CYCLOPIA IN FISH

The Italian observer, Paolucci (’74), has described a most
remarkable cyclopean ray. The monster was almost adult in
size, probably two years old, and measured 47cm. across the
pectorial fins and 20cm. in length not including the long whip-
liketail. Paolucci states that this cyclopean monster was captured
in the Adriatic Sea near the shore and had evidently been able
to cope with its surroundings and grow into a vigorous ray. So
far as the writer knows this case of a eyclopean monster in nature
being able to sustain itself and reach the adult stage, is unique.

Paolucei’s specimen proves the correctness of the writer’s
statement (’09) that the cyclopean eye is not necessarily associ-
ated with a single instead of a double brain, or with any other
serious defect in the brain region. This fact was clearly shown in
the brain structure of many of the cyclopean embryos studied,
as well as by their apparently normal behavior after hatching

‘from the egg.

The cyclopean Funduli have been kept living for more than one
month, which jis as long as the experiment was tried. They would
doubtless have lived much longer, as they were hardy and able
to obtain an abundance of food from the vegetable particles in
the sea-water. Paolucci’s observation would indicate that. the
Fundulus monsters might be reared to maturity and possibly
interbreed.

Gemmel (’06) has described four cases of cyclopia in newly
hatched trout collected from a fish-hatchery in England. The
conditions of the eyes and brains in these monsters are exactly
similar to those in the artificially produced Fundulus monsters.

The developing trout’s egg demands water of such high purity
that trouble is often experienced in the hatcheries, and monstrous
embryos commonly occur. These may result from weakened
developmental forces due to an insufficient oxygen supply or to
the accumulation of injurious chemicals about the eggs.

Gemmel (’06b) in describing cases of supernumerary eyes in the
trout embryos records that in one case of an aborted twin head
the lens alone of all the eye structures was present. Free lenses

390 Charles R. Stockard.

were also described and figured in other individuals. Free lenses
occur very commonly in heads showing various eye abnormalites;
a full consideration of these cases is recorded elsewhere.

SUMMARY

1. When the eggs of the fish, Fundulus heteroclitus, are subjected
during early stages of development to the action of weak solutions
of alcohol the resulting embryos show marked abnormalities in
the structure of their central nervous system and organs of special
sense.

The eyes in such individuals are either both small with poorly
differentiated retine, cyclopean, asymmetrically monophthal-
mic or entirely absent. These ophthalmic defects sometimes
occur in as many as 98 per cent of the specimens. Such anomalies
are closely similar to those previously induced with Mg, and in
both cases are probably due to the anesthetic property of the
substances acting upon the eggs.

Alcohol tends to suppress the development and differentiation
of the auditory vesicles. A few specimens are entirely without
ears, others have one ear more or less perfectly developed while
the opposite ear is scarcely formed at all and still other individuals
have both ears extremely defective. In all cases examined the.
better ear is invariably on the side with the better developed eye
if the eyes are also asymmetrically formed. The most persistent
portion of the internal ear, or that part which exists when all
other parts are wanting, is a cavity with an epithelial lining
resembling closely in structure an ampulla of the semicircular
canals. This fact may be interpreted to mean that the ampulla
is one of the most ancient or fundamental parts of the ear, and it
might further be considered indicative of the archaic function of
the ear as an organ of equilibrium since this is the chief function
of the ampulle.

The brain is usually narrow and pointed in embryos that have
been treated with alcohol. It occasionally has a dorsal hernia
and shows regions of poor differentiation. The cell arrangement
jn the spinal cord are abnormal in many cases and spina-bifida

Effect of Alcohol and Ansestheties. 391

is not infrequent. These conditions of the central nervous sys-
tem might result from any cause that tends to retard development
and are not particularly characteristic of anesthetic solutions
as the eye anomalies are; yet the defects of the central nervous
system are commoner in these anesthetics than in any other solu-
tions with which the embryos have been treated.

2. Chloretone, chloroform and ether induce much the same
structural deformities in these embryos as does alcohol. They
act, however, as more general anesthetics, causing a retardation
in development. The characteristic eye and ear defects are not
nearly so common, though they do occur as a result of treatment
with these three substances.

3. The effects of Mg on the developing fish’s egg have been
previously considered. This substance is even more local in its
action than alcohol, the principal defects resulting from its use
being various anomalous conditions of the eyes, whereas the ner-
vous system generally may be in many cases structurally normal.
The embryos on hatching from the egg are able to swim in the
usual manner and live for more than one month in aquaria, which
is as long as any effort was made to keep them. The latter fact
would seem to indicate that the nervous system also functionates
normally.

Magnesium was used to test at how late a period in develop
ment the eggs might be introduced into the solutions with the sub-
sequent development of the cyclopean condition. It was found
that after normal development had proceeded for two, four, six,
eight, ten, eleven, twelve or even fifteen hours, if the eggs were
then placed in MgCl,, solutions, many of the resultiny embryos
showed the cyclopean defect. At fifteen hours the eggs have
reached the periblast stage and the blastodem is flattening down
upon the yolk. The germ ring arises shortly after this time and
begins its downward growth over the yolk mass.

Whenever eggs are allowed to develop beyond the fifteen hour
period before being introduced into the solutions of MgCl, they
invariably give rise to normal two-eyed individuals. The occur-
rence of cyclopia is less frequent when eggs are subjected at later
stages than when introduced into the MgCl, solutions during the
four or eight-cell stage. This is doubtless due to the fact that a

. 392 Charles R. Stockard.

considerable period of time is necessary for the Mg to act upon the
substances of the embryo and influence the origin of the optic
vesicles. When an insufficient time intervenes between the
period at which the eggs are subjected to the action of the solu-
tion and that at which the optic vesicles are given off from the
brain the Mg is unable to influence the tissues so as to induce the
eyclopean condition.

The production of cyclopia by the action of Mg at such late
stages in development proves beyond doubt that this deformity
is due to the action of external or environmental conditions on
the developing animal. Any explanation of cyclopia based on
germinal hypothyses such as that recently advanced by H. H.
Wilder must: be reconstructed so as to conform to these facts.

Accepted by the Wistar Institute of Anatomy and Biology January 12, 1910. Printed June 21, 1910.

BIBLIOGRAPHY

GeEMmMEL, J. F. On cyclopia in osseous fishes. Proc. London Zodél. Society, i, pp.
1906 443-449,

1906b Notes on supernumerary eyes, and local deficiency and reduplication

of the notocord in trout embryos. Proc. London Zoél. Society, 1,

pp. 449-452.
Harrison, R. G. Embryonic transplantation and the development of the ner-
1908 vous system. Anat. Record, i, pp. 385-410.
Lewis, W. H. Experimental evidence in support of the theory of outgrowth
1907 of the axis cylinder. Am. Jour. Anat. vi, pp. 461-472.
1909 The experimental production of cyclopia in the fish embryo (Fun-

dulus heteroclitus). Anat. Record, iii, pp. 175-181.
Mau, F. P. A study of the causes underlying the origin of human monsters.

1908 Jour. Morph. xix, pp. 1-861.
Mayer, A. G. Rhythmical pulsation in seyphomedusae. Carnegie Institute of
1908 Washington, Pub. No. 102, pp. 118-131.
Paoxuuccr, L. Sopra una Forma Mostruosa dela Myliobatis Noctula. Atti della
1874. societa Italiana di Sc. Naturali. xvii, pp. 60-63.
SrockarD, C. R. The development of Fundulus heteroclitus in solutions of lith-
1906. ium chlorid, with appendix onits development in fresh water. Jour.
Exp. Zo@l., iii, pp. 99-120.
1907 The influence of external factors, chemical and physical, on,the
development of Fundulus heteroclitus. Jour. Exp. Zodl., iv; pp.
165-201.
1909 The development of artificially produced cyclopean fish, ‘the mag-

nesiumembryo.’’ Jour. Exp. Zodél., vi, pp. 285-338.
1909b The origin of certain types of monsters. Am. Jour. Obstetrics.
lix, no. 4.
Witprr, H. H. The morphology of cosmobia. Am. Jour. Anat., viii.pp. 355-
1908 440. i

THE INDEPENDENT ORIGIN AND DEVELOPMENT
OF THE CRYSTALLINE LENS

CHARLES R. STOCKARD

From the Department of Anatomy, Cornell Medical School, New York City

WITH TWENTY-EIGHT TEXT FIGURES AND TWO PLATES

RATE OCMLCRT Ets Ser ere mae ates te. ees CN ae Dye Gey nd is Geer Go Toe S898
Bae Vie hirer Gertaloner. tecyates wale gute ania he se ae ee bok bei oe 395
PEROT MN Me IMM MON awe Aoi... neice actin Stee beeen paca heals 397
Het ryOs SOLE CaM SOCOM sinicion ac qe fed Mac eee ces Pk eee ae bike 400

a Is the origin of the lens from the ectoderm dependent upon a contact
RGUMIMUSrOlet Me OPO ERVESTCIEL 2 ae <)! give he hres echelons Salt Wek ve cus enue 400

b Is the lens-plate or lens-bud capable of differentiating into a lens with-
Ouncontachauithan optic Cp? e .a.att. oe: ence Sees oleh cae ee 401

¢ Does the size of the optic cup regulate the size or shape of its associated
EIST Oh ae ie este ROSE PICS Ry PRA AR ee ee ee 404

d Is the optic vesicle, normal or defective, always capable of stimulating
lens formation from the ectoderm at some stage of its development? ... 405

e May the optic vesicle cause lens-formation from ectoderm other than
Pai Wiktela nOnM ally GrMs A LeWS! cee wacked «hobs vals 2e0 au owed aus o heeed 4 408

f Does a deeply buried eye have the power to regenerate or form a lens
RCOMIBUG AS WINGEISSUCH te chee ote, wenetie 2 oS FSR hagas Ae eee Be 410
5 Discussion of previous observations and experiments..................... 412
He ouaniet ya COMCINISION Rr: 6 s..o RUS 4 foe ule oh a ine 5 nv ose «see oars 419
ane SUL Opt ea Pinyin connie el L Pinyin C8. (Shee ai Se oe cl ky 421

INTRODUCTION

The crystalline lens normally arises in the embryo from ectoderm
overlying the optic vesicle and continues its development in
close association with the optic cup. This fact suggests that
some correlation exists between the manner of development of
the optic cup and the optic lens. In case such a correlation does
exist, to what extent is the optic vesicle and cup responsible for
the origin and subsequent development of the lens; and, on the
other hand, what influence, if any, does the presence of the lens

i

394 Charles R. Stockard.

exert over the development of the optic vesicle into the cup and
its subsequent development into the eye?

If it is proven that the optic vesicle and cup possess the power
to derive a lens from the ectoderm the further question then
arises is the ectoderm under any condition able to give rise to a
lens without the optic vesicle stimulus? If so, is this independent
lens-bud capable of self-differentiation to such an extent as to
form a perfectly constructed lens?

Many questions of detail, as, for example, the relationship
between the size of the optic cup amd the size of the lens, the
ability of partial or defective optic cups to stimulate lens forma-
tion, whether the lens may be derived only from certain areas
of ectoderm or from any ectoderm, and other problems which we
will presently consider, also present themselves.

The study of these questions has come to be known as the
lens problem. Experimenters have attacked the questions from
several sides during the last ten years and the lens problem has
in many ways been beautifully analysed, yet today much addi-
tional evidence is necessary to entirely clear the situation.

In a previous paper the writer (’09) briefly described the inde-
pendent origin and self-differentiation of the optic lens in the
fish embryo. Since that time further experiments have yielded
much additional material and evidence bearing on this subject.
The results of these experiments are considered in the present
contribution and show in a most convincing manner that the
crystalline lens is capable of originating independently from the
ectoderm and of subsequent self-differentiation. Definite proof
will also show that although the lens may arise independently,
nevertheless the optic vesicle invariably has the power to stimu-
late the formation of a lens from any overlying ectoderm with
which it may come in contact.

The action of the optic vesicle on the ectoderm is a much
stronger force for the production of a lens than is the innate
tendency of the ectoderm to produceanindependentlens. Slightly
injured ectoderm may be rendered unable to form a free lens
while the same weakened ectoderm will respond to a contact

Independent Development of the Lens. 395

stimulus of the optic vesicle by forming a lens. This point is
important, for herein the writer believes, after a study of his
own experiments and the results of other workers, lies the explana-
tion of many discrepancies in the operation experiments on the
lens. When the operation is so performed that the ectoderm
must be folded away in order to extirpate the optic vesicle and is
then returned to its place, free lenses have failed to occur, although
an optic vesicle may still have been able to derive a lens from
this replaced ectoderm. On the other hand, when the early open
medullary plate is operated on so as to remove the optic vesicle
areas, the ectoderm of the head wall is sometimes left uninjured
and from it may arise free lenses. The free lenses of King’s
experiments arose in embryos which were operated on dorsally
to burn out the optic vesicle areas of the partially open medullary
tube. The lateral head ectoderm was probably uninjured in
some of the specimens. In Spemann’s more recent experiments
the early open medullary plate was operated upon directly to
remove the optic vesicle areas; in such experiments free lenses
arose from the uninjured ectoderm. Experiments on other spe-
cies at such stages and in a similar manner will probably give like
results.

In the experiment of removing the optic cup and leaving a par-
tially differentiatied lens, this lens may have degenerated or
ceased to differentiate on account of the injury it suffered by the
operation, the absence of the optic cup not affecting it. It is
unquestionably true that in Some amphibians and fishes the lens
is capable of perfect self-differentiation.

The optic cup does not exert complete control over either the
size or shape of the optic lens. Numerous points of detail are
also elucidated by the study of the optic organs in artificially
produced fish monsters which are either blind or present various
eye defects.

The experimental part of this investigation was conducted dur-
ing the summer of 1909 in the Marine Bioligical Laboratory at
Woods Hole, Mass., while occupying one of the rooms of the
Wistar Institute.

396 Charles R. Stoekard.

2 METHOD AND MATERIAL

In all former experiments on the developing lens, except
those which the writer recorded (’09), mechanical methods have
been resorted to in destroying the early optic vesicle. This has
been accomplished by burning the region with hot needles or by
cutting away the tissue. It matters not how cleverly such
experiments may be performed they are often open to objection,
particularly when the experimenter has to remove or injure the
overlying ectoderm in order to reach and extirpate the optic
vesicle.

The present experiments have been conducted in an entirely
different manner. When developing fish embryos, Fundulus
heteroclitus, are treated with certain magnesium salts, alcohol,
chloretone or other anesthetic agents, the development of the
optic vesicles is prevented entirely in some cases, while in other
specimens only one vesicle forms on either the right or left side,
and finally a large majority of the embryos present the eyclopean
defect with a more or less double ventro-median eye. We have
here, therefore, an exceptidal opportunity to study the relation-
ship between the development of an optic vesicle and a lens. In
the first case, does a lens or do lenses ever occur in the eyeless
specimens? Does a lens ever appear on the eyeless side in the
single-eyed monsters? Finally do lenses ever arise in their usual
lateral positions when the embryo has a ventro-median cyclopean
eye? Allof these propositions are affirmatively answered without
an operation to injure in any way the ectoderm or tissues in the
primary lens-forming region. It may be thought that the action
of the chemical or anesthetic is as severe as an operation but
this is probably not true, as the embryos after being treated for
the necessary time with magnesium, on being returned to sea-
water develop, hatch and swim actively about, living in aquaria
as long as I have tried to keep them, more than one month.

The experiments of the past summer have convinced the writer
that his former idea that the eye defects are due to the anesthetic
properties of magnesium is correct; and there is no reason for
believing that certain tissues usually between the eyes are entirely

Independent Development of the Lens. 397

absent. These results are given in full in another paper. The
solutions employed act as anesthetics preventing the usual out-
pushing of the optic vesicles from the brain to a greater or less
degree, and give exactly the same condition so far as contact
influence of the optic vesicle on the lens is concerned, as though
the optic vesicle was cut away, without the disadvantages
accompanying the operation.

The experiments with anesthetics furnish a richness of material,
hundreds of specimens being obtained, which one would be unable
to duplicate from operations without spending days of tedious
labor. The crystalline lenses may be seen with the binocular
microscope as spherical refractive bodies in the living specimens.
The experimenter in this way is enabled to select various condi-
tion for study.

The embryos were best preserved for histological study in
picro-acetic though many fixitives gave good results. The lenses
stain equally well in eosin or picric acid used as a counter stain
after Mann’s hematein or Delafield’s heematoxylin.

3 THE LENS IN LIVING EMBRYOS

The living embryos at various stages when examined with a
binocular microscope show lenses isolated entirely from the optic
vesicle or optic cup. Figure 1 illustrates an embryo eighteen
days old that has no trace whatever of optic cups yet two per-
fectly developed transparent lenses occupy almost normal posi-
tions on the sides of the head. Sections of this specimen show the
two lenses with well differentiated fibers, fig. 5 and plate II,
fig. 3. _Mesenchymatous tissue lies between the lenses and the
brain. We must conclude that these lenses arose in lateral
positions and continued development and differentiation with no
direct influence whatever from either optic vesicle or the brain
tissue. The possibility of optic cups having arisen and degener-
ated is entirely out of the question, since, in the first place, this
has never been known to occur in any of the hundreds of Fundulus
embryos that the writer has studied. In the second place, plate I,
fig. 1, shows a lens arising from ectoderm on the eyeless side of

398 Charles R. Stockard.

a seventy-six hour embryo; it is scarcely conceivable that an
optic vesicle arose at about the thirty-fifth or fortieth hour,
came in contact with the ectoderm and entirely disappeared,
leaving no trace of itself by the seventy-sixth hour. Further,
the lens in these cases must continue to develop and differentiate
independently, which is contrary to the position held by Spemann
(05), Lewis (07) and LeCron (’07) for the frog and salamander.

Fig. 2 shows a fish nineteen days old with two well formed
lenses of normal size lying in contact with two small irregularly
shaped masses of heavy black pigment. In sections, figs. 23 and
24, the lenses are found to be perfectly formed and the two dark
spots are shown to be masses of choroid tissue or retinal pigment
without a definite retina or other associated eye parts. The lenses
may owe their existence to the choroid spots but the latter have
failed to influence the size or manner of development of the former.
If the origin of these lenses was due to the influence of the choroid
areas we have a striking example of how very small an amount of
optic tissue may call forth a lens. Many instances will be given
to show that extremely small amounts of optic tissue touching
the ectoderm will stimulate a lens to form, yet these cases will in
no way weaken the evidence that lenses do at other times form
entirely independently.

A fish is illustrated by fig. 38 with two small defective eyes
deeply buried in the head. The right eye possesses a lens but
the left faces ventrally into a mass of mesenchyme and is without
a lens. In front of the left eye is shown a lens in an extremely
anterior position but completely separated from the eye, and the
concave surface of the cup is not directed towards this lens. Fig.
4 proves the case by showing two slightly small eyeseach possess-
ing its own lens, while somewhat in front and between the two
eyes lies a perfectly isolated and independent lens. It is evident
that this supernumerary lens is independently formed and not
due to a stimulus trom either of the eyes, since each has its
own. lens.

Independent Development of the Lens. 399

CamerA Drawines or Livinc Funputus Empryos, Maeniriep 16 Times

Fic. 1 An embryo eighteen days old. First thirty-six hours after fertiliza-
tion were spent in7 per cent alcohol in sea-water. No optic cups formed, yet two
perfect crystalline lenses are shown in the sides of the head. See fig. 5 for a sec-
tion of these lenses.

Fig. 2. Anembryo nineteen days old, first thirty-six hours in 9 per cent alcohol.
Two perfect lenses near poorly formed eyelike choroid spots. See figs. 23 and 24
for sections of these lenses and eye spots.

Fic. 3 Anembryo of same age and lot as fig. 2; a free lens is seen in an extremely
anterior position in front of anill-formed eye. Lens. L.

Fra. 4 Another embryo of the same lot, showing a free lens between two some-
what defective eyes, each of whichcontainsitsownlens. Sections of thisembryo,
figs. 9 and 10, show also a second free lens which was hidden in the living specimen.

400 Charles R. Stockard.

4 THE EMBRYOS STUDIED IN SECTION

a Is the Origin of the Lens from the Ectoderm Dependent upon a
Contact Stimulus from the Optic Vesicle?

Spemann (’01) and Herbst (’01) first introduced the view that
the lens originates from ectoderm only when a contact stimulus
from the optic vesicle is present, although Spemann *(’07) has
since modified his opinion for one species of frog at least. Lewis
(04 and ’07) has found this to be true in his experiments on frog
tadpoles, and LeCron (’07) on salamanders. King (05) claims,
however, that such is not the case in her experiments and holds
the view that the lens arises independently of the contact stimulus
by the optic cup. Lewis (07) has brought objections to the
method employed by King and so criticises her results, but the
writer believes her method has a real advantage in that she burnt
out the optic vesicle areas of the still open medullary tube from the
dorsal side and thus may not always have injured the lateral
ectoderm of the future lens-forming region.

The writer’s experiments on the fish embryos clearly demon-
strate that the origin of the lens from the ectoderm may be entirely
independent of the contact stimulus of the optic vesicle. He
continues to use the expression ‘‘contact stimulus of the optic
vesicle” since this is what has been deemed necessary for the
origin of the lens, although he believes that a lens may arise with-
out any stimulus whatever from the optic vesicle either by con-
tact or from a distance.

In specimens lacking .optic vesicles entirely it is difficult to
imagine that tissues are present in the brain which possess the
power to form substances characteristically formed by optic vesicles
and that these substances diffuse until they reach the ectoderm and
stimulate it to formalens. In the case of isolated supernumerary
lenses the optic cups possess lenses but still other lenses arise at
a distance.

Again referring to fig. 1 of plate I, a section through the eye
region of a seventy-stx hour embryo, the ectoderm on the eyeless
side is forming alens which is somewhat slower in development

Independent Development of the Lens. 401

than the lens in the eye on the other side, yet this bud shows dis-
tinct lens character.

A perfect lens is seen in fig. 2, plate I, to be entirely separated
from the brain and no optic cup exists. Fig. 13 shows a lens in
a small choroid cup and a second free lens lying near. Fig 7
illustrates a similar case. Figs 8 and 12 show extremely anterior
lensesin eyeless individuals and again fig. 5and fig. 3, plate II, show
two beautiful lateral lensesin another eyelessspecimen. Fig.6shows
a section with three well differentiated lenses all free from contact
with an optic vesicle; a more posterior section of this embryo,
fig. 27 shows a choroid cup deeply buriedinbrain tissue and with-
out alens. This cup does not come in contact with either of the
three lenses shown in.the more anterior sections.

Finally, a most remarkable case of supernumerary lenses is
illustrated by figs. 9 and 10 and the outline fig. 11 shows the
position of these lenses in the entire head (see also plate II,
fig. 4). Two defective eyes each possessing a lens areshown in sec-
tion, fig. 10, and a third lens lies between the eyes. In a more anter-
ior region, fig. 9 and plate IJ, fig. 4, is found another section of this
third lens, C, and a fourth additional protruding lens lies below it.

These gases might be enumerated and illustrated until they ran
into the scores, but sufficient evidence has been given to prove
that the crystalline lens in these embryos does not depend upon
a contact stimulus of the optic vesicle for its origin from the ecto-
derm but originates independently.

6 Is the Lens-Plate or Lens-Bud Capable of Differentiating into a
Lens without Contact with the Optic Cup?

The above question is convincingly answered in the affirmative
by the evidence given in the foregoing discussion. In the older
embryos it is clearly shown that supernumerary lenses are as
highly differentiated and as perfectly formed in all respects as
are the lenses in the eyes. LEyeless individuals, as figs. 1 and 5
and plate II, fig. 3, indicate, may possess perfectly formed trans-
parent lenses which appear in the living specimen as clear refrac-
tive bodies.

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 3.

402 Charles R. Stockard.

Sections oF WruLuL DIFFERENTIATED FREE LENSES

Fiag. 5 A-section of two free lateral lenses in the nineteen day embryo shown by
fig. 1.; no trace of optic cups exists. This embryo has two ears of unequal size.

Fria. 6 Section of the head of nineteen day embryo, first thirty-six hours in 9
per cent alcohol. Three free lenses, A. B. and C are shown, a defective optic cup
completely separated from the lenses is deeply buried in the head tissues of a more
posterior region, see fig. 27.

Fic. 7 Section of the anterior tip of the head of a nineteen day embryo from 9
per cent alcohol. The upper right lens protrudes from a defective eye shown in
more posterior sections, while the lower lens, F’., is free, being in no way associated
with an eye.

Fic. 8 A somewhat sagittal section of a similarly treated embryo of same age,
showing another free lens, p, a pigment spot.

Fic. 9 An anterior and fig. 10 a more posterior section through the eye region
of a nineteen day embryo treated with alcohol. The diagram, fig. 11, shows the
plane of both sections. Fig. 4 shows the same embryo from life; the eyes in the
sections are reversed by the microscope. Two optic cups are present each with a
lens, A. and D, while two other perfectly differentiated lenses, B and C, are not
connected with an optic part.

Fria. 12. Asection through the anterior tip of a pointed-headed eyeless embryo
nineteen days old. The lens is well differentiated; the ears in this specimen are
scarcely formed.

Fra. 13. A section of a nineteen day embryo showing a small defective choroid
cup with a lens, and a second-accessory lens is near by.

403

Independent Development of the Lens

404 Charles R. Stockard.

The possibility of the action of some substance given off by a
distant optic cup is entirely aside, since other experimenters have
claimed that when the optic vesicle or cup is In any way separated
from the lens the latter organ begins to degenerate and usually
disappears.

Figs. 6, 7, 9, 10, 12, 13 and 21 and plate I, fig. 2, plate II,
figs. 3 and 4, all go to show that in the fish embryo the lens-plate
or lens-bud is capable of self-differentiation, finally forming a per-
fectly transparent refractive body even though completely isolated
from any other eye-like structure.

¢ Does the Size of the Optic Cup Regulate the Size or Shape of its
Associated Lens?

Lewis (’07) has stated in his more recent paper on the lens that,
“The lens is neither self originating nor self differentiating, but
is dependent for its origin, its size, its differentiation and its
growth on the influence of the eye.”? The writer had also inde-
pendently been led to think from his first experimental study of
eyclopia (’07), which was based on a limited supply of material,
that the size of the lens varied directly with the size of the optic
cup. He is now able to show that while normlly the lens and
optic cup are properly adjusted as to size this is not by any means
constantly true of ill-formed eyes. Here the size and also the
shape of the lens is often greatly out of accord with that of the
optic cup. In normal eyes the optic cup has a definite size and
so does the lens. The sizes accord, yet this may be incidental or
entirely without correlation, as is suggested by the fact that optic
cups of unusual shape and sizeare not able to regulate the develop-
ment of the lens so as to adjust it to their strange proportions.

Many of the illustrations of cyclopean eyes given in the writer’s
recent paper (’09) show misfits between the cups and lenses.
Remarkable cases are also shown by figs. 9 and 10 and plate II,
figs. 4 and 5, in which the lenses are clearly too large for the asso-
’ ciated cups. Fig. 15 shows the two components of an incomplete
cyclopean eye with one normally proportioned lens between them.
This lens is scarcely large enough to function with the unusually

Independent Development of the Lens. 405

wide double cup. In fig. 20 is seen a somewhat similar double
cup with an elongated and slightly constricted lens; fig. 18 also
shows one half of a double cyclopean eye with a double lens,
the other half eye isin amore posterior section. In figs. 14 and 17,
on the other hand, we find illustrated double lenses in one of
two closely approximated eyes. Fig. 14 also shows a tiny addi-
tional lens still further within the same cup which possesses the
large double lens. Fig. 16 shows the extreme anterior tip of a
cyclopean eye with two minute lenses protruding from it. This
section is only fifty micromillimeters from the anterior end of the
head. Numerous other examples of misfitting lenses might easily
be given.

These facts force us to conclude that the size of the optic cup
does not fully regulate either the size or shape of the associated
lens. It is, therefore, evident that the usual harmonious adjust-
ment between the optic cup and the lens may not be so entirely
due to a dominating influence of the optic cup on the lens as one
might be led to believe from previous contributions to the sub-
ject. That some influence or interaction exists; the writer does
not deny, and will show in the following parts of this paper the
remarkable ability possessed by the optic vesicle to obtain a lens
from any part of the ectoderm with which it may come in contact.

d Is the Optic Vesicle, Normal or Defective, always Capable of
Stimulating Lens-Formation from the Ectoderm at Some Stage
of its Development?

Of all the embryos which the writer has examined not one failed
to have a lens in a normal optic cup when the cup came in con-
tact with the ectoderm. If, however, the cup fails to reach the
outer body wall, although it may possess well differentiated reti-
nal layers and other parts, it is invariably without a lens, fig. 26.
The convex side of the choroid coat or pigment layer of the retina
does not cause a lens to arise even though it be closely applied to
the ectoderm, as is shown by fig. 26 and many other illustrations
in which the choroid touches the body wall.

Defective optic cups when deeply buried and separatea from

406 Charles R. Stockard.

ILL-ADJUSTMENT OF Optic Curs AND LENSES IN THIRTEEN Day FisH EmBryos.

Fic. 14 A section of an embryo treated with 5 per cent alcohol. Both optic
cups are defective, the left one contains a spherical lens, while the right cup has
a large lens, in shape a constricted oval, and a tiny spherical lens placed fur-’
ther within the same cup. E, ear.

Fic. 15 <A poorly formed eye of the incomplete cylopean type. Each com-
ponent faces in a ventro-median direction and a spherical lens hes between them
An ear, E. is shown on the side of the better component, the other ear is absent.

Fig. 16 Asection 50 microns from the anterior tip of anembryo. The anterior
border of a cyclopean eye is shown with two protruding lenses of very minute size.
ec, ectoderm.

Fic. 17 Section through the anterior tips of two small closely neighboring
eyes, Whose median planes are in more posterior sections. The smaller eye has a
protruding double lens. This embryo possesses only one ear located on the side
with the better eye.

Fic. 18 Part of an incomplete cyclopean eye (other half in more posterior
sections) containing a double lens.

Fic. 19 Two adjacent eyes facing ventro-medianly with two lenses.

Fic. 20 An ovoid lens in an incomplete cyclopean eye.

Independent Development of the Len

A407

S

408 Charles R. Stockard.

the ectoderm also lack a lens, as is illustrated on the left of figs.
27 and 28. On the other hand, it is remarkable how small and
ill-formed an optic cup-like structure has the power of stimulating
a lens to arise from the ectoderm. Figs. 2, 23 and 24 and plate
II, fig. 5, show small choroid cups with no retinal differentiation
whatever, yet closely associated with perfectly formed lenses. In
fig. 21 is seen an extremely defective cup with a small lens; a
larger independent lens is shown on the eyeless side. Fig. 22
illustrates an extremely insignificant eye-like body buried within
the brain, yet close by is a small crystalline lens; these are the
only eye parts found in thisembryo. In fig. 19, a section through
the eye of an incomplete cyclops, each component of the eye has
a lens, while in fig. 20 the eye components are closer together and
in fig. 15 further apart, yet each of these possesses only a single
lens, although it is elongate in fig. 20. It is difficult to say why
such eyes occasionally possess two lenses. After an examination
of a large number of such eyes no general rule is found. It may
be due in some way to the manner in which the optic cup periph-
ery meets the ectoderm, whether as a circle, an oval or at times
a much constricted oval so that two areas of ectoderm are sepa-
rately stimulated to form lenses.

This consideration forces the conclusion that an optic cup at
some stage in its development, whether normal or defective,
invariably possesses the power to stimulate lens-formation from
the ectoderm with which it comes in contact.

e May the Optic Vesicle Cause Lens-Formation from Ectoderm
Other than that which. Normally Forms a Lens?

This question is answered by the cyclopean monsters. It is
scarcely conceivable that the ectoderm which would normally lie
over the lateral eyes has the power to migrate, or follow the optic
vesicle so exactly as always to lie just over the vesicle wherever
it may chance to develop. Many embryos have eyes in unusually
anterior positions and derive their lenses from anterior ectoderm,
while others possess ventral eyes with lenses derived from ventral
ectoderm. It occurs in a few cases, as the writer previously

Independent Development of the Lens. 409

EXTREMELY DEFrEcTIVE Optic Cup-Like Bopies ASSOCIATED WITH PERFECT
LENSES

Fig. 21. An anterior section, as is indicated by its size, of a nineteen-day em-
bryo. A free lens is shown on the left while on the right a very small defective
eye contains a small lens. ;

Fig. 22. Asmall lensnear a vesicle-like structure, E, in the brain which might
represent an abortive optic body. F

Fias. 23 and 24 Sections through the embryo shown in fig. 2. In the diagram,
fig. 25, is shown approximately the planes of the sections. The large well-formed
lenses are associated with defective optic cups which lack any sign of differentia-
tion.

410 Charles R. Stockard.

recorded (09), that free lenses arise in their usual lateral positions
while the eyclopean eye possesses its lens of anterior origin.

In the fish embryo the optic vesicle may cause a lens to form
from ectoderm far removed from the usual lens-forming area,
and rarely in such cases it happens that free lenses may also
arise from the usual lens-forming region.

| Does a Deeply Buried Eye have the Power to Regenerate or Form
A Lens from its own Tissues?

It has been shown by many experimenters Colueci (91), Wolff
(95 and ’01), Miller (96) and Fischel (02), that the salamander’s
eye regenerates a new lens from the posterior surface of the iris
when the old lens is removed, or as Fischel found, if the old lens
be merely pushed back out of its usual placeinthe eye. When the
iris was injured in two places during the extirpation of the lens,
two lenses arose within the single eye, one growing from each
injured area of the iris. It has also been found that a fish’s
eye would regenerate a lens under certain conditions: when the
fish is young and when a sufficiently long time is allowed for the
lens to regenerate.

Lewis (04) finds that the deeply buried eyes in Rana palustris
which fail to come in contact with the ectoderm are unable to
form lenses from their own tissues. While on the other hand he
states that in a second species, Rana sylvatica, the optic cup
readily gives rise to a lens from its own tissues if prevented from
stimulating the formation of a lens from the ectoderm of the body
wall.

The fish embryos which are now being considered, act in a
similar manner to Rana palustris and are unable to form lenses
from the tissues of their optic cups. Whenever the optic cup
is deeply buried and fails to reach the ectoderm it also fails to
possess a lens, as is illustrated on the left side of figs. 26, 27 and
28. In this connection it may be mentioned that Morgan was
unable to obtain the regeneration of a lens in adult specimens
of Fundulus, although as mentioned above, lenses do regenerate
in the eyes of another species of fish.

Independent Development of the Lens. Al]

Dererpty BurtieED Eyres witHoutr LENSES

Fia. 26. A section of a thirteen day embryo which was treated with alcohol.
One eye faces outward and contains a lens, while the other faces in toward the
median plane of the head and is without a lens, probably never having come in
contact with ectoderm.

Fic. 27 A section of a nineteen day embryo more posterior in the same series
than section, fig. 6. The deeply buried defective eye has no lens.

Fig. 28 Section of thirteen day embryo after treatment with alcohol. Sec-
tion passes in front of the median plane of the right eye which is well differentiated
and contains a lens. The other eye is small and scarcely differentiated, buried in
the tissue below the brain and contains no lens. A normal ear exists on the side
with the large eye and a small defective ear is on the other side in more posterior

sections.

412 Charles R. Stockard.
5 DISCUSSION OF PREVIOUS OBSERVATIONS AND EXPERIMENTS

Before drawing final conclusions from the cases discussed above
it may be well briefly to review the modern work and experi-
ments which have been directed towards a solution of the so-
called lens problem.

Rabl (’98) in his study of the structure and development of the
lens, found in one case, at least, that a lens-like body was present,
although far removed from the optic vesicle. This observation
has been criticised by Lewis, who suggested that the ectoderm
with the newly forming lens had shifted away from the small
optic vesicle. Again, Lewis states that Rabl’s case, and also a
case to be considered below that was shown by Mencl, prove
neither one side nor the other, since the experimental evidence
is all directly for the idea that a lens will not arise from the skin
without the stimulus of the optic vesicle. At the present time,
however, the experimental evidence points in an opposite direc-
tion, and the cases of Rabl and Mencl can scarcely be disposed
of in so brief a manner. On the contrary Rabl’s example must
be considered the initial illustration of the origin of an early
lens without a stimulation from the optic vesicle.

Following this single case a strong tide turned towards the
idea of the dependent origin and development of the optic lens.
Herbst’s paper (’01) on ‘‘Die formative Reize in der thierischen
Ontogenese’’ and Spemann’s pioneer experiments (’01) on the de-
velopment of the lens seemed most convincing evidence in favor of
a correlation in development between the optic vesicle and the
lens, a correlation in which the latter played a dependent réle.
Herbst claimed the optic lens to be a ‘‘Thigmomorphose”’ origi-
nating only by a contact between the optic vesicle and the epi-
dermis. His reason for such a position being that in the case
of cyclopean monsters the median optic vesicle always derives
a lens from the overlying ectoderm, while no lenses arise in the
usual lateral positions. “If the lens is an independent organ why
does it not arise in the lateral eye region of cyclopean monsters?”
In former experiments the writer (09) produced just such a case
as Herbst thought necessary to show the independent origin of

@

Independent Development of the Lens. 413

lenses, that is, a cyclopean monster with lenses in the usual
lateral positions.

After a study of a great number of such monsters I am now able
to reaffirm that free lenses do occasionally arise in the lateral
eye regions. Herbst’s clever argument based on pathological
embryos is, therefore, rendered invalid.

Spemann (’01) was the first to clearly attack the problem experi-
mentally. He injured or destroyed the optic vesicles of Triton
embryos by means of hot needles and electric cauterizers. The
method was not altogether satisfactory since such an operation
often injures much of the surrounding tissue, yet Spemann’s
results were of the highest value, and stimulated an active interest
on the part of many experimenters. His conclusions furnished
strong support for Herbst’s idea of the dependent origin of the
lens. He found that whenever the optic vesicle was so injured
that it failed to come in contact with the head ectoderm, the
ectoderm failed to form alens. The lens was, therefore, depend-
ent for its origin on a contact stimulus of the optic vesicle upon
the ectoderm. Later Spemann (’05) also concluded that the lens
was not self differentiating but that a durable contact stimulus
of the optic cup was necessary for it to form lens fibers. Thus
the ‘‘Herbst-Spemann theory of dependent lens formation,”’
as Mencl has termed it, was developed. We shall see below,
since it seems best to consider these papers in a more or less chrono-
logical order, that Spemann’s own later work has helped materi-
ally to overthrow this theory.

Barfurth (’02) also operated witha hot needle to destroy the lens
anlage and the optic cup. The embryos were examined after
five or six days. One specimen showed on one side a poorly
regenerated optic cup that did not come in contact with the ecto-
derm and yet a lens still connected with the ectoderm was present
on this side. Barfurth was inclined to accept the Herbst-Spe-
mann idea of lens formation and so attempted to harmonize his
case with the theory as follows. Some sections of the embryo,
9 to 12, showed the optic vesicle lying very near the ectoderm
but not in contact with it. Barfurth supposes that at an earlier
period in development it may have been in direct contact with

&

414 Charles R. Stockard.

ectoderm and at such a time stimulated the lens to arise. This
is merely a conjecture and the more recent experiments of King
(05) and Spemann (’07) on the frog, and the writer’s on the fish,
would warrant equally well an interpretation of independent
origin for this lens.

Returning to observations on monsters we may consider the
case reported by Menel (’03) which has called forth so many
explanations and criticisms. This report described the existence
of an independent lens on each side of the head in an anadidymus
embryo of Salmo salar which he had possessed since 1899, his
interest in it being aroused by the contributions on the dependent
origin of the lens. One of the lenses in this monster was closely
applied to the brain wall, and in fact lay in a depression in the side
of the brain; the other lens, however, was completely separated
from the brain by mesenchyme. Spemann and Lewis have tried
to explain this case by assuming the existence in the brain wall of
some optic vesicle tissue or substance which when brought in
contact with the head ectoderm possessed the power to cause
the formation of a lens. There was absolutebLy no evidence of
optic tissue in this brain wall and further, the explanation could
apply only to one of the lenses, though it scarcely explains the
origin of either. The second lens, entirely free and with mesen-
chymatous tissue separating it from the brain wall, was as perfect,
though not so large, as the other which lay against the brain.
This lens probably arose and developed freely, just as did so many
lenses in the fish embryos the writer has studied.

Menel (’0S) has: more recently obtained other anadidymus
Salmo embryos and confirms his former observations, finding that
in such monsters free lenses are present in 25 per cent of the cases.

Gemmill (’06) has also recorded the frequent occurrence of free
lenses in the heads of monster trout embryos.

Schaper (’04) in considering a case of a typical lens develop-
ment comes to the theoretical conclusion that the lens is by
nature a primitive sense body similar to the sinnesknospe of
Amblystoma, and has secondarily taken on its present function.
It must, therefore, arise independently of the optic vesicle,
with which it is only recently associated. Such speculation

Independent Development of the Lens. 415

is not entirely out of accord with the present facts of lens
formation. |

Lewis thinks that the free rudimentary lenses in Schaper’s
experiments were undoubtedly caused by the shifting of the ecto-
derm and lens away from the optic vesicle. The writer is unable
to see.any reason why such a shifting is supposed to have taken
place. On the contrary, the facts seem to lend themselves more
readily to the interpretation of a free origin of the lens.

The experiments of Lewis (’04) seemed to show convincingly
that the lens was dependent for its origin and development upon
a contact stimulus of the optic vesicle on the ectoderm. Lewis
devised a method far more refined thar any previously used in
similar experiments. He operated on tacpoles under a binocular
microscope with needles and small scissors and was able to cut
the ectoderm and fold it forward so as to expose the brain and
early optic vesicle, which could now be cut away. The ectoderm
was then folded back inplace. Frcm these experiments the follow-
ing are some of the conclusions which were drawn:

Neither a lens nor a trace of a lens will originate from the ecto-
derm which normally gives rise to one, if the contact of the optic
vesicle with the skin is prevented.

There is no predetermined area of the ectoderm which must
be stimulated in order that a lens may arise. Various parts of
the skin when stimulated by optic cup contact may and do give
rise to a lens.

In normal development the lens is dependent for its origin
and differentiation on the contact influence or stimulation of the
optic vesicle on the ectoderm.

The conclusions are clearly stated and the experimental method
employed is most skillful, but not entirely free from objection.
The conclusion regarding the entire dependence of the lens on
an optic vesicle stimulus will not, the writer believes, be supported
by future experiments, and indeed at the present time such an
idea has a vast amount of evidence against it, unless the partic-
ular species experimented with shows a specific action in lens
formation. The later work of Spemann (’07) which contradicts his
former conclusion shows that in some species of frogs the lens

416 Charles R. Stockard.

may arise and differentiate independently of the optic vesicle
stimulus. In these experiments it must be noted that Spemann
operated with glass needles to remove the optic vesicle regions
from the open medullary plate. Such an operation does not
injure the lens-forming region of the ectoderm as Lewis’ experi-
ment does and it is on this account, the writer believes, that the
free lenses arise.

The tendency of the ectoderm to form a lens independently
is so delicately adjusted that a very slight injury or disturbance
may suppress it, yet the same ectoderm may still have the power
to form a lens in response to the stronger stimulus of the optic
vesicle. So in experiments where the ectoderm has been cut or
injured it loses the power to form free lenses even though the optic
vesicle can stimulate a lens to arise from it. When the ectoderm
is uninjured, as in some of King’s specimens, Spemann’s (07),
and the writer’s, then free lenses do occur.

We may imagine the lens to represent an ectodermal organ
formerly of independent importance. However, it has now be-
come so closely associated with the nervous portions of the eye
that it arises whenever such a part meets the ectoderm, yet the
lens retains to a feeble degree its impulse to arise independently
of other eye parts. When it has once arisen it is perfectly capable
of differentiation. Future experiments of removing the optic
vesicle without injuring the ectoderm will probably demonstrate
further this tendency of the lens to arise independently, just as
Mencl’s observations and the writer’s experiments show for the
fish, and many of the experiments mentioned show for amphibians.

King (05) states that she had begun her series of experiments in
1900. She destroyed the optic vesicles from the forebrain region
in the closing meduilary tube with hot needles. Many of the
embryos died as a result of theoperation. Theconclusionsreached
by King are entirely opposed to those of Lewis and Spemann’s
earlier results regarding the dependent origin of the lens. King
found that in some of the embryos a lens-like body arose from the
ectoderm on the side with an injured or unregenerated optic cup
which did not come in contact with either the ectoderm or the
lens-like body. Some of the specimens show a very suggestive

Independent Development of the Lens. 417

lens-like structure which is entirely free from any contact with
an optic vesicle. The inaccuracy of the operation may account
for these lenses, on the ground that in exceptional cases the outer
ectoderm of the lens region was too far distant at the early stage
of development to have been injured. This is probable when it is
noted that the operation did not enter through the lateral ecto-
derm, but through the partly open medullary tube from above.
The present evidence also goes to strengthen King’s position on the
subject.

Spemann (05), Lewis (07) and LeCron (’07) have all claimed
that the lens after its origin, was not self differentiating but
depended upon a durable contact with the optie cup for its
future development. Ail of the writer’s experiments and the
observations of Mencl are clearly contradictory to such a view,
and Spemann (’07) has more recently found the lens to be self-
differentiating in Rana esculenta.

In a former paper the writer (’07) described the development. of
the lens in the blind Myxinoid, Bdellostoma stouti. At that time
he presumed that this animal furnished support for the experi-
mental evidence that the lens was not self-differentiating. In
Bdellostoma embryos the optic vesicle is well formed at first and
reaches out from the brain to the ectoderm; at this time the ecto-
derm forms a localized thickening suggesting a lens-plate. The
optic vesicle then ceases to maintain its progressive development
and loses connection with the ectoderm, finally giving rise to
the poorly formed optic cup of the adult. The early lens thicken-
ing of the ectoderm degenerates and is entirely absent from older
embryos. The writer suggested that this degeneration was due
to the loss of contact with the optic vesicle, since such reasoning
seemed correct in the light of the experiments up to that time. He
has entirely changed his view, however, regarding the matter of
the lens in Bdellostoma and now believes that it represents a
rudimentary organ which has been lost in the adult and appears
only for ashort time during embryonic life. As Eigenmann (’01)
records of Amblyopsis, the lenses of other blind fishes appear at
certain stages in the developing embryo and then degenerate. A
‘case not so far advanced in the degeneration of the eye and lens is

THE AMERICAN JOURNAL OF ANATOMY, VOL, 10, No. 2.

418 Charles R. Stockard.

that of the Florida burrowing lizéard, Rhineura. In the adults of
Rhineura, Eigenmann finds lenses sometimes present and sometimes
absent and when present they are very variable. ‘Somany examples
are known of the traces of rudimentary organs in the embryo
which are entirely lacking in the adult that the above cases of
the lens are not at all strange and at present it seems that this
is the most satisfactory explanation of their behavior.

The more recent experiments of Spemann (’07) so frequently
referred to, may be briefly described. Embryos of Rana escu-
lenta were operated upon with glass needles so as to remove the
optic vesicle areas from the open medullary fold (his former
experiments were made with hot needles, which doubtless injured
more of the surrounding tissues).

After allowing these embryos to develop for several days and
then studying them in section it was found that in one case on the
side of the head lacking an optic vesicle a true lens-bud was still
in connection with the ectoderm. In an older embryo a free lens
vesicle was found, and finally, in a still older specimen a lens was
buried in connective tissue on the eyeless side of the head yet
it possessed fully formed lens-fibers. Thus, Spemann, five years
after his first paper on the dependent origin of, the lens, now con-
cludes that the lens in Rana esculenta is self-originating and
self-differentiating. He also accepts Mencl’s case of the inde-
pendent lens in Salmo salar.

We have thus seen in a rather cursory survey how the lens
problem has arisen and how experiments have built up first one
side of the question and then the other, and many may also feel
that the final word is yet to be added. Nevertheless, it is true
that at this stage of the investigation it has been clearly demon-
strated in several groups of animals that the ectoderm possesses
the power independently to originate a lens which subsequently
develops into the transparent refractive organ usually found
within the mature eye. The manner of origin and differentiation
of the lens may easily differ in different animal species, and the
statement would not be warranted that all of these facts apply
to animals in general.

The lens is self-originating and self-differentiating yet it is more

Independent Development of the Lens. 419

emphatically proven that the optic vesicle or optic cup has the
power without exception to cause a lens to form from the ecto-
derm with which it comes in contact, even though this ectoderm
may be far distant from the usual lens-forming area. There is
probably no strictly limited region of ectoderm from which the
optic vesicle must stimulate lens-formation. The self-originating
lenses, however, have invariably occurred in the head region,
though not always in their usual’ lateral positions. It would,
therefore, seem that the ectoderm of the head is more predis-
posed to the formation of lenses than that of the other body
regions.

6 SUMMARY AND CONCLUSIONS

1 When the developing eggs of Fundulus heteroclitus are
subjected to the action of Mg salts, aleohol, or other anesthetic
agents, the normal outgrowth of the optic vesicles is generally
inhibited. Embrvos are obtained either entirely without optic
cups, with small deeply buried eyes, with only a single eye on one
side of the head or, finally, with a median more or less double
cyclopean eye. These specimens furnish exceptional material
for a study of the relationship between the development of the
optic vesicle and the optic lens. The embryos with the nervous
eye parts entirely lacking are similar to specimens with the optic
vesicles mechanically cut out of the brain. At the same time,
the injury to the ectoderm which usually results from the opera-
tion is avoided, and this is a most important advantage. When
Spemann (’07) operated on Rana esculenta embryos with open
medullary plates he was able to remove the optic vesicle areas
from the future inner side of the tube without injuring the ecto-
derm of the lens-forming region and in such cases independent
lenses arose on the eyeless side. King (’05) obtained free lenses in
a somewhat similar experiment. Lewis’ embryos might possibly
respond in a like manner if operated upon in this fashion. The
optic vesicle may stimulate a lens from slightly injured ectoderm,
but the free origin of lenses is a more delicate process and one more
easily interfered with.

420) Charles R. Stockard.

2 The crystalline lens may originate from ectoderm without
any direct stimulus whatever from an optic vesicle or cup. These
self-originating lenses arise from regtons of ectoderm that are
not in contact with either optic vesicle, the brain wall, or any
nervous or sensory organ of the individual (see plate I).

3 The lens-plate or lens-bud is capable of perfect self-differen-
tiation. No contact at any time with an optic vesicle or cup,
is necessary. These lenses finally become typical transparent
refractive bodies exactly similar in histological structure to that
in the normal eye (see plate IT).

4 The size and shape of the lens is not entirely controlled by
the associated optic cup. Lenses may be abnormally small for
the size of the cup, or entirely too large, so that they protrude;
or, finally, peculiarly shaped oval or centrally constricted lenses
may occur in more or less ordinarily shaped optic cups. The lens
is by no means always adjusted to the structure of the optic cup
as has been claimed by some observers.

5 An optic vesicle or cup is invariably capable at some stage
of its development of stimulating the formation of a lens from
the ectoderm with which it comes in contact. It is remarkable
how extremely small an amount of optic tissue is capable of stimu-
lating lens formation from the ectoderm (plate II, fig. 5).

6 The optic vesicle may stimulate a lens to form from regions
of the ectederm other than that which usually forms a lens. This
is shown by the fact that a median cyclopean eye always stimu-
lates a lens to form from the overlying ectoderm. It is scarcely
possible that the lateral normal lens-forming ectoderm could
follow the cyclopean opti¢ cup to the many strange situations
it finally reaches.

7 The ectoderm of the head region is more disposed to the
formation of lenses than that of other parts of the body, since the
free lenses invariably occur in this region.

8 <A deeply buried optic vesicle or cup may fail to come in
contact with the ectoderm; in such cases it lacks a lens. The
tissues of the embryonic cup itself are unable to form or regenerate
alens. This is not true in all embryos, as Lewis has shown for
one species of frog.

Independent Development of the Lens. 421

9 The optic lens may be looked upon as a once independent
organ (possibly sensory or perhaps an organ for focusing light on
the brain wall, before the vertebrate eye had arisen) which
has become so closely associated with the nervous elements of
the eye that it has to some extent lost its tendency to arise inde-
pendently, although still capable of doing so under certain con-
ditions. The lens now arises much more readily in response to a
stimulus from the optic vesicle, a correlated adjustment which
insures the almost perfect normal accord between the optic cup
and the lens. In experiments on the origin of the lens one must
guard against disturbing the ectoderm from which an independent
lens might arise. Although an optic vesicle might have the power
to stimulate a lens from injured ectoderm, the innate tendency of
the ectoderm to form a lens is a more delicate impulse and may be
suppressed by a slight injury or disturbance of the ectoderm
during the operation.

7 BIBLIOGRAPHY

BarrurtH, D. Versuche iiber Regenerationdes Auges und der Linse beim Hiihner=
1902 embryo. Verh. d. Anat. Gesellsch. Halle.
Couucci, V.S., Rigenerazione parziale del!’occhio nei Tritoni. Mem. Accad. Se.
1891 Bologna, Series 5, Vol. i.
ErGgeNMANN, C. H. The History of the Lens in Amblyopsis. Proc. Indiana
1901 Acad. Se., 1901.
FiscHEL, A. Weitere Mitteilungen iiber die Regeneration der Linse. Arch. f.
1902 Entw’Mech., Bd. xv.
GemmiLL, J. F. Notes on Supernumerary Eyes, and Local Deficiency and Re-
1906 duplication of the Notocord in Trout Embryos. Proc. Zodl. Soc.
London, Vol. i.
Hersst, C. Formative Reize in der thierischen Ontogenese. Hin Beitrag zum
1901 Verstiindniss der thierischen Embryonalentwicklung. Leipzig,
H. Georgi. .
Kine, H.D. Experimental Studies on the Eye of the Frog. Arch. f. Entw’ Mech.,
1905 XX:
LeCron, W. L. Experiments on the Origin and Differentiation of the Lens in

1907 Amblystoma. Am. Jour. Anat., Vi.
Lewis, W. H. Experimental Studies on the Development of the Eye in Am-
1904 phibia. I. On the Origin of the Lens. Rana palustris. Am.

Jour. Anat., ili.

1907 Experimental Studies, etc. iii. On the Origin and Differentiation of
the Lens. Am Jour. Anat., vi.

Accepted by the Wistar Institute of Anatomy and Biology January 12, 1910. Printed June 21, 1910.

422 Charles R. Stockard.

Menci, E. Ein Fall von beiderseitiger Augenlinsenausbildung wihrend der
1903 Abwesenheit von Augenblasen. Arch. f. Entw’ Mech., xvi.
1908 Neue Tatsachen zur Selbstdifferenzierung der Augenlinse. Arch. f.
Entw’ Mech., xxv. :
Miu.uter, E. Regeneration der Augenlinse nach Extirpation be Tritonen. Archiv
1896 f. Mikrosk, Anat., xlvii.
Ras, K. Ueber den Bau und die Entwickelung der Linse, I. Zeitsch. f. Wiss-
1898 Zo6}\., Bd. 63.
Scuarer, A. Uber einige Fille atypischer Linsenentwickelung unter abnormen
1904 Bedingungen. Ein Beitrag zur Phylogenie und Entwickelung der
Linse. Anat. Anz., xxiv.
SPEMANN, H. Ueber Correlationen in der Entwickelung des Auges. Verhandl.
1901 der Anat. Gesellsch., 1901.
1905 Ueber Linsenbildung nach experimenteller Entferung der primiaren
Linsenbildungzellen. Zoél. Anz., xxiii.
1907 Neue Tatsachen zum Linsenproblem. Zoél. Anz., xxxi.
Srockarp, C. R. The Artificial Production of a Single Median Cyclopean Eye
1907a in the Fish Embyro by Means of Sea-Water Solutions of Magnesium
Chlorid. Arch. f Entw. Mech., xxiii.
19076 The Embryonic History of the Lens in Bdellostoma stouti in Relation
to Recent Experiments. Am. Jour. Anat., vi.
1908 The Question of Cyclopia. Science, n. s. xxviii.
1909 The Development of Artificially Produced Cyclopean Fish, ‘‘The
Magnesium Embryo.” Jour. Expr. Zodl., vi.
Wotrr, G. Regeneration der Urodelenlinse. Arch. f. Entw.’—Mech., I und xii.
1895 and 1901.

Accepted by the Wistar Institute of Anatomy and Biology January 12, 1910. Printed June 21, 1910.

Independent Development of the Lens. 423

EXPLANATION OF PLaTEs I anp II

Fig. 1 A photograph of a section of a seventy-six hour Fundulus embryo; the
brain is almost bilateral and perfect, with a well formed left optic cup, no indica-
tion of the right cup exists, yet the ectoderm on that side has formed a well
pronounced lens-bud, L.

Fig. 2 A section through the eye region of a thirty day embryo, there is no
optic cup in the specimen. A perfect lens is in the usual lateral position, L, a
band of muscle, M, lies between it and the brain.

Fic. 3 A section through the head of a Fundulus embryo eighteen days old.
The specimen is shown in text-figure 1, no eyes are present, yet two perfectly differ-
entiated crystalline lenses are seen in the sides of the head. A close examination
of the photograph will reveal mesodermal cells between the lens and the brain.

Fig. 4. A section through one defective eye which possesses a lens of dispro-
portionate size, while on the other side of the head two free lenses are seen, the
lower small one protruding from the head. A second defective eye with a lens
is found in a more posterior region of the head. Text-figure 4 represents this
embryo as it appeared in life.

Fic. 5 A section through one of the very defective optic vesicles shown in
fig. 2. The photograph illustrates the extreme lack of proportion which may
exist between the size of the optic vesicle and its associated lens. It indicates
also how small an amount of optic tissue may stimulate lens formation from the
ectoderm.

INDEPENDENT DEVELOPMENT OF THE LENS PLATE I
C. R. StockarD

2

THE AMERICAN JOURNAL OF ANATOMY, VOL, 10, No. 3.

INDEPENDENT DEVELOPMENT OF THE LENS PLATE II
C. R. SrocKarp.

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, No. 3.

THE DEVELOPMENT OF ISOLATED BLASTOMERES
OF THE FROG’S EGG

J. F. MCCLENDON

From the Zoélogical Laboratory of the University of Missouri and the Histological
Laboratory of Cornell University Medical College, New York City

WITH TWO FIGURES

After the numerous proofs furnished by various investigators,
there is no question that differentiation begins very early in
development, but the determination of the exact stage at which
various differentiations begin, has been hampered by technical
difficulties. The importance of overcoming these difficulties lies
in the fact that it is only by an exact study of the early develop-
ment that we can ever hope to know the mechanics of differen-
tiation. The later stages are so complex that it is doubtful that
they could be analysed even though the early stages were under-
stood.

The frog’s egg, owing to its large size, has been a favorable
object of study. O. Hertwig,! after pricking one blastomere,
found that a complete embryo was formed.

Roux,? in similar experiments obtained a half embryo from the
uninjured blastomere, and explains Hertwig’s observation by the
fact that some of these half embryos became complete larve
by a process of regulation which he called post-generation. This
might occur in the following different ways: first, the half embryo
might fold together on the side corresponding to the median plane
and be transformed into a whole embryo of half size; second, the
injured blastomere might recover from the operation and develop
into the missing half; and third, the injured blastomere might be

Arch. f. Mikroscopische Anat., 1893, XLII, p. 662.
?Ges. Abh. zur Entwicklungsmechanik der organismen, Leipzig, 1895.

426 J. F. MeClendon.

reorganized under the influence of the half embryo into the miss-
ing half. The occurrence of post-generation, and especially the
third type, has been denied by a number of investigators who
repeated this experiment. Laquer* has reinvestigated the ques-
tion.

This production of a half embryo from one blastomere is not
due to the inability of the egg to produce more than one embryo,
since Schultze produced double forms by inverting the egg in the
two-cell stage, and Morgan® produced a complete embryo from
one blastomere by pricking the other blastomere and then invert-
ing the egg. In order to determine whether this inversion of the
egg and consequent stirring up of its contents is necessary for the
production of a complete embryo from one blastomere, I attempt-
ed to find a frog’s egg that would permit the complete removal of
one blastomere without death of the egg. No one has hitherto
been able to do this, although Roux® observed a partial but very
abnormal development of extra-ovates of the frog’s egg.

Last spring at Columbia, Missouri, I located the breeding
places of Rana pipiens and Chorophilus triseriatus. The unseg-
mented eggs of both species could be collected in the ponds and
small streams, or in the frog cages in the laboratory, the eggs of
the latter species being more easily obtained.

I tried various methods of removing one blastomere in the two-
cell stage. It was possible to suck out one blastomere with a
capillary pipette connected with a rubber tube, one end of which
was held in the mouth, but it was very difficult to pierce the jelly
with the pipette without killing the egg, and the suction often
removed both blastomeres. A better way of making a hole in the
jelly was the time honored method of piercing it with a hot
needle. A very coarse needle was used and heated to a tempera-
ture that would coagulate a portion of the blastomere into which
it was thrust. On withdrawing the needle, the coagulated parts of
the jelly and blastomere were pulled out, thus leaving a large hole

“Arch. f. Entwicklungs. 1909, XXVIII, p. 327.

‘Arch. f. Entwicklungsmech., I. p. 298.

*Anat. Anz., 1895, X. p. 5238.

*Jahresbericht d. Schles. Ges. f. vaterl. Cultur, Juni, 1887.

Isolated Blastomeres of the Frog’s Egg. 427

through which the remaining part of the blastomere could be
extracted. The eggs of Chorophilus were more easily handled,
since the jelly was softer and the egg itself did not collapse readily.
This egg is not so small as to make operations difficult with the
low power of the binocular microscope.

When the first cleavage furrow had extended almost around the
egg, the greater part of the jelly was carefully removed with filter
paper and the puncture made. The egg was then allowed to rest
until the first cleavage plane had completely divided it. At this
time the remains of the punctured blastomere were removed
with the capillary pipette. In case the last remnants could not be
removed, the egg was allowed to rest again until the remnants of
the punctured blastomere turned pale, which was an indication that
they were more brittle and could now beremoved with a fine needle.

When the egg was punctured, the perivitelline space was oblit-
erated by the escape of the contained fluid and the jelly pressed
close to the egg, an opening was also made through which bacteria
might enter; therefore, the eggs laid in the laboratory under a
more sterile condition, gave better results than those laid in the
ponds. Care was taken to keep the egg normally orientated in
reference to the direction of gravity both during and after the
operation.

The operated eggs were observed with the binocular micro-
scope and the early cleavage noted. The gastrulation was watched
through the sides and bottom of the glass dishes in which the eggs
were kept. After the embryos became ciliated, some of them
turned in a normal manner so that they could be observed in all
aspects, from above:

A-large number of eggs of both Rana pipiens and Chorophilus
triseriatus were operated on. All of the former died, but of the
latter a considerable number gastrulated and several reached the
tadpole stage. Two were four days, three were five days and one
was eight days old when preserved for sectioning.

The control eggs hatched in less than eight days, but the operat-
ed eggs were retarded in development and the resulting tadpoles
were too weak to break through the jelly, although the twitching
of their tails showed that the'muscles were functional.

428 J. F. MeClendon.

The development of the operated eggs was very similar to that
of the normal, and quite different from that of the uninjured blasto-
meres in Roux’s experiment. The remaining blastomere rounded
up gradually after its partner had been removed, and the blastula,
gastrula and later stages were wholes of half size. As the mortality
was great, it is to be expected that defects would be observed in
many of the specimens, but these defective specimens could not
be interpreted as half embryos. The only defect that I could
observe in the oldest specimen was the small size of the suckers,

Fig. 1 Fig. 2

Fig. 1. Chorophilus triseriatus: a section of the left eye of a tadpole developed
from one of the first two blastomeres.

Fig. 2. A section of the right eye of the same tadpole as in fig. 1.

before the pericardium began to swell so that the heart beats
could no longer be observed.

Sections confirmed the observations on the living material
and showed that the embryos were complete and not half-
forms.

Figs. 1 and 2 represent the right and left eyes seen in transverse
sections of the eight day larva. The right eye lacks a lens, and
the left eye is not quite normal, yet these defects make no ap-
proach to those of a half-larva since they are not very extensive

Isolated Blastomeres of the Frog’s Egg. 429

and are not all on the same side. There is some displaced pigment
in the center of the lens, and the cells of the optic cup have not
preserved their normal arrangement. An examination of the
ectoderm showed more or less disintegration, and the defects
observed may be explained by the fact that the specimen was
dying when it was taken out of the water to be preserved.

These differences in the two eyes of the eighty-day embryo
from one blastomere, are the most striking differences between
the right and left sides. The ear vesicles are almost identical on
the two sides, and the same is true of the other paired organs.
Furthermore, no general defect of the anterior, posterior, dorsal
or ventral regions of the body could be found. The study of the
five other embryos developed from operated eggs, and cut in
serial sections also revealed complete forms.

Roux described right and left ‘‘hemiembryones iaterales’”’ and
also “‘hemiembryones anteriores”? developed from one blasto-
mere in the experiments mentioned above.

Since the anterior end of the neural folds lies on one side of the
egg between the equator and the animal pole, i. e., about the
region in which the gray crescent appeared, and the dorsal side
of the embryo is formed chiefly on this side, it is probable that in
case the first cleavage plane were transverse to the plane of sym-
metry, it would divide the egg into the halves that would form
the dorsal and ventral parts of the embryo respectively. There-
fore the ““hemiembryones anteriores” might better be described
as “‘dorsales” as indicated by Przibram. Roux obtained no
‘“posteriores” (‘‘ventrales” of Przibram) and since a number of
the eggs died, it is probable that those blastomeres which corre-
spond to the ventral part of the embryo, and therefore contain no
part of the gray crescent, are not so well adapted to independent
development. In my experiments the position of the grey crescent
as marking the future dorso-anterior region of the embryo was
not recorded, but it seems probable from the results of Roux’s
experiment, that those isolated blastomeres containing no part of
the grey crescent did not develop.

As noted above, Roux found that half-embryos sometimes be-
came complete embryos by post-generation.

430 J. F. MeClendon.

None of the three types of post-generation occurred in my
experiments. The isolated blastomeres rounded up in a manner
that might be compared to the first type of post-generation, but
this regulation occurred before an embryo was formed, and might
be distinguished as ‘‘immediate. ”’

It appears then that the egg at the time of the first cleavage is
differentiated in the direction of the primary axis, and further-
more in the direction of a plane passing through this axis and bi-
secting the grey crescent, so that the egg may be-divided in this
plane into two totipotent halves. The halves of the egg divided
in any other plane may or may not be totipotent. The formation
of a half embryo instead of a whole embryo from one blastomere
of the two-cell stage is due to the presence and mechanical inter-
ference of the other blastomere.

In this connection may be mentioned the results of Endres,
Herlitzka and Spemann on the development of the first two blasto-
meres of urodeles when separated by constriction with a fine hair.
If the constriction was slight, the hair merely marked the plane
of the first cleavage, if it was deeper, double monsters occurred,
and if it was complete, two perfect embryos resulte1.

In from 66 per cent to 75 per cent of the eggs, the first cleavage
plane became a frontal plane of the embryo, and in the remainder,
it became the median plane. In the latter case complete constric-
tion gave rise to two complete embryos, but in the former case,
complete constriction resulted in one complete embryo developed
from the prospectively dorsal half of the egg, whereas the pros-
pectively ventral half did not develop beyond gastrulation. Thus
it appears that the eggs of the urodeles possess the same organi-
zation as the eggs of the anura, with the difference that the first
cleavage plane is usually frontal in the former, and median 1 in the
latter in relation to the embryonic axes.

Accepted by The Wistar Institute of Anatomy and Biology, January 19, 1910. Printed June 21, 1910.

NOTES ON THE MYOLOGY OF ANTHROPOPITHECUS
NIGER AND PAPIO-THOTH IBEANUS

E. C. MacDOWELL

WITH FIVE FIGURES

Through the kindness of Dr. Spencer Trotter, professor of
biology at Swarthmore College, I have been afforded the oppor-
tunity of dissecting a female chimpanzee, Anthropopithecus niger,
which had just lost her milk dentition, and an adult male baboon,
Papio-thoth ibeanus. The notes from these dissections were used
by Dr. Trotter in May, 1908, as the basis of a communication to
the Academy of Natural Sciences of Philadelphia.

It is my plan to describe certain muscles in these two forms,
which are found to differ from the conditions normal in man, and
to compare these with the results of previous investigators.

I wish to express here my appreciation of the many sugges-
tions and the valuable assistance rendered by Dr. Trotter in the
preparation of this paper.

MUSCULATURE OF THE UPPER LIMB
A. MUSCLES OF THE SHOULDER

The Pectoralis Major in the chimpanzee, is divided into three
parts which are not absolutely differentiated: (a) pars abdominalis;
(b) pars sternalis; and (c) pars clavicularis. The first (a) arises
from an aponeurosis with the rectus abdominus at the level of
the seventh rib and from the sternum as far up as the third rib.
It is inserted into the humerus, 3 em. distal to its head, by a thin
tendon, composed of loose fibers. This is covered by the more
proximal border of the insertion of the pers costo-sternalis. The
second part (6) is the most highly developcd division of the group.
It arises from the sternum and the sternal ends of the costal carti-
lages of the first, second, and third ribs. Its fibers are inserted

432 E. C. MacDowell.

directly into the lip of the bicipital groove of the humerus, from
the tendon of the pars abdominalis to the insertion of the deltoid-
eus. The third division (c) which is, nearly as well developed
as b, arises from the mesial two-thirds of the clavicle. It is
inserted into the humerus for 5 em. below its head, and overlaps
the insertion of the second division (b). But slight differentia-
tion is found between the main masses of the second and third
divisions. There is an indication of a separation of a bundle of
fibers anteriorly, which may represent a pars sternalis, but this
bundle has no distinct insertion.

In the baboon, the pars clavicularis, is very poorly developed.
The pars abdominalis (fig. 1, 4) is a distinct ribbon-like band,
which arises from an aponeurosis with the external oblique 5 cm.
from the sternum, and extends from the seventh to the ninth rib.
In the axilla, it is entirely covered by other muscles. It is inserted
without a tendon, into the humerus close to its head. The pars
costo-sternalis (fig. 1, P) is highly developed. It arises and is
inserted asinman. The parssternalis is very closely intermingled
both with the deltoideus and the main portion of the pectoralis
major. It arises from the sternum (afew fibers are supplied from
the proximal end of the clavicle) and is inserted into the humerus,
proximal to the insertion of the pars costo-sternalis.

This condition bears out Bischoff’s statement, quoted by Prim-
rose, that the pars clavicularis is wanting inthelower apes. Hunt-
ington and Michaélis do not find it in Cynocephalus. Primrose
considers that the attachment of the great pectoral has slowly
crept out from the purely sternal origin, shown in the lower apes,
to the origin which extends nearly two-thirds the way along the
clavicle and joins the deltoideus as it does in my chimpanzee.
Champneys found that the pectoralis major in his chimpanzee
arose from eight ribs. The three divisions were not apparent,
although two slips were differentiated. These arose from the
fourth and fifth ribs respectively, and were fused with the main
part of the muscle at the level of the lower border of the axilla.
He describes the clavicular origin in the baboon as extending
one-eighth of the way from the sternal end of the clavicle. Vro-
lik describes this muscle in his chimpanzee as having a clavicular

The Myology of Chimpanzee and Baboon. 433

and a sternal part. In an orang, -Duvernoy describes two parts,
the sterno-clavio-humerale and the sterno-humerale, where as in a
chimpanzee he found no divisions. Beddard found no connection
with the clavicle in his orang. Owen describes three muscles in
place of all the pectorals, namely the sterno-humeralis, costo-
humeralis, and sterno-costo-humeralis.

The Pectoralis minor arises in the chimpanzee along a line
extending from the first to the fourth ribs, 2 em. from the sternum
at its anterior, and 5 cm. at its posterior end. The muscle is
divided into two separate digitations; the posterior digitation is
subdivided into three. All the fibers quickly converge to the
strong, round tendon, which is half as long as the muscular part.
This is inserted into the capsule of the shoulder joint.

In my baboon and in a small Macacus monkey, the pectoralis
minor is more developed. Distally it extends as far as the pec-
toralis major does in the -first case, and in the second, from the
second rib to the sixth. In both forms, it arises from the sternum
itself and from the sternal ends of the costal cartilages; whereas,
in man, it arises from the ends of the ribs themselves, thus form-
ing a much less powerful muscle.

Champneys and Hepburn describe this muscle in chimpanzees
as arising from the upper three ribs, and being inserted into the
shoulder joint. Duvernoy did not find it divided into distinct
parts. In the orang, Beddard found two parts, one from the third
and fourth ribs, the other from the fourth, fifth and sixth. Hart-
mann describes the condition in the chimpanzee very much as I
found it. His orang corresponded with Beddard’s description.
Primrose found the tendon of this muscle, in his orang, extending
to the coracoid process, whence two ligments were traced to the
head of the clavicle and to the acromion process, respectively.
This was a new condition. Bardeen considers the insertion into
the head of the humerus within the capsule, normal in primates
below man. Ina gorilla, Douvernoy describes this muscle (called
by him ‘‘costo-caracoidien”) as having two parts, one with six
digitations inserted into the coracoid process, the other with two
digitations joined to the short portion of the biceps after its inser-
tion into the coracoid process.

THE AMERICAN JOURNAL OF ANATOMY. VOL. 10, No. 3.

434 E. C. MacDowell.

The Chonro-epitrochlearis is found below the pars costo-ster-
nalis in the chimpanzee. It arises between the second and fourth
ribs from the fascia underlying the pars costo-sternalis, 2 em.
from its origin. It is composed of three strands lying parallel
to those of the superficial muscle. It is inserted into the humerus
by a thin strap-shaped tendon, which adheres closely to the ten-
don of the upper sheet at its insertion into the humerus.

Bardeen describes the muscle when found in man as follows:

This is a slip which springs . . . . from the thoraco-abdomi-
nal fascia beneath the pectoralis major . . . . and extends on
the inner arm to the intertubercular (bicipital) groove . . . . It

is found in 12 to 20 per cent of human bodies (Le Double) aud occurs
normally in many of the lower mammals.

Duckworth deseribes in a gorilla the separation of the posterior
fibers of the pectoralis major to form a.separate muscle, the “pec-
toralis abdominalis,” and identifies it with the chondro-epitro-
chlearis. However, it seems clear that the pars abdominalis in
my chimpanzee must correspond to his ‘‘pectoralis abdominalis.”’
So, if this slip under question corresponds to any muscle in man,
it must correspond to the chondro-epitrochlearis. This seems to
indicate that the abdominal portion of the pectoralis major and
the chondro-epitrochlearis should not be identified. In man,
this muscle has been found in one out of eight (Le Double).

In the baboon, I find a muscle that arises from the rectus
abdominis and the fascia and cartilage of the seventh rib, below
the origin of the abdominal portion of the pectoralis major. Its
fibers form a strip 5 em. wide, which runs parallel to the sternum.
At the level of the second rib, it becomes cartilaginous, and is
inserted into the costal cartilage and the surrounding fascia. It
seems probable that this muscle should be considered a portion
of the rectus abdominis that has been stranded in the thoracic
region upon the retreat of this muscle.

Wiedersheim states that the abdominis in Amphibians extends
from the tail to the head, and that this muscle crept back with
the development of the pectorals, which needed a solid frame
for their insertions. In the lower primates, this muscle regularly

The Myology of Chimpanzee and Baboon. 435

reaches well up into the thoracic region, while in man it has occa-
sionally been found to extend asfarup asthesecondrib. Bardeen
says that a more primitive condition, in which the muscle extended
to the neck, is frequently indicated in man by aponeurotic slips
or muscle slips on the upper part of the thorax. In my Macacus,
I find the rectus abdominis extending up to the fifth rib.

The Subclavius is found in the chimpanzee to arise from the
first rib 3.5 em. from the sternum, by a short tendon. The belly
is .75 cm. in diameter. Its fibers are inserted into the distal half
of the clavicle. Some of these fibers are closely related to the
conoid ligament. In the Macacus, this muscle is applied to the
clavicle for most of its length.

Vrolik found this as in man; Beddard describes it asinserted on
‘the proximal half of the clavicle and ensheathed by the coraco-clavi-
cular ligament. In his orang, Hepburn found an additional slip
from the second rib, while in the orang of Primrose it was in no
way continuous with the coraco-clavicular ligament, and was
poorly developed.

In the chimpanzee, I also find a Sterno-chondro-scapularis aris-
ing as a tendon inserted into the first rib, and affixed to the cora-
coid process internally to the attachment of the short head of
the biceps. ‘The only muscle fibers that occur are in a small
eroup at the coracoid attachment, and in a still smaller one at
the costal insertion. Champneys describes such a fibrous band
and notes a partial fusion with the subclavius at the anterior
end. He quotes Rolleston as calling this the representative of
the long coracoid of birds. Le Double reports it as occurring in
man in two out of forty cases.

The Deltoideus presents only slight irregularities. In the chim-
panzee and in the baboon, the connection between its most ven-
tral border and the clavicular portion of the pectoralis major is
unusually close. In the baboon, the short attachment of the
pectoralis to the clavicle gives the deltoideus a remarkable long
clavicular origin.

Champneys and Michaélis say that it arose from nearly all
the clavicle in their baboons, while Hepburn states that the close
joining of the deltoideus with the clavicular portion of the pec-

436 E. C. MacDowell.

toralis is common in all anthropoid apes. Beddard describes
this muscle in Anthropopithecus calvus as being relatively as
large as in man, and Hartmann says that it is well developed in
anthropoids generally.

B. MuvuscLES oF THE ARM

The Latissimus dorsi arises in my chimpanzee from the four
posterior lumbar vertebrae closely mingled with the posterior
border of the trapezius, from the crest of the ilium, and by inter-
digitations with the obliquus externus abdominis from the four
posterior ribs. The muscle is inserted into the bicipital groove
by a strong flat tendon 2 em. wide and 4 cm. long, superficial to
the proximal half of the broad insertion of the teres major. There
is no connecting fasciculus between these two muscles. The
fibers of the Latissimo-condyloideus (dorsi-epitrochlearis) arise
from the flat tendon of the latissimus dorsi at its distal end, and
continue down the arm in a belly 1.8 cm. in diameter to be firmly
inserted by a strong flat tendon 4 em. long into the internal con-
dyle of the humerus. This muscle receives a fasciculus from the
coracobrachialis.

In the baboon, the tendon of the latissimus dorsi (fig. 1, F)
is entirely distal to that of the teres major (fig. 1, R). <A large
fasciculus (fig. 1, V) is given off from the superior border of the
latissimus dorsi to be inserted into the lower border of the broad
flat tendon of the teres major before the insertion of the former
into the humerus. The latissimo-condyloideus (fig. 1, U) is
large and well developed. It- arises mainly from the muscle
fibers of the latissimus dorsi. Indeed, many of the fibers of the
latissimus dorsi seem to be continuous with those of this muscle.
It is inserted into the long head of the triceps and the brachial
fascia just above the elbow. In this way, the insertion of the
latissimus dorsi is practically extended to the olecranon process,
and a very long and powerful muscle is formed, which may be called
the Latissimo-dorsi-condyloideus. In the Macacus, the latissimo
condyloideus is very poorly developed. Its fibers are inserted
directly into the fascia over the triceps.

The Myology of Chimpanzee and Baboon. 437

In his orang, Primrose found a fasciculus from the latissimus
dorsi to the tendon of the teres major, similar to the one I have
described in the baboon. Hepburn describes a similar condition
in a chimpanzee and in an orang. He states that the tendons
of the latissimus dorsi and of the teres major are joined in the
gibbon. The fascicular connection in the higher forms denotes
an earlier joining of the teres muscles; however, in my baboon,

Fic. 1. The side view of the baboon; the left armis raised to show the insertion.
of the Latissimus dorsi. A, pars abdominalis of pectoralis major; B, biceps;
C, Coracobrachialis; ZH, Tendon of insertion of Latissimusdorsi; Z, Lattissimus
dorsi; P, Pars costo-sternalis of Pectoralis major; Rk. Teres Major; S, Serratus
anterior; 7’, Triceps; U, Lattissimo-condyloideus; Y, Fasciculus from Latissimus
dorsi to teres major.

where this is present, the insertions themselves are further sepa-
rated than in my chimpanzee, where it is absent. Vrolik, Chap-
man, Hepburn, Beddard, Duvernoy and so many others describe
the latissimo condyloideus that we may well believe that it is
present in all anthropoids. It is interesting: to note that it is
poorly developed in the chimpanzee, for in man it occurs only as

438 E. C. MacDowell.

an irregularity. Gray describes such a muscle in man, and says
that Dr. Struthers found it in 7.6 per cent of bodies. Macalister,
Le Double, Wood and Bardeen all found this in about 5 per cent
of cases. Patterson reports its occurrence as ‘‘occasional.”’
Bardeen says it is normally represented by a fascial slip. Hart-
mann describes two insertions of this muscle in the chimpanzee,
one into the condyle, the other into the middle or inner head of
the triceps. He quotes Bischoff as describing in the chimpanzee
an insertion into the fascia over the biceps; in the baboon, an
insertion into the intermuscular septum and the internal con-
dyle. Michaélis found it inserted into the triceps in his orang.
In the gorilla, Hepburn describes this muscle as arising from the
coracoid process.

In the chimpanzee, the Coracobrachialis has two divisions.
Their common origin is from the tip of the coracoid process, and
from the tendon of the short head of the biceps, with which it is
intimately related. The firsc division is inserted into the humerus
between the insertions of the pectorals and the latissimus dorsi.
This is accompanied by a strong tendon and tendinous bands,
which pass through the body of the muscle. At a point 5 em.
from the origin, the main division becomes separated from the
belly of the biceps. This division also has tendinous bands from
which several of the fibers of the biceps take their origin before the
bellies separate. The insertion of this division of the coracobra-
chialis extends from the distal limit of the insertion of the first
division to a point five-sixths of the way down the humerus.
Distally, it jos the medial head of the biceps by an aponeurosis.

In the baboon (fig. 1, C.) the proximal division is highly devel-
oped. It adheres closely to the humerus as a broad flat belly. In
the Macacus, also, this secondary head is well developed. It arises
as in the other forms and immediately leaves the common tendon
to be attached by a fleshy insertion close up under the head of the
humerus, its fibers being perpendicular to the long axis of the
humerus. There is complete separation of the parts. The lower
belly extends straight down to be inserted into the lower third of
the humerus.

From the comparison of all mammalian forms, Wood concludes

The Myology of Chimpanzee and Baboon. 439

that this muscle is typically composed of three parts, all of which
arise from the coracoid process. The upper head is regularly lack-
ing in man, and this seems to be the case in my chimpanzee.
However, it is present in the baboon and in the Macacus. Champ-
neys, Hepburn, and Beddard describes two divisions in the chim-
panzee, while Duvernoy and Vrolick describes one. Hepburn
found elements of all three divisions in his gorilla. Lubosch found
two divisions in a human body (see table I).

TABLE I
ANIMAL REPORTED BY CORACOBRACHIALIS

Chimpanzee Hepburn | 2divisions
Chimpanzee Champneys | 2divisions
Chimpanzee Beddard 2 divisions
Chimpanzee Duvernoy _ 1 division
Chimpanzee Vrolick | 1 division
Chimpanzee Author | 1 division

Baboon Author | 2 divisions
Macacus Author | 2divisions

Gorilla Hepburn | elements of 3 divisions
Orang Primrose 2 divisions

Typical Mammal Wood _ 3 divisions

Man Lubosch 2 divisions as a variation

The Triceps Brachii has but little variation from the muscle in
man. In the chimpanzee, the three heads are intermingled a
short distance from their origins. The long head arises from the
axillary border of the scapula for nearly its whole length, and dis-
tally from the deltoideus and the surrounding fascia. This origin
is semi-cartilaginous. Its belly is fiat and wide before its incor-
poration with the otherheads. It lies between the teres major and
the deltoideus. The long head in the baboon arises by a tendon
closely associated with the muscle fibers from the axillary border of
the scapula between the teres major and teres minor and is lost im-
mediately in the main belly of the triceps; thus it has a triangular
shape. A flat slip of muscle 1.5 cm. wide arises from the axillary
border of the scapula just beyond the distal limit of the origin of
the long head. This slip extends half way down the arm parallel
to the long head of the triceps and is inserted into the fascia cover-

440 EK. C. MacDowell.

ing the main belly of the triceps, just proximal to the insertion of
the lattissimo condyloideus. The whole muscle is characterized
by the close interlacing of the parts. “The Macacus shows a sep-
aration of three parts half way down the arm.

Hepburn found this muscle strongly developed in all anthro-
poids, and Beddard describes it asin man. Bardeen says a fourth
head is frequently found in man as a variation. Le Double re-
ported 20 such cases.

C. Mutscies or THE FOREARM

The Palmaris longus arises in the chimpanzee from the common
tendon from the internal condyle external to the flexor carpi
radialis, and is closely joined to it. It becomes separate 8 cm.
from its origin and is inserted into a slender tendon which is in-
serted into the proximal, radial aspect of the carpal ligament
and the neighboring palmar fascia.

Beddard, Vrolik, Champneys, and Rolleston (quoted by Champ-
neys) found the same arrangement in the chimpanzee. In the
orang, Duvernoy and Michaélis describe it as being very strong.
Primrose found a slip to the thumb from which arose a part of the
abductor brevis. It is wanting in the gorilla (Hartmann, Hepburn,
and Duvernoy). Forster found it well developed in his ‘‘ Papua
neugeborenen.”’ Bardeen saysthat it was oncea third flexor to the
digits (flexor longus superficialis). MceMurrich states that this
muscle seems to have been the first to separate from the common
flexor mass. Adachi reports it. lacking in 20.4 per cent of Euro-
pean men but only in 3.9 per cent of Japanese. Le Double did
not find it in 11.2 per cent of bodies.

The Flexor digitorum sublimis (perforatus) is characterized in
the chimpanzee by the extreme separation of the bellies going to
the different digits. Although closely bound together by fascia,
the bellies can be traced to definite and distinct origins.

A. The division for the index finger arises from the coronoid
process of the ulna and from the tendinous sheath of the flexor
carpi radialis by two groups of loosely bound fibers, which become
at once incorporated into the main belly. The fibers are inserted

The Myology of Chimpanzee and Baboon. 441

in a penniform manner into a tendon which extends nearly the
whole length of the muscle. One third of the way from its origin
to ius insertion, the tendon lies on its radial aspect, the fibers being
inserted upon the ulnar side of the tendon. Beyond this, other
fibers arise from the radial side of the tendon, and those on the
ulnar side twist around so that the tendon is on the ulnar side of
the belly. This gives the musclea digastric appearance. The ten-
don is inserted into the base of the second phalanx, as is the case
inman. All four bellies are inserted into their respective digits in
the same manner. Two fasciculi are given off at the poiat
where this belly is twisted, to join the belly for the third digit.

B. The division for the third digit arises (a) from the tendinous
sheath of the flexor carpi radialis, superficially to the origin of the
first head, by three separate divisions, each 1 cm. wide, at points
2, 4. and 5 em. respectively from the insertion of the flexor carpi
radialis into the internal condyle of the humerus; (6) along the
oblique line of the radius from a point 4 cm. from the distal end
of the bone to a point 6 em. from its proximal end, and from the
brachial fascia neighboring this line. The bundles of the proxi-
mal origin continue down the arm loosely joined. The radial
head joins this tendon obliquely on its radial border. This whole
belly is superficial to the first one, and lies on its radial side.

C’. The division for the fourth digit arises (1) most superficially
from the tendinous sheath of the flexor carpi radialis by three
strap-like divisions (a) 5 em. distal to the external condyle, (b)
more distal, 1 em. broad, (c) distal to (6) 2 cm. broad; a and b unite
but c does not join the other two for two thirds of the way down
the arm: and (2) from the intermuscular septum between it and the
flexor carpi ulnaris, near its insertion into the internal condyle.
All these fibers are inserted in a penniform manner upon the inner
surface of the tendon, which extends half way up the arm.

D. The division for the fifth digit arises from the intermuscular
septum between it and the flexor carpi ulnaris, at a point 6 em.
from its origin. This division is very small and is entirely covered
by the flexor-carpi ulnaris. The tendon islong and slender. From
this it will be seen that the independence of the origins is well
marked, and, with one exception, there is not the slightest con-
nection between any of the main divisions.

442 | EG) Macliaeall

In the right hand, there is a peculiar abnormality in the inser-
tion of the tendon to the middle finger. It is firmly affixed to the
base of the proximal phalanx. The tendon of the flexor profundus
is also inserted at this point, where the fibers of the two ten-
dons are closely interlaced. From here to the base of the sec-
ond phalanx extends a strong tendon, which is so short as not
to allow the first and second phalanges to form an angle greater
than 95°. A second tendon internal to this, extends from the base
of the first phalanx to the terminal phalanx. This tendon is long
enough to give the terminal phalanx free movement and, as there
are afew muscle fibers at its origin, it must have had a slight power
to flex the digit. From their relative positions and insertions, it
seems fair to suppose that these tendons correspond to the normal
tendons of the flexor digitorum sublimis and profundus.

In the baboon, the opposite extreme is found. The sublimis con-
sists of but one belly (fig. 2,8), which takes its origin from the com-
mon tendon attached to the medial epicondyle of the humerus.
The radial origin is not represented even by a fasciculus. The
belly is no longer than any of the other superficial flexors. The
special tendons to the digits do not separate until the common ten-
don has passed the carpal ligament.

In the chimpanzee, Champneys found an additional origin
from the radius only for the division to the second digit; Hep-
burn reports that the tendons for the second and fifth digits had
radial origins, while those for the third and fourth digits had ulnar
origins. Macalister, d’Alix and Gratiolet found no radial origin.
Chudzinski (quoted by Le Double) found that in all races of men
there was a digastric muscle in the deep part of this muscle.

TABLE II

ANIMAL REPORTED BY RADIAL ORIGIN OF FLEX. DIG. SUBLIMIS
Chimpanzee Macalister absent
Chimpanzee Gratiolet-Alix absent
Chimpanzee Hepburn present-supplies digits II-V
Chimpanzee Champneys | present-supplies digits II
Chimpanzee Author present-supplies digits IT, III, V
Baboon Author _ absent
Man LeDouble variable—may be absent

The Myology of Chimpanzee and Baboon. 443

MeMurrich (’03) shows that the sublimis has been differentiated
from a common flexor mass similar to that found in Monotremes
and, little by little, has developed until it includes all che condy-
lar portions of the original muscle. In the chimpanzee, the further
development of a radial origin does not seem to be constant.
Even in man, this origin is variable and may be wanting. Com-
plete separation of the divisions for the digits is also found (Le
Double). (See table IT.)

The Flexor digitorum profundus (perforans) arises in the chim-
panzee (I) (a) from the oblique line of the radius internal to the
origin of the second head of the sublimis, and (b) from the radial
half of the interosseous membrane; it also arises (II) (a), from the
dorsal surface of the ulna along its upper two-thirds by an aponeu-
rotic septum between it and the flexor carpi ulnaris, (b) from the
proximal half of the under surface of the ulna, and (c), from the
ulnar half of the interosseous membrane. The radial head (qa)
supplies a large perforating tendon for the second digit; the fibers
are inserted upon the radial border of this tendon. This belly cor-
responds in its origin to the flexor pollicis longus in man, although
in my chimpanzee, no connection can be found between this divis-
ion and the thumb. However, a fine tendon is found arising from
the annular ligament and from the surrounding palmar fascia,
which is inserted into the terminal phalanx. Such an insertion
would correspond to that of the flexor pollicis longus in man.
As there was no muscle connected with this tendon, it seems that
there could have been no flection of the terminal digit. Le Double
although never himself finding the flexor pollicis longus lacking in
man, cites Gruber, Wagstaffe, and Fromont as having found a
ease each where this muscle was represented by a thin tendon
from the sesamoid bone of the metacarpo-phalangeal articulation
co the terminal phalanx. The tendons for the third and fourth
digits extend far up into the fibers from the ulnar origin. They
show a tendency to divide into threads and two bellies are
closely associated all the way down. Just after leaving the annu-
lar ligament, the tendons are joined by a strong transverse sheet
of tendon 3 em. broad, from which arise two of the lumbricales.
The division for the fifth digit also arises from the ulna as a small

444 E. C. MacDowell.

independent head. Its tendon is divided all the way to the wrist.
The irregular insertion of the tendon to the middle finger in the
right hand has been previously described.

Fie. 2. Dissection of left forearm of the baboon, to show the arrangement of the
flexor digitorum profundus. The tendon of the Palmaris longus has been cut,
and the flexor digitorum sublimis has been pushed over to the right. A, Tendon of
Abductor pollicis longus; B, Biceps; C, Flexor carpi radialis; L, Palmaris longus;
M, Pronator quadratus; P, Flexor digitorum profundus; S, Flexor digitorum sub-
limis; U, Flexor carpi ulnaris; X, Internal condyle of humerus.

The Myology of Chimpanzee and Baboon. 445

The flexor digitorum profundus in the baboon (fig. 2, P) shows
great consolidation. It arises (a) from the oblique line of the rad-
ius as far as the pronator quadratus, (b) from the posterior bor-
der of the ulna, extending from the olecranon process to a point
8 cm. from the distal and, (c) from the common flexor tendon
from the internal condyle of the humerus; the fibers from these
various origins are joined in a broad flat tendon 2 em. wide, which
extends half way up the arm. It is heavily grooved, indicating
the elements of the tendons for the digits, but these tendons are
not differentiated until the tendon reaches the palm. The origin
(a) from the radius corresponds to that of the flexor pollicis
longus, which, as in the chimpanzee, is still undifferentiated from
the profundus. However, a strong round tendon, supplying the
pollex (fig. 3, 7’), is given off from the common tendon just after
passing the ligament.

In a gibbon, Hepburn found an origin of the profundus from the
internal condyle, which, he says, clearly shows the relation between
the two flexors. The condylar segment has become the sub-
limis. ‘The condition in my baboon shows the profundus, in-
cluding fibers which in the higher forms are included in the sub-
limis. The separation of the flexors into individual bellies is
considered, by Bardeen, to be associated with the refinement of
the movements of the fingers. Champneys, Beddard, and Hep-
burn found no tendon to the pollex, but name the division to the
index the flexor pollicis longus, and describe a small tendon given
off from this to the pollex. Testut calls the separated divisions
to the index in his chimpanzee a part of the profundus. This agrees
with MeMurrich’s conclusions that the flexor pollicis longus
should not be said to be missing in monkeys, nor be described as
fused with the profundus, but rather as still undifferentiated from
this muscle. In the chimpanzee, Keith found no tendon to the
pollex in ten out of twenty-five cases; Vrolik describes a tendon
from the flexor brevis digitorum that is inserted into the terminal
phalanx, which seems to take the place of the tendon of the flexor
pollicis longus. Bischoff found no tendon to the pollex in the go-
rilla. Henle, Wood, and Turner (quoted by Champneys) observed
in man a muscular slip from the flexor pollicis longus which sent

446 E. C. MacDowell.

a tendon to join that of the profundus for the index. Michaélis
describes three separate divisions of the profundus in his baboon.
A tendon for the thumb was given off from the head that supplied
the fourth digit. Champney’s description of his baboon corre-
sponds co mine. Testut claims that, with the exception of

Kia. 3. Dissection of the wrist and muscles of the thumb of the baboon, to
show the relations of the tendon of the flexor pollicis longus to the profundus.
The superficial muscles have been removed. A, Tendon of Abductor pollicis
longus; B, Abductor pollicis brevis; D, Abductor pollicis; 7, Provator quadratus;
P, Common tendon of flexor digitorum profundus; 7, Tendon to pollex, corre-
sponding to that of the flexor pollicis longus, in Man.

The Myology of Chimpanzee and Baboon. 447

nycticebus tardigradus, there is no separate flexor pollicis longus in
any of the monkeys. He believed that this is an important dis-
tinction between man and ape; however, he does not believe with
Gratiolet and Alix that its separation in man and blending in apes
is any hindrance to the evolution theory. For he had found in
human examples all the main types of the coalescence of this mus-
cle, as variations. Dally also answered Gratioles by arguing that
if man did not use it, this muscle would soon become undeveloped.
He believed that the development of this muscle has come with
civilization, and that it was far less developed in primitive man.

In the chimpanzee, the Abductor pollicis longus (extensor ossis
metacarpi pollicis) arises from the interosseous membrane, and the
intermuscular septa between it and the extensor carpi radialis
brevis and the extensor indicis proprius. It gives off two tendons:
one of these is inserted into the trapezium, the other into the base
of the metacarpal bone of the thumb. No sesamoid bone is found
in its tendon.

Beddard describes a similar division and insertion in his chim-
panzee. In man, these same conditions are found at times, which
closely connect the hand of man to that of the ape (Le Double).
In a gorilla, Duvernoy describes these divisions as the cubito-sus-
metacarpien and the cubito-sus-trapezien. Hartmann quotes
Bischoff as saying that the long abductor in the orang and in the
baboon is as in man, but that division is found in the gorilla,
chimpanzee, and macacus. It cannot be said that one of these
tendons belongs to the extensor pollicis brevis, for Hartmann, Duv-
ernoy and Vrolik have found an extensor pollicis brevis coexisting
with a ‘‘cubito-sus-metacarpien’’ which was inserted into the
metacarpal of the thumb.

The Ealensor digitt quints proprvus in the chimpanzee arises from
the ulna below the origin of the extensor digitorum communis.
The fibers are inserted into a strong, flat tendon that passes
through a special compartment of the annular ligament, to be
inserted into the fascia covering the second phalanx of the fifth
digit. It seems correct to name this the special extensor of the
fifth digit, since in its origin and in its passage through the liga-
ment, as well as in its insertion, it corresponds to this muscle in
main.

448 E. C. MacDowell.

Beddard says that this muscle was lacking in his chimpanzee, or
else the division of the extensor communis to the fifth digit was
lacking (which seems more reasonable, since he describes a ten-
don to the fifth digit that passed through a special compartment
in the ligament). As will be seen later, the division for the fifth
digit of the communis is present in my chimpanzee, as well as this
special extensor. Champneys found the fifth digit of the right
hand of his chimpanzee supplied only by a slip from the tendon
of the communis. The left hand wasasin man. Vrolik describes
the special extensor as the only tendon to the fifth digit, but Mich-
aélis claims that this is the ulnar belly of the extensor digi-
torum communis that has become entirely separated.

The Hatensor digitorum communis in the chimpanzee is separated
into four bellies half way down the arm. It arises by a strong
tendinous sheath from the external condyle, from the intermus-
cular septa between it and the extensor carpi ulnaris, from the
ulna, radius, and the interosseous septum. In the left hand, two
divisions of this muscle send tendons to the fourth digit. The
more radial and stronger arises from the condyle and is joined: to
the tendon of the third digit by a short tendinous fasciculus. The
other division arises from the whole length of the ulna. A slender
fasciculus arises from the ulnar border of this division 1.5 em.
from the annular ligament. It is provided with a thread-like
tendon which divides 5 cm. from the ligament. The radial half
re-divides close to the metacarpo-phalangeal articulation, and
sends one branch to the tendon for the fourth digit, while the other
rejoins the ulnar half of the tendon from the first division, which
is inserted along with the extensor digiti quinti proprius. This
slip represents the division of the communis for the fifth digit. In
the right hand, there are three separate tendons for the fourth
digit, besides the connection with the fasciculus for the fifth digic.

Vrolik and Macalister found no tendon for the fifth digit in their
chimpanzees. Primrose describes a slip in his orang that corre-
sponds to the one just described, but calls it the extensor digiti
quinti proprius (extensor minimi digiti). In man, the tendon for
the fifth digit is often absent or rudimentary. .

The Hatensor indicts proprius arises in the chimpanzee from the

The Myology of Chimpanzee and Baboon. 449

distal four-fifths of the ulna. It is divided into three parts which
give off three tendons. Two of these are large and are inserted
into the extensor expansion over the dorsal aspect of the proximal
phalanges of the second and third digits respectively, on the ulnar
side. ‘The third tendon is very small and is closely applied to
the tendon for the second digit, from which it becomes separated
1.5 cm. before its insertion into the radial side of the extensor
expansion of theindex. Similar conditionsare found in the baboon.

Beddard describes this muscle in a chimpanzee as having but
one strong tendon and that joining the branch of the communis
for that digit. Mayer found it small or entirely lacking in the
chimpanzee and in the orang. In the chimpanzees of Hartmann
and Macalister, an extra tendon for the third digit was found.
Champneys found such a condition in his Cynocephalus and
Le Double reports its occurrence in man. Duckworth describes
the double extensors to all the fingers (formed by branches of
the indicis proprius and digiti quinti proprius to the third and
fourth digits respectively) as being constant in many apes.
Again, Primrose quotes Huxley as stating that there were origi-
nally two sets of extensors, just as there were two sets of flexors,
and that this is indicated by the dividing of the special extensors
of the second and fifth digits. It may be remembered that. I do
not find a division of the tendon of the extensor digiti quinti
proprius, but I do find a division in the tendon of the fasciculus
of the communis for the fifth digit which could not correspond to
a deep flexor, as it was joined directly to the tendon of the com-
munis.

II. MUSCULATURE OF THE LOWER EXTREMITY
A. Muscies or THE Hip

The Scansorius is present in the chimpanzee. It arises from the
anterior border of the ilium below the anterior superior spine,
and its insertion is into the anterior border of the great trochanter,
together with the gluteus minimus. These two muscles are inde-
pendent for two thirds of the way from their origins.

In his chimpanzee, Beddard found the scansorius well devel-

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, No, 3.

450 E. C. MacDowell.

oped, but not separated from the gluteus minimus. Primrose
found it in his orang, and quotes Hepburn, Huxley, and Owen
as finding it separate inthechimpanzee. Bischoffand Champneys
describe it closely connected to the gluteus minimus. In For-
ster’s ‘‘Papua-neugeborenen”’ this muscle was well separated on
one side. Bardeen says that a special fasciculus from the anterior
margin of the gluteus minimus in men corresponds to the scan-
sorius and is frequently called invertor femoris or small anterior
gluteal. This muscle was discovered in man by Haughton
(Le Double); in the chimpanzee by Troill and in the orang by
Bischoff.

The Gluteus maximus in the chimpanzee arises (1) from the
crest of the ilium by a broad tendinous sheet from the sacrum and
coecyx and (2) from the tuberosity of the ischium, anterior to
the insertions of the semi-membranous and biceps femoris. The
fibers from the first origin converge towards the head of the femur,
over which they pass to join a tendon, which is inserted upon the
external aspect of the femur, half way down. The insertion of the
second part, ischio-femoralis (Duvernoy), is into the tendon just
described along the femur. Some of these fibers continue down the
leg all the way to the external condyle, and are connected to the
vastus externus by a septum.

Vrolick and Beddard describe the same structure in their chim-
panzees. Duckworth reports an insertion largely into the pos-
terior surface of the femur in the Lemuroidea. In the Cercopithe-
cidea, the femoral insertion is very small. Hartmann fourd a
tendon of insertion in Anthropoids descending far down towards
the knee. I find a very narrow insertion in my macacus. Wie-
dersheim states that the special development of this muscle is
peculiarly human.

B. Muscies oF THE LEG

In the chimpanzee, the Plantaris arises from the tendon of the
lateral head of the gastrocnemius and from the lateral line of the
bifurcation of the linea aspera, just proximal to the origin of the
head of the gastrocnemius. The belly is slender, 6 cm. long and
passes into a slender tendon that lies beneath the medial head of

The Myology of Chimpanzee and Baboon. 451

the gastrocnemius to be inserted into the medial border of the
Tendo Achillis. In comparison with man, this belly is longer.
The tibia of the chimpanzee is 20 em. long, that of man from 45
to 50 em. (Gray). In both cases the plantaris is 6 em. long.
Beddard found this muscle present in the chimpanzee, while
Champneys did not. Michaélis says it was well developed in
the baboon but absent in the chimpanzee and orang. It was lack-
ing in the orang of Primrose. Duckworth says it is never found
in the gorilla, but frequently in the chimpanzee; it is normal in
the Cercopithecide and Lemuroides. Hartmann found it in his
chimpanzee, although he says Bischoff and Brihl at first denied
its existence. He did not find it in either the gorilla, the orang,
or the gibbon. Kohlburgge reports its absence in the chimpanzee
in 43 per cent of cases and Loth in 45.7 per cent. Le Double
found rather frequent cases of the absence of this muscle in man.
However, in colored races its absence is rare. The plantaris,
as well as the palmaris its homologue in the arm, exemplifies a
degenerating muscle. Wiedersheim (p. 109) states that ‘‘as
an original flexor (the plantaris) must have begun to degenerate
from the time the plantar fascia became secondarily attached to
the calcaneus, and helped in the formation of the foot as the latter
became transformed into a supporting organ.’ See table III.

TABLE III

ANIMAL | REPORTED BY OCCURRENCE OF PLANTARIS

Chimpanzee Beddard | present

Chimpanzee | Hartmann present

Chimpanzee | Champneys absent

Chimpanzee | Michaélis | absent

Chimpanzee Duckworth | frequently found
Chimpanzee Author present

Chimpanzee Kohlbrugge absent in 43 per cent
Chimpanzee Loth absent in 45.7 per cent
Man Le Double frequently absent
Orang Duvernoy absent

Orang Michaélis absent

Orang Hartmann absent

Orang Primrose absent

Baboon Michaélis present
Cercopithecidae Duckworth present

452 E. C. MacDowell.

The Soleus is the largest muscle that enters the Tendo Achillis,
It has no tibialhead. This is true for the chimpanzee, the baboon,
and the macacus. In the chimpanzee, the fibers are inserted all
along the inner surface of the Tendo Achillisas far asits insertion.
The tibial head was absent in the chimpanzee and in
the baboon of Michaélis. He said that it probably is included in
the gastrocnemius lateralis when itis said to belacking. MeMurrich
(04) suggests that the plantaris represents the plantaris pro-
fundus III of Amphibian, while Hisler had regarded it as a deriva-
tive of the gastrocnemius lateralis.

All the bellies joined to the Tendo Achillis are separate from
their origins to their insertions, a condition frequently found in
man (Bardeen). Muscle fibers are incorporated with the Tendo
Achillis for its whole length. Michaélis found this to be the
case in his chimpanzee. The calf is slim, presenting nomore thaa
a slight swelling. This is in sharp contrast to the well developed
human calf, which occurs as a result of man’s mode of locomotion.

The Tibialis anterior has two bellies in both chimpanzee and
baboon. In the former, it arises from the anterior external sur-
face of the tibia along its proximal two thirds. One third the
distance from its origin, the muscle divides. The internal por-
tion is the larger; its fibers are inserted into the deep aspect of
a strong, round tendon, which is inserted into the internal palmar
aspect of the distal third of the cuneiform bone. The external
portion, whose tendon is half as large as the first one, is inserted
into the internal palmar aspect of the base of the first metatarsal
bone. This separation of the tendon, inserted partly on the cunei-
form, and partly on the metatarsal, has been found as a variation
in man (Le Double). In his baboon, Michaélis, found that the
tendon divided just before being inserted upon these bones. In
his chimpanzee, it divided at the origin. Beddard and Vrolik
described such a separation. Owen describes the tendon to the
metatarsal bone as a separate muscle with no homologue in man.
Ruge says the division is well known and that Bischoff has found
three parts.

The Peroneus longus in the chimpanzee arises from the head of
the fibula, from the intermuscular septum between it and the

The Myology of Chimpanzee and Baboon. 453

extensor digitorum communis and from the internal border of the
fibula down to a point 4 em. from the distal extremity. Itis
attached to the first metatarsal bone alone. Primrose describes
a double attachment in the orarg.

The Peroneus brevis arises from the lower half of the fibula
between the excensor digitorum communis and the peroneus lon-
gus. It is connected with these muscles by septa. Its tendon
is inserted into the tip of the tuberosity of the fifth metatarsal
bone. From this point, a tendonous slip is continued along the
phalanges to be inserted cogether with the tendon of the extensor
digitorum communis into the second phalanx. Owen, in an
orang, and Vrolik and Beddard, in the chimpanzee, found this
same arrangement. This extended tendon corresponds to the
peroneus digiti quinti, a muscle found in the monkeys, which
in man is frequently found fused with the peroneus brevis, so that
only its tendon of insertion is apparent (Bardeen). Poirier
(Morris’s ‘““Anatomy”’ p. 459) considers that all the peroneals
are varieties of this muscle (peroneus digiti quinti), which in its
most simple form ‘arises from the distal fourth of the fibula and
is inserted by a tendon into the fifth toe.”

The Haiensor digitorum longus in the chimpanzee arises from
the tibia and fibula, from the interosseous membrane and the
intermuscular septa between it and the tibialis anterior and the
peroneals. The tibial belly separates half way down the leg. The
muscle fibers are inserted into a tendon, which, after passing
through the second tarsal ligament, divides, sending one branch
to the second, the other to the third digit. The portions supply-
ing the fourth and the fifth digits do not separace from each other
until after passing the tarsal ligament. The tendon for the fifth
digit is not free from muscle fibers until it passes the second liga-
ment. This is much like the conditions in man, where there are
two main bellies, each of which divides just after passing the
ligamert. Owen, in his orang, found no tendon to the secoad
digit. Beddard found all four tendons, but the one to the second
digit was small.

The Flexor digitorum longus (Flexor digitorum tibialis) arises
from the whole length of the tibia (fig. 4, D); proximally it anas-

454 E. C. MacDowell.

tomoses with the popliteus, distally it leaves the bone 4 em. from
its end. The tendon is flat. It does not become entirely free
from muscle fibers until after it divides. One tendon is sent to

Fia. 4. Dissection of the plantar surface of the right foot of the Chimpanzee.
To show the arrangement of the flexor tendons for the digits. .A, Adductor halluci
Bb, Flexor digitorum brevis; D, tendons of flexor digitorum longus; F, Flexor
hallucis brevis; H, tendons of Flexor hallucis longus; M, Abductor digiti quinti;
Y, cut ends of Abductor hallucis brevis.

the second digit, the other to the fifth. In the baboon (fig. 5,

D) the origins and insertions of this muscle are similar.
Champneys found the tendon of this muscle in his chimpanzee

fused with the flexor hallucis longus, otherwise it would have

The Myology of Chimpanzee and Baboon. 455

supplied only the second and fifth digits as it does in my animal.
In the baboon, he describes the same relations. In Beddard’s
chimpanzee and orang, this muscle sent perforating tendons to
the third, fourth, and fifth digits. In Keith’s chimpanzee this
muscle supplied the second and fifth digits. Forster said that
this muscle supplied the fifth digit in his “Papua neugeborenen”’
but also united with the flexor hallucis longus to form a tendon,
which supplied the other four digits. Owen found three tendons
in his orang: first to the second digit (perforans); second to the
fourth digit (perforatus); third to the fifth digit (perforans). In
my chimpanzee, there arises from this tendon at its division a
belly which supplies the perforatus tendon to the fourth digit.
However, as no fibers from the common belly are included in this
one, I have decided to include it with the flexor digitorum brevis
rather than with this muscle. (See table IV.)

TABLE IV
INSERTION OF
ANIMAL REPORTED BY

FLEX. DIG. TIBIALIS | FLEX. DIG. FIBULARIS
Chimpanzee Champneys *digits II, V | digits I, IIt; IV
Chimpanzee Beddard digits III, IV, V | digits I, II
Chimpanzee Keith digits II, V digits I, III, IV
Chimpanzee Michaélis digits II, V digits I, III,1V
Chimpanzee Duvernoy - no tendon to hallux
Chimpanzee Author digits II, V Va aieris Ie Te TV,
Baboon Champneys digits II, V | digits I, III, IV
Baboon Michaélis digits II, V | digits I, III, IV
Baboon Author *digits II, V | digits I, JOE IW
Orang Beddard digits III, IV, V | digits I, II
Orang Owen digits II, IV, V no tendon to hallux
Orang Hartmann | notendon to hallux
Orang Primrose | no tendon to hallux
Simiadee Dobson digits II, III, IV

The Flexor hallucis longus (flexor digitorum fibularis) (fig.
4, H) arises from the shaft of the fibula extending from below the
insertion of the soleus to a point 6 cm. from its distal extremity,

456 BE. C. MacDowell.

from the interosseous membrane, and from the intermuscular
septum between it and the peroneus longus. The tendon is
large and stronger than that of the preceding muscle. It is
free from muscle fibers before reaching the plantar surface. Here
it divides in half. One branch, passing under a ligamentous
band on the inner palmar aspect of the base of the first metatarsal
bone, is inserted into the terminal phalanx of the first digit. The
other branch divides, sending perforating tendons to the third and
fourth digits. The baboon presents the same arrangement
(fig. 5, H), excepting in the tendon to the first digit, which arises
as much from the flexor digitorum longus as from the hallucis.
Thus, the two tendons are firmly united at this point, allowing
far less independence in the flection of the terminal phalanges of
the thirdand fourth digits. Michaélis describes a similar arrange-
ment in his chimpanzee and in his baboon. Owen, Hartmann,
and Primrose describe no tendons to the hallux in their orangs.
Duvernoy found the same lack in a chimpanzee. Dobson says
that this muscle supplies the three middle digits in the Simiade.
The main variations that have been described refer to the
relation of the tendon to the first digit, the insertion into third
and fourth digits being constant. (See table IV.)

The Flexor digitorum brevis will be described as the muscle whose
bellies supply the perforated tendons to the digits. In the chim-
panzee (fig. 4, B), it arises from the calcaneum in close relation
to the abductor hallucis brevis and from the tendon of the flexor
digitorum longus. The tendon of the second digié arises from the
main belly from the calcaneus. The tendon from the third digit
arises from a belly that also takes its origin from the “longus”
tendon. A few of the fibers from this insertion extend so far
back along the tendon that they become continuous wich those
from the belly of the ‘“‘longus” that have crept far down the ten-
don. However, as most of the fibers arise from the tendon just
as it divides, it seems fair to have called this belly a separate mus-
cle and not to have described its tendon as one from the flexor
digitorum longus. The perforatus tendon for the fifth digit is
lacking. Hartmann considers the division arising from the ten-
don of the flexor digitorum longus as a part of that muscle. He

The Myology of Chimpanzee and Baboon. 457

describes the flexor brevis digitorum in the gorilla as displaying
perforated tendons for the second and third digits, while those for
the fourth and fifth digits are provided by the flexor digitorum
longus.

Fia. 5. Dissection of the plantar surface of the right foot ot the Baboon, to show
the distribution of the bellies of the flexor digitorum brevis. A, Adductor hallucis
B,‘ Flexor digitorum brevis; D, tendons of Flexor digitorum longus; F, Flexor
hallucis brevis; H, tendons of Flexor hallucis longus; L, Lumbricals; P, Abductor
hallucis brevis.

458 E. C. MacDowell.

The baboon (fig. 5, B) shows a greater complexity in the arrange-
ment of the bellies of the flexor digitorum brevis. In general, it
arises from the plantar fascia covering the caleaneus, and from
the tendons of the flexor digitorum longus and hallucis longus.
The first belly arises from the palmar fascia and is situated near
the heel. Its tendon is long and slender and is inserted as a per-
forated tendon into the second digit. The second belly arises
from the plantar fascia further from the calcaneus than the first.
It swells out as the first one tapers down to receive its tendon.
Before its insertion into the third digit the tendon from this
second belly is connected by a ladder of muscle fibers to the per-
forating tendon of the flexor hallucis longus which lies directly
below this. The third belly comes partly from the tendon of the
flexor digitorum longus for the fifth digit, and partly from the
tendon of the hallucis longus for the fourth digit. ‘This belly
lies far out from the heel and thus has but a short tendon. The
fourth belly has its origin on the tendon of the flexor digitorum
longus and in the intermuscular septa between it and the neighbor-
ing muscles. This belly is nearer the digits than the third belly.

The flexor digitorum brevis acts more as a set of accelerator
for the long flexors than as an independent muscle from the cal-
ecaneus. However MeMurrich (’07) says that this is anintrinsic
muscle, which is homologus with the main mass of theflexor
brevis superficialis str. superficiale. The greater part of its fibers
arise from the tendon of the flexor longus. It may be noticed
that this corresponds to the condition found in the chimpanzee,
although there was no origin from an hallucis longus tendon.

Champneys describes a muscle in his chimpanzee which is even
more complicated than the one I describe. Owen and Beddard
found more simple ones in their orangs. The latter found but
three tendons. Primrose found four, of which the tendon for the
third digit was thestrongest; those to the fourth and fifth toes were
extremely small. The divisions for the fourth and fifth digits
arose as in my chimpanzee, largely from the tendon of the flexor
digitorum longus.

All the short muscles of the hallus and pollex are found in both
forms with but slight variations from their human homologues.

The Myology of Chimpanzee and Baboon. 459

In the chimpanzee, the flexor pollicis brevis is-very closely joined
with the oblique head of the adductor pollicis. The opponens is
but feebly developed. In both forms the transverse head of the
adductor pollicis is well developed and extends well over into the
palm.

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Apacut, B. ‘‘Anatomische Untersuchungen an Japanern.”’ III. Muskelvarie-
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BarDEEN, C. R. Morris’ Anatomy, 4 ed., p. 369.
1907

Beppparp, FRANK E. Contributions to the Anatomy of the Anthropoid Apes.
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Brscuorr, Th. L. W. Beitraige zur Anatomie des Gorilla. Abhandl. der math.
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CuampPNneEys, F. On the muscles and nerves of a chimpanzee and acyanocephalus

1872 anubis. Journ. of Anat. and Phys., vol. 6.

Cuapman, H. C. On the structure of the orang-outang. Proc. Acad. Nat. Sci.,
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CunnineHaM. A textbook of anatomy.

1905

Dautuiy, M. E. L’ordredes primatesetletransformisme. Bull. dela Soc. d’Anthro-
1868 pologie de. Paris. 4° série, III, p. 696.

Dosson, G. E. The homologies of the long flexor muscles of the feet of mammalia,
1882-83 with remarks on the value of their leading modifications in

classification. Journ. of Anat. and Phys., vol. 17, p. 174.
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1898

1904 Morphology and anthropology. Cambridge, Eng.
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Gray A text-book of anatomy. i
1870

HarTMANN, R. Die menschenihnlichen Affen und ihre Organisation in Vergleich
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Hersurn, D. The comparative anatomy of the muscles and nerves of the superior
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460 E. C, MacDowell.

Huntineton, G.S. Present problems of myological research and the significance
4 £

1902-03 in classification of muscular variations. Amer. Journ. of Anat.,
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Keritu, A. Notes ona theory to account for the various arrangements of the flexor
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«

Accepted by the Wistar Institute of Anatomy and Biology January 2, 1910. Printed June 21, 1910.

STUDIES ON THE CHIEF VEINS IN EARLY PIG
EMBRYOS, AND THE ORIGIN OF THE VENA
CAVA INFERIOR

DAVID M. DAVIS
From the Anatomical Laboratory, Johns Hopkins University

WITH THREE TEXT FIGURES AND SIX PLATES

INTRODUCTION

The vena cava inferior has always excited a good deal of interest,
on account of its unusual history and great importance. When
its channel first becomes patent, a great change takes place in the
circulatory system of the embryo, and in a very short time—a few
hours, in fact—the new vein is carrying practically all the blood
from the posterior part of the body. Owing to this circumstance,
the vein, by the time the older investigators observed it in stained
sections, had already attained considerable size. It was thought,
therefore, that an attempt to correlate its earliest origin with the
new theory of capillary or capillary plexus anlagen for the vessels,
as recently presented by Dr. Evans, would be of interest. The
notions formerly held will be understood from the following short
historical sketch.

HISTORICAL

In 1888 the result of the work that had been done upon the
development of the cava to that time, was summed up by Lock-
wood and Humphry in these words—‘‘It (the vena cava inf.)
commences a little in front of these organs (kidneys) by two blind
symmetrical rootlets (the ‘‘renal’’ veins) situated at the base of
the mesentery and in front of the aorta. After communicating
with both posterior cardinal veins, especially with the right, it
runs forward by the side of the aorta, and enters the liver and runs
through the substance of that organ to empty into the r. vitelline
vein in front of the junction with theductusvenosus arantii. The

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10. No. 4.

462 David M. Davis.

vena cava enters the liver through a junction—the caval junction
—which that organ previously acquires with the tissues at the
base of the mesentery.” This is practically the description
given by Kolliker, and His seems to confine his attention to the
liver veins, commenting only sparingly upon the vena cava. It
is noticed that little effort is made to explain the exact means by
which the changes take place.

The next important writer on the subject was Hochstetter,
whose first work appeared in about 1888, and covered a period of
many years. The technique of the time made certain mistakes
unavoidable, but his descriptions of the morphological changes
occurring in the already formed veins can scarcely be surpassed.
Regarding the origin of the cava, he practically reversed the proc- —
ess, speaking apparently, of it as ‘growing down”’ from the liver
into the Wolffian ridge, and there receiving bilateral veins drain-
ing the kidneys and Wolffian bodies.

As is well known, the veins of the very young embryo are quite
symmetrical. There are three systems—the vitelline, the umbil-
ical, and the cardinal, each consisting of two bilaterally symmet-
rical veins, the three of either side uniting at the duct of Cuvier to
form the sinus venosus. For reasons as yet unknown, the stom-
ach shifts to the left in development and the liver confines itself
mainly to the right side. The stomach, according to F. T. Lewis,
presses upon and obliterates the left ala pulmonalis (Ravn) where
it extends into the future abdominal cavity, while the liver
approaches the corresponding structure upon the right, and
appropriates it by sending first capillaries, then hepatic tubules,
into it. This bridge between liver and mesenchyme was called by
Hochstetter the Hohlvengekrése, and may be designated as
‘“‘caval mesentery’’ (Lewis). See fig. 2. Hochstetter’s view was
that the cava grew down through this caval mesentery, and gave
rise to three branches—more properly roots. One ofthese extended
caudally upon the right Wolffian body, the other two caudally and
cranially, respectively, upon the left Wolffian body. These two
roots upon the left united into a common stalk before crossing,
ventral to the aorta, to join their fellow of the right. (These veins
were called the vv. revehentes posterior by Hochstetter, and are

Veins in Pig Embryos. 463

identical with the bilateral renal veins of His, and the subear-
dinals of Lewis.) Communications were set up through the sub-
stance of the Wolfhan body with the cardinal veins; especially the
right; later with the kidneys, and so the ground plan of the adult
cava was laid. For more detailed accounts of these conditions as
given by Hochstetter, one is referred to his own articles and. to
various anatomies and embryologies.

In considering these points, we must not forget Zumstein,
whose work, criticised adversely by Hochstetter, has enjoyed little
recognition. He studied rabbit and mole, as well as human
embryos, and while his reconstructions are diagrammatic in the
extreme, he observed that the subcardinals exist before the vena
cava, and communicate with the liver veins by small vessels.
(“‘Gegen der Kopfende der Urniere treten beiderseits medial und
ventral von den Cardinalis kleine Venenlumina aus, welche mit
den Kardinal-Venen sich verbinden. Die rechten lassen sich
an die Lebergefaisse heran verfolgen. Sie sind noch kapillar.

In der Leber selbst tritt noch kein deutlicheres Gefass-
lumen hervor, dass man als cava inferior ansprechen kénnte.”’
Anat. Hefte, I. 1898, 8. 311. The quotation, given also by F. T.
Lewis, concerns an embryo mole, 3 mm. in length). As Lewis
observes, he did not appreciate the point, and did not push it. It
will be seen by this time that the question is almost entirely one of
interpretation.

It remained for F. T. Lewis (01) to describe the subcardinal
veins as such. He saw that they antedated the cava, and that
they approach very close to the venous spaces of the liver at the
time when the cardinal system is ‘‘tapped by the hepatic.”’ He
did not enter into details as to the exact manner in which the tap-
ping took place, except to remark that ‘‘the right subeardinal and
hepatic sinusoids approach one another and unite, thus forming a _
new access to the heart.”

Dating from Thoma’s paper upon the vascular area of the chick,
a new idea of the development of vessels has arisen—namely, that
they develop from pre-existing capillaries or capillary plexuses.
Thus every advance of the vascular system is preceded by a skir-
mish line of growing capillary buds. The work of Aeby and of

464 David M. Davis.

Baader, as early as 1866, had shown the earliest form of this idea.
Although they stated their belief that vessels arose from capil-
lary nets, adverse criticism prevented the adoption of this view
for many years. It was advanced by Mall in 1898 and again in
1905, in his work upon the liver. Miller and Rabl have also
supported this view, and lately the work of Evans upon this
subject has been very convincing. His résumé will give addi-
tional information on the subject. (Anat. Record, 1908, p. 411).

PRESENT INVESTIGATION

When the time is ripe for the formation of the vena cava, the
situation is such that, for its establishment, it is only necessary for
a channel to penetrate a very narrow bridge of tissue between two
already functioning venous trunks. These two trunks are, first,
a branch of the hepatic vein, and second, the right subcardinal
vein. To show that this penetration takes place through the
medium of capillary outgrowths, is a purpose of this study.

The material used consisted of pig embryos. The following
table gives data concerning the specimens reconstructed, and the
numbers by which reference is made to them.

Table of embryos used in this work

THICKNESS OF
SECTIONS IN
MICROMILLIMETERS

NO. OF | LENGTH OF
EMBRYO EMBRYO IN INJECTED BY | INJECTION MASS STAIN
(MILLIMETERS)
i | . .

73) 0) as eee 7.5 Dr. Evans. | Indiaink ~—-50 Haematoxylin
2302 ee sae 8. Dr. Evans. | India ink 100 None

23090, Seas 9.5 Dr. Evans. India ink 30 Alum Cochineal
5} Vere amet 5 illo, 74 writer | India ink 100 None

The reconstructions were made by the profile method, in which
tracings upon transparent paper are superposed. ‘This gives a
flat reconstruction, from which the drawing is worked up.

A number of other specimens, either in the collection of the
Anatomical Laboratory of the Johns Hopkins University, or
injected for the purpose, were examined in less detail.

Veins in Pig Embryos. 465

Fig. 1. Embryo 8m.m. No. 2302. Sagittal section. 1004. Showing one
of the subcardinal-portal anastomoses. The veins are ruled horizontally, the
arteries in white, the tissues dotted. At the left is seen one of the small aortic
mesonepbric branches.

It has been stated above that the vena cava inferior is formed
through an inosculation between one of the posterior hepatic veins
and the right subeardinal vein. This needs no proof, having been
shown clearly by Lewis. An idea of the topography of the viscera
at this time will be had by reference to Plates I and II, which
represent a 7.5 mm. embryo, and where, it will be seen, the liver is
very small, and the subeardinals are fully formed. In spite of the
undoubtedly distinct character of these veins, no channels are yet
to be made out entering them from the mesentery. This would

seem to contradict the theory of Lewis concerning their origin.
_ They have not, however, reached their full extent in an anterior
direction, and for that reason approach nowhere close to the liver.
Notwithstanding this fact, there is open communication between
the subeardinal system and the hepatic system. A channel is seen

466 David M. Davis.

extending from the left subcardinal into the omphalo-mesenteric
vein (Plate II). This channel lies in the mesentery, and extends
in a ventro-mesial direction. It apparently has no relation what-
ever to the future cava, and seems to be a representative of a more
or less numerous system of channels in this region. A later stage
may be seen in Plates III and IV, showing an 8 mm. embryo.
Here the channels are multiple, and pass from both subeardinals
into the omphalo-mesenteric vein. One of them is seen in section

in fig. 1. The one figured here is of almost capillary dimensions. °

In this specimen, similar vessels are seen in the mesentery caudal
to the inferior mesenteric artery, and joining the subcardinal sys-
tem to the inferior mesenteric vein. AIl these channels appear
later to be completely obliterated, and take no part in the forma-
tion either of the vena cava, in the upper part, or of the veins of
Retzius, in the lower part. Their significance is conjectural,
although they seem to convey a considerable amount of blood to
the liver. I have been able to find no previous reference to them.
It is interesting to note that, in the adult human, veins may pass
from the duodenum directly to the vena cava, as a rare anomaly.

The venous changes in the liver during its growth are well de-
scribed by Hochstetter, whose work has been amplified by recent
writers. The growth of the liver itself, in the region with which
we are concerned, is well discussed by Lewis, whose conclusions
have been referred to. The area designated in Plate III, as ‘‘Site
of Vena Cava Inferior, ’’is shown as it appeared in section in fig. 2.
One sees the liver just after it has touched and fused with the
‘‘caval mesentery.’’ At the top of the drawing are the liver cells
and liver capillaries, growing downward into the caval mesentery.
In advance are capillary sprouts—extending out toward other
capillary sprouts from the somatic area. To understand the
origin of these second sprouts, another system must be studied.

In a 7.5 mm. embryo, Plates I and II, the subeardinals, at their
anterior ends, are still in the growing state. Numerous fine,
irregular capillaries extend forward, both in and beyond the Wolf-
fian body. As they pass beyond the confines of the gland, they
grow into the adjacent tissue, which is, on either side, the abdom-
inal extension of Ravn’s ala pulmonalis—on the right side we may

ee)

Veins in Pig Embryos. 467

o

ie. aby? Ba

Gtom ecylye
of Mesonephros.

AORTA

Fig. 2. Embryo 8 mm. No. 2302. Sagittal section. 100”. Showing the
capillary sprouts in the caval mesentery. The veins are ruled horizontally, the
arteries in white, the tissues dotted. Within the liver the capillaries are
sinusoidal in character.” The capillaries of the more ventral parts of the liver
are not drawn in.

b)

speak of it as the ‘‘caval mesentery.’”’ Other capillaries also grow
into these ridges—across the front of the aortic roots from the two
posterior cardinal veins, from the sinuses of Cuvier, and from the
bronchial veins in the developing lungs. At this point the sym-
metrical development ceases, and the two ridges have very dif-
ferent histories. On the left, the stomach lies upon the ala of that
side, but no fusion occurs, and the ridge later disappears. The
vessels of the stomach wall and oesophagus anastomose through
the dorsal mesentery of the oesophagus with the anterior branches
of the subeardinals. This suggests the oesophageal collateral
path in portal obstruction. On the right, however, the ridge
remains, as the ‘‘caval mesentery’’—consequently the branches
of the r. subeardinal draining it are somewhat larger than those on
the left. This is the condition seen in Plates III and IV.
Depicted in Plate IV are the anastomoses between this cephalad
extension of the subeardinal and the stomach veins; and in both

468 David M. Davis.

plates vessels extending still farther and draining the oesophagus
and other tissues between the two aortic roots. This area about
corresponds with the posterior mediastinum in the adult. In
Plate III, the slightly greater size of that portion of the right sub-
cardinal draining the caval mesentery is apparent. Now, return-
ing to fig. 2, we can understand the situation. The factors are,
first, the hepatic capillaries reaching down into the caval mesen-
tery, draining their blood into the hepatic veins, vena hepatica
communis, and sinus venosus; and, second, the subcardinal capil-
laries, pushing out through the caval mesentery toward the liver.
The consequence is obvious; growing toward each other, they
come into contact, and inosculate. It was at this period of devel-
opment that Zumstein made his observation—and Plate V shows
the* condition when the new channel has become somewhat
enlarged. The sectional appearance is shown in fig. 3, where it is
seen that hepatic tubules still obstruct the caval lumen to some
extent. The changes at this period, both progressive and regres-
sive, are very rapid. The division of the posterior cardinals into
anterior and posterior portions has already commenced. It soon
becomes complete (Plate VI), all the blood from the posterior por-
tion of the body pouring through the new channel. The enlarge-
ment is rapid, the condition shown in Plate VI (16.2 mm.) being
fully reached in a remarkably short time—about two days; a
closer estimate of the period of change is difficult to arrive at in
the case of pigs. In Plate V (9.5 mm.) the ultimate course of the
complete cava is well seen—it is formed by the subcardinal for
about half the length of the mesonephros; from there caudad it is a
part of the right posterior cardinal. The designations of pars
hepatica, pars subeardinalis, and pars cardinalis, are self-explana-
tory. Of course, the relative length of the pars subcardinalis
becomes greatly reduced as the liver grows downward and the
thorax expands.

The connections of the veins of the caval mesentery, which aided
in the formation of the cava, remain as they were originally—as a
result we have a good sized vein flowing from the oesophageal
(posterior mediastinal) region into the cava just where it bends
ventrally to enter the liver (Plates V and VI). The network of

S

a, =e

i

Veins in Pig Embryos. 469

Pre-aortic
\ Capillaries

Ventricle

Coelom

. A.Subclavia
Dextra

Coelom

Glomerular
spaces of Wolffian
body.

Fie. 3. Embryo 9.5 mm. No. 2309. Sagittal section. 304. Showing the
lumen of the newly-formed vena cava inferior still partly obstructed by liver
cylinders. Just ventral to the aortic root are to be seen the forward prolongations
of the subeardinal communicating with the peri-oesophageal plexus.

veins surrounding the oesophagus has been called in this paper the
peri-oesophageal plexus. In tracing the history of this peri-oeso-
phageal plexus, it will be noted, first, that, while the subcardinal-
portal anastomoses mentioned above, which lie caudal to the cir-
cum-intestinal rings of the portal system, become completely
obliterated before the vena cava appears; second, these other
anastomoses, by way of the peri-oesophageal plexus and gastric
veins, remain patent during the entire period, and do not show any
special enlargement. This would indicate some inherent advan-
tage in the route through the caval mesentery. These connections
through the gastric veins are also noteworthy as the forerunners of
the vessels making up the gastro-oesophageal collateral channel
in portal obstruction.

It would, then, appear that the vena cava is no exception to the
rules governing the development of other vessels—having its
anlage in capillary sprouts at the critical point in the caval

470 David M. Davis.

mesentery. Since the subcardinal takes so prominent a part in
the synthesis of the cava, its own origin must be of interest.
Lewis spoke of it in rabbits as the result of longitudinal anasto-
moses between small veins entering the cardinal from the mes-
entery and mesonephric tributaries. This must be disputed, at
least in the pig, since in a number of early injected specimens,
the writer has seen distinct subeardinals, having no tributaries
whatsoever from the mesentery. These tributaries appear later.
At the earliest stage, the mesonephros is seen filled with a net-
work of capillaries springing from the cardinal vein; soon some
of them ventral to the mesonephric arteries dilate and form the
longitudinal channel of the subcardinal. The paper of Grafe,
written of the chick’s mesonephros, bears eloquent witness on
this point, even though he supposed some of the vessels to arise
in situ. Thus we have the history of the caval channel arising
everywhere from capillary anlagen. At later stages, numbers of
channels are formed in the same way in the mesonephros. An
example of them is a great ventro-lateral vein, appearing in the
different drawings. It is seen in pigs as small as 7 mm., and draws
blood from the periphery of the gland at its ventro-lateral angle.
It retains connections with the anterior portion of the posterior
cardinal vein until a very late stage, when, with the regression of
the Wolffian body it disappears, taking no part in the formation
of any important permanent vein.

The manner of origin of the various trunks of the vascular
systems has long interested embryologists, and twe sorts of expla-
nation have been advanced: according to one idea, arteries and
veins grow out as such to their end territories, and it will be
recalled in this connection that Hochstetter spoke of the vena
cava as “growing down”’ into the region of the Wolffian body.
According to the other idea of the development of the vascular
system, arteries and veins exist originally in the form of simple
capillaries, usually in the form of a typical plexus. rom this
netlike or plexiform condition the trunks of the adult are differ-
entiated. The latter idea, suggested by Aeby and Baader, has
recently been proven to be the correct one in the history of vari-
ous vessels, Evans having been able to show that the very largest

Veins in Pig Embryos. A471

trunks, such as the pulmonary, the carotid, and the limb arteries
can be traced to their origin from a capillary plexus. From the
same injections Miss Smith has shown clearly that the thoraco-
epigastric vein is formed from certain early body wall capillaries.

It is evident that the final act in the formation of the vena
cava inferior furnishes another clear instance of a capillary plexus
ancestry, for this portion of this great vessel. It has already been
pointed out that Zumstein deserves the credit of first having seen
these capillary connections, which are the precursors of the cava
here, and more recently F. T. Lewis has described and figured
the invasion of the caval mesentery by hepatic sinusoids. No
one, however, has interpreted these phenomena in the cava’s
development in accordance with the theory of the development
of the vascular system from capillary plexuses. These injections
show conclusively that here we are dealing with the fusion of
two capillary plexuses, one, as already mentioned, from the he-
patic vessels, the other from the cephalic portion of the right sub-
cardinal vein; and the writer has chanced upon the stage in which
the fusion has just taken place, and in which, consequently,
for the first time a complete vascular path is afforded from the
subeardinal to the hepatic channels.

CONCLUSIONS

The conclusions drawn from this study are as follows:

1. Open connections exist between the subcardinal veins and
portal system at an early stage, but they reach their maximum
and are obliterated before the vena cava is formed.

2. At the time that subecardinal capillaries form the anlage
of the cava in the region of the caval mesentery they have pre-
viously proliferated in a cephalic direction surrounding the ceso-
phagus; consequently before the cava is formed this peri-cesphageal
plexus drains into the cephalic tip of the v. subeardinalis, and
with the formation and rapid enlargement of the cava it naturally
comes about that this plexus is now drained into the latter vessel.

3. At the critical period in the history of the vena cava, its
channel is definitely formed by the fusion in the caval mesentery of
capillary sprouts from the hepatic and subeardinal vessels, respec-

472 David M. Davis.

tively. Circulatory conditions, as yet not clearly understood,
almost immediately force upon these minute capillaries a speedy
and tremendous enlargement, so that they quickly constitute
the vena cava inferior in this region.

My thanks are due Professor Mall, for the suggestion of this
research as well as for his generous supervision. I also wish to
express my appreciation of the constant interest and invalu-
able suggestions and criticisms of Dr. Evans.

BIBLIOGRAPHY

AgeBy. 1871 Der Bau des menschlichen Ké6rpers.
Baaper. Uber die Varietiiten der Armarterien des Menschen, Inaug. Diss. Bern.
1866
Euze, C. 1907 Beschreibung eines menschlichen Embryo, Anat. Hefte.
Evans, H. M. On the development of the aort, cardinal and umbilical veins,
1909 and the other blood vessels of vertebrate embryos from capillaries,
Anat. Record, —
Evans, H. M. An instance of two subclavian arteries in the arm bud of man,
1908 Anat. Record, p. 411.
GraFre, E. Entwicklung der Urniere, etc., bei Hiithnchen, Archiv. fiir Mikr.
1906 Anatomie, Bd. 67.
Hocustetter, F. Uber den Einfluss der Entwicklung der bleibenden Nieren auf
1888 die Lage des Urnierenabschnittes der hinteren Cardinalvenen,
Anat. Anzeiger, Bd. 3, 8. 938.
Hocustetrer, F. Beitraige zur Entwicklungsgeschichte des Venensystems der
1893 Amnioten, III, Sauger, Morph. Jahrbuch, Bd. 20, 8. 548.
K6.uikerR, 1870 Entwicklungsgeschichte, S. 915.
Lewis, F. T. The development of the vena cava inferior, Amer. Journ. of Anat-
1901-02 omy, p. 229.
Lockwoop AND Humpury. Qn the development of the pericardium, diaphragm,
1888 and great veins, Philosophical Transactions of the Royal Society of
London, vol. B, p. 377.
Matt, F. P. 1898 Johns Hopkins Hospital Bulletin.
1905-06 A study of the structural unit of the liver, Amer. Journ. of Anat., vol. 5,
DOs 3, Pp. 2ol:
MUuuer. 1903-04-08 Anat. Hefte.
Rasy. 1907 Arch. fur. Mikr. Anatomie, Bd. 69, p. 340.
SmirH, H.W. On the development of the superficial veins of the body wall in the
1909 pig. Amer. Journ. of Anatomy.
Tuoma, R. Untersuchungen iiber die Histogenese and Histomechanik des Gefiss
1893 Systems.
Zoumstrern, Uber die Entwicklung der vena cava inferior bei dem Maulwurfe u.
1898 bei dem Kaninchen, Anat. Hefte, Abt. 1, Bd. 10, 8. 309.

ai = ‘ “ A
; reels et = .

% Ls » ’ |

ne) ALY -aat tS) [

=e. oY _ ‘ ?

V.Card.Ant.Dex.

, i | 4,

Aorta

Ne Ductus Cuvier

| Sinus Venosus
Liver
Ventricle /]
q V.Portae

V.Ventro-lat. of W.B

V Vitellina

Anastomosis
- V.Omphalo-Mesenterica

A.Omphalo-Mesenterica

O°

<a)
Aa.Mesonephricae 440° |] ji ) Les
IN V.Umb. Dex.
Aorta , \ |
wh
V.Subcard. Dex. y oy. AUmb.Sin,
a 7

PuaTE1. Pigembryo, length 7.5-Smm. No. 2301. Reconstruction of venous
system of right side, viewed from the right. The right umbilical vein is repre-
sented as transparent, with the subeardinal, cardinal, etce., shining through. Note
the large size of the anastomosis between the subeardinal and portal veins; fenes-
trated character and large size of the right subcardinal; and the ventro-lateral vein
of the mesonephros. X 19.0,

VEINS IN PIG EMBRYOS. PLATE I.
DAVID M. DAVIS.

THE AMERICAN JOURNAL OF ANATOMY, Vou. 10. No. 4.

Aorta

Lf Atrium

Sinus Venosus

| Af

V.Ventro-lat, of W.B
V.Umbilicalis Sin,

Liver

V.Vitellina
V.Omphalo-Mesenterica

A.Omphalo-Mesenterica

Aa.Mesonephricae

V.Cardinali\Post.Sin.
V.Subcatdinalis Sin.

Aorta

PLATE 2, Same Embryo as Plate 1. Reconstruction of venous system of left
side, mesial view from the right. The aorta is represented as transparent, with
the cardinal shining through. The white interrupted line gives the outline of the
left Wolffian body. The caudal part of the embryo is somewhat twisted, so the
small vessels entering the subeardinal caudal to the mesenteric artery are lateral
(mesonephric) not ventral tributaries. > 19.0.

VEINS IN PIG EMBRYOS. PLATE II.
DAVID M. DAVIS.

THE AMERICAN JOURNAL OF ANATOMY, VoL. 10. No. 4.

PLATE ITI.

VEINS IN PIG EMBRYOS.
DAVID M. DAVIS.

A.Vertebralis Dex.

Art.Pulm. Dex.

Te NG
a =e AN Ze Vv.Bronchiales

Way Auricle
Ductus Cuvieri @ ‘)
oe @; é Peri-Oesophageal

¢ plexus

Sinus Venosus
Ventricle

V.Portae

entrallaniqen of
Pan¢reas
4 Dorsal

Yolk sac
V.Umbilicalis De

Aa.Mesonephricae

Ventré-lateral vein
of Mesonephros

VW/Subcardinalis Dex.

PLaTE 3. Pig Embryo, length 8-8.5mm. No. 2302. Reconstruction of venous
system of right side, mesial view from the left. The aorta, excepting its two
extremities has been omitted. Text fig. 3 represents a section at the point
“Site of V. Cava Inf.’”’ The position of the omphalo-mesenteric artery is shown

by dotted lines. »X 15.5.

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10. No. 4.

VEINS IN PIG EMBRYOS.
DAVID M. DAVIS.

$4

oY. ag

Sil $4 > Safe Lingual-facial vein
A.Pulmonalis “ °
V.Puimonalis —s VG

Peri7Desophageals=
plexus

uctus Cuvieri

oortvle

G

“ww

Peri-gastric plexus Yolk sac

Subcaydinal-Portal ¥ A
Anastdmoses

Ava.Mesagnephricae >
C

of Mesonephros

PLATE

BYic

Sinus Venosus

PuiatTEe 4. Same embryo as Plate III. Reconstruction of venous system of left
side, mesial view from the right. The aorta, excepting its two extremities, has
been omitted. The position of the omphalo-mesenteric artery is shown by dotted
lines. The portal vein has been cut off where it passes to the right dorsal to the

intestine. X 15.5.

THe AMERICAN JOURNAL OF ANATOMY, VOL. 10. No. 4.

Puate 5. Pig Embryo, length 9.5mm. No. 2309. Reconstruction of venous
system of right side, mesial view from the left. Only the upper end of the right
aortic root is shown. The ductus venosus has been cut off shortly before entering
the common hepatic vein. The position of the omphalo-mesenteric artery is shown
by dotted lines. The head of the Wolffian body would extend as far up as the level
of the subclavianartery. Theconnections between stomach veins and peri-oeso-
phageal plexus are not shown. 15.2.

VEINS IN PIG EMBRYOS. PLATE V.
DAVID M. DAVIS.

A Peri-oesophagea| Plexus

V.Pulmonalis
Sap Vv.Bronchiales
H=NDuctus Venosus\Arantii

Ages
| Ee yo — A.Subclavia

Vy.Hepaticae

y

py)

<

V.Cava Inferior

V.Vitellina Stomach /Veins

Iw

A.Omphalo-Mesenterica
Cardinalis Dex.

Subcardinal portion

of Vend@ Cava
-Lateral Vein

esonephros

V.Omphalo-Mesenterica

V.Umbiliéalis Dex

@-= Aa.Mesonephricae

THE AMERICAN JOURNAL OF ANATOMY, Vou. 10. No. 4.

V.Jugularis Ext. V.Jugularis Int.

A.Pulmonalis

Peri-Oesopnagael Pléxlus V/.Cephali
ica

Ventrigkt—— =
Auricle i A.Subclavia
ME aipovells V.Basilica

Oesophagus

A.Coeliaca

XS
G2

»»MeSenterica Sup.

HE Subeardinal portion

By
0 |
<4 f C
Re BB oo

Aa. Mesonephricae

Cardinal portion
of Vena Cava
A.Mesenterica Inf.

R.Kidney

Puate 6. Pig Embryo, length 16.2 mm. No. 21. Reconstruction of venous
system of right side, mesial view from left. The omphalo-mesenteric inf. mesen-
teric and coeliac arteries have been cut off not far from their origins. A portion
of the umbilical vein has been removed. The stomach has been removed from its
position, over the venacava. The right kidney is shown in section. The stumps
of the left Wolffian duct and of the left ureter are shown. ‘The three cross anasto-
moses of the venous system are shown at zz, x. X 11.1.

VEINS IN PIG EMBRYOS.
DAVID M. DAVIS.

PLATE VI.

THE AMERICAN JOURNAL OF Anatomy, Vou, 10. No. 4.

!

ON THE HISTOGENESIS OF GASTRIC GLANDS

EDWIN G. KIRK
From the Hull Laboratory of Anatomy, The University of Chicago

With Twenty-six FIGURES

CONTENTS
Peealinitin occ tiom caret wee. occas hee ears cet ici See oak are A Sa eh Se ora 473
OL Paes Felli sf Oia Call eaten enon a Sheer ck Bias Ree forse ees Stel canes oly. asia ste aa ome 474
ites Vicrpertaleanditechimt ete o ~s-rad capers Sans figs cei oe aosncs ees ga mlevoieeaoase Wtee, LE val dut 477
Wes Eistocenésis on tiercasiic elandsiof the piensa eet dee eee 480
IPS ere amv s bal Comer mettre pus ste, amet et Cae e CAS SECS need 480
ee Ep OLARS Eo: Aer eI ota a Myer NA Ne, ey Sead wean Ceca} hokiee eestaes 499
ME LUT CUS Pier wee pea een en nny mana. WAM ANC AE Cc aR ot a eh elle 500
4b, (CRN ISIE vay ae B/C VEVONDT GIT. DO ireh sian ake tier URANO ee RAL Te 508
a VAESLOCSO MaDe alee Mae hae parte er erran Stee ae UR icine ims Gnd Cad oR 5 TEE 513
6. Conclusions and theoretical deductions. ............:2.<se:.+-++<: 514
(a) As to homologies and phylogeny of the cardia............ 514
(O)eeANsttoycelllspeciticitiye cs. cnet inten c urnae noes Sie ale 516
Siem eS aU) Dales Io Vere pn sta vert Acs Leah cea yee, Peat coal! a MG e os a aha wos Seas 519

I. INTRODUCTION

This paper represents an attempt to throw some light on the
fundamental processes in the cytomorphosis of mammalian gas-
tric glands. A study of the literature convinces one that, in
spite of numerous researches, there still persists a marked lack
of agreement even as to the broader outlines, which becomes,
with regard to more minute details, a chaotic mass of contra-
diction, partially referable to the normal generic variations in
the forms used, and partially, perhaps, to misinterpretation.

It was decided, on the following grounds, to limit this investi-
gation to the study of a single species, the pig:—First, theoret-
ical considerations of a general, embryologic nature, lead one to
infer that, aside from slight generic differences, the same essen-
tial processes are probably concerned in the gastric adenogenesis

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, No. 4.

474 Edwin G. Kirk.

of the whole mammalian series. Second, one may disentangle
from the generally confused data of the literature, certain con-
cordant results, which confirm the validity of this assumption
as to the essential unity of the process throughout mammals.
Third, and very obviously, a limitation to one form means the
possibility of more detailed work than is practicable in a compara-
tive study. The present need seems to be for minute work, like
that of Toldt on the cat, but utilising valuable, recently developed
technique, and also one old technique, evidently very often
neglected nowadays, namely the patient use of serial reconstruc-
tions in place of the easier reliance on occasional isolated sections.
Finally, and herein lies the proximal motive in selecting this
particular species—an abundance of fresh material of every
stage has been readily procurable.

I wish to express my sincere gratitude to Dr. R. R. Bensley,
at whose suggestion this work was undertaken, and under whose
euidance it has been carried on. Without the aid of certain meth-
ods devised by him, much of the work on cyto-differentiation
would have been technically impossible. I am indebted to Miss
Katharine Hill for the accurate drawings.

II. HISTORICAL

Essentially all authors agree that the gastric epithelium is
derived from the endoderm. Brand ’77, Sewall, ’78 and Kolli-
ker, ’84 describe a transformation of the original single endodermic
layer into a stratified epithelium, which later, bysome unexplained
mechanism, again becomes simple. All recent writers who men-
tion the subject at all (Toldt ’81, Salivoli 90, Ross ’03) agree
that the epithelium remains simple from the first, although the
disposition of nuclei in several planes simulates stratification ;
of course this does not apply to areas which become stratified
and remain so, é. g., the left compartment of the field mouse
stomach (Tépfer. 91), or the pars oesophagea.

Conéerning the origin of the rudimentary gland tubules, there.
is great diversity of opinion. The various views may be classi-
fied under three general groups, depending on whether the tubules

On the Histogenesis of Gastric Glands. 475

are believed to develop (1) from the surface epithelium, (2) from
special embryonic cells interpolated between the basal parts
of the ordinary surface epithelium, or (3) from mesodermic cells,
such as leucocytes.

Group 1. Of those who describe the glands as originating as
downgrowths of the surface epithelium, several have considered the
all-important factor in early adeno-genesis to be irregular growth
of the mesoderm, first manifesting itself as upgrowths, in the form
of either villi or ridges. These coalesce or intersect, thus giving
rise to intervening cul-de-sacs, the rudimentary glands. Of course
the surface epithelium multiples to keep pace with the increasing
area of mesoderm it must cover, but, by convention, this activity
is considered secondary to that of the mesoblast. (For details
vide Laskowsky ’68, Schenk ’74, Brand ’77, Sewall ’78, Baginsky
’82, Kolliker ’84, p. 360, Negrini ’86.) Observations recorded
in part 2 of this paper explain, we believe, the discrepancies of
opinion regarding the exact nature of these mesodermic irregu-
larities, 7. e., whether villi, papillae or ridges. Ko6lliker (’52-’61)
Toldt ’81 (pylorus of cat) Patzelt ’84, Griffini and Vassale ’88,
and Salivoli ’90 believe that the epithelium displays the initial
activity, and that the glands may be said to have an intra-epi-
thelial origin, irregular growth of mesoderm later becoming a fac-
tor in bringing the glands to their definitive form. Thus, Kolli-
ker, in ’52—’61, described the downgrowth of solid plugs of epi-
thelium, which later acquire a lumen; he discarded this view
(84), nor has it since been advocated. Griffini and Vassale des-
cribe the gland as originating from the surface epithelium, as a
funnel-shaped evagination, sinking down into the mesoderm.
The deepest cells, being in continual mitosis, constitute an apical
growth point. Only at a comparatively late stage does unequal
mesodermic growth participate. According to Salvioli (90,
rabbit), at an early stage (4 em.), while the basement membrane
is yet perfectly level, intersecting ridges of epithelial cells rise
above the general level of the epithelium. The intervening
depressions are the rudimentary tubules. Later (6 cm.), these
epithelial ridges are reinforced from below by cores of connec-

476 Edwin G. Kirk.

tive tissue. Toldt (’81, cat) describes a similar process in the
pyloric region. ‘The process is wholly confined to the epithelial
layer.”’ Patzelt (84) describes the same process in the formation
of the glands of the large intestine.

Group 2. Toldt(81 cat) working on the fundic region, and Ross
(1903, pig), describes the rudiments of the glands as large,
coarsely granular, eosinophile cells interpolated between the basal
parts of the surface epithelium. Each divides into a cell group,
which, by central liquefaction acquires a lumen, the latter sec-
ondarily coming into communication with the surface. Toldt is
sure these cells are of epithelial origin, but believes they at no
time reach the surface, being always shut off from the latter
by the overhanging distal ends of the tall pyramidal surface
epithelium; he suspects that they arise from young Ersatzzellen
His Ersatzzellen have almost certainly been shown by the work
of Stohr (1882) and Bizzozero (1888) to be ‘‘Wanderzellen.”’
Griffini and Vassale maintain that Toldt’s figures and text har-
monize remarkably with their own findings (V. supra), except
that Toldt, through use of oblique sections, erroneously concluded
that these primary gland cells do not reach the surface, and that
their lumen is thus not at first continuous with the stomach
lumen. Griffini and Vassalefound many such groups with lumina
apparently shut in on all sides, but reconstruction always demon-
strated continuity with the stomach lumen from the first.

Group 3. Sewall (’78) believes that, once the original hypoblast
has differentiated into ovoids (parietals) and central (chief) cells,
new ovoids “‘originale’’ by differentiation of mesodermic corpuscles.
In embryo cats (13 em) mesoblast cells are found, presenting all
transitions from connective tissue corpuscles to parietal cells
(Toldt says that Salvioli simply cut the eccentric parietals tan-
gentially from adjacent tubules, and hence misinterpreted them to
be cells lying free in the lamina propria). The reader must be
referred to the recent work of Strecker (’08), which is too elab-
orate to be reviewed here. His conclusions are diametrically
opposed to mine. The diversity of these views as to the early
formation of gland tubules make evident the need of further

On the Histogenesis of Gastric Glands. 477

investigation of the two separate problems involved: (1), from
exactly what cells do the glands arise. (2), to what extent are irreg-
ular growth of mesoderm and epithelium, respectively, factors.

Compounding of the tubules is unanimously described as due to
the upgrowth, at the bottom of the simple tubules, of partitions
which never reach the surface. Toldt, ’81, believes that these are,
at first, proliferating solid ridges of epithelium, later reinforced
from below by a core of mesoderm. Sewall, ’78, Negrini, ’86, and
Ross, ’05, hold that the upgrowths contain mesodermic cores from
the first. According to Salvioli, 90, the process described in the
original formation of the foveola is repeated at the bottom of the
foveola. By mitosis, intersecting epithelial ridges arise; later
these are reinforced from below by mesodermic cores.

Toldt, Salvioli, Griffini and Vassale (opera cit.) also describe
a second method of compounding by ‘‘side-buds.” Lateral
diverticula appear, originating through local proliferation of a
cell-group, which starts from a single parietal cell. I have con-
firmed this for the pig (vide part IV).

As to cytologic differentiation, Laskowsky, Sewall, Toldt, et al.,
describe an early departure of the tubule cells from the sur-
face epithelial type, the former becoming polygonal or ovoid,
their cytoplasm granular, and their nuclei large and vesicular.
Toldt, ’81, Negrini, ’86, and others demonstrate that parietals
appear in the depths of the tubules by differentiation of the em-
bryonic tubule cells. Bensley, ’03, finds that mucus cells appear
in the pylorus and along the lesser curve in the 6 cm. pig. Parie-
tals appear in cardia and fundus at 7.5 em., inter-cellular duc-
tules being present from the first. Zymogenic (serous chief)
cells appear, at about 21 cm., at the bottom of the fundic glands.
A few, as Sewall, ’78, and Strecker, ’08, believe in a possible meso-
dermic origin of parietals, or of the whole gland anlagen, either
from leucocytes or from mesodermic corpuscles, fibroblasts etc.

III. MATERIAL AND TECHNIQUE

The embryos were obtained in an absolutely fresh condition
through the kindness of Swift & Company, of the Chicago Stock

478 Edwin G. Kirk.

Yards. Half a minute after the instantaneous death of the mother,
by cerebral concussion, the uterus was cut out, and immediately
opened. The embryos were removed, and their greatest length
in the natural attitude along a straight line (Minot’s System.
Vide Minot, ’03, p. 356) determined by calipers, and recorded in
centimeters. The stomach was then rapidly removed, the subdi-
phragmatic oesophagus and also a small bit of the duodenum
having been left attached in the earlier ones for orientation. The
stomach was then slit open along either anterior or posterior
side, and filled with fixing fluid. Stomachs from embryos of over
6 cm. length are distended with a clear, glairy, mucoid fluid,
often of a greenish tinge. This must first be allowed to escape, as
otherwise fixation is unsatisfactory. After being filled with fixing
fluid, the stomach is immersed in it and placed in the dark. After
a little practice the whole operation need take no longer than
half a minute. This fixation immediately after death is of great
importance in obtaining accurate pictures of intracellular condi-
tions, especially of the zymogenic, mucus and parietal cell gran-
ules. In dealing with embryos of over 12 cm. some of the stomachs
were treated as described above, others were subdivided, great
care being taken to preserve the identity and orientation of the
pieces. The series included embryos of 2 to 29 em. length at inter-
vals of 4 cm. Generally, several stomachs of each stage were
used; some for sagittal sections, thro’ the whole length where
practicable, some for cross sections. In all cases the sections were
cut serially, and this is very necessary, as will be seen later, for
the correct interpretation of certain appearances.

I tried many methods of fixation, but finally settled down to the
use of Bensley’s fluid as giving the best general results.'. Modified

1 This consists of equal parts of 3 per cent aqueous K2Cr207 and sat. HgC12 in
100 per cent alcohol, mixed just before using. The cloudy precipitate is to be dis-
regarded. The material is fixed in the dark from 30 minutes to 2 hours, according
to thickness. It is then transferred to50 per cent alcohol, in which, with frequent
changes, it remains several days; then to 70, 80 and 95 per cent a day or two, with
several changes in each; finally to 100 per cent and then bergamot, bergamot and
paraffine, and finally paraffine. At no time until after impregnation with paraffine
should the material be subjected to the action of water. Otherwise, as Bensley
pointed out (1903-4) the zymogen granules and mucigen are preserved poorly or
not at all.

On the Histogenesis of Gastric Glands. 479

K6pschis a good fixative for Bensley’s three color stain (see below).
Controls were often made with Zenker and 5 per cent freshly
distilled formalin, but these only served to emphasize the superior-
ity of Bensley’s fluid, especially for the neutral gentian and copper
chrome stains. The eosinophile granules of the parietals, the
zymogenic granules and the contents of mucigenous cells are
especially well preserved by this fluid.

Sections were cut 4 “ thick, and fixed to slide by the water
method, or by Mayer’s albumin. Demercurization was completed
by immersion of slides in iodine-alcohol.

Hematoxylin and eosin stains were used in each stage for com-

parison, but the main reliance was placed upon the four stains
named below, serial sections of each stage being stained with
each of these:

1. Bensley’s three color stain as described by Klein (06) p.
323. The granules of parietal cells are stained a vivid cherry-red
and the intercellular ductules are brought out well. This has
proved the best all around stain for stomach material.

2. Neutral gentian (Bensley ’00) in 20 per cent alcoholic
solution was invaluable in determining the appearance of zymo-
genic granules. It is also, as far as my experience goes, the best
cement line stain, and is thus of great value in the study of the
parietal ductules, and of cell boundaries. The material should
never be brought into water, as the zymogen granules then disap-
pear. I stained two to four days.

3. Bensley’s Copper chrome hematoxylin as described by Har-
vey (07, p. 209). The granules of the parietals are stained a
steel blue or deep blue black. The chromatin stains black, but
is more readily destained than are the parietal granules.

4. Mayer’s Muchematin, as modified by Bensley (’03, p. 11) for
the detection of mucigen. It is best to avoid the water and heat
method in fastening sections to slide, as the granular form of
the mucigen is thereby altered.

480 Edwin G. Kirk.
HISTOGENESIS OF THE GASTRIC GLANDS OF THE PIG
1. The early stages

The stomach of a 2 em. pig is about 2 mm. long. The cephalad
part just left of the oesophagus has already been partially folded
off as the secondary cardiac pouch (Coecum) being now separated
from the main lumen by a fold which isolates it.2. This ridge is
an infolding of all the layers, including the tunica muscularis,
the latter and the connective tissue coat being also thickened at
this point.

A definite basement membrane separates epithelium from
mesoblastic coats. This stains densely with Rubin §, and has a
fibrous structure.

The mesoblast has already differentiated into the connective
tissue layer, the tunica muscularis and the serosa. Only the first
concerns us. It is a typical mesenchyme, with numerous stellate
and spindle cells, anastomosing by their processes. The nuclei
are spherical, ovoid or elongated and many are in mitosis. The
ground substance is transparent and gelatinous. Already this
connective tissue coat has definitely differentiated into two
strata (fig. 1).

The cells and nuclei of the inner layer, just beneath the base-
ment membrane, are very closely set, almost touching. Mitoses
are very numerous. There is here but little of the mucoid, inter-
cellular matrix, and it is dense, staining somewhat reddish or
purplish with the three color or H & E. The blood vessels are of
capillary size only. The endothelial cells of the vessel walls are
very frequently mitotic. This layer is the young lamina propria
mucosae.

Outside this is a broader zone with cells and nuclei sparsely
distributed, and a corresponding predominance of intercellular
substance, the latter staining hardly at all, and being very clear
and transparent. Mitoses occur, but are rare. Blood vessels
are very numerous and many of them are of large size. This coat

2 For the topography of the adult stomach, reference may be made to Green-
wood’s accurate diagram, Fig. 252, Oppel, 1896.

On the Histogenesis of Gastric Glands. 481

Fig. 1. Earliest or preglandular stage. Epithelium and underlying mesoblast
6 cm. embryo. Three color stain.

Fig. 2. Young parietal cells, showing intercellular ductules. 6 cm. Fundus.
Three color.

Fig. 3. Intra-epithelial gland stage. 6 ecm. fundus. Three color.

Fies. 4 and 5. Young glands cut obliquely. 6 ecm. fundus. Three color. 5a
and 5b represent adjacent sections; similarly 6a and 6b. These illustrate the true
nature of the interbasal groups of Toldt and Ross.

Fig. 6. Fundus gland. 6 cm. parietal cells were present in other sections of
this same tubule. Three color.

482 Edwin G. Kirk.

is the young tela submucosa. The boundary between these two
layers, while not absolutely sharp, is quite well marked, being
definable within the limit of one or two cells breadths.

There is yet no trace of a muscularis mucosae, this first appear-
ing about 9 or 10 em. at the boundary between lamina propria
and tela submucosa. It originates through the elongation of
mesenchyme nuclei to a torpedo shape, and the condensation of
the protoplasm about each nucleus, as a highly eosinophile sub-
stance. The primitive syncitial anastomoses are retained, the
new muscle cells being thus joined inter se by delicate prolonga-
tions of the finely fibrous stroma.* In some stomachs isolated
muscle fibres apparently appear somewhat earlier (8-14, 9 cm.)
At 15-16 em. a definite, fairly compact muscularis mucosae is
present. The fibres of both coats of the tunica muscularis are
well defined at 2 cm.

The diversity of opinion as to the part taken in the inception
of the glands by unequal mesodermic growth, and as to the nature
of the first mesodermic irregularities, is partially referable, no
doubt, to differences in the forms used, but there has often been
the lack of a definite criterion of distinction between such up-
growths as simply minister to an increase of surface, and such as
have a definite réle in glandular development. By reference to
the adult structure, it is seen that only such mesodermic eleva-
tions can have taken an active part in adenogenesis as are constituted
by local thickenings of the lamina propria alone. At 2 em. the
later is of uniform thickness throughout, the basement membrane
presenting no irregularities of contour, and the epithelium being
everywhere of the same height. But the tela submucosa has
thickened in linear ridges, so as to produce folds or rugae, several
of which run almost the length of the stomach.

The epithelium is a single layer of high columnar cells, as shown
in fig. 1 of the 6 em. stage The nuclei are arranged in several
rows; this, in connection with the small diameter of the cells,
readily gives rise, in even slightly oblique sections, to a stratified

3’ This confirms MeGill, 1907.

On the Histogenesis of Gastric Glands. 483

appearance. At no stage does the epithelium acquire stratification
except in the pars oesophagea.?!

The nuclei really occur at all heights in the cells, but, may, for
convenience, be arbitrarily grouped into those occupying the distal
middle and basal parts. At 2-3 em. almost every nucleus of the
distal row is in mitosis, and all stages of this process are, of course,
represented. The nuclei lying at deeper levels are all resting.
Whenever the mitosis is in such a stage that its direction may be
determined, its axis is invariably parallel to the surface of the
epithelium. The nuclei are oval, saccular, and so broad as to
almost touch the sides of the cell. They have a moderate amount
of chromatin, arranged as a net, and with nodal karyosomes.
There are usually two nucleoli, metachromatic or slightly acid-
ophile. The cytoplasm is very finely granular in the epithelium
of the coecum, cardia and cardiac part of the fundus; in the
plyorus, pyloric part of the fundus and pars oesophagea it is
clear and transparent. Aside from this, no cyto-differentiation is
discoverable by the techniques employed.

At 2% em. irregular thickenings of the tela submucosa in the
coecal pouch begin to give rise to the characteristic grosser
ridges folds and papillae of that region.

About 214-3 cm. the surface line of the epithelium, hitherto
level, becomes, in some parts of the stomach (pylorus and fundus
along the greater curve near the pyloric region), undulatory,
displaying alternate very slight elevations and depressions. The
basement membrane shows no corresponding waviness, and the
lamina prop. no irregularities. On reconstruction, the elevations
are found to be short ridges, intersecting in all directions, the de-
pressions thus representing a slight pit bounded on all sides by the
ridges. It is readily determined that the elevations consist of
cells slightly taller and narrower than those of the depressions
(fig. 3), with the nuclei in the distal end; the cells of the depres-
sions have central or basal nuclei, and have retained their primi-
tive height and breadth, or, sometimes increased slightly in breadth.
Mitoses are very frequent in the ridge cells. All the cells
reach from basement membrane to surface. These slight intraepi-

‘ This finding agrees with those of all the later investigators, as Toldt, Salvioli,
Ross.

484 Edwin G. Kirk.

thelial depressions, due to irregularity in height of the epithelial
cells, represent the beginnings of the glands, as I have satisfied
myself by tracing, in unbroken series, all steps from the adult
glands back to these. It will be convenient, hereafter, to refer
to this as the intra-epithelial stage of gland formation.

Nevertheless, itishardly possible to say that the mesoderm plays
no part in even the initial stages, for these epithelial ridges have
scarcely appeared before mesodermic buds begin to push up into
them (fig. 4). These latter are constituted by local thickenings
involving only the lam. propria, and are true mesodermic gland
processes.> This latter condition is found rarely at 214 em. but
often at 3 em. in the precocious regions. As soon as mesodermic
cores appear, the basement membrane is seen to be sharper in
outline, and more compact, beneath the pit or gland cells than
beneath the ridge cells, being frayed out in the latter locality
into fibres which penetrate beween the bases of the cells. Fig.
6, indicates this same condition in a later stage. Possibly this
has some influence on the supply of nutrition to the epithelium,
and consequently on local differentiation of the latter(?).

The process just described is, in essence, the same as Salvioli’s
finding in the rabbit, but there are minor differences:

1, In the pig the cells of elevation and depression do not
diverge quite so widely in form, in these earliest stages. But at
the stage represented in fig. 3 the form divergence seems quite as
great.

2. Inthe pig, thereis yet no differentiation, as to the cytoplasm
into two types of cells, as Salvioli’s clear cells of the elevations,
and granular cells of the depressions. Instead all the cells in the
areas of gland formation, possess, as yet, a finely granular, slightly
acidophile cytoplasm. The cells of the fundus and pylorus were
non-granular at 2 em. but, shortly before the appearance of the
intra-epithelial gland anlagen, have acquired the fine cytoplasmic
granulation.

° This term of Sewall has the priority. His gland processes undoubtedly corre-
spond to these, and despite Ross’ criticism, certainly play an integral part in gland
development.

On the Histogenesis of Gastric Glands. 485

3. In the pig, the mesodermic cores appear relatively much
earlier—in fact so early that it was only after careful reconstruc-
tions that the appearance of epithelial ridges slightly in advance
was established. Toldt’s description for the pylorus is very
similar. He admits that in this region there is no independent
origin of the gland cells, with, later, secondary communications
between these and the lumen.

From 3 cm. on, the ridges of lamina propria grow rapidly up
into the epithelial ridges (figs. 4 and 6), causing a correspondingly
rapid heightening of the latter, and deepening of the glands. For
the correct interpretation of certain papilloid structures seen in
all stages of the gastric development up to about 23-27 em., it
js essential to understand the exact way in which the irregularity
of mesoblastic growth manifests itself. As has been indicated,
the early, intra-epithelial ridges intersect in all directions. They
are not straight, nor do they intersect often at right angles; rather
they are short, very irregular, curved or angular, and they inter-
sect at all angles. Now, when the mesoblastic ridges begin to
reinforce them from below, the mesoblast pushes up much more
rapidly at the nodal points of intersection of the ridges than along
the intervening parts. The result is that in a sagittal section
taken (fig. 7A) through the line a, b, c, d, the appearance seen
in fig. 7B results. That is, we have in addition to the ridges the
papilloid elevations a, c, and d, situated at the intersections of
the ridges, and higher than the rest of the interglandular ridge.
In cross or oblique section of the upper level of the mucosa, they
are, of course very numerous, and appear like true papillae. One
might readily gain the impressions, as did Brandt and others, that
in the early stages there are no gland processes, but simply papillae,
the depressed areas between them communicating with each
other. Such a condition would be represented by fig. 7A, with
all elevations except the nodal points effaced. But this is very
different from the condition actually found—namely, that ridges,
not so high, it is true, as the papillae, but still high in comparison
with the depressed areas, run across from the base of each papilla
to the adjacent one. This papilloid condition is marked from the

486 Edwin G. Kirk.

time of the appearance of the mesoblastic reinforcements to the
20-26. em. stage, but all this time there is a very gradual increase
in height of intervening ridges, until finally at 24-28 cm. the defini-
tive condition is reached, in which there are no longer any
papilliform elevations, all the ridges having grown up to the
same height as the papillae.

The nodal papillae are often expanded or club-shaped at the
top, resembling greatly intestinal villi, but of course shorter. By
reference to fig. 7B, it will be seen that in any section through the
mucosa the elevations and depressions must necessarily present
great irregularities, as e. g., the epithelial line may dip into a
gland pit, then perhaps ascend to the top of a nodal elevation,
then descend to the level of an interglandular ridge.

Moreover, these ridges and pits cannot be interpreted as mere
folds of the mucosa, having no permanent significance, nor any
relationship to the adult gland. Once established, these pits
indeed deepen and compound at the bottom, many of them even
divide into two glands; but their continuity with the adult gland
is established by unbroken series. Moreover, we shall see that
soon after this formation, certain cells in their depths differentiate
into recognizable elements of the adult gland. Finally, the meso-
blastic cores of the ridges are, as shown, true thickenings of the
lamina propria, not mere foldings of a |. p. of uniform thickness.

It is now easy to understand the many contradictory accounts
of the nature of the primary gland processes. Brandt described
papille or ville, which later fuse at their bases. Sewall, Salvioli
and many others described only the interlacing ridges, and failed
to explain the very frequent appearance of villoid cross sections.

As growth proceeds, all the epithelial cells increase in absolute
size. Those on the ridges become very high, inverted pyramidal
in shape, the proximal part being compressed into a narrow, caudal
process, which attaches to the basement membrane (figs. 6 and
11). The nuclei are elongated and laterally compressed. The
cells in the pits, or embryonic gland cells® increase in breadth, but

° T retain Sewall’s term, as it has priority, and is self-explanatory. Toldt calls
these “‘adelomorphs,”’ but applies this term at a later stage to the mucus and ser-
ous chief cells,

On the Histogenesis of Gastric Glands. 487

-
2

Fie. 7. Aand B. Diagrams (vide text).

Fic. 8. Fundus gland. 7 cm., showing one parietal cell with ductules. The
other cells are ‘‘Embryonic gland cells,’’ 7.e., undifferentiated; at least they present
none of the characteristic staining-reactions of any of the adult cell-types.

Fie. 9. aandb. Fundus epithelium 7 cm. Copper chrome hematoxylin.

Fig. 10. Tracing of mesial sagittal section through 7 em. stomach enlarged
x8.6. BtoC, Caecal pouch. Cto4, Cardiaandfundus. 4 to pyloric aperture, and

latter to 7, pyloric gland area. 7 to B, pars oesophagea. B, Caecal ‘‘Wulst’’ or
ridge.

488 Edwin G. Kirk.

notin height, so that they become thick, short and cylindric, or
pyramidal, with broad, basal ends. The nuclei are ovoid, or often
almost spherical and moderately chromatic. In glands of the
stage indicated in figs. 6 and 11, the gland cells average 16u in
length, 8u breadth; the ridge cells, 32u height, 3.24 breadth at mid-
dle, 6.4 breadth at the broadest part ordistalend. Up tothestage
of fig. 3 the only distinction between the epithelial cells of the
pits and ridges, are those just described, of size and of contour of
cell and nucleus. The cytoplasm of all these cells is quite refractile
owing to the presence of very fine, slightly acidophile, granules.
Up to this stage, then, we have found two factors coéperating
in the deepening of the primitive gland tubules. (1) Upgrowth of
the mesodermic gland processes, (2) increased height of the ridge
epithelial cells, which are now over twice as high as the gland
cells.

But as the mesoblastic buds reinforce the epithelial gland
processes mitoses become relatively fewer in the ridge cells, and
appear inthe pit cells, and it soon becomes apparent that the gland
tubules elongate by interstitial growth, i.e., by mitoses of the gland
cells. ‘Thus the epithelial part keeps pace with the growth of the
lamina propia..

The chronological priority, however slight, of the epithelial
ridges to the mesoblastic cores, seems, to indicate that, from the
first, certain epithelial cells possess an inherently different po-
tentiality from others. For the primary ridges and depressions
are so minute that it is inconceivable that differences in the
distribution of vascular supply in the underlying mesoblast
could account for this appearance. But, waiving this, at present,
fruitless discussion of the predetermination of the cells, it is
obvious that the grosser differences in the external form of the
ridge and pit-cells are largely a function (in the mathematical
sense) of the formation of ridges and pits, i. e.,—the inverted
pyriform contour of the ridge cells is largely the mechanical re-
sultant of the compressive forces exerted at their base end and of
tensile force exerted at their distal end; and the short, pyramidal
form of the gland cells, of compressive forces exerted at their
distal end, and of tensile force applied at the base. And these

On the Histogenesis of Gastric Glands. 489

compressive and tensile forces are obviously referable, in great
measure to the uneven mesoblastic growth.

However, we shall see below that other differentiative forces,
intrinsic to the cell, and of a metabolic nature, are at work, as
is indicated by the early departure of certain of the gland cells
from the primitive embryonic type.

These glands appear earliest in the following localities:

(1) The pyloric portion of the greater curve; (2) the part of
the fundus adjacent to (1); (8) the caeecal houch; (4) a narrow,
peripheral zone of the pars oesophagea. There is some doubt as to
whether these latter become true glands.

From these primary regions they extend to all other regions,
with the exception of the major part of the pars oesophagea, i.e.,
that epithelial territory which later becomes stratified squamous.
Thus, gland development is most retarded in the cardiac end, over
the facies anterior and posterior, and especially over the caecal
ridge. Moreover, apart from this general, regional precocity or
retardation, several slightly different stages of development may
be represented in each region.

At 4-5 em. the pyloric glands are mostly in the stage of
fig. 4 the processes having cores of mesoblast. Some are
advanced almost to the stage of fig. 6. Those of the fundus are
of the stage of fig. 3, while the cardia displays a slightly undulating
epithelium, with numerous mitoses in the upper row of nuclei
of the elevations. There is, as yet, no sharp boundary between
fundus and pylorus, as to size and form of the glands, but we shall
see below that already. through cyto-differentiation, these two
regions are marked off from each other.

Between cardia and fundus, no definite boundary of any sort
is discoverable, since the change from the shallower (vounger)
glands of the cardia to the deeper (older) of the fundus is very
gradual.

The developmental retardation, as intimated above, finds its
extreme expression over the coecal ridge, where the epithelium is
at this stage, level, with very numerous mitoses in the distal row
of nuclei.

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 4.

490 Edwin G. Kirk.

All the epithelial cells, both ridge and gland, with the exception
of those of the pars oesophagea, present at the first appearance of
the glands, the same type of cytoplasm,—highly refractile
from the inclusion of a very fine, closely set, slightly acidophile
granulation. These embryonic gland cells do not all remain
undifferentiated very long after the appearance of the gland anla-
gen. At 3 ecm. some of the glands of the stage of fig. 3 or. even
slightly earlier, situated in the fundus, along the greater curve,
display one or two cells, which have acquired a cytoplasmic
granulation somewhat coarser than that of the primitive cells,
and very highly eosinophile. These cells are at first of the same
size and contour as the others (figs. 4 and 9) but they soon enlarge
somewhat, the basal end becoming broader and rounded, while
the distal end narrows, often becoming caudate (fig. 2). Later,
the narrow distal end is lost (retracted?), but at all times, up to
24 em. the cells reach the surface (fig. 8, etc.). The nucleus, at
first ovoid, later rounds up, and approaches the spheroid form.
These cells are the earliest parietals. Even at their first appear-
ance, one often finds, in relation to them, minute intercellular
ductules, always continuous with the lumen of the gland, or, in
the more shallow glands, opening directly into the stomach
lumen (figs. 2, 4,5). These pass down between the parietal and
the adjacent primitive cells. The almost absolute constancy
with which these early parietals display the ductules is striking.
As already intimated, parietals sometimes appear in insinkings
so slight that they seem almost to have differentiated in level
epithelium. This occasionally occurs as late as 7-8 em. (fig. 9)
in small patches of epithelium which remain almost level in the
midst of active gland formation on all sides, as also in the ex-
treme cardia at a stage when the glands are just appearing.
Nevertheless, even these parietals are probably always situated
within the territory of a gland-to-be, for after the 10 cm. stage,
parietals are found only in bona fide glands, and no primary
gland-pits appear later than this stage. Transitional forms be-
tween embryonic gland cells and parietals are also found, their
cytoplasmic granules being intermediate in size aud avidity
for acid stain. ‘These occur not only in the early stages, but even

—

On the Histogenesis of Gastric Glands. 49]

at 11-12 em. and probably up to 19 em. A point which I wish
to emphasize is that the glands are largely constituted from the
very first, of adelomorphs.?. Some glands have parietals almost
from the first; some do not develop any for some time. Thus, as
late as 6-7 cm. glands occur, especially in the cardia, but also
in the fundus, in which all the cells are undifferentiated. And by
the side of these will be found glands of the same age (size) but
with one or more parietals. I am able to confirm Toldt’s state-
ment that parietals do not appear in the pylorus at any stage.
However, once or twice,—so very rarely that even to mention
it is to give an exaggerated idea of the importance,—I have found
a single parietal in a young gland from the undoubted regio
pylorica; never, however, a group of parietals. Also, while in
later stages, the boundary between fundus and pylorus is very
sharp on the greater curve, there is, at some points on the facies
anterior and posterior a zone, two or three tubules wide, in which
occur mucus chief and parietal cells, but no zymogenics.

Thus, with the differentiation of parietals, we have at once a
rough divergence of left gastric region, corresponding to caecal
pouch, cardia and fundus, where they appear, and right gastric
region, the pylorus, where they never appear. Merely a rough
divergence at first, because, in the earliest stages, not all fundic
tubules have parietals. Thus the parietals arise at first, by the
differentiation of certain of the adelomorphs; but, as early as
4-5 cm. they increase also by mitosis (fig. 5). This method of
division in par etals has been denied or doubted by many observ-
ers, but I can say positively that it frequently occurs up to rather
late stages of glandular development, certainly as late as 16
em. The eosinophile granules remain intact, but a clear zone,
free from them, surrounds the chromatic figure. As soon as
groups of 2 or 3 have thus arisen the ductules are formed not
only between the parietals and adjacent embryonic cells, but also
between adjacent, parietals (Figs. 2, 4, 5). Thus, by mitotic
division of preéxisting ones, and by differentiation of new ones

' This convenient term, used by Toldt in the same connection, may, for brevity,

be used interchangeably with primitive gland cell, care being taken not to apply
it, as did Toldt, to the specialized mucous and serous chief cells,

492 Edwin G. Kirk.

from adelomorphs, larger and larger groups and rows appear in
later stages. I do not believe that new ones develop from adelo-
morphs after 13-14 em. and it seems almost certain that this does
not occur after 19-20 em., as all the adelomorphs have at tnat time
differentiated into other types, mucus or zymogenic. Thus, in
the latest stages, the parietals probably arise only by division of
parietal predecessors, unless there is a genetic relation between
parietals and zymogenic or mucus cells, of which I found no
evidences.

If the section of the early gland be even slightly oblique the
parietals, whether isolated or in small groups, appear as conspicu-
ous, large, granular cells, lying apparently between the bases
of the surrounding adelomorphs and often as if far removed from
the surface. If cut obliquely, the cells appear of ovoid contour,
if cut transversely, of circular outline, hence sometimes inter-
preted as referable to a spherical shape (Ross, ’03). Figs. 4b, 5a,
and 9a illustrate this condition, as do Ross’ (’08) figs. 25, 26,
and 24. Byreconstruction it is found that they reach the surface.
Vide figs. 4a representing the section adjacent to 4b, 5b adjacent
to 5a; 9b where obliquely cut parietals are shown, including the
distal end of one. These obliquely cut parietals are much more
conspicuous objects than are the obliquely cut bases of the sur-
rounding adelomorphs, owing to their larger size, granulation
and deep red color. This probably accounts for the inconsistency
of the interpretations of Ross, whose figures show the obliquely
cut bases of parietal cells surrounded on all sides by undifferen-
tiated cells, likewise cut obliquely. The parietal cell bases are
then described as basal gland anlagen, which have never yet
approached the surface of the epithelium; curiously, this inter-
pretation is not extended to the obliquely cut bases of the adelo-
morphs. Ross’ figs. 28 and 29 are especially significant as
showing the true condition, and are in perfect accord with my
findings. The little depressions mentioned by Toldt and Ross
(Ross ’03, fig. 24d) and interpreted as surface insinkings which
later communicate with the fundus of the gland, are shown in
my fig. 4b etc., but a glance at adjacent sections (as 4a) always
shows that they are merely part of the shallowlumenof the gland,

On the Histogenesis of Gastric Glands. 493

i. e., have been from the first in communication with the Jumen of
the gland fundus. Toldt and Ross have both spoken of the appear-
ance of a minute vacuole between the cells of a group; thus Ross:
“A central lumen, small, to be sure, but still a lumen” (fig. 26).
This is said to have arisen independently within the group, and
to have secondarily communicated with the surface. They
figure and describe very accurately the appearance of the inter-
cellular parietal ductules, as seen in cross or oblique sections
(Vide my figs. 2, 4, 5).

Salvioli, too, (90) has pointed out the source of Toldt’s
error (’81)—namely, oblique sections, with no reconstruction
control —but this warning was ev’dently overlooked or disre-
garded by Ross (’03). Ross pictures similar groups of basal
cells for Amblystoma and the pig but the two are different in
nature. The basal group figured for Amblystoma are undoubt-
edly cells of the fundus segment of the glands the latter possessing
but one type of cell,—namely, those which become, in the adult,
zymogen cells. But Ross homologises with these groups, those
of somewhat similar appearance in the pig; and the latter are,
as I have shown,—simply certain very conspicuous ones ot the
fundic segment cells,—namely, the newly differentiated parietals.

How may we know that these are really young parietals?
Because they can be traced, in unbroken lineage, from the 3 em.
stage to the stage just before birth (29 em.), long before which they
are definitive in size, position and all morphologic characters;
because from their first appearance, they exhibit the slight enlarge-
ment as compared with the adelomorphs, the conspicuous intercel-
lular ductules and the cytoplasmic granules, the latter, at all stages,
showing a characteristic affinity for Rubin 8 and Kosin, and stain-
ing black or steel blue with copper chrome hematoxylin, and cop-
per red with neutral gentian. Certain other characteristics are
not so marked until the later stages; such are the polygonal,
spherical or lenticular shape, which succeeds the earlier piriform
shape. The nucleus of the early parietal is ovoid, as are those of
all the gland cells, but by 6 em. many of the older parietals have
spheroidal nuclei. This latter, however, is not a constant char-
acter, as, even in the adult, parietals are found with ovoid nuclei.

494 Edwin G. Kirk.

We have seen, then, that all the cells of the gland tubule, are,
from the first, on the same morphologic level; all reach the surface
and the basement membrane in the early stages; there are no
“basal cells’? in the sense of cells which are shut off from the
stomach lumen by higher cells. Moreover, the parietals are the
first of the adult gastric cell-types to be differentiated. We
shall find that, in the earlier stages,’ they appear only in the lower
parts of the gland tubule.

While the epithelium of all other parts of the stomach has
become granular and refractile, that of the pars oesophagea has
remained absolutely clear and transparent. As had been antici-
pated, no glands appear in the major portion of this territory,
which at 4 em. yet presents a simple columnar epithelium. But
a small peripheral one has already developed glands, by the same
general process described for other regions. Their cells, like
those of the rest of the pars oesophagea, are absolutely clear, and
the distinction from adjacent regions is very sharp, so abrupt that
one is able to compare the last transparent cell with the adjacent
finely granular cell of cardia, fundus or pylorus.

The caecal ridge is covered by simple, granular epithelium which
displays the slight waviness and the numerous distal mitoses
seen in the 2 cm. stage of the fundus and cardia.

In the caecal pouch, development is exceedingly erratic. At
2-3 em. slight irregularities occur; some of these elevations are
broad and being due to unequal growth of the submucosa, cor-
respond, when linear, to the gross rugae, and, when circumscribed,
to the gross papillae or villi, of the adult caecum; others are due
to a thickening of the lamina propria alone, but are very broad,
ar very high, and manifestly have no direct relation to gland
formation, helping to increase the surfacearea. Otherstretches are
yet level and present numerous mitoses in the distal row of nuclei.
Atother places, the first, minute, intraepithelial glandular depres-
sions are appearing. At 4 em. some of these glands are in the
stage of fig. 3, some of fig. 6. They are rather sparsely dis-
tributed. Thus, the small caecal pouch shows a developmental

* Thus up to about 7 em., in the fundus, and about 12 cm. in the cardia.

On the Histogenesis of Gastric Glands. 495

picture as varied as that of all other parts of the stomach together,
including within its limits all types of glandular development
found at these stages in the stomach. It is thus defined by its
gross boundaries, rather than by any well marked histologic or
glandular type. By 7 cm. the rugae and papillae have by the suc- .
cessive appearance of smaller ones on the larger, acquired com-
plex, fantastical shapes, like those seen in a papilloma. The
comparatively few glands appear both on these rugae and in
the depressions between them. Some of them have progressed
to the stages of Figs. 6 and 11, and hence resemble the pyloric
glands in size and form, but have developed some parietal cells.

Mucus cells appear, for the first, in the 6—6.5 cm. stomach, at
which time all the cells in the lesser curve (pyloric) area stain
very definitely and deeply with muchematin. In general, the
surface cells and those near the top of the gland have but a
shallow, distal rim of mucus, the rest of the cytoplasm being of
the ordinary, granular type. In the deep cells of the tubules, the
cytoplasm distal to the ovoid nucleus is of alveolar structure, the
alveoli being filled with mucin. If care is taken to preserve the
tissue from the action of water, the mucus occurs in the form of
spherules, one in each alveolus (fig. 12), and enclosed by the slen-
der alveolar wall. With the three color, or hematoxylin and eosin
stain, the cytoplasmic part of these cells, surrounding the nucleus,
and in the first mentioned type, occupying the whole cell except
the distal rim, presents the ordinary, finely granular, slightly
reddish appearance of the ordinary embryonic cells; the mucus
part appears clear, and has an alveolar network. In the other
parts of the pylorus (greater curve and facies anterior and pos-
terior) no mucus cells have appeared, all the cells being of the
primitive type. As to size and form, the glands are uniform in
all parts of the pylorus.

At 7 cm. the region from C to 4 (fig. 10) represents the terri-
tory of the adult cardia and fundus. There is a gradual and pro-
gressive increase from C to 4 in the age of the glands. Those
toward C are in the intraepithelial stage (fig. 3). They occur
clear up to the caecal ridge, where the epithelium becomes
level. Toward 4 the glands approach the type of fig. 7, but are

496 Edwin G. Kirk.

not quite so deep, the mesoblastic cores of the processes being
smaller. Many glands in the whole area C to 4 possess parietals,
which in the part toward C are isolated, groups being often found
toward 4. Thus cardia and fundus merge insensibly as yet, the
fundus being as always, more precocious. The fundic glands, from
about D to 4, present at 7 cm., the following measurements: The
lumen depth® averages 25y; the process height, 35-40u. Mitoses
now occur in the tubules, mostly about one quarter the distance
from the surface, i.e., intheregion corresponding to the adult neck.

‘At point 4 an abrupt change occurs, the glands from 4 to the
pyloric aperture being almost twice as deep as the fundie glands,
the average lumen depth being 40-50u, the process height,
50-70u. The gland processes are broader (32—40u) than in the
fundie glands. These pyloric glands also occupy the lesser curve,
from the pylorus to point 7, where the pars oesophagea abruptly
begins. The pyloric glands and processes are strikingly sym-
metrical, the former having the form of simple tubes. Many
mitoses are present at all depths, not so many on the surface.

It must not be forgotten that in the cardia, fundus and pylorus
longitudinal rugae occur, some running the whole length of the
stomach. They are not so large as those of the caecum, but are
readily visible with a hand lens. In the cardia, they are especially
large (4-t mm. in diameter), and often papilliform or highly
irregular. The caecal ridges are very high.

Parietals now occur in caecum, cardia and fundus; in the latter,
groups of them. We have seen that the first to appear, came to
occupy only the deeper parts of the tubule. But now, in the deeper
tubules (from 3 to 4, fig. 10) new ones are differentiated farther
up along the sides, at first as single cells, which, by mitosis, soon
give rise to groups in this new position. They never appear in the
upper quarter of the tubule.

At 7 em. the mucous cells have spread down to point 5 on the
greater curve, the pyloric glands from 5 to 4, yet containing no
trace of mucin. The boundary between the mucous and non-

* For brevity the distance, fig. 6, from A to B will be called ‘ ‘lumen depth;” from
A to C “process height.’

497

On the Histogenesis of Gastric Glands.

Three color.

cm.

e
‘

Tubules of cardia near fundus.

isis iil,

10 em. Muchae-

Mucous chief cell from bottom of a pyloric tubule.

Hines 12.
matin.

Adjacent serial sections through a tubule of fundus region. 10 em.

lerees BE
Three color.

498 Edwin G. Kirk.

mucous pyloric tubules is absolutely sharp. Many of the mucous
cells in the gland tubules are mitotic, as are also the embryonic
cells. The transition, then, between fundus and pylorus is
absolute (point 4) and marked by the sudden difference in size of
tubules, and by absence of parietals in the pylorus. Rarely, the
surface fundie cells show a very slight line of mucous stain, very
inconstant, and probably absorbed from the general stomach
content.

From now on, mitotic activity is incessant at all depths of all
the gland tubules. Surface mitoses also occur, but are much rarer.

In the pylorus of 7-8 cm., and fundus of 9-10 cm., secondary
gland processes appear at the bottoms of the primary tubules.
They are of the same general nature as the primary processes,
being intra epithelial for a short time, but being very soon rein-
forced by mesoblast. By this upgrowth they at first render the
lower portion of the tubule compound, but as they progress until
they reach the surface, the result is not a compound tubule,
but two independent glands. This mode of increase of the glands
is very common in fundus and pylorus, in these earlier stages.
In the cardia, the same process occurs later (14—15em.). These
upgrowths are of the same nature as the primary ones, being intra-
epithelial for ashort time, but being speedily reinforced by meso-
blast.

The true compounding occurs somewhat later, with the fail-
ure of these secondary processes to reach the surface. This begins
in the pylorus about 17 cm.,in the fundus about 19-20 cm.,in the
cardia about 21-22 em. During this compounding, there is enor-
mous proliferative activity in the depths of the tubules, as evinced
by the numerous mitoses, and by the rapid downward elongation
of the glands. This applies especially to fundus and pylorus,
but many mitoses are found even in the cardia.

After the gland-compounding has been largely completed (25—
29 cm.), the mitoses are somewhat less frequent, but are still
present in all parts of the tubule, especially in the elongating fun-
dus segments.

1° In the compounding of the cardiac tubules we shall find a complicating factor,
the evagination of parietal tubules.

——— a lo

On the Histogenesis of Gastric Glands. 499

It was deemed best to deal with the stomach as a whole in
the discussion of these earlier stages, up to and inclusive of
7cm. But inasmuch as certain definite gastric regions have now
differentiated, the development of these will, in the interest of
clearness, be followed separately.

2. Pylorus

At 9 cm. all the pyloric epithelium, both surface and glandular,
consists of mucous cells. The deeper tubule cells are of the mu-
cous chief type, those higher in the tubule have shallow thecae,
while the surface cells display a mere distal rim of mucus. No
parietals are present, except in two or three tubules next to the
fundic zone, and in these only a very few scattered ones. Now,
as earlier the fundo-pyloric boundary is very definite. If, as
seems justifiable in the light of the adult structure, our criterion of
a pyloric tubule be the mucous chief cell-lining of the deep seg-
ment, then the fundo-pyloric boundary is absolutely sharp. For,
if adjacent sections stained with muchematin and three color be
compared, we find that the fundic segment of the fundus glands,
even the last, has no mucous cells, while, the fundus of the pyloric
tubules is lined exclusively by mucous chief cells, except for the
scattered parietal or two in the last two or three tubules toward
the fundus.

The pyloric processes are, in general, broader (40-54) than
those of the fundus (25-40u), but resemble the latter in often
possessing expanded or clubbed tops. The pyloric tubules usually
present a very symmetrical test-tube form. Their appearance at
9 cm. is indicated by fig. 21, for although the latter is taken from
a much older embryo (19 cm.) yet the change in the pyloric glands,
after 9 cm. are mainly those of size andof the upgrowthof the en-
closing ridges rather than any fundamental alterations in form and
cytology. In the pyloric tubules, as in those of the fundic region,
there are from now on, many mitoses, and at all depths. They
also occur in the ridge cells, although not so frequently.

At 10-11 cm., the deep cells are so packed with mucus that
the nuclei are often flattened; the goblets of surface and foveola
cells are also becoming deeper.

500 Edwin G. Kirk.

At 15 em., many of the surface cells have gobletsreaching clear
to the nucleus, while some show merely a superficial rim, there
being no regularity in the distribution of these types.

At 18 em., the mucous chief cells line the lower third or
quarter of the tubule, the goblets the upper 3 or #, the deeper in
position having, as hitherto, the shallower goblets. At 26 em., all
the surface and foveolar cells have very deep goblets.

Thus cyto-differentiation in the pylorus is practically com-
plete at 9em. The size of the cells, the depth of the goblets and
other details alter somewhat later, but, microchemically and pre-
sumably, metabolically, the cells have then reached their definitive
condition. I do not mean to imply that they have reached a ter-
minus of potential differentiation. Harvey’s work (’07) shows
that very probably even the adult gastric cells, highly specialized
as we are accustomed to consider them, may under changed condi-
tions, pathologic or experimental, assume other forms and func-
tions. But this seems not to occur in normal, embryological
cyto-differentiation in the pylorus.

Measurements were taken of glands from each region at all
stages but inasmuch as such figures have no general embryo-
logical value, I quote only afew, and these for the purpose of com-
paring the rapidity of growth in these regions. Such figures are
necessarily only approximate, as there are considerable differences
between those from embryos of the same length, and also between
adjacent tubules.

Sem. 16cm. 19cm 23cm. 29cm.

Cardiac ies sete oat 35-50u 80u 105u 112u 160.

EIN CUS er eee 70u 110u 140u 165u 210u

Pvlorasteneciocetcor tees 80-110u 130u 160u 208u 240u
3. Fundus

From the first we have found considerable difference between
cardiac and fundic tubules, but the two districts have always
merged insensibly and the distinctions have been those of size and
age, depending on the general precocity of the right and retarda-

a

On the Histogenesis of Gastric Glands. 501

Fic. 14. Cardia. 15cm. Three color.

Fig. 15. Fundustubules. 15cm. Note ductules of the parietal cells and ‘‘lat-
eral parietal rows.’’ Three color.

Fic. 16. Cardiac tubules. 18 ¢m. Three color.

Fig. 17. Cardia. 19cm. Three color.

502 Edwin .G. Kirk.

tion of the left region, rather than on any true, differential diver-
gence. Nor, indeed, shall we expect to find, at any time, a sharp
boundary comparable to that between the fundus and pylorus—
for such does not exist in the adult pig. (Vide Greenwood ’85;
Bensley ’02, p. 128) . Hence, the somewhat arbitrary nature of
the separation for descriptive purposes, of these two regions,
should be constantly borne in mind.

After 7 cm., the parietals are no longer confined to the deepest
part of the fundic tubule, but invade all but the upper quarter.
Some groups contain 10-12 cells. Especially characteristic from
this time on is what we may call the “lateral parietal row.’’ This
is a row of parietals, one or two abreast, extending, often, from a
point # up the tubule, either to the very bottom, or only part
way down (fig. 15). The occurrence as a row is, of course, deter-
mined only by reconstruction and by cross sections, and has been
often verified thus. In cross section such a tubule will show one
or two parietals, all the others being adelomorphs. These lateral
rows occur even in the last stages before birth. However, the
haphazard distribution of parietals ordinarily described is also
found, isolated parietals, or groups of 2-3, or even 10-12, occurring
from now on, not only in rows, but also in superficies. Fig. 13
represents serial sections through a gland-tubule of 10 cm., and
illustrates the partial differentiation of the fundic elements into
parietals, staining a brilliant red in fuchsin §.

The ductules in relation to the parietal cells are present at all
subsequent stages, and hereafter, in this paper, their presence will
be taken for granted. Often a ductule ramifies, sending branches
between three or four parietals, or two branches between the same
two parietals. These intercellular ductules are limited by cement
lines, as shown beautifully by neutral gentian. Most of the parie-
tals, at 8 cm. and later, display clear vacuoles, sometimes as
clear areas surrounding the nucleus, but oftenlyingin the periph-
eral cytoplasm. Harvey (’07) Hamburger and others describe
these in the adult parietal (vde figs. 18, 20). In general, they
become more constant, numerous and large inthelaterstages. Of
course the obvious inference is that they represent droplets of
secretion to be poured into the intercellular ductules. [havenever

Fie. 18. Cardia. 19 cm. Three color. The cells represented in outline are
surface mucous cells.

Fig. 19. Fundus. 19 em. A complete tubule. Three color.

Fie. 20. Fundus. 19 em. Bottom of a tubule, showing parietals and adelo-
morph cells. Copper chrome hematoxylin.

504 Edwin G. Kirk.

been able, by Golgiimpregnation, although I have tried many times,
to demonstrate, even in the latest embryonic stages, any contin-
uity between these vacuoles and the ductules.

After 7 cm. mitoses are very frequent in the fundus tubules,
occurring at all depths, but especially at the very bottom and at
the juncture of the first (upper) and second quarter, 7. e., in the
territory corresponding in the adult, to the lower part of the fove-
ola. Most are in theadelomorphs, nevertheless parietalsare often
caught in mitosis. At 8-9 em., every tubule of the fundus
region has acquired some parietal cells. At 9 cm., and later, pari-
etals are found with constricted nucleus. These I interpreted
as examples of amitosis. Possibly they presagea cessation of cyto-
plasmic division. In later stages (20-29 em.) multinucleate parie-
tals appear. After 10-11 em. the actual and relative increase of
parietals in the cervical region, and relative decrease in the bottom
of the tubule is marked. The latter is due to the proliferative
activity of the undifferentiated gland cells, by which the tube
is constantly lengthening downward. Mitoses also occur on the
processes but not so frequently.

At 9-10 cm., no mucus is present in any fundic cell. At 12-
13 em., this is yet true for the large part of the fundus, but, near
the pylorus, along the greater curve, very shallow, distal goblets
of mucus appear in the cells of the surface and upper quarter of
the tubule. At 15 cm. in this part of the fundus, the surface
cells have acquired quite deep goblets, while in the upper third
of the tubule are shallower goblets, and a few mucous chief cells.
In the stages 15—20em. this mucous differentiation of surface and
foveolar cells spreads rather gradually to the other parts of the
fundus.?!

Thus, even at 17 em. we find a condition not far advanced,
in this respect, over that of 13 em.,—the surface and foveolar
cells of some areas presenting but narrow rims of mucus, and the
neck chief cells staining but faintly, or not at all, in muchematin.

‘This occurs with striking slowness as compared with its rapidity in the pylo-
rus. Moreover, in the latter, the gland cells at the very bottom were the first to
manufacture mucin, the pyloric surface cells of some embryos showing no mucus
as late as 12-18 cm, but the gland mucus chief cells staining heavily.

On the Histogenesis of Gastric Glands. 505

Fig. 15 represents the appearance of an average fundic gland
of 12 to 16 em., but it must be remembered that reconstruction
shows a preponderance of parietals in the 2d and 3d quarters,
over the number in the deepest fourth, also that in some parts
of the fundus the surface and upper tubule cells may have
acquired shallow goblets, which appear clear and transparent in
the three color stain. It will be seen that the tubule has been
lengthened by upgrowth of the gland processes. The latter are
often clubbed or expanded at the tops and sometimes irregular.
The upper + or } of the tubule devoid of parietals, and with cells
already sometimes bordered with mucus, represents the future
foveola. The rest of the tubule contains the cells from which
will be derived all the epithelium of the adult cervix and fundus.
Already (12-16 em.) the parietals are preponderating in the middle
third of the tubule, while the embryonic gland cells, in the deep
+, are multiplying rapidly mitotically, thus deepening the tubule,
and incidentally furnishing the material from which, as we shall
see later, the zymogenic cells are to differentiate. Forthe present,
these adelomorphs are, in every way, as far as the staining methods
at my disposal show, the same as the primitive, embryonic gland
cells. They have become however, somewhat more definitely
cubic or low cylindric.

This description of tubules applies to the large part of the
fundus, including those partsnearthepylorus. Inthe fundic areas
toward the cardia, we find that after 11-12 cm. not only are the
foveola and surface mucous cells present, but occasionally cells
in the deeper parts of the tubule tinge definitely with muchematin.
At 14-15 em. some of the tubules are lined to the bottom by mucous
and parietal cells, the mucus cells in the depths being of the mucous
chief type, those in the upper third shallow or deeper goblet cells.
Interspersed with such tubules are some with three cell types—
mucous, parietal and adelomorphs; and these tubules predom-
inate more and more in progressing toward the right, until at
last the typical tubules are reached. These left fundic tubules
have as far as cytodifferentiation is concerned, reached the de-
finitive condition. They constitute in embryos as in adult, a
zone transitional, in all respects, between cardia and fundus.

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 4.

506 Edwin G. Kirk.

In embyro and adult there is found, in passing through this zone
from left to right, a progressive decrease in mucous chief cells,
an increase of embryonic gland cells (or, inthe adult, of serous chief
cells) a gradual lengthening of the whole gland, and a gradual
relative shortening of the foveola and lengthening of the fundus
segment.

Thus at 17-19 em., in the typical fundus area, the bottom of
the tubule is lined by adelomorphs and parietals, the former
preponderating. The former do not tinge with muchematin, up
to the time of their final conversion into zymogen cells, for such
will soon be their fate. In other words, those cells destined to
become the serous chief cells, do not pass through a mucous stage,
but pass directly from the embryonic form to the complete zymo- .
genic cell. Up to 19 em., neutral gentian, aside from tinting the
cytoplasm of all the cells a rather faint, diffuse violet or blue,
stains no granules, except the moderately course ones of the pari-
etal cells. At 19 cm. granules of a new type appear in the distal
zone of those cells of the fundic segment of the fundic glands,
which have hitherto preserved the embryonic type. These
granules stain, from the first,.a dense blue. At the time of their
appearance they are slightly larger than the parietal cell granules.
They do not appear simultaneously in all the adelomorphs, but
at first in scattered ones, spreading rapidly to the other cells of
this type, so that, at 25 em. no embryonic gland cells remain in
the fundus, all having differentiated into serous chief or zymo-
genic cells. The granules increase rapidly in size, so that they
are soon much larger than those of the parietals (fig. 22). In
some cells, from the first, they are not limited to the distal zone,
but occur throughout that part of the cytoplasm distal to the
nucleus. The cytodifferentiation of the fundic glands seems now
complete. |

The remaining changes are merely such growth processes and
shifts in the relative size and position of parts, as are necessary to
bring about the definitive form and positions. As these undoubt-
edly display much generic, or even specific variation, they can
have no general developmental significance, such as possessed by
the details of cytodifferentiation, and will, therefore be described
but briefly.

On the Histogenesis of Gastric Glands. 507

Fig. 21. Pylorus. 19 cm. Three color.

K

508 Edwin G. Kirk.

In cross sections of tubules at 19 em. most of the parietals are
shown to be in line with the other cells (fig. 23) but here and there
a few are assuming the true parietal position. At all stages of this
process, the ductules preserve their direct continuity with the
gland lumen. The parietals here, as in the cardia, are, by the
24 cm. stage, much larger than the chief cells and of a rounded or
ovoid form.

At 19-20 cm., by the progressive upgrowth of the gland proc-
ess the foveola (part above the region of parietals) has length-
ened very perceptibly, so that now it constitutes about one-half
the total tubule length (fig. 19). The deep goblets of the foveola
(a, fig. 19 are replaced below, in the cervical region, with mucous
chief cells (b), with which parietals (c) are interspersed. The
lower third is lined by serous chief cells, adelamorphs and a few
parietals. The adelomorphs and serous chief cells are not dis-
_ tinguishable in the three color stain of fig. 19. Both are in fre-
quent mitoses.

After 19 em. by elongation of the tubule part (cervix and fun-
dus segments) the proportions of tubule length and foveolar
length are rapidly altered so that at 25 cm. the foveola : tubule :
1: 2. This proportion is preserved up to the time of birth.

4. Cardia and caecum

At 7-9 em. the cardiac glands are mostly either in the intra-
epithelial stage or in stages between those of figs. 3 and 6. Al-
most all, except the very youngest, have one or two parietals.
The cardiac glands haye now extended to the summit of the caecal
swelling, being here of the intra-epithelial type. At 7 cm. all the
cells are either adelomorphs or parietals. The later are at first,
as in the fundus, confined to the lower part of the tubule (fig. 11)
but at 10-11 cm. appear higher up, as in the fundic tubules of
7-8 cm. Lateral parietal rows soon become very common
(13 cm.)

At 11-12 em. and thereafter, differentiation of the surface cells
into mucous cells occurs. Some acquire a mere distal rim of
mucus, some a deep goblet. At 13-14 cm. the mucous change

On the Histogenesis of Gastric Glaads. 509

extends into some of the tubules, sometimes to the very bottom.
These deep mucous cells are then of the mucous chief type, the
whole cytoplasm distal to the nucleus being infiltrated by the
stainable substance. Thus, mucous differentiation occurs much
later than in the pylorus, and somewhat earlier than in the fundus.
However, the process lacks uniformity, for, as late as 14-15 cm.
the cardia of some embryos exhibits mucous cells only on the
surface, and a short distance into the tubule, with only an occa-
sional one in the deeper parts of tubules.

The parietals, are, of course, unstained in muchematin, but
are, as always, readily distinguishable (as, indeed, even in un-
stained specimens, fixed in Bensley’s fluid) by high refractility,
absolute opacity, and glistening shiny appearance. Moreover
the ductules are very apparent. The adelomorphs are clear and
transparent, though very finely granular.

At 17-18 em. all the cardic cells are either mucous or pari-
etal. The cells of the surface, for a little way into the tubule,
possess shallow to deep goblets. The deep cells, aside from parie-
tals are all of the mucuos chief type. Fig. 14 illustrates the
typical surface goblet cell (a) and the shallow goblets (b), which
go over by gradual transitions into the deeper mucous chief cells
(ec). In some tubules the goblet cells extend quite to the bottom
but it should be noted that, in embryonic stages, the distinction
between mucous chief and goblet cells is often not so sharp as in
adult life, for, in the surface and upper tubule cells, the mucus
often seems not to be homogeneous, but separated into minute
goblets by a cytoplasmic mesh. Karyokineses are frequent in
the mucous chief and goblet cells. Even the deep surface goblet
cells are found in mitosis frequently.

From 14 cm. on, a marked form-divergence js added to the
difference in size between cardiac and fundic tubules, the former
now becoming shallower (wider relative to the depth), while the
fundic tubules preserve their accustomed narrower, deeper form,
so that, after 14 cm. it is easily possible to distinguish the two
under low power (compare figs. 16, cardia, with 15 and 22,
fundus). The cardiac epithelium has, from now on, a curious
tendency to shrink away from the underlying mucosa, surface

510 Edwin G. Kirk.

epithelium and tubules cohering so that the whole epithelial area
loosens en masse. This tendency is observable from 14 to about
20 cm. in almost all stomachs, and is doubtless referable to the
shallowness of the pits, taken in conjunction of course, with the
fixation shrinkage.

The upper } or + of the primary cardiac tubule, devoid of parie-
tals and lined by goblet cells, represents the adult foveola, for
the lineage may be traced, without any break of continuity, to
the 29 em. stage, long before which all the cena parts of the
tubule are readily distinguished. The lower 3 or #, lined by groups
of parietals, interspersed with a large amine of mucous cells,
gives rise later (20-22 em.) to the tubules proper.

At 15 em. certain of the groups of parietals begin to push
outward slightly (fig. 16). These invaginations, at first shallow,
may occur wherever groups of parietals occur,—namely, in
the depths of the tubule, and in all but the upper quarter. A¢ all
stages they consist entirely of parietals. They appear with in-
creasing frequency in the later stages, and become definite,
secondary tubules (figs. 17 and 18) by 18-19 em. Those from
the sides of the pomary tubule are directed outward and down-
ward; those from the bottom, straight downward. These sec-
pada tubules are narrower than the primary ones, and at 19
em. vary from the merest incipient outpouchings to complete,
though short, tubules. No similar process occurs in the fundvs
or pylorus at any stage.”

About 20 em., the compounding of the lower part of the prmary
tubule occurs in the usual way, so that, at 22 em., several tubules
open into a foveola. Some of these tubules are purely of parietal
cells; these were partially derived from the parietal evaginations.
Some are purely of mucus cells. Many display both types, and in
all proportions. These three types of cardiac tubules persist up to
the time of birth (29 em). I do not believe that the formation of
these parietal tubules differs in essential mechanism from the or-

12 This unexpected finding agrees absolutely with Toldt’s observation in the cat,
that some tubules arise as lateral buds, which grow out and downward. They arise
at a parietal cell, and the latter by division goes over into the new tubule.

On the Histogenesis of Gastric Glands. 511

Aten

y

Fig. 22. Complete tubule of fundic region. 23 cm. Neutral gentian. Cyto-
differentiation is practically complete. (29 cm is about the birth-length).

Fria. 23. Cross section of fundic tubule. 23 cm. Three color.

Fig. 24. Cardia. 25 em. a. Foveolae. Mucous chief and goblet cells. 6.
Tubules. Mucous chief and parietal cells.

Fias. 25 and 26. Cross and longitudinal sections taken at the bottom of a
cardiac tubule. 29cm. (just before birth). This shows the parietals, which later,
by the third week of post-uterine life, disappear or become converted into mucous
cells.

512 ; Edwin G. Kirk.

dinary process of compounding, found in fundus, pylorus and also
in the cardia. In each type, the epithelium seems to display the
initial irregularity, but soon mesoblastic growth seems to become
the active factor. I wish to emphasize the point that there are
never, at this time, any transitions between the parietals and the
mucous cells, each of which are, in the cardiac tubules, sharply
distinguishable by the criteria mentioned above.

While the lower part of the tubule lined by mucous chief and
parietals, is being compounded, a very rapid growth of the gland
processes elongates greatly the wide, upper part lined by goblet
1.e., the foveola (19-25 em.) Thus at 18 cm. (fig. 16) the foveola
constitutes only the upper third or fourth. At 22.5 cm. the meas-
urements are, foveola 48y, tubule 64y, total 1124 or foveolar
length to tubule length as 3 to 4. At 25 em. the foveolarlengthis
to the tubular length as 5 to 8, and the total length is 140y, so
that the tubule portion has scarcely lengthened since22cm. Thus
the definitive form and proportions are practically reached at
25 em. (fig 24) and from now on, foveola and tubules grow with
about equal rapidity.

As in all earlier stages, there is, at 25 to 29 cm., a gradual
transition from undoubted cardiac to undoubted Prete tubules,
the foveolae, in approaching the fundic region, becoming relate
and absolutely shorter, but the tubules so much longer that the
whole gland length is greater.

At 29 em. the cardic tubule measures 160 in length, the un-
doubted fundic 210, and the pyloric 240.

In stages 26-29 cm. the parietals are just as numerous and
just as well defined (figs. 25 and 26) as ever, staining with nor-
mal brilliancy, and possessing ductules and all the characteristic
structures. But the cardiac glands of the adult pig contain no
parietals, only mucous chief and thecal cells. Thus the parietals
must either degenerate, or become transformed into mucous cells,
during the intervening period. I have been unable to obtain
stomachs of young pigs. I suspect, from the results of Cade and
Harvey (’07) that the parietals may transform into mucus cells.
I have to report on this in a subsequent paper.

One more point in the cytodifferentiation of the cardia should

On the Histogenesis of Gastric Glands. 513

be mentioned. The neutral gentian stain was used on every stage
of cardiac development. This was done purely as a matter of
routine, as I never anticipated finding anything in the cardiac
tubules staining specifically with it, except the copper red granules
of the parietals. But at 25 cm. and thereabouts granules resem-
bling zymogenic granules appear in the deeper cells of undoubted
cardiac tubules. Adjacent sections stained with muchematin
show these same cells tobe mucous chief cellsof the ordinary type,
so that the granules lie within the mucous parts of the cell. These
granules are found intracellularly for one or two (em.) stages,
and after that disappear. I did not find them in the extreme left
cardiac region.

At birth, the cytodifferentiation of the cardia is incomplete, or,
rather, that involution required to bring about the adult cyto-
logic status, with its single type of gland cell, has not yet begun.!

Caecum. At 10 cm. the glands are rather sparsely distributed.
Many are as deep as the pyloric tubules, but contain few or many
parietals. By 15 cm. the glands are almost as thickly distributed
as in cardia or fundus, the parietals are abundant, and the tubules
have the general characters of those of cardia and fundus, ap-
proaching more the deeper form of the fundic tubules. In later
stages this region undergoes the same change as the general
cardia.

5. Pars oesophagea

The greater part of this region gradually becomes stratified
during the 6 to 10 cm. stages, the cells still remaining clear and
transparent, except those of the deepest layer, which have acquired
a finely granular cytoplasm of the ordinary type. The clear cells
of the superficial strata have already begun to flatten out. At
the periphery of the stratified area, the columnar cells constituting
the deepest layer, are continuous with the simple columnar epi-
thelium of the surrounding zone. We have seen that at 4-6 cm.
a narrow, peripheral zone of this region has developed gland

8 Negrini, 1886, noted that parietals are present in the foetal, but absent in
the adult cardia; also Bensley, 1903.

514 Edwin G. Kirk.

tubules. These glands display the same form-development as
the others. They are more precocious than the cardiac tubules,
and deeper progressively, keeping pace with the pyloric tubules
in development. At 11 em. they still consist of clear, trans-
parent cells. At this stage, then, we find the major and central
portion of the pars oesophagea to be of stratified squamous epi-
thelium; outside this is a zone of simple columnar epithelium
which displays the glands at the very periphery. At 11 cm. the
boundary between the clear cells of pars oesophagea and granu-
lar cells of adjacent cardia, fundus and pylorus is sharp to a cell.
I have not been able to settle definitely the fate of this glandular
zone in later stages. It seems, by differentiation of the cells,
to merge with the adjacent cardia.

6. Conclusions and theoretical deductions

a. Homologies and phylogeny of the cardia

We have seen that fundic and cardiac glands display an abso-
lutely parallel development in the earlier stages, the cardia, how-
ever, lagging behind, from the first. In each the parietals appear
at a corresponding stage. It is impossible to distinguish an iso-
lated 8 em. fundic from a 12-13 em. cardiac gland. After 138-15
em. however, development in the cardia is retarded; while the
fundic tubules elongate rapidly and become narrow, the cardiac
tubules lengthen but slowly, and assume a shallow contour.

At the same time a significant parallelism appears in the cyto-
differentiation. The foveolar region of each, with goblet cells,
appear to be, at all stages, homologous. The lower part of the 16-
17 cm. cardiac tubules corresponds, cytologically, to the neck
region of the young fundus tubule, each being composed of mucous
chief and parietal cells. The length of the whole cardiac tubule at
this stage, is about that of foveola plus the neck of the fundus
gland.

In the fundice gland of 20 em. the body and neck segements
become compound. In the cardiac gland of 25 em. the body seg-
ment is evidently absent, barring the very temporary appearance

On the Histogenesis of Gastric Glands. 515

of a few zymogenic cells in the deepest part, but the neck segment,
like the corresponding part of the fundic gland, has become split
up into several tubules, which display the two cells characteristic
of the neck of the fundic gland. While the aberrant development
of zymogen granules is confirmatory, I lay most emphasis on the
early parallelism in form-development, and especially on the
presence, distribution and remarkable persistence of parietals
in the cardiac tubules, as demonstrating their affinity with the
fundie tubules.

By all these findings we are inevitably brought to one conclusion
with reference to the genetic relation of cardia and fundus. The
cardia represents a part of the fundus which has undergone partial
involution. The regressive changes have manifested themselves
first in the general chronologic retardation of ontogenetic develop-
ment in the left part of the old cardio-fundic region; second, in
the phylogenetic disappearance of the fundic segment of the tubule
with its zymogenic cells,—the curious, delayed appearance of
zymogenic granules at one stage of the developing cardia repre-
senting, probably, the last vestige of this segment; third, in the
ontogenetic transformation of the cervix of the old fundus gland,
with its mucous and parietal cells, into the tubule of the adult
cardia with its mucous chief cell. This last process I have not
seen, but it must needs occur, for we have the first and last terms
of the series before us.

The zone of fundic tubules, lying between undoubted cardia
and the ordinary fundic zone, represents an intermediate stage
in the regressive process.

The facts of development, then, show that the cardiac glands
are not primitive, but regressive structures. For the early stages
do not resemble, in any way, the condition found in lower verte-
brates, where, for example, even the fundus glands possess no
definitely identified parietals. Moreover, no fact in the develop-
mental history so much as suggests a possible derivation from
the oesophagus.

Thus, our embryological findings seem to confirm strongly
the conclusion as to the origin of the cardiac glands reached by
Bensley (’02), working from the standpoint of adult comparative

516 Edwin G. Kirk.

anatomy and histology. It has been pointed out that general,
mucous differentiation occurs slightly earlier in the cardia than
in the fundus, although it is less uniform. The cardiac glands, in
their whole development, display a very decided mucous char-
acter, as manifested in the discarding of the deep, zymogenic
segment of the fundic tubule, in the involution of parietal cell,
and in the slight acceleration of the mucous differentiation. This
must be a response to a functional demand. Bensley has already
suggested (02 p. 147) that in these retrogressive glands, the
cellular types disappear in the order of their specialization,—
zymogenics first, parietals next. We find that not only is there
no tendency on the part of the mucous cells to disappear, but
that they even differentiate at a. slightly earlier stage. Probably
the altered food, or the other conditions which have brought
about the involution of fundie to cardiac glands, have also de-
manded an increased mucus secretion.

It will be recalled that, while part of the caecal glands were
as retarded as those of the cardia proper, others kept pace with
the pyloric tubules, both types, however, developing parietals.
Later, all the caecal glands, whether precocious or retarded,
shared the fate of the cardiac tubules. This would seem to in-
dicate that seclusion in the secondary pouch has tended to parti-
ally protect these galnds from the regressive changes which have
attacked the old left fundic area.

Bensley’s comparative work on the mammalian stomach (Op.
cit. ’02) led him to the same conclusion,

b. Cell specificity

(

At 2 cm. all the cells are of the type described as ‘‘embryonic
gland cells.” From 3 em. on, some of these are constantly dif-
ferentiating into parietals, others retaining their undifferentiated
character for a shorter or longer period. Both types multiply
mitotically.

From 6 em. on, certain of the adelomorphs differentiate into
mucous cells, both mucous chief and goblet. New parietals and
new mucous cells arise, differentiating from adelomorphs. The

On the Histogenesis of Gastric Glands. aay

residual adelomorphs of the deepest part of the fundic tubules
differentiate at 19-20 cm. into zymogenic or serous chief cells,
and then multiply mitotically.

Thus the parietals are the first of the adult cell types to appear.
Since the completion of this work, Dr. Bensley has pointed out
that we have here a very remarkable and extreme instance of
the throwing back of a coenogenetic character (parietals are not
definitely identifiable in lower vertebrates) into the earliest
ontogenetic stage,—a process called by Cope and Hyatt ‘‘accel-
eration.”

Phylogenetically, zymogen cells (wde Amphibia), and espe-
cially mucous cells, are much more primitive, yet they here
appear in ontogeny later than the parietals. Dr. Bensley has
suggested that the early appearance of the intracellular ductules
perhaps points to a functional explanation of this otherwise puz-
zling ontogenetic anticipation.

Taken per se, the facts of cytodifferentiation, as recounted
above, seem to throw no definite light on the problem of cell
specificity. But studied in conjunction with Harvey’s findings
(07) they point, I believe, very definitely to certain conclusions.

The cytodifferentiation proceeds, as if the cells all started
with the same potentialities or character complexes. These
might be represented by a, b, and c. In some cells those meta-
bolic character complexes represented by a become, as develop-
ment proceeds, dominant, while 6 and c are dormant. In other
cells, character complex b may dominate, and so on. But the
dormant character may, under experimentally (Harvey) or pathol-
logically (Cade, ete., cited by Harvey) altered conditions, become
the dominant ones.

The results of the present work seem to indicate that this lat-
ter process does not occur in the normal course of histogenesis.
The embryonic cells differentiate directly into the definitive types
which then give rise, by division, to cell generations of the same
specialized type. Thus, if the zymogenic characters have become
dominant in a given cell, during the cytodifferentiation, then
the off-spring of this cell are al! zymogenic, but, nevertheless,
the dormant parietal and mucus characters are transmitted to

518 Edwin G. Kirk.

each new generation and are available, if altered conditions make
it desirable or imperative that they should become dominant.
This view of the morphologic flexibility of ‘‘specialized”’ cells,
under changed conditions will be recognized as O. Hertwig’s
doctrine of the nature of cell specialization, under a slightly
different guise. But this conception is also expressable in terms
of Weismann’s Keim-plasma theory, provided sufficient emphasis
be laid on the accessory Keim-plasms.

It seems probable that the mucigenic character of the large
proportion of cardiac mucus cells;—namely, those derived from
the mucus cells of the foveolar and cervical segments of the old
left fundus glands, is palingenetic. On the other hand, it is prob-
able that some of the deeply situated mucous cells are coenogen-
etically mucous, having passed, in race history, through the zymo-
genic or parietal stages.

The temporary occurrence of zymogen in the deeper mucous
cells seems to indicate this, and to suggest that here the reserve
zymogenic characters have not yielded, without a struggle, to
the mucigenic, the old dominance of the zymogenic having become
through the habit of ages, too strongly impressed on the cell
metabolism to be rapidly effaced by the coenogenetic dominance
of the mucous characters.

Bensley (00) found, in the developing of gastric glands of
Amblystoma, cells which contained for a brief period, both zymo-
genic and mucigenic granules. Thus it seems that sometimes, in
the embryonic and possibly in the adult stages,—the cell meta-
bolism may be controlled by two distinct sets of determinants,—
through the manifestation of either of which we are ordinarily
inclined to consider a cell as ‘‘specialized.’’ Such instances seem
to be uncommon, and it is probable that sooner or later,—one
metabolic complex takes a subordinate place,—at least in the
metazoa.

The fate of the cardiac parietals, after birth, promises to be of
the greatest interest in this connection, as there are only two
possibilities open to these cells —degeneration, or conversion into
mucous cells,

On the Histogenesis of Gastric Glands. 519
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1903 Differentiation of Elements in Gastric Glands. Am. Jour. Anat. vol.
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CoupDEREAU. Structure et fonctions des glandes de l’estomac, etc. Travaux
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Fiscuu. Beitrige zur normalen u. pathologischen Hist. des Saéuglingsmagens.
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GREENWOOD. Observations of the Gastric Glands of the Pig. Jour. of Physiol.,
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GRIFFINI AND VASSALE. Sulla riproduzione della mucosa gastrica. Imprimerie
1888 de la Société Typographique. Modéne. Also, ‘‘Ueber die Repro-

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Harvey. Structure of Gastric Glands of Dog after Gastroenterotomy, etc.

1907 Am. Jour. Anat. vol. 6, no. 2, pp. 207-243.

KaLopotHakts. La structure normale del’estomac chez le foetus. Bull. de la
1894 Soc. Anat. de Paris. Année 69, s. 5, tome 8, fse. 19, p. 685-696.
Kouuiker. Mikr. Anat., bd. 2—2 Halfte, Abt. 1, s. 199, or Handbuch der Gewebe-

1852 lehre des Menschen, Erste Aufl.
1861 Entwgesch. des Menschens u. d. Héh. Thiere. Leipzig. s. 368.

1884 Entwgesch. des. Menschens u. d. Hoh. Thiere. 2ndAufl. s. 360.
Laskowsky. Ueber die Entwickelung der Magenwand. Sitzuwngsber. der Wiener

1868 Akad. Math. naturw. Klasse. bd. 58, Abt. 2.
MonranierR. De la Différenciation des éléments des glandes gastriques chez
1889 le foetus. Comptes rendus de la Société de Biol. Par. Tome 1, série

9, no. 16, p. 314-316.
NecRINI. Intorno allo sviluppo e struttura della mucosa gastrica del majale.
1886 Giorn. di anat., fisiol. e patholog. degli animali. 18, 121, Pisa.
NeEuMANN. Flimmerepithelim Oesaphagus menschlicher Embryonen. Archiv. f.
1876 Mikr. Anat. 12, s. 570-574.
OprEeL. 1896. Lehrbuch d. vergl. mikr. Anat. d. Wirbelth. Erster Teil. Jena.

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1886 cardiaque. Compt. rend. Soc. de. Biol. Paris. Juillet, Tome 3,
série 8,

PituintT. Sur l’évolution des cellules glandulaires de l’estomac chez l’homme

1887 et les vertébrés. Journ. de l'anat. et de la physiol. +. 23, No. 5,

pp. 463-497.
Remak. 1855, Untersuchungen iber die Entwickelung der Wirbelthiere. Berl.s. 113.
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1903 Amblystoma and Pig.”’ Biol. Bull. of Marine Biol. Lab. Woods’
Hole, Mass. vol. 4, no. 2, Jan. pp. 66-95.
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bd. 7, Arch. ital. de biologie. Tome 14.
ScHENK. 1874. Lehrbuch der vergleichenden Embryologie, s. 117.
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1878 Jour. Physiol. vol.i, pp. 321-334, pl. 12.
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1876 f. Biologie. 12 bd., s. 225. 1876.

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1888 u. des Epithels des Magendarmkanlas. Anat. Anz. 3, Jahrg.
no. 26, s. 781-784.

HampBurGcer. Beitriige zur Kenntniss der Zellen in den Magendriisen. Archiv
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1906 :

McGitu. Histogenesis of Smooth Muscle in Pig. Internat. Monatsschrift f. Anat.
1907 u. Physiol. bd. 24, Hefte 4-6, p. 209. pl. 7-11.

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Sr6ur. 1882. Zur Physiologie der Tonsillen. Biol. Centralbl. 2, s. 368-370.

THE DEVELOPMENT OF THE MUCOUS MEMBRANE
OF THE GSOPHAGUS, STOMACH AND SMALL
INTESTINE IN THE HUMAN EMBRYO

FRANKLIN P. JOHNSON

From the Department of Comparative Anatomy, Harvard Medical School

WITH TWENTY-FOUR FIGURES

SEVEN PLATES

The development of the mucous membrane of the digestive
tract, although studied for many years by competent observers,
still affords opportunity for further investigation. The present
work was undertaken for the purpose of studying the development.
of the structures found in the digestive tract, and to obtain a com-
parable series of wax reconstructions illustrating the changes
that take place in the form of the mucosa. Especial stress has
been laid on the development of vacuoles and diverticula, villi,
glands, and folds. It was originally intended to include the de-
velopment of the mucous membrane of the vermiform process
and the large intestine, but for lack of favorable material, this
has been omitted in the present paper. This work was suggested
by Dr. F. T. Lewis, and has been done in connection with his
chapter on the digestive tract for Keibel’s Text Book of Human
Embryology. A more complete review of the literature than is
here given, will be found in Dr. Lewis’ chapter.

The material used was obtained from human embryos in the
Harvard Collection. The earlier stages were already prepared in
the embryological collection. The crownrump lengths and series
numbers of these are as follows:

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10, NO. 4.

522 Franklin P. Johnson.

LENGTHIN MM. Series No. LENGTHIN MM. Serres No.
Oe MA eR ae ORE TES 256 DS Sh OR AE Se Rene ee 181
LOR Lee a ae ee ats a 1000 A ay: RE Te a 24
LO retour ee ee ee is tee 1322 DONS eos PRE OTE 914
LO aera ee ee ee 819 BO oe et RR eee ee 913
1.Qis, 2 ie eal stern ee pe aa da i ee 828 RY PARE I RE CM nik it colt ee 649
DO ae be REE Sa ee Cee tee SET BY (ae Wig Re amare) eo bh ee 820
DOS ON eR Vey alee eae eee 871 7 ae es Se ge ree PRT) Re a 888
Dats Pet et ede cin Bers Rotvat ocean Ne 737 78 (incomplete series).....723,724

The crown-rump lengths and numbers of the older embryos used
are as follows:

LENGTH IN MM. SERIES No. LENGTHIN MM. SERIzEs No.
515 RE Cn (Tee, At Eee en a eat 249 20 aos wing So ee Oe Oe 203
Ta een tart he. arititas oes pg Ole IS ae 116 BAN rere eR yee, aa eae en eee 30
Otten Raya PE ergs 142 WA eaten do hee secant teres Woke 131
Oe hts Ac Np eae oben te rot: 224 oY Peles ein nrreds Ine Eiat oe eps shen <A 3 315
ON tat eases ine Oe Leyes pepe cen Ne 340 DAO eek EMy paket et een te ee ea 186

LO as ena yet Cie dye Pe 342 ING WORM cinatee cee eee 341

From the older embryos, the different parts of the digestive
tract were separately removed, imbedded in paraffin, and cut in
serial sections of 8 microns in thickness. The sections were stained
by different methods, among which were Heidenhain’s iron hema-
toxylin, Hansen’s iron hemotoxylin, and Mallory’s aniline blue
connective tissue stain. Both eosin and orange G were used as
counter stains. From the preparations thus made, certain stages
of development were selected, and a number of wax reconstructions
made. The models have in most cases been made to the magnifi-
cation of 145 diameters, so as to be easily compared. However,
in some of the gland models, this magnification was doubled, and
in others trebled.

GSOPHAGUS

Early Stages. In an embryo of 7.5 mm., the cesophagus is a
cylindrical tube of epithelium extending from the pharynx to
the stomach. The lumen is round and relatively large at its upper
end, but tapers gradually until at its middle it is quite small.
Passing downward from this region, the cesophagus gradually
increases in size and at its lower end leads into the stomach, there

Mucous Membrane in the Human Embryo. 523

being no sharp line of demarcation between the two. The indis-
tinctness of the cell boundaries makesit very difficult to determine
definitely the number of cell Jayers in the epithelium. Because of
this, different observers have not agreed upon the number of cell
layers present in the early stages. It can, however, be said with
reasonable certainty that the epithelium at 7.5 mm. is strati-
fied and in the upper and lower thirds of the cesophagus shows
three or four rows of nuclei, in the middle third, two or three rows.
The basal cells are the more columnar and are characterized by
having their nuclei in that end of the cell which is toward the
lumen. Surrounding this central tube of epithelium is mesen-
chyma, which although undifferentiated is slightly condensed.

In an embryo of 16 mm. the oesophagus has increased in length
and breadth. The upper end is both larger and more irregular
in outline. The epithelium contains three or four rows of nuclei
and lies on a very distinct basement membrane. Here again, the
basal layer of celJs is the more columnar and its rounded nuclei
lie in the upper ends of the cells, that is, away from the basement
membrane. In its lower portion, the cesophagus is smaller and
ovalin cross section. Throughout its entire extent the cesophageal
epithelium is surrounded by mesenchyma which is limited exter-
nally by a narrow ring of deeply staining myoblasts, elongated
in form, and so placed as to form the circular layer of the muscu-
aris.

Vacuoles. The next stage examined was an embryo of 19 mm.
(series 819). In this only about the lower half of the cesophagus
was available for study. The epithelial tube is now irregular
in outline, and presents within its walls numerous small cavities
or vacuoles. Some of these are minute, others have a diameter
0.051 mm., which is greater than the diameter of the lumen itself,
making it difficult to determine from cross sections which is lumen
and which vacuole. In some places the vacuoles open directly
into the lumen, in other places they are separated from it by a
partition of epithelium. Fig. la is of a model of the cesophageal
epithelium at this stage and shows its vacuolated structure.
The portion of the oesophagus modelled was selected at a place
just below the bifurcation of the trachea. In a second embryo

rm

524 Franklin P. Johnson.

of 19 mm. (series 829) similar vacuoles are found, but are less
numerous than in the above embryo of the same length. They
are also more numerous in the lower portion of the cesophagus
than in the upper.

At 22.8 mm. (series 871) the diameter of the cesophagus is
nearly double that at 19 mm. The vacuoles are more numerous
and, as shown in fig. 1b, communicate with one another and with
the main lumen to a greater extent. In the upper part of the figure
the cesophagus appears to have a double lumen. This appear-
ance is due to the fusion of a number of vacuoles. The vacuoles
are about the same size as those found in the preceding embryo
but the lumen has increased in diameter. Although vacuoles
also occur in the dorsal and ventral walls of the cesophagus, they
are less numerous there than in the lateral walls. The few vacu-
oles found in the dorsal and ventral walls do not show in the figures;
they are, however, of the same general character as those described.

In other embryos of 22 mm., 22.8 mm. (series 737) and 23 mm.
the vacuoles are found to be less numerous in the upper half of
the cesophagus than in the lower half. At 29 mm. and 30 mm.
they have become very few; at 32 mm. only slight traces of them
are found; at 37 mm. they have entirely disappeared. ‘

Vacuoles of the nature described were noted by Schultz in
1897. Kreuter (05) believed that they cause a temporary occlu-
sion of the cesophagus, but Forssner (07) showed by means of
models that the main lumen is not obliterated. Schridde (’08)
failed to find an occlusion at any stage and denied the presence
of vacuoles but described epithelial bridges which arise by epithe-
lial proliferation in circumscribed places. From a study of these
structures by means of wax reconstructions, the writer is able to
confirm the results of Forssner inasmuch as the lumen is not
entirely occluded and vacuoles are present.

The exact cause and significance of the vacuoles are difficult
to determine. Similar vacuoles are found in the cesophagus of pig,
rat and rabbit embryos. They have been reported by Forssner
in the hedgehog, where in a certain stage they lead to a complete
occlusion. Vacuoles similar to those of the oesophagus are found
in the walls of the stomach and the duodenum of human embryos,

Mucous Membrane in the Human Embryo. 525

Kreuter states that there is no reason to believe that the vacuoles
are areas of degeneration for he was nowhere able to demon-
strate evidences of a degeneration of cells followed by resorp-
tion. There can be seen, nevertheless, a few nuclei lying within
certain vacuoles which stain less intensely and are more indistinct
than the nuclei of the epithelium. It does not appear, however,
that these occasional degenerating nuclei are responsible for the
production of the vacuoles. It seems obvious from the arrange-
ment and crowded condition of the surrounding nuclei that the
vacuoles increase in size. As they become larger the epithelium
between the vacuoles and the lumen, and between adjacent vac-
uoles becomes reduced to a thin partition. This partition
eventually breaks through and makes the cavity of the vacuole
and the lumen continuous.

Measurements of the thickness of the epithelial wall of the
cesophagus are interesting in connection with the vacuoles. Ata
time when vacuoles begin to form, the epithelial wall of the
cesophagus is composed of apparently three or four layers of low
columnar cells. While vacuoles are present the number of cell
layers varies from one to four. By a breaking through of the
vacuoles into the lumen and a disappearance (or migration) of
the more superficial cells, the epithelium is reduced in thickness.
At 19 mm. and 22.8 mm. the epithelium averages .051 mm. while
at 30 mm. only .030 mm. in thickness. This loss in thickness is
soon compensated by a growth in the height of the individual
cells, for at 42 mm. there are only two layers of cells, the aver-
age thickness of the epithelium being .043 mm. Two results of
vacuolation are, therefore, a destruction of the more superficial
layers of cells, and an increase in the size of the lumen of the
cesophagus. This increase in the size of the lumen is propor-
tionately great in the early stages but less in the older ones.

Folds. At an early stage the epithelium of the cesophagus be-
comes thrown into large longitudinal folds which are usually
four in number. Later smaller folds develop at the bases of, or in
between, the larger ones. These folds give to the cesophagus in
cross section the appearance of a Maltese cross as was first noted
by Koelliker (’61). Certain of the folds make their appearance

526 Franklin P. Johnson.

before others and show different stages of development in different
regions of the same oesophagus. The exact time and order of the
appearance of these structures was studied in a number of embryos
with the following results.

As stated before, the epithelial tube of the cesophagus at 7.5
mm. is cylindrical in shape, very small at its mid-region, but
gradually becoming larger when followed either up or down.
In embryos of 10 mm. and 16 mm. the upper portion of the
cesophagus is flattened ventro-dorsally, the middleregion is circular
but now more expanded, and the lower portion is flattened later-
ally. In the upper part of the cesophagus there is a slight infold-
of the ventral wall of the epithelial tube.

In the upper part of the cesophagus at 22.8 mm. (series 871)
there is a ventral infolding of the epithelium. This is the direct
downward continuation of that fold of the pharynx which gives
to the latter, when seen in cross section, its characteristic cres-
centic shape. Following downward this fold is soon lost and the
cesophagus becomes rounded. Another fold appears in the mid-
region of cesophagus, this being an infolding from the dorsal side.
Still further caudally, as the stomach is neared, a third infolding
is found, this being left lateral in position. This lateral fold in
an older embryo is seen to be directly continuous with the dorsal
fold above. The folds found in this embryo are, therefore, really
two in number, a ventral one which appears in the upper third.
of the cesophagus, and a dorsal one which is seen in the middle
third and again as a lateral fold in the lower third. Since the.
study of the order of the appearance of these folds was, made.
by wax reconstructions in a series of embryos, it was necessary. to
secure for modelling approximately the same level of the ceesopha-
gus in each specimen. The region selected was that immediately
below the bifurcation of the trachea. ‘4

The cesophagus at 37 mm. in the region of the bifurcation of
the trachea (fig. 2) is somewhat crescentic in cross section. Dor-,
sally, facing the dorsal aorta, there is a deep fold which continues,
upward and downward and passes directly onto the stomach,
In the lower third, however, it deepens and changes its position.
by moving through an arc of about 90° to the right. At 42 mm,

Mucous Membrane in the Human Embryo. 527

in the same region as the above (fig. 3) there are two distinct
folds, a dorsal and a ventral. These folds, which I have designated
as the first pair of primary folds, are continued downwards onto
the stomach, but before reaching it are twisted to the right and
occupy positions on the left and right sides of the cesophagus
respectively. In the figure a second pair of primary folds can be
seen, which, although indistinct at this level, are well marked below.
In the region modelled and below it, the fold on the left side is
the more marked, but above the right fold alone is seen, and the
ventral and dorsal folds approach one another on the left side,
making a three sided figure. At the extreme upper end of the
oesophagus these folds become irregular and are broken up by the
formation of other smaller folds. All the folds shown in fieeD
are twisted as the stomach is neared, each occupying a position
of about 90° to the right of its original position.

At 55 mm., both pairs of primary folds are well developed
(fig. 4) and give to the lumen of the cesophagus in cross section
the appearance of a Greek cross. These folds are also seen in em-
bryos of 30 mm. and 42 mm. in the lower parts of the oesophagus.
As seen in fig. 5, at 134 mm., these primary folds are augmented
by secondary folds which develop at the bases of the former.
Three such folds are seen in fig. 4, one of which is quite large.
The same folds are found in embryos of 187 mm., 240 mm., and
at birth.

By way of summary concerning the development of folds of
the mucous membrane of the cesophagus, it may be said that the
dorsal one is the first to develop completely. This is followed
closely by the ventral fold. The second pair of primary. folds
develop from below upwards and have reached to the bifurcation
of the trachea in,an embryo of 42 mm. The secondary folds
are less constant structures. They begin as thickenings of the epi-
thelium between the bases of adjacent primary folds. In the upper
part of the cesophagus, especially in the older stages, folds are
very irregular. The reader is referred to Table 1, page.532. for
measurements of folds of the cesophagus in different stages of
development. |

The fact that the epithelial folds of the cesophagus rotate

528 Franklin P. Johnson.

through an angle of about 90 degrees and always in the direction
of the hands of a clock, seems to have some significance. Just
what the cause of this rotation may be is debatable. Kreuter
discards the idea that the lower part of the cesophagus may share
in the rotation of the stomach. However, this may be its cause.
This view is supported by the arrangement of the branches of
the vagus nerves. These branches are also twisted at the same
level as the cesophageal folds. It would, therefore, not seem
improbable that the lower part of the oesophagus had been
twisted along with the nerves by the stomach. Since the twisting
of the stomach takes place before the appearance of the folds, it
would necessarily have to be assumed that the lines along which
the first eesophageal folds develop are marked out before the actual
appearance of the folds themselves.

Ciliated cells. The presence of ciliated cells in the cesophagus
was noted in 1876 by Neumann, who observed them in em-
bryos of from 18 to 32 weeks. He described the epithelium
as stratified, the superficial layer being made up of patches of
both ciliated columnar and squamous cells. Schaffer (04), con-
firmed by Jahrmaerker (’06) and Schridde (07), states that the
ciliated cells may extend through the thickness of the epithelium
down to the basement membrane. Jahrmaerker has recorded the
presence of ciliated cells in the cesophagus of an embryo at 44
mm. In an embryo of 42 mm. no ciliated cells could be found,
but the preservation of this embryo was poor and the staining
unfavorable for the recognition of cilia. In a specimen of 55 mm.
ciliated cells are abundant. Here the epithelium of the cesophagus
is composed of from two to four layers of cells with distinct cell
boundaries. The basal cells are columnar and have their oval
nuclei away from the basement membrane. In those places where
there are not more than two layers of cells, the superficial cells
are columnar, very granular, and ciliated. Where there are three
_or four layers of cells, only the lower layer is columnar, the upper
layers being composed of polygonal or flat cells.

In order to study the distribution and number of the ciliated
cells, wax reconstructions were made, the areas of ciliated cells
being carefully painted on each wax section before piling. Fig.

Mucous Membrane in the Human Embryo. 529

6, from an embryo of 55 mm., shows one half of a model viewed
from its inner surface. The stippled areas represent those covered
with cilia. As can be seen, the larger areas of ciliated cells are
on the folds and these areas include but few islands of polygonal
cells. Between the folds there is a preponderance of squamous
cells, which are interrupted by a few small ciliated patches. In
the upper part of the cesophagus the ciliated areas are slightly
larger than in the region modelled. A similar reconstruction was
made of the epithelium of the cesophagus at 99 mm. (fig. 7). It
shows that the amount of ciliated surface has actually and rela-
tively increased and there are now more islands of polygonal cells
on the folds.

In embryos of 120 mm., 134 mm., and 187 mm., ciliated cells
in the cesophagus are observed to be distributed in about the same
proportions as at 99 mm. At eight months the upper part of the —
cesophagus shows only a few small patches of ciliated cells. In
a specimen at birth the areas are abundant but proportionately
smaller than in the earlier stages, and the islands are now more
widely separated. The islands consist of distorted columnar cil-
iated cells, the distortion probably being due to the pressure of
the rapidly increasing polygonal cells, which are seen crowded
closely against the islands and in places partly undermining them.
In a child of two weeks (7 months premature birth) no ciliated
cells can be found in any part of the cesophagus. Jahrmaerker
and Schridde were unable to find ciliated cells at birth.

Glands. In regard to the glands of the cesophagus, there are
two distinct types to be considered; the true or deep cesophageal
glands, and the cardiac or superficial cesophageal glands. The
former are found in the adult all along the cesophagus, with the
exception of the lower 2 to 4 mm. of its length; the latter in the
lower 2 to 4 mm. and also (70 per cent of cases examined by Schaf-
fer) in that portion of the upper part of the cesophagus which lies
between the level of the cricoid cartilage and the 11th tracheal
ring.

In an embryo of 78 mm. (incomplete series 723 and 724) small
patches or islands of glandular epithelium are found in both the
upper and lower ends of the cesophagus. Schaffer (’04) described

530 Franklin P. Johnson.

similar groups of cells in an embryo of four months, at the level
of the third or fourth tracheal cartilage, and Schridde (’07) found
a group inanembryo of 105-110 mm. (16 to 17 weeks) at the level
of the ericoid cartilage. Areas of glandular cells are abundant in
the lower part of the oesophagus at 120 mm. Some groups are
arranged as islands, others are evaginated so as to form small
pockets. From the bottoms of these pockets budding glands are
seen. In the lower part of the cesophagus at 240 mm. the glandu-
lar cells are more conspicuous. On the sides of the large folds,
but chiefly at their bases, patches of secreting cells are seen form-
ing small isolated islands, pockets, and grooves. The glandular
epithelium is high columnar and is everywhere surrounded by
stratified squamous and ciliated cells. The pockets form the am-
pulle of the cardiac glands, from which numerous small buds
have grown out. A model of such a gland, found at the bottom
of a fold at the lower end of the cesophagus, is shown in fig. 8.
The surface which is ruled in the figure indicates the simple glandu-
lar epithelium; the unruled surface, the stratified squamous epi-
thelium.

The presence of the deep cesophageal glands was first noted in
an embryo of 240 mm...Their beginnings are seen as small out-

growths of the stratified epithelium. These buds are found at,

the bases of the primary folds and upon the outer surface of the

secondary folds. They are lined by a stratified squamous epithe;,
lium similar to that lining the cesophagus., In a specimen from.

the upper part of the cesophagus at.8 months, a.gland was, found

which extended through the muscularis, mucosae into. the; jsub-,
mucosa, Its. walls were composed. of, one, and.,two. layers of.

cuboidal. cells, but nowhere Vhroushout its entire extent could
secreting cells be seen,

At birth the cesophageal glands (fig. 9) Re grown Cath aoe |

the muscularis mucose and lie in the inner part,.of the sub-
mucosa. The mouths of the glands.are Jined with a stratified

:

squamous epithelium of three or four layers of cells which. is.

directly continuous with the oesophageal epithelium.,.; Gradually

the stratified squamous epithelium passes, into the doublelayer (an,
some places single), of low cuboidal cells which lines the excretory,

Mucous Membrane in the Human Embryo. 531

duct. The excretory duct of the gland passes through the thick
muscularis muscosz at nearly right angles to the surface epithe-
lium. On reaching the outer border of the muscularis mucosae,
it bends to either side. Instead of pointing toward the stomach,
as has been described in the adult cesophagus (Goetsch), glands
may extend either upward or downward. The distal termina-
tions of most of the glands are expanded and lined by a simple
glandular epithelium of low columnar cells. In the gland A
(fig. 9) the end piece shows a number of bud-like processes which
are the beginnings of branches. The end piece of gland B is a
simple curved expanded tube, the expanded portion being for
the most part glandular. In the gland C there is no end piece at
all, the whole gland being lined by two layers of non-secreting
low cuboidal cells. In a study of the cesophageal glands of the
pig, Flint has described a similar early stage, in which the glands
contain no secreting cells. However, glands of this type are
rather rare in the human embryo at birth, there being a prepon-
derance of those with secreting cells and beginning branches.
In summarizing the development of glands of the cesophagus,
it may be said that the first evidence of the cardiac glands is the
appearance of small patches of glandular cells in the surface
epithelium. These patches occur at both the upper and lower
ends of the cesophagus and are seen as early as the third month.
Later, by evagination, small pockets are formed from these patches.
This.is followed by a subsequent budding from the outer; surface.
Thus are formed,a number of branches which open into a single
pocket. or ampulla... Cardiac glands never extend. through the
muscularis, mucosae... The deep cesophageal glands, first seen, in
an embryo of 240 mm., are outgrowths of the stratified squamous
epithelium, which after piercing the muscularis mucosae, bend
co either side, and lie-in the submucosa close to the muscularis
mucosae.. The,seereting cells do not appear until after the gland
is well formed, and.are found only near its distal termination.
Here at, birth a number of budding processes, lined with a secre-
tory epithelium, are seen arising from the end pieces. Thus the
cardiac glands and the cesophageal glands differ in that, in. case
of the cardiac glands the secretory epithelium appears before the

az Franklin P. Johnson.
gland formation begins, while in the cesophageal glands it does

not appear until after the excretory ducts are formed.

TABLE 1

OESOPHAGEAL EprtHEettum (Mip Rearon)

THICKNESS! OF DIAMETER OF

EMBRYO SURFACE EPITHELIUM | EPITHELIAL PRIMARY FOLDS SECONDARY FOLDS
TUBE
= path oes In mm.” In mm. Number Holeht. Number Holes
7.5 | 3-4 (?%) .020 .043 0
16.2 3-40) 1) 2048 148 0
19. 1—-4* .051* . 163 0
22.8 1-4* lee e0ol= 223 1 .010
30 2-3 | .030 306 2 .056
42 2-3 048 .398 3-(4) .089
55 2-4 .051 ol a 094 1 012
99 3-4 051 1.01 + .3l a 081
120 4-5 048 .90 + .29 + 054
187 4-5 051 1.63 4 AT 3 091
240 1.81 4 58 a 127
Birth 5-6 | .056 2.26 4 .68 3-4 .163

1 On tops of folds when present.
2 Average of a number of measurements.
* Vacuolated stages.

STOMACH

For this portion of the present paper only the fundus region
of the stomach has been studied, the development of the gas-
tric pits (foveolae gastricae) and the gastric or peptic glands
(glandulae gastricae propriae) being given chief attention. Koelli-
ker (’61 and ’79) regarded the gastric pits and glands as hollow
epithelial processes which grow downward into the mesenchyma.
Laskowsky (’68) confirmed by Schenk (’74) and Brand (77),
described glands as epithelial in origin, but believed that an up-
ward growth of the underlying mesenchyma was the active factor
in their production. Sewall (’79), working on sheep embryos,
concluded that the first chief and parietal cells were epithelial
in origin, but were later replaced by others of mesenchymal origin.

Toldt (’81) in a noteworthy study of the histogenesis of the gas-

Mucous Membrane in the Human Embryo. 533

tric mucosa in eat, pig, and human embryos, concluded that the
glands are formed exclusively by downgrowths from the epithe-
lium. Minot (’02) likewise describes an epithelial origin for the
glands. Later, Strecker (’08) returned to the old idea of a mesen-
cymal origin for the gland cells. The results of the present work
are largely in accord with those of Toldt. However the present
study deals mainly with the form of the mucous membrane and
is based upon a series of wax reconstructions, while Toldt was con-
cerned chiefly with detailed histogenesis.

Early Stages. In an embryo of 7.5 mm. the stomach is a wel!
marked, spindle-shaped swelling, about .132 mm. in transverse
diameter. It is placed with its long axis parallel to the long axis
of the body of the embryo. Its epithelium is about .046 mm. in
thickness and at the extremities of the stomach is composed of
two or three layers of low columnar cells, at the mid region of
higher columnar cells. As in the oesophagus, the basal layer of
cells is the tallest, and the nuclei are placed at the upper ends of
the cells. In an embryo of 8 mm., described by Jahrmeerker,
the nuclei of this strata were found in the basal ends of the cells,
and this he pointed out as a distinction between cesophagus and
stomach at this stage. The epithelium of the stomach is sur-
rounded by loose mesenchyma which is limited, except at its
mesenteric attachments, by the mesothelial lining of the coelom.

In an embryo of 10 mm. the stomach is more pyriform in shape
and is now undergoing rotation to the right on its long axis.
The lumen has greatly increased in diameter but the epithelial
wall is of about the same thickness, measuring .048 mm. The
number of strata of cells varies from two to three. Both the
basal and the superficial layers of cells are distinctly columnar
the latter showing distinct top plates. The nuclei of the basal
layer are again in the upper ends of the cells, those of the super-
ficial layer at the lower ends. The surrounding mesenchyma is
slightly condensed, and a basement membrane is distinct in some
places.

Gastric Pits. At 16mm. the stomach is lined by an epithelium
of high columnar cells of two or three layers, the entire thickness
of the wall measuring .054 mm. The nuclei of the basal layer of

534 Franklin P. Johnson.

cells are again in their upper ends. The epithelium on the side
toward the bursa omentalis shows several irregularities. These are
vacuoles and pit-like depressions. The vacuoles are similar to
those found in the cesophagus, but are smaller and far less num-
erous. The pit-like depressions are surrounded on their sides by
the columnar cells of the epithelium. The nuclei of the cells
which line the pits are rounded and are quite regularly placed
around the lumen, so that the whole structure has somewhat the
appearance of a taste bud. The pits measure about .009 mm. in
width and about .028 mm. indepth. There are no corresponding
outbulgings of the basement membrane made by these pits on
the mesenchymal surface of the epithelium.

In an embryo of 19 mm. the epithelium of the stomach is of
the same type as in the preceding embryo, having two or three
layers of cells and a thickness of .048 mm. It contains a few small
vacuoles. The pits are more numerous than before and some are
elongated so that now they form short grooves. These are most
numerous in the lower end of the stomach. Again there is nothing
on the mesenchymal side of the epithelium to correspond to these
depressions. In this embryo the beginning of the tunica muscu-
laris is first seen.

The epithelium of the stomach at 22.8 mm. measures on the
average .055 mm. in thickness. A few vacuoles, similar to those
found in the cesophagus of this embryo, were observed. They
were, however, much smaller than those in the cesophagus. The
pits are more numerous and distinct than in the preceding embryo.
Again some are rounded and some groove: like in form, but still
they produce no swellings on the under side of the epithelium.
At 42 mm. the pits are abundant. The epithelium measures .055
mm. in thickness and shows for the first time slight bulgings into
the mesenchyma.

In an embryo of 55 mm. the epithelium of the stomach shows
well defined pits and furrows on its inner surface. A surface
view of a portion of the epithelium of the fundus of the stomach
at this stage is shown in fig. 10. The cylindrical pits are irregularly
scattered about, and the grooves are placed parallel to the long
axis of the stomach. The epithelium, which was previously

OF

Mucous Membrane in the Human Embryo. Sao

stratified, is now becoming one-layered. This change is brought
about by a readjustment of the cells, caused—first, by the devel-
opment of the pits; second, by the development of somewhat
similar pits or furrows on the outer surface of the epithelium.
These latter structures are found alternating with the pits of the
inner surface and are in different stages of development. Some
appear as mere slits in the epithelium, others are already broader
than many of the gastric pits themselves. Whether these pits
are formed by an ingrowth of the mesenchyma, or whether they
are formed independently of mesenchyma by a readjustment of
the epithelial cells, is difficult to determine. In the specimens
studied, the mesenchyma had shrunken away from the epithe-
lium, but from their extreme depth and narrowness it is-hardly
probable that the mesenchyma could have extended into the nar-
rowest of these furrows. However formed, these furrows cause
the epithelium to be transformed into a simple epithelium. The
cells of the surface epithelium and those about the pits give off
slender basal processes. These cells have been described by
Baginsky (82) at 7 months, and by Fischl (91) at birth. The
cells have distinct cell boundaries, rounded nuclei, are granular,
and show distinct top plates.

At 91 mm. the grooves on the mesenchymal side of the epithe-
lium are well developed. In a model of the epithelium at this
stage (not figured) these mesenchymal grooves have about the
same appearance as the gastric pits, so that one can hardly deter-
mine which is the internal and which the external surface. The
epithelium is composed of a single layer of cells. The cells are now
higher, with more elongated nuclei, and have long basal processes.
Some of the cells are clearer than others and seem to be filled
with mucous secretion.

At 120 mm. the gastric pits have increased in number and in
size, and are slightly more separated from one another than before.
The growth of the epithelium has taken place both by an enlarge-
ment of the pits already formed and by the addition of other
pits. A wax reconstruction of the epithelium of the fundus of
the stomach at this stage (fig. 11) compared with fig. 10 (55 mm.)
will give a clearer idea of the growth of the pits. The arrangement

536 Franklin P. Johnson.

of the parallel grooves is beginning to be lost, and the whole
surface of the mucous membrane appears to be cut up by the
network of anastomosing pits. The areas marked off by this
network are slight elevations (much higher in the pyloric region),
and have been considered by Brand as villi. Those of the fundus,
however, are much broader and more irregular at their bases
than the intestinal villi. Moreover, they have an entirely differ-
ent developmentalhistory. It isdoubtful, therefore, whether these
structures of the fundus can be considered as villi. The tunica
propria, composed of developing reticular tissue, extends up
between the pits to the surface epithelium. The surface epithe-
lium iscomposed of a single layerof columnar cells, which are filled
with mucous and contain at their lower ends oval or irregular
elongated nuclei. The basal processes are not so elongated as
in the preceding stage. At the bottom of the pits the cells are
cuboidal.

Gastric Glands. A view of the under surface of the epithelium
at 120 mm. (fig. 12), shows the beginnings of the gastric glands.
They appear as knob-like outgrowths from the bottoms of the
gastric pits. As many as three glands are sometimes seen arising
from a pit at the same level. The small buds are for the most
part as yet simple glands, although several were seen which had
begun to branch. They are of different lengths, the longest
being about .048 mm.,and their diameters at the necks measure
about .036 mm. The glands show small but well defined lumina
and are lined by asimple cuboidal epithelium. The parietal (de-
lomorphous) and the chief (adelomorphous) are easily distinguish-
able. The parietal cells stain more intensely with eosin than do
the chief cells, and their protoplasm is slightly more granular.

Cross sections through the middle of the stomach at 187 mm.
show the mucous membrane to be thrown into a number of large
folds. The gastric pits are of about the same character as de-
scribed for the 120 mm. embryo but they have increased in size.
Some of the largest now measure about .089 mm. in depth. The
surface epithelium is higher than in the preceding embryo and is
composed of cylindrical cells which are mucous in character. The
cells contain oval nuclei which lie in the lower halves of the cells

Mucous Membrane in the Human Embryo. 537

but not against their bases. At the bottoms of the pits the glands
are seen. These are now longer and more branched, the longest
measuring about .12 mm. The diameters at the neck vary some-
what, but most are about .0838mm. The diameters of the branches
are smaller than the main stem of the gland. The glands are lined
by a simple epithelium of irregular or polyhedral cells. Both
the chief and the parietal cells are abundant. The bases of the
glands lie close to the muscularis mucosae, which at this stage is
well developed and consists of several distinct strands of smooth
muscle fibres.

The changes that take place in the stomach epithelium from
the 187 mm. stage are merely those in the increase in numbers
and growth of the structures already formed. At 240 mm. the
gastric pits measure from .102 to .158. mm., and the glands about
85 mm. to .186 mm. in length. From this it can be seen that
the growth of the glands has been more rapid than that of the
pits, some of the glands now being longer than some pits. Ina
model of the epithelium of the fundus of the stomach at 240 mm.
(not figured), gastric pits are more irregular in form but are
broader across their tops than those of the 120 mm. stage. They
are, moreover, further apart than before. Fig. 13 shows gastric
glands modelled under higher magnification. Two types of glands
are seen, the smaller simple ones and the larger branched. In the
compound glands the branching is not confined to any particular
level, but branches are given off at the neck or any level below it.
The cells lining the pits are like those found in the preceding
stages, the chief and parietal cells being more numerous than
before.

At birth the structure of the mucosa of the stomach is very sim-
ilar to that found in the adult. The gastric pits are only slightly
deeper than at 240 mm. The surface epithelium is composed of
high cylindrical mucous cells with oval nuclei and basal processes.
At the bottoms of the pits the cells are lower but are still distinct.
As seen in fig. 14 the glands are longer and more branched than
before. They have small lumina which are lined with chief and
parietal cells.

THE AMERICAN JOURNAL OF ANATOMY, VOL. LO, No. 4.

538 Franklin P. Johnson.

The growth of the mucous membrane of the stomach is per-
haps best shown in table 2. From the earliest stage up to 42 mm.
there is but a slight increase in the thickness of the epithelial
wall. At 16 mm. the pits begin to form, and rapidly increase in
number and in size. At 55 mm., due to the presence of pits on
the inner surface and alternating grooves on the outer (mesen-
chymal) surface, as described above, the epithelium becomes
reduced to a single layer of low cells. When measured as in the
preceding stages, the epithelium averages .057 mm. in thickness.
When determined as in the following stages, 7.e., the epithelium
which surrounds the individual pit, it measures .012 mm. Thus
by the readjustment of the cells which is completed at this stage,
the epithelium is reduced in thickness from .057 mm. to .012 mm.
At 91 mm. the surface epithelium is much thicker because of
the extremely long basal processes of its cells. At 120 mm.
the basal processes are shorter, but from this stage up to birth
there is a gradual increase in the thickness of the epithelium.
The gastric pits rapidly increase in depth but there is a more
gradual and variable increase in their width and the distance
between adjacent pits. The glands also increase rapidly in length,
but slowly in diameter. |

Folds. Folds of the mucous membrane of the stomach are more
variable in number than in the cesophagus. In an embryo of 10
mm. the epithelial wall of the stomach is perfectly smooth through-
out. At 16 mm. there are two or three slight folds on the side of
the greater curvature, and indications of about the same number
of the side of the lesser curvature. At 19 mm. there are 2-3
large longitudinal folds on the side of the greater curvature, while
the opposite wall is rounded, with no indications of folds. At
22.8 mm. there are 7-10 folds, including both walls; at 42 mm.
10-12; at 55 mm. about the same number but they are not as
high. At 120 mm. and 187 mm. large folds are found, while at
240 mm. and at birth the folds are very much higher. In the last
four cases, the number of the folds was not obtained. J’rom these
and other observations it would seem that the folds are rather
inconstant structures, their number and size depending on the
state of contraction of the muscular walls. Brand (77) found

Mucous Membrane in the Human Embryo. 539

12-15 longitudinal folds of the stomach mucosa in an embryo of
two months. Koelliker (’69) found in an embryo of four months,
3-4 folds; in another embryo of four months, 11-12; at three
months, 5-6. He therefore regarded their number as variable.
Toldt (81), in a study of cat embryos, considered the folds of
the stomach as unimportant, being formed by the contraction of
the musculature.

TABLE 2

Sromacu EprtHenium (Funpvus)

THICKNESS OF
EMBRYO SURFACE GASTRIC PITS GLANDS FOLDS
EPITHELIUM
ree , 2 Diam- |
een, | Eageell [tm mm. Depth! | "ope) in apartin | Lenath' eter (8t ayuouter Hele
. 7 mm. |
Te We hs 1046
Hoe" 3r 51) 048 |
16 2-3 054 028 O09. | 2f 5-6 | .042
19 223 |=. :048-|- 5-031 009 t 2-3 | .067
D218. 9-31 \o55. |. 1048 O12 t V=109\18
42 1-3 | .055| .041 |.012-.017|.036-. 048 10-12 | .25
55 1 057 |
or
012° .048 —.007-. 012, .024~. 036) | 10-12 | .11
91 1 .048| .072 |.012+.030!.048-.084
120 1 .029 .084.019-.036|.054-.108| 048 | .036 | 36
187 1 .036 | .089 |.019-.038.068-.170| .120 | .038 | 88
240 1 088 | 106 |.919-.038|.102-.238 .136 | .040 | ee 88
Birth 1 048 108.02 ~.048.102-.176| 255 .048 70

' Average of a number of measurements.
t Irregularly placed.

SMALL INTESTINE

Vacuoles and Diverticula. In the following section of the present
paper the development of the intestinal villi and glands has been
given especial attention. The vacuoles of the solid stage of the
duodenum and intestinal diverticula are briefly described, par-
ticularly, because of a probable relation which they bear to the
vacuoles found in the cesophagus and the stomach, and because
they are closely associated with the development of villi.

540 Franklin P. Johnson.

The small intestine in the early stages of development of the
human embryo is lined with an epithelium which is similar to
that of the cesophagus and the stomach. This epithelium is
composed of from two to four layers of cells surrounding the cen-
tral lumen, which in the earlier stages is pervious throughout.
The portion of the small intestine which is to become the duode-
num begins to grow more rapidly than the lower portion, and in
any single embryo may be seen to have attained a higher degree
of development. In an embryo of 7.5 mm. the diameter of the
epithelial tube of the duodenum is about twice that of the ileum,
and its epithelial wall is much thicker. In the older stages,
however, this advance in growth of the duodenum is not re-
tained.

In an embryo of 10 mm. the posterior wall of the duodenum is
thicker than the anterior and contains for a short distance both
above and below the entrance of the ductus choledochus, numerous
small vacuoles. Beyond this opening the lumen of the duodenum
becomes extremely small, and only a few vacuoles are present.
The remainder of the intestine is smaller than the duodenum at
this stage, and is more regular and round. Its epithelium, which
is composed of three to four layers of cells and is about half the
thickness of the duodenum, does not contain. vacuoles.

Tandler (00) described similar vacuoles in the duodenum of
embryos of from 30 to 60 days. He found that their formation
in some cases led to a complete occlusion of the duodenal lumen,
and believed that failure of the lumen to open again caused the
well-known anomaly of duodenal atresia. Minot (’00) described
a like condition in the large intestine of the chick. It is evident
from his drawings (figs. 3-5) that the lumen of the large intestine
of the chick is more completely occluded than that of the human
duodenum. Forssner (’07) found similar vacuoles and duodenal
occlusions in human embryos of about 20 mm. and also in the
duodenum of rat embryos.

At 16 mm. the epithelium throughout the entire extent of the
duodenum is filled with vacuoles which are larger and more
numerous than in the former stage. Immediately below the
openings of the ducts, the vacuoles are most numerous and the

Mucous Membrane in the Human Embryo. 541

lumen of the duodenum is totally occluded. Below this region
the vacuoles are fewer and the lumen from this point on is open.

A few vacuoles are present in the upper part of the jejunum
at this stage (16 mm.) but these are not as large or as distinct
as those found in the duodenum. The lumen of the jejunum and
ileum, unlike that of the duodenum, is pervious throughout its
entire extent, although in the ileum it is much smaller. There
have developed in the epithelial wall at this stage a number of
outgrowths, which have been described as ‘‘intestinal divertic-
ula.”’ These rounded, bud-like. structures are constricted at
their necks, and extend into the surrounding mesenchyma. They
are found in different stages of development and are variable in
size. The smallest have diameters of .043 mm., the largest of
about .072 mm., which is about 4 that of the small intestine itself.
The older pockets are well marked and are six in number, the
younger ones are not as distinct and a few are doubtful. These
were 11 in number, making a total of 17 pockets in the small
intestine of this embryo.

Keibel (05) noted the presence of similar pockets in embryos
of man, Tarsius, pig, and deer, and considered it strange that
these structures were not mentioned by Voigt (99) and Berry
(02). Lewis and Thyng (08) described diverticula which they
found along the small intestine in human, pig, and rabbit embryos.
In a human embryo of 23 mm. they counted 33 pockets; at 22
mm. (H. E. C. series 851), 48 pockets. Elze (’09) found in various
mammalian embryos, rounded and elongated intestinal pockets
similar to those described by Lewis and Thyng. He observed
that the pockets are always on the wall of the intestine away from
the mesentery, and that those pockets which were elongated and
extended along the intestine fora short distance, invariably pointed
down the intestine (ab-orally). In the human embryos of the
Harvard Collection, so far as they have been examined, the diver-
ticula present the same characteristics. Because of differences
in their arrangement and position, Elze puts the diverticula of the
duodenum in a separate class from those of the jejunum and ileum.

At 19 mm. (series 819) the vacuoles in the duodenum are larger
than those at 16 mm. The lumen is continuous throughout and

542 Franklin P. Johnson.

is larger both in actual measurement and in proportion to the
size of the vacuoles. In the lower part of the duodenum, these
vacuoles are somewhat different from those found in the upper
part and from those found in the cesophagus. Many of them have
corresponding outbulgings of the epithelium on the mesenchymal
surface. Some of these are constricted at their necks and form
small side pockets, but for the most part they do not open into
the main lumen of the duodenum. These ‘‘duodenal pockets,”
if such they may be called, are presumably different from those
referred to by Lewis and Thyng, which give rise to an occasional
accessory pancreas. They differ also from the intestinal pock-
ets in their position on the walls of the duodenum, their constric-
tions are not always so well marked, and their cavities only occa-
sionally open into the main lumen. However, so far as size and
general appearance are concerned, they resemble the pockets of
the jejunum and the ileum. Besides distinct pockets, all grada-
tions between the simple vacuole of the interior and constricted
side pockets are found in the walls of the duodenum. ‘These pock-
ets of the duodenum, therefore, probably represent transition
forms between the vacuoles of the upper part of the digestive
tube and the intestinal pockets of the lower part.

In the jejunum and the ileum at 19mm., there are 41 divertic-
ula. These have the same general character as those of the 16
mm. stage, but some, and especially those in the lower part of
the ileum, are much larger. They are not equally distributed
throughout the whole gut but are more numerous in the ileum.
None are present in the upper part of the jejunum.

In the duodenum of an embryo of 22.8 mm. the vacuoles are
few above the opening of the ductus choledochus. Below this
level, fig. 16, they are numerous and larger in proportion to the
size of the tube than at 19 mm. Moreover, they do not form du-
odenal pockets as in the 19 mm. stage. They cause the whole
lumen to be broken up, and, as shown by modelling, the lumen
in this region is occluded. This stage, therefore, as far as the vac-
uoles are concerned, resembles the condition found at 16 mm.
more than that found at 19mm. In the jejunum and ileum 33
pockets were counted. Again they vary in size, the largest meas-

Mucous Membrane in the Human Embryo. 548

uring .084 mm. Three of these pockets are seen in fig. 17. The
first one (lettered a) is constricted from the intestine on all sides.
The other two (6 and c) are elongated forms, and are not entirely
constricted at their necks, b being a surface view, and ¢ an end
view. The two latter extend along the intestine and their free
ends point ab-orally.

At 24 mm. the vacuoles have disappeared from the upper
half of the duodenum, but they are still present in the lower half.
In the upper half, well formed villi are present; in the lower half,
villi are seen which are fused together at their apices, but spaces
are seen between adjacent villi. These spaces are the above de-
scribed vacuoles. It is evident from this that the disappearance
of the vacuoles is associated with the formation of villi. The
exact part played by the vacuoles in the formation of villi will be
dealt with in connection with the latter structures. In the duo-
denum of an embryo of 30 mm. the vacuoles have entirely dis-
appeared.

Because of the development of villi, the intestinal diverticula
are difficult to count at 24.mm. It is due to these beginning
villi that the pockets disappear. Villi arise around the mouth
of a pocket, and by their subsequent growth and enlargement,
the pocket is gradually lost by being absorbed in their walls.
Some pockets are more elongated than others, and on either side
of these short longitudinal folds are seen. By a growth of these
folds, pockets are likewise obliterated. As the villi develop from
above downward, the pockets disappear in the same order. Con-
sequently, in embryos of 29 mm., 30 mm., 32 mm., and 42 mm.,
the number of intestinal diverticula becomes smaller and smaller.
Those in the ileum attain the highest development. As they in-
crease in size their outer surfaces become flattened and later may
even become concave. At 30 mm., the diameter of one of the
largest pockets measures .108 mm ; at 37 mm. .176 mm.
- The significance and fate of the diverticula have been discussed

by Lewis and Thyng. They conclude that some elongated forms,
which are usually found in the duodenum, are the source of an
occasional accessory pancreas. The more rounded intestinal
pockets usually degenerate, but some may remain to form cysts

544 Franklin P. Johnson.

and nodules. It has been my fortune to find, in the mid region
of an embryo of 134 mm., a large persistent intestinal diverticu-
lum, fig. 28. It resembles in shape the pockets found in the lower
part of the ileum in the 30 mm. and 37 mm. stages. It is connected
to the intestinal epithelium by a constricted neck. At its widest
place it measured .50 mm. in transverse diameter, and at its
neck .21 mm. Extending outward from its somewhat concave
outer wall are a number of intestinal glands which are similar
to those found in other parts of the intestine. Extending into
its cavity from the internal surface of the outer wall are a few villi,
one of which is large and extends through the neck of the pocket
into the lumen of the intestine.

In the duodenum at 134 mm., several epithelial cysts were
found, which are entirely cut off from the epithelium of the duo-
denum. They are spherical in shape, and each has several glands
extending from its external surface. The flattened cells which
line these cysts give evidence of an internal pressure. In what
manner these cysts are formed, I will not venture to say. There
is, however, the probability of their having developed by the
persistence of the early duodenal vacuoles or pockets.

Development of Villi. Koelliker (’61 and ’79) states that at
the end of the second month and in the third, the beginnings of
separate villi appear. Barth (’68) and Brand (’77) confirmed this
view. Voigt (99), working on pig embryos, describes an irregular
breaking up of the heretofore smooth inner surface of the gut wall
by means of depressions and furrows. ‘These furrows increase
in numbers and run together to form a net-work of canals. From
the fields thus marked off arise slight elevations, the first traces
of the villi. Berry (’00), studying pig and human embryos,
writes:

“The summary given by Oppel shows beautifully the compara-
tive anatomy of the villi and their evolution. In vertebrates of
low order, the intestine is smooth, no villi being present. Then
appear longitudinal folds, and then all gradations between folds and
villi, and finally villi. It is interesting to note that in those intes-
tines in which folds of che mucous membrane are present, they are
more numerous and prominent in the upper part than in the lower

Mucous Membrane in the Human Embryo. 545

part of the intestine. As villi are developed they again appear in
the upper part of the intestine first, so stages are found with villi
in the duodenum, and only folds in the ileum. Furthermore,
when villi alone are present, they are more numerous in the duo-
denum than lower down. The most striking result in the follow-
ing out the development of the villi in the human intestine is that
they repeat all the stages found from the standpoint of compara-
tive anatomy.”

He concludes, ‘‘ The villiappear first aslongitudinal folds. These
folds grow larger and then break up into villi.”’

Forssner (’07) in a study of cesophageal and intestinal atresiae,
while not mainly concerned with the development of villi, con-
firms the work of Berry in that the villi of the human small intes-
tine appear first as longitudinal folds.

In the human embryos I have studied, only in the lower part of
the small intestine was I able to find distinct longitudinal folds
preceding the formation of villi. In the upper two thirds of the
small intestine, villi are found developing as knob-like invagina-
tions of the epithelium.

The first evidences of villi found are in an embryo of 19 mm.
Immediately below che pyloric end of the stomach, the duodenal
epithelium shows, besides its vacuoles, other irregularities. These
are thickenings of the epithelium and invaginations of the epithe-
lium into the lumen of the intestine, and represent different
stages in the development of villi. The thickenings vary in size
and height, some being twice as thick as the epithelium of the in-
testine in other places. The invaginations are slight rounded
elevations of epithelium, and have developed from the thickenings
by a pushing in of the mesenchyma, which now forms the cores
of these small villi. That these are villi and not folds is evident
because they can be followed through only four to six 23 » sections
and in breadth at their bases they measure from .10 to .14 mm.
From these measurements the bases of these elevations are seen
to be approximately circular in cross section. Villi are of all
gradations in height, one of the tallest measuring .09 mm. (The
height of a villus has in every case been measured on the mesen-
chymal surface, v.e., the depth of the hole which is filled with mes-

546 Franklin P. Johnson.

enchyma.) Although the villi are somewhat more numerous in
the duodenum, where the diameter of the epithelial tube is large,
they are also foundinthe greater part of the jejunum. Following
down the jejunum, the larger villi become fewer and fewer until
a region is reached which shows only thickenings. Below this
point the lumen of the intestine is smaller and rounded and re-
mains this way until the large intestine is reached.

At 22.8 mm. the villi are larger and more numerous than at
19 mm. They are, as shown in fig. 15, irregularly shaped
processes. Below the openings of the ducts, villi—if it can be
assumed that they are present—are not recognizable from an
inner view Of the gut cavity. This is due to the occlusion of
the duodenal lumen and to the occurrence of the large vacuoles.
However, the outer surface of the epithelium of this region is
marked with pits filled with mesenchyma. These pits are simi-
lar to those forming the cores of distinct villi found in the other
parts of the intestine. It can be inferred from this, therefore,
that the formation of villi is taking place while the vacuoles
are present’in the epithelium, but the individual villi are not
distinguishable during the solid stage of the duodenum.

Just below the vacuolated region of the duodenum, true villi
are found at this stage (22.8 mm.). They extend into the first
loop of the jejunum, but the villi of the duodenum, having devel-
oped first, are the larger. As the jejunum is followed down, the
villi become smaller and smaller, until a level is reached in which
villi are absent. The villi developing in this lower portion of the
small intestine are as young as any in the whole gut. In order to
makeanaccurate study of their development, a wax reconstruction
was carefully made of thisregion. Fig. 18 shows half of the model,
a, looking at the outer surface of the epithelium; 6, at its inner
surface. An examination of the inner surface shows the presence
of a number of rounded, knob-like structures, the beginnings of
the villi. These are only seen in the upper part of the model.
Thickenings of the epithelium are seen in a region below the villi,
but at the extreme end of the tube there are no evidences of villi
at all. By comparing the external surface of the epithelium, a,
with the internal surface, b, it isseen that there are depressions on

Mucous Membrane in the Human Embryo. 547

the outer surface to correspond with the villi. The thickenings
of the epithelium may or may not have corresponding slight
depressions. This model indicates that each villus originates as
a thickening of the epithelium, and that later this thickening is
invaginated.- Still later the epithelium of the invagination becomes
thinner and is reduced to a single layer of cells. Below the
region of thickenings, the lumen of the intestine becomes smaller
and rounded. Inno place could longitudinal folds of the epithe-
lium be found.

The idea that villi formation goes on simultaneously in that
portion of the small intestine (duodenum) which is vacuolated
and in that portion which is not, is further supported by the condi-
tion found in the duodenum at 24 mm. In this embryo, only the
lower portion of the duodenum shows vacuoles. In theupper part
of that region which was previously occupied by vacuoles, villi
are easily recognized. Further down there is a region in which
are seen a number of villi which are fused together at their apices,
that is, end to end. In some places this fusion has the form of
thin membranes extending from villus to villus. The formation
of the villi seems to be taking place by a pulling apart of preformed
villi, and in this process the vacuoles are lost by becoming a part
of the lumen. The process of separation of the villi progresses
from above downward.

In the mid region of the small intestine at 24 mm. (fig. 19),
the villi are seen as separate knob-like elevations of various shapes
and sizes. For the most part they have nearly circular ‘bases,
but some are elongated. Their apices are rounded. The average
of a number of measurement shows the breadth of a villus to
be about .11 mm. The tallest villi found in this region measured
12mm. The epithelium on the tops of the villi is single layered,
although in places it appears pseudo-stratified, and is .020 to
.025mm.in thickness. Between the villi are epithelial thickenings
which measure .064 mm. in thickness and are composed of from
3 to 5 layers of cells. An examination of the whole of the model
(half of which is seen in fig. 19) shows that the villi are arranged
in five longitudinal rows. In the lower part of the intestine the
villi are smaller; still lower down only epithelial thickenings occur.

548 Franklin P. Johnson.

Some of these are more elongated than the thickenings found above
and form slight irregular ridges. These ridges have as yet no
corresponding depressions of the mesenchyma. Below this,
there is a diminution in the size of the intestine, and the ridges—
2—4 in number—are irregular and changeable in position. Before
the large intestine is reached, the ridges are lost and the lumen
of the small intestine assumes a more rounded appearance.

The vacuoles have entirely disappeared from the epithelium
of the duodenum at 30 mm. and the lumen is pervious throughout.
The villi are even more numerous than at 24 mm. and vary in
size, the tallest now measuring about .211 mm. The diameters of
their bases vary within the limits of .089 and .146 mm. The villi
are very irregular in shape, some flattened at their bases, others
rounded. The arrangement of villi in rows is more pronounced
than at 24mm. Additional villi have developed either in between
the villi of the rows already present, or by forming additional
rows. In some places the rows of villi have the appearance of
irregular longitudinal folds, suggesting a fusion of adjacent villi.
This apearance is perhaps due to the rapid development of new
villi between the older ones.

In the jejunum at this stage a similar arrangement of villi is
found, but in its lower part the developing villi are further sepa-
rated.. Following down the ileum, they become smaller and
smaller. Then comes a region in which there are only a very few
villi and thickenings, these being more scattered than before.
Following this, there is a region in which the diameter of the tube
diminishes, and in this portion longitudinal folds are present.
Further down it widens out again and the folds are lost; this is
followed by another diminution. Several of these narrowed places
are found in this part of the small intestine, and between them the
tube regains its former diameter. In the narrowed portions, rather
irregular folds are found, varying from 2 to 4 in number. In the
widened portions, very young villi are present, but no folds.
Fig. 20 is of a wax reconstruction of a portion of the epithelium
at a place where there is a diminution in the diameter of the ileum.
In the widened portion, the beginnings of a few villi are seen;

Mucous Membrane in the Human Embryo. 549

in the narrowed portion, two distinct folds, the tops of which
are irregular in height, are present.

In the jejunum at 37 mm., the villi are similar to those at 30
mm., but are now regular in shape. They appear to be fused to a
greater extent, and there are more longitudinal rows. In shape
the villi vary from small low swellings to rounded or elongated
processes, some of which are .214mm. in height. They are only
a little taller in the duodenum than below. Further down, at
the entrance of a persistent yolk stalk, the intestine becomes wide.
In this widened portion both villi and thickenings are seen. Be-
low this region the diameter of the epithelial tube becomes smaller
and irregular longitudinal folds are found. Still further down
only low ridges are found which have no corresponding depres-
sions for the mesenchyma. These ridges extend as far as the ileo-
eaecal valve. Practically the same conditions are found in an em-
bryo of 42 mm., there being a few folds in the lower part of the
ileum where the lumen is narrow. Where it is wide, villi are found.

In the mid region of the small intestine at 55 mm. (fig. 21),
the villi have greatly increased in size and in number. They are,
for the most part, rounded conical processes but a few are ir-
regular and blunted. A most striking fact is that the villi are all
very near the same size. Since additional villi develop as separate
invaginations, one would expect to always find a greater number of
small villi than large ones. But the small villi have always
been found to be less numerous. It follows, therefore, that their
growth must be more rapid than that of the older villi. Additional
rows of villi have developed between those already present.
At 55 mm. there are about 13 rows in the mid region of the small
intestine. The villi of the new rows are as tall as those of the older
ones, in fact, the later developed rows cannot be distinguished
from the earlier ones. The arrangement of the villi in rows is
best seen in a view of the external wall of the epithelium (fig. 22).
In the extreme lower end of the ileum at this stage (55 mm.)
two or three slight folds are found.

Regarding the further development of villi, there is but little
of interest. They gradually increase in number and in size through-
out stages from 73 mm. to birth (Tables 3 and 4). In the later

550 Franklin P. Johnson.

stages they present a variety of different shapes in different parts
of the intestine. The villi of the duodenum at 78 mm. are cylin-
drical, with rounded ends, averaging .23 mm. in height. In the
upper part of the jejunum they are blunter and measure .20 mm;
in the mid region elongated and slender, .25 mm. in height; while
in the lower part of the ileum they are swollen at their apices and
measure about .28 mm. In the mid region at 134 mm. (fig. 23),
the villi are pointed processes; at 187 mm. and 240 mm., they are
‘more elongated. In the mid region at seven months (premature
birth) irregular villi are found, some of which are foliate in shape
and grooved on their sides. At birth, the villi of the mid region
are likewise very irregular in shape.

In briefly summarizing the development of villi, it may be
said that the general tendency throughout the whole of the small
intestine is for villi to develop as separate invaginations of the
epithelium. Owing, however, to the occurrence of transitory
structures (vacuoles, diverticula, and folds) their development is
manifested differently in different parts of the intestine. In the
duodenum the early growth of the villi is closely associated with
and concealed by the vacuolated condition of the epithelium.
While the vacuoles are present, the apices of the villi of one wall
seem fused with those of the opposite wall. They become distinct
when this fusion breaks down. At the same time the vacuoles
are lost by entering into the formation of the lumen. Through-
out the upper two thirds of the jejuno-ileum, villi are first seen as
separate thickenings of the epithelium. These thickenings become
invaginated, and the thickened plate of epithelium becomes
reduced to a single layer of cells. In the lower third of the small
intestine, longitudinal folds precede the formation of villi. How-
ever, since the folds are present only in those regions of the intes-
tine which are not expanded, they are probably formed because
the outer coats of the intestine have not grown as rapidly as the
epithelium. The irregularities seen along the tops of the folds,
which are of about the same size as the slight swellings in the ex-
panded portions of the gut, are probably developing villi. The
folds may later disappear by a general expansion of the epithelial
tube, or by being absorbed by the rapidly growing villi. I find

Mucous Membrane in the Human Embryo. 551

no evidence that there is a mechanical cutting up of the folds into
villi, such as has been described.

The villi of all parts of the intestine later become arranged in
more or less regular longitudinal rows. Additional villi develop
as separate growths by forming new rows, and also in between
the villi of the older rows. In some places their development
has gone on so rapidly that the villi of a single row are not entirely
~ separate, and give the appearance of short, irregular longitudinal
folds. This appearance is lost in the later stages.

In any single embryo, the larger villi, which are all approxi-
mately the same size, are always found to be more numerous than
the smaller ones. This indicates that the growth of the younger
villi is more rapid than that of the older ones. After they have
attained the size of the older ones, however, they develop at a
corresponding rate.

The growth of the villi in the duodenum and mid region of
the small intestine is shown in Tables 8 and 4. The villi of the duo-
denum, having developed first, are the taller in the earlier stages.
At 55 mm. the villi in the mid region have attained the height
of those in the duodenum and from this stage on are larger. The
bases of the villi remain approximately constant in size but show ,
a slight initial increase and later a diminution in their diameters.
The epithelium on the tops of the villi, which is at first stratified,
then simple columnar, and finally simple cuboidal, is much re-
duced in thickness.

Intestinal Glands. According to Koelliker (61 and ’79) the
intestinal glands (Lieberkiihn’s crypts) arise as hollow out-
growths of the epithelium which push their way into the meso-
derm. Barth (’68) did not accept this view of the origin of the
glands, but believed that they are formed by the spaces left in
between the villi. Brand (77) believed that an upward growth
of the mesenchyma was the active factor in the development of
the glands. Minot (92) and Kollman (’98) agree with Koelliker.
Voigt (99) studied the development of the intestinal glands in
the pig embryo, and likewise concluded that they originate as
hollow outgrowths of the epithelium which are given off between

552 Franklin P. Johnson.

the bases of the villi and extend into the mesenchyma. This
view was later accepted by Hilton (’02).

So far as can be observed from sections, the results of the present
work are in accord with those of Voigt and Hilton. In a wax
reconstruction of the intestinal epithelium at 837 mm. (not figured)
no evidences could be seen of the glands from an external view of
the epithelium. At 55 mm. (fig. 22) the exterior of the epithelium
in the mid region of the small intestine shows small knoblike out-
growths. These I have considered to be the very first appearances
of the intestinal glands. The cells of these knoblike processes are
very granular, distinctly different from the clearer cells on the
tops and sides of the villt. In amongst the granular cells can be
seen a few scattered goblet or beaker cells. The protoplasm of
the goblet cells is clear, their flattened or crescentic nuclei staining
deeply. In the duodenum of this stage (55 mm.), the beginning
glands are somewhat more pronounced. In embryos of 73 mm.,
91 mm., and 120 mm., the glands gradually become more distinct
and longer, and the goblet cells which line both the glands and the
villi become more numerous. The intestinal glands at 134 mm.
(mid region of the small intestine) are shown in fig. 23, and at 240
mm. (duodenum) in fig. 24. Throughout the older stages, the
glands increase in length andin number. They are usually simple
tubular in form, but some are branched. Measurements of the
glands in different stages of development are given in Tables3
and 4.

Duodenal Glands. Regarding duodenal (Brunner’s) glands
Koelliker states that their formation begins in the fourth month
and that they are originally the Lieberkiihn glands. Later they
send branches into the submucosa, which, at the end of the sixth
month, reach to the muscular layer. Barth found that in the
rabbit the duodenal glands sprout off from the Lieberkiihn glands
as a double outgrowth. Brand (’77) observed Brunner’s glands
first in a human embryo of three and one half months.

In the duodenum of an embryo of 55 mm. no traces of the duo-
denal glands could be found, while at 78 mm. (Embryo 142)—
three months—a few could be seen in the upper third of the duo-
denum. They arise from the bottoms of the intestinal glands,

Mucous Membrane in the Human Embryo. 553

as described by Barth, but may or may not be forked at their
origins. They can be distinguished from the intestinal glands by
the clearer protoplasm of their cells, by their branching, and by the
greater distance that they extend into the mesenchyma. <As the
muscularis mucosae has not developed, there is as yet no line of
division between the mucosa and the submucosa.

In the middle third of the duodenum at 91 mm., no duodenal
glands are present. At 120 mm. they are present in the whole
of the duodenum. They aremorenumerousin the upper third than
lower down, and branching has gone on to a greater extent. In
the upper third of the duodenum at 187 mm. the glands are much
larger and the branches of each gland are arranged in groups.
At the base, toward the muscularis, there is a slight condensa-
tion of the mesenchyma, which appears to have been caused by
the downward growth of the gland. Only a few glands are found
in the middle third of the duodenum at this stage (187 mm.).
Fig. 24, from a wax reconstruction, shows the structure of the
duodenal glands at 240 mm. The intestinal glands are the short
tubular ones arising from around the bases of the villi; the duo-
denal glands are larger and more branched. The fewness of the
primary sprouts in the area modelled is striking, there being only
four for the entire number of branches shown, two of which are
quite small. The branches of each gland are arranged in groups,
lettered in the figure A, B, C, and D.

Folds. As stated before, irregular longitudinal folds of the
mucous membrane are found in the ileum in stages ranging
from 24mm. to 55mm. These folds, however, are only transient,
and are in no way related to the folds found in the later stages and
in the adult. They have practically disappeared at 55 mm.

Circular folds, which have been studied from longitudinal sec-
tions, are found in the small intestines of the older embryos.
Whether these folds are formed by the contraction of the muscu-
lature, or whether they are the non-effacable plicae circulares
(valvulae conniventes) could not be positively determined from
cut sections, but from their regularity and constaacy, I have con-
sidered that they are the latter structures. Their presence was
first noted in the mid region of the small intestine at 73 mm., but

THE AMERICAN JOURNAL OF ANATOMY, vor. 10, No. 4,

554 Franklin P. Johnson.

here they are rather indefinite and uncertain. In the upper third
of the duodenum and in the uppermost part of the jejunum at
78 mm. no evidences of the circular folds could be found, but
they are distinct in the mid region of the small intestine. They
are quite regularly placed, averaging .78 mm. distant from each
other. In height they vary from .17 mm. to .23 mm. At the
extreme lower end of the ileum at this stage, no folds are present.
None were found in the duodenum at 120 mm. (embryo 203).
Slight folds are present in the lower end of the ileum at 134 mm.
In a distended portion of the small intestine at 145 mm. (mid
region), circular folds were found which measure .06 mm. in
height, and at an average distance of 1.87 mm. apart. In the
duodenum at 240 mm. two circular folds were found at distances
of 4.7 mm. and 7.1 mm. below the pyloric orifice. In the mid
region of the small intestine at this stage, folds were found which
vary from .25 mm. to .51 mm. in height, and at a distance averag-
ing 1.3 mm. apart.
TABLE 3

DvuoDENAL EPITHELIUM (UPPER THIRD)

ng val
THICKNESS! OF DIAMETER OF)

EMBRYO SURFACE EPITHELIAL | VILLI Yea poten
EPITHELIUM TUBE
Length In Gell |i mm.2| In mm. Height? eye Length‘ vane | Length® | ers
in mm. Layers in mm. bases” in mM. | jn mm. in mm. | in mm.
|
7:5| 23 | 086| 077 | |
16. a “ Mle © | |
19. 24 | .048|. .194 088 | .11
22.8; 12 | .019| .299 1250 NEA oae
24 i! .024 »36 .193 mp a |
30 ? a ee oid gals
37 1 .024 | 58 23 13.
42 1 021": 565 £2) ea
55 1 .019 .95 A260) eal 048 06
78 1 .016 | 1.038 .29 .11 | .088 04 07 | .033
120 1 L015 \ ale 14 ol | es) 09 .06 12 | 031
187 1 O15 | 2h AT | .09 14 05 .22 | 0381
240 1 (O15 |) 3.5 Gtaye o) lll 12 05 .30 .036
|
10n tops of villi when present. at ;
2 Tallest.
3 Average of a number of measurements.
4 Longest.

5 Vertical distance below bottoms of intestinal glands.
* Vacuolated stage.

Or
ey |

Mucous Membrane in the Human Embryo. Di:

TABLE 4

EPITHELIUM OF SMALL ImTEsTINE (Mrp Rearon)

THICKNESS! OF DIAMETER OF

EMBRYO SURFAGH EPITHELIUM EPITHELIAL VILLI INTESTINAL GLANDS
TUBE
; j 2 Diameter®
Length In Cell Taek? Tagen Height? (at bases) Length‘ Diameter?
In mm. Layers in mm. in mm. in mm. in mm.
lee 2-3 .024 .052
16 2 024 101
19 3-4 .052 141
22.8 3-5 052 . 158
24 1 024 135 12 sill
30 ? t 28 18 ll
37 1 024 39 .20 14
42 1 .021 44 .18 as
55 1 020 io .26 males .036 .06
73 1 .019 1.09 100 15 .048 06
91 1 .024 1.28 of 14 .050 .06
120 1 015 1.40 56 ll .060 .06
134 1 015 2) .63 sal - .065 06
187 1 015 Zo .70 alin .088 .06
240, 1 .012 one .70 07 10 04
Birth 1 .012 4.9 .70 sil 12 04
1 On top of villi when present.
2 Tallest.
3 Average of a number of measurements.
4 Longest.
CONCLUSIONS
(E'sophagus

1. The cesophagus is at first a simple epithelial tube, the
walls of which contain three or four rows of nuclei.

2. In embryos of about 20 mm., vacuoles form in the epithe-
lium but the lumen remains pervious throughout. These vacuoles
disappear by breaking into the lumen. This causes the epithelium
to become thinner and the lumen to increase in size. The cause of
the formation of vacuoles has not yet been determined.

3. Longitudinal folds of the mucosa are constant structures
in the cesophagus. In the upper third of the cesophagus the folds
are irregular and variable. In the middle and lower thirds there

556 Franklin P. Johnson.

are four large primary folds. Of these the dorsal and ventral
(left and right respectively in the lower part of the cesophagus)
develop first; the left and right (ventral and dorsal below) develop
soon afterward. Smaller secondary folds, variable in number,
appear later at the bases of the primary folds. In the lower
part of the cesophagus both primary and secondary folds are
twisted through an are of about 90 degrees in the direction of the
hands of a clock. It is probable that this twisting is due to the
early rotation of the stomach.

4. Areas of ciliated cells are found in the epithelium of the
cesophagus in embryos ranging from 55 mm. to birth. There is
both an actual and a relative increase in the amount of surface
covered by ciliated cells in embryos up to 187 mm. At birth
these areas are relatively smaller. Ciliated cells are absent in
the cesophagus of a child of 14 days (seven months premature
birth).

5. Cardiac glands are found in both the upper and lower ends
of the oesophagus. They begin as small areas of glandular cells,
which were first seen in an embryo of 78 mm. Later these areas
evaginate, forming small pockets and grooves. Later, a number
of tubular glands grow out from these pockets.

6. Qisophageal glands were first observed in an embryo of
240 mm. They grow out from the epithelium through the mus-
cularis mucosae and lie in the submucosa. In contradistinction
to the cardiac glands, their glandular epithelium does not develop
until after the excretory ducts are formed. At birth the end pieces
of the glands have begun to branch.

Stomach

1. <A few vacuoles, similar to those of the cesophagus, are found
in corresponding stages in the stomach.

2. At16mm., the heretofore smooth epithelium shows a num-
ber of pit-like depressions, the first appearances of the gastric
pits. These rapidly increase in number and many become elon-
gated to form grooves.

ee ee ee ae ee

Mucous Membrane in the Human Embryo. 5di7

3. At first the basal surface of the epithelium shows no irreg-
ularities due to the gastric pits. Then appear slight swellings
into the mesenchyma. Still later depressions or furrows, alter-
nating with the gastric pits, extend inward into the epithelium
from the mesenchymal surface. This brings about a readjust-
ment of the cells, and causes the heretofore 2 to 3 layered epithe-
lium to become single layered.

4. The groove-like pits anastomose with one another to form
a network. This network marks out irregular areas of the surface
epithelium, which have been described as villi.

5. Growth of the mucous membrane is accompanied by an
increase in the number of pits and by an increase in their size.
The additional pits develop in between those already formed.

6. Glands, first seen at 120 mm., bud out from the bottoms
of the pits. These rapidly increase in number and give off
branches. Parietal cells are first distinct at 120 mm.

7. Large longitudinal folds of the mucous membrane occur in
the stomach, but are variable in position, number, and size.
Their occurrence is probably due to the contraction of the mus-
cularis.

Small intestine

1. Vacuoles occur in the epithelium of the duodenum of em-
bryos ranging from 10 mm. to 24mm. Inembryos of 16mm. and
22.8 mm. the vacuoles were found to lead to a complete occlusion
of the duodenal lumen. In an embryo of 19 mm. some of the vac-
uoles were found to be arranged so as to form small ‘duodenal
pockets.” These resemble the intestinal pockets of the jejunum
and the ileum. It is probable that these duodenal pockets are
transition forms between the vacuoles of the upper part of the di-
gestive tube and the intestinal pockets of the lower part. In the
duodenum at 134 mm. several small epithelial cysts were found.
It is possible that these are vacuoles or duodenal pockets which
have persisted.

2. In the jejunum and ileum intestinal pockets, similar to
those described by Keibel, Lewis and Thyng, and Elze, were found.

558 Franklin P. Johnson.

Most of these disappear by becoming incorporated in the walls
of the developing villi, but some may persist. A large persistent
diverticulum was found in an embryo of 134 mm.

3. The vacuoles of the csophagus, stomach and duodenum,
and the pockets of the duodenum, jejunum, and the ileum are
related structures. They are all the result of a similar process
of growth, which for some undetermined reason, manifests itself
differently in different parts of the digestive tube.

5. In the upper part of the duodenum, in the jejunum,
and in the upper part of the ileum, the villi originate singly as
thickenings of the epithelium which are not preceded by longitudi-
nal folds. These thickened plates of epithelium are invaginated,
and later reduced to a single layer of cells.

5. In the remainder of the duodenum (approximately the
lower two-thirds) the villi are developing while vacuoles are pres-
ent, but are not recognizable because of the ‘‘solid’’ condition
of the duodenum. The apices of the villi of one wall are adherent
to those of the opposite wall. The spaces in between the adjacent
fused villi are the vacuoles. By a separation of the apices, the
individual villi appear, and the vacuoles become confluent with
one another to form a continuous lumen.

6. In the lower part of the ileum low longitudinal folds occur.
These folds, varying from two to five in number, are irregular in
height and position. Their presence is probably due to the fact
that the outer coats of the gut do not grow as rapidly as the epi-
thelium. The irregularities along the tops of the folds are develop-
ing villi. Later the folds disappear by an expansion of the epi-
thelial tube or by being absorbed by the rapidly growing villi.

7. As the villi develop they become arranged in longitudinal
rows of varying regularity.

8. Additional villi develop as separate growths, either forming
new rows between those already present, or arising in the older
rows.

9. The growth of the younger villi, up to the size of the larg-
est, is more rapid than that of the older ones.

10. Intestinal glands develop as separate hollow outgrowths
of the epithelium. They were first seen in an embryo of 55 mm.

Mucous Membrane in the Human Embryo. 559

11. Duodenal glands originate as outgrowths from the intes-
tinal glands. They branch repeatedly and the branches of each
become arranged in groups. They were first observed in an em-
bryo of 78 mm.

12. Circular folds of the mucous membrane make their ap-
pearance in the mid region of the small intestine at 73-78 mm.
They become more numerous, more regularly placed, and taller
in the older stages.

560 Franklin P. Johnson.
BIBLIOGRAPHY
Baainsky. Untersuchungen iiber den Darmkanal des menschlichen Kindes.

1882 Arch. fiir path. Anat., Bd. 89, p. 64-94.

Bartu. Beitrige zur Entwickelung der Darmwand. Sitzungsber. d. Wien. Akad.
1868 d. Wiss., Bd. 58, p. 128-136.

Berry. On the development of villi in the human intestine. Anat. Anz., Bd.
1900 17, p. 242-249.

Branp. Beitrige zur Entwickelung der Magen- und Darmwand. Verh. der phys.
1877 med. Gessellsch. zu Wiirz., Bd. 9, p. 243-256.

Eze. Beitrag zur Histologie des embryonalen Siugetierdarmes. Inaug. Diss.
1909 Freiburg.

Fiscuu. Beitrige zur normalen und pathologischen Histologie des Sauglings-
1891 magens. Zevt. fiir Heilkunde, Bd. 12, p. 395-446.

Fuinr. Organogenesis of the oesophagus. Anat. Anz., Bd. 30, p. 442-451.
1907

ForssnER. Die angeborenen Darm und (£sophagusatresien. Anat. Hefte,
1907 Abt. 1, Bd. 34, p. 1-168.

GortscH. Structure of the mammalian cesophagus. Am. Jour. Anat., 10, p.

1910 1-40.

Hitton. The morphology and development of intestinal folds and villi in verte-
1902 brates. Am. Jour. Anat., 1, p. 459-504.

JAHRMAERKER. Ueber die Entwickelung des Speiserdhrenepithels beim Menschen.
1906 Inaug. Diss., Marburg.

Kriset. Zur Embryologie des Menschen, der Affen, und der Halbaffen. Verh. d.
1905 anat. Gesellsch., p. 39-50.

Kor.iurker. Entwickelungsgeschichte des Menschen und der héheren Thiere.
1861 Leipzig.
1879 Ibid. Aufl. 2.

KoLuUMANN. Lehrbuch der Entwickelungsgeschichte des Menschen. Jena.
1898

Kreuter. Die angeborenen Verschliessungen und Verengerungen des Darm-
1905 kanals im Lichte der Entwickelungsgeschichte. Deutsche Zeitschr.
jf. Chir; Bd: 79; p. 1-89)

Laskowsky. Ueber die Entwickelung der Magenwand. Sitz. d. Wien. Akad. d-
1868 Wiss., Bd. 58, p. 137-148.

Lewis AND Tuyna. The regular occurrence of intestinal diverticula in embryos
1908 of the pig, rabbit, and man. Amer. Jour. Anat., 7, p. 505-519.

Mucous Membrane in the Human Embryo. 561

Minor. Text-book of human embryology. New York.
1892
1900 On the solid stage of the large intestine in the chick. Jour. of the
Boston Soc. Med. Sci., 4, p. 153-164.

NeEuMANN. Flimmerepithel im Gisophagus menschlicher Embryonen. Arch. f.

1876 mikr. Anat., Bd. 12, 570-574.

ScHaFrer. Die oberen cardialen Gisophagusdriisen und ihre Entstehung. Arch.
1904 f. path. Anat., Bd. 177, p. 181-205.

ScHENK. Lehrbuch der vergleichenden Embryologie der Wirbelthiere. Wien.
1874 p. 1-198

ScurippE. Ueber Magenschleimhautinseln im obersten Msophagusabschnitte.
1904. Arch. f. path. Anat., Bd. 175, p. 1-16.
1905 Weiteres zur Histologie der Magenschleimhautinseln in obersten

(Esophagusabschnitte. Jbid., Bd. 179, 562-566.

ScHrIpDDE. Die Entwickelungsgeschichte des menschlichen Speiseréhrenepithe-
1907 les und ihre Bedeutung fiir die Metaplaislehre. Wiesbaden. 101 pp.
1908 Ueber die Epithelproliferation in der embryonalen menschlichen

Speiseréhre. Arch. f. path. Anat., Bd. 191, p. 128-192.

Scuuttze. Grundriss der Entwickelungeschichte des Menschen und der Saéuge-
1899 thiere. Leipzig.

Sewauu. The development and regeneration of the gastric glandular epithelium
1879 during foetal life and after birth. Journ. of Physiol., 1, p.
321-334.

Tanpuer. Zur Entwickelungsgeschichte des menschlichen Duodenum in frihen.
1900 Embryonalstadien. Morph. Jahrb., Bd. 29, p. 187-216.

Toupr. Die Entwickelung und Ausbildung der Driisen des Magens. Sitz. Ber.
1881 Wien. Akad. Wiss., 82 Abth. 3. p. 57-128.

Voier. Beitrag zur Entwickelung der Darmschleimhaut. Anat. Hefte, Abth.
1899 1, Bd. 12; p. 49-70.

THE AMERICAN JOURNAL OF ANATOMY, VOL. 10. NO. 4.

PLATE I

EXPLANATION OF FIGURES

1. Wax reconstruction of epithelium of cesophagus, showing vacuoles. a,
human embryo of 19 mm., series 819. 6, embryo of 22.8 mm., series 871. X 89.

2 Wax reconstruction of epithelium of cesophagus (region immediately below
bifurcation of trachea). Human embryo of 37 mm., series 820. Showing single
dorsal fold. 89.

3. Same, human embryo of 42 mm., series 838. Showing dorsal and ventral
folds and beginnings of lateral folds. X 89.

4. Same, human embryo of 55 mm., embryo 249. Showing Greek cross stage.
X 89.

5. Same, human embryo of 120 mm., embryo 342. Showing four primary and
four secondary folds. Maltese cross stage. > 89.

MUCOUS MEMBRANE IN THE HUMAN EMBRYO
FRANKLIN P. JOHNSON

PLATE I

THE AMERICAN JOURNAL OF Anatomy, Vou. 10. No. 4.

PLATE II

EXPLANATION OF FIGURES

6. Wax reconstruction of epithelium of cesophagus at level of bifurcation of
trachea. Human embryo of 55 mm., embryo 249. Stippled areas represent those
covered with ciliated cells; non-stippled areas, those covered with squamous cells.
xX 89.

7. Same, human embryo of 91 mm., embryo 224. X 89.

8. Wax reconstruction of a cardiac gland from lower end of cesophagus.
Human embryo of 240 mm., embryo 186. Ruled surfaces represent those cov-
ered with glandular cells; non-ruled, those covered with squamous cells. X 89.

9. Wax reconstruction of single fold of epithelium of the cesophagus, viewed
from the exterior. Human at birth, embryo 341. Showing cesophageal glands.
x 89.

MUCOUS MEMBRANE IN THE HUMAN EMBRYO PLATE II
FRANKLIN P. JOHNSON

THE AMERICAN JOURNAL OF ANATOMY, Vou. 10. No. 4.

PLATE III

EXPLANATION OF FIGURES

10. Waxreconstruction of epithelium of fundus of stomach. Human embryo
of 55mm., embryo 249. View of internal surface. X 119.

11. Same, human embryo of 120 mm., embryo 342. » 119.

12. View of external surface of same model as shown in fig. 11. Showing gland
formation. X 119.

MUCOUS MEMBRANE IN THE HUMAN EMBRYO PLATE III
FRANKLIN P. JOHNSON

THE AMERICAN JOURNAL OF Anatomy, Vou. 10. No. 4.

PLATE IV
EXPLANATION OF FIGURES

13. Wax reconstruction of gastric glands of stomach. Humanembryo of 240
mm., embryo 186. X 267.

14. Same at birth, embryo 341. X 267.

15. Wax reconstruction of epithelium of upper part of duodenum. Human em-
bryo of 22.8 mm., series 871. Showing beginning villi and duct of the dorsal pan-
crease. X 89.

16. Wax reconstruction of epithelium of lower part of duodenum. Human em-
bryo of 22.8 mm., series 871. Showing vacuoles and occlusion of lumen. X 89.

MUCOUS MEMBRANE IN THE HUMAN EMBRYO PLATE IV
FRANKLIN P. JOHNSON

16

THE AMERICAN JOURNAL OF ANATOMY, Vou. 10. No. 4.

PLATE V
EXPLANATION OF FIGURES

17. a,b, andc. Waxreconstructions of intestinal pockets. Human embryo of
22.8 mm., series 871. X 89.

18. Wax reconstruction of epithelium of jejunum (first loop in umbilical
cord). Humanembryo of 22.8 mm., series 871. Showing developing villi. a, ex-
ternal view: b, internal view. 89.

19: Wax reconstruction of epithelium of small intestine (mid-region). Human
embryo of 24 mm., series 24. x 89.

20. Wax reconstruction of epithelium of ileum. Human embryo of 30 mm.,
series 913. Showing low longitudinal folds. > 89.

MUCOUS MEMBRANE IN THE HUMAN EMBRYO PLATE V
FRANKLIN P. JOHNSON

THE AMERICAN JOURNAL OF ANATOMY, Vou. 10. No. 4.

PLATE VI
EXPLANATION OF FIGURES

21. Wax reconstruction of epithelium of small intestine (mid-region). Human
embryo of 55 mm., embryo 249. Viewed from interior. X 89.

22. Same, viewed from exterior. X 8&9.

23. Waxreconstruction of small intestine (mid-region). Human embryo of 134
mm.,embryo 30. Showing villi, intestinal glands, and persistent intestinal pocket.
x 89.

MUCOUS MEMBRANE IN THE HUMAN EMBRYO PLATE VI
FRANKLIN P. JOHNSON

23

Tue AMERICAN JOURNAL OF ANATOMY, VoL. 10. No. 4.

PLATE VII

EXPLANATION OF FIGURES

24. Waxreconstruction of epithelium of duodenum (upper third). Human em-

bryo of 240 mm., embryo 186.
M178:

Showing villi, intestinal and duodenal glands.

MUCOUS MEMBRANE IN THE HUMAN EMBRYO PLATE VII
FRANKLIN P. JOHNSON

24

THe AMERICAN JOURNAL OF ANATOMY, VoL. 10. No. 4.

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