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THE JOURNAL

OF

COMPARATIVE NEUROLOGY

EDITORIAL BOARD

Henry H. DonaLpson ADOLF MEYER
The Wistar Institute Johns Hopkins University
J. B. JOHNSTON OuiveR 8S. StRoNG .
University of Minnesota Columbia University

C. Jupson HERRICK, University of Chicago
Managing Editor

VOLUME 33

APRIL—DECEMBER
1921

PHILADELPHIA, PA.

THE WISTAR INSTITUTE OF ANATOMY AND BIOLOGY

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CONTENTS

No. 1. APRIL

F. L. Lanpacre. The fate of the neural crest in the head of the urodeles. Four plates

eleven fPUTES). «2066.0. - occ os oe ep RMB: 26s See eee dens con ete he Be Eh Joe 1
Marcaret H. Coox anp H. V. Neat. Are the taste-buds of elasmobranchs endodermal

in origin? Four plates (twenty-nine figures)............--- 0s eee eee eee eee eee es 45
H. V. Neat. Nerve and plasmodesma. One plate (five figures)..............-.++-5-+- 65
S. E. Jonnson anp M. L. Mason. The first thoracic white ramus communicans in man.

TDA GET Tee eee ene Adin ac ae S30 eats Oe roe acne aie aneee SIE TY RCA! Seen ac 77

Sypnry E. Jonnson. An experimental study of the sacral sympathetic trunk of the cat,
with special reference to the occurrence of intrinsic commissural neurons. Seven

eee Anita MEH Dah eGo tafe wise cisrseaic, o's </aha,d eile alma sia/s nie wreeiw ohm eit e = Sinise sei sen eye 24 85
No. 2. JUNE
O. Larsetu. Nerve terminations in the lung of the rabbit. Fifteen figures............ 105
E. C. Casz. On an endocronial cast from a reptile, Desmatosuchus spurensis, from the
iaper briassre of western Vexas. Nine fsures.... 06.6.4 «ace se alas <a seas able es 5 133
J. M. D. Otmstep. Effect of cutting the lingual nerve of the dog. Six Porpee . 149
Howarp Ayers. Ventral spinal nerves in Amphioxus. Seven figures.................-- 155
Apa R. Hatu. Regeneration in the annelid nerve cord. Seventeen figures............. 163

No. 3. AUGUST

Epwarp Horne Craici£. The vascularity of the cerebral cortex of the albino rat. Five

BPRCOR REE Netes lie Pe 8). Oy fe Sprig Mats a Sate peter Shea ttat s Bla aces aces « Faden she 193
C. Jupson Herrick. The connections of the vomeronasal nerve, accessory olfactory
bulb and amygdala in amphibia. Thirty-seven figures.................¢. cesses eens 213

Cut Pine. On the growth of the largest nerve cells in the superior cervical sympathetic
ganglion of the albino rat—from birth to maturity. Six charts and one plate (seven
HURAURRE ERE eT ON alti Agen aie 5 <i Re chem area ete ake Cai fo, cry 2 paral @igli th ele) Atoe ot WAT & > 281

No. 4. OCTOBER

Cut Pine. On the growth of the largest nerve cells in the superior cervical sympathetic
ReanPiiencan THEUNOLWAY Tat, Bive CHATts, xc. 52.5 Sopcast: 5 -a/oncdsees ame Ma arines <4 313

Howarp Ayers. Vertebrate cephalogenesis. V. Origin of jaw apparatus and trigem-

inus complex—Amphioxus, Ammocoetes, Bdellostoma, Callorhynchus. Thirty-six

1V CONTENTS

No. 5. DECEMBER

Hatsert L. Dunn. The growth of the central nervous system in the human fetus as

expressed by graphic analysis and empirical formulae. Thirty-eight figures...... 405
S. R. DerwiterR anp Henry Laurens. Studies on the retina. Histogenesis of the
visual cells. in Amblystoma, - (hirteen fieures! 2.5.7 .% 20s. os = + ee see ee 493

O. LarseLL AnD M. L. Mason. Experimental degeneration of the vagus nerve and its
relation to the nerve terminations in the lung of the rabbit. Five figures.......... 509

Resumen por el autor, Francis L. Landacre.
El destino de la cresta neural en la cabeza de los Urodelos.

La cresta neural de Plethodon se incorpora primeramente al
tubo neural, y después emigra desde la superficie dorsal del
tubo en direccién ventral, a lo largo de la superficie lateral del
tubo, formando porciones de los ganglios de los nervios V, VII,
IX y X, continuando después su emigracién ventral hacia las
barras branquiales, arco mandibular y arco hioideo, en los cuales
forma mesenquima y cartilagos. El mesenquima ectodérmico
puede distinguirse del derivado del mesodermo (mesenquima
endodérmico) por el tamafio de las células, ntimero y tamafio
de los granos de vitelo, pigmentacién y por la continuidad de
las lAminas de células emigrantes. El] mesodermo produce todos
los musculos de la cabeza, mesenquima de la regién cefdlica
dorsal, y la porcién posterior de las trabéculas, los paracordios,
base del erdneo, arco occipital, cdpsula auditiva y segundo
basibranquial. La cresta neural produce la porcién anterior de
las trabéculas, cartilago de Meckel, palatocuadrado, y todos
los cartilagos branquiales con la excepcién del segundo basi-
branquial, asi como el mesenquima de la regién ventral de la
cabeza.

El ectodermo lateral de la regién oral produce un collar ecto-
dérmico, en el cual se desarrollan los dientes y tejidos adyacentes.
Con la excepcién del collar ectodérmico oral, el ectodermo
lateral no produce mesenquima, sino las placodas dorso-laterales,
placodas epibranquiales, parte de los ganglios de la linea lateral,
lineas laterales y ganglio del nervio profundo.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, MARCH 14

THE FATE OF THE NEURAL CREST IN THE HEAD
| OF THE URODELES

F. L. LANDACRE
Department of Anatomy, Ohio State University

ELEVEN FIGURES

CONTENTS
MAG rOCUCELOMI eee Noh tere Lt ET IST ESSE EM cae Wee e ela clotengs eroteleraetes 1
EISTORICATES KU CHirges ce ter esennes ote eae aes oes co Neuaraiolckal snares Sheie eueitionyaiecwrae was 3
INS SRCTON DY Geese, oes Sis ARS oe Leteoee Ia eRe t rre Pa 9
hevoriainromshesneuralaeresbers. ces ts otek. ne Fate 6 nae ate cee oe sae eeiele cies 11
The migration of the neural crest and its relation to the mesoderm....... 18
ahevderiwvativessomune) lateral egtouderm.. 1. ss ciaaeactec.esslscraciowe acl 4 ae oe 22
aM RO NA IME CUO ERIN eR Trina Wee vasa cues chav iopanel Bayshore tots. ofr evo) w et(arcemat onaTe ny hone evens se Beans 25
The differentiation of the mesoderm and neural crest in the head.......... 28
SUTIN yews API syn ew Pee CEN ahs: SEIU A 14h RAN YSN 2 Lights Mnaccihe ecg 32
ILS Aer CNL |v ee hes OA Mee Ot I ORE pee we he > an Rae Se rae ase oe 34
‘INTRODUCTION

During the course of an investigation of the origin of the
cerebral ganglia of the urodeles it became necessary to determine
accurately the ventral limits to which the neural crest migrates.
In the attempt to do this the distribution of the neural crest
was found to be so extensive and to involve so much tissue of a
non-nervous character that it was decided for the time being to
discontinue the study of the cerebral ganglia and follow the
history of that portion of the crest which is not concerned in the
formation of these ganglia.

. The ventral limit of ganglia into the composition of which the
neural crest enters, namely, V, VII, IX, and X, is at the level
of the epibranchial placodes, approximately at the dorsal border
of the corresponding gill slit. The neural crest in all vertebrate
_types, apparently, migrates ventral to this level beyond its
ganglion-forming region and further is distributed in the head
of the embryo anterior to the level of the gasserian ganglion.
1

2 F. L. LANDACRE

Only a relatively small portion of the neural crest in the head is
involved in the formation of cerebral ganglia.

In the urodeles the neural crest forms in the anterior head
region an almost continuous sheet of mesenchyme, lying lateral
to the axial mesoderm which is derived from the endoderm, and
is interrupted for a time by the olfactory bulb and optic stalks
only. In the region of the mandibular and branchial bars, the
neural crest migrates ventrally into these bars to the extreme
ventral level of the body and is prevented from forming a junc-
tion with the crest of the opposite side by the presence of the
heart and ventral aorta.

The possibility of determining the fate of neural crest cells
that do not enter into the composition of cerebral ganglia depends
upon conditions that exist in some types and are apparently
absent in others. It is the almost unanimous opinion of workers
that it is extremely difficult and often impossible to determine
the fate of all neural crest cells in the anterior head region. In
the branchial region of the lower vertebrates particularly, the
situation is very different. A number of reliable workers have
given detailed descriptions of the fate of the neural crest cells
and of the manner in which they enter into the composition of
structures usually considered as mesodermal in origin. As to
the migration of the neural crest beyond ganglion-forming regions
in all vertebrates, there is apparently little doubt. }

In the urodeles the determination of the extent of the migra-
tion of the neural crest and the differentiation of the neural
crest and its derivatives from those of the endoderm are com-
paratively easy except in the anterior region. The contrast
between the large, heavily yolk-laden, light staining endoderm
cells and their derivatives on one hand and the smaller, dark
staining, pigmented, slightly yolk-laden ectoderm cells and their
derivatives on the other hand is very striking. In addition to
these distinctions based upon size of cells, staining reaction,
pigmentation, and number of granules, there is also an actual
difference in the size of yolk granules carried by endoderm cells
as compared with those carried by ectoderm cells, those carried
by endoderm cells being larger. These distinctions, furthermore,

FATE OF NEURAL CREST IN HEAD OF URODELES 3

persist up to so late a stage in development in certain regions of
the head that it can be determined accurately that head struc-
tures, usually considered as of mesodermal origin, are derived
from the ectoderm of the neural crest.

HISTORICAL SKETCH

The idea that ectodermal cells, derived from either neural
crest or from the lateral ectoderm, can be traced into permanent
head: structures, other than ganglia, was first stated definitely
by Miss Platt in 1893. It was claimed in a preliminary paper
published by her at that time that branchial cartilages arose
from ectodermal cells. Before the appearance of her paper,
however, several papers appeared bearing more or less directly
on the fate of the non-nervous portion of the neural crest. Mar-
shall (78), working on the chick, and van Wijhe (’82), on the
selachians, both called attention to the fact that the neural crest
is present in the head anterior to the trigeminus ganglion, but
could not determine the fate of that portion of the crest which
does not enter into the formation of that ganglion.

Kastschenko (’88) went a step farther and stated that, in the
head of the selachian embryo, other layers than the endoderm,
particularly the ectoblast, take part in the formation of mesen-
chyme, and that in the formation of the neural crest some of the
cells detach themselves from the neural-crest mass, become
loosely arranged, and form mesenchyme, while other cells from
the same source are concerned in the formation of ganglia. The
further fate of the mesenchymal cells is not discussed, neither is
their relation to mesenchyme derived from endoderm treated.

Goronowitsch (’92) published a preliminary paper in which
he determined for birds not only that the neural-crest cells give
rise to mesenchyme in the head region, but that they fuse with
the axial mesoderm derived from the endoderm to form a homo-
geneous mesenchyme in which neural-crest cells can no longer be
distinguished. He also noted a _ proliferation of mesoderm
(mesenchyme?) cells from the ectoderm in the dorsal portion of

the gill.

4 . F. L. LANDACRE |

Miss Platt (’93), as noted above, published a preliminary
notice in which she traced the branchial cartilages definitely to
the ectoderm derived apparently from the lateral body wall
rather than from the neural crest. Her statement is rather hard
to follow and caused her work to be criticised and possibly to
be misunderstood. She seems to derive the mesoderm from two
longitudinal ridges which later break up into three vertical ridges.
The median portion of each vertical ridge proliferates mesoderm
into the gill arch to form the cartilage of the gill arch. Ventral
to this point the lateral ectoderm proliferates cells into the
branchial arch to form mesoderm. Miss Platt states that the
neural cells (neural crest?) break up into stellate mesoderm,
whose fate she is unable to follow. She does not in her pre-
. liminary paper introduce the terms ‘mesectoderm’ and ‘mesento-
derm’ for mesenchyme derived, respectively, from ectoderm
and entoderm.

Later in the same year Goronowitsch (’93 a and b) published
on the development of the neural crest in birds and fishes and
asserted again the origin of mesenchyme from both the neural
crest and the lateral ectoderm. He makes the surprising state-
ment that the neural crest is concerned neither in the formation
of ganglia nor nerves, but only in the formation of mesenchyme,
a part of which latter forms the sheath of Schwann and deter-
mines the course of growing nerves. The nerves.arise from
neuroblasts. Goronowitsch derives mesenchyme from the ecto-
derm in the region of the gill arch. The periaxial mesoderm
comes from both neural crest and from the lateral ectoderm
and is associated with the formation of the gill arch presumably,
but he does not derive the cartilages specifically from the ecto-
derm as Miss Platt had done.

In 1894 Miss Platt elaborated the idea contained in her pre-
liminary paper introducing the terms mesectoderm and mesento-
derm for mesenchyme derived from the ectoderm and endoderm,
respectively. She further recognizes the neural crest as con-
tributing to the formation of mesectoderm, though the lateral
ectoderm is its chief source. The axial mesoderm is recognized
in this paper as coming from the endoderm, but in later papers

FATE OF NEURAL CREST IN HEAD OF URODELES 5

(Platt, ’96, ’97) this is questioned, and as a consequence she
discards the term mesentoderm. In these later papers she
derives ganglia, nerves, mesenchyme, and branchial cartilages
and the dentine of the teeth from the mesectoderm, but neither
muscles nor embryonic nerve supporting tissue (sheath of
Schwann). In Miss Platt’s second paper (’94) she follows the
ectoderm cells into the gill bars, but not to their complete differ- -
entiation into cartilage. The later history of these cells and
their differentiation into cartilages was given in the paper pub-
lished in 1897. ‘

Later papers by Kupffer (95), by Lundborg (’99), by Dohrn
(02), and by Brauer (’04) support in the main Miss Platt’s
contention. However, Rabl (’94), Corning (99), Minot (01),
and Buchs (’02) do not agree with her interpretation. The
criticisms of Miss Platt’s theory will be given first.

Rabl (94) at the Strassburg meeting of the Anatomische
Gesellschaft criticised Goronowitsch’s description of the mode of
derivation of the mesenchyme from ectoderm in birds and teleosts
as an assumption, because Goronowitsch admitted that after
ectoderm cells fuse with mesoderm he could no longer follow
them, and consequently he had no right to assume that they
became a permanent part of the mesenchyme. ‘This objection
seems to be valid so far as birds and bony fishes are concerned.
His objection to Miss Platt’s mode of derivation of cartilage
is based not on a study of Necturus, which he admits he has not
examined, but on a study of Triton, salamander, and axolotl.
He thinks that the appearance, which Miss Platt finds in her
preparations, of.cells being proliferated from the ectoderm, can
be explained best as due to faulty fixation. Rabl makes a vigor-
ous defense of the idea of the integrity of the germ layers. Most
of his criticism is devoted to Klaatch’s (94) conception of the
origin of the skeleton of the fins of the fishes. Harrison (’95)
has since shown that Klaatch was wrong in his interpretation.

Corning (’99) derives, in the Anura, from the neural crest,
only ganglia and nerves, the ventral portions of which extend
well down into the corresponding branchial arches. The neural
crest is closely fused with the lateral ectoderm, deriving some of.

6 F, L. LANDACRE

its cells from that source, but in Rana it does not break down
into mesenchyme, according to this author. The mesenchyme
of the head is derived from the endoderm, and is consequently
mesentoderm in Miss Platt’s sense of this word or mesoderm
in the older sense.

Minot (’01), in an address before the New York Pathological
Society, makes a vigorous defense of the doctrine of the integrity
of the germ layers, in which he agrees with Rabl (’94), whose
opinion was expressed under somewhat similar circumstances.
He calls attention to Miss Platt’s work specifically and says,
“‘an examination of a number of series and stages has not enabled
me to find the slightest evidence in favor of Miss Platt’s con-
clusions.”” He says further that ‘‘we may, therefore, I think
safely regard this attempt to overthrow the morphological value
of the germ layers as unsuccessful. I know of no other attempt
of sufficient importance to be even mentioned.” He states in an
earlier paragraph that ‘‘the efforts to upset the validity of this
fundamental doctrine have failed to find support or recognition
from any leading embryologist.”” These statements of Doctor
Minot’s were made previous to the appearance of the work by
(Dohrn (’02) and Brauer (’04), but after the appearance of that
of Kupffer (95) which he must have overlooked. Both Minot
and Rabl seem, in the opinion of the writer, to have given too
much weight to the doctrine of the integrity of the germ layers
in their estimate of a question, which is purely one of accuracy
of observation and description.

Buchs (’02), after studying Necturus, disagrees with Miss
Platt’s conclusions, taking exception particularly to her state-
ments concerning the derivation of mesenchyme from the lateral
ectoderm and to her distinction between mesectoderm and
mesentoderm on the basis of the amount and size of the yolk
granules. Buchs’ opposition to Miss Platt’s interpretation of
the mesenchyme is based frequently on minor details and possible
ambiguities in statements, although he has worked over the
same type. He derives cartilage from mesenchyme which arises
from endoderm by a folding process of the endoderm and not
from ectoderm. He can find no evidence for the wandering of

FATE OF NEURAL‘CREST IN HEAD OF URODELES 7

the precartilage cells from the nerve anlagen or from the ecto-
derm. He denies the contribution of cells from the ectoderm
to the mesenchyme, but does not follow the fate of neural-crest
cells.

Of the writers opposing Miss Platt’s hypothesis, two only,
Corning (’99) and Buchs (’02), give sufficiently detailed descrip-
tions and figures to enable one to estimate the value of their
criticism. Corning certainly did not follow his stages far enough
to determine that the mesoderm of the branchial bars is com-
pletely surrounded by neural-crest cells. This can be seen on
any good series of the frog, Rana pipiens. Buchs, on the other
hand, contents himself with an effort at destructive criticism.
His actual evidence is of a negative character, since he does not
follow the fate of neural-crest cells that do not form ganglia.

Of the authors who support wholly or in part Miss Platt’s
~ contention, the earliest is Kupffer. Kupffer (95) described in
detail and figured the cartilages in Ammocoetes as arising from
the deeper layer of the ectoderm in the region of the branchial
bars. He had previously (94) designated this layer as neuro-
dermis, believing it to be concerned in the formation of the
branchial nerves, but here designates it as branchiodermis and
derives not only the cartilages, but goes a step farther than Miss
Platt and derives muscles also from it. He agrees fully with
Miss Platt’s interpretation after examining her preparations.
He further derives mesenchyme in the dorsal anterior head regions
from the neural crest.

Lundborg (99) derived the pterygopalatine cartilages in
Salmo salar and the trabeculae in Rana temporaria from the
ectoderm of the roof of the mouth. He also derived the ethmoid
cartilages in Salmo in the same manner. The anterior end of
these cartilages in Salmo are in process of formation in sixty-
eight-day-old embryos.

Koltzoff (02), in Petromyzon, derives mesenchyme in the
head from both the neural crest and lateral ectoderm, but is
unable to follow its fate beyond the point of the mingling of
- ectodermal cells with those derived from endoderm.

8 F. L. LANDACRE

Dohrn (’02) gives a full description with numerous figures of
the migration of the neural crest ventrally into the branchial
region and its metamorphosis into mesenchyme and cartilage of
the branchial bars in Torpedo ocillata. He agrees fully with
Miss Platt’s interpretation of the origin of branchial cartilages
from ectoderm, although he derives them from the neural crest
rather than from the lateral ectoderm. He does not exclude the
contribution of the cells from the lateral ectoderm in later stages,
although neither his figures nor text include such a contribution.
He adopts Miss Platt’s term mesectoderm and criticises the
effort of Corning to maintain the integrity of the germ layers in
the formation particularly of structures derived from the meso-
derm. Dohrn’s evidence for the derivation of connective tissues
and cartilage from ectoderm is more convincing, if possible, than
that of Brauer in the Gymnophiona and of Kupffer in Petromyzon,
both of whom give detailed descriptions. Dohrn’s figures taken
with Neal’s (’98) reconstructions of Squalus (figs. 7 to 21, pls.
3 and 4) furnish convincing evidence for the continuous migra-
tion ventrally (Neal) of the neural crest and its ultimate trans-
formation into permanent mesenchyme and cartilage of the
branchial bars (Dohrn).

Brauer (’04), working on the Gymnophiona, derives mesen-
chyme of the anterior region of the head from neural-crest cells
which later mingle with cells derived from endoderm to form
mesenchyme in which the two derivatives cannot be recognized
after this fusion. He can find no evidence for the disappearance
of these neural-crest cells, however, before their fusion with
cells derived from the endoderm. In the posterior head region
neural-crest cells, when not involved in the formation of ganglia,
grow ventrally into the branchial bars and surround the meso-
derm of the bar at first lying on its lateral surface, but finally
entirely surrounding it. He can find no evidence for the deriva-
tion of mesenchyme in the gill bar from the adjacent lateral
ectoderm, as Miss Platt had done, but derives it entirely from
the neural crest. He does not deny the possible later mingling
of cells derived from mesoderm with those derived from the
neural crest, but insists that the chief part in the formation of

FATE OF NEURAL CREST IN HEAD OF URODELES 9

the gill bar is performed by cells derived from the neural crest.
Brauer objects to Miss Platt’s terms mesectoderm and mesento-
derm which had been accepted by Dohrn and Koltzoff. He
reserves his description of the ultimate fate of the ectodermal
mesenchyme of the gill bar for a later paper and consequently
does not describe the origin of definite cartilage.

An impartial examination of the papers cited above furnishes
strong evidence for the formation of the mesenchyme in the
anterior head region in the embryo from both endodermal
(mesentoderm) and ectodermal (mesectoderm) sources. In all
cases the fate of these individual cells is lost and it is not possible
to determine the extent to which either or both of them is con-
cerned in the formation of adult mesenchyme in this region.
However, evidence for the disappearance of mesectoderm cells
in the anterior head region is conspicuously absent. The same
statement holds in the main for the dorsal mesenchyme in the
posterior portion of the head.

The fate of the ectodermal derivatives in the branchial regions
is much more definitely stated. Platt (93), Kupffer (95),
Brauer (’04), and Dohrn (’02), all derive either the cartilages
and mesenchyme or both cartilages and muscles in addition to
mesenchyme from the ectoderm. Platt (’93), Kupffer (95), and
Koltzoff (’02) derive the ectodermal cells in the branchial region
largely or even altogether from the lateral ectoderm, while
Brauer (’04), Corning (’99), and Buchs (’12) can find no evidence
for the proliferation of cells from the ectoderm, and Brauer (’04)
and Dohrn (’02) derive the structures in the branchial bar
entirely so far as they are ectodermic from the neural crest. _

MATERIAL

The material on which the work was‘done consists of three
series of urodele embryos collected in the same pond, but repre-
senting at least two different species. The youngest series
(series I) consists of thirty-five stages taken from two egg clusters
at intervals of five to five and one-half hours, reared at room
temperature, and covers an interval of eight days. The first

10 F, L. LANDACRE

stage of this series was fixed immediately after collection and the
medullary folds appear in stage 6. This series, judged by the
size of the egg clusters and character of development, is evidently
Plethodon glutinosus.

The second series is older and consists of thirty-two stages
taken from four egg clusters reared at room temperature and
fixed at intervals of about five hours. The total period covered
by this series is about eight days. The youngest stage of this
series corresponds closely to no. 32 of series I. The larvae were
reared and identified as Plethodon glutinosus. These larvae
hatched in the twenty-fourth stage, so that combining series
I and II there are fifty-six stages taken previous to hatching.

The third series (series III) consists of fifty-one stages taken
from five egg clusters reared at room temperature and fixed at
four-hour intervals. The total period covered is again about eight
days. ‘The first stage of series III corresponds closely to no. 10
of series I and the twenty-third stage of series III corresponds
to no. 1 of series II. ‘The larvae of this series were reared up to
metamorphosis, but escaped before an identification could be
made. Seventeen stages of this series only were cut.

Including duplicates, there are 292 slides in all. All stages
were cut transversely, and critical stages were cut in the sagittal
and coronal planes also. The material was fixed in Zenker and
stained in Delafield’s haematoxylin and counterstained in orange
G. The younger stages were covered with a film of celloidin. to
prevent the loss of yolk-laden cells which are likely to become
detached. ‘The closeness of the series and their sequence are
particularly important in the discussion of the problem involved
in this paper. Where a series is taken from more than one
cluster of eggs, care was used to insure that the series would be
continuous by having the eggs taken from the two lots overlap
in time. )

FATE OF NEURAL CREST IN HEAD OF URODELES iy

THE ORIGIN OF THE NEURAL CREST

There are in the vertebrates three modes in which the neural
crest is related to the neural tube and overlying ectoderm. Har-
rison (’01) has described and compared two of these. In the
first type, the neural-crest cells—selachians and other types—
represent the dorsal portion of the lateral walls of the neural
tube which is at first continuous with the ectoderm. The
neural crest is incorporated in the neural tube, forming a wedge-
shaped mass in its dorsal portion. This wedge-shaped mass
later becomes detached from the tube and migrates laterally
and ventrally. In the second mode the neural crest (teleosts)
lies between the dorsal border of the neural tube and the ecto-
derm, not being included strictly in either, but forming later a
cap over the dorsal border of the neural tube. In the third mode
(Ameiurus and other types (Landacre, ’10)) the neural crest or
cells homologous to the neural-crest cells remain in the ectoderm
lateral to the neural canal and are later detached from this
position to form parts of cranial ganglia and other structures.

The three series of urodeles studied correspond closely in the
behavior of the neural crest to the first type mentioned above
and do not require an extended description. The neural crest
is completely incorporated in the dorsal border of the neural
tube, but can be distinguished from the tube, usually, by its
looser structure. The following description is based on the
behavior of the crest at the level of the VII ganglion.

When the medullary plate can be identified first, it is very
broad and its lateral borders include the two portions of the
neural crest. Just before the closure of the neural canal (fig. 1)
the superficial layer of the ectoderm and much of what will
become neural crest are invaginated and included within the
limits of the neural groove. The superficial heavily pigmented
layer of the ectoderm forms the inner lining of the greater portion
of the neural groove. The looser texture and greater pigmenta-
tion of the dorsomesial portions of the walls of the neural groove
indicate roughly the position of the neural-crest cells.

11 F, L. LANDACRE

In the closure of the neural groove (fig. 2) the superficial
pigmented cells lining the dorsal two-thirds of the neural tube
come into contact and obliterate that portion of the canal lined
by flat cells. The line of juncture is indicated by the heavy
pigmentation of the cells. The dorsomesial portion of the wall
of the tube has a looser texture and less of a syncytial character
than the lateral and ventral portions. The neural tube as a
whole is well delimited from the ectoderm except at the dorsal
border.

The outline of the neural tube (fig. 3) as distinct from the
neural crest first becomes apparent when the cells of the tube
assume a syncytial character with their nuclei arranged with
their long axes toward the center of the neural canal. At the
same time the neural crest while still forming a conspicuous
wedge in the dorsal portion of the tube is evidently now largely
outside the limits of the tube. The dorsal third of the tube
becomes neural crest and presents the appearance of being
erupted from the tube. The tube is horseshoe-shaped with the
open dorsal portion filled with a wedge of neural-crest cells.
The tube except at the ventral border is of uniform thickness.

The next step (fig. 4) involves the further exclusion of neural-
crest cells from the dorsal wall of the tube. In this process many
of the loosely arranged, heavily pigmented cells are left for a
time in the position of the original wedge. In fact, after the
neural crest is well defined and has begun to migrate ventrally
(fig. 5) a few cells of this type still form the roof plate of the
neural canal. In this stage (fig. 5) the greater portion of the -
neural crest rests upon the dorsal portion of the neural tube,
but there are two prominent lateral extensions lying between
the dorsolateral border of the neural tube and the ectoderm.
At a slightly later stage (fig. 6) the neural crest is represented
almost exclusively by these lateral extensions. The large mass
of cells previously lying over the tube is now represented by a
few flat cells connecting the two lateral portions of the neural
crest. These flat cells disappear later. In figure 6 the original
wedge seems to be represented by a few irregularly arranged
cells.

FATE OF NEURAL CREST IN HEAD OF URODELES 13

Up to the last stage described there is practically no indication
that cells are added to the crest, as defined, from the lateral
ectoderm. The crest is continuous on its dorsal border with
the inner layer of the ectoderm, and undoubtedly receives cells
from this source (figs. 1, 2, 3, and 5), and in several series at the
age of that from which figure 3 is taken and the crest is continuous
ventrally with the inner layer (fig. 3). This continuity is usually
absent at the level under consideration as the lateral portions of
the crest become better defined (figs. 5 and 6) and also in the
earlier stages (fig. 4).

These conditions as described at the level of the VII sun
are duplicated at the levels of ganglia V, IX, and X. In the
intervals between these ganglia the neural-crest cells are less
numerous and the lateral extensions contain fewer cells and do
not reach so far ventrally. At the stage from which figure 5
was taken (fig. 7), the neural crest is continuous throughout the
whole region. In the stage from which figure 6 was taken
(fig. 8) the crest is interrupted, except for a few scattered cells
on the dorsal portion of the cord, between the V and VII ganglia
and again between the VII and IX ganglia.

THE MIGRATION OF THE NEURAL CREST AND ITS RELATION TO
THE MESODERM

The accuracy with which the migration of the neural crest
ean be followed depends upon histological differences between
cells derived from the ectoderm as distinguished from those
derived from the endoderm. ‘These distinctions have been stated
already in the introduction (p. 2). They will be assumed
for the present and the migration of the crest will be described
from the reconstructions given in figures 7 to 11. The discussion
of the basis of these distinctions together with the discussion of
two other disputed points, namely, the question as to the con-
tribution by the lateral ectoderm of cells to the mesenchyme
and the question as to the fate of cells in the anterior head region,
will be deferred to a later section.

As to the use of terms, Miss Platt rejected the term ‘mesento-
derm’ presumably because the term implies a derm or germ layer

14 F, L. LANDACRE

derived from entoderm, and of course this is equivalent to meso-
derm and therefore superfluous. There is in the head region,
however, a good deal of loose tissue quite similar to mesenchyme
found in the trunk and like that found in the trunk, derived from
mesoderm, which is itself a derivative of endoderm. Now, since
it is necessary, in describing the behavior of mesenchyme in
the head, to distinguish between that derived from the neural
crest, which retains most of its ectodermal characters, and that
derived from mesoderm, which for a long time retains its endo-
dermal characters, I have ventured to suggest the terms ‘ento-
dermal mesenchyme’ and ‘ectodermal mesenchyme’ for mesen-
chyme in the head where the two types of cells can be distin-
guished. When head mesenchyme becomes homogeneous, that
is when we can no longer distinguish two types of cells, it will
be referred to as mesenchyme, with the implication, however, that
it sometimes contains both ectodermal and entodermal cells.
While both types of mesenchyme have passed through inter-
mediate stages, the entoderm through a mesodermal stage and
the ectoderm through a neural crest stage, each retains the char-
acter of its more remote rather than of its immediate ancestor.
This seems to justify the terms ectodermal mesenchyme and
entodermal mesenchyme rather than neural-crest mesenchyme
and mesodermal mesenchyme. The term ectodermal mesen-
chyme is substituted for mesectoderm, which has become general
in the literature. The tissue we are dealing with is not a derm
or layer, but a true mesenchyme quite similar to mesenchyme in
the trunk, but coming from ectoderm rather than from mesoderm
as is in the trunk.

The use of the terms ectodermal mesenchyme and endodermal
mesenchyme is further justified by the fact that in the head all
branchial muscles come from the mesoderm, and show through-
out their earlier stages definite somatic and splanchnic layers,
indicating their relation to the lateral mesoderm of the body,
while the mesenchyme of the head is more or less loose and of a
syncytial character, like that derived in the body from the
sclerotomes and the lateral and splanchnic mesoderm. As the
mesoderm of the anterior trunk region grows forward into the

FATE OF NEURAL CREST IN HEAD OF URODELES 15

head, it gives rise to a) eye muscles from its dorsal or somitic
portion; 6b) branchial muscles from its ventral portion, and, c)
mesenchyme which retains it endodermal characters. That por-
tion of head mesoderm which gives rise to branchial muscles
does not become loose or syncytial in character as does the
mesenchyme, but retains its integrity to such an extent that
two layers, somatic and splanchnic, can for a long time be
identified.

The neural crest, on the other hand, gives rise to specific
cerebral ganglia such as V, VII, IX, and X and then migrates
ventrally into the ventral head region and into the branchial
bars and differentiates into cartilages and loose mesenchyme.
The neural-crest ectoderm furnishes three specific derivatives,
a) ganglia, b) cartilages, c) mesenchyme. Since the two types
of mesenchyme overlap and are histologically distinct for a
long time and since the ventral portion of the neural crest passes
from a mesenchymal stage to a cartilaginous stage, the terms
entodermal mesenchyme and ectodermal mesenchyme seem not
only to be justified, but to be absolutely necessary to accurate
description.

We shall first follow the migration of the neural crest and
mesoderm.

In the first stage plotted, which is 3 mm. in length (fig. 7),
the neural crest is continuous along its dorsal border throughout
the whole head region, beginning anteriorly at the level of the
middle of the eye horizontally and extending caudally into the
spinalmneural crest. It presents three conspicuous enlargements.
The anterior enlargement extends caudally to the vertical level
of the posterior border of the eye. The gasserian ganglion
differentiates out of the posterior portion of this enlargement
and can be identified at this stage by the slight condensation
of the cells. This enlargement is a ventral extension of the neural
crest, but owing to the flexure of the head it seems to extend
caudally.

_. The second enlargement or ventral extension is at the level
of the VII ganglion, the third at the level of the IX ganglion,
‘and the fourth inconspicuous enlargement at the level of the

16 F. L. LANDACRE

anterior division of the X ganglion. Posterior to this level the
neural crest gradually becomes narrower dorsoventrally and
passes into the neural crest of the spinal cord. In referring to
the V, VII, IX, and X ganglia it is to be understood that only
the general cutaneous and general visceral portions of these
ganglia are under consideration. These are homologous to
spinal ganglia. The special somatic and special visceral ganglia
are referable to other sources.

‘The endodermal derivatives at this stage fall into two regions.
First the posterior head region, which extends cephalad to the
level of the anterior end of the alimentary canal and is char-
acterized by the presence of somites and a two-layered lateral
mesoderm. The somites are not well defined at this stage and
the lateral mesoderm forms a broad sheet extending from a level
slightly dorsal to the notochord to the ventral limit of the body,
being absent only in the heart region and the region of the
stomodaeum.

The second, or anterior head region, lies anterior to the level
of the anterior end of the alimentary canal and forms a loose
mesial or axial mass of entodermal mesenchyme and two lateral
extensions. These lateral extensions contain cavities which in
the urodeles do not form definite head cavities or somites, but
their dorsal portions are undoubtedly homologous to the head
somites of selachians. The dorsal border of this entodermal
mesenchyme maintains the same relative level as the dorsal
border of the somites. It reaches cephalad to the vertical level
of the middle of the optic vesicle and ventrally it forms a promi-
nent extension, which is separated from the anterior border of
the lateral mesoderm by an area free from mesoderm cells. This
area’ corresponds roughly to the future position of the spiracular -
gill cleft. The ventral border of this extension reaches nearly
to the ventral limit of the optic vesicle. The whole of this
anterior head mesoderm consists of the large endoderm cells
containing large yolk granules and except those regions giving
rise to eye and branchial muscle has a loose arrangement presag-
ing its modification into typical mesenchyme. The neural crest
overlaps the cells derived from endoderm at two points only—

FATE OF NEURAL CREST IN HEAD OF URODELES 17

in the region of the V ganglion and at the ventral border of the
IX ganglion. In the overlapping areas mentioned the ento-
dermal mesoderm lies mesial to the neural crest.

In the second stage plotted, 3 mm. in length (fig. 8), but
six hours older than the last, the neural crest in the region of the
gasserian ganglion and anterior to this ganglion has migrated
ventrally and caudally and now forms two prominent extensions,
the one anterior to the eye and mesial to the olfactory capsule,
the other posterior to the eye and extending slightly into the
mandibular bar ventral to its ganglion-forming region. This
anterior neural crest is now completely detached from the neural
crest in the region of the VII ganglion, except for a few scattered
cells on the dorsal portion of the neural tube.

The neural crest at the level of the VII ganglion has grown
ventrally to the region of the dorsal border of the alimentary
canal, doubling its length as compared with the last stage. The
IX ganglion has also grown ventrally to the same extent and the
X is now represented by a conspicuous ventrocaudal extension
of the same general neural-crest mass from which the IX ganglion
forms. The neural crest of VII, IX, and X have not, as yet,
grown ventrally beyond their ganglion-forming regions. The
neural crest in the region of VII is connected dorsally by a well-
defined strand of neural crest cells with that of ganglia [X and X.

The lateral mesoderm lying posterior to the anterior end of
the alimentary canal at this stage is interrupted by the out-
growth of the endoderm to form the spiracular and hyoid pha-
ryngeal pouches. The interruption of the lateral mesoderm at
the level of the spiracular cleft is not formed entirely. by the
pharyngeal pocket. Its ventral portion represents the remains
of the prominent notch shown just caudal to the eye in figure 7.

The head mesoderm lying anterior to the alimentary canal
has extended in two directions, dorsally and anteriorly, until
it has reached the dorsal wall of the brain at the vertical level
of the anterior end of the optic vesicle. It has also extended
ventrally and caudally posterior to the optic vesicle. The
posterior boundary of this extension lies just ventral to the
spiracular pharyngeal pocket. The ventral limit of this extension

18 F. L. LANDACRE

reaches almost to the ventral border of the optic vesicle. The
region of the overlapping of neural crest and mesoderm is quite
extensive, as indicated in figure 8. In the region just posterior
to the eye the mesoderm is not completely covered by neural
crest. In the region of the VII, IX, and X ganglion the ventral
half of each ganglion overlaps the lateral mesoderm and lies
lateral to it.

In figure 9 from a stage 4 mm. long and ten hours older than
the stage from which figure 8 was taken, the neural crest has not
altered its relation greatly except that posterior to the eye it
has grown ventrally and caudally into the mandibular bar and
now almost covers, on the lateral surface, the primordium of the
mandibular muscles. This extension reaches almost to the
ventral limit of the body and far beyond the ventral limit of the
V ganglion. In the region of the VII and X ganglia, the ventral
extensions of the crest have moved into the hyoid and second
true branchial bars reaching, at least in the case of VII, to the
middle of the body and well beyond the ganglion-forming region.
The crest in the region of IX shows little change.

The lateral mesoderm in the pharyngeal region is now inter-
rupted by three visceral pouches, but is otherwise unmodified
so far as its extent is concerned. In the anterior head region
the mesoderm has extended cephalad over the eye and ventrally
at the same level so that now it lies between the dorsal border
of the optic vesicle and the brain. The increase in the amount
of overlap of neural crest is most marked in the regions of the
VII and X ganglia.

Figure 10 is taken from stage 12 of series III and is 43 mm.
long and approximately twelve hours older than the stage from
which figure 9 was taken. In this stage there are two striking
changes in the extent of the neural crest noticeable in the anterior
head region. The first is progressive and carries the neural
crest ventrally and slightly cephalad in the region of the olfactory
and optic vesicles. The olfactory capsule is now completely
separated from the brain wall by a sheet of ectodermal cells
which, extending ventral and caudal from the olfactory capsule,
reaches the ventral limit of the brain wall below the optic vesicle.

FATE OF NEURAL CREST IN HEAD OF URODELES 19

Much of the optic vesicle, aside from the region of the optic
stalk, is separated from the brain wall by this continuous sheet
of ectodermal mesenchyme.

The second change in this region is regressive and is indicated
by the absence of ectodermal mesenchyme in the region lying
vertically over the olfactory capsule. The question as to whether
this is due to a withdrawal by migration or to a disappearance
of the cells will be taken up in the next section. Althoughthe
anterior limit of the ectodermal mesenchyme is hard to determine
on account of the scattered neural-crest cells in this region, there
is no doubt that the process of disappearance of neural crest at
this point has taken place. It is much more marked in the
next stage plotted (fig. 11). Posterior to the eye, the change
in the extent of the ectodermal mesenchyme is slight as seen
from the lateral surface. It is almost coextensive with the pri-
mordium of the mandibular muscle mass. It surrounds this
mass completely, however, so that the endodermal. derivative
of the mandibular bar is inclosed by a sheet of ectodermal mesen-
chyme except at its extreme posterior tip.

In the posterior head region at the levels of the VII, IX, and
X ganglia the most marked changes are the ventral growth of
the neural crest into the hyoid and first two true branchial
bars, with the accompanying process of surrounding the endo-
dermal derivatives (branchial muscles) in these bars by ecto-
dermal cells. This last process is, of course, not indicated on
the plot. Posterior to the pharyngeal pouch of the second
true gill in the region of X the neural crest has not only grown
ventrally, but also caudally as a broad sheet which will be pierced
by the third true branchial pouch, thus setting off the second
branchial ganglion of X from the first. In this process the mode
of development is altered somewhat, since in the more anterior
gill bars the neural crest grows ventrally into the bars after they
are formed by the pharyngeal endodermic evagination, while
here the migration of the neural crest ventrally precedes the
formation of the pharyngeal evagination and must be displaced
by it. As in the more anterior ganglia, the dorsal portion of the
broad mass extending caudally from X forms ganglion, while

20 F, L. LANDACRE

the ventral portion takes part in the formation of the mesen-
chyme and cartilage of the gill bar.

In the last stage plotted (fig. 11) from stage 1 of series IJ,
6 mm. long, and approximately twenty-four hours older than
that from which figure 10 was taken, the ectodermal mesenchyme
has reached, except in the posterior pharyngeal region, its final
distribution. Its later history involves its differentiation as
distinct from its migration.

In the anterior head region, including the mandibular bar,
the changes in distribution are both progressive and regressive.
The most marked increase in extent of ectodermal mesenchyme
in this region is in the mandibular bar due apparently to the
growth of the bar. There isalsoaslight increase in the extreme
anterior end of the head or rather a residue in the form of spur
of ectodermal mesenchyme extending dorsally from the position
of the epiphysis.. This spur lies morphologically on the dorsal
wall of the midbrain. In the region of the olfactory capsule the
ectodermal mesenchyme forms a continuous sheet between the
forebrain and ectoderm. It also forms a continuous sheet
between the anterior border of the optic vesicle and ectoderm
with a second spur extending dorsally to the level of the pro-
fundus ganglion and a third extending ventrally under the olfac-
tory and optic capsules which is continuous with the ectodermal
mesenchyme of the mandibular bar. Except for the first two
spurs mentioned, the ectodermal mesenchyme is now confined
as a continuous sheet to the anterior and ventral head regions
anterior to the level of the gasserian ganglion. This represents
a rather marked decrease in the relative extent of the ectodermal
mesenchyme in the dorsal portion of the anterior head region.

In the posterior head region there is in this stage a marked
ventral extension of the crest at the levels of V, VII, IX, and X
ganglia. The ectodermal mesenchyme at these levels has grown
into the corresponding gill bars and now reaches to the level of
the heart and ventral aorta, covering from the lateral view the
whole of the entodermal derivatives of these bars. Except in
the posterior bar, the ectodermal mesenchyme has further
completely surrounded the endodermal derivatives. Posterior

FATE OF NEURAL CREST IN HEAD OF URODELES . ral

to the last pharyngeal pouch, the neural crest forms a broad
sheet which will be perforated by the pharyngeal pocket of the
third true gill. In this stage the neural-crest portion of the X
ganglia possesses two ventral extensions or pharyngeal ganglia
in addition to the third broad band lying behind the last pha-
ryngeal pouch.

In the lateral mesoderm posterior to the anterior end of the
alimentary canal there is no change except that it is perforated
by an additional pharyngeal pouch. The withdrawal of the
lateral mesoderm in the region of the last pharyngeal pouch
is more extensive than the contact between the ectoderm and
endoderm, due apparently to the fact that this pouch excludes
both ectodermal mesenchyme and entodermal mesenchyme in
reaching the ectoderm. Mesoderm is certainly absent over a
much larger area after the contact is formed than is ectodermal
mesenchyme. Anterior to the anterior end of the alimentary
canal the entodermal mesenchyme occupies the whole head
region except the area ventral to a line drawn from the region
of the epiphysis to the base of the hypophysis. In this region
at this stage the mesenchyme is derived from ectoderm. ‘The
area of overlap in the anterior head region is slight and can be
best understood by reference to the reconstruction (fig. 11).
In the posterior head region the ectodermal mesenchyme and
mesoderm are coextensive except in the regions between the
ganglia V, VII, IX, and X and in the region posterior to the
X ganglia. ?

The general result of the migration of the neural crest may
be stated as follows: it furnishes first the general somatic and
general visceral portions of the V, VII, IX, and X ganglia.
Whether it furnishes other portions of the V, VII, IX, and X
ganglia must be deferred to a later paper. It also migrates
into and furnishes mesenchyme in the early stages in the mandib-
ular, hyoid, and pharyngeal gill bars. The mesenchyme at
first lies lateral to the endodermal derivative which is always
more dense in the bars, but later comes to completely surround it.
~The neural crest also furnishes a complete sheet of mesenchyme
in the extreme ventral head region. The crest disappears as a

oe F, L. LANDACRE

continuous sheet in the region of the midbrain except for the two
dorsally directed spurs shown in figure 11 and between the V
and VII ganglia, and as a continuous sheet disappears in the
region immediately dorsal to the hypophysis. The identifica-
tion of the crest in these regions rests on certain histological
characters of its cells, but to some extent on the continuity of
the sheet of cells which everywhere grows continuously in a
ventral direction morphologically. The continuity of this sheet
of ectodermal mesenchyme derived from the neural crest needs
to be emphasized, since it facilitates greatly the identification
of the progressive changes in extent of the crest. At points
where the neural crest is diminishing in extent the problem of
describing its boundaries is much more difficult. |

The behavior of the head mesoderm, in addition to the facts
shown by the plots, consists in the formation of branchial muscles
from the lateral mesoderm, which do not lose their entodermal
characters, the formation of eye muscles from the dorsal or
somatic region of head mesoderm, and the formation of mesen-
chyme, particularly in the anterior and dorsal head regions, which
mingles with detached cells derived from the neural crest to
form later homogeneous head mesenchyme.

THE DERIVATIVES OF THE LATERAL ECTODERM

Among authors who derive mesenchyme from ectoderm, one
of the most debated points is concerned with the origin of mesen-
chyme from the lateral surface ectoderm of the head as compared
with the origin from the neural crest. Miss Platt, in her earlier
paper (’94), derived ectodermal mesenchyme entirely from
lateral ectoderm, but in her later paper seems to include the
neural crest also a source of this tissue. Kupffer (’95) also, in
Petromyzon, derives branchial cartilages and mesenchyme from
the lateral ectoderm, which he designates as branchodermis,
and later as neurodermis. Lundborg (’99) and Koltzoff (’02)
both derive mesenchyme from lateral ectoderm. Dohrn (’02),
however, working on Torpedo ocellata, and Brauer (’04), working
on the Gymnophiona, can find no evidence that anything but

FATE OF NEURAL CREST IN HEAD OF URODELES 23

neural crest is concerned in the formation of ectodermal mesen-
chyme and cartilages.

This problem must be settled finally, of course, by observa-
tion; but since my results agree with those of Dohrn and Brauer,
and since I have examined carefully types in which some authors
derive mesenchyme from lateral surface ectoderm, it is interesting
to examine conditions in the lateral ectoderm, which might have
led to a misinterpretation of the facts by earlier authors.

The lateral ectoderm is concerned in the formation of a number
of structures which do not form mesenchyme and their number
and significance are impressive. These structures, taken approxi-
mately in the order of their sequence in development, are as
follows:

a. In the early stages of the formation of the neural plate a
few scattered cells lie between the lateral border of the neural
plate and the thin surface ectoderm, but they are few in number
and show every appearance of detached neural-crest cells. In
the later migration ventrally of the neural crest they are either
lost or incorporated with the neural crest. They certainly fur-
nish no evidence for a general contribution of cells to the mesen-
chyme from the lateral ectoderm.

b. The profundus ganglion arises from the lateral ectoderm
anterior to the gasserian ganglion and is quite extensive both
in its longitudinal and its vertical diameters and shows at times
the appearance of an extensive delamination. A careful study
of its development shows, however, that the delamination of
the ectoderm at this point is concerned solely with the formation
of the profundus ganglion and not with mesenchyme.

c. The lateral ectoderm next gives rise to the lateral-line
ganglion of nerves VIJ, IX, and X in certain types, and espe-
cially in those types in which the homologue of the neural crest
is not included in the neural canal (type III, p. 11). These
forms present the appearance of contributing cells to the mesen-
chyme. Their true fate, however, can be determined to be the
formation of lateral-line ganglia, if one has a complete series
- taken at close intervals.

24 F, L. LANDACRE

d. The lateral ectoderm also gives rise to the lateral-line
primordia (placodes) which are distinct structures, although in
some types closely related in time of appearance and sometimes
even in position with other structures derived from the lateral
ectoderm. If followed carefully, however, their identity can be
established with certainty. Their thickenings or placodes are
usually accompanied by detached cells and should not lead to
confusion as to their fate.

e. Next the lateral ectoderm gives rise to the epibranchial
placodes which furnish the special visceral or gustatory portions
of the VII, IX, and X ganglia. Since these placodes become
completely detached from the lateral ectoderm and added to
the neural-crest portions of the VII, IX, and X ganglia and do
not always have definite boundaries, they might be misinterpreted
as contributions of the lateral ectoderm to mesenchyme in the
branchial region. If these placodes are followed carefully they
are found to have nothing to do with mesenchyme, but are
ganglion-forming structures.

Aside from the oral region, which must be taken up more in
detail, these five structures are all that might be mistaken for
contributions to the mesenchyme. ‘The point to be emphasized,
aside from our own observation, is this: any claim that the lateral
ectoderm contributes to the formation of mesenchyme that does
not take into consideration and account for the structures
mentioned above is open to serious criticism. We believe that.
the disagreement among authors as to the exact condition in the
lateral ectoderm is due to failure to account for the structures
enumerated above before assuming that cells delaminated from
the lateral ectoderm went into the formation of mesenchyme.

With the facts mentioned above in mind, we have examined
carefully our series and, aside from the oral region yet to be
described, we can find no evidence for the contribution of lateral
ectoderm cells to mesenchyme. In fact, the ectoderm at all
points except those mentioned is intact and shows no indication
of liberating cells, and we find ourselves in the closest accord
with the statements of Dohrn (’02) and Brauer (’04), who can
find no such contribution.

FATE OF NEURAL CREST IN HEAD OF URODELES 2S

THE ORAL ECTODERM

Lastly, the oral region in the urodeles presents a curious con-
dition with reference to the positions of ectoderm and endoderm.
None of the authors interested primarily in the origin of head
mesenchyme has described this region carefully. This is all the
more striking, since, on superficial examination, it would seem
to furnish the best evidence for the derivation of mesenchyme
from the lateral ectoderm. Johnston (’10), working on the fate
of the oral entoderm in Amblystoma, gives an accurate descrip-
tion of the oral region. He does not, however, follow the fate
of the mesenchyme. My results agree with his, except in one
particular.

I find, in agreement with Johnston, that the anterior end of
the archenteron is open at first well forward to the position of the
future oral opening. ‘This open oral and pharyngeal cavity next
becomes closed and forms a solid, flat column of entoderm abut-
ting cephalad against the oral ectoderm, which at this point
consists of the usual two layers, a superficial epithelium and a
deeper layer of nervous ectoderm. The next change consists
in the formation of a collar of ectoderm around the solid column
of oral entoderm. ‘This collar grows caudally from the anterior
oral region and mesially from the lateral oral region as far as the
position of the vomerine teeth, which is approximately the
posterior boundary of the solid column of the oral entoderm.
This ectodermic collar is derived from lateral ectoderm as distinct
from neural crest. During the formation of the ectodermal col-
lar the nervous layer of the ectoderm disappears at the line of the
future opening, but we are unable to verify Johnston’s statement
that the ectoderm disappears entirely, leaving the endodermal
column exposed at the oral region. In our preparations the
surface ectoderm is never absent at this point before its rupture
to form the oral opening.

- The next change consists in the formation of a definite oral
cavity in the previously solid column of oral endoderm. It
splits from behind forward, and after the endodermal column
splits, the thin layer of superficial ectoderm ruptures, thus forming

26 F, L. LANDACRE

a true oral cavity lined entirely by entoderm, but possessing
a collar of ectoderm, dorsal to its roof and ventral to its floor,
This curious relation of the ectoderm to the entoderm is to be
interpreted as a modification of the usual process of producing
an oral cavity or stomodaeum by the invagination of ectoderm.
In the urodele the solid column of oral endoderm which abuts
against the ectoderm cephalad prevents the formation of the
usual ectodermal «invagination and we have the ingrowth of
ectoderm to form a collar around a solid column of endoderm,
which later splits to form an oral cavity lined by endoderm
instead of by ectoderm as in other vertebrates. The ectoderm
which in other vertebrates lines the oral cavity is in the urodeles,
consequently, separated from the oral cavity by endoderm.
The ectodermal collar is the structure of greatest interest in
the present discussion. The shape of the mouth in the urodeles
is indicated during the solid stage of the oral endoderm by the
great breadth of the endodermal column; consequently, the
ectodermal collar is also broad and appears in transverse sections
as broad plates of ectoderm lying dorsal and ventral to the endo-
dermal column. These plates are curled at the lateral borders
of the solid endodermal column, bending dorsolaterally and
ventrolaterally around the lateral borders of the endodermal
column and projecting slightly between what will be the roof
and the floor of the oral cavity. These infolded edges cover the
future lips of the oral cavity laterally with ectoderm. The
ingrowth of the ectodermic collar begins first in the 6-mm.
larvae dorsal to the solid endodermic oral collar and later the
ventral portion of the collar is formed in the same manner.
Preceding the formation of the ventral portion of the collar,
there is mesenchyme between the ventral surface of the endo-
dermal oral column and the ectoderm of the mandible, which is
of ectodermal origin. This statement is made in spite of the
fact that occasionally an endodermic cell is found here. Many
of these can easily be’shown to be walls of blood-vessels or isolated
blood-cells which retain their endodermic characters, while
others can be shown to be solid extensions of blood-vessels, and
it is probable that all of them are concerned with blood-vessel

FATE OF NEURAL CREST IN HEAD OF URODELES 20

or lymphatic-vessel formation. However, when the ectodermal
collar forms, its distinctness from the mesenchyme is quite evi-
dent, since the mesenchyme is loose in structure, while the
ectodermal collar is dense and continuous with the deeper layer
of the surface ectoderm. As to the later differentiation of the
ectodermic collar, we find ourselves in agreement with Johnston
(10), who states that it gives rise to the dental ridges and teeth,
while the endodermal lining of the oral cavity gives rise to the
taste buds and oral epithelium.

Concerning the fate of the remaining cells of the ectodermic
collar, we cannot make such definite statements. Aside from
the teeth, the ectoderm gives rise to the dense tissue in which
the teeth are imbedded. ‘The dental ridges and teeth, however,
disarrange the cells of the ectodermic collar where it comes into
contact with mesenchyme, and the collar may possibly contribute
to the mesenchyme, but the indications are against such an
interpretation. We have followed the ectodermic collar up to
the time when membrane bone begins to form around Meckel’s
cartilage and about the bases of the teeth. At this late stage
one is forced to the uncertain method of tracing derivation by
position mainly. Based on this criterion, the bone that forms
about Meckel’s cartilage comes from the mesenchyme of both
ectodermal and endodermal origin, while the membrane sur-
rounding the teeth seems to come from ectoderm only. It is
certainly formed in the same dense tissue of the collar from
which the teeth arise, although we cannot exclude the possibility
of the migration of mesenchyme cells of mixed origin into the
ectodermal collar. The late stage at which true membrane bone
forms renders it unsafe to make definite statements concerning
its derivation from ectodermal mesenchyme as distinct from
entodermal mesenchyme because of the fact that both types
are similar in structure, owing to the loss of yolk granules by the
endoderm and the reduction in size of entodermal mesenchyme
cells.

_ My conclusion, therefore, concerning the contribution of the
lateral ectoderm to mesenchyme is that the lateral ectoderm,
aside from the oral ectodermic collar and its derivations, is not

28 F, L. LANDACRE

concerned in the formation of mesenchyme and that any delami-
nation of cells from the lateral ectoderm is readily explained as
concerned in the formation of other well-known structures.
Dorsal and ventral to the oral cavity ectoderm forms the dense
connective tissue in which the teeth are imbedded, but I find
no evidence for the derivation of loose mesenchyme from the
lateral ingrowth of ectoderm.

THE DIFFERENTIATION OF THE MESODERM AND NEURAL CREST
IN THE HEAD j

The possibility of following the differentiation of mesoderm
into muscles, cartilages, and entodermal mesenchyme, and of
neural crest into ganglia, ectodermal mesenchyme, and cartilages
depends, as indicated earlier in this paper, upon easily recognized
histological differences in ectodermal and entodermal cells.
Ectodermal cells are smaller than entodermal cells, are usually
pigmented, containing brown pigmented granules, and usually
contain few yolk granules, or, if loaded with yolk granules, the
granules are small and fairly uniform in size. Entodermal cells,
on the other hand, are large and contain large numbers of large
granules of varying sizes, and are not pigmented. When an
embryo is stained with Delafield’s haematoxylin and orange G,
the contrast between neural crest and mesoderm is very striking.

These differences between ectodermal and entodermal deriva-
tives can be followed in the urodeles up to the stage where the
neural-crest ganglia, branchial muscles and eye muscles, branchial
and cranial cartilages can be readily identified. One can follow
the differentiation of the ectoderm and entoderm into mesen-
chyme with ease where the two types do not overlap. In areas
of overlap even, the two types can be followed easily in the earlier
stages when they lie in continuous sheets of cells. In the loose
mesenchyme where the ectodermal and endodermal mesenchyme
cells are mingled one can for a long time identify with certainty
ectodermal and endodermal mesenchyme cells. ‘The loose mesen-
chyme finally becomes homogeneous, entodermal cells assuming
the appearance of ectodermal cells, and after this stage it is
impossible to say which type of cell predominates.

FATE OF NEURAL CREST IN HEAD OF URODELES 29

It is a rather striking fact in the distribution of ectoderm and
endoderm in the head that, in general, ectoderm gives rise to
mesenchyme and cartilages located ventrally, that is, farthest
from its source of origin, while mosoderm gives rise to mesen-
chyme in the dorsal regions of the head, which makes its location
farthest from its source of origin. The loose entodermal mesen-
chyme of the head is derived in part from the mesoderm, which
also gives rise to branchial muscles ventrally and to eye muscles
dorsally. The actual formation of mesenchyme in these loca-
tions consists in the detachment of individual cells from the
mesial surface of the more anterior and ventral muscle primordia,
such as the masseter and temporalis, as well as from the more
dorsal mesoderm from which eye muscles arise.

That portion of mesoderm which migrates beyond the muscle-
forming area breaks down completely into loose mesenchyme.
The mesoderm of the head, therefore, shows two types of tissue,
a) dense masses of cells which can be followed into definitive
branchial and eye muscles, retaining all the time their ento-
dermal character up to the time muscle fibrils appear in the case
of branchial muscles, and, b) loose mesenchyme which gradually
loses its entodermal character and assumes the form of ectodermal
mesenchyme.

The formation of ectodermal mesenchyme and cartilages from
neural crest is really easier to follow than the mesodermal deriva-
tives, except in the dorsal head regions from which the ectodermal
mesenchyme withdraws as a continuous sheet. The neural
crest, in its ventral migration, forms a continuous sheet of cells
readily distinguished from mesoderm and lying lateral.to the
mesoderm. This continuous sheet of neural-crest cells gives
rise, in its dorsal region, to the neural-crest ganglia and in other
dorsal regions disintegrates into mesenchyme, which disappears
as a continuous sheet in certain regions as indicated in figures
9, 10 and 11.

That portion of the neural crest which migrates into the ventral
head region, that is, dorsal and ventral to the oral region and
into the branchial bars, presents a quite different history. This
ventral portion of the neural crest does not disintegrate. It not

30 F. L. LANDACRE

only maintains its continuity, but grows extensively, surrounding
the branchial muscles and filling the areas between muscles and
epithelial structures. In this rather dense and continuous sheet
of neural-crest cells there can be identified the still more con-
densed areas where the cartilages, such as Meckel’s and the
branchial cartilages, will form. Coincident with the formation
of procartilage, there is going on a disintegration of the remaining
neural crest to form loose ectodermal mesenchyme in much the
same way as loose entodermal mesenchyme is formed from the
mesoderm in more dorsal regions. Consequently, in the ventral
head and branchial regions where the mesoderm gives rise to the
muscles, most of the loose mesenchyme is derived from neural
crest and is therefore ectodermal, while in the dorsal head regions,
from which neural crest has disappeared as a continuous sheet,
it is chiefly entodermal (mesodermal) mesenchyme.

Our conclusions concerning the metamorphosis of neural-crest
cells into cartilage (excepting Miss Platt’s conception of the
behavior of lateral ectoderm) are so closely in accord with those of
Platt, Dohrn, and Brauer that it is not necessary to give a detailed
discussion of the process. The branchial cartilages all differ-
entiate out of the ventralmost extension of the neural crest.
They all show uniformly a stage where the neural crest is
in the form of a sheet of cells surrounding the corresponding
mesodermic branchial muscle primordium. Since this sheet of
neural-crest cells forms a syncytium, it is true mesenchyme;
although somewhat more compact than entodermal mesenchyme.
Throughout the whole ventral head region and branchial region
the differentiation of this primitive mesenchyme shows two
types of behavior: a) Where no cartilages form its cells become
detached from the sheet of neural-crest cells and form loose
mesenchyme surrounding muscles and cartilages. ‘This is prac-
tically pure ectodermal mesenchyme. The few entodermal cells
found may prove usually to be growing blood-vessels or blood-
cells; b) where a cartilage forms the neural-crest cells first increase
in number, then condense, passing through typical procartilage
stages, and finally form true cartilage. We find, in agreement
with Miss Platt, that the anterior portion of the trabecular

FATE OF NEURAL CREST IN HEAD OF URODELES Sik

bars, the palatoquadrate and Meckel’s cartilages, all the branchial
cartilages except the second basibranchial are formed from ecto-
dermal mesenchyme, while the posterior portion of the trabecular
bars, the parachordals and basal plate of the chondrocranium
along with the occipital arch arise from entodermal mesenchyme.
The roof of the chondrocranium arises very late and is assumed
to come from entodermal mesenchyme on the basis of the distri-
bution as shown in figure 11, although at the stage when the
lateral walls of the cranium appear the mesenchyme is homo-
geneous. I am unable to determine the composition of the brain
membranes and their homologues, the choroid and _ sclerotic
coats of the eye, on account of the lateness of their formation,
which occurs after the mesenchyme from which they form has
become homogeneous. ‘The cartilages listed above as ectodermic
or entodermic in origin show, in addition to the fact that their
development from ectoderm and entoderm can be followed,
quite definite ectodermic and entodermic characters after they
have assumed their definitive forms and after their cells are
surrounded by hyaline material so dense that cell division within
the cartilage has practically ceased.

The formation of the second basibranchial from entoderm
needs a word of explanation. The second basibranchial of Miss
Platt is the urohyal of some authors. It is the most caudal of
the median branchial cartilages and lies ventral to the other
branchial cartilages, except at its anterior attached extremity.
It develops from mesoderm along with the paired geniohyoid
muscles which are attached to its extremity. The mesoderm
from which these three structures develop is continuous at first
with the lateral borders of the alimentary canal from which other
muscle primordia form. It retains for a long time its ento-
dermal type of cell, so that there can be no question as to its
origin. No explanation is offered of this curious fact further than
that suggested by its position, which, as shown by a reference
to figure 11, is at a point where ectoderm does not extend, that
is, on the midventral line ventral to third, fourth, and fifth
branchial bars.

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 1

a2 F. L. LANDACRE

The same explanation is offered for the double composition
of the trabeculae. The anterior portions of the trabeculae form
in a region dominated by ectodermal mesenchyme. ‘This region
lies between the two dorsal spurs just over the eye in figure 11,
while the caudal portions of the trabeculae and the parachordals
form in a region containing axial entodermal mesenchyme.

SUMMARY

1. The neural crest in the urodeles is incorporated in the
neural canal at first and later erupted and then grows ventrally
as a continuous sheet of cells. This ventral migration carries
it into the ventral region of the anterior portion of the head and
into the ventral portions of the mandibular and branchial bars.

2. The dorsal portion of the neural-crest sheet gives rise to
the general cutaneous and general visceral portions of ganglia
V, VII, IX, and X cranial nerves. The dorsal portion of the
neural crest not concerned in the formation of ganglia disinte-
grates to form mesenchyme, which becomes mingled with ento-
dermal mesenchyme in the dorsal head region.

3. The ventral portion of the continuous neural-crest sheet,
after migrating into the ventral head region and into the mandib-
ular and branchial bars, surrounds the primordia of branchial
muscles growing from their lateral surfaces around to their
mesial surfaces. This ventral portion of the neural crest later
differentiates into loose ectodermal mesenchyme and into dense
cartilage primordia, which pass through procartilage into typi-
cal cartilages.

4. The greater portion of the mesenchyme in the ventral
head region and in the branchial regions is consequently ecto-
dermal mesenchyme.

5. The cartilages arising from ectoderm. are the anterior
portion of the trabeculae, Meckel’s cartilage, the palatoquadrate
bar, and all the branchial cartilages except the second basi-
br Sneinal or urohyal.

6. The mesoderm in the head gives rise to the beeen
muscles, to the eye muscles, and to loose entodermal mesen-

FATE OF NEURAL CREST IN HEAD OF URODELES 33

chyme, which retains for a long time its entodermal character-
istics. Entodermal mesenchyme later differentiates into carti-
lages forming the posterior portion of the trabeculae, the para-
chordals and base of the cranium, the occipital arch, the auditory
capsule, and probably the lateral walls of the cranium.

7. The neural crest and mesoderm, where not concerned in
the formation of ganglia and muscles, respectively, shift their
positions extensively, but the net result of their migrations is to
leave entodermal mesenchyme in the dorsal head regions, ecto-
dermal mesenchyme in the ventral head regions, and mixed
mesenchyme in the median longitudinal axis of the head. Where
this overlapping occurs ectodermal mesenchyme is lateral in
position, while entodermal mesenchyme is mesial in position.
The loose axial mesenchyme lying in the median line between
the brain floor and the dorsal surface of the mouth and pharynx
is almost pure entodermal mesenchyme.

8. The lateral ectoderm, aside from the oral region, is not
concerned in the formation of mesenchyme, but does give rise
to the profundus ganglion, to lateral-line ganglia, to lateral-
line organs, and to epibranchial ganglia.

9. The lateral ectoderm in the oral region gives rise to an
ectodermic collar, which surrounds the solid entoderm of the
oral region, and in this collar arise dental ridges and teeth and
dense connective tissue in which the teeth are imbedded. We
can find no evidence for the formation of loose mesenchyme
from this ectodermic collar.

10. The histological grounds on which the distinctions in the
behavior of ectoderm and entoderm rest are as follows: entoderm
cells are larger than ectoderm cells: entoderm cells are rarely
pigmented, while ectoderm cells usually are. Ectoderm con-
tains small, fairly uniform-size yolk granules and loses them
early, while entoderm contains large granules of irregular size
and retains them much longer. In addition to these histological
differences, the continuity of sheets of ectoderm and of entoderm
cells is a valuable aid in following their distribution. The
distinctions mentioned above can be observed after definitive
ganglia, muscles, cartilages, and mesenchyme are formed.

34 F, L. LANDACRE

LITERATURE CITED

Braver, Aucust 1904 Beitrige zur Kenntnis der Entwickelung und Anatomie
der Gymnophionen. IV. Die Entwickelung der beiden trigeminus
ganglion. Zoolog. Jahr., Suppl. Bd. 7.

Bucus, Gnorc 1902 Uber den Ursprung des Kopfskelets bei Necturus. Mor-
phol. Jahrb., Bd. 39, H. 4.

Cornine, H. K. 1899 Uber einige Entwicklungsvorgiinge am Kopfe der Anu-
ren. Morph. Jahrb., Bd. 27.

Dourn, A. 1902 Studien zur Urgeschichte des Wirbeltierkérpers. XXII.
Weitere Beitrige zur Beurteilung der Occipitalregion und der Gan-
glionleiste der Selachier. Mitteil. aus der Zoolog. Station zu Neapel,
Bd. 15H. 4,

GoronowiTscH, N. 1892 Die axiale und die laterale Kopfmetamere der Vogel-
embryonen. Die Rolle der sog. ‘Ganglionleisten’ im Aufbaue der
Nervenstamme. Anatomischer Anzeiger, Bd. 7.

1893 a Untersuchungen iiber die Entwickelung der sog. ‘Ganglion-
leisten’ im Kopfe der Vogelembryonen. Morph. Jahr., Bd. 20.

1893 b Weiteres tiber die ectodermale Entstehung von Skéletanan-
lagen im Kopfe der Wirbelthiere. Morph. Jahrb., Bd. 20.

Harrison, R. G. 1895 Die Entwickelung der unpaaren und paarigen Flossen
der Teleostier. Arch. f. Mikr. Anat., Bd. 46.

1901 Ueber die Histogenese des peripheren Nervensystems der Salmo
salar. Arch. f. Mikr. Anat., Bd. 57.

JOHNSTON, J. B. 1910 The limit between ectoderm and entoderm in the mouth
and the origin of the taste buds. Am. Jour. Anat., vol. 10, no. 1.

KAsTscHENKE, N. 1888 Zur Entwickelungsgeschichte des Selachier Embryos.
Anat. Anz., Bd. 3, No. 16.

Kuaatrcu, H. 1894 Ueber die Herkunft der Scleroblasten. Morph. Jahrb.,
Bd. 21, H. 2. me

Kuprrer, C. v. 1894 Studien zur vergleichenden Entwickelungsgeschichte des
Kopfes von Ammocoetes planeri. Miinchen und Leipzig, 1894.

1895 Uber die Entwickelung des Kiemenskelets von Ammocoetes und
die organogene Bestimmung des Exoderms. Verh. d. Anat. Ges. a. d.
g. Vers. in Basl. Erginzungsheft 2. Bd. 10, d. Anat. Anz. Bd. 3, s. 589.

Kourzorr, N. K. 1902 Entwickelungsgeschichte des Kopfes von Petromyzon
planeri. Bull. Soc. Natural Moscou, vol. 15.

Lunpsporec, H. von 1899 Studien iiber die Betheiligung des Ektoderms an der
Bildung des Menenchyms bei den niederen Vertebraten. Morph.
Jahrb., Bd. 27, H:-2.

Lanpacre, F. L. 1910 The origin of the cranial ganglia in Ameiurus. Jour.
Comp. Neur., vol. 20, no. 4.

Marsuatu, A. M. 1878 The cranial nerves in the chick. Quar. Jour. Micros.
Sci., vol. 18.

Minot, C. 8. 1901 The embryological basis of pathology. Science, vol. 13.

Nea, H. V. 1898 The segmentation of the nervous system in Squalus acan-
thias. A contribution to the morphology of the vertebrate head.
Bull. Museum of Comp. Zool. at Harvard, vol. 31.

FATE OF NEURAL CREST IN HEAD OF URODELES 315)

Pratt, Jutta B. 1893 Ectodermic origin of the cartilages of the head. Anat.
Anz., Bd. 8.
1894 Ontogenetische Differenzierung des Ektodermis in Necturus.
Arch. Mikros. Anat., Bd. 45.
1896 Ontogenetic differentiations of the ectoderm in Necturus, St. II.
Quar. Jour. Micros. Sci., N. S., vol. 39.
1897 The development of the cartilaginous skull and of the branchial
and hypoglossal musculature in Necturus. Morph. Jahrb., Bd. 25,
He 3:

Ras, C. 1894 Ueber die Herkunft des Skelets. Verh. Anat. Ges., 8. Vers.

WisHE, J. W. vAN 1882 Ueber die Mesodermsegments und die Entwicklung der
Nerven des Selachierkopfes. Verh. d. Konikl. Akademie (Amsterdam).

PLATE 1
EXPLANATION OF FIGURES

Figures 1,.2, 3, 4, 5 and 6 are transverse sections of Plethodon glutinosus at
the level of the VII ganglion. The figures were drawn at a mangification of 275
and reduced one-half in reproduction. Figures 1, 2, 3, 4 and 5 are from larvae
between 2 and 3 mm. in length. Figure 6 is from larvae 3 mm. in length.

1 Open neural canal stage.

-2. Closed neural canal stage with the neural crest included in the neural tube.

3 Shows the ‘eruption’ of the neural crest. The neural crest forms part of
the neural tube, but is easily distinguished from the tube.

4 The neural crest is largely outside the neural tube, but still forms a plug
in its dorsal border.

5 Shows the neural crest almost completely outside the neural tube.

6 Shows the neural crest present in the form of typical cerebral ganglia.

36

1

PLATE

Ss

LE

JRODE

T IN HEAD OF

ES

2
L

FATE OF NEURAL CI

F. L. LANDACRE

a
& |

v)

09.6

MLAKD “
v
v

S)

PLATE 2

EXPLANATION OF FIGURES

Figures 7 and 8 are flat reconstructions of the neural crest and mesoderm made
by projecting the camera figures on coordinate paper. The plates give an accu-
rate picture of the extent of the neural crest (diagonal lines) and mesoderm (stip-
ple). The neural canal, notochord, alimentary canal, and sense organs are repre-
sented by lines. Both figures are from larvae 3 mm. in length, but the larva
from which figure 8 is taken is six hours older than the larva from which figure
7 is taken.

FATE OF NEURAL CREST IN HEAD OF URODELES PLATE 2
F. L. LANDACRE

\
Sse

NSS
Sa
Se

RRR

wee O pt Vesicle
Mand ae ar.

PLATE 3

EXPLANATION OF FIGURES

Figures 9 and 10 are reconstructions similar to figures 7 and 8. Figure 9 is
from a larva 4 mm. long and figure 10 from a larva 44 mm. long. The neural
crest is in diagonal lines and mesoderm in stipple.

PLATE 3

FATE OF NEURAL CREST IN HEAD OF URODELES

F. L. LANDACRE

Motochord =
R\vmentary

Canal

5 SAC N
oe
‘ Ny
S

Oo EI
rea ag
Ouee 2 Lo
Lod 12 Bed
D5 V te
o Oo
y_ O E

gee

—_—
je

Zo\

Vesicle

‘ Q t ete

41

PLATE 4

EXPLANATION OF FIGURE

Figure 11 is a reconstruction similar to figures 7, 8,9, and10. This larva was
6 mm. long. Cerebral ganglia are represented by cross-hatched lines; neural
crest by diagonal lines, and mesoderm by stipple. The neural canal, notochord,
alimentary canal, and sense organs are represented by lines. The gill slits are
unshaded. All a the neural crest below the ventral border of the corneas gan-
glia will be converted into mesenchyme or cartilages.

PLATE 4

FATE OF NEURAL CREST IN HEAD OF URODELES

F. L. LANDACRE

ROR Ap,
SSR
\' SON we :

‘Ort. Vesicle

ars
I

ye

SS
AW

‘ Branchial

Comal Be

olin

PG | L
g¢ 3 5
SON eG +2
wc oO 9]
a +6 E
Z =

43

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 1

Resumen por los autores, M. H. Cook y H. V. Neal,
Tufts College, Massachusetts

gSon de origen endodérmico los botones gustativos de los
Elasmobranquios?

El presente trabajo representa un intento de determinacién
del orfgen ectodérmico o endodérmico de los botones gustativos
de Squalus acanthias. Los autores han llevado a cabo un estudio
de cortes de embriones de 7 a 80 mm., suplementado con disec-
ciones de los estados jévenes y adultos. En esta especie los
botones gustativos estan limitados a la regién de la faringe, que
en todos los estados de la ontogénesis esta tapizada por endo-
dermo. Nose puede observar un avance marcado del ectodermo,
ni siquiera en la regién bucal. En el piso y techo de la faringe
se forman escamas semejantes a las que caracterizan a la piel
en los estados avanzados de la ontogénesis. De este modo
parecen originarse en Squalus acanthias dos clases de. 6rganos
que generalmente se consideran como ectodérmicos, esto es,
los botones gustativos y las escamas placoides.

Afirmar que los botones gustativos faringeos y las escamas
de Squalus son de origen ectodérmico implica la admisién de
que el revestimiento endodérmico de la faringe desaparece por
completo durante la ontogenia y es reemplazado por el ectodermo.
No existe prueba alguna de tal sustitucién. Estos resultados
extienden a los Elasmobranquios la conclusién de Johnston
(98, ’05, ’10) de que los botones gustativos de los Teleésteos
y Anfibios derivan del endodermo. También suman a las
estructuras derivadas del endodermo las escamas faringeas,
que hasta el presente han sido consideradas como ectodérmicas,
y asi afiaden otra excepcién a la ley de la especificidad de las
capas germinales.

Translation by José F. Nonidez
Cornel! Medical College, New York

AUTHORS’ ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, FEBRUARY 28

ARE THE TASTE-BUDS OF ELASMOBRANCHS
ENDODERMAL IN. ORIGIN?

MARGARET H. COOK ann H. V. NEAL
Tufts College, Massachusetts

FOUR PLATES (TWENTY-NINE FIGURES)

Scattered over the surface of the pharynx of the common
spiny dogfish (Squalus acanthias) are numerous sense organs
(taste-buds and placoid scales, figs. 28 and 29)—structures which
morphologists generally have regarded as ectodermal. More-
over, the pharyngeal epidermis resembles that of the skin rather
than epithelium such as that of the stomach and intestine, which
is known to be of endodermal origin. Yet, as is perfectly well
known, the pharynx of vertebrate embryos is primarily exclu-
sively lined by endoderm. Are we to believe that the primary
endodermal lining is secondarily replaced by ectoderm? That
the stomodaeal ingrowth extends to the oesophagus? Or that
the specificity of germ layers is not so precise as was formerly
assumed? That endoderm may under certain conditions give
rise to sense organs and placoid scales? Is it possible to settle
these questions? The present paper is an attempt to do so on
the basis of observations upon embryos of Squalus acanthias.

Taste-buds are known to be present in the pharynx of all
vertebrates from cyclostomes to man. They were first described
for fishes by Weber (’27) in the carp, and after him for other fishes
by a number of observers. All of these correctly inferred their
function. Some distinguished between pharyngeal taste-buds
and organs of similar structure situated on the external surface
of the body called ‘end-buds’ or ‘terminal-buds.’

In many fishes and some amphibia terminal-buds occur over
the whole outer surface of the body, extending even to the tail.
This condition Herrick (’04), who demonstrated a similar function
for terminal-buds and taste-buds, correlates with sluggish habits

45

46 MARGARET H. COOK AND H. V. NEAL

and poor vision. In Squalus acanthias, a very active form,
taste-buds are limited to the mouth and pharynx. Here they
arise in the epithelial lining of the floor, the roof and the sides,
including the visceral arches. They are first visible when the
embryo is about 40 mm. in length. They are numerous and
extend forward on the roof and floor of the pharynx to the region
of the upper and lower jaws and posteriorly to the papillae of the
oesophagus. Their distribution shows clearly in a ‘pup’ stage
such as is represented in figure 28 of this paper.

The structure of taste-buds in Squalus acanthias is similar to
that described for other vertebrates. They occur upon small
papillae projecting slightly above the surrounding epithelium.
The papillae are covered with a many-layered epithelium similar
to that which covers the surface of the pharynx. At the apex
of the papilla, however, the epithelial cells are modified into
long, slender ‘sense-cells,’ each of which terminates externally
in a short hair-like projection. Nerve fibers may be traced to
the bases of such cells. Figures 1, 2, and 3 of this paper show
three stages in the differentiation of taste papillae and ‘buds.’

Bateson (’90) and Nagel (’94) were the first to attempt to
demonstrate the function of these structures by experiment.
They worked on a number of forms, including two dogfishes,
Scyllium canicula and 8. catulus. Bateson describes the distri-
bution of taste-buds in the dogfishes as limited to the pharynx,
although he adds that he would not presume to say that they may
not be found also on the outer surface of the body. He failed
to demonstrate that they play any important part in food-getting.
This process, according to his observations, is controlled by the
olfactory organ. Nagel, using dilute solutions of sour, bitter,
and salty substances for taste, finds that the sense of taste is
but feebly developed in Scyllium. He concludes that a sense
of taste, such as in most animals is located in the mouth, is absent
from the outer skin of all fishes.

Recent work by Sheldon (’09) on the smooth dogfish, Mustelus
canis, which resembles 8. acanthias in having the taste-buds
limited to the pharynx, shows that these organs are concerned
in reactions to the stimulus of bitter substances. To quinine

ORIGIN—TASTE-BUDS OF ELASMOBRANCHS - BG

this form reacts in the mouth and spiracle only, that is, in the
region bearing taste-buds, as was discovered by Parker (’08)
for the catfish. But the whole surface of the body is sensitive
to the stimulus of sour, salty, and bitter substances. By oper-
ations Sheldon was able to determine that these reactions of the
general body surface are due to stimulation, not of the gustatory
nerves which supply the taste-buds, but of the nerves of general
sensation. ‘These same nerves of general sensation were shown
to give rise when stimulated to responses even in the mouth,
where also taste-buds and their nerves take part in reactions.
From these results Sheldon concludes, and in this he agrees with
Herrick (’08), that in the dogfish the more specialized senses
of smell and taste (as in the taste-buds) are derived from the
primitive general chemical sense. Parker (712), on the other
hand, considers the olfactory sense the more primitive and
derives from it the chemical sense which later gives rise to the
taste-buds.

Just how much the gustatory nerves are stimulated through
the secretory action of the so-called sense-cells no one has ever
been able to determine. It has, however, been suggested by
Botezat (710) and Parker (712) that these modified epithelial
cells to which the name taste-buds is given may be primarily
secretory, and that the nerves receive their stimulation through
the response (secretion) of these cells to the stimulating sub-
stances. Materials, then, like quinine, would be tasted by the
dogfish, because the cells of the taste-buds react to the quinine
and secrete a substance which in turn stimulates the associated
nerve. Such an explanation would rule out the term ‘sense-cell’
as applied to the groups of slender cells making up the taste-buds.

Turning, now, to the problem of the endodermic origin of the
taste-buds in vertebrates, we find that Johnston (’05—’10) has
been the chief proponent of the view that they are endodermal,
and it is he who has given the most positive and convincing
evidence in support of this view. Johnston (’05) finds that in
Petromyzon and two bony fishes the taste-buds develop first
in the endodermal lining of the pharynx and make their appear-
ance on the outer surface of the body later at the time of hatch-

48 MARGARET H. COOK AND H. V. NEAL

ing. In Catostomus and Coregonus taste-buds arise in the
endodermal lining of the pharynx and are distributed over its
surface and as far posteriorly as the opening of the air bladder.
Johnston (’09, 710) finds even more positive evidence of the
endodermal origin of the taste-buds in Amblystoma, in the
embryos of which the pharynx is lined with endoderm ‘to the
very lips.’ Taste-buds make their appearance within this endo-
dermal-lined pharynx several days before the mouth breaks
through. More direct evidence of the endodermal origin of
the taste-buds could hardly be expected. Yet in this connection
and in direct conflict with Johnston’s assertion that an ectodermal
stomodaeum is wanting in Amblystoma, we have the evidence
given by Kingsley and Thyng (’04) of a stomodaeal ingrowth in
Amblystoma and of a considerable ectodermal invagination
even before the mouth breaks through. While this evidence
by no means disproves Johnston’s main contention that the
pharyngeal taste-buds are endodermal in origin, it is sufficient
to make doubtful the exact amount of ectodermal ingrowth and
to this extent to open up the possibility of an ectodermal origin
of the taste-buds. Landacre (’07) supports Johnston’s view
of the endodermal origin of the taste-buds on the basis of obser-
vations upon Ameiurus embryos. He finds, however, that the
‘terminal-buds’ of the skin, which are so similar to the taste-
buds, are ectodermal in origin. Keibel (712) holds that in man
the taste-buds are “probably derived from the entoblast. The
majority of the taste-buds lie undoubtedly within the ento-
blastic territory, and even although epithelial encroachments
are possible, yet it seems difficult to suppose that the ectoblast
has penetrated into the region of the larynx.”’

The opinion that taste-buds are endodermal in origin seems
strengthened by the evidence presented in sections of Squalus
embryos. An examination of sections of the pharynx of this
form shows that the whole pharyngeal cavity is endodermal in
its origin and that there is little or no inward migration of the
ectoderm into the pharynx except in the region of the upper and
lower jaws, the epidermis of which is ectodermal. Until the
time of perforation of the mouth and visceral clefts it is possible

ORIGIN—TASTE-BUDS OF ELASMOBRANCHS 49

in this form to distinguish ectoderm and entoderm by differences
in staining properties and the presence of large yolk granules
in the endodermal cells. After this rupture the limits of the
two are more difficult to determine. Evidence, however, of
the degeneration of the endodermal lining of the pharynx or
of active extension of the ectoderm is wholly lacking. The
hypophysis soon loses its primary connection with the ectoderm
from which it arises and from it therefore evidence of ectodermal
ingrowth into the mouth is not obtainable as in Amniote embryos.
As the embryo continues to grow the upper lip region is pushed
backward slightly and the ectoderm is carried with it forming
the dental ridge, as is shown in figure 16.

Throughout the formation of the pharynx and of pharyngeal
structures, such as the thyroid and gill pouches, it is the endo-
derm which is the active layer. And it is within the endoderm
that the taste-buds first make their appearance in a 45-mm.
embryo by the local thickening of the epidermis and the differ-
entiation of cells of the stratum germinativum. To assert that
the taste-buds in the pharynx of Squalus are ectodermal would
necessitate the assumption—for which there is not a particle
of direct evidence—that the primary endodermal lining of the
pharynx is completely supplanted by ectodermal ingrowth.

In connection with the study of the histogenesis of taste-buds
in Squalus another problem presents itself in the appearance in
late embryonic stages of pharyngeal placoid scales, the distri-
bution of which, as is shown in figure 29, is somewhat more
restricted than that of the taste-buds. There are relatively
few scales upon the roof of the pharynx, while they are abundant
upon its floor, where in the region of the basibranchial cartilages
they tend to conceal the taste-buds. The number of scales in
the pharynx varies considerably in different individuals. In one
individual examined there were so few that they could be detected
only by boiling the epithelium of the roof of the pharynx in
strong KOH solution. Yet in some individuals they may be
easily distinguished with a hand lens or by rubbing the fingers
anteriorly over the surface of the pharynx, since the scales like
the pharyngeal teeth of teleosts are directed backward.

50 MARGARET H. COOK AND H. V. NEAL

Pharyngeal scales in Squalus are similar to, although not identi-
cal in structure with, the placoid scales of the outer skin. They
differ from them, however, in their origin. The placoid scales
of the outer skin and the teeth of the upper and lower jaws
derive their enamel layer from the embryonic ectoderm. Pha-
ryngeal scales, however, form their enamel layer from the endo-
derm. The reasons for this conclusion are identical with those
which convince us of the endodermal origin of the taste-buds.
The pharyngeal scales, however, arise later than the taste-buds.
No developed pharyngeal scales appear in the pharynx of a
‘pup’ stage in which the taste-buds are well developed and numer-
ous (fig. 28). Sections of such a stage, however, show the anlagen
of the pharyngeal scales (figs. 4 and 9). In such a section as
that shown in figure 4 it may be seen that the same layer of the
epidermis gives rise to the sense-cells of the taste-bud and to the
enamel layer of the pharyngeal scale. Entoderm, therefore, it
would appear, may give rise not only to sense organs, but to
scales such as usually are conceived as ectodermal, that is to say
cutaneous, In origin.

Against such a conclusion two chief objections may be urged:
first, that it is highly improbable, since it conflicts with the
principle of the specificity of the germ layers, and, second, that
the acceptance by morphologists of the annelid ancestry of
chordates justifies the assumption of an ectoderm-lined pharynx
in the latter comparable with the stomodaeum of annelids. It
is very doubtful, however, that such considerations will seem to
many morphologists to outweigh the direct evidence from onto-
genesis presented by Johnston and Landacre and in the present
paper. It is hardly necessary to suggest that a biological prin-
ciple is neither a self-evident truth nor a universal law, but a
generalization or hypothesis usually formulated before a com-
plete knowledge of the evidence. In the phenomena of budding
in colonial tunicates and in the regeneration processes of chordates
may be found exceptions to the principle of the specificity of
germ layers. That endoderm may produce taste-buds and
placoid scales is surely not more surprising than that muscle may
be regenerated from ectoderm or the lens of the eye from mesen-

ORIGIN—TASTE-BUDS OF ELASMOBRANCHS Dil:

chyma, as has been demonstrated (Morgan, ’01). In our con-
clusions concerning organic processes we have to reckon with the
fact of organic plasticity as well as with that of specificity of
tissue.

In the light of the considerable doubt that attaches to the
annelid hypothesis of chordate ancestry, it is perhaps sufficient
simply to suggest that such an hypothesis forms a most insecure
foundation for deductions concerning the derivation of the lining
of the chordate pharynx. The annelid hypothesis certainly seems
less convincing than it did formerly. Yet it seems as if in our
thinking we had tacitly assumed on the basis of that hypothesis
a considerable ectodermal invagination into the pharynx. Such
a prejudice is not supported by the ontogenetic evidence.

A brief summary of the arguments which have been presented
for and against the endodermal origin of taste-buds and pharyn-
geal scales may now be given. In favor of the ectodermal
derivation of these structures it may be urged that:

1. The resemblance of the histological structure of the pharyn-
geal epidermis to that of the mouth and skin—which are known
to be of ectodermal origin—suggests a similar derivation for
both. Furthermore, taste-buds and pharyngeal scales in Squalus
structurally resemble cutaneous organs of known ectodermal
origin. Such an argument, however, leads logically to the con-
clusion that the mucous lining of the esophagus is also ectodermal
—a conclusion contradicted by the ontogenetic evidence. Are
we to conclude, also, that because the epidermis of the gills of
fishes does not resemble that of the pharynx and skin it is there-
fore not ectodermal in origin?

2. Taste-buds occur in the skin of some fishes in regions where
their ectodermal origin seems indisputable (Herrick). It may
be urged that it is highly improbable that the same sort of
structure should arise independently from different germ layers.
This argument seems strengthened by the fact that associated
with the taste-buds in Squalus are placoid scales structurally
comparable with those derived from the outer skin. Is it pos-
sible to believe that the specificity of the germ layers is so slight
that identical structures may develop from both endoderm and
ectoderm?

52 MARGARET H. COOK AND H. V. NEAL

3. Since all other nervous receptor cells are ectodermal in
origin, the presumption is wholly in favor of the ectodermal
origin of the hair-cells of the taste-buds. This deduction, how-
ever, is based upon the assumption that the hair-cells of the
pharyngeal taste-buds are nervous receptors. If they are glan-
dular, as Botezat (’10) and Parker (’12) have assumed, the deduc-
tion is a logical non-sequitur.

4. That an ectodermal stomodaeal invagination occurs in
vertebrate embryos is an established fact. The persistence in
some animals of the connection of the ectodermal hypophysis
with the roof of the embryonic mouth makes it probable that
in these forms the ectodermal ingrowth extends as far poster-
iorly as the eustachian tubes. A relatively slight continuation
of this process of ingrowth would line the pharynx with ectoderm.
In the light of this positive evidence of extensive invagination
of the ectoderm, disagreement with the opinion of Keibel (’12,
p. 183), that “it is difficult to suppose that the ectoblast has
penetrated into the region of the larynx,’ might seem not
unreasonable.

5. The divergence in the conclusions of Kingsley aid Thyng
(05) and Johnston (’10) regarding the amount of the ectodermal
invagination in Amblystoma might appear to justify some doubt
as to the certainty of the conclusion reached by the latter.
Johnston seems not to have taken into consideration vine facts
presented in the former paper.

The following considerations, however, favor the wonelon
that the pharyngeal lining with its associated taste-buds and
placoid scales are derived from the entoderm:

1. The similarity of structure of the pharyngeal lining and
the skin by no means proves a similar genesis or derivation.
The similarity may be the result of convergence. Many instances
of the convergence of organic structures from dissimilar begin-
nings are known. The force of the doctrine of the specificity
of the germ layers has been greatly weakened by abundant
evidence of the plasticity of regenerating tissue.

2. The pharynx of all vertebrate embryos is lined primarily
with endoderm (figs. 5 to 27 of this paper). Direct evidence

ORIGIN—_TASTE-BUDS OF ELASMOBRANCHS 53

that by a process of substitution this endodermal lining is second-
arily replaced by ectoderm has never been given. The demon-
stration of an ectodermal dental ridge cannot be considered to
be a demonstration of an ectodermal lining of the pharynx.
It is equally fallacious to conclude that because annelids have
an ectodermal foregut vertebrates must have an ectodermal
pharynx. On the contrary, direct observation proves that the
lining of the pharynx of vertebrate embryos is entodermal in
origin.

3. “The presumption, from the standpoint of nerve distri-
bution, is all in favor of the origin of taste-buds from endoderm”’
(Johnston, 10, pp. 64, 65), since they are ‘‘innervated by the
facial nerve which is strictly a nerve related to entodermal sur-
faces in all vertebrates except those fishes in which the taste-
buds spread into the outer skin.”’

CONCLUSIONS

1. Taste-buds in Squalus acanthias are limited to the pharynx,
where they are distributed over the floor, the roof, and the gill
pouches.

2. The structure of the taste-buds resembles that of taste-
buds in other forms. They are groups of slender cells slightly
raised above the surface into a papilla. Each cell bears, exter-
nally, a hair-like process, and is connected internally with a nerve
ending.

3. Taste-buds in S. acanthias are derived from the endoderm.
They develop from the epithelial lining of the pharynx which at
all stages shows itself as endodermal, there being no indication
at any period of development of a migration inward of the ecto-
derm, except the slight invagination which forms the dental
ridge.

4. The pharyngeal scales arise in late embryonic stages. They
resemble placoid scales in structure, but are derived from the
endodermal lining of the pharynx.

54 MARGARET H. COOK AND H. V. NEAL

BIBLIOGRAPHY

Batrour, F. M. 1878 The development of elasmobranch fishes.

Bateson, W. 1890 The sense organs and perceptions of fishes, with remarks on
the supply of bait. Jour. Mar. Biol. Asso., London.

Cook, M. H. 1915 Are the taste-buds of elasmobranchs endodermal in origin?
Science, N. S., vol. 41.

Bornzat, E. 1910 Ueber Sinnesdriisenzellen und die Function von Sinnes-
apparaten. Anat. Anz., Bd. 37.

Go6prerT, E. 1906 Die Entwickelung des Mundes und der Mundhohle mit Drii-
sen und Zunge, u. s. w. Handb. Vergl. Exp. Entwickl. Wirbeltiere,
Bd. 2

GRABERG, J. 1898 Beitrige zur Genese Geschmacksorganes des Menschen.
Schwalbe’s Morph. Arbeit, Bd. 8.

Herrick, C. J. 1904 The organ and sense of taste in fishes. U.S. Fish Com-
mission Bull.
1908 On the phylogenetic differentiation of the organs of smell and
taste. Jour. Comp. Neur., vol. 18.

JOHNSTON, J. B. 1905 The cranial nerve components of Petromyzon. Morph.
Jahrb., Bd. 34.
1906 The nervous system of vertebrates. Philadelphia.
1909 The limits between ectoderm and entoderm in the mouth and the
origin of taste-buds. Anat. Rec., 1909..
1910 The limits between ectoderm and endoderm in the mouth and
the origin of taste-buds. I. Amphibians. Am. Jour. Anat., vol. 10.

KeiBeL, F. 1912 Development of the sense organs. Manual of Human Embry-
ology, vol. 2.

Kinestey, J. S., anD Tuyne, F. W. 1904 The hypophysis in Amblystoma.
Tufts College Studies, vol. 8.

Lanpacre, F. L. 1907 On the place of origin and method of distribution of
taste-buds in Ameiurus melas. Jour. Comp. Neur., vol. 17.

Morean, T. H. 1901 Regeneration. New York.

Naas, W. A. 1894 Vergleichend physiologtsche und anatomische Untersuch-
ungen ueber den Geruchs- und Geschmackssinn und ihre Organe, u.
s.w. Bibliotheca Zool. Stuttgart, Bd. 18.

Parker, G. H. 1908 ‘The sense of taste in fishes. Science, N. S., vol. 27.
1912 The relations of smell, taste, and the common chemical sense
in vertebrates. Jour. Acad. Nat. Sci. Phila., vol. 15.

ScHWALBE, G. 1883 Lehrbuch der Anat. der Sinnesorgane. Erlangen.

SHELDON, R. E. 1909 Reactions to chemical stimuli. Jour. Comp. Neur., vol.
19.

SmirH, P. E. 1914 Some features in the development of the central nervous
system of Desmognathus. Jour. Morph., vol. 25.

Strona, O. 8. 1898 Review of Johnston on the cranial nerves of the sturgeon.
Jour. Comp. Neur., vol. 8.

WeBER 1827 Ueber das Geschmacksorgan der Karpfen. Meckel’s Arch. f.
Anat.

PLATES

ABBREVIATIONS

Arc.vsc.1—-6, visceral arches 1-6
ar’ent., archenteron
brs.vsc.1—6, visceral pouches 1-6
cd., chorda dorsalis

cl.n’bl., neuroblastic cell

d.r., dental ridge

ec’drm., ectoderm

en’drm., entoderm

fil.br., branchial filaments
fis.vsc.1-6, visceral clefts 1-6
gl.th., thyroid gland

hyp., hypophysis

vfb., infundibulum

m., mouth

m.pl., mouth plate

mand., mandibular cartilage
n’v., nerve

olf., olfactory pit

ph., pharynx

pl.sc., placoid scale

sp., spiracle

st.germ., stratum germinativum
t.b., taste-bud

t.pap., taste papilla

tb.n., neural tube

vil.oes., oesophageal villi

55

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 1

PLATE 1

EXPLANATION OF FIGURES

Figures 1 to 10 were drawn with oil-immersion objective and reduced a little
over one-half in reproduction. They illustrate various stages in the develop-
ment of taste-buds, pharyngeal scales, and the formation of mouth and gill-slits.

1 An early stage in the development of a taste-bud and papilla as seen in a
section of a60-mm. Squalus embryo. Differentiation begins as a process of elon-
gation and aggregation of a group of cells in the stratum germinativum of the
pharyngeal epidermis which is of endodermal origin.

2 A section of an older taste papilla taken from a 90-mm. Squalus embryo,
showing a portion of the associated sensory nerve.

3 Asection of a fully differentiated taste papilla and ‘bud’ as seen in the ‘pup’
stage of Squalus.

4 A section of the pharyngeal lining of a ‘pup’ stage, showing to the left
the anlage of a pharyngeal scale and to the right a (somewhat damaged) taste
papilla.

5 A portion of a median sagittal section of a Squalus embryo (Balfour’s stage
K), showing the relations between ectoderm and endoderm in the region of the
degenerating buccal plate (‘preoral lobe’). The presence of large yolk granules
in the endodermal cells serves at this stage to distinguish them from the ecto-
dermal cells of the hypophysial invagination (above).

6,7, and8 Successive stages in the formation of a visceral cleft, showing the
ectodermal-endodermal relations in frontal sections of a Squalus embryo (stage
K). It is the endoderm which persists longest in the region of perforation of
the cleft. At this stage the ectoderm stains more intensely than the endoderm
so that their limits are easily distinguishable. The endoderm is the more active
layer of the two in the process of visceral cleft formation. :

9 A developing scale as seen in a section of an older ‘pup’ stage through the
pharyngeal epidermis.

10 Anenlarged portion of the upper jaw of a60-mm. Squalus embryo, showing
the region of transition between ectoderm:and endoderm. At neither this nor
any other stage it is possible to discover evidence of degeneration of the endo-
derm or of the rapid proliferation of ectoderm which would manifest itself if the
ectodermal ingrowth were to supplant the primary endodermal lining.

ORIGIN—TASTE-BUDS OF ELASMOBRANCHS PLATE 1
MARGARET H. COOK AND H. V. NEAL

PLATE 2
EXPLANATION OF FIGURES

Figures 11 to 17 are drawn from median sagittal sections of Squalus embryos
of different stages showing the limits and mutual relations of the ectoderm and
endoderm in the region of the mouth and pharynx. ‘The visceral pouches and
clefts were superimposed from in toto preparations. Endodermal structures are
cross-hatched and nervous system stippled in all drawings.

lland12 Median longitudinal sections of Squalus embryos in Balfour’s stages
G and H, showing the relations of the endoderm and ectoderm before the per-
foration of the buccal plate.

13 A median longitudinal section of stage I following the partial perforation
of the buccal plate. The deeply staining properties of the ectodermal cells and
the coarse yolk granules in the endodermal cells make easy the determination
of the boundaries between ectoderm and endoderm at this stage. Two visceral
clefts and two pouches formed.

14 In the region of the hypophysial invagination a remnant of the double-
layered buccal plate remains as the so-called preoral lobe. There are three vis-
ceral clefts and two pouches. Stage K.

15 Stage L. The hypophysis still retains in connection with the ectodermal
stomodaeum. The pharynx still retains its endodermal lining. The ectodermal
ingrowth shght.

16 A median longitudinal section of a 60-mm. embryo in which taste papillae
and buds are distinguishable in both floor and roof of the pharynx. The ecto-
dermal ingrowth does not extend farther than the limits of the upper and lower
jaws.

17 A more enlarged drawing to show the relations of ectoderm and endoderm
in the region of upper and lower jaws. The taste-buds are limited to the epi-
dermal surfaces formed from endoderm.

en
a)

PLATE 2

ORIGIN—TASTE-BUDS OF ELASMOBRANCHS

MARGARET H. COOK AND H. V. NEAL

PLATE 3
EXPLANATION OF FIGURES

Figures 18 to 27 are taken from cross-sections and frontal sections of Squalus
embryos, stages G to L, to show the relations of the ectoderm and endoderm
in the pharyngeal region. Subsequent to the perforation of the visceral clefts
the limits of the ectoderm and endoderm are difficult to determine and the dia-
grams are to be understood as only approximately accurate.

18 A cross-section of a Squalus embryo (stage G) through the first (spiracular)
pouch.

19 A cross-section through the same pouch (stage H). In the region of the
future cleft the ectoderm thins out more rapidly than does the endoderm.

20 A cross-section (stage I) through the first and second pouches.

21 A cross-section (stage K) through the first and second pouches and the
mouth which has broken through at this stage. .

22 A frontal (horizontal) section of stage H, showing the endodermal out-
pocketings to form the first, second, and third visceral pouches.

23 A frontal section of a Squalus embryo (stage I) cut dorsal to the hypophy-
sis, showing the formation of the first four visceral pouches.

24 A frontal section of a Squalus embryo (stage I) at the level of the hypophy-
sis, showing the formation of two visceral clefts and two pouches.

25 A frontal section of a Squalus embryo (stage K) dorsal to the level of the
hypophysis, showing the ectodermal-endodermal relations in the regon of the
pharynx.

26 A frontal section of a Squalus embryo (stage L) cut at the level of the
hypophysis, showing the exent of the pharyngeal epidermis.

27 A horizontal section of the same stage as figure 26, but cut at a higher level.

60

PLATE 3

ORIGIN—TASTE-BUDS OF ELASMOBRANCHS

NEAL

MARGARET H. COOK AND H. V.

TT
LL

Ks)
rs)
n
re
é
2
o

PLATE 4

EXPLANATION OF FIGURES

Figures 28 and 29 show the distribution of taste papillae and pharyngeal scales
in the region of the mouth and pharynx of Squalus acanthias.

28 The left half of a dogfish, ‘pup’ stage, showing the distribution of taste
papillae over the floor, roof, and sides of the pharynx.

29 The pharynx of an adult spiny dogfish laid open so as to show the distri-
bution of taste papillae and pharyngeal scales. The floor of the pharynx is
reflexed to the left in the figure.

ORIGIN—TASTE-BUDS OF ELASMOBRANCHS PLATE 4
MARGARET H. COOK AND H. V. NEAL

vil.oes!

63

Resumen por el autor, H. V. Neal,
Tufts College.

Nervios y plasmodesmos.

1. Antes del estado de 4.5 mm. no existen conexiones proto-
plismicas entre el tubo neural y los miotomos en los embriones
de Squalus acanthias. 2. La substancia neurofibrillar aparece
en las primeras conexiones protoplasmicas entre el tubo neural
y el miotomo. En estas conexiones primarias aparecen neuro-
fibrillas intensamente tenidas que pueden seguirse hasta los
neuroblastos bipolares situados en el tubo neural. La idea de
que las conexiones protoplasmicas primarias estan constituidas
por plasmodesmos no diferenciados est’ basada en métodos
neurologicos inadecuados. 3. En estados anteriores al estableci-
miento de conexiones entre el tubo neural y el miotomo, ciertas
células medulares, en zonas en las cuales mds tarde aparecen
los esbozos de los nervios, presentan tenidas con el método de
Bielchowsky-Paton un neuro-reticulo intensamente coloreado.
En estados un poco mis avanzados, cuando se establece la
conexidn neuromuscular se encuentran células neuro-reticulares
semejantes, unidas con neurofibrillas que se extienden en los
esbozos de los nervios del modo earacteristico de los neuroblastos
medulares tenidos con los métodos neurofibrillares especificos.
La evidencia sobre la presencia de una substancia neurofibrillar
semejante en todas las partes del esbozo del nervio confirma
la inferencia de que las conexiones neurofibrillares se establecen
no mediante células indiferentes, sino por neuroblastos medulares,
segtin mantienen los partidarios de la teoria de Bidder-Kupffer.
Las células indiferentes solamente participan en la formacion
de los esbozos nerviosos en los estados mis avanzados mediante
un proceso de emigracién desde el tubo neural.

Translation by José F. Nonidez
Cornell Medica! College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, FEBRUARY 28

NERVE AND PLASMODESMA

H. V. NEAL
Tufts College, Massachusetts

FIVE FIGURES

That neuraxones develop as processes of ganglion cells scarcely
admits of reasonable doubt in the light of the evidence now in
our possession. Few today would challenge the truth of Har-
rison’s (’13) assertion that ‘‘the work on the cultivation of tissues
may be said without reserve to have completely proved the
correctness of the conception of His and Ramon y Cajal.” The
long controversy between the adherents of the cell-process theory
and the supporters of the cell-chain hypothesis of neurogenesis
has been finally settled in favor of the former. To say this,
however, is by no means to assert that all problems of neuro-
genesis have been solved. Such has never been the claim of
Harrison or any other supporter of the Bidder-Kupffer hy-
pothesis. There yet remain unsolved a number of controverted
questions of great interest, some physiological and some morpho-
logical, which neither tissue cultures in vitro nor observations
upon sectioned material have been able to solve. Familiar
examples of such are the problems of the genesis of the nerve
sheaths and of the sympathetic cells. Further questions are
suggested by the phenomena of nerve regeneration (Boeke, ’16).

One of the disputed points in the histogenesis of nerve is
whether or not there exists previous to nervous (neurofibrillar)
connection between neural tube and myotome a connection by
means of undifferentiated protoplasmic threads, or plasmodes-
mata, or ‘fasernetz’ of Szily (04). Among those who assume
the existence of such plasmodesmata it is a matter of dispute
whether the plasmodesmata are primary—the result of incom-
plete cell division, as first stated by Hensen (’64)—or secondary
the result of protoplasmic outflow of medullary cells as first

65

66 H. V. NEAL

stated by Bidder and Kupffer (57). Furthermore, among those
who hold that neuromuscular connections are secondary there
is disagreement as to whether such connections are effected by
indifferent (glia or neurilemma) cells, as suggested by the Hert-
wigs (’78) and recently maintained by Held (’09), or by the out-
growth of the neuraxone processes of medullary neuroblasts,
as believed by advocates of the Bidder-Kupffer hypothesis.
Not until these questions are adequately answered is it likely
that the investigation of neurogenesis will cease.

The invention in recent years of specific neurofibrillar stains
and the demonstration of neurofibrillae in the earlier stages of
neurogenesis have led to the general adoption of the aphorism
of Apathy that “there is no nerve without neurofibrillae”’ as a
criterion of nervous structure. How greatly needed such a
criterion has been is only too well known to those familiar with
the literature of neurogenesis. In the absence of such a criterion
of nervous structure, it has been hitherto possible to find in any
cellular strand extending to or from the nervous system the
evidence of ‘primitive nerves’ wherever they were demanded for
schemes of ancestral metamerism. However, notwithstanding
the application of improved methods of demonstrating neuro-
fibrillae in embryonic nervous tissue, it has been found impossible
up to the present to demonstrate the presence of neurofibrillae
in the primary connections between nerve and muscle, that is
to say, between neural tube and myotome. The existence of
such neurofibrillae in the plasmodesmata of Squalus embryos
is asserted in this paper for the first time in vertebrate embryos.!

The present paper raises three controverted problems in nerve
histogenesis:

1. Are the connections between nerve and muscle primary
or secondary?

2. Are neuromuscular connections primarily undifferentiated
plasmodesmata, as asserted by Paton (’07) and Held (’09), or
are they primarily neurofibrillar?

1A preliminary report of the conclusions reached in this paper was made
before the American Society of Zoologists at Philadelphia, December 29, 1914, and
an abstract reprinted in Science, 1915, vol. 41, pp. 485-486. War work has delayed
the appearance of the final paper.

NERVE AND PLASMODESMA 67

3. Are neuromuscular connections effected by indifferent—
neurilemma or glia or mesenchyma—cells or by medullary
neuroblasts? All three questions have been discussed at length
in papers by Paton (’07), Held (’09), and the writer (14), in
which the voluminous literature has been reviewed. ‘The reason
- for raising again the three questions stated above is that, while
none of the investigators up to the present has been able to
demonstrate neurofibrillae in the primary protoplasmic con-
nections between the neural tube and the myotome, the writer
has been able to discover them in preparations made by the
Bielschowsky-Paton process. The evidence is presented in the
five figures of this paper. The divergence in the conclusions
reached by Paton and Held on the one hand and by the writer
on the other are essentially the result of the capriciousness of
special neurofibrillar stains as applied to the earlier stages of
nerve histogenesis. Not until after some years of experimen-
tation with the Bielschowsky method and not until after the
publication of the 1914 paper on the morphology of the eye-
muscle nerves did the writer succeed in staining the neuro-
fibrillae in the plasmodesmata. The present paper is therefore
to be considered as a supplement to the one published in 1914.

Turning, now, to the first of the questions raised above, the
writer can only reiterate the assertion made in earlier papers
that in embryos of Squalus acanthias of stages previous to 4.5
mm. there is not the slightest evidence of protoplasmic con-
nection between neural tube and myotome. Figures 1 and 2
of this paper reproduce faithfully the relations which obtain
previous to the appearance of definitive nervous connection.
These figures represent sections cut transversely- through the
middle of a myotome where in the later stages the anlagen of the
motor nerves make their appearance. In some preparations it
is possible to demonstrate in the plasma-filled space between
neural tube and myotome a minimal amount of vacuolated
coagulable material, invisible in most preparations made by the
usual methods of fixation and staining, and lacking the staining
properties of cell protoplasm. The ‘Fasernetz’ of Szily (04)
and the plasmodesmata of Paton (’07) and of Held (’09) make

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 1

68 H. V. NEAL

their appearance later in the manner described below. Proto-
plasmic connection between tube and myotome in Squalus is
secondary and not primary.

The failure of Paton, Held, and the writer to discover the
presence of neurofibrillae in the first protoplasmic connections
between neural tube and myotome led all three to assert that
neurofibrillas are secondarily differentiated within non-nervous,
undifferentiated ‘plasmodesmata.’ The divergent views as to
the details of this process have been summarized by the writer
(14, pp. 35-41) and need not be repeated here. In that paper
the writer asserted that “medullary cells by a process of out-
growth, form the first protoplasmic connection between tube
and myotome” and that later within these processes the neuro-
fibrillae make their appearance. Bielschowsky-Paton prepara-
tions also showed the presence of a neuroreticulum within bipolar
neuroblasts lying in the somatic column of the spinal cord and
in the zones opposite the middle of the somites where later the
plasmodesmata make their appearance. Similar neuroblasts
containing a deeply staining reticulum are shown in figures 1
and 2 of this paper. But at the time the 1914 paper was pub-
lished no neurofibrillar structures were seen in the plasmodes-
matous connections at the time of their first appearance. Since
that time, however, they have been seen in a number of series
prepared by the Bielschowsky-Paton method. Figures 3 to 5
of this paper show the presence of neurofibrillar substance within
the ‘plasmodesmata’ of Paton and Held. The justification for
calling the deeply staining material of the plasmodesmata
‘neurofibrillar’ consists in the fact that it stains precisely like
the neurofibrillar network present within the neuroblasts which
by their outgrowth form the plasmodesmata and in the further
fact that it is possible to trace the neurofibrillar network of the
neuroblastic cells from the tube into the plasmodesmata as is
seen in the figures mentioned. Its granular appearance within
the plasmodesmata may be interpreted as the result of cutting
a fibrillar network transversely. In those instances where the
neuraxone processes of the medullary neuroblasts are cut length-
wise, as in figures 3 and 4, the neurofibrillar substance appears

NERVE AND PLASMODESMA 69

as heavy ‘fibrils’ as described by Paton. The affinity for neuro-
fibrillar stains is especially marked at the lower, advancing
extremity of the nerve anlage where Paton (’07) found the first
indications of neurofibrillar substance in his preparations. From
this evidence the conclusion seems warranted that the primary
connections between neural tube and myotome are truly nervous,
just as the experiments of Paton (’07) upon the reactions of
elasmobranch embryos at the time the plasmodesmata are formed
would lead us to infer.

The answer to the third question is already implied in the fore-
going discussion. As has already been stated, in stages before
protoplasmic connection between tube and myotome is effected
certain medullary cells in zones where later nerve anlagen make
their appearance show in Bielschowsky-Paton preparations a
deeply staining neuroreticulum. Such cells show a pronounced
polarity and tend to assume a spindle shape. The staining
material of the reticulum is peripheral in position and extends
throughout the entire length of the cell, showing a tendency to
become more densely aggregated at the outer extremity of the
cell. Not infrequently the end of the cell appears to extend
along the inner surface of the external limiting membrane of the
tube just as if the neuroblastic cell were in the process of elonga-
tion in the direction of its long axis and the movement of proto-
plasm were deflected in direction by the resistance offered by
the limiting membrane of the medullary wall (fig. 1). The
neuroreticulum is densely aggregated in the elongated peripheral
portion of some cells so as to appear in such as a heavy deeply
stained neurofibril. 3

In slightly later stages, when protoplasmic connection with
. the myotome has been established in the manner described in
detail by the writer in earlier papers (’14), similar neuroblastic
cells with deeply staining processes are seen to extend into the
nerve anlagen (figs. 3 to 5), that is to say into the so-called
plasmodesmata. That these plasmodesmata are primarily
formed by the protoplasmic outflow of similar neuroblastic
cells has already been asserted by the writer (14. Note espe-
cially figs. 4 to 7, pls. 1 and 2). The plasmodesmata, in other

70 H. V. NEAL

words, are not ‘paths’ utilized by the neuraxones as a means of |
conduction and material for growth, as assumed by Held (’09),
but are ab initio nervous. The participation of indifferent
(neurilemma or glia) cells in the formation of the motor nerve
anlagen in elasmobranchs by means of the migration of medullary
cells along the nerve anlagen (‘plasmodesmata’) has been de-
scribed by many, including the writer (’03—’14), but it is not such
cells which form the primary plasmodesmata. Admitting the
possibility that in the plasmodesmatous connection shown in
figures 3 to 5 of this paper processes of indifferent, non-neuro-
blastic cells occur, nevertheless the important fact remains that
the primary neuromuscular connections were not formed by
these indifferent, non-nervous elements. Their relations to the
nerve anlagen are secondary and not primary.

In the light of the evidence that neuromuscular connections
are ab initio nervous, there seems to be little justification for the
application of the term ‘plasmodesmata’ to them. What we
are actually dealing with in the case of such structures are nerve
anlagen, or neuraxones and their sheaths. Why then term them
plasmodesmata, thereby suggesting that they are merely non-
nervous, undifferentiated intercellular connections?

SUMMARY

1. Previous to the stage of 4.5 mm. there are no protoplasmic
connections between tube and myotomes in Squalus embryos.

2. Neurofibrillar substance is present in the primary connection
between tube and myotome. Within the plasmodesmata of
Paton (’07) and Held (’09) may be found in adequately stained
Bielschowsky-Paton preparations deeply stained neurofibrils
which may be traced to bipolar neuroblasts within the neural
tube. The assertion that the primary connection between
muscle and nerve is non-nervous, i.e., non-neurofibrillar, is based
upon incompletely stained preparations.

3. In stages before protoplasmic connection between tube and
myotome is effected certain medullary cells in zones where later
the nerve anlagen (‘plasodesmata’) make their appearance
show in successfully stained Bielschowsky-Paton preparations

NERVE AND PLASMODESMA tl

a deeply stained neuroreticulum. Similar cells containing a
neuroreticulum are the first to become connected by protoplasmic
outflow with the adjacent myotome. In all later stages of
neurogenesis similar neuroreticular cells within the neural tube
are connected by neuraxone processes with the nerve anlage
and the myotome. Glia or neurilemma cells participate only
secondarily in the formation of neuromuscular connections. The
assumption of the Hertwigs (’78), later supported by Paton
(07) and Held (09), that neuromuscular connections are pri-
marily effected by indifferent cells and that the ‘plasmodesmata’
thus formed are utilized as ‘paths’ by the growing neuraxones,
does not accord with the observed facts. The term ‘plasmo-
desma’ should therefore be discarded as unnecessary and mis-
leading.

LITERATURE CITED

ApAruy, 8. 1907 Bemerkungen zu den Ergebnissen Ramén y Cajals hinsicht-
lich der feineren Beschaffenheit des Nervensystems. Anat. Anz., Bd.
30.

BippeEr, F., aNnD Kuprrer, C. 1857 Untersuchungen iiber das Riickenmark.
Leipzig.

Boeke, J. 1916 Die Regeneration der motorischen Nervenelemente und die
Regeneration der Nerven der Muskelspindeln. Verhandelingen der
Koninklijke Akademie van Wetenschappen te Amsterdam, Deel 18.

CasaL, Ramon y, 8. 1907 Nouvelles observations sur l’evolution des neuro-
blasts avec quelques remarques sur l’hypothese neurogenique de Hen-
sen-Held. Trav. d. lab. de rech. biol., Bd. 5. Also1908. Anat. Anz.,
Bd. 22.

Dourn, A. 1888 Studien, u.s. w. 14. Ueber die erste Anlage und Entwicklung
der motorischen Riickenmarksnerven bei den Selachiern. Mitt. Zool.
Stat. Neapel, Bd. 8.

Frorisp, A. 1901 Ueber die Ganglienleisten des Kopfes und des Rumpfes und
ihre Kreuzung in der Occipitalregion. Arch. f. Anat. u. Physiol.,
Anat. Abth.

Harrison, R.G. 1913 The life of tissues outside the organism from the embryo-
logical standpoint. Trans. Cong. Am. Phys. and Surgeons, vol. 9.

Heitp, H. 1909 Die Entwicklung des Nervengewebes bei den Wirbeltieren.
Leipzig.

HENRIKSEN, P. B. 1913 Nye Unders kelsen over Nervenregeneration. Kris-
tiania. Stunske Bogtrykkeri.

HewnseNn, V. 1864 Zur Entwickelung des Nervensystems. Arch. f. path. Anat.
u. Physiol., Bd. 30.

Hertwic, O. unp R. 1878 Das Nervensystem und die Sinnesorgane der Medu-
sen, Monographisch dargestellt. Leipzig.

fe H, V. NEAL

His, W. 1890 Histogenese und Zusammenhang der Nervenelemente. Arch.
Anat. u. Physiol., Anat. Abth., Suppl.

Lewis, W. H. 1906 Experimental evidence in support of the outgrowth theory
of the axis cylinder. Proc. Am. Asso. Anat., Ann Arbor, 1905. Am.
Jour. Anat., vol. 5.

Neat, H. V. 1903 The development of ventral nerves in Selachii. I. Spinal
ventral nerves. Mark Anniv. Vol.
1914 The morphology of the eye muscle nerves. Jour. Morph., vol.
25.

Paton, 8. 1907 The reactions of the vertebrate embryo to stimulation and the
associated changes in the nervous system. Mitt. Zool. Stat. Neapel,
Bd. 18.

Szrty, A. 1904 Zur Glaskérperfrage. Anat. Anz., Bd. 24.

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PLATE 1
EXPLANATION OF FIGURES

All of the figures of this plate were drawn with Abbé camera, one-twelfth homo-
geneous oil-immersion objective and no. 6 compensation ocular of Zeiss. In
reproduction the mangification has been reduced by one-half. The series illus-
trates stages immediately preceding and following the appearance of the so-called
plasmodesmatous connections between neural tube and myotomes in the trunk
region of Squalus embryos. In reality such ‘plasmodesmata’ are the anlagen of
somatic motor nerves as demonstrated by their genesis, neurofibrillar structure
(when adequately stained) and later histogenesis. As is well known to all stu-
dents of elasmobranch embryos, there is no difficulty whatever in the determina-
tion of stages as earlier and later, since such successive stages appear in succes-
sive metameres of each embryo. In all metameres the somatic nerve anlagen
make their appearance opposite the middle of the myotome. The embryos from
which the drawings were made were prepared by the Bielschowsky-Paton method
to demonstrate neurofibrillar structures.

1 and 2 Portions of cross-sections of a 7-mm. Squalus embryo cut in the
middle of metameres lying near the cloacal region. In the metameres immedi-
ately anterior to the ones shown protoplasmic connection between tube and myo-
tome has already been established by the migration of medullary protoplasm.
In more posterior segments there is no protoplasmic connection whatever. The
sections demonstrate the presence in the medullary wall of neuroblastic cells
(cl.n’bl) containing a deeply staining neuroreticulum extending throughout the
entire length of the cell. Such cells, as stated by the writer in an earlier paper
(14), are the first to effect neuromuscular connections by means of protoplasmic
outflow. Their nervous (neuroblastic) character is attested by their polarity
and neuroreticular structure even before their neuraxone process extends outside
the limiting membrane of the neural tube. They contrast in form, color, and
structure with the adjacent cells of the medullary wall. In successful Bielschow-
sky-Paton preparations the neuroreticulum is stained a deep blue-black, while
the cytoplasm of adjacent cells is reddish-yellow in color.

3, 4 and 5 Portions of cross-sections of Squalus embryos treated by the
Bielschowsky-Paton method in metameres in which neuromuscular connections
(‘plasmodesmata’) are already established. The nervous nature of the so-called
plasmodesmata is evidenced by the presence of neurofibrillar material within
them and the extension of neuraxone processes of neuroblastie cells into the
‘plasmodesmata.’ Such evidence shows that instead of being undifferentiated,
non-nervous bridges of protoplasm, into which or along which neurofibrils or
neuraxones grow in order to effect neuromuscular connection, the ‘plasmodes-
mata’ in reality are ab initio nervous structures. Non-nervous material is
secondarily added to the nerve anlage by migration or outflow from the neural
tube.

ABBREVIATIONS

ax,, neuraxone process new’ret., neuroreticulum
cd., chorda dorsalis _ pl’sm., plasma

cl.n’bl., neuroblast cell scl., sclerotome
ms’ench., mesenchyma so., somite

mb.cl., cell membrane tb.n., neural tube
mb.lim., limiting membrane vac., vacuole

my., myotome
74

NERVE AND PLASMODESMA PLATE
H. V. NEAL

foe--Ms’ench.

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Resumen por los autores, 8. E. Johnson y M. L. Mason,
Northwestern University Medical School, Chicago.

El primer ramo comunicante blanco del t6rax del hombre.

En los libros de texto de Anatomia humana y en la literatura
corriente se encuentran muchas afirmaciones incompletas y
contradictorias sobre la presencia de un ramo comunicante blanco
en relacion con el primer nervio espinal tordcico. Los autores
del presente trabajo, al coleccionar material fresco en autopsias
con el propésito de llevar a cabo un estudio general de la estruc-
tura de los tronecos del simpdtico en el hombre, han encontrado
en todos los sujetos uno 0 mais ramos comunicantes blaneos que
reunen el primer nervio espinal tordcico con el primer ganglio
tordcico o ganglio estelar.

Una diseccion cuidadosa de doce sujetos did los mismos
resultados (tinendo y seccionando los ramos, los autores han
podido en la mayor parte de los casos distinguir entre los ramos
grises y los blancos). Los ejemplares frescos fueron tefidos con
Acido 6smico. En un ramo blanco han podido contar un total
de 1252 fibras meduladas, que varian entre 1.9 a 15.9 micras
de diimetro, la mayor parte de ellas oscilando entre 2 y 4 micras.
Teniendo en cuenta el tamano relativo de las fibras, estiman
que por lo menos el 16 por ciento de las fibras meduladas en
este ramo eran de caracter aferente.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHORS’ ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, FEBRUARY 28

THE FIRST THORACIC WHITE RAMUS COMMUNICANS
IN MAN

S. E. JOHNSON anp M. L. MASON

Anatomical Laboratory of Northwestern University Medical School!

FIVE FIGURES

In text-books of human anatomy and in current literature
there is evident lack of agreement and uncertainty in the various
statements regarding the occurrence of a white ramus com-
municans in connection with the first thoracic spinal nerve.
In Cunningham’s text-book of human anatomy (’18) we read
that ‘“‘Each thoracic nerve, with the probable exception of the
first, sends a visceral branch (white ramus communicans) to
join the ganglhated trunk in the thorax.’ Piersol (’18) says,
“from the first or second thoracic to the second or third lumbar.”’
In Lewis’ edition of Gray’s Anatomy (18) is this statement:
“Two rami communicantes, a white and a gray, connect each
ganglion with its corresponding spinal nerve.” According to
Morris-Jackson (714), ‘““Each ganglion, with the possible exception
of the first, receives a white ramus communicans from a thoracic
nerve. . . . . The statement as worded does not commit
the author as to the source of the white ramus which runs to the
first thoracic sympathetic ganglion. It might come from the
first, second, or third thoracic nerve, or even from the eighth
cervical. In Poirier, Charpy et Cunéo (’08) we find the state-
ment that the thoracic sympathetic ganglia are connected to
the thoracic nerves by one or two rami communicantes. This
statement is rather typical of the ones found in foreign texts
generally. There is seemingly a desire to avoid statements
warranting specific interpretation as to the nature of the rami
which are described. It is quite obvious that the mere number
of rami implies nothing as to their actual nerve constituents.

1 Contribution no. 81, January 1, 1921.

ae

78 S. E. JOHNSON AND M. L. MASON

As further examples of this non-commital sort of statement,
we may quote from Chiarugi (’10—’17), “each ganglion is con-
nected by one or two rami communicantes with a thoracic nerve.”
Similarly in Rauber-Kopsch (’07) no definite statement as to
the upward extent of the white rami can be found. Spalteholz
(20) in four separate statements does not attempt to distinguish
between the white and gray rami of the thoracic nerves. ‘The
following quotation is typical: ‘“‘Jedes Ganglion (sympathicus)
ist durch ein oder mehrere Rami communicantes mit den Nn.
thoracales verbunden”’ (p. 791).

Literature consulted shows that little has been done to deter-
mine the nature of the rami communicantes in man, conclusions
derived from other mammalian work being read directly into
human anatomy. Langley (’94, p. 235) says, “The uppermost
white ramus in man should come from the first thoracic, 1.e.,
the first thoracic is probably the highest spinal nerve by which
motor and sensory fibers run to the viscera. But it seems not
unlikely that a posterior arrangement of the brachial plexus
occurs sometimes in man in which the first thoracic has not a
white ramus.” Langley at another time (’00) states that there
is more or less evidence to substantiate the statement that the
first thoracic nerve is in all mammals the first to give off a white
ramus. And again (’03) he states that afferent fibers which
accompany the sympathetic arise from the nerves which give
off efferent fibers and from these only, in man they (efferents)
arise from the first thoracic to the second or third lumbar.

In R. L. Miiller’s paper (’09) we could find no definite state-
ment as to the superior limit of the white rami. His claim that
in all mammals without exception, the inferior cervical ganglion
is fused with the first one or two thoracic to form the ganglion
stellatum, we are unable to verify as regards man.

In collecting fresh autopsy material for the purpose of making
a survey of the structure of the sympathetic trunks in man,
the authors found in all cases examined at least two rami com-
municantes arising from the first thoracic nerve. The number
varied from two to five in different subjects and often the number
was not the same on opposite sides of the same subject. These

THORACIC WHITE RAMUS COMMUNICANS IN MAN 79

rami did not always run to the first thoracic sympathetic ganglion.
They were not infrequently distributed to the inferior cervical
(when separate), the second, and even the third, thoracic ganglion.
As a rule the autopsy material had to be removed rather
hurriedly, and for this reason it was thought advisable to supple-
ment these findings with careful dissections of laboratory speci-
mens. The first thoracic nerve and its associated rami com-
municantes were dissected out on both sides of twelve cadavers.
The number of rami vaiied from two to five and was frequently
unequal on opposite sides. Examples of the various conditions
found are shown in the accompanying photograph, figures 1 to
4. The specimens illustrated were taken from opposite sides
of four cadavers. Figure 1, 7, shows a section of the right sym-
pathetic trunk with the inferior cervical and first thoracic ganglia
appearing as distinct enlargements. ‘The internodal segment
joining the ganglia, however, was short and thick, and probably
contained ganglion cells throughout its length. The eighth
cervical spinal nerve (C. 8.) is seen to be connected with
the inferior cervical ganglion by two rami, medial to its
junction with the first thoracic nerve (7.1.) to form the lower
primary trunk of the brachial plexus. Three rami connect the
thoracic nerve with the first thoracic sympathetic ganglion.
Figure 1, /, from the left side of the same cadaver, shows a more
complicated arrangement of the rami. Two run from the
eighth cervical nerve to the inferior cervical ganglion, one joins
the two nerve trunks, one runs from the first thoracic nerve to
the inferior cervical ganglion, and a fifth runs from the first
thoracic nerve to the first thoracic sympathetic ganglion. In
the remaining specimens shown in the accompanying photo-
graph the inferior cervical and first thoracic sympathetic ganglia
are united to form more or less symmetrical enlargements of the
sympathetic trunks. Each of these ganglionic masses is con-
nected with its related first thoracic spinal nerve by short nerve
strands which vary a great deal in size as well as in number.
Naturally, the presence of two or more rami does not mean
anything unless the nature of the fibers which they contain can
be determined. Fresh material was stained in osmic acid, and

80 Ss. E. JOHNSON AND M. L. MASON

Figs. 1, 2,3, and 4 Photograph of right (r) and left (1) dissections from four
cadavers in the anatomical laboratory. In each dissection there are shown short
stretches of spinal nerve and of sympathetic trunk with the associated rami com-
municantes. C.8., eighth cervical spinal nerve; G., gray ramus communicans;
Gn.crv.inf., inferior cervical ganglion; Gn.7.J., first thoracic sympathetic gan-
glion; l., left; L.7., junction of eighth cervical and first thoracic spinal nerves
to form lower primary trunk of brachial plexus; /., ramus communicans (mixed)
which contains large proportions of both medullated and non-medullated fibers;
r., right side; 7'’.J., first thoracic spinal nerve; Tr.sym., sympathetic trunk.
Photograph blocked out, but not retouched. Reduced 1/4.

THORACIC WHITE RAMUS COMMUNICANS IN MAN 81

in figure 5 we show a camera-lucida sketch (cross-section) of a
first thoracic white ramus communicans which is typical of our
series. In the subject from which this section was taken, two
well-defined rami ran from the first thoracic nerve to the first

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Fig. 5 Transverse section of a first thoracic white ramus communicans.
This ramus ran from the first thoracic spinal nerve to the first thoracic sympa-
thetic ganglion. It was removed and fixed in osmic acid two hours after death.
Note the separate fasicle of large medullated fibers at the upper margin of the
drawing. Camera sketch. X 227.

thoracic ganglion. The latter was connected with the inferior
cervical ganglion by a short constricted portion of sympathetic
trunk which could hardly be considered an internodal segment.
The two rami were of unequal size, the smaller white ramus
measuring approximately 0.6 mm. in diameter, and the gray
ramus about 1.5 mm. The gray ramus was composed almost

82 S. E. JOHNSON AND M. L. MASON

wholly of non-medullated fibers, there appearing only about 150
small medullated fibers, scattered irregularly over the field of
a cross-section.

In a cross-section of the white ramus we counted a total of
1252 medullated fibers of various sizes from approximately 1.54
up to 16.94. The results of a differential count may be tabulated
as follows:

Ac OpOT LESS erie ee one a ees rr Ee ES Oe ete ae 422
DIPS Psa iON PCa es | aa OR RE ane en oe Coe ee WO RAS ORT Ne aii an tt Mewes 506
SLU TAM KO} URS G) Pie Seeuehe es pee nen tee Pee Reh gh Rr ORG arab Ae Ame Sete tp 151
DOM ALO EO Oiarg. pote see ob oece Br tpey Sen e hk Sri PEA By VATE OO REE Pea as 40
ca! ADAGE N27 err ysan cscaey A UET SUS BGM nS ea Reve nT tee ee ea 74
DAG OMI Bis 3. deotrarte eee cusee so POT Oe oO EL EO ee eae 35
Joey HO) IRON G6 oa dacoes Ps ike Ie et is ii MEIN Grn any ciomnts Cito e 10
IS TPA STO) HD UT esr eee ee eS AN) 4 Pe eRe Erneta inet Sete EEE I 13
Dalit OWN: Deco aero Susy aie) coeae See ugh oS eee tceeae FO A eRe ee sR il

OLAS, Joes ahi bs acts Soe ee reeds fre RO OC CR ne 1252

In the above tabulation of fibers the feature that stands out
most prominently and unexpectedly is the presence of such a
relatively large number of sensory fibers. It will be noted that
there is an abrupt diminution of fibers of sizes between 5.6 and
7.du. The fibers of this size and under are undoubtedly of two
types (Ranson and Billingsley, ’18), visceral efferent (the major-
ity) and small afferent fibers. These cannot at present be differ-
entiated microscopically. The remaining 193 fibers range from
5.6 to 16.94, and probably all of these are afferent fibers. Two
hundred (approximately 16 per cent) would be a low estimate
for the total number of afferent fibers in the ramus. As shown
in figure 5, many of the large medullated fibers are grouped in
definite fascicles. It will be a matter of the greatest interest
to trace out the course and distribution of these well-defined
bundles of large medullated fibers.

The rami of the laboratory specimens shown in figures 1 to 4
were run through various fixing solutions, stained in iron haema-
toxylin, and sectioned, with the hope that we might be able to
identify the white rami. This we found to be quite possible in
the majority of instances. The nature of the rami is indicated

THORACIC WHITE RAMUS COMMUNICANS IN MAN 83

in the figures (figs. 1 to 4). Rami which were undoubtedly white
are marked with the letter w. Others are gray (g); mixed (m),
or unidentified as to the nature of contained fibers. In all of
the figures with the exception of figure 1, r, the first thoracic
nerve gives off one or more white rami which in most instances
connect with the stellate ganglion. In.the case of the specimen
shown in figure 1, 7, the nature of all of the rami could not be
determined. °

To summarize, we have found, in each of twelve laboratory
bodies, as’ well as in fresh autopsy subjects, one or more white
rami communicantes arising from the first thoracic spinal nerve
and connecting, as a rule, with the stellate ganglion. These
rami contain a relatively high percentage of large medullated
fibers. We have not found the inferior cervical and first thoracic
sympathetic ganglia invariably fused, as they are stated to be
by Miller, although this was the usual condition. Of the
numerous rami which we have seen connecting the eighth cervical
nerve with the inferior cervical or the stellate ganglion, none
have been shown to be either white or mixed in character. How-
ever, it does not seem impossible that in an occasional subject
the highest white ramus may arise from the eighth cervical nerve
or, with a posterior arrangement of the branchial plexus, as
Langley has suggested, from the second thoracic nerve. While
admitting these possibilities, we feel justified in concluding that
the normal condition in man is that which we have described
above.

84 Ss. E. JOHNSON AND M. L. MASON

BIBLIOGRAPHY

Curaruci 1910-1917 Istituzioni di Anatomia Dell "Uomo. Societi Editrice
Libraria, Milano.

CunniINGHAM’s Textbook of human anatomy, 5th edition, edited by ARTHUR
Rosinson, 1918. Wm. Wood & Co., New York.

Gray’s Anatomy of the human body, 20th edition, edited by Warren H. Lewis,
1918. Lea & Febiger, Philadelphia.

Lane ey, J. N. 1894 The arrangement of the sympathetic nervous system,
based chiefly upon observations upon pilo-motor nerves. Jour. Phys-
iol., vol. 15, p. 176.

1900 Article on the sympathetic nervous system in Schaeffer’s text-
book of physiology.
1903 The autonomic nervous system. Brain, vol. 26.

Morets’ Human anatomy, 5th edition, edited by C. M. Jackson, 1914. P.
Blakiston’s Son & Co., Philadelphia.

Miter, L. R. 1909 Studien itiber die Anatomie und Histologie des sympa-
thischen Grenzstranges, insbesondere iiber seine Beziehungen zu dem
spinalen Nervensysteme. Deutscher Kongress fiir innere Medizin,
Wiesbaden, S. 658.

Prersou’s Human anatomy, 6th edition, 1918. J. B. Lippincott Co., Philadel-

phia.

PorripR, CHARPY ET Cunfo 1908 Abrégé d’Anatomie, T.2. Maisson et Cie.,
Paris.

QuaIn’s Elements of anatomy, 11th edition, 1908. Longmans, Green & Co.,
London.

Ranson, 8S. W., AND Biuuinestey, P. R. 1918 The thoracic truncus sym-
pathicus, ete. Jour. Comp. Neur., vol. 29, p. 419.

RavusBer-KopscH 1907 Lehrbuch d. Anat. d. Mensch., 7th edition, part 5.
Theime, Leipzig.

SPALTEHOLZ, WERNER 1920 Handatlas der Anatomie des Menschen, 9th edi-
tion. S. Hirzel, Leipzig.

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86 SYDNEY E. JOHNSON

ganglia, various possibilities have been suggested. These have
been considered by Langley (’03), and his observations have
been often referred to. Langley has produced much evidence
against the existence of such connecting neurons. His results
and conclusions which have been reviewed recently by Ranson
and Billingsley appear to be supported and confirmed by an
increasing amount of purely anatomical investigation. If com-
missural neurons are present their terminations would be expected
to form some part of the intercellular plexus of the sympathetic
ganglia, but upon degenerative section of the cervical trunk,
Ranson and Billingsley (’18 a) found that the fine axonic rami- —
fications of the intercellular plexus completely disappeared,
indicating that it is derived wholly from ascending preganglionic
fibers. Two distinct functional types of autonomic cells such
as might be anticipated from Dogiel’s report could not be demon-
strated by Carpenter and Conel (’14) on the basis of cell structure.
The writer (18) has produced experimental evidence against the
occurrence of commissural neurons in the sympathetic ganglia
of the frog.

Suggesting the occurrence of commissural neurons are obser-
vations of a rather fragmentary character which, however, must
be considered. The more important of these observations have
been reported by v. Lenhossék, Dogiel, and Huber.

Lenhossék’s (’94) report is substantially to the effect that he
observed, in Golgi preparations of fourteen-day chick embryos,
sympathetic fibers entering a sympathetic ganglion from the
periphery and there terminating in simple end-brushes. The
fibrillae of these end-brushes are said often to terminate on
autonomic cells in small end bulbs.

Observations of a somewhat similar nature are recorded by
Huber, who, in his 1897 ‘‘Lectures on the Sympathetic Nervous
System” wrote that ‘ain methylen-blue preparations of the
ganglia of the chain taken from mammalia and birds, I have
often observed a free ending of branches of non-medullated
nerve fibers in sympathetic ganglia.”’ These endings are different
from those described by v. Lenhossék, inasmuch as they are on
the dendrites of sympathetic neurons rather than on the cell

SACRAL SYMPATHETIC TRUNK OF CAT 87

bodies. Huber states further that the fibers thus ending did
not seem to enter the ganglion from the periphery, i.e., did not
seem to be terminations of neuraxes whose cell bodies were
situated distal to the ganglion under observation. Huber’s later
observations (’99, ’13) do not appear to alter his position with
reference to these fibers.

More far-reaching are the observations of Dogiel, who in
1896 described two types of sympathetic cells, 1) the commonly
recognized postganglionic motor neuron with cell-body located
in a sympathetic ganglion and, 2) another type of cell with
usually a larger cell body and much longer dendrites. The
dendrites branch less and were often traced beyond the limits of
the ganglion. The neuraxes of these cells were sometimes
traced into another ganglion, where they were seen to divide
repeatedly and take part in the formation of the intercellular
plexus. Dogiel expresses the belief that these cells are sympa-
thetic and of sensory function. He refers further to the proba-
bility that the pericellular baskets about spinal ganglion cells of
type II take their origin from sensory sympathetic cells.

While the observations of the three authors last mentioned
have not been materially extended or substantiated by subse-
quent investigations, it is also true that anatomical and physio-
logical evidence is hardly sufficient as yet to rule out the possi-
bility of the existence of sympathetic commissural neurons.

The writer’s contribution to this phase of the structure of the
sympathetic trunk is based on an experimental study of the
sacral and anterior coccygeal parts of the sympathetic trunk
of the domestic cat. As stated previously, this part of the
trunk in the cat is especially favorable, as it is connected with
the spinal nerves only by gray rami communicantes, the lowest
white ramus being that of the fourth or the fifth lumbar nerve
(Langley, 92). No preganglionic efferent fibers run from the
spinal cord to the sympathetic through the gray rami (Langley,
96). It follows that all preganglionic efferent fibers in the sacral
trunk must run through the lower thoracic and lumbar white
rami and descend in the trunk to the sacral and coccygeal region.
Division of the trunks caudad to the lowest white rami should

88 SYDNEY E. JOHNSON

therefore produce easily interpreted results, as the field is cleared
of the terminations of all preganglionic efferent fibers, leaving
only the postganglionic fibers and the terminations, connections,
or fibers which are intraganglionic, interganglionic, or reach the
trunk by way of the gray rami. ‘The fine axonic ramifications
of the intercellular plexus, if formed entirely of preganglionic
efferent terminations, should completely disappear, but if formed
in part by intraganglionic or other intrinsic commissural con-
nections, these should remain undisturbed. One would also
expect marked changes in the internodal segments. As they are
largely composed, in the sacral region, of anterior and posterior
root fibers, which descend from a higher level, there would be
expected to follow practically complete dropping ‘out of the
medullated fibers as well as the non-medullated fibers of central
origin. ‘The consideration of these facts led to the experiments
described in this paper.

MATERIAL AND METHODS

Cats were used for the experiments. In a series of twelve
the sympathetic trunks were divided between the seventh lumbar
and the first sacral ganglion. For a second series of experiments
two cats were employed and each received a double operation.
The first operation was the same as that described above. The
second operation, performed a week later, consisted of an exten-
sive laminectomy and, in one of the two specimens, removal of
the entire cauda equina and conus medularis, and in the other,
division of the spinal nerve roots (on the right side) cephalad
to and including the seventh lumbar. The dorsal roots were
divided distal to the spinal ganglia. All specimens were kept
for a degeneration period of from twenty-five to thirty days..
At the end of this time the success of the operation was verified
and the portions of the trunks under investigation were removed
and stained with osmiec acid or by the pyridine-silver method.
Transverse serial sections were cut at 10x.

In addition to the experimental material, several normal
series of slides of the same parts of the sympathetic trunk were
made to serve as controls and for comparison.

SACRAL SYMPATHETIC TRUNK OF CAT 89

The writer is indebted to Dr. P. R. Billingsley for doing the
operation on several specimens and for the dissection illustrated
in figure 1.

OBSERVATIONS

Normal specimens

As a continuation of the lumbar trunks the sacral portions
converge rapidly and lie in close contact with the caudal blood-
vessels. Each trunk possesses, as a rule, three ganglionic enlarge-
ments, but internodal ganglionic swellings were frequently seen.
The first sacral ganglion is the largest of the three and was found
constantly present in all specimens examined. The second
sacral ganglion is much smaller than the first and occasionally
its cells were scattered through the internodal segments so that
there was not present any single distinct ganglionic enlargement.
The third pair of ganglia was present in all specimens examined
and the ganglia were of slightly larger size than the second. The
coceygeal trunks were followed caudad to the eighth ganglion
and appeared to be more constant in macroscopic structure than
the sacral.

The internodal (or interganglionic) segments of both sacral
and coceygeal portions of the trunk were found to be most vari-
able as to size and number. In most cases the internodal segment
was represented by two or three separate nerve strands and any
number from one to five was not infrequently seen. Fusion of
opposite ganglia and connection of opposite trunks by nerve
strands was found in practically all specimens, and in many
cases it was impossible to say with certainty which strands
belonged to the right internode and which to the left.

The coccygeal trunks diminish rapidly to mere thread-lke
filaments, and in their dissection a binocular microscope of low
power was a most valuable aid. Gray rami were dissected out
as far caudad as the fourth coecygeal ganglion.

The gray rami of the sacral and coceygeal region leave the
trunks, as a rule, near the caudal poles of the gangla, but a
good many were seen which followed the internodal segments
for half their length or even more before diverging from the
trunk.

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 1

90

SYDNEY

Int. seg. Saas

KE.

Int. seg. (cy Poe ee

Tat. seg. C32

Fig.

JOHNSON

SACRAL SYMPATHETIC TRUNK OF CAT 91

It is not the writer’s purpose to present in this paper a complete
account of the normal microscopic structure of the sacral and
eoceygeal sympathetic trunks. Only observations which are
essential to a correct interpretation of the experimental results
are included. This applies particularly to fiber counts. The
counts given below for normal specimens were not planned to
be more than approximately correct. For the purpose of this
paper, perfect counts would be no more useful as they would
not alter the significance of the experimental results which were
obtained.

Three cats were used for normal preparations. The right
trunks were stained by the pyridine-silver method and the left
were fixed in osmic acid. The preparations of one of these
specimens will be described and used as a basis for comparison
and contrast with the experimental material.

The ganglia. The sacral and coccygeal ganglia present the
same general features microscopically as the ganglia of the more
cephalad portions of the trunk. The nerve cells, however,
appear to be more irregularly and more widely scattered through
the internodal segments. Serial sections show that many
internodes contain ganglion cells throughout their entire length.
Figure 4 shows a section taken approximately midway between
the first and second sacral ganglia and it contains five nerve
cells.

The intercellular plexus has been described fully and the
literature pertaining to it considered by Ranson and Billingsley
(18a, p. 337, et seq.). In the writer’s preparations. of the
sacral and coccygeal ganglia this plexus is well stained and shows

Fig. 1 A diagram of the lower part of the sympathetic trunk, the associated
spinal nerves, and the gray rami communicantes as far caudad as they were
dissected. From a drawing by Dr. Paul R. Billingsley. The line at A shows
the level of division of the trunk in the first operation. The lines B1 to B7?
indicate the points at which the spinal nerve roots were divided in the second
operation. Gn.L7, seventh lumbar ganglion; Gn.S1 to Gn.S3, the sacral ganglia;
Gn.C1 to Gn.C4, first to fourth sacral ganglia; Int.seg.L7 to Int.seg.C3, internodal
segments from the seventh lumbar to the third coccygeal; Rm.S1 to Rm.C4,
gray rami communicates, first sacral to fourth coceygeal; Sp.gn.S1, first sacral

spinal ganglion; Sp.N.S1 to Sp.N.C4, spinal nerves, first sacral to fourth cocey-
geal; V.R., ventral root of spinal nerve; D.R., dorsal root of spinal nerve.

92 SYDNEY E. JOHNSON

* Fig. 2. Transverse section of the seventh lumbar internodal segment of a
normal specimen. There is no apparent special grouping of the medullated fibers,
and no preponderance of any one size. There are a few fibers less than 1.54 in
diameter, a great many from 1.5 to 7y, and a small number from 7 to10u. Camera
sketch. Osmie acid. X 440.

Fig. 3 Transverse section through the first sacral ganglion. Same specimen
as illustrated in figure2. Most of the medullated fibers are grouped in a crescent-
shaped bundle in the periphery of the ganglion. A detached fascicle (a) could
be traced through the entire ganglion. A fair-sized bundle of fibers (b) runs from
fascicle (a) to the forming ramus (c). A few small medullated fibers are seen
scattered throughout the ganglion. Osmic acid. Camera tracing. X 136.

Fig. 4 Transverse section of the first sacral internodal segment, approxi-
mately midway between the first and second sacral ganglia. Ganglion cells were
scattered throughout the length of the internode, and five cells appear in this
section. Osmic acid. Camera sketch. X 300.

SACRAL SYMPATHETIC TRUNK OF CAT 93

the same general features. In pyridine-silver preparations the
dendrites radiating out from the nerve cells as well as the post-
ganglionic axones appear rather lightly stained and can be traced
for only a short distance away from the cell body, usually not
more than a distance equal to the diameter of a nerve cell multi-
plied by approximately one to four. The greater part of the
plexus is formed by fibers of lesser diameter and of much darker
staining properties. The fine axonic ramifications of this plexus
give the same appearance as those shown by Ranson and Billings-
ley (18 a) to disappear from the superior cervical ganglion upon
degenerative section of the cervical trunk. A section of normal
ganglion stained by silver nitrate is shown in figure 5.

Osmic-acid preparations of normal ganglia show chiefly the
variously sized medullated fibers. The majority of these fibers
occupy distinct bundles which lie at the periphery of the ganglion.
Small medullated fibers (1.5 to 4u) are scattered throughout the
ganglion and occasionally a fair-sized bundle containing larger
fibers was seen in the interior of the ganglion. At intervals in
serially mounted sections small bundles of variously sized medul-
lated fibers were seen to separate from the main group and form
part of the gray rami (fig. 3).

The internodal segments. A considerable number of non-
medullated fibers was seen in pyridine-silver sections of the
sacral and coccygeal internodes. Small groups of these fibers
are scattered irregularly among the medullated fibers throughout
the section. Neither the size of the individual groups of fibers
nor their distribution appeared to bear any constant or suggestive
arrangement in different specimens. The crescent-shaped fasci-
cle observed by Ranson and Billingsley (18 b, p. 426) in the
thoracic trunk does not appear to be present in the sacral and
coccygeal portions.

The medullated fibers, afferent and preganglionic efferent,
are best seen in sections stained by osmic acid. In the internodes
at some distance from the ganglia these fibers are distributed
evenly over the field of a cross-section (fig. 2). As they pass
through the ganglia the medullated fibers occupy a periphera!
position. A gray ramus may be given off from the lower pole

94 SYDNEY E. JOHNSON

of a ganglion or may arise from the internode somewhat more
distally.

The size of the fibers in cross-section varies from 1.5 to 9 or
more micra. A small number of medullated fibers less than

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Fig. 5 A small portion of the second sacral ganglion. Normal specimen.
The axons and dendrites of the autonomic cells are lightly stained, while the ter-
minations of the preganglionic efferent fibers are finer and much darker. Com-
pare this camera-lucida sketch with figure 6. Pyridine silver. X 500.

Fig. 6 A section of the third sacral ganglion, made thirty days after division
of the sympathetic trunk between the seventh lumbar and the first sacral gan-
glion. The intercellular plexus of fine fibers seen in normal ganglia has completely
disappeared. Pyridine silver. Camera-lucida sketch. X 500.

1.5u in diameter was seen in the lower poles of the ganglia and
in the gray rami. Occasionally a few, never more than four or
five, of these very small medullated fibers were seen in an inter-

SACRAL SYMPATHETIC TRUNK OF CAT 95

nodal segment caudad to the origin of the associated gray ramus.
Fibers of this type were the only medullated fibers seen in certain
of the experimental material.

As to the total number of medullated fibers at various levels
in the sacral trunk, the numbers given in the succeeding para-
graphs are probably too low, as the oil-immersion lens was not
used to check up on the number of the smallest fibers. The
enumerations were carefully made under a 4-mm. objective and
a 10X-ocular, and the results are sufficiently accurate to serve
as a basis for comparison with the degenerated specimens. The
figures given are all for the same normal specimen.

In a cross-section of the seventh lumbar internodal segment
(i.e., the segment connecting the seventh lumbar with the first
sacral ganglion) a total of 444 medullated fibers was counted.
Of this number there were counted seven fibers 94 in diameter,
eighty fibers 6 to 7u, and 357 of various sizes from 1.5 to 5z.
The fibers are evenly distributed over the entire field of the
section without any apparent grouping as to size (fig. 2). The
very largest fibers, however, appear to be more frequently located
near the periphery of the trunk.

Figure 3 shows the arrangement of the medullated fibers as
they pass through the first sacral ganglion. There are about
300 medullated fibers in this section, and as the trunk just above
the ganglion contains approximately 450, it seems probable
that the decrease in number is due to the termination in the
ganglion of a proportionate number of preganglionic efferent
fibers. The ramus which is seen in the process of formation in
this section contains, a few sections caudad where it is completely
formed and separated from the trunk, 107 medullated fibers of
all sizes seen in the trunk.

In the first sacral internodal segment (fig. 4) were counted
313 medullated fibers, a decrease of 131 from the number in the
seventh lumbar segment. This segment was represented by a
single nerve strand, but it contained nerve cells throughout its
entire length.

Right and left second sacral ganglia were fused together in
this specimen, a condition commonly seen in the sacral and

96 SYDNEY E. JOHNSON

coccygeal region. As in other ganglia, the majority of the
medullated fibers are located peripheral to the ganglion cells.
The ganglion in this case is larger than usual and apparently
receives the terminations of approximately 185 medullated
preganglionic fibers.

The second sacral internode consisted of three fine nerve
strands with a total of 178 medullated fibers. Sixty-seven of
these fibers run into a ramus a short distance caudad to the
ganglion.

The third sacral ganglion in this specimen was exceptionally
small, producing only a slight enlargement in the diameter of
the trunk. The ganglion was fused with its fellow of the opposite
side and four nerve strands connected the united ganglionic
mass with the first coccygeal pair of ganglia. These four nerve
strands comprised the internodal segments of both trunks.
Together they contained a total of 254 medullated fibers. This
would allow an average of half that number for each internodal
segment.

The coccygeal trunks present the same general characters as
the sacral. Sections were made to and including the third
coceygeal internodal segment, and the outstanding feature in
these specimens is the presence of a relatively large number of
fibers (sensory) with diameters ranging from 5 to 9 or more
micra.

Results of the experiments

As described in a previous section, the experimental material
was obtained, 1) by dividing both sympathetic trunks between
the seventh lumbar and the first sacral ganglion and allowing
twenty-five to thirty days for degeneration to take place before
removing the sacral trunk for staining and, 2) by dividing the
trunks as stated above and in addition dividing or destroying
the spinal nerve roots distal to the spinal ganglia. These animals
were kept alive from twenty-five to thirty days from the time
of the second operation.

For the first series of experiments twelve cats were operated,
one after another as time permitted over a period of almost two

SACRAL SYMPATHETIC TRUNK OF CAT 97

years. In these specimens there constantly appeared a greater
number of large medullated (dorsal root) fibers than was antici-
pated would enter the trunks through the gray rami. This large
number of specimens was used to assure the writer that these
fibers could not possibly be due to failure of degeneration or to
regeneration through union of the divided trunk ends. The
results obtained in all of these specimens are characteristically
uniform and it will therefore be sufficient to describe the changes
from normal in one example.

In the specimen to be described sections were cut serially
beginning with the first sacral ganglion, the trunk having been
divided just cephalad to the ganglion. The left trunk was
stained with osmic acid, the right with silver nitrate.

In the pyridine-silver preparations of the ganglia the nerve
cells and their dendrites are stained as in normal specimens,
although the dendrites can be followed for a greater distance
owing to the absence of axonic terminations. The characteristic
change is in the intercellular plexus. The fine axonic termina-
tions seen in normal pyridine-silver preparations have entirely
disappeared. The few fascicles of fibers which remain do not
stain as darkly as the fibers of the intercellular network and the
majority can be traced into the gray rami. A few appear to
form small but definite fascicles in the internodal segments and
apparently enter successive rami or branches of distribution.

Figure 6 shows a section of sympathetic ganglion after degen-
eration of the preganglionic fibers. The elimination of the inter-
cellular plexus by section of the sympathetic trunk caudad to the
lowest white ramus was anticipated in view of the results obtained
by Ranson and Billingsley (’18 a) in the superior cervical ganglion.
In osmic-acid preparations, however, the results appeared at
first more difficult to explain.

A section through the upper third of the first sacral ganglion
showed sixteen small medullated fibers, all under 3.5u. Near
the lower pole of the ganglion a ramus containing eleven medul-
lated fibers of various sizes from 1.5 to 9u separated from the
_ trunk. There was only one fiber of 9u in diameter, one 8u, one
6u and all of the others were under 5uy.

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL, 33, NO. 1

98 SYDNEY E. JOHNSON

In the first sacral internodal segment twenty-nine medullated
fibers were counted. ‘The proportion of the various sizes was
practically the same as given above. A few are scattered over
the entire field, but the majority occur in small groups at or
near the periphery of the section.

In the gray ramus associated with the second sacral ganglion
twenty-five medullated fibers were counted. These are of
various sizes from about 1.5 up to 9 or more micra. A cross-
section of the second sacral ganglion contained fifty medullated
fibers. Most of the fibers are located near the periphery of the
ganglion, but a good many of less than 4.54 in diameter are
scattered over the entire section.

Fifty or more fibers were counted in the second internodal
segment. Classified as to size, there were two fibers 9u in diam-
eter, ten fibers 8u, fifteen fibers 64, and the rest ranged from 1.5
to 5y. All except a few small fibers occupied a crescent-shaped
bundle in one side of the trunk.

Caudally in the trunk the medullated fibers decrease slowly
in number to the third coccygeal internodal segment (as far
caudad as sections were cut) where the number of large medul-
lated fibers varied in different specimens from three to twenty
with a somewhat greater number of smaller medullated fibers.

Throughout the trunk the largest of the medullated fibers are
seen at the periphery of the ganglia and internodal segments and
the majority can be traced into the rami. The smallest of the
medullated fibers, many less than 1.5u, do not appear to have a
definite arrangement in the ganglia, but as they enter the inter-
nodes most of them join the fascicles of larger fibers and enter
the rami. It seems probable that these small fibers are post-
ganglionic axones which have acquired thin medullary sheaths.
The large and intermediate fibers remain to be accounted for.
Their size and heavy myelinization would appear to preclude
the possibility of these fibers being either the axones of intrinsic
commissural neurons or the axones of postganglionic nerve cells.
Many of the fibers cannot be readily distinguished micro-
scopically from the axones of preganglionic efferent neurons, but
the only pathway for anterior root fibers was interrupted by

SACRAL SYMPATHETIC TRUNK OF CAT 99

division of the seventh lumbar internodal segments, since pre-
ganglionic fibers are absent in the gray rami (Langley, 796).
The most probable explanation for the presence of these fibers
appears to be that they are afferent (dorsal root) fibers which
enter the trunks by way of the gray rami. Such a large number
of dorsal root fibers entering the trunks by this route was not
expected, and in order to make certain as to the source of these
fibers the second series of experiments was carried out.

In the first cat operated for this experiment both trunks were
cut between the seventh lumbar and the first sacral ganglion.
A week later the lamina of the sacrum and the sixth and seventh
lumbar vertebrae were resected and the entire cauda equina
and conus medullaris removed. White ramus fibers descending
in the trunk were eliminated by the first operation, while the
second operation removed the source of any dorsal root fibers
which might reach the trunk through the sacral gray rami. The
cat was killed thirty days after the operation. On dissection
the trunks and rami were found to be much more reduced in
diameter than was usual in the singly operated specimens. The
trunks were stained in osmic acid and serial sections cut from
the first sacral to the third coccygeal ganglion. <A section of
lumbar trunk was run as a control and short pieces of the sacral
nerves were stained and sectioned in order to check up on the
degeneration of dorsal and anterior root fibers in the spinal
nerves. ‘These sections showed practically complete degenera-
tion of all medullated fibers.

In a section through the first sacral ganglion there were found
only five small medullated fibers ranging from about 1.2 to 2u
in diameter. They occupy a small part of a fairly large fascicle,
the other medullated fibers of which have undergone degenera-
tion. A few sections caudad this fascicle separates from the
trunk to form a gray ramus. In sections through the internodal
segment just caudad to the ganglion no medullated fibers could
be found, although it was expected that a few of the same type
as found in the ramus would be present. The conditions found
in the rest of the trunk may be summed up briefly in the state-
ment that all large and intermediate fibers as well as the majority

SS a

ee ee ee ce ee

SSS

meee see eet ee

_-Int.seg. Sule i

ee SS Se SE oe

--Gn.9.2.

ie Post ganglionic efferent
# Preganglionie efferent

ee Afferent

Broken line indicates
degeneration of nerve fiber

Jai 7
100

SACRAL SYMPATHETIC TRUNK OF CAT’ 101

of the small fibers have completely degenerated, and that only
an insignificant number of the smallest medullated fibers found
in the trunk remain. The few small medullated fibers which
do remain appear to be practically limited to the gray rami and
the lower poles of the ganglia. In only one instance (the second
coccygeal internode) were any of these fibers found in the inter-
nodal segment distal to its related gray ramus.

In a second specimen the operation was modified to include
only the spinal nerve roots on the right side in addition to intra-
abdominal division of both sympathetic trunks as previously
described. Both trunks were stained, and in the right the results
are identical to those described above, in the left, to the results
obtained in the singly operated specimens, there appearing the
usual number of variously sized medullated fibers.

It is hardly necessary to say that in the few ganglia of these
specimens which were stained by the pyridine-silver method no
trace of the intercellular plexus could be found.

SUMMARY AND CONCLUSIONS

The sacral trunk presents many variations in macroscopic
structure. Asa rule, it consists of three small ganglionic enlarge-
ments, each ganglion connected with an adjacent one by one
nerve strand termed by Ranson and Billingsley (18a) the
internodal segment. The number of ganglia, however, may be
reduced or increased or the ganglion cells may be scattered
throughout the length of the trunk. Adjacent ganglia are often

Fig. 7 A diagrammatic sketch to illustrate the relations and paths of degen-
erated fibers in the sympathetic trunks of cats which have been operated upon
as described in the text. The broken lines indicate degenerated fibers. A, level
of division of the sympathetic trunks; Ant.rt.N.L.4, anterior root of fourth lum-
bar nerve; B1 to B3, points at which spinal nerve fibers were interrupted by the
second operation; G.rm.L.4 to G.rm.S.2, gray ramicommunicantes, fourth lumbar
to second sacral; Gn.L.4 to Gn.S.2, sympathetic ganglia, fourth lumbar to second
sacral; Int.seg.L.4 to Int.seg.S.1, internodal segments, fourth lumbar to first
sacral; N.L.4 to N.L.7, fourth to seventh lumbar nerves; N.S.1-N.S.2, first and
second sacral nerves; Sp.gn.L.4, fourth lumbar spinal ganglion; W.rm.L.4, white
ramus communicans of the fourth lumbar nerve. Note that no undegenerated
fibers of central origin can reach the sympathetic trunk caudad to the seventh
lumbar internodal segment.

102 SYDNEY E. JOHNSON

connected by two or more strands representing the internodal
segment. Fusions between opposite ganglia of the same segment -
are common. By degenerative section of the trunks and dorsal
roots the internodal segments are reduced to the finest thread-
like filaments.

The number of medullated fibers contained in the first sacral
internodal segment varies a great deal in different specimens.
The numbers observed ranged from 400 to 700, although counts
were not made with the greatest possible care and a good many
of the smallest fibers may have been missed. The fibers are of
all sizes from about 1.2 to 9 or more micra in diameter. The
majority of these fibers degenerate when the trunks are divided
caudad to the lowest white ramus. In view of this fact and other
evidence previously considered, it seems practically certain that
the fibers in question are dorsal and ventral root fibers which
have descended in the trunk from the lower white rami. The
very large fibers are obviously sensory fibers from the spinal
ganglia. That a good many of the small and intermediate
fibers must be from preganglionic efferent neurons is indicated
by the disappearance of the intercellular plexus in these speci-
mens. The fibers which remain after the single operation
described above number approximately from ten to fifty in
different specimens and are of all sizes seen in the normal trunk.
Section of the dorsal roots in addition to the first operation
caused all of these fibers to degenerate, except an insignificant
number which were thinly medullated and mostly under 1.5u
in diameter. All fibers which remain after the first operation
except these few very small ones appear therefore to be dorsal
root fibers which have reached the trunk by way of the gray
rami. Langley (’96) admits the possibility of the gray rami
containing a few dorsal root fibers, but such a relatively large
number was not anticipated. The small fibers which remain
after the double operation do not occur constantly in different
specimens nor regularly in successive internodes of the same
specimen. Five was the largest number seen in the cross-section
of any internodal segment. In most instances they could be
traced into the gray rami. It seems probable that these fibers

_ SACRAL SYMPATHETIC TRUNK OF CAT 103

are axones of postganglfonic cells which are in the process of
myelinization.

The sacral and coccygeal ganglia possess a rich intercellular
plexus which, so far as comparison has been made, does not
differ in any essential detail of structure from the intercellular
plexus in the cervical and thoracic ganglia. It completely
disappears following descending degeneration of preganglionic
axones in the lower lumbar and sacral trunk. It must therefore
be formed by the terminations of preganglionic efferent axones
which run to the trunk through the lower white rami.

Regarding the occurrence of commissural neurons in the sacral
and coccygeal ganglia, it is hardly necessary to add that no evi-
dence of the presence of such structures has been found. On
the contrary, all nervous elements stained by the methods em-
ployed have been traced to some other source or otherwise
accounted for. While it may be argued reasonably that possibly
there are nerve elements present which do not stain by the
methods employed for these experiments, it is to be remembered
that such structures have never been satisfactorily demonstrated
to exist by any other method. Further, the extensive observa-
tions of J. N. Langley have shown that not only are commissural
fibers not necessary to explain the physiology of the sympathetic
- system, but that their presence, in the case of certain limited
reactions, would introduce real difficulties for explanation.

104 SYDNEY E. JOHNSON

LITERATURE CITED

CARPENTER, F. W., AND ConEL, J. L. 1914 A study of ganglion cells in the sym-
pathetic nervous system with special reference to intrinsic sensory
neurons. Jour. Comp. Neur., vol. 24.

DoaieL, A. 8. 1896 Zwei Arten sympathischer Nervenzellen. Anat. Anz.,
Bd. 11, No: 22.

Huser, G. Cart 1899 A contribution on the minute anatomy of the sympa-
thetic ganglia of the different classes of vertebrates. Jour. Morph.,
vol. 16.
1913 The morphology of the sympathetic system. XVIIth Interna-
tional Congress of Medicine, London.

Jounson, SypNEy E. 1918 On the question of commissural neurons in the

_ sympathetic ganglia. Jour. Comp. Neur., vol. 29, no. 4.

Kuntz, A. 1913 On the innervation of the digestive tube. Jour. Comp. Neur.,
vol. 28.

Lanaury, J. N. 1892 The origin from the spinal cord of the cervical and upper
thoracic sympathetic fibers, with some observations on the white and
gray rami communicantes. Phil. Trans. Roy. Soc., London, vol. 183.
1896 Observations on the medullated fibers of the sympathetic sys-
tem and chiefly on those of the gray rami communicantes. Jour. of
Physiol., vol. 20.
1900 Remarks on the results of degeneration of the upper thoracic
white rami communicantes, chiefly in relation to commissural fibers
in the sympathetic system. Jour. of Physiol., vol. 25.
1903 The autonomic nervous system. Brain, vol. 26.
1904 On the question of commissural fibers between nerve cells having
the same function. Jour. of Physiol., vol. 31.

v. LennossiéK, M. 1894 Beitraige zur Histologie des Nervensystems und der
Sinnesorgane. Wiesbaden. 1894.

Micuattow, 8. 1911 Der Bau der zentralen sympathischen Ganglien. Inter-
nat. Monat: f. Anat. u. Physiol., Bd. 28.

Ranson, 8S. W., AND Bruuinestey, P. R. 1918 a The superior cervical ganglion
and the cervical portion of thesympathetictrunk. Jour. Comp. Neur.,
vol. 29, no. 4.
1918 b The thoracic truncus sympathicus, rami communicantes and
splanchnic nerves in the cat. Jour. Comp. Neur., vol. 29, no. 4.
1918 ¢ An experimental analysis of the sympathetic trunk and greater
splanchnic nerve in the cat. Jour. Comp. Neur., vol. 29, no. 4.

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Resumen por el autor, O. Larsell,
Northwestern University.

Terminaciones nerviosas en el pulmén del conejo.

En el epitelio de los bronquios intrapulmonares primarios § y
en los puntos de divisién de los 6rdenes consecutivos de bron-
quios existen terminaciones nerviosas sensoriales. Los puntos
mas distales en los cuales se han encontrado son las paredes de
los atrios del pulmén. Las diferencias estructurales relacionadas
con la posicién indican la existencia de tres tipos funcionales
de terminaciones nerviosas. Las células ganglionares de los
pulmones estan rodeadas por redes pericelulares intracapsulares
caracteristicas, que representan los procesos terminales de las
fibras nerviosas, originadas en apariencia en el vago. Estas
células emiten axones que se distribuyen por la musculatura
lisa del Arbol bronquial. Existe de este modo una disposicién
tipica preganglionar y postganglionar. La arteria pulmonray
sus ramas, incluso las arteriolas, presentan una rica inervacion
de fibras amielfnicas que terminan en relacién con las células
musculares lisas de la tunica media. Existen también unas
cudntas fibras nerviosas en las paredes de las venas pulmonares.

Translation by José F. Nonidez
Cornell Medical College. New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, APRIL 11

NERVE TERMINATIONS IN THE LUNG OF THE
RABBIT!

O. LARSELL

Department of Zoology, Northwestern University, Evanston, Illinois
FIFTEEN FIGURES

The innervation of the mammalian lung appears to be little
understood by either anatomists or physiologists. The average
text-book description states that the lungs receive their nerve
supply from the sympathetic system by way of the pulmonary
plexuses, and that the vagus nerve also contributes fibers.
- Usually references, more or less vague, are made to motor termi-
nations in the bronchial musculature and in that of the pulmonary
artery, and also to. sensory terminations in the parenchyma of
the lung. Physiologists have evidence for the presence of both
sensory and motor fibers in the bronchi and their branches,
but concerning the vasomotor control of the pulmonary circu-
lation they have obtained discordant results.

It was with the hope of throwing further light on this subject
that the present investigation was undertaken at the suggestion
of Dr. W. 8. Miller, to whom the writer is greatly indebted for
assistance with difficult points of the anatomy of the lung.

The general problem undertaken has to do with the distribu-
tion and arrangement, as well as the source, of the nerve fibers
which enter the lung, in addition to the types of nerve termi-
nations and the position of the latter within the organ. A
description of the fibers, their origin and the relation of the
intrapulmonary ganglia is reserved for a subsequent report,
and these points will be touched upon at present only to the
extent necessary to make clear the relations of the various
sensory and motor terminations which are described.

1 Contribution from the Anatomical Laboratory of the University of Wiscon-
sin and the Zoélogical Laboratory of Northwestern University.

105

106 O. LARSELL

MATERIAL AND METHODS

Rabbits weighing 3 to 4 pounds were used for the greater part
of the present investigation. Various modifications of the
methylene-blue technique were tried. The best results were
obtained by using 0.05 per cent methylene-blue in Locke’s
solution or in 0.9 per cent NaCl solution. Both Griibler’s and
Harmer’s methylene-blue were employed with about equal
satisfaction.

The best staining of the nerve terminations in the bronchial
tree resulted from injecting the warm stain into the pulmonary
blood-vessels, through the right ventricle. On the other hand,
the best preparations showing the innervation of the pulmonary
vessels were obtained by filling the lungs through the trachea,
by means of a funnel, with the warm stain. The lung was filled
to about normal maximum distention, when the stain was intro-
duced through the trachea, or was inflated with air to about the
same degree of distention after the stain was injected into the
pulmonary vessels, when that procedure was adopted.

Following either method of introducing the stain, the lung
was allowed to lie undisturbed in situ for ten minutes. In those
cases in which the stain had been introduced by way of the
trachea, the excess stain was then drained off. In the other
cases in which it had been injected into the blood-vessels this
was not necessary. The lungs were then alternately inflated
with air and deflated, at the rate of twelve to fifteen times per
minute. This was accomplished by means of a rubber-bulb
hand blower which was connected to a cannula inserted into the
trachea by means of a rubber tubing and a Y-glass connection.
A short rubber tube with a pinch-cock was attached to one arm
of the Y connection. The blower and the cannula were con-
nected by tubes with the other two arms, respectively. Defla-
tion of the lungs was accomplished by opening the pinch-cock
and pressing gently with the hand on the outer walls of the
thorax. The thoracic cavity had previously been opened to
allow of observation of the process of staining.

NERVE TERMINATIONS IN LUNG OF RABBIT 107

It was found that twenty to twenty-five minutes was the
optimum time for continuing the oxidizing of the stain in the
lung tissue in this manner. The lungs were then quickly extir-
pated and filled with 8 per cent cold ammonium molybdate, to
which two to five drops of 1 per cent osmic acid were added
per 100 cc. of the ammonium molybdate. The trachea was then
ligated and the lungs immersed in a considerable quantity of the
same mixture kept at a low temperature. This was allowed to
act overnight. The ammonium molybdate was washed out by
allowing the preparation to remain for an hour in running
tap-water. The lungs were then filled with 95 per cent alcohol,
which was changed several times within an hour, and also
immersed in alcohol to bring about fixation and hardening of
the tissues. This was followed by absolute alcohol for several
‘hours or overnight. The lungs were then cut into pieces 2 to 4
mm. thick, cleared in xylol, and embedded in paraffin. Some
of the lungs were treated with 95 per cent alcohol immediately
following the ammonium molybdate, without previous washing
in water. These showed the atria and air spaces to better
advantage, but many of the air passages were filled with a precip-
itate which often obscured the nerve fibers. Sections were cut
at 25 u to 100 w and mounted serially by the usual methods.
Some were counterstained on the slide with aurantia.

The pyridine-silver method also was tried on the lungs of
several kittens, but with unsatisfactory results.

SENSORY TERMINATIONS

Sensory nerve endings occur in the epithelium of the primary
bronchi, at the division points of the bronchi of the various
orders and also in the walls of the atria. Careful examination
of five pairs of lungs prepared by several modifications of the
methylene-blue technique failed to reveal any indubitable
nerve terminations resembling the sensory type in the walls of
the air sacs or pulmonary alveoli. Many of the preparations
appeared to be sufficiently well stained to warrant the expec-
tation of finding such endings in the air sacs and alveoli if they
are present.

108 O. LARSELL

The sensory nerve terminations in the epithelium of the primary
bronchi consist of elaborate ramifications of relatively large mye-
linated fibers (fig. 1). These fibers are given off from the nerve
trunks which course along the bronchi, forming a loose longitudi-

Fig. 1 Sensory nerve terminations in epithelium of primary bronchus within
the lung. Rabbit Rl. Methylene-blue stain. Camera lucida. X 300.

Fig. 2 Sensory nerve termination at the point of division of one of the larger
bronchi: Rabbit Rl. Methylene-blue stain. Camera lucida. 50u. X 280.

ABBREVIATIONS FOR ALL FIGURES

art.pul., pulmonary artery mu.bd., muscle band

br., bronchus mu.c., smooth muscle cell

brl., bronchiolus n.fi., nerve fiber

du.alv., ductulus alveolaris n.ter., nerve termination

epith., epithelium n.tr., nerve trunk

l.nod., lymph nodule po.fi., postganglionic fiber

lu., lumen pr.fi., preganglionic fiber

l.v., lymphatic vessel t.fi., terminal fiber to muscle cell

nal meshwork on the walls of the air tubes, and pass singly or in
small bundles of two or three fibers to the base of the bronchial
epithelium. When they occur in strands of more than one
fiber, these strands break up near the base of the epithelium
into single fibers which supply separate areas.

NERVE TERMINATIONS IN LUNG OF RABBIT 109

The individual nerve endings spread over a relatively large
area of epithehum. The termination illustrated in figure 1
is represented as viewed somewhat obliquely so that the relative
thickness of the epithelium in which it lay could not be indicated
in the figure. The greatest breadth of this termination is about
125 » and its length about 150 u. As will be noted in the figure,
the main nerve fiber breaks up into seven principal branches,
which in turn subdivide into secondary and tertiary twigs. All
are studded with varicosities of varying size and form, and the
tip of each of the terminal twigs is formed by a small knob-like
enlargement. These terminal twigs pass between the cells of
the columnar epithelium. Many approach the surface of the
epithelium, but most of the twigs appear to end between the
_ epithelial cells. These large, intricate nerve terminations
resemble in many respects the endings found by Wolff (02) in
the frog’s lung, with the difference that those described by Wolff
were found in the interalveolar walls.

The most characteristic position in which sensory endings are
found in the rabbit’s lung is in the area of epithelium which lies
between the bronchi of the various orders at the point of division
of the bronchi. The plexiform nerve termination is situated at
or near the apex of the point of epithelium in the crotch formed
by the branching of the bronchial divisions. The endings
which occur in the larger bronchial branches are of relatively
larger size than those which are present in the epithelium of
the lesser divisions. There does not seem, however, to be any
difference in the size or appearance of the nerve fibers which
lead to these terminations, no matter where located. With
the methylene-blue stain they all show varicosities of varying
size and form. ‘These fibers are given off from the nerve bundles
which run roughly parallel with the bronchi. In many instances
they may be traced individually for considerable distances before
terminating in the epithelium (figs. 4 and 5). On reaching the
epithelium they break up into a network which interlaces among
the epithelial cells, very much in the same manner as the larger
endings, above described, in the primary bronchi. The plexus
thus formed possesses numerous varicosities, the larger of which

110 O. LARSELL

appear to constitute nodal poits from which the secondary
and tertiary branches are given off. As in the case of the larger
terminations found in the primary bronchi, some of the tertiary
rami approach the surface of the epithelium, but others appear
to end between the epithelial cells. All are characterized by
small knobs at their tips. In thick sections of 504 to 75u, in
which these terminations can be studied to best advantage, the
greater number of terminal filaments appear to end between the
cells of the epithelium.

With this general description in mind, it will be of advantage
to turn attention to the peculiarities of the terminations at the
various orders of the bronchial branches. Figure 2 represents
a sensory ending located at the point of division of a large
bronchus. Comparison of this figure with figure 1, from the
epithelium of the primary bronchus, will show many points of
similarity between the two. The termination from the bifur-
eating portion of the bronchus has the same characteristic type
of ramification, its rami subdivide similarly, and in general
have a similar appearance. The principal difference between
the two consists in the smaller size of the termination from the
smaller bronchial division.

These endings in the larger bronchi appear to correspond
with the terminations described and figured by Berkley (’93) as
brought to view by the Golgi method of staining. Berkley also
figured enlargements on the fibers leading to some of the nerve
endings, and suggested that these might be nerve cells. Similar
enlargements have been observed in the present study, but they
appear to be unusually large varicosities rather than nerve cells.
This conclusion is based on the fact that the enlargements in
question are of considerably smaller size than any undoubted
nerve cells encountered in the lung. Also no nucleus was
observed in any of the enlargements of this type encountered,
although the method of staining employed would ordinarily
differentiate between the nucleus and the cytoplasm, while the
Golgi method might not do so.

A point of considerable interest in connection with the nerve
terminations at the points of branching of the larger bronchi

NERVE TERMINATIONS IN LUNG OF RABBIT Tf

consists in the fact that masses of lymphoid tissue are often
located between the bifurcating branches in such a manner as to
lie in close juxtaposition to the epithelium in which the nerve
endings are found. In many cases the fibers leading to the
nerve terminations pass through the borders of the lymphoid
mass, and the latter frequently extends to the base of the
epithelium.

Fig. 3 Sensory nerve termination at the division point of a small bronchus.
Rabbit R1, series D5. Methylene-blue stain. Camera lucida. 624. X 374.

As the bronchial tree is followed to its smaller branches, it is
easy in thick sections to trace individual fibers which are given
off from the nerve bundles. These pass either individually or
accompanied for greater or shorter distances by one or two
similar fibers, and make their way to the points of division of
the bronchioli and alveolar ducts. Such a fiber, with its termi-
nation at the division of a small bronchus, is illustrated in figure
3. It will be noted that the termination, which is located in a

LARSELL

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112

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NERVE TERMINATIONS IN LUNG OF RABBIT fis

mass of low columnar epithelium, shows much more limited
branching than do the two previously described. The general
type, however, appears to be the same. There is adaptation to
the more limited area of epithelium, and in this case, as in
those previously described, the terminal twigs radiate into the
surrounding territory of the epithelium.

A termination of somewhat different appearance is illustrated
in figure 4. In this instance, and in others resembling it, the

Fig.5 Graphic reconstruction of two sections cut at 62u, illustrating the rela-
tion of a nerve fiber and its sensory termination to a respiratory bronchiole and
two alveolar ducts which arise from the bronchiole. -Rabbit R1, series D4.
Methylene-blue stain. Camera lucida. 60.

Fig.6 The nerve termination represented in figure 5, highly magnified. Cam-
era lucida. X 660.

terminal arborization presented as a whole an ovoidal appearance
with the ultimate branches curved. These terminal twigs were
found in a nodule-like mass of epithelium, which was located at
the point of bifurcation of a small bronchus into a respiratory
bronchiolus.

In figure 5 is represented a graphic reconstruction from two
sections of a series cut at 62 yu, to illustrate the relation of a fiber
and its termination to a respiratory bronchiole and two alveolar
ducts which arise from the latter. The wail of the bronchiole

114 O. LARSELL

is represented as translucent, as it appears in a balsam mount,
so that the nerve fiber shows through it. The fiber pursues a
tortuous course outside the epithelial wall of the bronchiolus
to the point where the latter divides to give rise to the alveolar
ducts. At this point, between the openings into the alveolar
ducts, there is a nodule of somewhat thickened epithelium.
The nerve fiber enters this nodule and breaks up into numerous
fine branches, each of which terminates as a small knob. All
the twigs, as well as the entering fiber, have numerous vari-
cosities.

This same termination is represented at greater magnification
in figure 6, in which the details of structure are more clearly
shown. As represented in the figure, a central filament may be
followed for a considerable distance. This filament gives off
the numerous branches to which reference has already been made.
These branches interlace in the most confusing manner. Many of
them subdivide into smaller rami which terminate in minute
knobs. The larger mass near the center of the figure indicates
a varicosity of unusual size, which is somewhat foreshortened
in the figure, and on that account appears larger than it does
in the preparation itself.

The terminal fibers in the two terminations last described,
and in numerous others which were observed to occupy a corre-
sponding position, are curved, as already noted, and appear
somewhat drawn together, in contrast with the extended and
radiating ultimate processes of those previously described.
Whether or not this difference is of significance is problematical.
The suggestion presents itself that these terminations located
at the openings into the respiratory portion of the lung may
represent a somewhat different functional type than those
located in the larger air passages. It appears possible that the
latter represent a type of tactile terminations which may be
stimulated by foreign bodies or by masses of mucus within the
bronchi. ‘Their location at the portals to the smaller-air passages
appears admirably adapted to guard against the entrance of
such objects from the larger bronchi by causing stimulation at
the points such objects would naturally strike, and thus produc-

~

NERVE TERMINATIONS IN LUNG OF RABBIT 115

ing reflex contraction of the bronchial musculature, constricting
the smaller air passages still further, and possibly also initiating
reflex coughing to expel the object.

The terminations at the openings into the _ respiratory
portion of the bronchial tree might likewise serve the purpose of
guarding against the entrance of foreign objects into the atria
and air sacs by initiating reflex constriction of the small sphincter-
like muscle bands at the openings into the atria. The difference
in form, however, of these endings, which appear better adapted
to react to pressure stimuli than to touch, suggests the possibility
that such terminations may be stimulated rather by the partial
collapse of the lung during expiration. When the lung is dis-
tended the alveolar ducts are doubtless spread apart at relatively
greater angles with each other than are the other subdivisions
of the bronchial tree. On expiration they probably approach
relatively closer together, thus affording sufficient pressure to
stimulate the terminations in question.

The most distal point at which nerve endings have been found
in the epithelium is just inside the atria as they are entered
from the alveolar ducts. These terminations (fig. 7) are of
small size and are covered by a delicate capsule, formed in part
by the squamous epithelium, beneath which they lie in the wall
of the atrium. The nerve fibers leading to these terminations
are branched, as shown in the figure, so that each fiber probably
terminates in a number of sensory endings. On entering the
capsule the fiber divides into several branches, which in turn
subdivide into small twigs. The latter terminate in small knobs.
All the branches have typical varicosities. The whole struc-
ture has a flattened appearance, roughly elliptical in outline.

It appears possible that this type of termination may be
influenced by stretching of the atrial walls during inspira-
tion, and may represent the nerve terminations which many
physiologists have maintained are responsible for the inhibition
of the inspiratory movements and excitation of expiration.

Retzius (93) found fibers and terminal branches in a human
foetus of 15 em. reaching to ‘‘the necks of the alveoli,”’ as a
rule, with an occasional fiber spreading over the rounded alveoli.

116 O. LARSELL

Ponzio (’06) has described and figured fibers in the lung
parenchyma which he designates as interalveolar, intercellular,
and intracellular nerve filaments, respectively, and he also
describes an intracellular nerve reticulum. Ponzio employed the

5

Fig. 7 Sensory nerve terminations in the wall of an atrium. Rabbit R1,
series;H3. 62u. Camera lucida. X 880.

methylene-blue and Cajal methods of staining on the lungs of
newborn kittens and puppies.

In my preparations, both those stained with methylene-blue,
and those prepared by the pyridine-silver method, there exist
fibers and filaments which closely resemble those illustrated by
Ponzio and in corresponding positions. I am inclined to believe
that they are elastic fibers rather than nerve fibers. This

NERVE TERMINATIONS IN LUNG OF RABBIT fly

conclusion is based on a detailed comparison of such fibers, as
demonstrated in methylene-blue and silver preparations of the
lung, with the elastic fiber network brought to view by the
specific elastic fiber stains, such as orcein and resorcin-fuchsin.
Undoubted nerve fibers elsewhere in the lung, even those of the
smallest size, have a different appearance from the fibers under
consideration. The difference consists in the much darker stain
which the nerve fibers take and in the fact that the latter always
present varicosities and have a much more irregular course than
do elastic fibers.

A reexamination of my preparations after reading Ponzio’s
article, and a comparison of his figures with the fibers in these
preparations, has strengthened the conclusions already reached.
namely, that the network of delicate fibers in the walls of the
air sacs is composed of elastic fibers. Ponzio’s figure 9 should
have separate comment and will be referred to again in connection
with the innervation of the pulmonary blood-vessels.

There appears little room for doubt that the several termina-
tions described are of sensory type. Their position in the epi-
thelium, particularly at the points which appear most subject
to irritation, their connection with the relatively large, myeli-
nated nerve fibers, and the character of their terminal arboriza-
tions, appear to point to the conclusion that they are sensory
rather than motor. The present writer has not attempted any
experimental studies to test these conclusions, but the results
_ obtained by Molhant (713), who studied the distribution of the
sensory fibers of the vagus nerve in the rabbit, agree very well
with the interpretation expressed.

Molhant extirpated the superior and median lobes of the right
lung in the rabbit, and after allowing time for the injury of the
nerve fibers involved in the trauma to affect the cells of origin
of these fibers, he stained the peripheral ganglia of the vagus
by the Nissl method. He found that ganglionic cells in the
external portions of the median and inferior segments of the
ganglion nodosum showed the typical degeneration of the Nissl
granules as a result of the injury suffered by their peripheral
processes. Molhant concluded that the cells thus affected

118 O. LARSELL

were the ganglionic cells of the sensory fibers which supply the
two lobes of the lung in question. .

Similar results had previously been obtained in the dog by
Ikegami and Yagita (’07), also by experimental methods. They
concluded that from about 12 per cent to 13 per cent (1/8.3 to
1/7.5) of the total number of cells of the right nodose ganglion
are in connection, in the dog, with the pulmonary parenchyma.

Chase and Ranson (714), who studied the finer structure of
the vagus nerve in the dog, cat, rabbit, rat, and human, state
that the bronchial rami of the vagus contain large numbers of
myelinated fibers and take from the vagus a considerable propor-
tion of the myelinated fibers which have continued in it down to
this level. They do not make any specific statement as to the
size of the myelinated fibers in the bronchial rami of the vagus,
but in a preceding paragraph they state that this nerve, just
above its pulmonary branches, contains a few myelinated fibers
of large size, although the majority are of small or medium size.
Below the pulmonary rami of the vagus nerve relatively few
myelinated fibers are present.

The results of these various investigators indicate that periph-
eral fibers from cells in the ganglion nodosum pass by way of
the vagus nerve and its pulmonary rami into the lung. It
appears reasonable to assume that the sensory terminations
above described are the terminal processes of these fibers.

In addition to the sensory terminations in the bronchial tree,
the writer is able to confirm the observations of Schemetkin,
reported by Dogiel (’98), that there are sensory terminations in
the walls of the pulmonary artery. So far as the writer has
observed, these are found only near the base of this artery,
close to the hilum of the lung.

MOTOR TERMINATIONS

Motor terminations are found in the rabbit’s lung, both in
the smooth muscle of the bronchial tree and in the muscular
coat of the pulmonary vessels.

NERVE TERMINATIONS IN LUNG OF RABBIT 119

Plesehko (’97), by staining with methylene-blue, found motor
terminations in the smooth muscle of the trachea. He also
figured and described ganglion cells with processes which pass
to the smooth muscle bundles, there to divide into fine fibers
which terminate in the individual muscle cells. These ganglion
cells are surrounded by pericellular plexuses of fine varicosed fibers
formed by the splitting up of nerve processes which apparently
are given off from the vagus nerve. ‘There is present, therefore,
in the trachea the typical relation of preganglionic (vagus)
fibers to the terminal ganglionic cells with their postganglionic
fibers. The latter are represented by the axones which pass to
the trachealis muscle, there to ramify in the manner above
described.

According to Miller (18), the musculature of the bronchi is a
direct continuation of the musculature of the trachea. It will
appear that the same pattern of innervation also holds for the
smooth muscle of the bronchi and their branches that is present
in the trachea.

Clusters of ganglionic cells (figs. 8 and 9) are found along the
primary bronchi and their larger branches. These clusters
constitute small ganglia which are usually located along the
course of the larger nerve bundles on the wall of the bronchial
tree. The larger clusters of cells, which are located nearer to
the hilum of the lung, are surrounded by a connective-tissue
capsule. This capsule also contains frequently a fairly large
nerve trunk, which is composed of both myelinated and unmye-
linated fibers. The myelinated fibers are of three sizes, corre-
sponding to the description given by Chase and Ranson (’14)
of the fibers in the vagus nerve. There are a few myelinated
fibers of large size and more numerous myelinated fibers of small
and medium size.

It has been long known that in the walls of the larger bronchi
the muscle occurs in bands. In the smaller subdivisions where
no plates of cartilage are present, the muscle is said to constitute
a continuous sheet. No muscle fibers are found distal to the
alveolar ducts, but there isa sphincter-like band at the extremities
of these ducts, surrounding the openings into the atria, as was

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 2

120 O. LARSELL

pointed out to me by Doctor Miller. These various muscle
bands have a very rich innervation. The fibers which pass to
them are easily distinguished from those which have been
described. above as sensory fibers, by reason of the smaller size
of the former and the fact that they are unmyelinated.

Many of these fibers are direct processes from the ganglionic
cells, as shown in figures 8 and 9, and are therefore postgan-

Fig. 8 Cluster of ganglionic cells in the wall of one of the larger intrapul-
monary bronchi, showing preganglionic fibers terminating about the nerve cells
in pericellular, intracapsular networks, and postganglionic fibers as processes
from the nerve cells. A portion of a nerve trunk is also shown. Rabbit R3.
Methylene-blue stain. 50u. Camera lucida, stage level. X 400.

glionic fibers. Some, as in figure 9, can be observed to pass
directly from such a ganglion cell to the smooth muscle bands,
where they branch into numerous slender varicosed rami. These
rami run parallel to the smooth muscle bands between the smaller
bundles of muscle cells which compose the bands. At intervals
they give off short twigs which terminate as small knobs on the
muscle cells. The processes from other ganglionic cells, especially

NERVE TERMINATIONS IN LUNG OF RABBIT Lib

from clusters of such cells which lie along the nerve bundles,
become a part of these nerve bundles and cannot be followed
individually. There can be no doubt, however, that they have a
distribution and termination similar to that just described for
the processes of the relatively few isolated cells which it was
possible to follow.

Fig. 9 A single nerve cell in the wall of an intrapulmonary bronchus, showing
a fiber (preganglionic) from a nerve trunk, terminating about the cell as an intra-
capsular, pericellular network, and also showing the axone from the nerve cell.
The axone divides into two branches outside the capsule of the cell, each branch
passing to bundles of muscle. The manner of termination of one of these branches
in relation to two small muscle bundles, is indicated in the left-hand side of the
figure. The preganglionic fiber and one of the branches of the axone of the cell
have been fore-shortened, as represented by the broken lines, to save space.
Rabbit R38. Methylene-blue stain. 50u. Camera lucida. > 400.

22 O. LARSELL

At intervals small bundles of fine fibers are given off from the
main nerve trunks. These bundles pass to the muscle bands
and there break up by the separation from them of the individual
nerve fibers, as shown in figure 10. The individual nerve fibers,
on reaching the muscle bands, divide into numerous slender
filaments which run parallel between the muscle fibers, as illus-

Fig. 10. Muscle bands in wall of a bronchus, illustrating the manner in which
the nerve bundles break up into single strands and the manner in which these
strands are distributed to the muscle fibers. Rabbit R4. Methylene-blue stain.
60u. Camera lucida, stage level. X 150.

trated in the figure, and at intervals give off short twigs, better
shown in figure 11, which terminate near the nuclei of the indi-
vidual smooth muscle cells, as already described.

The recent work of Carlson and Luckhardt (’20) has demon-
strated the presence of inhibitory fibers only, in the lungs of the
axolotl and Necturus, while both inhibitory and motor fibers
reach the lungs through the vagus nerve in the frog. .There
appears to be an even larger number of motor fibers in the
reptilian lung, as represented by the turtle.

NERVE TERMINATIONS IN LUNG OF RABBIT 123

_ If these results can be applied to the mammalian lung, one
might expect to find two physiological types here also. It
appears possible that many of the fine nerve fibers which are
present in the intrapulmonary nerve trunks are not related to
the intrapulmonary ganglia, either as preganglionic or post-

A.

Fig. 11 Muscle bands from a bronchus, under higher magnification, illustrat-
ing the manner in which the nerve fibers terminate. Rabbit Rl, series Al.
Methylene blue, with aurantia counterstain. 25u. Camera lucida. X 560.

ganglionic fibers, but represent fibers from some source outside
the lung, which reach the lung by way of the vagus nerve and are
distributed to the smooth muscle within this organ. This
supposition may serve to account for the very rich innervation
of the muscles of the bronchi, particularly. This innervation
is much richer than the processes from the intrapulmonary
ganglionic cells would appear to account for.

124 O. LARSELL

The nerve bundles follow the bronchi in diminishing size to
the bronchioli and even to the alveolar ducts. They give off
branches which are distributed to the smooth muscle bands
around these smaller air passages, as shown in figure 12, which

Fig. 12 Nerve terminations on the muscle bands of a bronchiolus. Rabbit
Ri, series Al. Methylene blue, aurantia counterstain. 254. Camera lucida.
x 300.

NERVE TERMINATIONS IN LUNG OF RABBIT 125

represents one of the smaller bronchioles. The most distal point
to which fibers of the motor type have been followed is renre-
sented by the small sphincter-like muscle bands at the openings
into the atria from the alveolar ducts. Slender varicosed fibers
have been observed to run among the muscle cells of these bands
and to terminate in relation to them.

No ganglionic cells have been found beyond the larger bronchi,
so that the nerve bundles which pass to the smaller air passages,
must contain, in addition to the few sensory fibers, which can
be recognized by their large size, also the motor and inhibitory
fibers.

Innervation of the pulmonary vessels

The pulmonary artery and the vessels branching from it have
a much richer nerve supply than the usual anatomical descrip-
tion indicates or than might be inferred from the meager and con-
flicting results obtained in the past by physiological experiment.

Near the hilum of the lung relatively large nerve trunks
accompany the larger pulmonary arterial branches. No ganglion
cells have been encountered anywhere along the nerve trunks
which accompany these vessels. The source of these nerve fibers
isreserved for further investigation. It should, however, be stated
at this point that the smaller branches of the pulmonary artery,
which lie in close juxtaposition to small bronchi, receive fibers
from the nerve plexus around the latter. A similar observation
was made by Berkley (’93) in the gray rat. The recent physio-
logical results of Carlson and Luckhardt (’20) indicate that
these fibers are derived in large part, if not entirely, from the
vagus nerve, at least in Amphibia and reptiles.

The nerve bundles wind about the blood-vessels, giving off
individual fibers at more or less irregular intervals. These
fibers run roughly parallel with the artery, then turn nearly at
right angles to the longitudinal axis of the latter, and divide
into several main branches, as illustrated in figure 13. One of
these rami usually runs back along the artery, proximally, from
the point of primary division of the fiber, another runs distally
from this point. These larger branches give off numerous

126 O. LARSELL

slender varicosed fibers which in turn subdivide. These appar-
ently correspond to the fibers observed and figured by Karsner
(11) in the pulmonary artery of the dog. The final fibers pass
between the smooth muscle bands of the tunica media of the
arterial wall, and give off twigs which terminate in relation. to
the smooth muscle cells, as above described and as illustrated
in figure 15, which represents a portion of the wall of an arteriole.

Fig. 13 Distribution of nerve fibers in the muscular portion of the wall of
the pulmonary artery within the lung. Rabbit R38. Methylene-blue stain. 50z.
Camera lucida. X 75.

Nerve fibers are present in the walls not only of the larger
vessels, such as represented in figure 13, but also in the various
subdivisions of the pulmonary arterial system, including the
arterioles. This may be seen by examination of figure 14, which
represents several orders of branches of a pulmonary vessel, as
shown in a single section. That the blood-vessel is a branch of
the pulmonary artery can be judged from its relation to the
bronchus which is also shown in the figure. W. 5S. Miller has

Fig. 14 Pulmonary artery and branches, showing the relation of nerve fibers
to the muscular coat of the various branches of the blood-vessel. Rabbit R1,
series B2. Methylene blue, with aurantia counterstain. 60y. Camera lucida.

x 60.
127

128 O. LARSELL

repeatedly emphasized the fact that the pulmonary arterial
branches are located ‘‘as near as possible to the bronchi” which
they accompany, while the corresponding veins are as far
removed as the architectural plan of the lung will permit.

It will be observed that the nerve fibers are present all along
the larger arterial branch represented, as well as in the various
divisions of this branch. The definite relationship to the smooth
muscle bands of the arterial walls is not so clearly indicated in
all parts of the figure, but this is due to the variations of different
portions of the arterial wall with respect to the plane of section.

That these fibers terminate in relation to the smooth muscle
cells may be seen from figure 15, which pictures a portion of an
arteriole under high magnification. It will be noted that the
nerve fiber, which lies in the adventitious layer of the arteriole,
gives off lateral branches at short intervals. These branches
subdivide and send terminal twigs to the individual smooth
muscle cells of the muscular layer of the arteriole. These
terminal twigs end as small knobs, apparently on the surface of
the muscle cell and usually near the nucleus. In the figure,
which was sketched from a relatively thick section, the smooth
muscle cells are represented at various angles. Some were cut
transversely, some obliquely, and others curve upward on one
side of the arteriole, pass across the field and curve downward
on the opposite side.

In some of my preparations there are indications of a very
delicate nerve plexus on the blood capillaries which are located
in the walls of the air sacs and the atria. Extremely fine vari-
cosed fibers are present which both in their staming reaction
and in their general appearance bear a closer resemblance to
nerve fibers, aside from their small size, than to elastic fibers.
No indication of nerve cells in connection with this possible
plexus was observed, such as have been described by Dogiel
(98) and by Prentiss (04).

Attention has been already called to Ponzio’s figure 9, which
pictures a similar network on a blood capillary in the lung. The
further study of these fibers appears to the writer to present a
separate problem, which must be left for further investigation.

NERVE TERMINATIONS IN LUNG OF RABBIT 129

A few nerve fibers were observed in the tunica media of the
pulmonary veins. Berkley (93) made a similar observation.
They appear to have the same relation to the musculature of the
veins as the fibers above described have to the arteries.

Fig. 15 Arteriole from pulmonary artery, showing a single nerve fiber in the
adventitia which gives off short branches which terminate in relation to the indi-
vidual smooth muscle cells of the tunica media. Rabbit R1, series C3. Methyl-
ene blue, with aurantia counterstain. 254. Camera lucida. X 880.

130 O. LARSELL

SUMMARY

1. Sensory nerve terminations are present in the epithelium
of the primary bronchi within the lung and at the peints of
division of the succeeding orders of bronchi. The most distal
points at which they have been observed are in the walls of the
atria of the lung.

Differences in structure, correlated with position, suggest three
functional types of sensory terminations.

2. Nerve endings, probably motor and inhibitory, are present
in the smooth muscle fibers of the bronchial musculature.

3. The ganglionic cells of the intrapulmonary ganglia are
surrounded by intracapsular, pericellular networks which are
the terminal processes of nerve fibers, apparently of vagus origin.
These cells give off axones which are distributed to the smooth
muscle of the bronchial tree. There is a typical preganglionic
and postganglionic fiber arrangement.

4. The pulmonary artery and its branches, including the arte-
rioles, havea rich innervation of fibers which terminate in relation
to the smooth muscle cells of the tunica media.

A few nerve fibers are also present in the walls of the pulmonary
veins.

NERVE TERMINATIONS IN LUNG OF RABBIT ial

LITERATURE CITED

Berkey, H. J. 1893 The intrinsic pulmonary nerves in mammals. Jour.
Comp. Neur., vol. 3, pp. 107-111.

CaRLson, A. J., AND LuckHarpT, A. B. 1920 Studies on the visceral sensory
nervous system. I. Lung automatisms and lung reflexes in the frog
(R. pipiens and R. catesbiana). Amer. Jour. Physiol., vol. 54, pp.
55-75.
Idem. IJ. Lung automatisms and lung reflexes in the salamanders
(Necturus, axolotl). Amer. Jour. Physiol., vol. 54, pp. 122-137.
Idem. III. Lung automatisms and lung reflexes in reptilia (Turtles:
Chrysemys elegans and Malacoclemmys leseurii. Snake: Eutenia
elegans). Amer. Jour. Physiol., vol. 54, pp. 261-306.

Cuase, M. R., anp Ranson, 8. W. 1914 The structure of the roots, trunk and
branches of the vagus nerve. Jour. Comp. Neur., vol. 24, pp. 31-60.

Doatet, A. 8. 1898 Die sensiblen Nervenendigungen im Herzen und in den
Blutgefissen der Siiugethiere. Archiv fiir Mikros. Anat. und Entwick.,
Bd. 52, S. 44-70.

IkEGAMI UND YaaiTa 1907 Ueber den Ursprung des Lungenvagus. Sonder-
abdruck aus den Okayama-Igakkwai Zassbi, no. 206, 32 Marz, 1907.
(Cited from Molhant, 713.)

Karsner, H. T. 1911 Nerve fibrillae in the pulmonary artery of the dog.
Jour. Exper. Med., vol. 14, no. 3, pp. 322-325.

Miuier, W. 8. 1918 A study of the nerves and ganglia of the lung in a case of
pulmonary tuberculosis. The Amer. Rev. of Tuberculosis, vol. 2, pp.
123-139.

MotuHant, M. 1913 Les ganglions périphériques du vague. Le Nevraxe, T.
15, pp. 525-579.

Pioscuxo, A. 1897 Nervenendigungen und den Ganglien der Respirationsor-
gane. Anat. Anz., Bd. 13, S. 12-22.

Ponzio, F. 1906 Le terminazioni nervose nel polmone. Anat. Anz., Bd. 28,
S. 74-80.

Prentiss, C. W. 1904 The nervous structures in the palate of the frog; the
peripheral networks and the nature of their cells and fibers. Jour.
Comp. Neur., vol. 14, pp. 93-117.

Rerzius, G. 1893 Zur Kenntnis der Nervenendigungen in den Lungen. Biol.
Untersuchungen, N. F., Bd. 5, S. 11.

Witson, J. Gorpon. 1910 Intra vitam staining with methylene blue. Anat.
Ree., vol. 4, pp. 267-277.

Wo.trr, Max 1902 Ueber Ehrlich’sche Methylenblaufirbung und iiber Lage
und Bau einiger peripherer Nervenendigungen. Arch. fiir Anat. und
Entwick., S. 155-183.

Resumen por el autor, E. C. Case,
University of Michigan.

Sobre un vaciado endocraneal del reptil tridsico Desmatosuchus
spurensis, del tridsico superior del occidente de Texas.

El vaciado endocraneal descrito en el presente trabajo corre-
sponde a un individuo de un nuevo subdrden de reptiles, el de
los Desmatosuchia, descrito en un trabajo preliminar publicado
en el Journal of Geology, vol. 28, Niim. 6, 1920. Mediante un
vaciado, el autor ha conseguido obtener una reproduccién muy
perfecta de la cavidad endocraneal. El vaciado demuestra la
existencia de una ep’fisis 0 pardfisis casi vertical, con procesos
laterales que se extienden de la misma regién cerebral. Los
procesos laterales son los vaciados de dos fosas profundas, tal
vez orificios, que marcan la pared interna de la caja cerebral a
cada lado de la profunda fosa que deprime la pared del craneo
en la posicién del orificio pineal, sin llegar a perforarle.

La hip6fisis es muy grande y esta ligeramente inclinada hacia
atrais; la porcién anterior no es visible por no estar osificadas
anteriormente las paredes de la caja cerebral. Apenas existe
una expansi6n de los l6bulos cerebrales y no se presentan estruc-
turas que indiquen la posicién de los nervios 6pticos y talamo.
El tracto olfatorio es voluminoso y ancho; los nervios II, HI y
IV pasan a través de escotaduras formadas por la aproximaci6n
de los alisfenoides u orbgitosfenoides; el nervio V sale al exterior
por un ancho orificio de las paredes laterales de la caja craneal
y no se divide hasta después de abandonar esta. El nervio
VI pasa exteriormente a través de la base de la caja craneal.
La region 6tica esta’ indicada solamente por un proceso, puesto
que las otras estructuras fueron destruidas por la fosilizaci6on.
Los nervios VII y VIII abandonan la cavidad cerebral cerca
del orificio 6tico. Los nervios IX, X y XI salen por un solo
orificio grande que también aloja la vena yugular. El XII
pasa al exterior a través de un orificio del exoccipital.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, MAY 9

ON AN ENDOCRANIAL CAST FROM A_ REPTILE,
DESMATOSUCHUS SPURENSIS, FROM THE
UPPER TRIASSIC OF WESTERN TEXAS

E. C. CASE
University of Michigan
NINE FIGURES

As has been shown by the author! in a preliminary paper, the
remains of Desmatosuchus indicate a new suborder of phyto-
sauroid reptiles. In cleaning the skull of this specimen it was
found that the brain cavity was practically undistorted and
that it was possible to obtain a plastic cast of the endocranium.
As is well known, any endocranial cast does not reveal the true
shape of the brain, and this is particularly true of the reptiles
where the brain is surrounded by a mass of connective tissue or
by a large space between the pia and dura mater which is crossed
by fibers of connective tissue. Nevertheless, such casts give
an idea of the form and relative size of brain and the location
of the cranial nerves and blood-vessels. Such casts from mam-
malian skulls are not uncommon in some localities as the Oligo-
cene deposits of the Big Bad Lands of South Dakota, but very
few have been found from the lower vertebrates. Even skulls
so well preserved that the cavity may be cleaned out and casts
made are not common. Moodie? has figured such a cast from
the Pennsylvanian deposits of Kansas without assigning any
taxonomic position to the specimen and has briefly reviewed
the literature of endocranial casts of fossil forms up to the time
of his paper. To his publication the reader is referred for a
historical account of the subject and a discussion of such casts
as have been found or made. Moodie mentions two papers by

1 Journal of Geology, vol. 28, no. 6, 1920, p. 524.
2 Moodie, R. L., Jour. Comp. Neur., vol. 25, no. 2, 1915.

133

134 E. C. CASE

Cope* describing such casts, one from a cotylosaurian reptile,
Diadectes, from the Permian of Texas, and one from a phytosaur,
Belodon, from the upper Triassic of Texas, but he neither dis-
cusses these papers nor reproduces the figures given by Cope.
This is understandable, as the figures are very poor and difficult
to interpret and the subject-matter is only distantly related to
the material of his paper. As the casts figured and deseribed
by Cope are from forms more nearly related to the one described
in this paper, the figures are reproduced in line drawings as
accurately as they may be made out.

The cranial cavity of Desmatosuchus was completely cleared
out, leaving the surface of the bone in good condition with all
the pits and foramina clearly marked. As may be seen in the
figures, the cast as finally secured shows the general form and
proportions of the brain cavity and the positions of the main
outlets. The anterior wall of the cavity was entirely carti-—
laginous or membranous, and as this portion was not preserved
in the fossil the opening was stopped with plastic clay which is
easily detected in the figures; for this reason, the form of the
olfactory tract and of the pituitary body is not completely shown.

The olfactory tract was evidently large, as in most of the
primitive forms, and extended well forward directly beneath
the upper wall of the skull. The cerebral portion was relatively
small, scarcely any swelling being revealed in this part of the
cast except at the posterior end of the prosencephalic region.
The anterior-lower part of this region was enclosed by the al.-,
orbitosphenoid bones, and the approximation of the bones of
the two sides forms notches in two places which indicate the
points of escape of the nerves which supplied the eye. No
indication of the origin of the II, III, or IV pair of nerves is
shown on the cast and no outlets except the notches mentioned.

It is impossible in the cast to distinguish between the dience-
phalic and the mesencephalic regions of the brain, but the area

3 Cope, E. D., Proceedings American Association Advancement of Science,
Ann Arbor meeting, 1885, and Proceedings American Philosophical Society,
1886, p. 234.

Proceedings American Philosophical Society, 1887, p. 219, and American
Naturalist, vol. 22, 1888, p. 914.

ENDOCRANIAL CAST FROM A REPTILE 135

occupied by these two is marked by a slight but distinct depres-
sion, which is outlined by definite elevations; the posterior one
amounting to a low, sharp ridge. From this depression rise the
processes, above and below, which may be referred to in general
terms as the epiphysis and ae pituitary body.

The upper process is complex and is, perhaps, composed of
two parts. Just posterior to the edge of the prosencephalic
portion there are two protuberances which mark the position
of a pair of deep pits in the upper wall of the brain case. In
cleaning the skull it was impossible to be certain that the bottom
of these cavities had been reached, but it seemed probable that
it had. With the aid of a dentist’s mouth-mirror and fine
curved awls the pyrite filling was picked out until it seemed that
the bottom had been reached, but because of the inaccessibility
of the cavities and their small diameter it is possible that the
cavities may have been deeper and even that they may be the
entrances to foramina. Cope, in describing the endocranial
casts of a phytosaur, Belodon, and of a cotylosaurian reptile,
_Diadectes, speaks of the ‘lateral processes of the epiphysis’ and
in describing the skull of Belodon speaks of the process as lying
in a “large canal which enters the posterior part of the orbit.”
To this canal he gave the name of the orbitopineal canal and in
certain papers speaks of the orbitopineal process on the casts.
The function of this canal he was unable to determine, but
suggests that it carried a nerve or blood-vessel.4 In his earlier
papers he was inclined to the belief that Diadectes was blind
because he could find no outlet for the- optic nerve and because
the structure of the animal suggested that it was burrowing in
habit; as the parietal foramen is exceptionally large in this form,
he was inclined to believe that the orbitopineal canal might
have carried a nerve from the large, probably functional, eye
which occupied the parietal foramen, which in part supplied
the necessary vision. It is impossible to tell whether such a
canal existed in Desmatosuchus, but if it was present it was
very small, and the author of this paper is inclined to believe

4Dr. R. L. Moodie, in conversation with the author, has suggested that these
processes may indicate a portion of the course of the ductus endolymphaticus.

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 2

136 E. C. CASE

that it did not exist. Moreover, there is a decided difference
in the endocranial casts in this region. In Belodon and Diadectes
the processes are large and rise from the sides of the epiphysis,
in Desmatosuchus they are small and are entirely anterior to
the epiphysis. It is possible that if a true cast of the brain could
be obtained the origin of the processes might be found “to be
the same in all, but as the casts were all made from empty
cavities a similar origin should be apparent. '

The epiphysis is very different in form. In Belodon and
Diadectes there is a strong posterior process, and Cope describes
the epiphysis of the former as ‘subquadrate.’ The orbitopineal
process extends either directly outward from the side, Diadectes,
or outward and forward, Belodon. In Desmatosuchus the
epiphysis is erect and narrow anteroposteriorly with no posterior
process. In both Belodon and Desmatosuchus the processes
referred to as the epiphysis are casts of a deep pit on the under
side of the skull in the exact position of the pineal foramen in
other reptiles, but in neither of these is the roof perforated. . In
looking up this matter the author has found that much uncer-
tainty exists as to the exact character of this process in the brain;
it is known that both the epiphysis and the paraphysis may reach
large size and that either one may terminate in a functional
eye; at least, either one may carry organs which possess the
histological structures of the retina and the crystalline lens.
In some forms there has also been found a third evagination of
the brain, posterior to the epiphysis, called the pineal organ,
which has a similar histological structure. Wilder, in his History
of the Human Body, states that it is the paraphysis which was
developed in the extinct Stegocephalia and filled the parietal
foramen and the epiphysis which was developed in the reptiles,
birds, and mammals. On the other hand, it is known that the
epiphysis is not developed in the modern alligator. The term
epiphysis is used in this paper only in a general sense and without
knowledge of its true nature.

On the lower side of the diencephalic region of the cast is the
second process; this represents the combined infundibulum and
the saccus vasculosus, or the pituitary body. Only the posterior

ENDOCRANIAL CAST FROM A REPTILE 137

and lower borders are shown, for the anterior part was enclosed
by the cartilaginous anterior wall of the skull which was lost in
fossilization. The process extended directly downward by a
narrow neck which passed through a narrow notch formed by
the approximation of the alisphenoid bones at their lower borders.
Its posterior face lay against the basisphenoid bone, not pene-
trating it, and its lower surface is an excavation on the upper
surface of the posterior part of the parasphenoid. The lower
part ot the process was enlarged and the posterior face, at least,
extended backward at a sharp angle. The lower end terminates
in a bifureate extension which is formed by the casts of the
beginnings of two foramina which open outwardly and down-
ward in an excavation on the upper surface of the parasphenoid.
These foramina continue and terminate in deep grooves on the
side of the basisphenoid. On the posterior face of the process
are two small prominences which mark the position of two fora-
mina on the lower face of the basisphenoid, evidently the openings
for the internal carotid arteries.

The posterior part of the depressed area mentioned above
must also include the mesencephalic portion of the brain, but
there is nothing to mark the presence of either optic lobes or
optic thalami. This does not, however, suggest either their
absence or relatively small size, for if a cast were made of the
brain cavity of Sphenodon or of an alligator no evidence of these
structures would appear, though they are of large size.

Posterior to the depressed area the whole cast is curved sharply
downward and then straightened out horizontally in the metence-
phalic region. On the lower edges of the anterior part of this
region there are large prominences which mark the position of
the large foramina for the passage of the V pair of nerves. There
is no indication in the cast of the division of this nerve into its
parts; this must have taken place external to the cranial wall.
Within and a little posterior to these prominences is indicated
the position of a pair of small foramina in the floor of the skull,
evidently the outlets for the V pair of nerves. Posterior to the
V and at about the middle of the posterior part of the cast there
are a pair of processes on each side, one almost directly above the

138 i c. CASE

other. The upper pair are the casts of the otic cavities and mark
approximately the position of the VIII, and probably, also, the
VII nerves, for these two pairs escape from the skull of the
Crocodilia in almost the same place. ‘The otic cavities were
injured in fossilization both by pressure and by the erystalli-
zation of the gypsum and pyrite which filled the cavities of the
skull. It is apparent that there was a thin wall between the
otic cavity and the brain cavity, but this has been so injured that
it is impossible to determine the original form of the otic cavity
or the form and position of the semicircular canals.

Below are the large cylindrical projections which filled large
foramina carrying the IX, X, and XI nerves and the jugular
vein. All of these must have escaped through a common open-
ing, as the walls of the brain are very perfect in this place and
no other openings are present.

Near the posterior end of the cast are slender processes which
mark the position of the XII nerves. Above these processes
there are small prominences which filled pits in the inner walls
of the exoccipital bones. ‘These pits were entirely cleared, and
it is certain that they were not the beginnings of foramina;
their meaning is unknown.

The whole metencephalic portion of the cast is rather high
and narrow. It is possible that this is due in some degree to
crushing, but there is no indication of such crushing in the skull,
and it is probable that it is the true form. The whole cast is
very small relative to the size of the animal, and even assuming
that the brain occupied the whole cavity its size would be remark-
able, though after all it is not much smaller, relatively, than the
brain of Sphenodon or of an alligator.

It is difficult to make any satisfactory comparison of this
endocranial cast with the one made by Cope from the specimen
of Belodon buceros because of the unsatisfactory nature of his
figures, but some points can be made out. As can be seen by
the figures in this paper, the whole shape is different, the cast
from Belodon does not show the sharp downward curve posterior
to the middle region. The cast of Desmatosuchus is thinner
for its height and does not have so long a metencephalie portion.

ENDOCRANIAL CAST FROM A REPTILE 139

The epiphysis lacks the posterior prolongation, certainly it is
not ‘subquadrate’ in form, and the lateral processes rise in front
of, not at the sides of, the epiphysis. The olfactory tracts were
much broader. The optic nerves did not escape through distinct
foramina. Only the origin of the pituitary body is shown in
Cope’s figures, but he describes it as small and occupying a
fossa in the base of the cranial cavity. These characters support
the evidence afforded by the bones of the skull that Desma-
tosuchus must be placed, at least, in a distinct suborder from
the Phytosauria.

In considering the endocranial casts from the four very different
primitive reptiles figured in this paper, it is certain that the brains
of all had certain very distinctive characters in common. ‘The
brain cavity was relatively long and narrow with small devel-
opment of the cerebral hemispheres. The optic lobes and tracts
were too small to leave any distinct marks on the casts, though
they were probably well developed. The brain was sharply
elevated in the middle portion with large epiphysial or para-
physial processes. There was a sharp downward bend in the
posterior portion. There is a considerable degree of constancy
in the location of the origin of the cranial nerves. One thing
is especially noticeable—the large size of the pituitary body in
the giant forms. Tyrannosaurus was the largest of the cainivor-
ous dinosaurs; Diplodecus was one of the largest creatures that
has lived upon the earth; Triceratops was elephantine in size;
Desmatosuchus, though not so large as the dinosaurs mentioned,
was 10 or 12. feet long and probably a giant of its kind. In
all of these forms the brain is exceedingly small relative to the
body, but the pituitary body is very large relative to the size of
the brain. The connection between the size and activity of
the pituitary gland and the size and the proportions of the body
in mammals is well known. MHyperpituitarism results in large
size or the over-development of certain structures, as the fingers,
features, etc. It is equally well known to paleontologists that
one of the common variations among those which occur so
abundantly in the senile stages of any phylum is giantism.
The suggestion naturally rises that the disturbances which

140 E. C. CASE

arose in the phylum coincident with the development of adverse
conditions were primarily of a physiological character affecting
the deep-seated organs of the nervous system and through them,
and only secondarily, the superficial structures. The terato-
logical affections of the pituitary body which produced abnormal
structures in a normal phylum may have gradually become a
fixed character and resulted in normal giantism in certain groups.
Similar correlations between other glands of the body and such
characters as spines, horns, tusks, etc., which finally developed
into excessive overgrowths in the senile stages of various phyla
may have arisen in the same way.

Since this manuscript was prepared two excellent figures of
endocranial casts have been published: Osborn and Mook,
Memoirs American Museum of Natural History, New Series,
volume 3, part 3, plate LXIV, Endocranial cast of Camarasaurus
supremus; Lambe, Canada Department of Mines, Memoir 120,
fig. 27, Endocranial cast of Edmontosaurus.

PLATE 1

EXPLANATION OF FIGURES

la Lateral view of endocranial cast of Belodon buceros. After Cope.
1b Upper view of the same. Both figures X 4.

ABBREVIATIONS |
Ep., epiphysis Olf., olfactory tract
Orb.Pin., orbitopineal process Cb., cerebellum
Cer., cerebrum II,V, cranial nerves

2a Upper view of endocranial cast of Diadectes phaseolinus. After Cope
2b Lateral view of the same. Both figures X 4.

ABBREVIATIONS
Olf., olfactory tract Cb., cerebellum
Cer., cerebrum V, cranial nerve
Ep., epiphysis Ot., otic tract

3a Lateral view of endocranial cast of Tyrannosaurus rex. After Osborn.
3b Upper view of same. Both figures X }.

ABBREVIATIONS
Olf., olfactory tract I to XII, cranial nerves
Cer., cerebrum hi and h2, epiphysis

Opl., optic lobe

ENDOCRANIAL CAST FROM A REPTILE PLATE 1
E, C. CASE

(onto
Hill
u

143

PLATE 2

EXPLANATION OF PLATES

4 Lateral view of endocranial cast of Diplodocus longus. After Osborn.
She

ABBREVIATIONS

Sac.End., saccus endolymphaticus II to XII, cranial nerves
Int.Car., internal carotid arteries

5a Upper view of endocranial cast of Triceratops serratus. After Hatcher

from Marsh. .
5b Lower view of same. Both figures X }.

ABBREVIATIONS
Olf., olfactory tract Ve., vein
Cer., cerebrum Pit., hypophysis
Cb., cerebellum IT to XII, cranial nerves

6a Lower view of endocranial cast of Triceratops serratus. After Hay.
This and the two following figures give Hay’s interpretation of the casts, as
opposed to that given by Hatcher.

6b Upper view of same.

6c Lateral view of same. All figures X i.

ABBREVIATIONS
Olf., olfactory tract Car., carotid artery
Cer., cerebrum Ju.v., jugular vein
Ve., vein IT to XII, cranial nerves
Pit., hypophysis Ep., epiphysis

Op.a., base supposed ophthalmic artery

144

ENDOCRANIAL CAST FROM A REPTILE PLATE 2
E. C. CASE

145

PLATE 3

EXPLANATION OF PLATES

7 Lateral view of endocranial cast of Dematosuchus spurensis.
8 Upper view of same. ;
9 Lower view of same. All figures X }.
ABBREVIATIONS
Ep., epiphysis Ca., carotid artery
Lat.proc., lateral processes Hyp., hypophysis

Olf., olfactory tract II to XII, cranial nerves

146

ENDOCRANIAL CAST FROM A REPTILE PLATE 3
E. C. CASE

Vit VU.

typ.

IX,X,Xl.

147

Resumen por el autor, J. M. D. Olmsted,
University of Toronto.

Los efectos de la reseccién del nervio lingual del perro.

Los botones gustativos desaparecen de las papilas fungiformes
en la parte anterior de la lengua del perro como resultado de la
resecciOn del nervio lingual. Solamente desaparecen los botones
gustativos del lado de la lengua inervado por el nervio que se
ha cortado, mientras que los del otro lado persisten sin cambio
alguno. Este resultado se debe a la reseccién del nervio y no
a otros trastornos producidos por la operacién. El proceso que
sigue a la seccién de dicho nervio es un proceso de degeneracién
acompanhada de la accién fagocitica de los leucocitos, no un
proceso de desdiferenciacion o metamorfosis. El punto que
ocupaban los botones gustativos viene a ser reemplazado por
células epiteliales.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, MAY 23

EFFECTS OF CUTTING THE LINGUAL NERVE OF THE
DOG

J. M. D. OLMSTED
Department of Physiology, University of Toronto

SIX FIGURES

It has been found that cutting the glossopharyngeal nerve of
the dog results in the disappearance of the taste buds in the cir-
-cumvallate papillae on the side of the tongue innervated by the
severed nerve (Vintschgau und Héningschmied, ’76; Vintschgau,
80; Drasch, ’87; Sandmeyer, 795; Semi Meyer, 796). Those
investigators who studied the manner of disappearance of the _
taste buds, Vintschgau and Meyer, were convinced that degenera-
tion did not take place, but rather a process of dedifferentiation
or metamorphosis transposed the sense cells into indifferent
epithelial cells. Ranvier (’88) performed this sane operation
on the rabbit and claimed that the sense cells did undergo degen-
eration and were completely destroyed on the spot. In another
paper (Olmsted, ’20) I have described the changes occurring in
the taste buds situated on the barbels of the fish, Amiurus nebu-
losus, after cutting the appropriate branches of the facial nerve.
With the fish, as with the rabbit, there were distinct degenera-
tive processes observable at a certain time after the operation,
accompanied by phagocytic action of leucocytes. I suggested
that similar degenerative changes would probably be observable
in the dog if one could happen upon a preparation taken at
just the correct interval after severing the nerve.

The mammalian tongue is innervated by two nerves, the
glossopharyngeal, supplying the posterior third (including the
circumvallate papillae), and the lingual, supplying the anterior
two-thirds. All the operations on mammals to which reference
has been made were on the glossopharyngeal nerve. This paper

149

150 J. M. D. OLMSTED

gives the results of severing the lingual nerve and shows the
correctness of. the supposition that the taste buds would dis-
appear through a process of degeneration—not dedifferentiation
or metamorphosis.

The dogs were anesthetized, and after shaving the region
under the jaw, a slit was made through the skin at the inner
border of the jaw, usually on the right side. The thin mylo-
hyoid muscle was cut through and the fascia separated so as to
disclose the loop of the lingual nerve just peripheral to the
branch to the submaxillary gland. This can be done with prac-
tically no interference to the blood supply of the tongue. Only
a small amount of bleeding takes place in the skin. A small
piece of the nerve, about 5 mm. in length, was excised. The
cut edges of the mylohyoid muscle and the fascia were sewn
together with sterile gut, and some six stitches were sufficient to
close the skin over the wound. Often the dogs began to eat
within six hours after the operation and none of them seemed
seriously inconvenienced, being able to move their tongues in a
fairly normal fashion. Theré was never any infection or sup-
puration. Swelling did occur, but this disappeared before the
end of a week.

When sufficient time had elapsed after an operation the dog
was killed and the tongue removed. The taste buds on the
anterior part of the tongue are situated on the fungiform papillae.
These papillae occur singly and at irregular intervals over the
surface of the tongue, being more numerous at its tip. With a
a little care they can be recognized, especially with the aid of a
hand lens, and removed from the tongue together with a few of
the encircling filiform papillae. Ten or more fungiform papillae
were taken at random from the dorsum and sides of the anterior
half of the tongue on the side corresponding to the severed nerve,
and also for control a similar number from the unoperated side.
These excised papillae were fixed in Zenker’s fluid containing
acetic acid, in formol-Zenker, or Heidenhain’s osmic-acetic-sub-
limate mixture (Heidenhain, ’14). Sections were cut 8 y» in
thickness. The usual haematoxylin stains were used.

LINGUAL NERVE OF THE DOG PE

No pronounced changes in the taste buds from the operated
side could be detected after twenty-four hours, nor even after
three days, except perhaps in the latter case there might be a
diminution in size of the taste bud, as if the cells were not so
well filled out as normally. In one dog which was killed the
eighth day after operation there were found eighteen fungiform
papillae without a taste bud and with no particular arrangement
of the epithelial cells to even suggest where taste buds had
been; three papillae each with the remains of one taste bud, as
in figure 4; and two papillae each with one perfect taste bud,
and one of them having also the remains of a second. On the
unoperated side ten fungiform papillae were examined. Each
of them had two to five taste buds in perfect order, with an
average of three to a papilla. When these were compared with
taste buds from the tongues of normal (unoperated) dogs no
differences could be detected. Forty fungiform papillae from
unoperated dogs were examined, and with one possible excep-
tion none contained less than two taste buds, and there was an
average of four to each. This possible exception was a very
small papilla resembling a filiform papilla in the thickness of its
cornified layer, but in its shape a fungiform papilla. This par-
ticular papilla was the only one of the forty which possessed no
taste buds.

In one very large dog the following record was obtained fifteen
days after operation. On the operated side there were six fungi-
form papillae without sign of a taste bud; three with the arrange-
ment of epithelial cells showing the former position of taste buds,
and two having the remains of one taste bud and also a single
perfect one in each. On the unoperated side five papillae were
examined. Hach had taste buds in perfect order with an aver-
age of five to a papilla. Two other dogs gave similar results.

To test whether the disappearance of the taste buds was due
to the injury incident on the operation, one dog was treated
exactly as the others, 1.e., the lingual nerve was disclosed, etc.,
but the branch to the submaxillary gland was cut between the
lingual nerve and the salivary gland, and the lingual left intact.
Kleven days after the operation, seven fungiform papillae from

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 2

152 J. M. D. OLMSTED

All the figures were drawn under a camera lucida at a magnification of 1500
and reduced one-half in reproduction. The boundary of each taste bud is marked
by a heavy line. c.l., cornified layer of epidermis; g./., germinative layer of epi-
dermis; g.m., granular mass of extruded cytoplasmic debris; l., leucocyte; s.c.,
sense cells.

LINGUAL NERVE OF THE DOG 153

the operated side were examined, and each possessed taste buds
in good order, having an average of four to a papilla.

The process by which the taste buds disappear is shown in the
figures. One finds in many of the taste buds on the tongues of
unoperated dogs leucocytes which resemble the large lympho-
cytes of the blood. In a very few cases I observed polymor-
phonuclear leucocytes within the taste bud, as in figure 2. The
first portion of the taste bud to disintegrate is the region just
above the nuclei of the sense cells (figs. 3, 4). This is exactly
similar to what I found in the fish. Furthermore, the debris
seems also to be extruded through the pore by which the taste
bud communicates with the surface, for, as figure 5 illustrates,
there was found at the pore of one taste bud a mass of granular
material, evidently cytoplasmic, since it stained faintly with
eosin, exactly similar to that inside the taste bud. Within this
mass were two nuclei, one evidently belonging to a leucocyte.
In the dog whose taste buds were examined on the eighth day
after operation there were many cases of mitosis observed, in
one instance three in a single field. In the forty fungiform
papillae from normal dogs there was not a single case of mitosis,
nor were there any in the neighboring filiform papillae in the
operated dogs. ‘The final process consists in filling up the site
of taste buds with epithelial cells (fig. 6).

In a few cases where degeneration had evidently occurred
recently, the boundary of the taste buds was still fairly evident
in the form of elongated cells, as contrasted with the more or

Fig.1 Longitudinal section through normal taste bud from fungiform papilla
of dog.

Fig. 2 Tangential section through normal taste bud, showing presence of
leucocyte.

Fig. 3 Longitudinal section through taste bud of dog whose lingual nerve
had been cut eight days before. The area surrounding the leucocyte is entirely
empty. The whole bud is considerably shrunken.

Fig. 4 Longitudinal section similar to figure 3. Distal portion of taste bud
empty except for a leucocyte and a disintegrating nucleus. The more proximal
part is filled with granular material.

Fig. 5 Tangential section through same taste bud as figure 4. Mass of
granular material lying at the neck of the pore of the taste bud.

Fig. 6 Fifteen days after operation. Taste bud indicated by arrangement
of cells. Possible boundary indicated by broken line.

154 J. M. D. OLMSTED

or less hexagonal cells inclosed by them. In other cases there
were no evidences whatever of taste buds. In all respects is
this similar to what was found in Amiurus.

SUMMARY

1. Taste buds disappear from the fungiform papillae on the
anterior part of the dog’s tongue as a result of cutting the lingual
nerve.

2. Taste buds disappear only on the side of the tongue corre-
sponding to the severed nerve, those on the other side remaining
in perfect order.

3. The process by which the taste buds disappear is one of
degeneration with the aid of phagocytic leucocytes, and not one
of dedifferentiation or metamorphosis.

4, Epithelial cells take the place of the former taste buds and
during this process there is marked proliferation by mitosis from
the germinative layer.

BIBLIOGRAPHY

Drascuo, O. 1887 Untersuchungen iiber die Papillae foliatae et circumvallatae
des Kaninchen und Feldhasen. Abhand. Sachs. Gesellsch. Wissensch.,
Bd. 24, S. 229-252.

Heipennain, M. 1914 Uber die Sinnesfelder und die Geschmacksknospen der
Papilla foliata des Kaninchens. Beitrige zur Teilkérpertheorie III.
Arch. f. mikr. Anat., Bd. 85, 8S. 365-479.

Meyer, 8. 1896 Durchschneidungsversuche am Nervus glossopharyngeus.
Arch. f. mikr. Anat., Bd. 4, 8. 96-110.

Ousted, J. M. D. 1920 The results of cutting the seventh cranial nerve in
Amiurus nebulosus (Lesueur). Jour. Exp. Zoél., vol. 31, pp. 369-40.

Ranvier, L. 1888 Traite Technique d’Histologie. Paris, 1109.

SanpMeyerR, W. 1895 Uber das Verhalten der Geschmacksknospen nach Durch-
schneidung des Nervus glossopharyngeus. Arch. f. Anat. u. Physiol.,
physiol. Abt., Jahrg. 1895, S. 268-276.

Vintscueau, M. von 1880 Beobachtungen tiber die Verainderungen der
Schmeckbecher nach Durchschneidung des N. glossopharyngeus.
Arch. f. gesam. Physiol., Bd. 23, S. 1-13. .

VintscHGau, M. von, unp HéniascumiepD, J. 1876 Nervus glossopharyngeus
und Schmeckbecher. Arch. f. gesam. Physiol., Bd. 14, S. 443-448.

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Los nervios espinales ventrales del Amphioxus.

Los nervios espinales ventrales del Amphioxus son nervios
delgados y largos, y no estructuras cortas en forma de pincel o
ramillete, extendiéndose a lo largo de todo el miotomo. Cuando
los fasciculos terminales mds pequefios abandonan el nervio,
se curvan fuertemente para entrar entre las placas musculares
y se separan sobre la superficie de las fibras musculares formando
las Ultimas fibrillas nerviosas que penetran dentro del musculo.
Los nervios espinales dorsales emiten ramas que entran en los
espacios inter-miot6émicos y envian finos nervios dentro del
cuerpo de los miotomas, probablemente de naturaleza sensorial.
La semejanza de los nervios espinales ventrales del Amphioxus
con los nervios ventrales motores de las formas mds superiores
es, pues, bien manifiesta.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHOR'S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, MAY 9

VENTRAL SPINAL NERVES IN AMPHIOXUS
HOWARD AYERS

SEVEN FIGURES

The spinal cord of Amphioxus presents a sinuous outline,
when viewed either from the dorsal or ventral face. This
sinuosity is due to the curved projections of the cord (fig. 6)
where the motor roots issue, and since they emerge, as a rule,
alternately on the right and left sides, the superficial curvatures
result. The main structures of the cord are not affected thereby.

The motor roots are composed of fibers which lack protective
covering of any kind. The number of fibers varies from about
100 to 250, in keeping with the variation in the size of the myo-
tomes. The larger the myotome the greater the number of
muscular elements requiring nerve control. The terminal myo-
tomes in head and tail are the smallest of the lot and the size
increases in both directions toward the middle of the body.
The motor nerves increase in size in the same way—the increase
in size being due solely to the addition of nerve fibers.

It has been stated that no motor nerve has been found for
the first myotome. I have dissected the first myotome and
find the nerve supplying it small, and could locate with cer-
tainty only two branches, one for each fork of the myotome.

Rhode’s description of the motor nerve is more complete and
accurate than his figures, but is far from correct either as to the
size and the course of the nerve or its method of branching. It
is, however, the best I have seen.

He describes three sections of the motor nerve, an anterior
which spreads like a fan on the dorsal part of the myotome, a
median section composed of ventral fibers which run to the
rectus abdominis and the ventral portion of the outer part of
the myotome, and a posterior section or bundle of fibers which

155

156 HOWARD AYERS

passes in between the rectus abdominis and the longus dorsi
and supplies both these muscles.

The current opinion seems to be that ‘‘the motor nerves
spread out like a fan and terminate upon the muscle fibers of
the myotomes,” to use Willey’s phrasing. This is not correct,
for the ventral spinal nerves of Amphioxus are relatively long
nerves and the muscle fibrils most distant from the cord are
controlled by means of nerve fibers which span the interval
between the spinal cord and muscle. The longest of these
fibers are, consequently, as least as long as the vertical diameter
of the animal, since the fibers run ventrocaudad in the ventral
fork of the myotome.

Another point should be kept in mind—the motor root divides
into nerve branches, distinct and compact bundles which, while
without noticeable sheaths, nevertheless hold their fibers firmly
together as nerves, to give them off as they pass along the terri-
tory of their distribution by the familiar method of nerve branch-
ing until the ultimate nerve filaments are set free singly or in
small groups of two to eight fibrils. These single fibers or small
groups are the elements which pass to the muscle for contact or
fusion and also for the formation of nerve nets upon the surface
of the muscle plates. .

It is common experience of those who have tried to lay bare
the connection of the ventral spinal nerves with the cord, that
the root too easily tears away from the cord at its junction
therewith. A dissection of a motor nerve with the parts in
place is shown in figure 1. The fibers issue from the ventro-
lateral angle of the cord as a wide, thin plate of fibers, which, as
Rhode has shown, is composed of groups of bundles of fibers.
This differentiation in the structure of the root I have followed
back into the spinal cord. It indicates a specialization of the —
nerve supply assembled from several parts of the cord for the in-
nervation of the structures under the control of the ventral roots.

The motor roots have been thought to belong exclusively to
the striated muscles of the myotomes but, as indicated in
figures 1, 2, and 3, there is a small bundle of fibers (relatively
too large in the figures) marked 5, which runs out with the

VENTRAL SPINAL NERVES IN AMPHIOXUS 157

large bundle 3, but which does not enter the myotome, passing
along its inner face to join the nerve plexus of the aortic trunk.
It is a splanchnic nerve. All the other branches of the ventral
nerve, so far as I have observed, are distributed to the myotome.

The four branches to the myotome are, counting from before
backward: nerve 1 leaves the anterior part of the root and runs
dorsolateral and enters the dorsal fork of the preceding myotome;
nerve 2 is the main supply of the dorsal fork of the myotome
belonging to the root, it runs between the two sections of the
muscle and extends the entire length of the muscle; nerve 3,
the largest and longest given off by the ventral root, enters the
posterior edge of the ventral fork of the myotome belonging to
the root, between the closely applied surfaces of the longus dorsi
and the rectus divisions of the muscle ventrad of the junction
of the two forks and runs the entire length of the ventral fork;
nerve 4 runs caudad and enters the tip of the ventral fork of the
next succeeding myotome; while nerve 5, as stated above, runs
into the body to join the nerve plexus associated with the cen-
tral vascular mechanism. ‘The ventral spinal nerves are there-
fore composite nerves supplying not only the trunk muscles, but
also visceral organs.

In life, the muscle plates are semifluid and are applied close
to the connective structures forming the walls of the base of the
dorsal fin, the spinal cord, the notochord and the sheet that
forms the supporting wall of the body cavity. The trunk muscle
is attached to this connective-tissue skeleton and some of the
fibers in the vicinity of the motor nerve interdigitate among the
fibers of the issuing ventral root.

The fact that muscle fibers are attached to the dural sheath
and penetrate among the bundles of nerve fibers of the ventral
root was largely responsible for the controversy as to whether
the motor fibers are striated like the muscle fibers. None of the
nerve fibers show striation. Rhode’s figure 34 shows two bun-
dles of muscle fibers which he calls striated nerve fibers. These
belong to the group of muscle fibers indicated in figure 4, M’’,
some of which attach direct to the dura.

158

HOWARD AYERS

ABBREVIATIONS

A-B line indicates the position of the
aorta with reference to the myotomes

C, sp:nal cord

D, dorsal spinal nerve

L, subdural lymph space

M, body of the myotome adjacent to
the ventral root

M’, muscle fibers inserting close upon
the motor root

M’’, muscle fibers inserting on the dura
and among the fibers of the ventral
root

V, ventral spinal nerve

d, ramus dorsalis of dorsal spinal nerve

1, first branch of motor root, runs to
preceding myotome

2, second branch of motor root, runs to
dorsal horn of myotome

3, third branch of motor root, runs to
ventral horn of myotome

4, fourth branch of motor root, runs to
succeeding myotome

5, fifth branch of motor root, runs to
aortic plexus

VENTRAL SPINAL NERVES IN AMPHIOXUS 159

The neuromuscular mechanism thus arranged insures coérdi-
nation of the contraction of the myotomes of one side as a physi-
ological unit. Besides the innervation by the motor root there
is a further innervation by the branches of the dorsal spinal
nerve through the ramus ventralis. These branches pass into
the myotome where the visceral ramus of the dorsal nerve crosses
the interspaces between the myotomes (fig. 7) and enters the
muscle. These nerves probably terminate in the connective
tissue, but I have-not seen their endings.

Another point of interest in connection with the dorsal roots
is the group of branches given off between the so-called dorsal
branch and the point where they pass lateral of the myotomes
(fig. 7). These branches I have not followed to their endings;
they are mainly distributed to the connective-tissue structures
between and about the mesial surfaces of the myotomes.

Here is material out of which the dorsal nerves of the higher
vertebrates could be made up. By a reduction of the branches
outside the myotomes accompanied by an increase in these
mesial branches, the characteristic relation of the dorsal root in

Fig. 1 Dissection showing spinal cord of Amphioxus with dorsal and ventral
roots and the branching of the motor nerve. The cord is represented as trans-
parent to show the sweep of the motor fibers as they assemble to leave the ventral
angle of the cord. The horns of the myotome are placed so as to show the motor
nerve to advantage.

Fig. 2 Dissection of myotome of Amphioxus seen from mesial face to show
motor nerve and its branches1to5. The rectus (mesial) portion of the myotome
is removed. The drawings fail to reproduce the translucent, silver-gray color
of the nerves and the delicate outlines of the thin plates before they break up
into the smaller nerve bundles. The branches shown in the figure are only the
main bundles which lie between the lateral and mesial halves of the myotome.
The innumerable branches given off from these to pass immediately among the
muscle plates to their motor endings are not shown.

Fig.3 Dissection of motor roots of Amphioxus, showing three adjacent nerve
roots and their course in the muscles. The myotomes and ventral roots are sep-
arated longitudinally, hence the figure shows the nerves connecting adjacent
myotomes as stretched to the extent of the separation of the myotomes. The
double line A-B, indicates the position of the aorta in this region, to which
branches of the motor roots make their way. The arrow points cephalad. The
heavy lines represent the motor roots as they issue from the dura. They are
displaced to the right, their normal position is along the line of junction of the
ventral and dorsal forks of the myotome.

160 HOWARD AYERS

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Fig. 4 Dissection of motor root of Amphioxus to show the motor fibers leav-
ing the spinal cord, C, to pass out to their endings. M, group of muscle plates;
M’, muscle elements inserting among the nerve fibers; 1/’’, muscle elements in-
serting on and near the dura.

Fig. 5 Four illustrations of the motor nerve terminals in Amphioxus. A .
shows a group of fine terminal fibers crossing a muscle plate; striation of muscle

VENTRAL SPINAL NERVES IN AMPHIOXUS 161

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fibrils indicated. A four-branched end-plate. is shown on one fibril. B shows
approach and terminal branching of motor nerves on ends of muscle fibers, also
one end-plate of nerve with lateral approach. C, muscle fibers more highly magni-
fied, showing motor end-plate formed by one branch of motor nerve which broke
up into six terminal fibers. D, nerve net formed by terminal fibrils of motor
nerve on surface of muscle plate.

Fig. 6 Dorsal view of dissection of spinal cord of Amphioxus, showing two
dorsal and one ventral root. Of special interest’ is the dorsal root which issues
from the cord, dorsal to the ventral root and on the same side of the cord. The
bulbous swelling of the cord at the place of issue of the ventral root is shown.

Fig. 7 Dissection of three myotomes, two dorsal nerves and one ventral, to
show relation of the latter.to spinal cord and to the muscle segments. The
group of branches from the dorsal nerve given off near the place of issue between
the myotomes is largely submyotomic, only the distal parts arriving at the
surface of the myotomes. The branches from the nerve given off where the
nerve trunk crosses over the interseptal spaces are indicated; some of these go
to the body of the myotome.

162 HOWARD AYERS

the higher forms would come into being. I think this transfor-
mation has taken place in the evolution of the vertebrate stock.

Regarding the termination of the fibers leaving the cord in the
ventral root, it is plain that not all of them form end-plates on
the muscle (fig. 5). Some of them form nerve nets which lie
superficially upon the muscle plates and are apparently inter-
muscular fibers. There are, however, gradations between a few
anastomoses between adjacent fibers of a bundle and the extreme
of complicated nerve net. In the former condition I have found
anastomosing fikers to terminate in end-plates, but where an
extensive net is formed I have not seen terminals leaving it to
enlarge into end-plates. They may occur, nevertheless, as in a
few instances fibers from the superficial net seemed to penetrate
into the muscle plate (fig. 5, D). The figures of Kutchin and
Dogiel drawn from methylen-blue and Golgi preparations do not
show the actual structure of the end-plates.

My observations of the neuromotor mechanism of Amphioxus
indicate that, while both nerve and muscle structure is much
less differentiated than in the higher forms, it is doubtless an
ancestral stage. The addition of sheaths would convert the
nerve fibers into the structures we find higher up. Bdellostoma,
Ammocoetes and Petromyzon furnish some of the intermediate
stages to this process of acquisition of protective coverings by
the nerve fibers. The same is true as regards the relation of the
dorsal and ventral roots. Amphioxus has them as separate and
distinct bundles throughout their root territory, but they are
found ending peripherally in the same territory (the myotomes),
but on different business. Again, Bdellostoma, Ammocoetes,
and Petromyzon present us with stages which show how the
condition in the higher forms has been arrived at.

The ventral root is relatively as extensive as in the higher
forms, 1.e., it runs to the extreme limit of the muscle organ to
which it is assigned and is a long branching nerve, not a brush
of fibers.

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March 21, 1921

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Resumen por la autora, Ada Roberta Hall,
University of Oregon.

Regeneracion del cord6én nervioso de un gusano anélido.

La autora ha llevado a cabo diversos experimentos sobre el
cordén nervioso de la lombriz de tierra, Helodrilus caliginosa,
mediante simple seccién y también mediante reseccion de varios
segmentos, con el fin de estudiar los tejidos comprendidos en
el proceso de la regeneracion, el papel de las células sanguineas
y el orfgen de las fibras nerviosas regeneradas. Las conclusiones
de estos experimentos son las siguientes: 1) Existe una rapida
formacion de material de cicatrizacion, el cual procede de las
células sanguineas, un tejido facilmente obtenible para llenar el
espacio producido por el corte. 2) Existe un crecimiento definido
de las fibras nerviosas a partir de los extremos del cordén hacia
la banda formada por el tejido de cicatrizacién. ‘También
pueden emigrar dentro de esta banda las grandes células gang-
lionares, si el corte se ha practicado a través de un ganglio,
reemplazando de este modo el tejido en su condicién normal.
3) Sise extirpan varios ganglios se forma una banda de cicatri-
zacion del mismo modo que en la regeneracién normal, para
llenar el espacio producido por el corte, y las fibras y células
ganglionares son reemplazdas por el crecimiento de los extremos
cortados. Después de dejar pasar un largo periodo (dos a tres
meses) para que contintie el crecimiento después de haber vuelto a
presentarse los movimientos normales, todas la células superfluas
son reabsorbidas de tal modo que el gusano aparece completa-
mente normal. Aunque el tiempo necesario para la regeneracion
parece notablemente rapido, las medidas tomadas por la autora
tienden a indicar que cuando se comparan con la regeneracion
de las fibras de los vertebrados, este tiempo tan corto en el
anélido se debe a la escasa longitud de la fibra mas bien que
a su crecimiento mas rapido.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, MAY 9

REGENERATION IN THE ANNELID NERVE CORD

ADA R. HALL
Zoological Laboratory of the University of Oregon

SEVENTEEN FIGURES (SIX PLATES)

CONTENTS

RTNECUITCHLON PSE HeNet aye ee. coe CASALL. Mehta PARES Meta Se Bae aco ASEM oe nee ies 163
DiscissrOnwoisuhesprOWleMminme a. kes er ie eee eter etnies olage cis pes cyeiscyens Sk 164
JN. IMeNiGribletnrcl aero lay, ocamer oscmucce poo ecue.s LoOmb ceo Sescesad Sc 164
COHEN CE cee epee ts Okan etre eats oe Bln. cemn ye oe wee ain nicioe © earns & 165
Geherenernvionraiter SSimiplereMts ool: 82 se ets kle ed «cee eantde sin 167
(PA SOUTCeLOR ileucleaiinix CCllSmpr ere. cee. 8 ous, 5 fers aisycbets ciojererere = apete 167

PR ECrOWUNROMMeTE VeRO CTSio® sakes arare ke dies 2 oe cieays S-evothe teats a ole wes 172

Ser SUMMA nyRO MSM plese yenecaGlOles e:- te yeris an es oeeya tase 173

D. Regeneration when two to four segments are removed............... 173
foe NOLEVETAWOLKG yer ats Hein hie. 4 syd WS? Chern con tae ts See daie oe beat et 173
PemresentmexpenilmM ets seas ve thie oe orate acs s. ced cious eas) Stoaeus ted 174

3. Summary of regeneration—several segments removed........... 175
GEnerAINCONCIISTONS A ate Falah ete heh Fore alas oy ANd Sette Sab Memes sees 176
ID ISEUSSTONBOMTESUMGS sf a.crec ic) 6 oon yes chs ore O cick? NA NIEISS IS SIP iy hoe Se 176
DSL AGRI TT EN A htc eer ones BIA Recon. eee ise ae a wee ear 178

INTRODUCTION

The earliest work on regeneration in the earthworm is con-
cerned with the general growth of an individual or with the
grafting together of parts of the same or different species of
worms. Heschler (98) and Rand (’08) have outlined the
steps in the process when a new head is formed, four or five
segments having been removed. They consider that the ecto-
derm is responsible, at least in part, for the growth of the new
nerve material. Friedlander has shown that the pull of the
longitudinal muscles will cause coordinated rhythmical move-
ments in two pieces of worm when their only connection with each
other is a piece of string. Biedermann shows that the nerve
cord is important in that it may carry impulses through several

163

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 2

164 ADA R. HALL

segments when the muscles are dead and are fixed to prevent
‘pull.’ Bovard (17) says that not only does the nerve cord
carry impulses through several ganglia, but that muscle pull
reinforces these impulses in each segment so that the message
goes a long distance without losing its force. He has also in
his histological studies shown that when the nerve cord is cut
regeneration is very rapid, and that the fibers which are regen-
erated bear a very direct relation to the movements of the
worm, the locomotor movements depending on the fine central
fibers, which regenerate first, and the general collapsing move-
ments depending on the giant fibers which recover about twenty-
four hours later. My problem has been to discover in the proc-
ess of regeneration, a single cut having been made: 1) what
cells form the cicatrix region and unite the ends of the nerve
cord and, 2) whether nerve fibers actually grow through this
region from the ends of the nerve cord before impulses can pass
from one end of the worm to the other, or whether the muscle
layers serve as a bridge to get the impulses past the gap. In
connection with this, I have also studied different periods of
regeneration in worms from which two or three segments of
cord have been removed, in order to see if this process will throw
any light on simple regeneration.

DISCUSSION OF THE PROBLEM

A. Materials and methods

The worm used in these experiments is Helodrilus caliginosa,
commonly found in our fields and meadows, especially where
there is turf. They are kept several days between moist cloths
to free them from as much grit and dirt as possible. Cloths are
preferable to filter-paper, as they can be sterilized frequently
and they also prevent the fillmg of the digestive tract with
paper pulp—a hindrance in sectioning. When ready to operate,
the worms are anesthetized by placing them in 5 per cent alcohol
until movements practically cease; left longer they are not
strong enough to recover and may die before regeneration is
complete. The worm is next laid over a rounded surface, such

REGENERATION IN ANNELID NERVE CORD 165

as a cork, under a binocular microscope. A small obliquely
transverse slit is cut in the epidermis, a few metameres posterior
to the clitellum. Then with a needle sharpened to a knife
edge on one side, the nerve cord is picked up and cut.. Care
must be taken not to sever the ventral blood-vessel, as too
severe bleeding is injurious. The worm is then replaced in the
moist cloths.

When removing several segments of the cord, a transverse
slit is cut in the ventral epidermis as before, and then the skin is
carefully lifted up with the point of the scissors and cut longi-
tudinally for several segments. When placed under the binoc-
ular, the nerve is seen lying in the furrow made by the cut.
The sharpened needle is inserted and the cord cut. One can
then count down as many segments as he wishes, insert the
needle and cut again. The section of cord is removed with the
forceps and dropped into alcohol, which causes the ganglia to
form contraction knots. These may be counted as a check on
the number of segments removed. The skin wound usually
closes in from two to four hours. The jars and cloths in which
the worms are kept are thoroughly cleaned with hot water each
day to prevent infection.

B. Technique

In order to study the process of regeneration, a large number
of worms were operated on and then killed at intervals of fifteen
minutes for the first six and a half hours. This series was
stained by the Hardesty method no. 1, which gives good fiber
differentiation and is supposed to show mitosis, if any be present.
Erythrosin and toluidin blue were the stains used. Although
cellular and intercellular structures are clearly contrasted by
this method, the killing agent used is a poor one where delicate
scar tissue is involved, as the violent action of the absolute
alcohol tends to pull the cicatrix loose from the wound on one or
both sides. Another series gives fifteen-minute intervals of
regeneration, up to an hour and ten minutes, then for periods of
several hours up to four days. This series was killed in Perenyi’s
fluid and stained with iron haematoxylin and orange G.

166 ADA R. HALL

For a study of the time of regeneration ordinary haematoxylin
and eosin give very good slides. For a further study of cell
and fibrillar structure, different staining methods were tried.
Picrosulphuric acid or Perenyi’s fluid for killing agent, with iron
haematoxylin and orange G, gives good cell and fiber differen-
tiation. The slides should be very heavily stained and then
differentiated slowly for the best results. Another method
which gives good fiber stain is Lewis’ pyroligneous acid method.
Tissues are killed in von Rath mixture for eight days. They
are then washed in methyl] alcohol, placed in 50 per cent pyro-
ligneous acid solution for forty-eight hours, then washed in
absolute alcohol for several days. Paraffin sections were cut
10 vu thick.

Several silver-nitrate methods were tried, but with little
success. It was thought that the intravitam methylene-blue
method would give clear fiber areas. This method was tried
repeatedly, but the penetrability of the substance is poor where
the central nerve cord and its sheath are concerned. Even
when injected into the coelom and left for varying lengths of
time, there seemed to be no reaction to it in the cord.

For comparative study smears were also made. ‘These were
of three kinds: 1) pure blood from the ventral vessel at the ante-
rior end; 2) body fluid; 3) cicatrix fluid drawn from a cut which
has been open a few seconds. To get only pure blood it was
necessary to make a fine capillary pipette, lift the vessel on the
forceps away from all other fluids, and press the pipette into it.
The contraction of the vessel will force the blood up in the tube
rhythmically. The other fluids were easily obtained. These
smears were dried; some were stained by Wright’s method for
blood corpuscles, and others directly with erythrosin and toluidin
blue or Hardesty’s method no. 1. Films were also made and
allowed to stand for half an hour in a moist place in order that
the corpuscles might expand. These were then killed in cor-
rosive-acetic fixing fluid and stained with eosin and methylene
blue.

REGENERATION IN ANNELID NERVE CORD 167

C. Regeneration after a simple cut

1. The source of the cicatrix cells. In studying the source of
the plug cells formed when a cut is made in the body wall, all
of the tissues of the normal worm should be considered (fig. 1)
and comparison made with these same tissues in the regenerat-
ing worms.

The epithelium of the normal worm is of the simple columnar
type. Its appearance varies somewhat according to its position
on the metamere. On the outer rounded parts of the metamere
is a high columnar epithelium with many gland cells, large
rounded bodies closely packed with granules. In the furrows
are the low columnar type with few or no gland cells. When a
cut is made in the body wall the epithelium is one of the first
tissues to regenerate. Worms killed within a few minutes after
section show the edges of the epithelium turned inward (fig. 2).
This is due to the contraction of the circular muscles. In a very
short time—forty-five minutes to two and a quarter hours—the
wound is filled with a mass of cicatrix cells which extends up,
nearly to the digestive tract and fills all the space between the
cut ends of the different layers (fig. 3). The inturned edges of
the epithelium may be imbedded in this mass, but the epithelial
cells which are covered thus break down and degenerate without
leaning toward or migrating into the plug. Figure 13 shows
_ such a condition (D. Ep.). The cells at the outer edge lean out,
and by proliferation give rise to the new epithelium (R. Ep.).
More often the skin is not turned in so deeply and the growth
is from the cut end of the epithelial layer (fig. 14).

An early stage in the process is shown in figure 12. Here the
plug (P1.) fills the cut area, with a few strands of circular muscle
imbedded in it. The new low epithelial cells extend out a short
distance, but the central part of the plug is directly exposed.
As regeneration progresses the whole plug shrinks and the
epithelial cells grow entirely across (figs. 14 and 15). Rand (01)
has shown that these arise from the old epithelium and migrate
across the cicatricial area. Although the old cells tend to lean
in the direction of growth, yet there seems to be no proliferation

168 ADA R. HALL

of these cells inward through the plug toward the nerve cord.
We may conclude therefore that the epithelium does not give
rise to the cicatrix cells nor to the regenerating nerve cord, but
merely covers over the exposed area formed by the cut.

There are two muscle layers: the circulars, lying just beneath
the epithelium, forming a band around each metamere (fig. 1,
C. M.), and the longitudinals, which extend lengthwise, forming
a muscular cylinder just inside the circulars (ZL. M.). As stated
before, the first reaction of the muscles when a cut is made is
the contraction of the circulars. This tends to draw the cut
edges of epithelium together and also holds the. longitudinal
muscle ends toward each other. The formation of the plug is
very rapid; at first it merely fills the spaces, showing no direct
connection with the muscles. It is easily seen that these cells
are not proliferations of the muscle layers. As regeneration ©
progresses, the plug cells elongate and orient themselves with
their long axis in the direction of the muscle pull.

In figure 15 longitudinal fibers are seen between the cut ends
of the longitudinal muscles (L. M. F.), and between these and ©
the epithelium is an area of cut ends (C. M. F.), showing that
the circulars are also connected by these fibers. Friedlander
sugcests that the plug cells come from the muscles, and from the
shape of the nuclei after a day or two of regeneration one might
be led to think so. However, from the rapidity of formation
and the appearance of the cells in the earliest stages this is not ~
probable. At a very early stage (two and a half hours, fig. 3),
the plug is formed and closely packed with round nuclei, but
there is no particular direction to the fibers. At ten hours
(fig. 12) the nuclei are still large and round. Only in the later
stages do the nuclei elongate and the fibers show a definite
orientation. I believe that this is a mechanical effect, in part
at least, due to the pull of the old muscles after a definite amal-
gamation has taken place between their raw edges and the cells of
the plug. Figure 6 shows the close relationship between the
plug and the muscle fibers.

Heschler believes that the subsequent growth and regenera-
tion comes as a proliferation of the old muscle cells or of undif-

REGENERATION IN ANNELID NERVE CORD 169

ferentiated cells scattered through these layers, but since the
plug cells are mesodermal in origin, as I shall show later, and are
capable of changing shape under the influence of a steady pull
exerted along one axis, it seems probable that these plug cells
might also develop contractile fibrils and fill the gap caused by
the cut. More work is needed to determine whether this latter
step actually takes place or not. The early stages of plug for-
mation and cell elongation and orientation in response to muscle
pull, are clearly shown in many of my slides.

The digestive tract lies as a smaller tube within the cylin-
drical body wall. As a rule, it is not cut during an operation
and does not enter into very close relationship with the plug
cells. Occasionally small fragments of chlorogogue cells may be
distinguished, by their granular nature, in the plug, but their
position here is accidental rather than purposeful.

Between the digestive tube and body wall lie two other struc-
tures, the nerve cord and blood-vessels (fig. 1, N. C. and Sub.).
When the nerve is first cut the elasticity of its substance causes
the ends to draw as far apart as the mesenteries will permit,
sometimes even slipping through the sleeve in the septum into
the next segment (fig. 3). This causes the mesenteries to assume
a slanting position. In addition to the small vessels of the skin
and muscles, the subneural vessel is cut and there may even be
breaks in the ventral vessel. This allows blood to escape into
the segment where the cut is. Experiments have been made
with this blood to determine the part which it may play in the
regeneration process. When a cut is made in the body wall,
very little bleeding occurs, except from the large vessels. The
small vessels are apparently closed by the first drop of blood
which comes from the cut end. The closure of even the largest
vessels, dorsal and ventral longitudinal vessels, is very rapid.
We may say, then, that very little bleeding occurs when an
earthworm is cut.

If the ventral vessel be raised and a clean smooth capillary
pipette inserted, a large amount of blood may be drawn without
any closure of the cut. If this is run out under a cover-glass on
a clean slide, the cover being slightly raised by a paraffin V open

170 ADA R. HALL

at the point, the drop remains liquid for a long time. A fiber
inserted at the point will cause a current in the drop and the
action of the cells may be studied. There are a large number of
cells in earthworm blood which seem to have an amoeboid action
and may be likened to the leucocytes of higher forms. These
cells, when contracted, are fairly regular, but long delicate arms
of cytoplasm may be thrown out. ‘These cells tend to aggregate
in the liquid around any dust particles or fibers which may be on
the slide. As other corpuscles come along in the stream and
strike these clumps they also cling fast. Drew (710), in his
study of the molluse blood (Cardium norvegicum), finds that
agglutination takes place there also. He has caused these
amoeboid cells to agglutinate on cotton fibers. As the cells
strike each other, protoplasmic strands are formed between
them, and each cell may then attach to a different fiber. Grad-
ually shrinkage takes place and the fibers are drawn together.
The cells of the earthworm blood agglutinate around dust par-
ticles or fibers in a drop or with each other in smears, since
friction seems to be the causative factor in this process. Fig-
ure 7D shows such an agglutination taken froma smear. Figure
7B shows a contracted corpuscle with eosinophile granules; C is
such a cell in the expanded condition. There are also present
in the smears, cells whose cytoplasm shows basophile granules
(fig. 7A). :

The question then arises as to just what takes place in the
blood stream and wound area when a cut is made in the body
wall. Blood may flow on to a glass slide from a clean, smooth
pipette without forming a compact plug, but if the same blood
be poured from a blood-vessel into a cut in the body wall, a plug
is formed which prevents further flow. We know that mam-
malian blood is caused to clot by the action of the thrombokinase
in the cut tissues. If clam blood be brought in contact with
friction surfaces or cut tissues, its corpuscles agglutinate. Drew
believes that not only does friction of the cut surface cause this,
but that some substance formed by the injury hastens the proc-
ess. The earthworm blood seems to be very like the mollusc
blood in its power of agglutination, so that in all probability plug

REGENERATION IN ANNELID NERVE CORD 171

formation is due to similar conditions. The molluse plug is due
to the thick mass of agglutinated cells holding back the further
flow of liquid, rather than to the formation of a true clot.

The plug which forms at a cut in the earthworm may be
thought of, then, as a large mass of agglutinated cells, filling
all the wound region and attached to all raw surfaces through
this same power of agglutination. The subsequent shrinkage
of the cytoplasm brings the nuclei closer together, giving the
plug a denser appearance and also draws the cut edges toward
each other through shortening of the cytoplasmic strands.

A comparative study of cells from the cicatrix of different
stages shows some very interesting results. The cells from an
early cicatrix (fig. 9, forty-five mimutes’ regeneration) show a
marked similarity to the agglutinated cells of the blood smear,
both in size and staining reactions. With two days’ regenera-
tion (fig. 10) the cells are smaller and more compact, while the
cytoplasm shows definite fiber formation. With further growth,
the ‘cells show still more condensation, and the fibers become
orientated in the direction of pull of the muscle layers and nerve
cord. A comparison of figures 12, 18, 14, and 15 shows this
continuous growth of cicatrix cells and fibers and their relations
to other layers of the body. Figure 12, of ten hours’ growth,
shows large round nuclei. Figure 13, a day and six hours’
regeneration, shows a condensation, with definite fiber growth
and orientation. At one day and twenty-one hours the cells are
still smaller (fig. 14), while at three and a half days (average
time for recovery of normal movements after section of the
cord) the fiber growth and orientation show a distinction in form
for the two muscle layers (fig. 15, L. M. F. and C. M. F.).

A study of blood smears and films and a comparison of the
cells found there with the cells of the cicatrix at different stages
of regeneration lead to the conclusion that the plug is a mass of
agglutinated blood corpuscles whose cytoplasm forms definite
fiber connections with all raw surfaces and, through subsequent
shrinkage and orientation, like tissues become united and func-
tional in a very short period.of time.

ee ADA R. HALL

2. Growth of nerve fibers. ‘The next point to be considered is
the further growth in connection with the nerve cord. Does
the cicatrix strand, mesodermal in origin, differentiate into
nerve material, thus bringing about the early functioning of
that organ, or is there a growth of nerve fibers from the cord
itself? This mesodermal strand does form a bridge between
the cut ends of the cord which rapidly shortens through shrink-
age, but it seems more probable that the original nerve should
furnish the material for nerve connections. In a study of the
cord we find confirmatory evidence for this. The first step in
the process is seen in figure 4, two and a quarter hours’ regen-
eration. Here the cord has been cut between ganglia and is
now united by a definite strand. The cells of the ganglia at
either side have the appearance of leaning toward the cut region
and the fibers are thinner at each end of the cord where the
strand attaches, as if they were stretching also to fill the gap.
This region is characterized by few nuclei. The central part of
the strand, however, shows a large number of round nuclei,
indicating a comparatively small amount of fibers. This is the
region of the strand formed by the agglutinated blood-cells,
while the clearer, non-nucleated strand region is formed by
fibers from the nerve cord. As regeneration progresses the
strand shortens through shrinkage, and these fiber areas extend
farther and farther across this bridge. Now the nuclei, of
necessity, appear fewer, since the fiber growths have usurped
their place. In figure 5 this further fiber growth is seen, causing
the nerve part of the strand to appear lighter in color than the
muscle part. ‘The fibers in the cord also show varicosities which
are characteristic of nerve but not of muscle. When the cut is
through a ganglion the large ganglion cells move out into the
cicatrix at the same time as the fiber growth takes place, thus
building up again the ganglion which was destroyed. This
process shows even more clearly in the study of the regenera-
tion, when several segments are removed. This will be dis-
cussed more fully later. When the normal functions have been
recovered, the fibers extend clear across through the central
region and a different more hollow part appears dorsally connect-
ing the ends of the giant fiber (fig. 11).

REGENERATION IN ANNELID NERVE CORD Rio

The regeneration in the nerve cord consists, then, of two steps,
first, the formation of a strand of plug cells which unites the
ends of the cord and, second, a growth of fibers from the original
cord through the strand uniting the cells of the two ganglia
which were separated by the cut.

3. Summary of sumple regeneration. We may then summarize
the steps in the process when a simple cut is made in the body
wall of the earthworm and the nerve cord sectioned.

1. In a very short time (forty-five minutes to two and a
quarter hours) the wound is closed. This is accomplished by
means of a plug which fills all the spaces between the cut ends of
the tissues. The meshes of the plug are filled with corpuscles
and large clumps of agglutinated cells.

2. In this same short period of time a strand of the cicatrix
material unites the ends of the nerve cord, which may be as
much as the width of two metameres apart, due to the elasticity
of its fibers.

3. Shrinkage of the cells of the strand draws the oe of the
cord nearer and nearer together.

4. A growth of characteristic fibers takes place from both
ends of the cord as regeneration progresses, finally bridging the
gap. If the cut be through a ganglion, large nerve cells may
migrate into the connecting material reconstructing the destroyed
ganglion.

5. In the final stages the different layers are connected and
functioning, the muscle layers by a strand of the differentiated
plug cells, and the nerve stumps by a growth of fibers from the
old cord.

D. Regeneration when two to four segments are removed

1. Former work. In connection with my former study of the
time required for a simple cut to regenerate, I operated on sev-
eral worms, taking out from two to four segments of the cord.
Some very striking results weré apparent, but it was impossible
to verify these on account of lack of material. At that time I
allowed regeneration to progress to the point at which loco-

174 ADA R. HALL

motor movements were recovered. This required fourteen
days. The worm which regenerated for eleven days shows a
very curious manner of growth. <A strand of cicatrix cells has
united the cut ends of the longitudinal muscles. This strand
shows the first steps in the regeneration of that muscle layer.
But further, these same cells have entirely filled the space up to
the nerve cord, have become attached to its cut ends, and have
drawn together across the three-metamere gap. Thus the nerve
cord is united and lies on a definite strand of cicatrix tissue. The
circular and part of the longitudinal muscles around the cut
have been massed together between the strand and the epithe-
lium. The digestive tract also is folded and the mesenteries of
these three segments are pulled together so that they lie at an
angle. This would seem to indicate that the first step in the
regeneration process is the uniting of the cut ends by scar tissues.

In the worm which regenerated for fourteen days this drawing
together was not so pronounced, a longer stretch of strand lying
between the cut ends of the cord. The mesenteries were washed
away in staining the slide, so that proof of the length of the cord
was lacking: However, there was some bunching of the muscle
fibers between the cord and epithelium and the cord appeared
shorter than normal.

2. Present experiments. In my present work I have attempted
to verify these results, and also have allowed worms to live for
several months in dirt after recovery from such an operation, in
order to see if complete regeneration of all parts takes place.
Before placing them in the dirt the exact position of the cicatrix
is established by counting the segments from head to cut.

In these experiments worms were killed at different intervals
to get an idea of the steps in the process. Recovery of the
movements occurs in from eleven to fourteen days in most cases.
One worm killed after fourteen days had not recovered, but
shows some interesting facts. The whole ventral part of the
worm for five segments was filled in with cicatrix cells (fig. 16).
These show differentiation for the muscle layers, which is appar-
ent in the orientation of the cell nuclei. A delicate epithelium
covers the lower border. Contrary to expectation, however,

REGENERATION IN ANNELID NERVE CORD 175

there is no bunching apparent. The strand (S) is nearly as
long as the part of the cord removed and seems to have taken its
place. The septa (Mes.) are also practically in their proper posi-
tions, the middle segment of the three removed being shorter
than normal. In the strand (S) of the cicatrix lying between
the ends of the cord there are fewer nuclei and also there is a
distinct difference in the staining reaction, this part being lighter
than those each side of it. A closer study reveals the fact that
it is the fibrillar structures which stain differently, those of the
strand being more of a yellow with fine varicosities, while those
of the surrounding cicatrix are brown. Comparison of the
fibrillar structure of the strand and of the ends of the cord which
we know to be nerve fibers shows that color and structure are
the same, which seems to indicate a growth of nerve fibers.

After three months’ growth others of these worms were killed
and sectioned (fig. 17). The pieces are about ten to twelve seg-
ments long, the cicatrix being at the center of the piece. Such
worms show no sign of scar in epithelium, muscle, or nerve
tissue. The septa are all nearly parallel and the coelom free
from extra cell structures. The new ganglia are normal in
every respect except length, two of the regenerated ganglia
being shorter than normal. I know that two and three segments
were removed, from actually counting the ganglia on the bit of
cord excised. I also know, by counting back, that the part of
the worm which was sectioned contains the excised region of the
cord. Thus it seems that actual growth of nerve fibers takes
place, and that all superfluous cells are again absorbed, leaving
all the structures normal. Since the epithelium does not grow
inward when the body wall is cut, but merely covers the plug,
these replaced ganglion cells as well as fibers must come from
growth in the cord.

3. Summary of regeneration when two to four segments are
removed. 1. There is a massing of cicatrix cells into all parts of
the cut region as in simple regeneration. These connect all cut
layers and either draw the ends of the cord directly together by
means of a strand so that all the mesenteries and muscle layers
become massed or the strand fills in the vacant place in the cord
leaving the other layers in their normal positions.

176 ADA R. HALL

2. Differentiation of this plug material takes place. The
nuclei elongate and orient themselves in the direction of the
muscle pull. Definite fibers with a dark-staiming reaction are
formed here also. In the region of the nerve strand the nuclei
are fewer and the fibrillar material has a different staining
reaction—one which is similar to that of the fibrillar portion of
the original cord. This may occur as early as fourteen days
and is present before recovery of movements takes place, indicat-
ing the actual presence of nerve fibers before functioning can
take place.

3. Complete recovery takes place, with ganglion cells formed
in'each metamere of the completed cord and with the coelom
entirely free from extra cell structures.

GENERAL CONCLUSIONS

1. There is a rapid formation of cicatrix material which closes
the wound. This comes from the cells in the blood, a tissue
easily brought in to fill the gap.

2. There is a definite growth of nerve fibers from the ends of
the cord into the strand formed by the cicatrix material. Large
ganglion cells may also migrate into the strand if the cut is
through a ganglion, thus restoring the tissue to its normal
condition.

3. If several ganglia are removed, a strand is formed as in
simple regeneration to fill the gap, and fibers and ganglion cells
are replaced by outgrowth from the cut ends. If a long period—
two to three months—is allowed for further growth after the
normal movements return, all extra cells are absorbed, so that
the worm appears perfectly normal in every respect.

DISCUSSION OF RESULTS

This growth and regeneration of the annelid nerve cord seem
at first glance remarkable for the speed of recovery. The cord
is cut and in from four to six days the normal movements and
impulses are again present in the worm. From a study of the
process we may conclude that the central nervous system has

REGENERATION IN ANNELID NERVE CORD EE

regenerated nerve fibers which connect the ganglia in front of,
and behind the cut, so that normal impulses may pass. Experi-
ments and observations on the higher forms or in man show that
when a nerve is cut recovery of the lost function is very slow.
Does the annelid tissue have a more rapid rate of growth than
mammalian tissue? I believe not. In the earthworm the ganglia
lie close together, 0.4 mm. to 0.45 mm. apart. The nerve cell
processes extend from one ganglion to the next. When these
fibers are cut the axon to be regenerated is very short, therefore
recovery is rapid. In regenerating a simple cut, the fibers must
grow from one ganglion to the next—a distance of 0.4 mm. or
less. This takes four days, as a general rule, making a rate of
growth about 0.1 mm. a day. In the nerve regeneration of
higher forms the part cut is usually a peripheral nerve which
may have its ending several feet from the nerve cell, conse-
quently growth from the cut to the ending ‘takes a long time.
Regeneration has not been successfully brought about in the
central nervous system, presumably on account of the lack of
neurilemma sheaths, which, in the case of the peripheral nerve,
furnish a directing path. In the earthworm the plug, with its
fibrous connection, forms a path for the developing fibers through
practically the entire distance traveled.

Ranson (710), working on peripheral regeneration in the
sciatic nerve of the dog, has carefully outlined the steps in degen-
eration and regeneration of the nerve fibers. He reports a
growth of fibers for a distance of 4 mm. up the nerve and 13 mm.
down from the cut during a period of twenty-five days. He has
not worked out the length of time necessary for completion of
the process and the return of function. This shows a rate of
growth of 0.65 mm. a day. If such a growth of the peripheral
fibers of the dog can be compared to the growth of the central
nervous system of the earthworm, there is here a slower rate of
growth for the invertebrate than for the vertebrate.

Thus, although the process of regeneration in the annelid
appears at first to be remarkably rapid, the early recovery is
due to the short length of fiber replaced rather than to a more
rapid rate of growth.

178 ADA R. HALL

This work was undertaken at the suggestion of Dr. J. F.
Bovard at the zoological laboratory of the University of Oregon.
I wish to express my appreciation to him for his kindly. interest
and many helpful suggestions both in the experimental work and
in the revision of this paper. I am also indebted to Dr. C. H.
Edmondson for the assistance and inspiration which he has
given me.

BIBLIOGRAPHY

Bovarp, J. F. 1917 The function of the giant fibers in earthworms. U. of Cal.
* Pub. in Zool., vol. 18.

1917 a The transmission of nervous impulses in relation to locomotion
in the earthworm. U. of Cal. Pub. in Zool., vol. 18.

Drew, G. H. 1910 Some points in the physiology of Lamellibranch corpuscles.
Quart. Journ. of Micr. Sci., vol. 64.

FRIEDLANDER, BuNnepict 1894 Uber die Regeneration herausgeschnittenen
Theile des Centralnerven System von Regenwiirmern. Zeitsch. wiss.
Zool., Bd. 60.

Hauu, A. R. 1917 Regeneration in the nerve cord of an annelid worm. Senior
thesis. University of Oregon (unpublished).

Morean, T. H. 1901 Regeneration. New York.

Ranp, H. W. 1901 The regenerating nervous system of Lumbricidae. Bull.
Mus. Comp. Zool. Harvard Coll., vol. 37.
1901 The behavior of the epidermis of the earthworm in regeneration.
Arch. Entw.-Mech., Bd. 19.

Ranson, 8. W. 1912 Degeneration and regeneration of nerve fibers. Jour.
Comp. Neur., vol. 22.

PLATES

ABBREVIATIONS

B.M., basement membrane

C.M., circular muscles

C.M.F., plug cells oriented to circular
muscle pull

Ciz., cicatrix

D.Ep., degenerating epithelium

Ep., epithelium

Gang., ganglion cells

G.C., gland cells

G.F., giant fibers

Int., intestine

L.F., central longitudinal nerve fibers

L.M., longitudinal muscles

L.M.F., plug cells oriented to longi-
tudinal muscle pull

Mes., mesenteries

N.C., nerve cord

Pl., plug cells

PI1.F., plug fibers

R.C.M., regenerated circular muscles

R.L.M., regenerated longitudinal mus-
cles

R.Ep., regenerated epithelium

Sh., sheath of the cord

S., strand

Sub., subneural vessel

Vent., ventral vessel

G.Nep., glandular part of the nephrid-
ium

S.Nep., saecular part of the nephridium

179

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 2

PLATE 1

EXPLANATION OF FIGURES

1 Longitudinal section of normal earthworm, showing relations of epithelium,
muscle layers, nerve, digestive tract, and blood-vessels. Heidenhain’s haema-
toxylin and eosin. %X 107.

2 Longitudinal section of earthworm killed immediately after section of the
nerve cord, to show the relation of the cut ends of the cord before the regenera-
tion process starts. Iron haematoxylin and orange G. X 107.

3 Longitudinal section of earthworm killed thirty minutes after section of
nerve, to show early formation of plug and its attachment to the ends of the cord.
Tron haematoxylin and orange G. X 107.

REGENERATION IN ANNELID NERVE CORD PLATE 1
ADA R. HALL

‘f° IVa 8 ~ yy

} ly “Vh\ SS en,
La ps4
o) alo) i OS ee

rg PEE
Gang. oe - ign od Gathe S€OE6 9 9

Sut tlh ee ae

—

(Fit, aN

PLATE 2

EXPLANATION OF FIGURES

4° Longitudinal section of earthworm killed two hours and fifteen minutes
after section of the nerve. The plug has formed a definite strand between the
ends of the cord, and the beginnings of the migration and growth of nerve ele-
ments from the ends of the cord are apparent. Preserved by the Hardesty method
no; IF GK 107

5 longitudinal section of earthworm killed one day and twenty-one hours
after section of the nerve. This shows growth of the plug in relation to the
various layers of the body. The wound is entirely covered by the epithelium.
Iron haematoxylin and orange G. XX 107.

REGENERATION IN ANNELID NERVE CORD: PLATE 2
ADA R. HALL :

183

PLATE 3

EXPLANATION OF FIGURES

6 Detail of a cross-section of earthworm, regeneration complete, showing
the epithelial growth and the intimate relations of the fibers of the plug to the
muscle layers. Iron haematoxylin and orange G. XX 750.

7 Camera-lucida drawings of cells from blood smears and films, stained with
Wright’s stain for blood corpuscles. X 1700. A. Contracted corpuscle with
basophile granules. B. Contracted corpuscles with eosinophile granules. C.
Same as B showing cytoplasm expanded. D. Clump of agglutinated corpuscles.

184

PLATE 3

REGENERATION IN ANNELID NERVE CORD

ADA R. HALL

185

PLATE 4

EXPLANATION OF FIGURES

8 Clump of agglutinated blood corpuscles taken from a smear stained with
erythrosin and toluidin blue. X 1700.

9 Camera-lucida drawing of plug cells from the cicatrix of a worm killed
45 minutes after section of the cord. Stained with Hardesty’s method no. 1.
xX 1700.

186

REGENERATION IN ANNELID NERVE CORD ; -) Jeb tay!
ADA R. HALL

187

PLATE 5

EXPLANATION OF FIGURES

10 Camera-lucida drawing of plug cells from the cicatrix of a worm killed
two days after section of the nerve. Stained with iron haematoxylin and orange
Ge XS £700:

11 Detail of the nerve cord of a regenerated worm; time, four days. This
shows the position of ganglion cells and the characteristic varicosities on the
fibers. Iron haematoxylin and orange G. X 170.

12 A portion of the body wall was removed, 0.5 mm. wide and three meta-
meres long, in order to watch regeneration in the muscle layers and epithelium.
Worm killed after ten hours’ regeneration, showing the size and character of the
plug cells and the epithelial growth. X 480.

13. Same as figure 14. Worm killed after one day and six hours’ regeneration.

14 Cicatrix region after one day and twenty-one hours’ regeneration. This
worm had only a simple cut in the body wall. X 480.

188

REGENERATION IN ANNELID NERVE CORD

PLATE 5
ADA R. HALL

ie.

—" “ — . a — - we — =
en a te eigen = So a
ie ee neesiil he meee me ace =
— oe i ab ; FE

i
I 3

PLATE 6

EXPLANATION OF FIGURES

15 Same as figure 14. Worm killed after three days and twelve hours’ regen-
eration.

16 Worm killed fourteen days after removal of three segments of the cord.
Regeneration not complete. X 107.

17 Worm having three segments removed, grown in dirt for three months
after normal movements had been recovered. All tissues appear normal except
that two ganglia are shorter than the rest. X 107.

190

REGENERATION IN ANNELID NERVE CORD PLATE 6
ADA R. HALL

, 7
hy CMF <2 7

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THE JOURNAL OF COMPARATIVE NEUROLOGY,
VOL.33, NO..3, AauacusT, 1921

Resumen por el autor, Edward Horne Craigie.
La vascularidad de la corteza cerebral en la rata albina.

La vascularidad relativa de las cinco laminas que se disti-
nguen en una misma regién cortical es semejante en todas las
Areas corticales examinadas y en todos los casos la l4mina granular
interna (IV) eslamasrica. Las capas supragranulares presentan
una tendencia a ser mids ricas que las infragranulares, y la capa
mas pobre es la lamina multiforme (VI). Parece probable que
las capas granular y supragranular sean receptivas y asociativas
en funcion, mientras que las infragranulares dan lugar a fibras
coritcifugas. Esta particularidad invita a una comparacién con-
los centros inferiores, en los cuales los ntcleos sensoriales y
los de correlacién han sido encontrados mds vascularizados que
los nucleos motores. La vascularidad media de las cinco capas
es la misma en las dreas oecipital y temporal, y es solamente un
poco menor en la regién precentral. El drea parietal es clara-
mente mds rica en capilarres que las otras, mientras que la
corteza insular es la mds pobre. La rasculavidad de las cinco
laminas de la corteza insular es semejante en su extensién a la de
los diversos centros motores estudiados en el tallo cerebral y
médula espinal, mientras que la de las otras dreas corresponde
aproximadamentea la de los nticleos sensoriales y correlativos.
La vascularidad de la cortezacerebelar es préximamente del
mismo orden de magnitud de la de la corteza cerebral. En la
corteza las diferencias sexuales y raciales aparecen mds marcadas
que las otras parte sdel sistema nervioso central estudiadas pre-
viamente. El drea insular parece diferir de las restantes en
algunos puntos mucho mas que las demas dreas entre si.

Translation by José F. Nonidez
Cornell Medical College, New York

/

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, JUNE 27

THE VASCULARITY OF THE CEREBRAL CORTEX OF
THE ALBINO RAT

EDWARD HORNE CRAIGIE
Department of Biology, University of Toronto

FIVE FIGURES

The work upon which the present paper is based is an exten-
sion of that previously reported by the writer (’20). In the
former publication an account was given of the relative vascular-
ity of various parts of the spinal cord, brain stem, and cerebellum
of the albino rat, and this account is extended in the present
communication to include the cerebral cortex of the same ani-
mal. In such a connection, there comes up for consideration
not only the richness of the capillary supply of the cerebral
cortex as compared with that of the parts previously studied,
but also the relative vascularity of the various regions into which
the cortex has been divided by physiologists and histologists.
Also, little, if at all, behind these problems in interest comes the
question of the conditions in the various layers which character-
ize the cortex.

The more important accounts of the cortical localization in
rodents, in so far as they are based upon histological studies,
are reviewed by Sugita in his paper (717) on the growth in thick-
ness of the cortex. He also correlates the various regions in
which his measurements were made with the detailed description
by Fortuyn (’14) of the conditions in the Norway rat and with
the anatomico-physiological terms of Brodmann (’09).

Sugita found the cortex of the albino rat, as Fortuyn had
found that of the Norway rat, to be divisible, in typical localities,
into five laminae. His description of these laminae is as follows:

The cerebral cortex of the albino rat has five cell layers if a typical
locality be taken. The most external layer is the lamina zonalis
(1), which has a few scattered glia-cells. Under this, there is the lamina

193

194 EDWARD HORNE CRAIGIE

pyramidalis (III) consisting of typical, deeply-staining, pyramidal cells
lying closely together, which corresponds to the third layer of Brodmann
(09). In the rodent brain, the lamina granularis externa (II), or
second layer of Brodmann, is always indistinct, and it is almost im-
possible to distinguish it from the lamina pyramidalis (III). Beneath
the lamina pyramidalis, the lamina granularis interna (IV) is situated,
composed of crowded, deeply-staining, small granules, somewhat re-
sembling glia-cells. Below this layer, there is the lamina ganglionaris
(V), which has dispersed, large-sized, deeply-staining pyramids. Next
to the lamina ganglionaris, there is the lamina multiformis (VI) with
polymorphous cells.

MATERIAL AND METHODS

The material consisted of eight of the ten brains used in the
previous study (’20), to which reference is made above. These
were numbered 16, 23, 24, 26, 31, 55, 56, 58, for the last four
of which the writer is indebted to The Wistar Institute. The
forebrains of the remaining two specimens used in the earlier
work were, unfortunately, not in good enough condition for
study. A full account of the animals from which these brains
were obtained, as well as of the technic employed in preparing
them and of the method of measuring the capillaries, is given
in the former paper. It will be sufficient to repeat here that
nos. 23, 24, 31, and 58 were male albino rats, nos. 16, 26, 55,
and 56 females. Numbers 16, 23, 24, and 26 were obtained in
Toronto, while the remaining four, as mentioned above, were
secured at The Wistar Institute. The microscope, lenses
(Leitz no. 7 objective and no. 3 ocular), and square-ruled mi-
crometer employed in making the measurements were the same
ones as were used before, but owing to the thinnesss of the
cortical laminae, it was found much more convenient to measure
the capillaries enclosed in only one half of the large square on
the micrometer. Hence the measurements made were the total
length of the capillaries enclosed in an area of 3 x 189? sq. u
in each section, the length of one edge of the micrometer square
being 189u. As before, such measurements were made in the
same region of each of ten successive sections, and as these
sections were 20, in thickness, the sum of the (10) readings
gave the total length of the capillaries in a block of tissue of
the volume } x 189? x 200 ¢. u. These results are shown’in the
accompanying table (table 1).

VASCULARITY OF THE CEREBRAL CORTEX 195

The cortical localities selected for study are those general
regions in which Sugita’s measurements were made. The fol-
lowing is a list of these localities, using Brodmann’s terminology,
with the number of Sugita’s region to which the position of the
present measurements in each corresponds approximately, and
the designations of the areas used in figure 1.

BRODMANN’S TERM SUGITA’S NUMBER reser Neg ae
ERG MATA COMETS sah ain accyirvetas A fiee don odie oe II A
REPTONOCOlPIbaet a... ooh ness ones eed nde Ramen IV D
veriompamletalligt ctr cm staleam aac se one tetre sateen Vil B
PPLE PTOI AT IS 5 oa. aes secertecr hos domed winged gae VIII E
IREHio reMOORAIK Aadoae moter ooo. Gee on ene aero. XI C

Fig. 1 Lateral and dorsal views of the left half of the brain to show the
regions in which the measurements of vascularity were made. The exact loca-
tion and relations of each of the areas represented is based on a comparison of
figures by Fortuyn and Sugita, the names being applied in accordance with
Sugita’s correlation of Brodmann’s terminology with these figures. Comparison
with Brodmann’s own figures and description seems to indicate that the anterior
portion of area B is equivalent to the regio postcentralis. A, regio praecentralis;
B, regio parietalis; C, regio temporalis; D, regio occipitalis; E, regio insularis.

EDWARD HORNE CRAIGIE

196

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198 EDWARD HORNE CRAIGIE

The accompanying illustrations (fig. 1) show these areas with
the points where the measurements of vascularity were made
indicated approximately by the hatched spaces. The outlines
of the cortical regions are based upon a comparison of Sugita’s
reproduction of Fortuyn’s figures with the former author’s own
illustrations of sections, and his correlation of both these with
Brodmann’s terminology. The outline of the brain was taken
in the first place from two injected specimens which had been
carried only as far as the 70 per cent alcohol stage in preparation.

The laminae of the cortex were by no means easily distin-
guished in many cases, the simple picric-acid stain which was
employed not being well adapted for such a purpose. By careful
comparison, however, with a series of sections stained with
toluidine blue and erythrosin and with the specimens in which
the layers were more distinct, they were located fairly accurately,
it is believed, even in the more difficult cases.

Figure 2 shows the cell lamination and the vascular supply
in the occipital cortex of one rat.

OBSERVATIONS

The measurements obtained are presented in table 1. The
results for the eight brains have been averaged and the probable
error of the averages calculated, these figures being given at the
right-hand side of the table. The probability of error tends,
of course, to be a little higher than was the case in the previous
study on account of the use of only eight brains instead of ten.
The ratios of the averages to the readings for the ventral white
funiculus and the ventral gray cornu of the spinal cord in the
same individuals! are given in the last columns of table 3. The
sex of each rat has been indicated in table 1 by placing M. or
F. in front of its identification number, while its locality is
shown by adding ‘Toronto’ or ‘W. I.’ (The Wistar Institute).

In the preparation of the tables the mean values for the five
laminae in each area, as shown in table 1, were averaged, and
the areas were arranged in order of increasing vascularity, as
thus determined. These averages are recorded in table 2. The

1 See Craigie (’20).

199

OF THE CEREBRAL CORTEX

VASCULARITY

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Fig. 2 On the right, a drawing of the blood vessels in a small part ofjthe
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same piece of cortex.

iV,

lamina zonalis; JJ, lamina pyramidalis;

Sermo I

VI, lamina multiformis.

lamina granularis interna; V, lamina ganglionaris;

200 EDWARD HORNE CRAIGIE

average vascularity of the five layers is the same in the occipital
and temporal regions, and it is very little less in the praecentral
region. The parietal region is distinctly the richest, while the
insular is much the poorest.

With regard to individual variation, it is noticeable that the
same individuals tend to give rather high or rather low results
in all the cortical areas, but the values obtained in the lower,
subcortical centers of these individuals were not always similarly
high or low. These differences are reflected in the ratios of the
cortical values to those for the ventral white column and the
ventral gray cornu of the cord, which have been set down in a
detailed table, not published, from which table 1 has been con-

TABLE 2

Average vascularity of five areas of the cerebral cortex. (The averages of the values
shown for the five laminae of each area in the second last column of table 1)

CORTICAL AREA MPER 5 X 1892 X 200 ¢c.u
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vidual ratios referred to have all been averaged, and the results
are found to correspond so satisfactorily with the ratios of the
average readings, shown in table 3, that it is considered sufficient
to publish the latter.

The averages for the various layers of each area, as recorded
in next to the last column of the first table, are represented
graphically in figure 3, which illustrates the relation between the
different regions as well as that between the five laminae in each.

It will be observed that the relative vascularity of the five
laminae in the various areas studied is fairly constant, not only
as regards the averages, but even, though to a smaller extent,
in the different individuals, as shown in table 1. The greatest
irregularity which appears is in the case of R.56, in which the

201

VASCULARITY OF THE CEREBRAL CORTEX

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202 EDWARD HORNE CRAIGIE

lamina zonalis tends to be richer as compared with the other
layers than is the case in the rest of the brains studied. This
is believed to be due to an unexplained vacuolate appearance

Vascularity
mine Heme CBE gl
SEES a
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Re
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Fig. 3 Chart showing the relative vascularity of the five cortical areas
studied and of the five laminae in each.

T, lamina zonalis I, insular area

III, lamina pyramidalis O, occipital area i
IV, lamina granularis interna P, parietal area

V, lamina ganglionaris Pr

, praecentral area
VI, lamina multiformis

T, temporal area

in this specimen of. all the layers except the lamina zonalis.
It seems probable that this condition of the tissue has resulted
in the measurements for all the inner layers in R.56 being a
little lower than they should have been. As these results fell

VASCULARITY OF THE CEREBRAL CORTEX 203

within the range of variation of the other individuals, however,
it was decided not to discard them.

In all the five areas studied, the lamina granularis interna
(IV) is decidedly the richest, the lamina pyramidalis (III)
coming next, with the lamina ganglionaris (V) very little behind
it. The fourth in order of richness is the lamina zonalis (I),
while the lamina multiformis (VI) is the poorest in every region
except the insular. It may be remarked that the lamina granu-
laris interna, which is absent from the praecentral region of many
mammals, is distinct, though thin, in that portion of the cortex
of the rat, and it is interesting to note that it is not relatively
poorer in its capillary supply there than in the other areas.

These observations may be compared with the description of
Duret (’74) fifty years ago. He found that the outer 0.1 mm.
of the human cerebral cortex contained large quadrangular meshes
parallel to the surface, the next 2 mm. is filled with rather fine
polygonal capillary meshes, while the inner 1 mm. has a transi-
tional network with larger meshes, which, however, are much
less elongate than those of the white matter into which they pass.

There is little certainty at the present time regarding the
functions of the different layers of the cerebral cortex. Bolton
(00) noticed twenty years ago that, while the deeper layers did
not vary appreciably in thickness as a result of age or chronic
insanity, ‘‘there is an almost exact correspondence between the
thickness of the conjoined first and second layers? of the cortex
and the degree of amentia or dementia existing in the patient.”
Nine years later, Kappers (09) concluded upon the basis of
comparative evidence that ‘“‘the granular layer in the cortex
is primary in character, and has originally receptive functions,”’
that the subgranular layers (V, VI) have chiefly the functions
of projection and intraregional association, while the supra-
granular layers (II, III), which are the last to appear phyletically,
are concerned chiefly with association of a higher order (inter-
regional), including intellectual processes.

* The ‘second layer’ of Bolton appears to be equivalent to Brodmann’s laminae
II and III, i.e., the supragranular layers,

204 EDWARD HORNE CRAIGIE

Brodmann (loc. cit.) considers Kappers’ theory groundless,
describing it as a wild hypothesis which is quite untenable and
erroneous. Van Valkenburg (’13, 713 a[?]), however, has brought
forward certain evidence which seems to support Kappers’ view;
as have also Nissl, Nieuwenhuijse, and Bielschowsky, whose
contributions are summarized by Van’t Hoog (’20). The last-
mentioned author also adduces further evidence in favor of
the theory of Kappers, and stresses particularly the rdle of
the cells in the lamina granularis interna. These he considers,
emphasizing a point concerning which Kappers had been less
positive, to be ‘matrix cells,’ i.e., cells which have retained much
of their primitive character and potentialities, which are still
capable of a wide range of differentiation, and from which the
other layers have probably been derived phylogenetically. It
may be pointed out that, in the ontogeny of the cerebral cortex
of the rat, this layer is the last one to become distinguishable
according to Sugita (’17), but, on the other hand, it is said by
Tandler and Kantor (’07) to be the first to appear in the embryo-
logical development of the reptilian cortex. Thus practically all
the available evidence, whether it be conclusive or not, seems to
point in the direction outlined by Kappers.

It would be an interesting observation if it was really the
more recent and more highly specialized portions of the cortex
which were the less richly vascular, and we have possibly a
somewhat similar case among the lower centers studied. Refer-
ence to table 3 will show that the chief vestibular nucleus is
more highly vascular than the cerebellar cortex, the dentate
nucleus, or Deiters’ nucleus. Now the cerebellar gray matter
is a highly specialized derivative of the primitive acoustico-
lateral area, from which, of course, both the other nuclei are
also developed. Moreover, the chief vestibular nucleus is com-
posed of small, granule-like cells, most of the axones of which
are said to take up a longitudinal direction in the substantia
reticularis, so that it seems not unreasonable to suppose that
it may be less highly differentiated than either the nucleus of
Deiters or the cerebellum. This, however, is pure speculation
without definite authority or conclusive basis.

VASCULARITY OF THE CEREBRAL CORTEX 205

Kappers (loc. cit.) offers a suggestion, in another connection
altogether, which seems to provide a very plausible explanation,
at least in part, for the special vascular richness of the lamina
granularis, and perhaps also of the sensory nuclei. In discussing
the importance of the granule cells in relation to his principle
of neurobiotaxis, he points,out that, “while in projection cells
the nervous current is directly realized and led away, on the
contrary, in the granular cells with short axis cylinders forming
an intricate network the stimulation is kept within a certain
region.”’ He goes on to point out that the long ascending and
descending tracts very often end in relation with such cells,
citing among other examples the case of the sensory root fibers
ending in the medulla oblongata ‘‘and (less general) in the cord.”
Such a region of concentrated or localized activity might reason-
ably be expected to have a relatively rich blood supply, and no
doubt this is one at any rate of the factors giving rise to the
observed differences.

To facilitate comparison of the vascularity of the cerebral
cortex with that of the lower centers previously studied, table
3 has been prepared. In this table all the values have been
reduced to the basis of a cube of tissue of 100 » edge, so that
the figures given represent the total length of the capillaries in
a block of tissue of volume 1,000,000 c. u, or 0.001 ec. mm. This
not only makes easy the direct comparison of the figures in
the table, but also provides a unit with which comparisons
may readily be made in future studies. It may be remarked
that the ratios in the table were calculated from the original
readings, not from the reduced figures which are tabulated
beside them. These ratios show that, on the whole, the vascular
supply of the cortex is not much greater than that of the ventral
cornu of the gray matter in the spinal cord, but exceeds that of
the ventral funiculus of the white matter from three and a half
to over seven times.

It will be observed that the vascularity of the insular cortex
corresponds roughly with the values obtained for the motor
centers, while the various laminae in the other areas cover about
the same range as the sensory and correlation centers in the lower

206 EDWARD HORNE CRAIGIE

part of the brain, though the richest part of the cortex studied
is slightly poorer than the richest center lower down—the dorsal
cochlear nucleus. May it be that, great and intense as the
activity of the cortex probably is, that activity is nevertheless
more intermittent in any one area than is the activity of the
lower sensory nuclei? The latter, as was pointed out in the
previous paper, are in more or less constant receipt of stimuli,
but only a small proportion of these give rise to any reflex,
and only a still smaller proportion, when any, reach the cerebral
cortex. Many impulses are, no doubt, generated within the
cortex itself, and such generation may possibly involve a greater
expenditure of energy than the mere passage of an impulse
caused by a stimulus somewhere else, so that the cortex may
reasonably be expected to be relatively rich in most areas; but
the activity in any one portion of the cortex is probably less
constant than that in a sensory nucleus, so that greater vas-
cularity is not required.

It might perhaps be mentioneu here that what appears to
be a clear example of a direct relation between vascularity and
functional activity has been described recently in Cajal’s labo-
ratory by De Castro (’20), who found such a relation distinctly
shown in comparing the vessels belonging to the olfactory
glomeruli in man with those in macrosmatic animals.

It may be remarked that, while the ‘motor cortex’ (regio
praecentralis) ranks low among the five areas studied, it is very
little poorer than the oceipital and temporal areas, and is con-
siderably richer than the regio insularis.

Finally, we note that the values obtained for the vascularity
of the lamina zonalis in the four richer areas are very similar
to the figure representing the condition in the molecular layer
of the cerebellar cortex.

As in the previous study, the results for-the two sexes have
been separated and averaged, and the comparison of these is
made in the charts in figure 4. The difference between the
sexes is much more definite than it was found to be in the lower
centers, the vascularity in the males being greater in every
lamina of the parietal, temporal, occipital, and praecentral

VASCULARITY OF THE CEREBRAL CORTEX 207

areas, though the females surpass the males in three of the
laminae of the insular cortex. In the subcortical regions the
tendency was for the females to be richer than the males. It
is perhaps hardly justifiable to base any conclusion upon the
averages of so few individuals, especially when the difference is

Vascularlily

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Cortical saminae

Fig. 4 Graphs showing the relative vascularity of the cerebral cortex of the
two sexes in each lamina of the five areas studied. Male @. Female +.

~ not very great, being less than the amount of individual varia-
tion, and when the poorer group includes the specimen (R. 56)
for which the results are suspected of being rather low. Never-
theless, it seems distinctly suggestive that the sexual difference
should be so uniform and so much more definite in the cortex
cerebri than in the lower centers studied. :

208 ; EDWARD HORNE CRAIGIE

A comparison of the groups according to locality has also
been made, the average of the results for the four Toronto
animals being compared with that for the four specimens from
the standard colony of The Wistar Institute. The differences

Vascularity
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Fig. 5 Graphs showing in the same way as in figure 4 the relative vascularity
of the cerebral cortex in rats from two different localities. Toronto @. The
Wistar Institute +.

between these groups are illustrated graphically in figure 5.
Here again, the difference is a little more decided than in the
lower parts, though less definite than that between the sexes.
In this case, however, the suspected specimen (R. 56) belongs
‘to the group which is richer in most layers (the group from The

5000

VASCULARITY OF THE CEREBRAL CORTEX 209

Wistar Institute). It may be noticed that, in all cases where
the Wistar average is less than the Toronto one, it is the deepest
layers which are involved, the lamina multiformis being poorer
in the Wistar group in four of the five areas, the lamina gangli-
onaris in three, the laminae granularis interna and pyramidalis
in only one, and the lamina zonalis in none. It should be
pointed out in this connection that each local group contains
two specimens of each sex.

Evidently the difference between the regio insularis and the
other four regions is either of some other nature or of a much
greater degree than their differences from each other. It is
this area which is so much more poorly vascularized than the
others; it is this area in which the female average exceeds that
of the males in three laminae; and it is this area in which the
average for the Wistar animals is poorer in four laminae than
that for the Toronto animals.

It should be remarked here that it is possible that some of the
differences in the measurements of vascularity are possibly
attributable to differences in brain weight. The situation in
this respect is summed up by Donaldson? as follows:

If the vascular supply grows in proportion to the brain—within the
limits of variation of brain size in full grown rats—then a sample of a
fixed volume of tissue from one brain will be comparable with that from
another.

If the increase in the volume of the entire brain is more rapid than
that of the vascular supply, then the supply in a fixed volume of tissue
will appear less in the larger brain—and vice-versa.

Which of these conditions obtains, the writer is not at present
in a position to state definitely, but he is inclined to believe that
the second supposition is the one which represents the facts.
This would not account, however, for the differences in the rela-
tions seen in the different parts of the central nervous system.
Unfortunately, the weights of the brains employed in the present
study were not recorded, but the body weight and body length
of the four rats from The Wistar Institute are known. From
these data the probable brain weights may be determined from

3 Personal communication.

210 EDWARD HORNE CRAIGIE

the normal tables for the albino rat (Donaldson, 15, table 68),
which give the following results:

BRAIN WEIGHT | BRAIN WEIGHT

RAT SEX BODY LENGTH | BODY WEIGHT eye ges las Seen one
LENGTH WEIGHT
mm. grams grams grams
3l of 200 _ 228.4 1.858 1.900
55 9 185 159.2 1.782 1.773
56 je) 198 197.4 1.841 1.835
a8 rot 210 234.8 1.903 1.907

It is thus evident that the female brains may be regarded as
slightly smaller than those of the males. Since the vascularity
of the cerebral cortex is poorer in the female brain, rather than
richer, the sexual difference cannot be explained upon the basis
of size in the manner suggested above.

Since measurements were not recorded for the Toronto ani-
mals, a similar scrutiny cannot be applied to the comparison of
local groups. It seems not improbable that the difference in
this case may be capable of explanation on the above basis,
though this would not account for the greater difference found
in the cortex as compared with the subcortical regions.

While one must hesitate to generalize upon only two com-
parisons with the limitations already pointed out, these data
seem to suggest a greater susceptibility in the cortical vasculari-
zation than in that of the more ancient portions of the central
nervous system to differences either within or without the body
—sexual, hereditary, or environmental.

The writer is much indebted to Dr. H. H. Donaldson, of The
Wistar Institute, who kindly read the manuscript and made a
number of valuable suggestions regarding the presentation of
the material.

SUMMARY

1. The vascularity of each of the five distinguishable layers
in five different areas of the cerebral cortex has been measured
in eight albino rat brains out of the ten which were used in the
prosecution of a study, already reported (Craigie, ’20), upon

VASCULARITY OF THE CEREBRAL CORTEX AlN

the relative vascularity of various parts of the cerebellum,
medulla oblongata, and spinal cord.

2. The relative vascularity of the five laminae in a single
region is found to be similar in all the cortical areas examined,
the lamina granularis interna (IV) being the richest in every
case.

3. The supragranular layers show a tendency to be a little
richer than the infragranular ones, the poorest layer being the
lamina multiformis (VI) in every area except the insular, where
the lamina zonalis (I) is very slightly poorer.

4. It seems probable that the granular and supragranular
layers are receptive and associative in function, while the infra-
granular layers give rise to corticifugal fibers. This suggests a
comparison with the lower centers, where the sensory and cor-
relation nuclei were found to be more richly vascular than the
motor nuclei. Moreover, it has been suggested that the lamina
granularis interna is composed of relatively less highly differ-
entiated cells than the other layers, which gives rise to specula-
tion as to why it should be more vascular than the remaining
laminae.

5. The average vascularity of the five layers is the same in
the occipital and temporal areas, and is only slightly less in the
praecentral region. The parietal area is distinctly richer than
the others, while the insular cortex is much the poorest.

6. The vascularity of the five laminae in the insular cortex
covers about the same range as that of the various motor centers
studied in the brain stem and spinal cord (table 3), while the
vascularity of the other areas corresponds approximately to
that of the sensory and correlation nuclei.

7. The vascularity of the cerebellar cortex is of about the
same order of magnitude as that of the cerebral cortex taken
as a whole.

8. Sexual and racial differences appear to be more marked in
the cortex cerebri than in the parts of the central nervous system
previously studied, suggesting that the vascularization of the
more recently evolved centers is more susceptible than that of
more ancient regions to sexual, hereditary, or environmental
influences.

212 EDWARD HORNE CRAIGIE

9. The fact that the vascularity of the regio insularis not
only is much poorer than that of the other four areas, but also
differs from them in its sexual and racial characteristics, seems
to indicate that this area differs from the rest more than they
do from each other.

LITERATURE CITED -

Asterisk (*) indicates papers inaccessible to the present writer

Bouton, J. S. 1900 The exact histological localisation of the visual area of the
human cerebral cortex. Phil. Trans. Roy. Soc. London, Ser. B, vol.
193, pp. 165-222.

BropMaNNn, K. 1909 Vergleichende Localisationslehre der Grosshirnrinde, 8.
324. Leipzig.

Craicin, E. Horne 1920 On the relative vascularity of various parts of the
central nervous system of the albino rat. Jour. Comp. Neur., vol.
31, pp. 429-464.

De Castro, Fernanpo 1920 Estudios sobre la neurolgia de corteza cerebral
del hombre y de los animales. I. La arquitectonia neuroglica y vas-
cular del bulbo olfativo. Trab. del Lab. de Inv. biol., T. 18, pp. 1-35.

Donautpson, H. H. 1915 The rat. Memoirs of The Wistar Institute of Anat-
omy and Biology, no. 6.

Durer, H. 1874 Recherches anatomiques sur la circulation de l’encéphale.
Arch. de Physiol. Norm. et Path., ser. II, T. 1, pp. 60-91, 316-353,
664-698, 919-957.

Ferrier, Davip 1886 The functions of the brain. 2nd ed London.

*Fortuyn, A. B. Droocurrver 1914 Cortical cell-lamination of the hemi-
spheres of some rodents. Arch. Neurol. and Psych., Path. Lab.
London County Asylums, vol. 6.

Kapprrs, C. U. Artins 1909 The phylogenesis of the palaeocortex and archi-
cortex compared with the evolution of the visual neo-cortex. Arch.
Neur. and Psych., Path. Lab. London County Asylums, vol. 4, pp.
161-173.

Lewis, W. Bevan 1881 On the comparative structure of the brain in rodents.
Phil. Trans. Roy. Soc. London, Ser. B, vol. 173, pp. 699-746.

Sucira, Naoxr 1917 Comparative studies on the growth of the cerebral cor-
tex. II. On the increase in the thickness of the cerebral cortex during
the postnatal growth of the brain. Albino rat. Jour. Comp. Neur.,
vol. 28, pp. 511-591. ;

TaNnpDLER, J., UND Kanror, H. 1907 Die Entwickelungsgeschichte des Gecko-
Gehirns. Anat. Hefte, Bd. 33, S. 555-665.

Van’? Hooe, E. G. 1920 On deep-localization in the cerebral cortex. Jour
Nerv. and Ment. Dis., vol. 51, pp. 313-829.

Van VaLKensurec, C. T. 1913 Experimental and pathologico-anatomical re-
searches on the corpus callosum. Brain, vol. 36, pp. 119-165.

*Van VaLKEenBuRG, C. T. 1913a On vertical localization in the cerebral
cortex. Proc. Netherland Medico-physical Congress, 1913. (Author’s
abstract, Jour. Nerv. and Ment. Dis., vol. 52, p. 459. 1920.)

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Resumen por el autor, C. Judson Herrick.

Las conexiones del nervio vOmero-nasal, bulbo olfatorio accesorio
yamigdala en los anfibios.

En los urodelos toda la pared lateral del hemisferio cerebral
retiene el cardcter del nicleo olfatorio lateral, dentro de cuya
parte ventral existe un primordium estrio-amigdaloide no
diferenciado. En los anuros esta pared se ha diferenciado en:
1) Un nticleo lateral combinado, formado por el nicleo olfatorio y
el l6bulo piriforme; 2) El cuerpo estriado; 3) La amigdala. Los
anuros poseen un 6rgano vOmero-nasal bien definido, un nervio
vomero-nasal y una formacién vomero-nasal en el bulbo olfatorio.
La diferenciacion de este sistema comienza en los urodelos, pero
no ha llegado a consumarse. La amigdala de los anuros se ha
desprendido del complejo estrio-amigdaloide de los urodelos bajo
la influencia de la diferenciacién periférica del é6rgano voémero-
nasal y las vias nerviosas especfficas relacionadas con él. En los
peces no existe una verdadera amigdala, aun cuando algunos de
los elementos representados en este complejo de los cerebros mas
superiores puede estar presente en varias combinaciones. En
los vertebrados mds superiores diferentes sistemas funcionales
se han agregado a la amigdala primitiva, produciendo complejos
amigdaloides de formas diversas, en la mayor parte de los cuales
existe un componente olfatorio, aunque puede haberse supri-
mido como ocurre en el delfin, sin destruir la unidad del complejo.
La relacién del 6rgano vémero-nasal con este complejo en los
amniotos niotos no ha sido investigada.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, JUNE 27

THE CONNECTIONS OF THE VOMERONASAL NERVE,
ACCESSORY OLFACTORY BULB AND AMYGDALA
; IN AMPHIBIA

C. JUDSON HERRICK
Anatomical Laboratory, The University of Chicago

THIRTY-SEVEN FIGURES

CONTENTS
Heep TOUN CHLOE Ake. 5 Serra Lanh AAMT eet OSE Ne oat ad che ae aya Me RTEN tae olet 213
GLUT AS erties cer tal EAC ese peel eine miele crap ers 1g MO TENS Ra ound as ow emg 916% 217
HME: Cl tate A eee er pe ive, Bass arsine A cers eta eyahe moda aheud wi HS sgiehs im Sig Elece 238
IV. Morphogenesis of the urodele strio-amygdaloid complex............... 255
V. The comparative anatomy of the strio-amygdaloid complex.......... 263
VERE Str ATV Se TT PERE. cots nisl. Naa Sahat CRIS Soeid See OR awa s ae Etieoehie 276

I. INTRODUCTION

It has long been known that in various mammals, reptiles,
and amphibians the olfactory bulb is divided into two parts or
lobules, of which the smaller, usually called the accessory bulb,
occupies various positions, generally near the caudal border of
the main bulb. It was early recognized by a few students of
of this region that the accessory bulb is related by a special
strand of nerve fibers with the vomeronasal organ (Jacobson’s
organ) of the nose; and this morphological relationship has more
recently been emphasized by McCotter (12). In 1917 McCotter
extended his previous observations on the mammals to include
the turtle and the frog.

From all of these studies it is clear that the neuro-epithelium
of the vomeronasal organ is similar in character to that of the
remainder of the olfactory mucosa; that from it there arises in
amniote vertebrates a special nerve, the vomeronasal nerve,
which is morphologically and probably physiologically a special-
ized part of the olfactory nerve, and that the vomeronasal nerve
terminates in a differentiated portion of the olfactory bulb, the

213

214 C. JUDSON HERRICK

accessory bulb. The nervus terminalis is distinct from both
the olfactory nerve and the vomeronasal nerve, as has been
clearly shown by McCotter (’13) and others.

The portion of the olfactory bulb which receives the fibers
from the general olfactory epithelium is termed by McCotter
(713) the olfactory formation (regio bulbaris principalis of Gaupp,
’°99, in the frog); the portion which receives the vomeronasal
nerve (i. e., the accessory bulb) is the vomeronasal formation
(Zuckerkandl, 710, p. 11). The latter is usually small, but in
the turtle (Chrysemys) it is nearly as large as the olfactory
formation, occupying the entire dorsal surface and upper half of
the medial surface of the olfactory bulb (McCotter, 717). The
vomeronasal nerve and formation appear to vary in size in
proportion to the extent of the sensory epithelium of the vome-
ronasal organ (Zuckerkandl, 10 a, p. 839).

In a review of the relations of the vomeronasal organ to the
respiratory mechanisms of Amphibia, Bruner (’14) has summa-
rized some of the literature and discussed the functional relations.
He supports the belief of Seydel that in the higher amphibians
the nasal organ consists of two functionally and morphologically
distinct parts; a true olfactory cavity which is used primarily
for testing the inspiratory current, and an organ of Jacobson
which receives its stimuli through the medium of the expiratory
current. His detailed examination of the respiratory mechanisms
and their functional workings in various amphibians shows that
this relationship prevails whether the olfactory medium be air
or water.

Bruner distinguishes two types of respiration in Amphibia:

a) In the first the respiratory medium passes freely inward through
the nasal cavity to the mouth, but its return to the nasal cavity from
the mouth is prevented by a mechanical valve at the choana. b) In
the second type the respiratory mechanism is wholly under muscular
control and the olfactory medium passes freely in and out through the
nasal cavity.

Corresponding to these two types of the respiratory mechanism, we
can distinguish a) Monosmatic forms (single smellers), including
Necturus and the larvae of Amblystoma and Rana, in which the olfactory

organ is used to test only the external medium.’ For this condition I
propose the name monosmesis. b) Diosmatic forms (double smellers),

THE AMYGDALA IN AMPHIBIA 215

including Siren, Cryptobranchus, Amphiuma, larvae of lungless salaman-
ders, and the adult stage of higher amphibians. In these forms,
ingoing and outgoing currents bear odorous matter to the olfactory
organ, which is accordingly used to test both the external medium
and the contents of the oral cavity. This condition, which is common
to all higher vertebrates, may be called diosmesis.

In (adult)! single smellers (Necturus)! Jacobson’s organ is wanting
and the olfactory organ has a very simple condition. In double smellers
the olfactory organ is complex and Jacobson’s organ is present.

For the development of a complex olfactory organ, with an organ
of Jacobson, the nature of the olfactory medium is of less importance
than the question whether the animal is a single or a double smeller.

The more recent observations of Broman (’20) on reptiles and
mammals are commented upon on page 265.

in the frog the vomeronasal formation (bulbulus olfactorius
accessorius of Gaupp, 799, p. 99) forms a distinct accessory
bulb. on the lateral surface of the cerebral hemisphere farther
caudad than the olfactory formation. Its relations to the
vomeronasal nerve and organ have been fully described by
McCotter (17).

In 1910 in connection with a brief review of the forebrain of
the frog I described a tract of unmyelinated fibers, the ventro-
lateral olfactory tract (’10, pp. 422, 444 and figs. 40, 41), which
passes from the accessory olfactory bulb to the so-called corpus
striatum, or lateral basal nucleus of the hemisphere. In view of
the peripheral relationship of the accessory olfactory bulb to the
vomeronasal organ, it is clear that the ventrolateral olfactory
tract can be nothing other than the central-conduction pathway
of the vomeronasal system, and the terminal nucleus of this tract,
a part of the so-called corpus striatum of the frog, is similarly
related to the vomeronasal apparatus.

The fiber connections of the region hitherto called corpus
striatum in the frog show that it must be separated into two
clearly distinct parts: 1) a true corpus striatum (paleostria-
tum) which is very simply developed in the frog, and, 2) farther
caudad a definite nucleus which is comparable with a part of the
mammalian nucleus amygdalae. The latter nucleus receives

1 The words in parenthesis are added with a pen in the copy of the author’s
reprint sent to me,

216 C. JUDSON HERRICK

the ventrolateral olfactory tract from the vomeronasal formation
and will be here termed the amygdala. Our attention in this
paper will be chiefly directed to the structure and connections
of the amphibian amygdala.

The terminology of the amphibian forebrain is in great con-
fusion. The usage here employed conforms in general, except
as noted, with that of my contribution published in 1910.

The cerebral hemisphere of the frog is divisible into five regions,
the olfactory bulb and four quadrants termed dorsomedial, ventro-
medial, ventrolateral, and dorsolateral. The dorsomedial quad-
rant is the primordial hippocampus; the ventromedial quadrant
is the septum, using this term in its broadest sense; the ventro-
lateral quadrant includes the very undifferentiated corpus
striatum and the amygdala, which is a sharply defined nucleus;
the dorsolateral quadrant comprises the lateral olfactory nucleus
and the pyriform lobe, which are relatively undifferentiated and
not clearly separable from each other.

All four quadrants converge into an undifferentiated anterior
olfactory nucleus at the base of the olfactory bulb, and the two
dorsal quadrants converge into an undifferentiated area at the
posterior pole of the hemisphere. Comparison with higher
brains shows that the dorsal quadrants are primordial pallium
(the dorsolateral incompletely so), but the ventral quadrants are
essentially basal nuclei and do not in any vertebrate give rise to
true cerebral cortex. The general relations of the amphibian
cerebral hemispheres to those of other ichthyopsid forms are
reviewed in a recent paper (Herrick, ’21).

This account is based upon a collection of several hundred
brains of urodele and anuran Amphibia prepared by various
methods by Dr. Paul 8. McKibben, upon about 150 brains of
larval Amblystoma prepared by the Golgi method by Dr. Charles
Brookover, and upon about 150 brains of adult and larval
Amblystoma variously prepared in this laboratory by my assist-
ant, Miss Jeannette B. Obenchain. To these careful workers
I acknowledge my deep indebtedness.

THE AMYGDALA IN AMPHIBIA yal 7

Il. ANURA
The olfactory bulb

In the frog the internal structure of the vomeronasal formation
is, so far as observed, essentially similar to that of the remainder
of the olfactory bulb. The fibers of the vomeronasal nerve
enter glomeruli, where they engage dendrites of the mitral cells
in the usual way. Beneath the mitral cells is a layer of ies
cells bordering the ventricle (figs. 1, 19).

The accessory bulb not only forms ie familiar eminence on the
lateral surface of the cerebral hemisphere, but also projects into
the lateral ventricle as a well-defined ventricular swelling (figs.
1, 35). This is shown, somewhat imperfectly, in horizontal
section in figures by P. Ramén y Cajal (’05, pl. 15, fig. 3, and
pl. 17, fig. 5). Through the deeper part of this thickening there
pass fibers of the dorsolateral olfactory tract (a few of which are
myelinated) on their way from the ventral part of the olfactory
formation to their definitive position in thedorsolateral quadrant
of the hemisphere. Toward the caudal end of the accessory
bulb these fibers form a distinct daseicke at its dorsomedial angle
(fig. 1).

In the olfactory formation of the bulb the layer of granule cells
is separated from the layer of mitral cells by a distinct molecular
layer; this latter layer, however, is absent in the vomeronasal
formation (fig. 19), and here there is no clear boundary between
mitral cells and granule cells.

The ventrolateral olfactory tract

The course of the ventrolateral olfactory tract, its termination
in the amygdala, and the course of the dorsal olfactory projection
tract from the latter organ to its nucleus in the chiasma ridge
as seen in cross-sections of the brain of the frog, Rana pipiens,
are illustrated in figures 1 to 12. These figures are drawn from
a series of Weigert sections and for purposes of orientation all
myelinated fiber tracts are entered on the right side of the draw-
ings. The greater part of the courses of the ventrolateral olfac-
tory tract and the olfactory projection tract can be followed in

ABBREVIATIONS FOR ALL FIGURES

amg., nucleus amygdalae

b.ol.ac., bulbus olfactorius accessorius

c.f., columna fornicis

c.gen.lat., corpus geniculatum laterale

ch., chiasma opticum

ch.r., chiasma ridge

com.amg., commissura amygdalarum

com.ant., commissura anterior

com.hab., commissura habenularum

com.hip., commissura hippocampi

com.po., commissura postoptica

c.8., corpus striatum

d.amg.hab., decussation
amy gdalo-habenularis

d.B., diagonal band of Broca

d.f.lat.t., decussation of lateral fore-
brain bundles

d.f.med.t., decussation of medial fore-
brain bundles

d.ol.p.tr., dorsal olfactory projection
tract

d.tr.th.h.c., decussation of tractus tha-
lamo-hypothalamicus cruciatus

em.th., eminentia thalami

ép., epiphysis

epen., ependyma

F., foramen interventriculare

jim., fimbria

f.lat.t., fasciculus lateralis telencephali
(lateral forebrain bundle)

f.med.t., fasciculus medialis telen-
cephali (medial forebrain bundle)

f.ol., formatio olfactoria

f.retr., fasciculus retroflexus Meynerti

f.vn., formatio vomeronasalis

g.c., granule cells

gl.ol., glomeruli olfactorii

hab., habenula

hyp.g., hypophysis, pars glandularis

hyp.n., hypophysis, pars nervosa

inf., infundibulum

lob.p., lobus piriformis

l.t., lamina terminalis

m.b., midbrain

m.c., mitral cells

mol., stratum moleculare

..ol., nervus olfactorius

..ol.lat., lateral division of olfactory

nerve

.op., nervus opticus

.term., nervus terminalis

of tractus

> 2

= 2

nuc.ac., nucleus accumbens septi

nuc.ol.ant., nucleus olfactorius anterior

nuc.o.p.tr., nucleus of dorsal olfactory
projection tract

nuc.po., nucleus preopticus

nuc.v.l., nucleus ventrolateralis hemi-
sphaerii

p.d.hyth., pars dorsalis hypothalami

p.f., prominentia fascicularis

p.hip., primordium hippocampi

pol.post., posterior pole of cerebral
hemisphere

p.v.hyth., pars ventralis hypothalami

p.v.l.h., pars ventrolateralis hemi-
sphaeri

p.v.th., pars ventralis thalami

r.po., recessus preopticus

s.a., stratum album

sac.v., saccus vasculosus

sep.ep., septum ependymale

s.g., stratum griseum

str.med., stria medullaris

str.t., stria terminalis

tect., tectum mesencephali

thal., thalamus

tr.amg.hab., tractus amygdalo-habenu-
laris

tr.amg.p., tractus amygdalo-piriformis

tr.c.hab.l., tractus cortico-habenularis
lateralis

tr.ol.c.s., tractus olfacto-corticalis septi

tr.ol.d.l., tractus olfactorius dorsolat-
eralis

tr.ol.hab.ant., tractus olfacto-habenu-
laris anterior

tr.ol.v.l., tractus olfactorius ventro-
lateralis

tr.op., tractus opticus

tr.s.hab., tractus septo-habenularis

tr.th.f., tractus thalamo-frontalis

tr.th.h.c., tractus thalamo-hypothal-
amicus cruciatus

ir.th.p.1., tractus
laris intermedius

v.l., ventriculus lateralis

v.ol.p.tr., ventral olfactory projection
tract

v. 3, third ventricle

z.lim.lat., zona limitans lateralis

z.lim.med., zona limitans medialis

thalamo-peduncu-

THE AMYGDALA IN AMPHIBIA 219

these sections, though their fibers are unmyelinated. These
are sketched in on the left side of the drawings, their terminal
relations (which are not demonstrated by the Weigert sections)
being added from data derived from, Golgi sections in various
planes.

These connections are also shown as seen in horizontal sections
by the Golgi method of the cricket frog, Acris gryllus, in figures

p. hip.
lob. p. LE Ol. G45
z. lim. med. 7 ey vl.
tr ol.vl. i! a a2: Sptr od f

m.C.

g.c.
septum

xxxi- 197

Figs. 1 to12 Aseries of transverse sections through the brain of Rana
pipiens. X12. The outlines and arrangement of cells and myelinated fibers are
drawn to scale from a series of Weigert sections. The sections are somewhat
compressed dorsoventrally and are slightly oblique, the right side being farther
rostral than the left. The arrangement of cells is necessarily somewhat conven-
tionalized on:account of the small scale of the drawings.

These sections were prepared by the Weigert method, fixing in formalin and
potassium bichromate, mordanting the sections in copper acetate, and destaining
in potassium permanganate and the oxalic potassium sulphite mixture. The
destaining was arrested at a point which differentiated the myelinated fibers,
but left considerable brownish-yellow color in the background. All cell bodies
are clearly visible and their arrangement was further controlled by comparison
with sections stained with toluidin blue. Some of the unmyelinated tracts are
also differentiated by the ‘brown reaction.’

The myelinated fibers are entered only on the right side of the drawings.
On the left side are entered the ventrolateral olfactory tract and the dorsal
olfactory projection tract, both of which are unmyelinated. The courses of
these tracts can be followed in the Weigert sections throughout their length
except near their ends, but the details of these unmyelinated fibers are drawn
in by comparison with various other series of transverse and longitudinal sections
prepared by the methods of Golgi and Cajal.

Fig. 1 On the left the section passes through the extreme caudal end of the
vomeronasal formation, showing its layers of granule cells and mitral cells.
There are no glomeruli at this level; on the right these are present. The un-
myelinated fibers of the ventrolateral olfactory tract have assembled from the
vomeronasal formation to form a compact bundle in the deeper part of the zona
limitans lateralis.

220 Cc. JUDSON HERRICK

19 to 21. We have sections of the brains of several other speci-
mens of different species of frogs prepared by the methods of
Golgi and Cajal and cut in various oblique longitudinal planes
in which each of these tracts is clearly demonstrated for practi-
cally its entire length in a single section.

Since the cell bodies from which these fibers arise are not
impregnated in these preparations, the direction of conduction
is not demonstrated. The indications are, however, clear that
the ventrolateral olfactory tract is mainly (probably exclusively)
a descending system from the vomeronasal formation to the
amygdala and free terminal arborizations of these fibers are seen

lob. p. trol. dil.
p. hip. z. lim. med
troles.t+ce.f
trol. v1. Zin ae
septum fateh
Cus: f.medt

nuc. ac, :
XXX1- 169

lob.p.
p. hip.

trol. v.I.

ety ag tela * AXKII9Y

Fig. 2. Through the cerebral hemisphere between the vomeronasal formation
and the lamina terminalis. The ventrolateral olfactory tract lies close to the
ventricle in the zona limitans lateralis.

Fig. 3. Through the rostral border of the lamina terminalis. On the left the
ventrolateral olfactory tract is seen entering the rostral end of the amygdala.

THE AMYGDALA IN AMPHIBIA Zak

in the amygdala. There is no evidence of physiological connec-
tion by collateral branches or terminals with any other neurons
between its origin in the vomeronasal formation and its termina-
tion in the amygdala. That is, the true corpus striatum of the
Anura does not receive any appreciable number of olfactory
fibers, a relation in marked contrast with that of Urodela (p. 242).

The ventrolateral olfactory tract rises up slightly dorsalward
from its origin and then passes directly backward in the deeper
layers of the zona limitans lateralis between the dorsal and the

XXX/- 20!

lob. p.
p. hip.

amg.
C. S.
Nuc. po.

XxXX1- 209

Fig. 4 Through the lamina terminalis immediately rostral to the interven-
tricular foramen, showing terminals of the ventrolateral olfactory tract in the
dorsorostral part of the amygdala.

Fig. 5 Through the interventricular foramen on the right side and immedi-
ately behind it on the left, the section being slightly oblique. At this level the
amygdala attains its maximum size and its neurons are very-numerous; those
of the corpus striatum are few in number.

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 3

222 Cc. JUDSON HERRICK

ventral lateral quadrants of the cerebral hemisphere. Upon
reaching the amygdala its fibers turn ventralward to arborize
freely among the neurons of the dorsolateral part of this nucleus.

1b. p. . ate SFE. _\-Z. Im. med.
ray We ht com. hip.
p. hip. tr. amg. hab.
d. amg. hab.
ASUS d. f lat. t.
dol. p.tr. frtait
Gas d. f. medt.
nuc. po.

Fig. 6 Through the middle of the anterior commissure complex. At this
level there are represented in this commissural complex the decussation of the
medial forebrain bundles, the decussation of the lateral forebrain bundles, the
commissura hippocampi, and between the last two a mixed system of unmyeli-
nated fibers containing the commissure of the two amygdalae and habenular
fibers (cf. fig. 17 and p. 227). Unmyelinated fibers of the dorsal olfactory projec-
tion tract are present in the lateral part of the amygdala. A few myelinated
fibers of the habenular tract of the amygdala (tractus amygdalo-habenularis) are
seen on the right.

Fig. 7 Through the caudal end of the anterior commissure ridge, containing
unmyelinated fibers of the hippocampal commissure. The fibers of the dorsal
olfactory projection tract are leaving the ventral border of the amygdala. The
amygdalo-habenular tract is directed upward toward the stria medullaris. The
section passes through the extreme caudal end of the corpus striatum, repre-
sented by a few neurons embedded in the lateral forebrain bundle.

THE AMYGDALA)‘ IN! '‘AMPHIBIA 223

The structure and connections of the amygdala

The amygdala is anatomically one of the most clearly defined
‘regions of the anuran brain. Externally it participates some-
what in the formation of the ventrolateral Tidge termed by
Gaupp prominentia fascicularis, though this eminence is formed
chiefly by the lateral forebrain bundle and the associated corpus
striatum. The amygdala is somewhat pear-shaped, the stem
of the pear (which is directed forward). receiving the ventro-
lateral olfactory tract. Internally it is ovoid in cross-section
and projects into the lateral ventricle at the level of the interv ven-
tricular foramen (figs. 5 and ali

INOS (SOOM: asl } DJONITSS

d.ol.p. tr
f. lat.t.
nuc. po.

Fig. 8 Through the extreme caudal end of ‘the amygdala.
Fig. 9 Through the rostral end ae the Gulch re eke sues ee
amygdala. BIOS

224 Cc. JUDSON HERRICK

This part of the frog brain has been called corpus striatum by
some previous writers (including myself, ’10). But not by
Gaupp (99, p. 107), who with his usual acumen recognizes
that only the undifferentiated middle part of the ventrolateral
area which is closely associated with the lateral forebrain bundle
is with certainity to be compared with the ‘basal ganglia’ of
higher vertebrates. Further than this he was not able to carry
his analysis of this region.

The true corpus striatum of the frog, as already indicated,
occupies the ventrolateral quadrant of the cerebral hemisphere,
chiefly rostrally of the interventricular foramen, and is charac-
terized by its functional connection with the lateral forebrain
bundle (figs. 2 to 6, 14, 15). Its more rostral portion is one of
the least differentiated regions of the hemisphere. Its caudal
end at the level of the foramen is expanded to form a highly
differentiated nucleus containing but few neurons and a very
dense neuropil (figs. 5, 6, 17, 20, 23) derived chiefly from the
lateral forebrain bundle. From this neuropil arises the decussa-
tion of the lateral forebrain bundle (figs. 6, 17), which is com-
posed largely of collaterals of the fibers of this bundle.

The cell bodies of the neurons of the amygdala tend to be
accumulated on the medial border, though some are distributed
throughout. They are sparse or absent on the lateral side.
This cellular area is confluent dorsally with that of the pyriform
lobe, ventrolaterally with that of the corpus striatum, and ventro- _
medially with that of the lamina terminalis and preoptic nucleus
(figs. 4, 5), though the limits between these regions are rather
sharply defined.

As seen in Golgi preparations (fig. 13), some neurons of the
amygdala have large rounded cell bodies and some are very
small. Their dendrites spread widely and irregularly throughout
the nucleus. The neurons of the adjacent corpus striatum tend
to be larger and more irregular in shape. In most of our prepara-
tions where the cell bodies and dendrites are impregnated their
axons are not, and conversely, so that it is difficult to be sure
of the connections of individual cells. - The impression gained
from the preparations is that the larger elements send their

THE AMYGDALA IN AMPHIBIA 225

axons downward into the olfactory projection tracts, while
the axons of the smaller ones are directed laterally and dorsally
into the pyriform lobe (tractus amygdalo-piriformis).

In Golgi preparations in which the cell bodies and dendrites of
none of the intrinsic neurons of the amygdala are impregnated
the entire structure is filled with a very dense neuropil of fine
fibers. These are axons, apparently chiefly of extrinsic origin,
_and this neuropil may be divided into two incompletely separable
parts, one at the rostral end related chiefly with the ventro-
lateral olfactory tract, and below this a larger part related with
the hypothalamic, habenular, and commissural connections.

com. hab.
d. ol. p.tr.

f lat.t
Ch.

Fig. 10 Through the habenular commissure and the rostral border of the
optic chiasma. The olfactory projection tract is passing ventralward, medial-
ward, and spinalward along the medial border of the lateral forebrain bundle.

Fig. 11 Through the caudal part of the chiasma ridge and postoptic com-
missure complex and the posterior poles of the cerebral hemispheres.

226 ©. JUDSON HERRICK

‘The amygdala. of the frog, as. we have seen, receives a direct
olfactory tract of the second order from the vomeronasal forma-
tion, the tractus olfactorius. ventrolateralis, and is therefore
to be regarded asa derivative-of the primitive lateral olfactory
nucleus. It has in addition the following fiber connections, all
of which are characteristic of regions of both lower and higher
ire related with the olfactory complex:. Bek bea pi

. A-commissural strand in the anterior commissure connected

ae the amygdala of the opposite side. r 5 fe gt et

dol. p. tr.

nuc. o. p. tr.
chr

XXXI-2ES

=- ae

Fig. 12. Through the caudal edge of the chiasma mUee including the nucleus
of the olfactory projection tract.

Fig. 13. Neurons of the amygdala of Rana catesbiana. Golgi method.
x 70. The section is from a horizontal series, left side, at the level of the inter-
ventricular foramen; cf. figure 19. The broken line marks the outline of the
amygdala, the arrow points rostrad, and the medial'surface is at the right.

THE AMYDGALA IN AMPHIBIA 227

2. An amygdalo-habenular connection by way of the stria
medullaris, some of these fibers decussating in the anterior
commissure.

3. An amygdalo-pyriform connection with the overlying
pyriform lobe.

4. The diagonal band of Broca, directed forward and medial-
ward under the lateral forebrain bundle to connect with the
septum.

5. The stria terminalis, directed forward and medialward above
the lateral forebrain bundle, to connect with the septum and
preoptic nucleus, some of its fibers probably decussating in the
anterior commissure.

6. The ventral olfactory projection tract, directed backward
and medialward under the lateral forebrain bundle.

7. The dorsal olfactory projection tract, directed backward
and medialward above the lateral forebrain bundle, to connect
with the hypothalamus.

These seven connections will next be described. They are
schematically indicated in figures 35 and 36.

The commissura amygdalarum. This is a very small compact
strand of unmyelinated fibers passing between the two amygdalae
in the anterior commissure (figs. 17, 20). It lies ventrally of the
decussations of the amygdalo-habenular tracts and lateral fore-
brain bundles and dorsally of that of the medial forebrain bundle.
In Golgi preparations its fibers are lost in the dense neuropil of
the amygdala and their cells of origin are not demonstrated.

Tractus amygdalo-habenularis. This is a system of fibers, partly
myelinated (figs. 6, 7) and partly unmyelinated (figs. 17, 18),
between the amygdala and the habenula. They accumulate
on the dorsomedial border of the amygdala and pass directly
dorsalward to enter the stria medullaris. Some of them appear to
decussate in the anterior commissure dorsally of the commis-
sure of the amygdalae and decussation of the lateral forebrain
bundles and ventrally of the hippocampal commissure (fig. 17,
d. amg. hab.). This component of the anterior commissure is
of open texture and its fibers are diffusely arranged, in marked
contrast with the other three components of this commissure
complex last mentioned.

228 Cc. JUDSON HERRICK

p. hip.
fim lob. p.
p. hip. lob. p. trol. d.}
com. hip. trol.d.! troles
Gar A trol.v I. corm. hip
z.lim. med. ‘Uz. lim lat. trolvl
se yy Gao:
C. S.
ae n. term. {lab
n. term. flat. £. ‘ : a
tr ol. hab. ant. f med.t ro aie
d. B. iB po.
ii 193
14 15
poe lob. p.
Ave trol.dl.
cor. hip.
amg:
d. f med.t. ste
n. term. flat.t
f.med.t ; d. B.
aa tr ol. hab, ant.
po. L ©
\ 111-209

16

Figs. 14 to 18 Five transverse sections through the brain of Rana pipiens
prepared by the silver method of Cajal. X 20. The specimens were much
distorted by shrinkage during preparation, as will be seen by comparison with
figures 1 to 12.

Fig. 14 Section taken a short distance rostral to the lamina terminalis be-
tween the levels of figures 2 and 3.

Fig. 15 Section through the rostral end of the lamina terminalis between the
levels of figures 3 and 4.

Fig. 16 Section immediately rostral to the interventricular foramen at about
the level of figure 4. The amygdala forms an eminence projecting into the lat-
eral ventricle. It is connected with the ventromedial olfactory centers by the
stria terminalis above the lateral forebrain bundle and by the diagonal band of
Broca below this bundle.

THE AMYGDALA IN AMPHIBIA 229

Mingled with the fibers of the amygdalo-habenular tract are
others passing between the septum and the habenula (fig. 18,
tr. s. hab.), the tractus septo-habenularis of my earlier account
(10, pp. 426, 428). Some of these fibers apparently decussate
in the anterior commissure in company with those of the tractus
amygdalo-habenularis. °

The tractus amygdalo-habenularis of this description was
designated in the former paper (710, pp. 429, 444) tractus habenulo-
striaticus. Our preparations do not indicate the direction of
conduction of its fibers. This tract contains the only myelinated
fibers related with the amygdala of the frog. The tractus strio-
thalamicus which I mentioned (’10, p. 444) as related with this
region is connected with the true corpus striatum (whose limits
at that time were not clearly defined) and has nothing to do with
the amygdala.

Note ON THE TRACTUS OLFACTO-HABENULARIS ANTERIOR. In all
species of Amphibia which I have studied there is a connection (which
was overlooked in my paper published in 1910) between the ventral part
of the septal region near its rostral end and the habenula, which I shall
term the tractus olfacto-habenularis anterior. This clearly defined
tract of unmyelinated fibers passes caudad along the extreme ventral
surface of the cerebral hemisphere, and at the rostral end of the lamina
terminalis occupies the angle between this structure and the ventral
border of the prominentia fascicularis (figs. 14 to 18). It then turns
abruptly dorsad and caudad to join the tractus cortico-habenularis
lateralis in the angle between the dorsal border of the prominentia
fascicularis and the posterior pole of the hemisphere. In the latter
part of its course it is a thin sheet of fibers forming the most super-
ficial layer of the prominentia fascicularis. The less densely stained
fibers of the diagonal band form an intermediate layer between these
and the longitudinally directed fibers of the lateral forebrain bundle.

Tractus amygdalo-piriformis. There is a broad connection
between the amygdala and the overlying pyriform lobe, probably
of mixed character. For the observed relations see the account
of the dorsal olfactory projection tract below.

The diagonal band of Broca. From the entire lateral surface
of the amygdala very fine unmyelinated fibers assemble in diffuse
formation and turn ventralward in a narrow zone external to
the lateral forebrain bundle and, farther forward, ventrally of it.

firm.
corn. hip,
“\tramg. hab,
“ “gtr amg. p.
ama,

z ant.

yell
ki y) Mii; -
d. f. lat.t. ue yy a) f. med.t.

d. f. med. t: S nuc, po.
com. amg: —r po.
n. Op. =
17
p. hip.
lob. p.
tr. ol. d.l.
fh amg.
} } tramq. p.
So tr amg, hab.
WARS a hab ant.
Biss anf / (1 "pi Esa ~ d. ol. p. tr
od ROASTS AN v. ol. p. th
com. hip. Ages, 1 ey pie
tr s,hab/ Wey : \f med. t.
r. po. 4

ch.

18

Fig. 17 Section through the interventricular foramen and anterior com-
missure; cf. figures 5 and 6. The amygdala is seen to be connected with the
pyriform lobe by amygdalo-pyriform fibers and with the opposite side by two
systems of fibers, one above and one below the decussation of the lateral fore-
brain bundles. The ventral system, commissura amygdalarum (com. amg.), is &
small compact fascicle of unmyelinated fibers which apparently form a true com-
missure. The dorsal system is a more diffuse collection of fibers of various
sorts, containing decussating fibers of the tractus amygdalo-habenularis (d. amg.
hab.), mingled with those of the tractus septo-habenularis and perhaps other
kinds of fibers. Amygdalo-habenular fibers are accumulating on the dorso-
medial border of the amygdala; these are both myelinated and unmyelinated.

Fig. 18 Section immediately caudad of the interventricular foramen at
about the level of figure 7. Fibers of the olfactory projection system are passing
ventralward, medialward, and spinalward from the amygdala, some dorsally of
the lateral forebrain bundle (d.ol.p.tr.) and some ventrally of this bundle
(v.ol.p.tr.).

230

THE AMYGDALA IN AMPHIBIA 231

Here they form a compact sheet of fibers directed ventralward,
medialward, and forward. Some of them apparently connect
with the preoptic nucleus; others extend farther forward to
connect with the septal nuclei (figs. 14 to 17).

Stria tetminalis. There is another connection of unmyelinated
fibers between the amygdala and the medial olfactory areas of
the septum and preoptic nucleus. These fibers leave the medial
surface of the amygdala near its rostral end and pass medialward
and forward dorsally and medially of the lateral forebrain bundle,
forming the stria terminalis (fig. 16). The diagonal band and the
stria terminalis are best seen in our Cajal sections. The direction
of conduction is not revealed in these preparations. The fibers
of both tracts probably run in both directions between the amyg-
dala and the medial olfactory areas. Some fibers of the stria
terminalis system probably decussate in the anterior commissure,
though our preparations do not demonstrate this.

The ventral olfactory projection tract. These fibers (fig. 18)
occupy the same position as the diagonal band of Broca farther
rostrad, these two systems-forming a single sheet of fibers running
superficially across the prominentia fascicularis and distinguished
‘only by the fact that the fibers of the diagonal band are directed
forward, while those of the projection tract are directed ventral-
ward and probably spinalward. The latter join the medial
forebrain bundle, and their destination is unknown. They
probably connect with the adjacent preoptic nucleus and perhaps
with the hypothalamus. The direction of conduction has not
been determined.

The dorsal olfactory projection tract. This small but very well-
defined tract takes a course similar to the last, except that it
runs dorsally and medially of the lateral forebrain bundle and
can definitely be related to a specific nucleus of the hypothalamus.

Almost the entire course of this tract can be seen in a single
section in some of our Golgi preparations, as illustrated in hori-
zontal section of Acris gryllus in figure 20 and in an oblique
plane in Rana pipiens in figures 22 and 23. For the relations
in transverse sections see figures 6 to 12. Its fibers take a very
direct course between the amygdala and the gray matter near

232 C. JUDSON HERRICK

ol.
df. lat.t.

ud &!

Ry

* a
NEE is! Ba Xt
a gh

Figs. 19 to 21. Three horizontal sections through the brain of Acris gryllus.
Golgi method. X 35. Figure 19 is from a specimen in which the ventrolateral
olfactory tract is impregnated for its entire length, and figures 20 and 21 are
from another specimen cut in the same plane, in which the olfactory projection
tract is impregnated, in both cases on the right side only. The cell bodies of
but few neurons are impregnated in these preparations. Those which are
unimpregnated, however, are clearly visible and their arrangement is indicated
somewhat conventionally by stipple.

THE AMYGDALA IN AMPHIBIA » 233

the dorsal border of the chiasma ridge, which is termed the nucleus
of this tract. Our preparations indicate that this tract includes
both ascending and descending fibers in both Anuraand Urodela.
Free terminal arborizations are abundant at both ends of the
tract and in urodeles axons of neurons of its hypothalamic nucleus
are directed dorsally into it.

The nucleus of the olfactory projection tract is not clearly
defined in cell preparations (figs. 12, 20, 21), but in Golgi prepara-
tions of Rana it is seen as a horseshoe-shaped mass of dense
neuropil occupying the most dorsal part of the chiasma ridge
immediately caudad of the decussating chiasma fibers and
extending across the median plane (figs. 22, 23). In Anura it
appears to occupy the extreme dorsal border of the chiasma
ridge, but in urodeles it lies considerably farther caudad and
ventrad in the hypothalamus. We have no satisfactory impregna-
tion of its neurons in the Anura.

Following the dorsal olfactory projection tract spinalward from
the amygdala (figs. 7 to 12), its fibers are seen to assemble from
the entire caudal part of this structure on the medial border
of the lateral forebrain bundle, to accompany this bundle for

Fig. 19 Section through the olfactory and vomeronasal formations of the
olfactory bulb and the amygdala. The ventrolateral olfactory tract, which
curves slightly dorsally in its course from the vomeronasal formation to the
amygdala, does not lie in the plane of this section. On the right side it is
sketched in by projection from the three adjacent sections farther dorsal. In
the region of the vomeronasal formation the stratum moleculare is absent and
the mitral cells are more or less mingled with the granule cells.

Fig. 20 The section passes considerably farther ventral than that shown in
figure 19, below the level of the interventricular foramen. The amygdala lies
for the most part dorsally of this level. From its ventral border the olfactory
projection tract passes backward and medialward to its nucleus at the level of
the chiasmaridge. The brain being small and the sections thick, almost the whole
length of this tract lies in the plane of a single section. At its caudal end it dips
ventralward for a short distance. The commissure between the amygdalae
and the decussation of the lateral forebrain bundles are sketched in from the
adjacent section ventrally.

Fig. 21 Three sections ventrally of the plane of figure 20, passing through
the dorsal border of the chiasma ridge. The nucleus of the olfactory projection
tract lies mostly dorsally of this plane, probably including some neurons in the
position indicated (nwuc.o.p.tr.) at the level of this section. Its boundaries are
nowhere clearly defined in this series,

BoA: Cc. JUDSON HERRICK

a short distance and then diverge from it medialward and ventral-
ward to connect with its nucleus in the chiasma ridge. Its
fibers are probably derived in part from the large neurons of the
amygdala (fig. 13) and in part from the nucleus at its lower end.
Free terminal arborizations are found in both of its terminal
nuclei.

tect.
tr. amg. p.
pe eee en ce gE =e
z. lim. lat. sige eZ ZZ SS th op.
tr. hab. |.

amg.

crabs oi Z LE
SOO aN a S,
nue, po. ~ Hf :
hr po. by
P Y, aS Xxu- 64
ch.

Figs. 22 and 23 Two adjacent longitudinal sections through the brain of
Rana pipiens by the Golgimethod. > 20. The plane of section is inclined about
45° to the horizontal, as indicated by the diagram, figure 23 A, and the caudal
ends of the sections figured are, on the right side, slightly nearer the median
plane than the rostral ends. The sections are taken from the right side, including
a portion of the left side below.

The plane of these sections is so chosen as to show the entire extent of the dor-
sal olfactory projection tract and its nucleus, a horseshoe-shaped neuropil lying
on both sides of the meson in the dorsal part of the chiasma ridge. The arbori-
zations of this tract in the dorsolateral quadrant (pyriform lobe) of the cerebral
hemisphere are shown. There are also included a portion of the lateral fore-
brain bundle, the neuropil of the corpus striatum from which its decussating
fibers arise, the forward extension of this bundle into the ventrolateral quadrant
(corpus striatum), and a portion of the medial forebrain bundle extending for-
ward into the ventromedial quadrant (septum).

THE AMYGDALA IN AMPHIBIA 235

Some of the ascending fibers of this tract end by free arboriza-
tion in the amygdala, but many of them pass along the caudal
border of this nucleus, giving off collaterals to its neuropil, to
terminate by widely expanded free arborizations in the adjacent
parts of the pyriform lobe (figs. 22, 23). This connection between
the amygdala and the pyriform lobe I call the tractus amygdalo-
piriformis, and I believe that it contains, in addition to the

BLL¢y. Op,
Ye ~~ ——~tr.c. hab. |.
ie amg. p.
amg.
d.ol. p. tr.

filet:
d.ol.p.tr

Fig. 23 One section ventrolaterally of figure 22.

fibers just mentioned, axons of the small intrinsic neurons ofthe
amygdala, and fibers passing in the reverse direction from the
pyriform to the amygdala, though the last two types of fibers
are not clearly demonstrated.

The olfactory projection tracts of the Anura, the diagonal band
of Broca, and the stria terminalis may be regarded as parts of a
single complex system of correlation fibers which put the lat-
eral olfactory area into physiological relationship with the ven-

236 C. JUDSON HERRICK

tromedial areas of the septum, preoptic nucleus, and hypothala-
mus. In the frog the lateral connections of this system center
in the amygdala, with more or less extensive connectio~s with the
overlying pyriform lobe. Its fibers pass partly ventrai'v of the
lateral forebrain bundle and partly dorsally of it.

The fibers of the ventral component which can be followed
forward from the amygdala to connect with the nuclei of the
septum and rostral end of the preoptic nucleus form the diagonal
band. The ventral fibers which are followed spimalward form
the ventral projection tract, whose medial connections are
-unknown, probably chiefly with the caudal end of the preoptic
nucleus.

In a similar way the fibers of the dorsal component of this
system form a continuous sheet. Those which are followed for-
ward from the region of the amygdala passing above the lateral
forebrain bundle to connect with septal areas form the stria ter-
minalis; those which are followed spinalward to connect with the
hypothalamus form the dorsal olfactory projection tract. It is
probable that both dorsal and ventral components contain corre-
lation fibers passing in both directions between the lateral and
medial terminal nuclei throughout their entire extent.

The term olfactory projection tract was first used by Cajal as
a synonym for the stria terminalis of lower mammals. The inti-
mate relation of the diagonal band of Broca with this system was
recognized by Johnston in the turtle brain (15, p. 407), and
Crosby (’17) described relations of these tracts to the amygdaloid
complex in the alligator which are very closely similar to those
of the frog. Both of these authors describe in reptiles a com-
ponent of the amygdaloid complex (medial large-celled nucleus
of Johnston, ventromedial nucleus of Crosby) whose topographic
relations and fiber connections resemble those of the amygdala
of the frog as described above (see beyond, p. 269).

Crosby separated the olfactory projection tract of Cajal into
two systems: (1) a system related on the medial side with the pre-
optic nucleus (and septum?), and (2) a system directed farther
caudad to connect with the hypothalamus. To the first she ap-
plied the old term stria terminalis; for the second she adopted

THE AMYGDALA IN AMPHIBIA Qk

Cajal’s name, olfactory projection tracts, and she distinguished
the dorsal and ventral components of the latter. The relations
in the frog are in principle identical with those described in the
alligator, and Miss Crosby’s nomenclature is here adopted.

The dorsal olfactory projection tract is one of the oldest and
most conservative fiber systems in the vertebrate brain, having
long been known and described in various types of fishes under
the name tractus pallii.

Larval Anura

In half-grown tadpoles of the bullfrog, Rana catesbiana, 30 mm.
long, horizontal sections show a well-formed vomeronasal for-
mation of the olfactory bulb. The amygdala is clearly recognized
as an area of neuropil distinct from that of the striatum. In
sections stained with hematoxylin and erythrosin the related cells
cannot be separated from those of the striatum and other adja-
cent parts. The commissure of the amygdalae is large and
readily recognized and is quite distinct from the decussation of
the lateral forebrain bundles. The other fiber tracts are not
revealed by these preparations.

In the brains of much older tadpoles of this species approaching
the metamorphosis (150 mm. long), without forelegs and with
hind legs 65 mm. long, cut in the horizontal plane and stained
with hematoxylin and eosin, the relations of vomeronasal forma-
tion and amygdala are essentially as in the adult. The vomero-
nasal formation is large and very far caudad and is entered by a
large and well-isolated vomeronasal nerve. The amygdala is also
large and its neurons are clearly separated from those of sur-
rounding regions.

Weigert sections of a slightly younger tadpole of this species,
145 mm. long without fore legs and with hind legs 35 mm. long,
show a few myelinated fibers in the tractus amygdalo-habenularis.
The ‘brown reaction’ reveals the course of the ventrolateral
olfactory tract clearly and the dorsal olfactory projection tract
obscurely.

Our collection contains sections through the entire head of old
tadpoles of Pickering’s treetoad, Hyla pickeringii. These are

238 C. JUDSON HERRICK

about 15 mm. long, with well-developed fore legs, and the meta-
morphosis was imminent, Dr. McKibben’s notes stating, “these
toads would have hopped from the water 2 to 4 days after they
were killed.”

In horizontal sections stained with hematoxylin and eosin the
large vomeronasal nerve can readily be followed from the vom-
eronasal organ back to its termination in the vomeronasal forma-
tion, crossing the ventral surface of the main olfactory nerve
close to its junction with the olfactory bulb in the way described
by McCotter (’17, p. 67) for adult Rana catesbiana.

The vomeronasal formation is well differentiated, but is not so
large or so sharply separated from the olfactory formation of the
bulb as in the tadpoles of Rana catesbiana of corresponding age
nor are the neurons of the amygdala so clearly separated from
those of adjacent areas. In similar sections stained in various
ways these relations are confirmed, the ventrolateral olfactory
tract can be followed clearly and the dorsal olfactory projection
tract obscurely. The nucleus of the latter tract is not well dif-
ferentiated from the surrounding gray matter in any, of these sec-
tions of larval forms.

Our material of larval Anura is not extensive; the observations,
though fragmentary, are sufficient to indicate that the vomero-
nasal formation, ventrolateral olfactory tract, and amygdala are
organized essentially as in the adult.

Ill. URODELA
The olfactory nerve of Amblystoma

From our collection of urodeles I shall select for first considera-
tion the brain of adult Amblystoma, of which we have abundant
material variously prepared. Here we find a very different con-
dition from that described in the preceding section and one that
is very instructive from the standpoint of the functional factors
which have operated in the morphogensis of the vertebrate cere-
bral hemisphere.

In the frog the vomeronasal organ as defined by Gaupp is a
small diverticulum from the medial side of the inferior chamber

THE AMYGDALA IN AMPHIBIA 239

of the nasal sac. In urodeles the supposed representative of.
this organ is a lateral pouch from the much simpler nasal sac.
In both cases the recess in question is lined with sensory olfactory
epithelium. For figures of the nasal organ of Amblystoma see
Bawden (94) and Zuckerkand1 (10).

It has long been recognized that in some urodeles there is an
accessory olfactory bulb, less clearly separated from the remainder
of the bulb than in the frog, which receives a separate posterior
and lateral division of the olfactory nerve. In Amblystoma
Coghill (’02, pp. 209 and 253) demonstrated the course of the
posterior division forward, first lying laterally of the anterior
division, then ventrally of it, and finally turning abruptly lateral-
ward to connect with the sensory epithelium of the lateral pouch
termed Jacobson’s organ. He does not exclude the possibility
of a certain: amount of interchange of fibers between the poste-
rior and anterior divisions, and our Golgi preparations in fact
do demonstrate some mingling of these fibers.

I have verified Coghill’s description on old larvae of Ambly-
stoma punctatum taken a few days before the metamorphosis,
save that I have not been able on this material to trace the fibers of
the posterolateral division of the olfactory nerve separately from
the others of the common trunk for a short distance. These
fibers course, as described by Coghill, along the ventral border
of the common trunk. Farther rostrally, immediately caudad
of the level of the choana, this common trunk breaks up into
numerous branches, several of which from the ventral border
of the trunk turn laterally along the ventral surface of the nasal
sac. One of these supplies the olfactory epithelium of the lateral
diverticulum (the supposed vomeronasal organ), others supply
the ventral olfactory epithelium medially and caudally of this
organ. The peripheral twig which supplies the lateral diver-
ticulum is much smaller than the posterolateral division of the
intracranial course of the nerve which terminates in the accessory
bulb, and the indications are that this division contains fibers
from some of the other peripheral lateral branches referred to
above.

240 Cc. JUDSON HERRICK

Coghill is correct, I have no doubt, in tracing the fibers from
the lateral diverticulum of the nasal sac into the accessory bulb,
for in well-preserved material this fascicle can probably be
readily followed separately; but my material suggests that the
incompletely differentiated accessory olfactory bulb receives also
many fibers from other parts of the nasal sac.

From these observations and from the accounts in the litera-
ture it may be concluded that the nasal sac is much more simply
organized in Urodela than in Anura, and that if a*true vomerona-
sal organ is present in the former it is a small lateral diverticulum
of the simple sac. In Amblystoma the olfactory fibers from this
diverticulum terminate centrally in an incompletely differentiated
accessory olfactory bulb which also receives other fibers probably
derived from the ventral wall of the nasal sac.

These relations strongly suggest the partial homology of the
posterolateral division of the olfactory nerve with the anuran
vomeronasal nerve and the accessory bulb with the vomeronasal
formation; but these names will not be applied to the urodeles
in the following description for reasons which will appear in the
discussion on pages 255 and 264.

The olfactory bulb and lateral olfactory tract

In adult Amblystoma the olfactory bulb is strictly lateral in
position (Herrick, ’10, figs. 8 to 11; Bindewald, ’14, figs. C to F)
and occupies the whole thickness of all or part of the lateral wall
of the cerebral hemisphere for about half the distance from the
rostral end to the level of the interventricular foramen. The
caudal end of the bulb (bulbulus accessorius of Bindewald, 714, |
fig. B) is not so clearly separated from the remainder as in the
Anura.

The olfactory bulb as a whole forms a considerable eminence
on the external surface of the brain and also a projection into
the rostral end of the lateral ventricle. The ventricular eminence
is in some preparations separated by a slight internal sulcus into
larger rostral and smaller caudal portions, thus marking on the
ventricular surface the rostral boundary of the accessory. bulb.
This sulcus is visible only in well-preserved material free from

THE AMYGDALA IN AMPHIBIA 241

distortion by shrinkage. The relations of the posterolateral
division of the olfactory nerve to the accessory bulb have been
described above.

The olfactory bulb on the whole is more simply organized than
in the frog. The layer of granule cells is well formed throughout
the bulb and in front it curves around the rostral end of the
hemisphere to the medial surface. In the latter region there
are no mitral cells or glomeruli and the granular layer reaches
the external surface (fig. 24). The mitral cells form a compact
layer in the rostral part of the bulb, but in the accessory bulb
tkey are more scattered, as in the frog. Nowhere is there devel-
oped a definite molecular layer between the granule and the
mitral cells.

Except as just noted, the internal structure of the accessory
bulb seems to be essentially similar to that of the remainder of
the bulb. Mitral cells and subglomerular cells are freely impreg-
nated in our Golgi preparations and the relations of terminals
of the posterolateral division of the nerve to glomeruli and mitral
cells are similar to those of the anterior division throughout. '

From the whole extent of the bulb fibers pass backward into
the lateral wall of the cerebral hemisphere as tractus olfactorius
lateralis, whose dorsal and ventral subdivisions are not so distinct
as in the frog. These are mostly unmyelinated, with a few
myelinated fibers among them which run for a short distance
only (Herrick, ’10, p. 422). Some of the myelinated and unmye-
linated fibers from the ventral part of the bulb pass dorsocaudad
through the deeper layers of the accessory bulb to join the dorso-
’ lateral olfactory tract, as in the frog.

The unmyelinated fibers of the dorsolateral olfactory tract are
very numerous and form a wide compact superficial fibrous layer
over the dorsolateral quadrant of the hemisphere. These second-
ary olfactory fibers come chiefly from the rostral part of the bulb
and are mingled with fibers of the third order (tractus olfacto-
corticalis lateralis). The dorsolateral quadrant also receives
numerous unmyelinated fibers from the accessory bulb which
form a zone of open neuropil between the compact dorsolateral
tract already mentioned and the stratum griseum. These
fibers have been mentioned by Bindewald (14, p. 38).

242 Cc. JUDSON HERRICK

Unmyelinated fibers from all parts of the olfactory bulb also
pass backward in diffuse formation into the ventrolateral part
of the hemisphere. These come from both the anterior and
posterior parts of the olfactory bulb and pass caudad, partly
mingled with but chiefly superficially of the fibers of the lateral
forebrain bundle (fig. 25).

Here I must correct the account of the ventrolateral olfactory
tract given in my previous description of the brain of Ambly-
stoma (10, p. 422), which was based on material inadequate
for accurate study of the unmyelinated fibers. This tract does
not arise exclusively from the accessory bulb and pass “directly
back close to the ventricular ependyma to end in a cellular thick-
ening at the caudal end of the pars ventro-lateralis opposite
the anterior commissure,’’ as in the frog. There is no such
specific relation to the accessory bulb or to the amygdala here.
Bindewald (14, p. 39) very correctly failed to confirm my descrip-
tion. The ventrolateral tract occurs, but not as I formerly
described it.

Clearly the accessory olfactory bulb is by no means so specific,
a structure asin the Anura. It is not so well separated anatomi-
cally from the rostral part of the bulb and. the ventrolateral
olfactory tract arises from both of these parts of the bulb and
not exclusively from the accessory bulb, as in the frog. This
ventrolateral tract, accordingly, is not the exact equivalent of
the tract so named in the frog, though the latter is derived from it
by the suppression of the fibers from the olfactory formation and
further specialization of those from the vomeronasal formation.

The ventrolateral area of the cerebral hemisphere

The forebrain of Amblystoma has been several times described,
the most important papers being those of Stieda (75), Herrick
(10), and Bindewald (14). Figures showing the general struc-
ture will be found in the two papers last cited.

The structure of the urodele cerebral hemisphere is much more
generalized than that of the Anura, though the more important
anuran regions can be recognized. The olfactory bulb makes
up the entire lateral wall in front and part of it for a considerable

THE AMYGDALA IN AMPHIBIA 243

distance farther caudad. The four quadrants into which the
remainder of the hemisphere is divided in Anura can be recognized
in Urodela; the two medial quadrants are structurally very
distinct, but the two lateral are incompletely separated. The
lateral forebrain bundle is the chief distinguishing character of
the ventrolateral area and the dorsolateral olfactory tract of the
‘dorsolateral area. ;

In Amblystoma, as in the frog, the olfactory bulb extends
backward into the ventrolateral area, so that this area does not
extend so far forward as does the dorsolateral (’10, figs. 10, 11).
The arrangement of cells in the lateral wall of the hemisphere
of Amblystoma is indicated schematically in figures 8 to 20 of
my 1910 paper and in figures A to L of Bindewald’s description
(14).

The ventrolateral quadrant is much more simply organized
than in the frog. The anuran corpus striatum and amygdala
are here completely merged, though the fiber connections of this
generalized region show that it contains the primordia of both
of these structures.

A thickening of the ventrolateral wall of the hemisphere at the
level of the interventricular foramen, less sharply defined than
in the frog, contains a considerable collection of nerve cells and
a dense neuropil related chiefly to the lateral forebrain bundle
and its commissure (figs. 24, 37). This clearly is the urodelan
representative of the two separate areas in this region of the frog
which have been already described. The diffuse ventrolateral
olfactory tract cannot be traced definitely to any part of it, but
its fibers are distributed apparently to the entire ventrolateral
area, especially its rostral end. This thickening is the only
well-differentiated structure in the lateral wall of the urodele
cerebral hemisphere behind the olfactory bulb. Not to preju-
dice its morphology at the start, it will here be termed the
ventrolateral nucleus of the hemisphere.

The rostral end of the ventrolateral area of the hemisphere
appears to be physiologically dominated by the secondary olfac-
tory fibers of the ventrolateral olfactory tract, that is, it is a por-
tion of the lateral olfactory nucleus. The caudal end of this

244 Cc. JUDSON HERRICK

area, the ventrolateral nucleus, is clearly dominated by the lateral
forebrain bundle, whose fibers spread freely throughout its
stratum album, as well as dorsally into the the overlying dorso-
lateral area (figs. 24 to 29, 37). Toward the rostral end of the
ventrolateral area these fibers tend to lie deeper than those of
the olfactory tract, though the terminals of the two systems are
freely mingled. .

gl. ol
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Mm. c
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Fig. 24 Horizontal section through the brain of adult Amblystoma tigri-
num. Weigert method. X17. The section is inclined so that the left side is
farther ventral. It passes through the ventral border of the interventricular
foramen and ventrally of the accessory olfactory bulb. On the left side unmye-
linated fibers of the dorsal olfactory projection tract (d.ol.p.tr.) are demon-
strated by the ‘brown reaction,’ embedded within the lateral forebrain bundle.
In neighboring sections these fascicles can be followed forward to their rami-
fication within the ventrolateral nucleus. On the right side the unmyelinated
thalamo-frontal tract (tr.th.f.) is seen immediately dorsally of its entrance into
the lateral forebrain bundle. Its fascicles can be followed forward to ramify
within the ventrolateral nucleus, in general dorsally of those of the olfactory
projection tract.

THE AMYGDALA IN AMPHIBIA 245

The dense neuropil of the ventrolateral nucleus is composed
chiefly of terminals and collaterals of fibers of the lateral fore-
brain bundle (fig. 28). These are both ascending and descending
fibers. Within it are found also the fibers of the dorsal olfactory
projection tract, as described below. Myelinated fibers (crossed
and direct) connect this neuropil with the habenula in the same
way as similar fibers in the frog are related to the amygdala

H. 2t14-3- 1-2

Fig. 25 Horizontal section through the brain of adult Amblystoma tigrinum
at the level of the interventricular foramen. «Golgimethod. X 12. Thesec-
tion illustrates the origin of the ventrolateral olfactory tract from all parts of the
olfactory bulb; also the course of the more dorsal fibers of the lateral forebrain
bundle and the mingling of these fibers with those of the olfactory tract, in
marked contrast with the condition in the Anura, figures 19 and 20.

(tractus amygdalo-habenularis). These are the fibers termed
tractus habenulo-striaticus in my earlier paper (’10, figs. 16, 17,
18, tr. hab. st.). A large fascicle of the anterior commissure
containing both myelinated and unmyelinated fibers connects
the two ventrolateral nuclei (figs. 24, 28, and Herrick, ’10, fig. 16,
c. a. lat.). These include decussating fibers of the lateral fore-
brain bundles and probably also representatives of the several
systems found in the frog connecting the two amygdalae.

246 C. JUDSON HERRICK

The olfactory projection tracts

In urodeles it is probable that there are fiber connections
comparable with the stria terminalis, diagonal band of Broca,
and ventral olfactory projection tract as described above for
Anura; but, if present, these fibers are diffusely mingled with

Figs. 26 to 31 A series of consecutive horizontal sections through the brain
of a half-grown larva of Amblystoma, arranged in order from dorsal to ventral.
Golgi method. X74. In this series the dorsal olfactory projection tract is
impregnated for its entire length and can readily be followed.

Fig. 26 Section through the wide interventricular foramen dorsally of the
anterior commissure ridge. Within the ventrolateral nucleus of the cerebral
hemisphere some of the more dorsal fibers of the lateral forebrain bundle are
impregnated. Among these are slender wisps of the very fine fibers of the dorsal
olfactory projection tract which are directed- medialward from the tract (cf.
fig. 27) to reach the deeper part of the nucleus. Unimpregnated neurons of the
stratum griseum fill the medial and caudal parts of the nucleus back to the poste-
rior pole of the hemisphere. Medially of the latter is the eminentia thalami,
the. axons of whose neurons form a short tract (tr.th.p.i.) ending in the pars
ventralis thalami, as in Necturus (Herrick, 717, p. 291 and fig. 48).

THE AMYGDALA IN. AMPHIBIA 247

others and they have not been separately followed in our prepara-
tions. ‘The dorsal olfactory projection tract, comparable with
the tractus pallii of fishes, is, however, clearly defined. The
observed relations are indicated diagrammatically in figure 37.

The entire course of the dorsal projection tract is visible in the
Golgi preparations from which figures 26 to 31 were drawn.
These are horizontal sections through the brain of larval Ambly-
stoma, and numerous other preparations by various methods
show that the relations of the tract are essentially similar in
the adult. These figures are drawn from consecutive. sections
arranged in order from dorsal to ventral.

Beginning the description at the caudoventral end of the tract
and following it forward, we notice that the hypothalamic nucleus
of the tract lies relatively farther ventral and caudal than in
Anura. It is an ill-defined group of neurons (none of which are
impregnated in this preparation) lying in the central gray of the
hypothalamus at the caudoventral border of the chiasma ridge.
Some of its neurons are seen in figure 32. The nucleus is filled
with a very dense neuropil derived from several sources and
takes the form of a crescent which crosses the midplane in the
chiasma ridge, the horns being directed caudad into the hypo-
thalamus (figs. 31 to 34). Within this neuropil are free end-
ings of the fibers of the projection tract, many of which
decussate just before terminating.

From the nucleus the tract passes dorsally and slightly laterally
across the posterior border of the chiasma ridge, here being
surrounded by fibers of the medial forebrain bundle (fig. 30).
Upon reaching the level of the lateral forebrain bundle the tract
turns abruptly forward embedded within the other fibers of the
ventral border of this bundle (figs. 28 to 30), and at the lateral
border of the ventrolateral nucleus of the hemisphere its fibers
scatter throughout the neuropil of this nucleus (figs. 26, 27). —

In none of the urodeles which I have examined, either larval
or adult, is the olfactory projection tract related to a special part
of the ventrolateral nucleus of the hemisphere, but its fibers are
mingled with those of the lateral forebrain bundle in a common
neuropil. On the other hand, the tract itself is as large and

248 C. JUDSON HERRICK

clearly defined as in any of the Anura and its hypothalamic
nucleus is apparently relatively larger and a more important
component of the hypothalamus.

The neurons of this nucleus (fig. 32) are of the same peculiar
forms as those which border the infundibulum farther ventrally

/ f iy
seplep / We
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Lee haibale
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Fig. 27 This section passes immediately dorsally of the anterior commissure
ridge. The dorsal olfactory projection tract is a compact fascicle of very fine
fibers in the midst of the less densely crowded fibers of the lateral forebrain
bundle. As shown by the preceding figure, these fibers turn medialward to enter
the deeper part of the ventrolateral nucleus, whereas the greater number of fibers
of the lateral forebrain bundle continue forward laterally and dorsally of this -
neuropil to reach the more rostral region of the ventrolateral part of the hemi-
sphere. Fibers are converging medialward from the neuropil of the ventro-
lateral nucleus to enter the decussation of the lateral forebrain bundles, most of
these fibers being apparently collaterals of those of the forebrain bundle.

THE AMYGDALA IN AMPHIBIA 249

and posteriorly.. Each is connected with the ventricular sur-
face by a thick process which may join the cell body or the base
of the principal dendrite. They are of quite different form
from the ependyma cells of this region (fig. 32). These ependyma

Ke

str med.

Fig. 28 Section through the dorsal border of the decussation of the lateral
forebrain bundles. That segment of the dorsal olfactory projection tract which

accompanies the lateral forebrain bundle is visible for its entire length i this
section.

cells, moreover, are slender, much branched and varicose, in

marked contrast with the mossy ependymal elements found
elsewhere in the brains of these larvae. The axons of these
neurons, so far as observed, are directed dorsalward and forward

250 C. JUDSON HERRICK

toward the projection tract. The rough and slightly thorny
dendrites spread widely forward and lateralward among the termi-
nals of the descending fibers of the projection tract, the medial
forebrain bundle, and the most caudoventral component of
the postoptic commissure, the tractus thalamo-hypothalamicus
cruciatus.

Br LIll-] \ 1-2-

Fig. 29 Section through the ventral part of the decussation of the lateral
forebrain bundles. The dorsal olfactory projection tract is turning ventrally
and medially to leave the lateral forebrain bundle. The detached portion of the
projection tract at the lower and is entered from the adjacent section ventrally,
vy hich is otherwise omitted from the series of drawings.

The intimate relationship of the tract last mentioned with
this nucleus in Necturus I have previously commented upon
(17, pp. 260, 269). In adults and old larvae of Amblystoma
the same relations prevail (figs. 32, 33). The decussation of the
thalamo-hypothalamic tract lies immediately rostrally of the
nucleus and is penetrated by dendrites of its neurons. Terminals
and collaterals of the tract fibers also turn caudad into the
hypothalamus and here engage dendrites of other neurons of
the nucleus.

THE AMYGDALA IN AMPHIBIA 251

The tractus preopticus of R6éthig (11) reaches the vicinity of
this nucleus and may effect functional connections with it. In
Amblystoma this is a slender strand of a very few myelinated
fibers which run longitudinally in the midplane ventrally of the
preoptic recess for its entire length. Apparently they are accom-
panied by a larger number of unmyelinated fibers, but these
have not been clearly demonstrated. The myelinated tract

Br, L{tl-2- 1-2

Fig. 30 Section immediately dorsally of the chiasma ridge, showing the
olfactory projection tract turning: ventrally along its caudal border.

at its caudal end rises up along the rostral aspect of the chiasma
ridge and crosses its dorsal surface to disappear at its caudal
border. In Necturus the fibers take the same course and are
clearly traced to a region immediately dorsally of the hypothala-
mic nucleus of the olfactory projection tract.

Immediately ventrally of the nucleus of the olfactory projec-
tion tract in the hypothalamus are neurons of much the same
type, whose dendrites probably make the same functional connec-
tions as do those of this nucleus, but whose axons appear to be

D2 C. JUDSON HERRICK

directed caudad (fig. 33). Many Golgi preparations show that
the region reached by these axons is also reached by fibers passing
between the wall of the infundibulum and the pars nervosa of
the hypophysis, including the saccus vasculosus; but actual
continuity of these neurons with the hypophyseal fibers has not
been observed. This, however, has been demonstrated by

Br. Lit 2- 1-3

Fig. 31 Section through the nucleus of the olfactory projection tract, show-
ing terminals of fibers of the tract. The more caudal of these terminals are
added from the section next ventral to the one drawn.

Bochenek (’02) in Salamandra. A similar tract to the saccus
vasculosus has been described by others (e.g., Johnston in Acipen-
ser, 01, p. 67), and Cajal (711, p. 489) describes in the mouse
what is probably the same connection, the fibers arising from
a cluster of neurons immediately behind the optic chiasma.

THE AMYGDALA IN AMPHIBIA 253

Necturus and Amphiuma

In Necturus maculosus the relations are broadly similar to
those of Amblystoma, though there are important differences
which have not been fully worked out. There is an accessory
olfactory bulb very poorly differentiated from the rostral portion
of the bulb. The fibers of the lateral olfactory tract arising from
it are distributed to the entire lateral wall of the hemisphere
mingled with those from the rostral part of the bulb. There is
no separately differentiated amygdala, but a very clearly defined
dorsal olfactory projection tract passes between the ventro-
lateral nucleus of the hemisphere and the hypothalamus, as in
Amblystoma.

In Amphiuma means the relations are much as in Necturus.
The large olfactory bulb is fairly uniformly developed in the
rostral third of the lateral wall of the cerebral hemisphere. The
caudal part of it receives the more lateral bundles of fibers of the
olfactory nerve, but there is no clearly defined accessory bulb.
Our material does not permit determination of the peripheral
relations of the fibers related with the caudal end of the bulb,
nor can the unmyelinated fibers of the lateral olfactory tract
be analyzed.

The dorsal olfactory projection tract is large and can readily
be followed in horizontal sections for its entire course. Its
hypothalamic nucleus lies very far ventrally at the caudal border
of the chiasma ridge. The fibers of the tract enter the nucleus
in many small fascicles, most of which decussate as the most
caudal component of the postoptic commissure complex and then
spread widely caudolaterad in the deepest part of the stratum
album of the infundibulum.

Following the fibers of the olfactory projection tract forward
from their nucleus, they are seen to converge into one or several
compact fascicles which pass dorsalward along the caudal border
of the chiasma ridge, then turn slightly lateralward and abruptly
forward embedded among the fibers of the lateral forebrain
bundle. Their further course can readily be traced into the
neuropil of the ventrolateral nucleus of the cerebral hemisphere,

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, No. 3

' 254 Cc. JUDSON HERRICK

where they again break up into slender strands which spread
out forward and Jlateralward. No silver preparations of this
brain being available, it is impossible to determine the ultimate
connections of these fibers. They certainly spread throughout
the ventrolateral nucleus and probably reach considerable
distances farther forward in the ventrolateral area of the hemi-
sphere and dorsalward into the dorsolateral area and posterior
pole.

—--—-—-—
= ae

Sg

/
H. 2168- 2-1-4

Fig. 832. Three neurons of the nucleus of the dorsal olfactory projection -
tract of an old larva of Amblystoma tigrinum. Drawn from a horizontal section
by the method of Golgi. The left side is slightly farther dorsal than the right.
x 74. These neurons are in connection with the ependymal surface by thick
smooth processes. This process was probably present in the neuron at the
left, but is not included in the plane of the section. Many neurons similar to the
two at the right are seen in our preparations. Their dendrites are widely
branched and somewhat thorny. The smooth axon arises from the base of the
dendritic arborization and is directed toward the olfactory projection tract,
which it probably enters to ascend to the ventrolateral nucleus of the hemi-
sphere. The dendrites of these neurons ramify in a dense neuropil derived
chiefly from the medial forebrain bundle and the tractus thalamo-hypothalamicus
cruciatus, the latter only being impregnated in this preparation. This neuropil
is greatly simplified in the drawing and is omitted altogether in the regions
surrounding the axons of the neurons. A single ependyma cell is impregnated.

THE AMYGDALA IN AMPHIBIA 255

IV. MORPHOGENESIS OF THE URODELE STRIO-AMYGDALOID
COMPLEX

From the relations .described it is evident that the urodeles
possess no amygdala as a separate nucleus differentiated from
the other elements of the ventrolateral part of the hemisphere.
The fiber connections of the ventrolateral nucleus which forms
a thickening of the wall in the caudal part of this area show,
however, that this nucleus combines the functions of corpus
striatum and amygdala, both in a very unspecialized form,
that it is, in short, a strio-amygdaloid body.

inf.

7
H. 215 9-2- 2-21.

Fig. 33 Horizontal section through the most ventral fibers of the postoptic
commissure of an old larva of Amblystoma tigrinum. Golgi method. X 74.
The single neuron impregnated lies considerably ventrally of those shown in
figure 32 and is of different type. As in the other case, the principal dendrite
extends forward into the decussation of the tractus thalamo-hypothalamicus
cruciatus; but the axon, instead of passing forward into the olfactory projec-
tion tract, breaks up into a loose arborization whose longest branches are directed
caudad. In other preparations of similar larvae fibers connected with the pars
nervosa of the hypophysis extend forward into this position, and this neuron
may be the source of such a fiber. It lies ventrally of the nucleus of the olfac-
tory projection tract, though fibers of that tract may extend into this region.
Its chief source of excitation is clearly the thalamo-hypothalamic tract, free
terminals derived from which reach back almost to the caudal end of the hypo-
thalamus, as shown on the right side of the figure (on the left this neuropil is
present, but is omitted from the drawing). ‘

256 C. JUDSON HERRICK

The corpus striatum of the frog is in an exceedingly rudimen-
tary condition, but it is a true striatum (paleostriatum), that is,
it is a portion of the lateral wall of the cerebral hemisphere
relatively free from olfactory influence. The ventrolateral
olfactory tract passes it by toconnect withits own specific terminal
nucleus farther caudad in the amygdala.

In urodeles, on the other hand, secondary olfactory fibers reach
all parts of the lateral wall of the hemisphere, including the

H. 2114-1-2-5>

Fig. 34. Horizontal section through the hypothalamus of adult Amblystoma
tigrinum, from the same specimen as figure 25. Golgi method. X12. The
section passes through the nucleus of the dorsal olfactory projection tract,
which is filled with a dense neuropil of fine fibers, which extend caudad through
almost the entire length of the pars ventralis hypothalami. The nerve fibers
of the saccus vasculosus in the thin roof of the infundibulum and of the pars
nervosa of the hypophysis are richly impregnated. Other preparations: show
that these nerve fibers connect with the hypothalamus, some through the mem-
branous roof of the infundibulum and some through the floor and side walls, and
that they extend forward into the region here filled by neuropil from the nucleus
of the olfactory projection tract, but mostly farther lateral and caudal.

distribution area of the lateral forebrain bundle in the ventro-
lateral quadrant. There is, therefore, neither a true corpus
striatum nor a true amygdala in the urodele, but a relatively
undifferentiated area in the ventrolateral quadrant which retains
some of the characteristics of the lateral olfactory nucleus which
primitively occupied this region, and at the same time has
acquired, under the influence of its ascending non-olfactory
connections, the character of a common primordium of both the
corpus striatum and amygdala.

THE AMYGDALA IN AMPHIBIA 250

The dorsolateral quadrant of the amphibian hemisphere is
also a derivative of the primitive lateral olfactory nucleus. In
urodeles it is scarcely more than this; in anurans it has assumed
more of the characteristics of the mammalian pyriform lobe, but
even in mammals this area has deviated far less from the most
primitive condition (viz., lateral olfactory nucleus) than has the
corpus striatum.

The distinctive characteristics of the differentiated corpus
striatum of higher forms include neither olfactory nor hypothala-
mic connections. Its predominant functional relations are
rather with diencephalic sensorimotor systems termed by many
recent writers somatic, viz., the somesthetic complex and perhaps
the optic and auditory reflex systems. Its chief fiber connec-
tions are with the thalamus proper (pars dorsalis thalami, as I
defined that term in 1910) on the afferent side and with the
motor tegmentum of the thalamus and midbrain on the efferent
side.

The corpus striatum complex, however, makes its appearance
in lower vertebrates within the lateral olfactory area, and there
is evidence that, as its phylogenetic differentiation progressed
under the influence of increasing numbers of ascending thalamic
fibers, the receptive apparatus of the thalamic tracts and the
related motor neurons discharging into the tegmentum segre-
gated away from the apparatus receiving the descending second-
ary olfactory fibers, thus forming the true striatum.

The lateral olfactory area, moreover, in all vertebrates has an
important hypothalamic connection comprising both ascending
and descending fibers, the tractus pallii of fishes and the olfactory
projection tract of mammals. And this hypothalamic connec-
tion, unlike the thalamic apparatus, maintains throughout the
veretebrate series its physiological relationship with the lateral
olfactory centers.

The following physiological factors, accordingly, must be
recognized as influencing the morphogenesis of the lateral wall
of the vertebrate cerebral hemisphere:

1. The primary influence of the lateral olfactory tract.

258 C. JUDSON HERRICK

2. Ascending thalamic systems (somesthetic, optic, auditory).
These came forward originally into the lateral olfactory area,
thus effecting various somatic-olfactory correlations. This type
of correlation probably survives to some degree in the mamma-
lian pyriform lobe and in part of the amygdala. But the corpus
striatum, and in mammals the neopallium, are largely emanci-
pated from the olfactory influence. (The striatum is not entirely
so, as shown by Cajal’s description (’11, p. 723) of fibers arising

Fig. 35 Diagrammatic horizontal section through the brain of the frog at the
level of the interventricular foramén, to illustrate the relations of the ventro-
lateral olfactory tract, the dorsal olfactory projection tract, the amygdalo-
pyriform tract, and the lateral forebrain bundle.

from neurons of the cortex of the pyriform lobe and entering
into the complicated intrinsic neuropil of the lentiform nucleus.)

3: Ascending and descending hypothalamic connections. This
is a very primitive connection and apparently originally put the
non-somatic systems (as the term somatic is used above) of the
hypothalamus into relation with the lateral olfactory area.

4. The habenular connections. These are related to the
olfactory centers of the hemisphere only and so far as known are
efferent with reference to the hemisphere.

THE AMYGDALA IN AMPHIBIA 259

5. Commissural connections.

6. Correlation tracts to and from the medial wall.

In the course of differentiation of the cerebral hemisphere each
of these six factors may be variously subdivided and the com-
ponent parts related physiologically among themselves and to
other parts of the brain as required by the mode of life of each
species.

com.amg.-~

d. amg.hab~ he
tramghab-
hab.~

Fig. 836 Diagram of the frog brain similar to the last to illustrate additional
connections of the amygdala.

8} —— ote

The urodele hemisphere exhibits no regions in the lateral wall
behind the olfactory bulb which are related specifically to any of
these systems. The anuran dorsolateral and _ ventrolateral
parts of the hemisphere are here incompletely distinguishable
only by the internal connections of their dominant fiber tracts.
The entire lateral wall, except perhaps the caudal part of the
ventrolateral nucleus, receives fibers of the lateral olfactory
tracts, and hence is to be regarded as lateral olfactory nucleus,
though these fibers are very unevenly distributed within it.
Most of the fibers of the lateral olfactory tract enter the compact

260 Cc. JUDSON HERRICK

dorsal division of the tract, though more diffusely arranged fibers
are freely distributed throughout the lateral wall.

The rostral end of the hemisphere is least affected by the as-
cending diencephalic systems, and it is to this relatively unspecial-
ized part of the olfactory area that the term nucleus olfac-
torius anterior has been applied. In some animals it is an
extensive region bordering the olfactory bulb or even extending
out into it; in others the elaboration of special centers like the

Fig. 37. Diagrammatic horizontal section through the brain of Amblystoma,
to illustrate the connections of the ventrolateral nucleus of the cerebral hemi-
sphere. :

tuberculum olfactorium has invaded this territory so that but
little such undifferentiated secondary olfactory tissue remains.
In urodeles this anterior nucleus is fairly extensive on the
lateral wall, especially dorsally.

The ventrolateral area of the urodele hemisphere is dominated
(except at the rostral end) by the lateral forebrain bundle. This
contains, in all of the urodele species which I have examined,
an extensive system of fine unmyelinated fibers which ascend
from the pars dorsalis thalami, which I first called tractus
thalamo-corticalis (710, p. 434) and later tractus thalamo-frontalis
Cig, Dp. 208):

THE AMYGDALA IN AMPHIBIA 261

These fibers unquestionably are the precursors of the mamma-
lian thalamic projection fibers for somesthetic, visual, and audi-
tory sensibility. Within the hemisphere they terminate through-
out the ventrolateral area and to some extent at least in the
dorsolateral area, especially toward the posterior pole. The
return paths for these somatic systems are the strio-thalamic
and strio-tegmental fibers of the lateral forebrain bundle. These
are myelinated and unmyelinated fibers which arise (so far as
now known) from neurons of the ventrolateral area.

The same part of the urodele hemisphere which receives the
ventrolateral olfactory tract and the thalamic connections last
described is also functionally related with the hypothalamus by
the olfactory projection tract (tractus pallii), whose ascending
fibers also reach the caudal part of the dorsolateral area.

Fibers pass from the lateral wall of the hemisphere to the
habenula from the dorsolateral area by the massive tractus
cortico-habenularis lateralis and from the ventrolateral area
by the smaller tractus amygdalo-habenularis. The ventrolateral
area is connected with the opposite side of the brain through the
anterior commissure by decussating and commissural fibers.
And, finally, the lateral areas are ‘broadly connected with the
medial areas of the hemisphere by extensive systems of associa-
tional fibers, some of which form the four tracts already mentioned
(diagonal band, stria terminalis, and dorsal and ventral olfactory
projection tracts), while others in more diffuse formation cross
the dorsal and ventral angles of the hemisphere.

Briefly summarizing our analysis of the lateral wall of the
urodele hemisphere, we notice:

1. The olfactory bulb is lateral, with an imperfectly differenti-
ated accessory bulb posteriorly which receives fibers from the
supposed vomeronasal organ, as well as fibers from other parts
of the nasal sac.

2. The lateral wall of the hemisphere is thin and undifferenti-
ated except for a thickened region laterally of the interventricular
foramen, here termed the ventrolateral nucleus of the hemisphere.

3. The entire lateral wall, except perhaps a portion of the
ventrolateral nucleus, receives secondary olfactory fibers and

262 Cc. JUDSON HERRICK

therefore retains the primitive characteristic of the lateral olfac-
tory area. The differentiation within this area is chiefly correla-
ted with the entrance of various specific non-olfactory systems
of fibers.

4, The region bordering the olfactory bulb is least affected
by these immigrant systems and is termed the nucleus olfactorius
anterior.

5. Most of the fibers of the lateral olfactory tract enter the
dorsolateral area, which is therefore the chief lateral olfactory
nucleus.

6. Posteriorly this quadrant receives ascending thalamic fibers
from the lateral forebrain bundle, ascending hypothalamic
fibers from the olfactory projection tract, association fibers
relating it with the adjacent primordial hippocampus (dorso-
medial area), and commissural fibers from the hippocampal com-
missure; it discharges the lateral cortico-habenular tract to the
epithalamus and may also discharge into the hypothalamus.
These connections define this region as primordial pyriform lobe
(see beyond, p. 271).

. 7. The relations of the ventrolateral area to the ascending and
descending fibers of the lateral forebrain bundle indicate that
here is to be sought the functional equivalent of the corpus
striatum (paleostriatum); but since there is no group of neurons
related with the thalamic and tegmental fiber systems of the
lateral forebrain bundle which do not also receive secondary
olfactory fibers (by the ventrolateral olfactory tract), or ascend-
ing hypothalamic fibers (by the olfactory projection tract), or
both of these systems, it is clear that in the urodele there is no
anatomical structure to which the name corpus striatum can
properly be given.

- 8. The amygdaloid complex of reptiles (Johnston, ’15; Crosby,
17) is related to the lateral olfactory tract, the hypothalamic
olfactory projection tract, the stria medullaris for the habenula,
the stria terminalis, and the diagonal band of Broca. The first
three of these fiber tracts (and probably the other two) are related
with the ventrolateral nucleus of the urodele hemisphere, but
not with any specific group of neurons within this nucleus. Here

THE AMYGDALA IN AMPHIBIA 263

then, is the physiological primordium of the amygdala, but this
nucleus as a morphological entity has not emerged from the com-
mon olfacto-striatal matrix. In the Anura, on the other hand,
there is such a nucleus amygdalae with all of the characteristic
connections enumerated above.

V. THE COMPARATIVE ANATOMY OF THE STRIO-AMYGDALOID
COMPLEX

Returning now to the Anura, we find the lateral wall of the
hemisphere considerably more highly organized. The dorso-
lateral quadrant has assumed definite form as lateral olfactory
nucleus anteriorly and pyriform lobe posteriorly, much as in
lower mammals. It is sharply separated from the underlying
ventrolateral quadrant by external and ventricular sulci and
between these a histologically defined zona limitans lateralis.
The fiber connections are as in urodeles, though more clearly
defined.

In the ventrolateral quadrant at the rostral end immediately
behind the olfactory bulb there is an undifferentiated region
which may receive some secondary olfactory fibers, though
our preparations do not reveal them. Behind this is a true
corpus striatum, characterized by connection with the lateral
forebrain bundle with no olfactory component. ‘This striatal
region is enlarged at the level of the interventricular foramen.
Closely associated with it, but structurally distinct, is a true
amygdala not related to the lateral forebrain bundle, which is
characterized by: 1) a specific ventrolateral olfactory tract from
the vomeronasal formation of the olfactory bulb; 2) a specific
relation with both ascending and descending hypothalamic fibers
of the olfactory projection tracts; 3) a habenular connection
through the stria medullaris; 4) a dorsal connection with the
septum and preoptic nucleus through the stria terminalis; 5) a
ventral septal connection through the diagonal band of Broca;
6) a specific commissural connection with the opposite amygdala;
7) a connection with the overlying pyriform lobe through the
tractus amygdalo-pyriformis.

264 C. JUDSON HERRICK

The anuran amygdala in emerging from the urodele strio-
amygdaloid complex of the ventrolateral nucleus has assumed all
of the olfactory, habenular, and hypothalamic connections of
the ventrolateral area—connections which, however, it shares
with the overlying pyriform lobe. It possesses, moreover, some
features which are not shared with the pyriform lobe, viz.:

1. The commissural connection with the opposite amygdala
in the anterior commissure. The commissural fibers of the
pyriform lobes in mammals pass through the hippocampal
commissure, a strictly pallial formation (dorsal psalterium,
Cajal, 11, p. 715). In the frog also some fibers of the hippocam-
pal commissure pass through the primordial hippocampus,
around the dorsal angle of the hemisphere, to arborize throughout
the entire pyriform lobe (P. Ramén y Cajal, ’05, pl. 16, fig. 4).
I have seen similar fibers from the hippocampal commissure
curving around the posterior pole of the hemisphere to reach
the pyriform lobe.

2. In the frog the descending fibers of the olfactory projection
tract seem to come chiefly (perhaps exclusively) from neurons
of the amygdala, while the ascending fibers of this tract terminate
freely in both the amygdala and the pyriformlobe. In mammals,
on the other hand, Cajal (’11, p. 721) describes the fibers of this
tract as arising chiefly as axons of cortical neurons of the pyri-
form lobe, though he believes that some fibers arise also in the
amygdala. The absence of a true cortex in the amphibian
pyriform lobe probably accounts for this difference.

3. The secondary olfactory connection of the amygdala is
specifically derived from the vomeronasal formation. This point
suggests some further reflections. .

The absence of the vomeronasal organ in fishes, its very rudi-
mentary condition in urodeles, especially the lower forms, and
its close association in many types with the posterior nasal
aperture suggest that its primary function is in some way linked
with the reception of sensory substances coming from the mouth
cavity, though from the-relations in higher forms it may be
inferred that this primary function ‘has been subject to further
modification.

THE AMYGDALA IN AMPHIBIA 265

Broman (’20) in a reexamination of the morphological and
physiological relationships of the vomeronasal organs of mam-
mals and reptiles has shown that in both of these groups this
organ is normally filled with liquid, not air, and that there is a
pumping mechanism by which the liquid olfactory medium may
be alternately sucked into and expelled from it. This mechanism
is very different in the two cases, but in both it appears to be
under control and to act rapidly.

In lizards and serpents there is a mechanism by which the
liquids of the mouth cavity can be forced into and out of the
vomeronasal organ, thus providing “an ideal mouth-smelling
organ.” In mammals the pumping apparatus is much more
complex and diversified in, different groups, in some species
adapted to draw liquid olfactory media into the vomeronasal
organ from the mouth cavity, in others from the nasal cavity,
and in most cases from both of these cavities. Broman suggests
that the liquid media derived from the respiratory passages of
the nose enable Jacobson’s organ to function in macrosmatic
mammals in ‘tracking’ by odors (Spiirsinn). In this connection
it should be borne in mind that the ordinary olfactory epithelium
of mammals is not directly excited by gaseous media, as some-
times taught, but the odorous substances must first be dissolved
in the liquid which bathes the olfactory membrane. Neverthe-
less it is not improbable that the liquid of the respiratory pas-
sages may absorb a larger amount of the odorous substances and
this more concentrated medium, when sucked into the vomero-
nasal organ, would give to the latter an enhanced olfactory
efficiency as a distance receptor.

Broman has not investigated the Amphibia, but from the ob-
servations of Bruner already cited (p. 214) it seems very probable
that aquatic ‘double-smelling’ Amphibia developed first a lateral
diverticulum of the nasal sac in close relationship with the
choana especially adapted to serve as a mouth-smelling organ,
and that from this simple beginning the true vomeronasal organs
of Anura and Amniota have been derived. Broman (’20, p. 188)
himself reached a different conclusion, viz., “that the organon
vomeronasale Jacobsoni is nothing other than the old water

266 Cc. JUDSON HERRICK

olfactory organ of vertebrates adapted for life on land,” an un-
tenable position which he unfortunately attempted to support
by a reference to the nervus terminalis which reveals a total
neglect of the recent contributions dealing with the innervation
of this region.

The Anura possess a well-developed vomeronasal organ in
typical position with a specific vomeronasal nerve related to a
specific part of the olfactory bulb, which in turnis connected with
the amygdala by a specific olfactory tract. It is strongly sug-
gested that the emergence of a morphologically circumscribed
amygdala in Anura is directly correlated with the specificity of
its physiological relation with the vomeronasal organ.

All of the fiber-tract connections of the anuran amygdala are
probably present in some form in urodeles—certainly the most
important ones are clearly recognizable. In urodeles these
fiber systems converge into a single non-specific “ventrolateral
nucleus.” It is not improbable that in the more highly differenti-
ated brains of different types of fishes some of these elements are
present but dissociated in various ways. In the Anura the inte-
grating factor which has brought these elements together into a
single correlation mechanism and detached this center from the
more generalized ventrolateral nucleus of the ancestral form seems
to be the vomeronasal organ, for this specific connection is the only
obvious physiological factor which is added in the anuran brain.

If, as suggested by Seydel and others (p. 214), the vomeronasal
apparatus was differentiated in connection with the opening of
the posterior nasal aperture and the consequent passage of
olfactory media from the mouth cavity into the nasal sac, it is
obvious that from the beginning of this evolutionary process an
intimate physiological relationship existed between olfactory
excitations of this type and the gustatory excitations arising
within the mouth cavity.

Our knowledge of the ascending gustatory path in the brains
of vertebrates is very meager. In teleosts it has been shown
(Herrick, ’05, p. 415) to pass from a reflex correlation center in
the isthmus region (the Rindenknoten of Mayser, superior
secondary gustatory nucleus of later authors) to the region of the

THE AMYGDALA IN AMPHIBIA 1 207

hypothalamus, where its further connections are unknown. In
lower Amphibia a similar condition was described (Herrick,
17, p. 248), but the fibers could be traced forward only as far
as the level of the III nerve. They probably reach the hypothal-
amus.”

It is not improbable that there is a direct or indirect connection’
between the tertiary ascending gustatory system just mentioned
and the hypothalamic nucleus of the olfactory projection tract,
thus putting the gustatory system into physiological relationship
with the olfactory centers of the lateral wall of the cerebral
hemisphere. This latter connection is very primitive.®

Now in the Amphibia the opening of the internal nasal passage
(choana) introduced a new functional factor into the peripheral
olfactory complex, viz., the reception of odorous emanations from
the mouth cavity. The central correlation of these newly
acquired excitations with those received from taste buds in the
course of the feeding reactions naturally accompanied this change
in peripheral relations, the mechanism for this being already
present in generalized form in the broad connection between
the lateral olfactory area and the hypothalamus by way of the
olfactory projection tract. This is the condition in urodeles.
With the appearance of a vomeronasal organ in definitive form

2In the passage cited (’17, p. 249) I referred to the secondary visceral (and
gustatory) nucleus as belonging ‘‘in the midbrain, rather than in the medulla
oblongata.’’ This statement requires correction; for in view of the fact that the
nucleus in question lies behind the sulcus isthmi (as described on page 222 of the
same paper), it must be regarded as rhombencephalic if the sulcus isthmi is
correctly interpreted as marking the rostral boundary of the rhombencephalon.
The corresponding region in teleosts (Uebergangsganglion) has been shown to
be rhombencephalic by Palmgren (Acta Zoologica, vol. 2, 1921, p. 91).

’Dart (’20, p. 17) has asserted that the olfactory projection tract (tractus
pallii) ‘‘is the only ascending tract from the hypothalamus,”’ and that there are
no ascending fibers from this region in the medial forebrain bundle. In this he
ignores or rejects a considerable body of positive observation, some of it based
on ample material stained by the Golgi method, published by competent ob-
servers. From the study of my own preparations I have no doubt that there are
such ascending fibers in the medial forebrain bundle. The medial wall of the
cerebral hemisphere must, accordingly, be recognized as sharing with the lateral
wall in the reception of ascending hypothalamic tracts. The broad functional
differences between these regions must be interpreted in terms of other factors.

268 C. JUDSON HERRICK

in the Anura there followed differentiation of a specific vomero-
nasal nerve, vomeronasal formation, ventrolateral olfactory
tract, and amygdala, as these are found in the brain of the frog.

In the various fishes the relationships of the structures here
under consideration are exceedingly diverse. In ganoids and
elasmobranchs the available descriptions do not indicate the
presence of any differentiated center which can be compared with
the anuran amygdala with any very definite assurance, though in
some forms some of the corresponding functional connections
are present in a less specialized arrangement more nearly
comparable with that seen in the urodeles, and in sharks the
differentiation has advanced in a different direction.

In teleosts the nucleus teniae of the carp as described by
Sheldon (’12) resembles fairly closely in position and fiber connec-
tions the anuran amygdala. It receives fibers of the lateral
olfactory tract and sends fibers to the habenula. The tractus
pallii (tractus olfacto-hypothalamicus lateralis) reaches the
adjacent “nucleus pyriformis,” but is not described as entering
the nucleus teniae. Similar relations are described in other
teleosts by Goldstein (’05), Kappers (’06, p. 11), and others.
(The nucleus teniae and tractus teniae of Johnston’s descriptions
of fishes refer to quite different structures from those so named
by the authors just mentioned. Johnston, ’11, p. 35.)

The absence of a differentiated vomeronasal organ in fishes
raises the question how far these structures can be compared
with the anuran amygdala. It is probable that the primordial
elements of the amygdalo-pyriform complex of higher brains
(and of the hippocampus as well) are here represented in various
combinations; but that in none of these species are these elements
combined to form an amygdala of the type seen in the frog, for
in the absence of the vomeronasal apparatus the integrating
factor essential in the latter case is lacking.

Living forms of Amphibia, it is now generally agreed, cannot
be regarded as ancestral to any Amniota, so that the anuran type
of amygdala is not necessarily the point of departure for this
complex as found in‘higher forms. ‘The elements here associated
may be found dissociated or combined in other patterns in

THE AMYGDALA IN AMPHIBIA 269

amniote brains. But the specificity of the relationship between
vomeronasal organ and amygdala in the frog suggests further
lines of interesting inquiry.

In those reptiles and mammals which possess well-developed
vomeronasal organs is there a similar specific relationship with
any component of the amygdaloid complex?

In the course of amphibian evolution the first component of
the amygdaloid complex to emerge in morphologically recog-
nizable form is the anuran amygdala as described in the preceding
pages, and this apparently occurred under the specific influence
of the vomeronasal system of peripheral connections. In rep-
tiles and mammals, as indicated by Johnston, Crosby, Elliot
Smith, and others, other formations from different sources are
added to the amygdaloid complex in various patterns. Clearly,
then, the structure here designated in the brain of the frog as
amygdala cannot be homologized with the whole of the amyg-
daloid complex of mammals. This has various other elements
which may be represented in the urodele ventrolateral and dorso-
lateral areas in undifferentiated form.

Johnston (715, p. 419), in connection with his description of
the amygdaloid complex of reptiles, has shown that the mamma-
lian amygdala is a composite of elements of diverse origin. The
definition of the mammalian amygdala is, accordingly, very diffi-
cult, and in the present state of our knowledge it is of doubtful pro-
fit to attempt to fix the mammalian homologies of the anuran
amygdala. In reptiles, however, this can be done, as we have
seen (p. 236) with some measure of probability (cf. Johnston,
15, p. 418, and Crosby, ’17, p. 349), though the absence of the
vomeronasal organ in Crocodilia (Zuckerkandl, ‘10, p. 35) leaves
this question in some obscurity.

The amygdaloid complex of mammals has been the subject of
numerous investigations, of which the most extensive is that of
Volsch (06, 710). This author, like Cajal (11, p. 724), denies
direct connection of the olfactory tract with any part of the
complex in mammals. This, I think, is very questionable, for
my own observations, mentioned below, suggest that the lateral
olfactory tract does reach one part of it (the presubicular area),

270 C. JUDSON HERRICK

and such a connection has often been described by others (e.g.,
Edinger, ’11, p. 389.)

In any case, it is clear from Cajal’s description that the entire
amygdaloid complex of the mouse is under indirect, if not direct,
influence from the lateral olfactory tract, for it is enveloped by
cortex of the pyriform lobe and is penetrated by axons from
these cortical neurons, which in turn receive fibers from the lateral
olfactory tract directly. There is also a “tangential tract of the
amygdala” (Cajal, ’11, p. 725, fig. 463, d) between the distribution
area of the lateral olfactory tract and the presubicular area of
the amygdala.

Rothig (’09), in Didelphys marsupialis, describes a nucleus
amygdalae under the pyriform lobe and rostrally of this two
cellular areas which are probably to be regarded as parts of the
amygdaloid complex as this is usually defined in mammals. One
of these is a large-celled nucleus occupying the angle ventrally
of the pyriform lobe which receives fibers from the lateral olfac-
tory tract; this he terms nucleus of the tractus bulbo-corticalis
(our lateral olfactory tract). The second nucleus (his nucleus
taeniae semicircularis) lies dorsally and internally of the first
and in very close association with it. From it arise fibers of the
stria terminalis (his pars ventralis taeniae‘).

Kappers (’08, p. 241) describes in Hypsiprymnus an origin of
stria terminalis fibers from a similar nucleus of the lateral olfac-
tory tract and nucleus taeniae. These two nuclei in the aggre-
gate probably correspond pretty closely with the medial large-
celled nucleus of Johnston (’15, p. 415) in the turtle and the
ventromedial nucleus of Crosby (717, p. 348) in the alligator.

In rodents the relations are much the same. My observations
on the brain of the rat show (in conformity with previous de-
scriptions of rodents) that the most ventral component of the
amygdaloid complex is the area presubicularis. This is a sharply
circumscribed spherical nucleus of large cells lying close to the
surface at the rostral end of the pyriform lobe in much the same

4My own observations upon the opossum, Didelphys virginiana, suggest

that Réthig’s account of the relations of the pars ventralis taeniae requires
revision, but the discussion of this matter must be reserved until a later time.

THE AMYGDALA IN AMPHIBIA LA DAFAI|

relations as in the rabbit (Winkler and Potter, ’11). In Weigert
sections it appears to receive fibers from the overlying lateral
olfactory tract. It gives rise to a fascicle of myelinated fibers
of small size which enters the stria terminalis and can be followed
separately for the entire length of the stria. They form the
most ventral fibers in the dorsal loop of the stria, and many of
them can be followed into the anterior commissure. These
relations are confirmed in sections stained by Cajal’s method.
Stria terminalis fibers are related to the entire extent of the amyg-
daloid complex and the overlying cortex of the pyriform lobe.
Some fibers associated with these can be definitely followed into
the habenula, but I have not demonstrated that any of these
arise specifically from the area presubicularis.

So far as present information justifies conclusions, it may be
suggested that the structure here designated amygdala in the
Anura is represented in mammals by the presubicular area and
the adjacent region termed subiculum cornu Ammonis by Winkler
and Potter (’11, fig. xi) and the paleostriatal element of the
amygdaloid complex by Dart (’20), though the homology probably
is not exact. The remainder of this complex in the Amniota does
not seem to be so closely related with the olfactory system and is
not represented as a differentiated structure in the frog.

That the olfactory component is not essential to the integrity
of the mammalian amygdala is clear from the fact that in the
totally anosmic dolphin the amygdaloid complex is of large size,
in marked contrast with the atrophied pyriform lobe and hippo-
campus (Zuckerkandl, ’87, p. 113; Addison, 715). The fact that
in the dolphin the stria terminalis, though present, is very small
(Addison, ’15) supports the view that this tract is to be regarded
as largely composed in mammals of descending olfactory pro-
jection fibers.

The relation of the amygdala to the pyriform lobe requires a
further comment. Ihave earlier (’10, pp. 479, 487) compared the
dorsolateral quadrant of the amphibian cerebral hemisphere with
the mammalian pyriform lobe, stating, ‘‘it is represented in mam-
mals as one of the components of the pyriform lobe,” and ‘‘the
function of the amphibian dorso-lateral part, as of the pyriform

pat hoe C. JUDSON HERRICK

lobe of mammals, is evidently the correlation of olfactory with
other exteroceptive impressions belonging to the somatic sen-
sorysystems.” This conclusion was based chiefly on three ana-
tomical features of this part of the frog brain: 1) it receives the
strong dorsolateral olfactory tract; 2) it is broadly connected by
correlation fibers with the hypothalamus and primordial hippo-
campus; 3) it receives ascending non-olfactory fibers from the dor-
sal part of the thalamus. Kappers (Folia Neurobiologica, Bd. 5,
1911, p. 625) questions the validity of the last point on the ground
that the mammalian pyriform is not known to receive projection
fibers from the dorsal thalamus, and if such are present in the frog
their distribution area should be considered neopallial. No such
homology as I have suggested can be regarded as exact, for the
amphibian brain nowhere exhibits a degree of differentiation
which permits clear-cut definition of regions precisely comparable
with those of mammals; but the question raises a principle which
justifies a further inquiry.

The pyriform lobe of lower mammals is clothed with true cor-
tex, i.e., superficial gray matter, and rostrally this passes by in-
sensible gradations into the lateral olfactory nucleus, a part of
the primordial subcortical olfactory area. Immediately internal
to this pyriform cortex are the lentiform nucleus and the amyg-
dala. In lower mammals the lentiform nucleus is related by in-
numerable fibers (probably mostly descending) with the cerebral
peduncle, there is a dense neuropil between the lentiform nucleus
and the pyriform cortex (Cajal, ’11, p. 722, fig. 462), and the
nucleus is traversed by very numerous radiating fibers connected
with the pyriform cortex. So intimate, indeed, is the connection
between the nucleus and the overlying pyriform cortex that Cajal
(loc. cit., p. 514) says that the lentiform nucleus seems to be a
dependency of the pyriform cortex. This relation is probably
reciprocal.

Now in the frog there is no true cerebral cortex in the lateral
wall of the hemisphere, but the other structures mentioned are
related as just described. And ascending thalamic projection
fibers from the thalamo-frontal tract cross the zona limitans la-
teralis from the ventrolateral quadrant (primordial striatum) to

THE AMYGDALA IN AMPHIBIA 273

end in the distribution area of the dorsolateral olfactory tract.
It is true that the telencephalic distribution of these thalamic pro-
jection fibers in mammals is typically neopallial, and these fibers
might be regarded as neopallial in the Amphibia if only there
were any neopallium here. The fact is that they reach an area
which is dominated by the lateral olfactory tract and which con-
tinues to be so dominated even in mammals where it develops
true cortex of the pyriform lobe.

The process of the gradual unfolding of the neopallial cortex
under the combined influence of functional factors primitively
present in the paleostriatum and pyriform lobe has been briefly
sketched by Elliot Smith (’19) and need not be reviewed here;
but it is interesting to note that during the entire course of this
evolutionary history the cortex of the pyriform lobe retains a
very primitive character and even in the highest mammals is
structurally and physiologically transitional, on the one hand to
the subcortical lateral olfactory nucleus and on the other hand
to the subcortical corpus striatum and the amygdala. In high-
er brains the neopallial components of this undifferentiated com-
plex as seen in Amphibia have passed on to more elaborate evo-
lution elsewhere, but the corpus striatum (or part of it) and the
pyriform lobe remain as residual structures in practically their
primitive relationships.

In this confusing region it is very difficult to determine where
are the limits between cortical and subcortical structures. In
fact, the terms cortex and pallium are currently employed so
loosely that such a determination is impossible without a more
satisfactory definition of these terms than has hitherto been
published. A clear reformulation of these concepts in the light
of recent comparative and embryological studies is urgently
needed.

The amygdala of the frog is clearly subpallial in the sense that
Gaupp (’99) and I (10) have defined the pallium; in accordance
with the same criteria, the greater part, at least, of the mamma-
lian amygdala appears to be pallial. But careful embryological
work will be required before the mammalian complex can be re-
solved. If Iam correct in my belief that the amphibian amygdala

274 Cc. JUDSON HERRICK

is represented in the mammalian presubicular area (and some
contiguous tissue), evidently the name “pallial element of the
amygdaloid complex” given to this region by Dart (’20, p. 18)
is Inappropriate.

The anatomical configuration and connections of the amygda-
loid complex suggest that as a whole it possesses a certain physi-
ological unity and that the component parts of this integrated
complex are diversely represented in various vertebrate -types, in
accordance with their respective modes of life. The amphibian
relations suggest, further, that the correlation of olfactory, gusta-
tory, and perhaps other excitations arising from food within the
mouth was the original integrating physiological factor. The
complex of higher forms probably includes also connections of
the tactual and muscle senses.* Though the differentiation of the
vomeronasal organ peripherally was the initial point of departure
for the fabrication of the amygdala, the complex, once developed,
retains its individuality in the absence of the vomeronasal organ
(alligator, man), and even of the entire olfactory system (dolphin).

The mammalian amygdala, like many other complex correla-
tion centers, is a mechanism in which there converge into final
common paths numerous very diverse kinds of peripheral excita-
tion—some of visceral (interoceptive) type and some of somatic
(exteroceptive) type. In some species one of these types may be
dominant, in other species the other, and the analysis of these
components must be carefully worked out in each vertebrate
group before we can generalize profitably. In the Amphibia
the visceral components are clearly predominant, and this appears
to be the primitive situation.

The vigorous and rather temerarious attack of Dart (20, p.
18) upon this method of cerebral analysis is evidently based

5 The tactual factor may in some animals be very important. In this con-
nection it is interesting to note that Broman (’20, p. 178) considers that the
forked tongue. of serpents is accessory to the vomeronasal organs which here
are very large, the extruded tongue taking up odors and carrying them back to
the openings in the roof of the mouth of the ducts of these organs, which in-
deed the tips of the tongue may enter. The serpent’s tongue is generally re-
garded as a very efficient tactile organ, and this function also, accordingly,
would naturally be largely represented in the amygdaloid complex of serpents.

THE AMYGDALA IN AMPHIBIA 245

upon an inadequate appreciation of the underlying physiological
principles involved. It is generally recognized that the analysis
of the vertebrate nervous system in terms of the physiological
modalities of the related peripheral end-organs has greatly clari-
fied an obscure field. The detailed application of this method
of attack is the distinctive contribution of the so-called American
school, beginning with an adequate analysis of the functional
components of the peripheral nerves and carrying this analysis
more and more deeply into the intricacies of the higher correlation
centers.

The first generalizations to be derived from these studies
(Strong, 95) brought out in sharp relief the fundamental nature
of the distinction drawn by some of the early English physiologists,
notably Bell and Gaskell, between the somatic and visceral
systems of nerves, a conception later employed with brilliant
results by Sherrington.

This method of analysis has tones the central nervous
system as helpfully as the peripheral, and its value in the fore-
brain is not less than in the medulla oblongata. In fact, the
tardy progress in the comparative morphology of the forebrain
has been due chiefly to lack of exact knowledge in just this field,
to ignorance of just what peripheral systems are represented
in the higher correlation centers of the various animal types
under investigation. Before we can reach final conclusions
regarding the origin of the cerebral cortex we require much
more detailed information regarding the connections of the pre-
existing subcortical mechanisms of the forebrains of critical
types. This applies especially to the centers of correlation
between olfactory and non-olfactory systems.

What names are applied to these systems is a relatively unim-
portant matter, but it is necessary to bear in mind that the olfac-
tory system itself is a complex in which visceral (interoceptive)
and somatic (exteroceptive) elements are always present. To
deny or ignore this dual nature of the sense of smell is to close
the door to further progress.

The olfactory apparatus, whatever may have been its primitive
physiological character, has in fishes (as in most higher verte-

276 Cc. JUDSON HERRICK

brates) come to be one of the dominant exteroceptive systems,
that is, it functions as a distance receptor. Nevertheless,
throughout the entire phylogenetic history of this system it
exhibits pronounced interoceptive or visceral functions associated
with the selection of food, functions closely similar to those
of the. gustatory system (Herrick, ’08). In man, a micros-
matic animal, the sense of taste and the interoceptive aspects
of the sense of smell are so intimately related that we are unable
introspectively to separate them, and physiological experimenta-
tion is required to effect this analysis. One wonders whether,
if we were provided with a functional vomeronasal organ and
its specific central apparatus as this is seen in the frog, our sensory
experience would not thereby be enlarged. :

VI. SUMMARY

1. The vomeronasal organ (Jacobson’s organ) first appears
in Amphibia, apparently in correlation with the opening of the
posterior nasal aperture. In Urodela it is present (if at all) in
very rudimentary condition, but in Anura it has assumed defini-
tive form as a diverticulum from the medial side of the nasal sac
provided with its specific innervation, the vomeronasal nerve.

2. The vomeronasal nerve terminates in a specific part of the
olfactory bulb, the vomeronasal formation, and from the latter
a specific secondary path, the ventrolateral olfactory tract,
passes to a differentiated amygdala in the ventrolateral wall of
the cerebral hemisphere.

3. The other connections of the anuran amygdala are: with
the opposite amygdala through the anterior commissure; with the
adjacent pyriform lobe; with the medial olfactory areas (septum
and preoptic nucleus) ; with the habenula; and with the hypothal-
amus. The connection last mentioned is by descending and
ascending fibers of the olfactory projection tract, which is the
equivalent of the tractus pallii of fishes.

4. The frog possesses a true corpus striatum (paleostriatum)
specifically related with the lateral forebrain bundle and quite
separate from both the lateral olfactory nucleus and the amyg-
dala.

THE AMYGDALA IN AMPHIBIA Aut

5. In some urodeles the supposed primordium of the vomero-
nasal organ sends nerve fibers to the accessory olfactory bulb,
which also receives fibers from other parts of the nasal sac and is
therefore not the exact equivalent of the vomeronasal formation
of the frog. There is a ventrolateral olfactory tract whose
fibers arise from the accessory olfactory bulb and also from other
parts of the bulb and, accordingly, are not specifically related to
the vomeronasal organ, as in the frog.

6. The ventrolateral area of the urodele cerebral hemisphere
comprises an undifferentiated strio-amygdaloid primordium in
which the fiber connections described above as characterizing
the corpus striatum and amygdala, respectively, of Anura are
inextricably mingled. This is termed the ventrolateral nucleus
of the hemisphere. The hypothalamic connection of this nucleus
(olfactory projection tract) is large and well defined.

7. In both Urodela and Anura the hypothalamic nucleus of
the olfactory projection tract is in functional connection with the
septal areas through the medial forebrain bundle, with the dorsal
part of the thalamus through the tractus thalamo-hypothalamicus
cruciatus, probably with gustatory and other visceral systems
of the hypothalamus, and possibly with the pars nervosa of the
hypophysis.

8. The lateral wall of the amphibian cerebral hemisphere is
derived from the lateral olfactory area of more primitive verte-
brates. The course of differentiation of this region has been
largely determined by the penetration into it of ascending dience-
phalic fibers of two systems: 1) somatic fibers from the dorsal
part of the thalamus through the thalamic projection tracts,
and 2) visceral fibers from the hypothalamus through the
olfactory projection tracts. The first factor has dominated the
further differentiation of the striatal and neopallial complexes;
the second that of the amygdala and pyriform-lobe complexes.

9. The olfactory system in all vertebrates has a twofold func-
tional rdle: 1) visceral or interoceptive in relation with the
selection and digestion of food, and 2) somatic or exteroceptive
in relation with the adjustment of the organism to environmental
conditions. With the opening of the posterior nasal aperture

278 Cc. JUDSON HERRICK

in the Amphibia, the vomeronasal organ appears to have been
differentiated in connection with the first of these factors, namely,
for cooperation with taste and perhaps various tactual and other
general forms of sensibility within the mouth, in the analysis
and appropriate disposition of the contents of the oral cavity.

10. The anuran amygdala was apparently differentiated under
the influence of the specific sensory excitations coming from the
vomeronasal organ. So far as the pre-existing nervous pathways
connected with the ventrolateral area of the cerebral hemisphere
conducted excitations physiologically congruous with those from
the vomeronasal organ, these were integrated into a morphologi-
cal unity as we see it in the amygdala of the frog.

11. The amygdaloid complex having been thus integrated
under the dynamic influence of vomeronasal excitations, further
complication of the system progressed in higher vertebrates in
various directions by the incorporation of other related sensory
systems, with perhaps profound modification of the functional
aspect of the complex asa whole. So great may be this deviation
from the primitive physiological pattern in some cases that the
suppression of the vomeronasal organ, as in man, or even of the
entire olfactory system, as in the dolphin, does not destroy the
integrity of the surviving components of the amygdaloid com-
plex, which retains its individuality in modified form.

THE AMYGDALA IN AMPHIBIA 279

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Resumen por el autor, Chi Ping.

Sobre el crecimiento de las mayores células nerviosas del
ganglio superior cervical del simpatico de la rata albina desde
el nacimiento hasta la edad adulta.

Las mayores células nerviosas de este ganglio presentan un
rapido crecimiento durante los primeros veinticinco dias después
del nacimiento, seguido de una fase de crecimien to mds lento.
Después de la pubertad existe una tendencia en la hembra a pre-
sentar células mayores que las del macho dela misma edad. El
crecimiento de estas células esta mas intimamente relacionado con
la edad que con el peso del individuo. Unas pocas células en
fases avanzadas se presentan al tiempo del nacimiento; su
nimero aumenta lentamente antes del décimoquinto dia y
después fecha mds rapidamente. Relacionado con esto hay una
disminuci6én en el nimero depequefias células que se transforman
en células mayores. Estas pequefias células parecen transfor-
marse en mayores niimeros en la hembra durante la edad mds
avanzada. La relacién nucleo-plasmatica aumenta desde 1 hasta
4 en la época del nacimiento, hasta llegar a ser proximamente 1
a 12 durante la edad adulta. En la edad adulta las células
grandes pueden distribuirse en tres grupos segun la distribucién de
sus granos de Nissl. Existen unas cuantas células binucleadas
durante todas las edades y después de la pubertad también hay
células pigmentadas. Las observaciones anteriores se refieren a
ratas albinas standard. En los albinos cruzados entre si (inbred) e
existen siempre células mds pequefias, con una pequefia diferencia
sexual en el tamafo de las células y una relacion nucleo-plas-
matica menor. El autor no explica esta diferencia entre. las dos
castas de ratas albinas.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, JUNE 27

ON THE GROWTH OF THE LARGEST NERVE CELLS IN
THE SUPERIOR CERVICAL SYMPATHETIC GAN-
GLION OF THE ALBINO RAT—FROM BIRTH TO
MATURITY

CHI PING

The Wistar Institute of Anatomy

SIX CHARTS AND ONE PLATE
INTRODUCTION

- This paper contains observations on the largest nerve cells in
the superior cervical sympathetic ganglion of the albino rat.
The purpose of this study is to trace the growth of these cells
by their increase in diameter in relation to the age and size of the
animal. In order to compare the possible differences in
growth in the sympathetic nerve cells due to sex, a male and a
female rat of each age were used throughout the series of
observations.

The author desires to express his sincere appreciation and
gratitude to Dr. M. J. Greenman for granting him the privileges
and facilities of the Institute for this investigation, and to Dr.
H. H. Donaldson, under whose direction the work was carried on
and whose valuable advice and guidance enabled him to formu-
late his results.

MATERIAL

The material used for this investigation consisted of sixteen
pairs of albino rats, of known ages, from 1 to 365 days. Besides
these, two females of 540 days and 570 days, respectively, were
used for comparison. All these were obtained from the animal
colony at The Wistar Institute and belonged to the so-called
‘standard strain.’ In selecting the specimens, five-day intervals
were taken between each two ages from birth to thirty days,
but from this age onward greater intervals were used. The
body weight, body length, sex, and age of each rat were recorded.

281

282 CHI PING

For comparison and control a second limited series of inbred
albino rats was also used. The data for this series are given on
page 303. Up to the introduction of this series the paper deals
only with albino rats of the ‘standard’ strain.

TECHNIQUE

The rat was etherized and, after the necessary measurements
had been noted, was completely eviscerated. The superior
cervical sympathetic ganglion was removed from each side. In
the removal care was exercised to avoid distortion of the tissue,
for mechanical injury to the ganglion is likely to affect the size
and shape of its cells. As the ganglion is small, it was deemed
necessary to remove it in the mass of other tissues which closely
invest it.

Both ganglia from each rat were prepared, but only one was
used for measurements. No distinction between right and left
was made in the record.

Aiming at a satisfactory preservation of the natural size of the
cells, I followed King’s (?10) recommendation of Bouin’s solution
for fixation. The ganglia from older rats were fixed in the solu-
tion for twenty-four hours, while for those from the younger
ones—from birth to twenty-five days old—the period was reduced
to twelve hours. Such a reduction of the fixation period has
given satisfactory results.

The specimen was washed in different grades of alcohol, from
70 to 98 per cent, containing a small amount of carbonate of
lithium. By so doing the yellow tinge given to the tissue by the
fixation was completely removed. ‘The specimen stayed in the
alcohols of lower grades for twelve or more hours, and in the 90
and 98 per cent alcohol for about one hour. It was finally trans-
ferred to cedar oil for twenty-four hours for complete dehydra-
tion. Paraffin of 52° was used for imbedding. By employing
an electric bulb above the container the paraffin was kept melted
only in its upper layer in the jar, the specimen sinking to the
contact line between the melted and unmelted paraffin.

Under these conditions the specimen could be left in the
paraffin for thorough penetration as long as seemed necessary
without danger of overheating.

SYMPATHETIC CELLS: ALBINO RAT 283

Serial sections of the entire ganglion were cut 8 u in thickness.
Heat from an electric bulb was used in flattening the sections.
The slide was placed underneath the bulb, so that the water that
served to float the sections on the slide, also chilled them from
beneath, when they were spread by the heat.

The procedure in staining was as follows: The sections were
passed from xylol down through the graded alcohols to water,
and then put for five minutes in a saturated solution of lithium
carbonate, after which they were stained for two or three minutes
in a one-third saturated solution of thionin. They were then
passed up through the graded alcohols to xylol and mounted in
acid-free balsam. So far as possible, the plane of section was
made perpendicular to the short axis of the ganglion, thus giving
the maximum area.

MEASUREMENTS OF THE CELLS AND NUCLEI

The cells and nuclei of the ganglion were measured with an
eyepiece micrometer, using a Zeiss ocular no. 6, and objective,
4mm. Each division in the micrometer scale was equivalent
to 4.47 u. The measurements were made in the following way:
In the case of each specimen a section at the middle of the series
was selected. Starting both ways from this, four more sections
were selected, two in each direction, by skipping every other
section. In this manner five sections altogether were chosen
and marked for study. In each of the five sections the two
largest cells were measured; thus ten cells in all were measured
in each ganglion.

There were four principal points kept in mind when selecting
the cells for study: First, the cells must be the largest in the
section; second, they must be uninuclear; third, the nucleus
must be located at or near the center of the cell and must be
fairly large; fourth, in the nucleus at least one nucleolus must be
present.

Under these conditions, the measurements made on the cells
and nuclei are considered to represent the maximal longitudinal
and transverse diameters of each cell and nucleus taken close
to their median planes. It was often found in this study that

284 CHI PING

the boundaries of a cell body were obscure. Furthermore, the
distribution of the Nissl granules was rather irregular (as will
be described later), so that neither the longitudinal nor the
transverse diameter could be measured according to the extent
of the stainable mass.

After the measurements had been taken, a sketch of the section
with the two cells measured therein was made, and the nucleoli
in these cells were noted, so that in making measurements for
the second time the same cells could be identified by their location
and the number of the nucleoli. As a matter of routine, the cells
in each ganglion were measured twice, a considerable time being
allowed to elapse between the first and second measurements.
The procedure in measuring did not follow in the order of age or
of body weight of the animal, as given in the tables, but was
purposely haphazard, and in making measurements for the second
time, the records were taken without referring to those already
made. ‘The values used are the means of the two series.

By this procedure prejudice was avoided and a more accurate
determination of the size of the cells and nuclei obtained. The
records thus made were tabulated in detail, but the averages of
the values for the ten cells in each ganglion are those used for
the tables, charts, and discussion which follow. The individual
data have been filed in the archives of The Wistar Institute.

The square roots of the products of the longitudinal and trans-
verse diameters of the cells and of the nuclei, respectively, for
each ganglion were averaged, and the mean was multiplied by
4.47, the value in u of one division of the eyepiece scale. In
table 1 the diameters of the cells and nuclei thus computed are
arranged according to age, and in table 2 according to the body
weight of the animal.

Based on the records in tables 1 and 2, charts 1 and 2 were
plotted. Chart 1 shows on age the graphs for the diameter of the
cells and nuclei in micra and chart 2 the same relations on body
weight.

In the graphs for the cells in chart 1 we see in the increase
before puberty only chance variations between the male and the
female in diameter of the cell body, but after the rat has attained

SYMPATHETIC CELLS: ALBINO RAT 285

the age of eighty days (body weight about 100 grams) which is
the period of puberty (Donaldson, ’15, The Rat, p. 21) there
appears a tendency for the cells to be larger in the female than
in the male. It will be noted, however, that at the age of eighty-
nine days, and also at 250 days, the male exhibits larger cells than
the female of the same age. This discrepancy is explained when
we take the body weights of the males into consideration. As
given in table 1, the body weight of the male rat eighty-nine days
old is twice that of the female of the same age, and the dis-
crepancy is even greater in the case of the male at 250 days.

These males should be expected to have larger nerve cells by
virtue of their body weight, and when a correction is made for it,
the values for the male cells should fall below those for the female
at these ages also. In. general one may say that the female,
after reaching puberty, has these cells larger than the male, if
the body weight of the male does not too greatly exceed that of
the female. As regards the nucleus, however, chart 1 exhibits
a less clearly marked sex difference.

The fact that there is a better growth of the cell bodies in the
female is more clearly illustrated in chart 2, in which graphs for
the diameters of the cells and nuclei have been plotted on body
weight. From birth to the time just before puberty, the varia-
tions in the growth of these cells in the two sexes are similar to
those shown in chart 1.

Just before puberty, when the rat weighs about 60 grams, the
female becomes gradually more advanced in the growth of these
cells and overtakes the male of the same body weight. The
growth of the nerve cells in the female also shows a more regular
course than that of the male. The growth of the nuclei at the
corresponding ages of the two sexes follows in the same manner
as that of the cell bodies, although the difference is relatively
small.

It is proper to keep in mind, however, that when the compari-
son is made on the basis of body weight, the female is normally
the older, and, further, that in several other growth changes the
female tends to be precocious; both of these influences would
tend to produce larger cells in the female under these conditions.

286 CHI PING

TABLE 1

Computed diameters of the largest cells and nuclei according to age. From the
superior cervical sympathetic ganglion of the albino rat

I B c D BE F

sees ace BODY BODY Computed diameter in u SUE OR

WEIGHT LENGTH ————— NUCLEUS TO
Cell Nucleus | DIAMETER OF CELL
days grams mm,

rot 1 O20 50 19.5 11.4 isi
Q 1 6.3 51 19.8 10.2 1:1.93
oi 5) 9.0 63 22.1 NO 7 1:2.06
2 5 11.0 65 21.3 10.5 1:2.03

(of 11 15.0 UC 24.9 iGjp il 1:1.90 -
19 11 14.0 73 26.4 isi il EZ RO2:
Gt 16 18.9 83 2523 13.1 1:1.92
9 16 19.0 81 vase 11.2 1:2.06
of 20 Blo A 102 26.4 IPAS 1274501
ie) 20 29.5 99 23.6 11.8 1:1.99
ot 25 23.8 93 26.6 12.6 eee
2 25 25.0 95 27.3 1% 122.15
rou 29 40.7 112 PAVE 12.0 og
°) 29 16.4. 82 24.8 12.2 1:2.038
ol 42 61.4 129 27.0 13.4 Det
2 42 43.5 105 PH aes 13.2 1:2.05
GH 48 1O5 eee 156 29.0 13.4 SPE?
je) 48 49.7 120 27.0 11835 1L 12205
on 60 51.6 124 Dail 74 133524 1:2.06
ce) 62 53.8 iLike/ Pile Al 163, 1 12207
oe 81 63.3 128 27.4 13.3 1:2.06
¢ 80 83.7 142 26.6 12.8 1:2.09
oh 89 143.5 173 32.4 13.0 1:2.49
fe) 88 73.0 135 29.2 a2 1282
of 124 iis at 174 Df oll 13.0 1:2.08
i) 124 Oo 157 30.5 13.8 Lez 2
a 171 198.2 192 27.0 So Al LBP
Q 171 123.8 159 30.9 12.8 1:2.41
rot 250 230.0 207 36.8 15.4 1:2.38
2 250 98.0 160 30.6 14.2 Ze
ot 365 186.0 203 29.6 13.5 1:2.20
i) 365 170.6 186 31.4 118305; 2a
2 540 151.3 184 30.7 13.4 TER ad)
©) 570 Poll 169 33.4 14.3 1:2.34

SYMPATHETIC CELLS: ALBINO RAT 287

On examining the ratios between the values at one day and at
365 days, as shown in columns D and E of table 1, it is found that
the cells in the male have increased in diameter 1.55 times, and
in the female 1.58 times, while the nuclei in the male have increased
1.17 times and in the female 1.32 times. This shows that
the difference between the male and female in the growth of the
nuclei in the course of one year is greater than that in growth
of the cells, but the cells in both sexes have a greater rate of
growth than do the nuclei, as indicated in table 1.

Diameters of CellS and Nuclei in micra
1 a

ar]
0 20 50 100 200 300 350

Chart 1. Based on table 1 and giving the computed diameters of the cells
and their nuclei according to sex—on age in days. Males
Females ------------ ---

The graphs in chart 1 show that the increase in the diameter
of the cell body is rapid for the first twenty-five days and then
becomes slower. ‘There is a similar change in the nucleus, though
the change in the rate of growth in this case is less marked. From
25 to 365 days the diameters of the cells and of the nuclei of the
two sexes have increased as shown in table 3.

In column F of table 1 are given the ratios between the diameter
of the cell body and that of the nucleus. Generally speaking,
the cell has about twice the diameter of the nucleus throughout
the series of measurements as given in table 1, but if we consider
the ratios carefully, we see that there is an increase in the ratios

288 CHI PING

TABLE 2

Computed diameters of the largest cells and nuclei—on body weight—together with
the nucleus plasma ratios—from the superior cervical sympathetic
ganglion of the albino rat

A B c D E F

SEX fi BODY BODY Computed diameterin uw | nucLEUS PLASMA

WEIGHT LENGTH RATIOS

Cell Nucleus
days grams mm.

o 1 5.6 50 19.5 11.4 1: 4.0
g 1 6.3 | 51 19.8 10.2 LENGrS
oh 5 9.0 63 Be Il 10.7 1B ffete:
g 5 11.0 65 21.3 10.5 IE 8
1} 11 14.0 73 26.4 13.1 ME ies
rot 11 15.0 77 24.9 Seal Te be8
g 29 16.4 82 24.8 12.2 1: 6.8
oe 16 18.9 83 25.3 13.1 TieorZ
g 16 19.0 81 23). 1 1aS2 LE: Pah
ot 25 23.8 93 26.6 L226 1: 8.4
2 25 20.5 95 293 WET te Be)
2 20 29.5 99 23.6 11.8 ike eo)
rot 20 31.7 102 26.4 12.5 1: 8.0
rou 29 40.7 112 Pf od 12.0 ORS
g 42 43.5 105 Pile, 13.2 il carvends
Q 48 49.7 120 27.0 13.1 Ie fod
rou 60 51.6 124 Die 13.2 ILS Zo a
eo} 62 53.8 117 Zell 13.1 IS Cfcte:
fof 42 61.4 129 27.0 13.4 MS eres
rou 81 63.3 128 27.4 13.3 lieve
2 88 73.0 135 29.2 13.2 Le 9.8
g 80 83.7 142 26.6 12.8 Een
Q 250 98.0 160 30.6 14.2 L980
of 48 105.1 156 29.0 13.4 lie sOEd
2 124 107.1 157 30.5 13.8 TOES
g ileal 123.8 159 30.9 12.8 12 118i.4
g 570 1271 169 33.4 14.3 ILE le Tf
of 89 143.5 173 32.4 13.0 1:14.4
ioe 124 151.1 174 ira 13.0 aoe)
g 540 151.3 184 0). 13.4 ISIE
@ 365 170.6 186 31.4 13.5 LEG
CG 365 186.0 203 29.6 13.5 1 ee)
oh 171 198.2 192 27.0 13.1 UE ad
(ott 250 230.0 207 36.8 15.4 eG

SYMPATHETIC CELLS: ALBINO RAT 289

as age advances, as they are, respectively, 1 :1.72 and 1: 1.938
for the youngest male and female; 1 : 2.11 and 1 : 2.15 at twenty-
five days, and i : 2.34 for the oldest female.

This increase is therefore most marked during the first twenty-
five days. By comparing the progress from birth to twenty-five
days with that from twenty-nine days to 365 days, one can -
appreciate the rapid increase in amount of cytoplasm within the
former period, as contrasted with the slower increase in the course

Aco
: rch

a
race

COE

eee :
LI |

Body weight—gms.

ee
125 15 175 200 250

Chart 2 Based on table 2 and giving the computed diameters of the cells
and their nuclei according to sex—on body weight. Males
Females --------------- ‘

of a much longer time. It is fair to say, therefore, that the ratios
change but slowly after the first twenty-five days. This agrees
with the statement of Donaldson and Nagasaka (’18) on the
ventral horn cells, that after twenty-four days the nucleus-
plasma ratio increases but slowly.

VARIABILITY WITHIN A GIVEN GANGLION

The number of large cells examined in each ganglion is hardly
great enough to permit of satisfactory statistical treatment,
but it has been thought worth while to tabulate for each animal
the range and average of the diameters of the cells and of the

290 CHI PING

nuclei, entering these according to age as in table 1. In a fairly
graded series of measurements we may expect to find the average
for the series close to the mean of the limiting values, and a little
study of table 4 shows this to be the case.

MORPHOLOGY OF THE LARGE CELLS
Plate 1. (Figures 1 to 7)

In considering the morphology of the cells in the superior
cervical sympathetic ganglion, it must be recalled thatfrom its
cells arise several classes of fibers—pupillodilator fibers, motor,
vasomotor, pilomotor and secretory fibers. It is a priori possible,

TABLE 3

Increase in diameters of cells and nuclei from 25 to 365 days

GAIN

- DIAMETERS SEX 25 DAYS 365 DAYS
Absolute Percentage
ML Me a
nen J 26.59 29. 60 3.01 11.3
CS ee peatscooreyae chee a Penene elie 9 27.26 31.38 4.12 15.1
see o 12.60 13.45 0.85 6.31
i CR ME UURESE fe Wag Park 9 12.65 13.54 0.89 6.58

that the several functions thus indicated are correlated with cell
characters that are distinctive, but at the moment we have
nothing to contribute to the solution of this problem.

When young, the cells of the superior cervical ganglion are
very similar in appearance to those of the young spinal ganglion,
and practically all of them are more or less elongated with
processes at one or both ends. Each cell has a large clear nucleus
surrounded by a little cytoplasm. This cytoplasm is homogene-
ous in structure and stains uniformly. Those coarse Nissl
bodies, which are found in the cells at later ages, are totally lacking.
Usually each nucleus has a single, dark stained, nucleolus, but
occasionally there may be found more than one. ‘This condition
continues from birth to five or six days of age, when differentia-
tion begins in the cytoplasm of these immature cells.

SYMPATHETIC CELLS: ALBINO RAT 291

TABLE 4

Giving the ranges in the values for the diameters of the largest cells and their nuclet
in the superior cervical sympathetic ganglion of the albino rat—
arranged according to sex and age

CELLS NUCLEI
sex Rent edrer fades lad 8
Diameter Range Diameter Range
days grams rv vu ip rv

rou 1 5.6 19.5 22.0-18.0 11.4 14.0- 9.4
Q 1 6.3 19.8 24.0-17.4 10.2 1220 et
rou 5 9.0 poze 25.0-20.0 10.7 14.0- 9.0
Q 5 11.0 A-3 25.0-18.3 10.5 12.3- 9.0
9 11 14.0 26.4 30.0-23.0 igo 15.0-11.0
ou 11 15.0 24.9 30.4-22.0 13 Al 1327-16
ou 16 18.9 25.3 29 .0-22.0 13.1 14.0-12.0
9 16 19.0 3 51 25.0-20.3 ila 14.0-10.0
2. 20 295.0 23.6 27.0-19.4 fies 16.0-10.4
oe 20 317.0 26.4 29 .0-24.0 12.5 14.0-11.0
ou 25 238.0 26.6 31.0-23.0 12.6 14.0-11.0
Q 25 25.5 B73 31.0-24.7 1287. 14.0-11.0:
fe) 29 16.4 24.8 26 .0-22.7 TOR: 14.0- 9.4
ou 29 40.7 OT 30.0-24.7 12.0 14.0-10.0
Q 42 43.5 27.2 31.0-24.7 13.2 16.0-11.0
ou 42 61.4 27.0 30.0-24.7 13.4 16.0-12.0
Q 48 49.7 27.0 29 .0-24.7 1391 15.0- 9.0
ou 48 105.1 29.0 33.0-26.4 13.4 15.0-12.6
ou 60 51.6 272 30.0-24.0 1322 15.0-12.4
Q 62 53.8 27.1 30.0-24.0 13.1 15.0-11.6
fo) 80 S307 26.6 29 .0-25.7 1258 13.7-11.6
oe 81 63.3 27.4 31.5-25.7 133 15.6-11.0
fe) 88 73.0 29.2 31.0-26.0 12 14.6-11.0
ou 89 143.5 32.4 36. 7-28.5 13.0 14.6-11.0
9 124 107.0 30.5 33.0-29.0 13.8 16.0-13.0
roe 124 151.1 27 98522510) | 13.0 14.0-11.7
2 171 123.8 30:9 33.7-29.0 1258 13.7— 9.7
rou 171 198.2 27.0 32.4-24.7 1351 14.0-12.4
fe) 250 98.0 30.6 33 .0-26.6 1459 16.0-14.0
of 250 230.0 36.8 39.0-35.6 15.4 18.0-14.4
Q 365 170.6 31.4 38.0-28.5 13 755 lek 61248
roe 365 186.0 29.6 33.7-26.6 13.5 | 16.0-12.0
Q 540 151.3 30.7 34.4-28.3 13.4 | 14.7-11.0
Q 570 1275 33.4 36.7-29.0 14.3 | 16.0-14.0

292 CHI PING

At birth or during the first days of life there are found among
the young cells a few advanced cells which appear conspicuously
different from the rest. In these advanced cells the cytoplasm
may be already differentiated, even at birth. The stainable
Nissl granules, which are of course much finer than those found
at later ages, are evenly but distinctly distributed through the
entire contents of the cell. Among these granules some clear
spaces appear which seem to indicate the differentiation of the
homogeneous cytoplasmic mass, and this change in the advanced
cells must have commenced during fetal life.

When the young cells begin to develop, there is the same dif-
ferentiation of the cytoplasmic mass, and the stainable bodies
arrange themselves in the same way as those seen in the advanced
cells. Hereafter more differentaition will be found in them and
they grow to resemble the advanced cells in appearance.

Taking this as the starting-point in the morphological develop-
ment, we see among the comparatively large cells in the ganglion
four types which probably appear one after the other as here
given in the course of growth.

Type 1. The advanced cells and the cells which are trans-
forming into advanced cells, as described above, belong to this
type. ‘There is a beginning of aggregation of the Nissl granules
and a growth of the unstainable ground-substance in the cells.
This type is common during the first twenty days of postnatal
life (fig. 2). .

Type 2. The Nissl bodies are larger than in type 1 and
ageregated at the periphery of the cells, forming a ring within
which is a comparatively clear portion of the ground-substance
surrounding the nucleus. The Nissl bodies stain much darker
than in type 1. The nuclear membrane, the nucleoli, and the
reticular structure in the nucleus are distinctly visible. There
are frequently two or more nucleoli in one nucleus. This type
is common in the period between twenty and sixty days (fig. 3),
but may also be found at birth (fig. 1).

Type 3. Instead of being distributed at the periphery, the
Nissl bodies are aggregated around the nucleus, leaving a rather
clear space at the periphery of the cell. In some of the cells they

SYMPATHETIC CELLS: ALBINO RAT 293

are more crowded at certain regions close to the nucleus, forming
dark masses, but some of them may be loosely scattered toward
the periphery. It is in this type of cell that difficulties have
often been encountered in making out the boundary between
the cell wall and the supporting tissue, because the unstained
ground-substance is chiefly distributed at the periphery of the
cell. This type is common after twenty days of age, but is not
infrequently found after sixty days (fig. 4).

Type 4. The cells resemble the first type in the arrangement
of Nissl bodies, but the stainable bodies are much coarser. There
is a considerable evenness in their distribution, though here and
there we find a larger dark stainable mass resulting from their
aggregation. Whether this type is developed from the preceding
type through modifications in the course of development or
whether it is directly derived from type 1, without undergoing
the various changes as in types 2 and 3, is a matter to be settled
through more detailed investigation (fig. 5). This type is
characterized by the dense appearance of Nissl bodies throughout
the entire cell body, not leaving much space for the ground-
substance, and is common at the age of 124 days and later.

In interpreting these several types it is to be recalled that
the cells of this ganglion have severa] different functions and
there’ always remains the possibility of a correlation between
function and morphology.

Besides the four types of cells described above, binuclear cells
are found at all ages until the rat is very old. In recording
them, special care needs to be taken. As the cell wall of the
sympathetic cell is at times difficult to distinguish, two uninuclear
cells in close contract with each other may frequently resemble
one cell with two nuclei. In order to avoid error due to such
misleading appearances, the precaution has been taken to use an
oil-immersion lens in distinguishing the true binuclear cells
from those which resemble them. The cells which have their
cytoplasm discontinuous somewhere between the nuclei or a
constriction at the middle, either slight or pronounced, as the
one figured by Apolant (’96, Majer’s ‘cell bridge,’ fig. 8, pl.
XXIII), were not considered as of the true binuclear type.

294 CHI PING

TABLE 5

Giving the number of the cells with two nuclei and of the cells showing pigment, at
different ages. Superior cervical sympathetic ganglion—albino rat.
In each case the numbers are for one ganglion only

NUMBER OF BI-| CELLS WITH

SEX AGE BODY WEIGHT | BODY LENGTH | Oop ear CELLS Se
days grams mm.
oe 1 5.6 50 2 0
°) 1 6.3 51 2 0
Gi 5 9.0 63 1 0
2 5 11.0 65 2 0
of 11 15.0 Ue 1 0
2 11 14.0 73 2 0
St 16 18.9 83 il 0
2 16 19.0 81 1 0
of 20 eileerd 102 2 0
je) 20 29.5 99 4 0
ot 25 23.8 93 4 0
2 25 2020 95 3 0
St 29 40.7 112 3 0
©) 29 16.4 82 3 0
rou 42 61.4 129 5 0
Q 42 43.5 105 12 0
rot 48 105.1 156 a 0
ee) 48 49.7 120 2 0
Oo 60 51.6 124 5 0
Q 62 53.8 117 5 0
of 81 63.3 128 3 0
2 80 83.7 142 12 0
Co 89 143.5 173 a 0
2 88 73.0 135 1 1
ot 124 151.1 174 15 8
2 124 107.1 157 6 0
o 171 198.2 192 2 4
2 171 123.8 159 9 0
of 250 230.0 207 4 0
2 250 98.0 160 5 3
ot 365 186.0 203 6 3
2 365 170.6 186 + 2
Q 540 ely 184 2 4
2 570 127 169 3 13

{[% 4.0

Average of binuclear cere ° 44

SYMPATHETIC CELLS: ALBINO RAT 295

Every one of the cells recorded in table 5 had an unbroken layer
of Nissl granules around the two nuclei, and at: the middle of the
cell there existed absolutely no trace of any partition whatsoever
which might suggest the contiguous surfaces of two cells closely
erown together. Figures 6 and 7 show the binuclear cells in a
very young and in a comparatively old rat, respectively.

If we determine, by direct measurement, the nucleus-plasma
relation in this particular older cell (fig. 7), contrasting the volume
of both nuclei with that of the cytoplasm, we find a ratio of
1:5.0. This is almost as low as the ratio at birth, and indicates
that we are dealing with an increase in the nuclear mass not
accompanied by a corresponding increase in the cytoplasm.
This, so far as it goes, is an argument against the suggestion that
we have here two cells that are fused.

According to table 5, the occurrence of binuclear cells is not
related to sex. In many cases the numbers of these cells in both
sexes are equal or almost equal. There appears, however, to be
an increase in their number toward middle age, ranging from
sixty days to 365 days, with a possible decrease later.

Apolant found cells of the binuclear type in the superior
cervical ganglion of an embryo rabbit three weeks old, and
states that such cells persist in the older animal, when the cells
have been completed anatomically and physiologically. Accord-
ing to him, this is the result of direct nuclear division; about half
of the binuclear cells being formed during embryonic life and
the remainder later. It is not the purpose of this paper to deal
with the function and origin of this type of celis. Their appear-
ance in the postnatal stages of the rat, as recorded in table 5,
agrees with what Apolant points out as the course of the develop-
ment of the cells in the later ages of the animal. Carpenter and
Conel (714) noted this type of cells in considerable number in the
rabbit, guinea-pig, muskrat, and porcupine, but rarely, if ever,
did they find them in the sympathetic ganglion of the rat.

As these authors’ observations were made most probably on
one or on only a few stages of the rat, the small number of such
cells in the entire ganglion justifies their statement, in a way but
nevertheless the presence of the binuclear cells in the superior
cervical sympathetic of the rat is beyond question.

296 CHI PING

Incidentally, pigmented cells have been noted in the superior
sympathetic ganglion of the albino rat. The cells of compara-
tively young animals, from birth to eighty days, are entirely
free from pigment. At the beginning of puberty we occasionally
find pigment in one or two cells in an entire ganglion. The
number of the pigmented cells tends to increase as age advances,
as recorded in table 5. Some of the cells are only partly pig-
mented; a few are completely covered with these granules, the
nucleus remaining unaffected, while others are totally pigmented,
including the nucleus. The pigments appear yellow brown, or,
black in color, but whether this is merely a result of their relative
abundance or whether there are several sorts of pigment has not
been determined. The whole question of pigment in the Albino
nervous system seems worthy of a special investigation.

INCREASE IN THE NUMBER OF THE LARGE CELLS

The increase in the number of the large cells in the ganglion
during the first twenty days is an important event. This is
chiefly due to the rapid increase in diameter of the young cells
after ten or fifteen days of age. The large cells measure 19 to
25 uw in diameter, and are loosely scattered and intermingled
with small cells, as seen in each section. Disregarding their
finer differences, such a group of cells consists of three kinds:

1. The advanced cells. During embryonic development it is
known that the sympathetic trunks are formed through the
migration of some cells which pass from the spinal cord along the
paths of the communicating rami (Kuntz, 710). The advanced
cells in the superior cervical ganglion are the forerunners of the
neurones which come to this locality in “‘skirmish order—much
in advance of the others”’ and ‘‘they represent but a fraction of
the final number of large cells’? (Donaldson, 717). The number
of these cells during the first twenty days varies from one to
eight in the entire ganglion.

2. The moderately large cells. These cells constitute an
intermediate group between the advanced cells and the small
cells in the same ganglion during the first ten days of age. They
are not different from other younger cells in general structure and

SYMPATHETIC CELLS: ALBINO RAT 297

TABLE 6

Increase in number of large and advanced cells, 19 to 25 » in diameter, during the
first twenty days of life. “The ratios in the increase in the total number for both
sexes between one day and twenty days stand at the foot of the column. Superior
cervical sympathetic ganglion—albino rat

NUMBER OF LARGE
SEX AGE BODY WEIGHT CELLS AND OF
ADVANCED CELLS
days grams
o 1 9.6 188
Q 1 6.3 174
oO 5 9.0 289
2 5 iO 306
rot 11 15.0 291
2 11 14.0 301
fot 16 18.9 760
°) 16 19.0 584
fot 20 31.7 2508
& 20 29.5 2248
: o& 13.34
Ratios: ie 12.91

Number of large cells °

Chart 3 Based on table 6 and showing the number of large cells present in
the superior cervical sympathetic ganglion of the albino rat from birth to twenty
days of age. Males -—————-._ Females --------------- :

298 CHI PING

form, but they are distinguishable, owing to their larger size. ©
It is this group of cells which will appear later as the advanced
cells.

3. The growing small cells. These cells are small during the
first five days after birth, but some of them grow very fast toward
the end of twenty days, to a size equal to that of the other large
cells. There is a constant increase in the number of these smaller
cells which are growing. :

For the determination of the rate of increase in the number of
the large cells, 19 to 25 » in diameter, counting was undertaken.
The cells counted comprised those just described under | and 2.
Since the same large cell does not appear. in two successive sec-
tions, repetition in counting them is easily avoided. Table 6
gives the numbers of these cells. Based upon these numbers,
the graphs in chart 3 were plotted on age. In chart 3 the male
has a slightly higher rate of increase than has the female after
twelve days. When the animal reaches sixteen days, both sexes
show a more rapid increase, and the difference between them
becomes more evident. If the data are plotted in a like manner
on the body weight, they show similar relations. On the whole,
then, the data show that the increase of large cells during the
first sixteen days is relatively slow and afterward increasingly
rapid. Between the age limits here given the increase in the
number of large cells—sexes combined—is about thirteen-fold.

THE TRANSFORMATION OF THE YOUNG CELLS

During the later period of development there remain in this
ganglion a number of young cells which, in contrast with the
large cells, are slow in growth and which retain their neuroblastic
appearance for a considerable length of time (fig. 1). As already
noted, some nerve cells are precocious and many of them have
attained their maximum size at the end of twenty to twenty-five
days. It is most probable that the young cells found after
twenty-five days of age are largely rudimentary elements, and
some of them will never grow to the same size as the others.
Yet some development is going on in both their structure and
size, as is indicated by the constant decrease of their number

SYMPATHETIC CELLS: ALBINO RAT 299

TABLE 7

Showing the changes in the number of young cells from the period just prior to puberty
to the end of one year. Superior cervical sympathetic ganglion—albino rat

NUMBER OF YOUNG

SEX AGE BODY WEIGHT Raine
days grams

of 60 51.6 470
°) 62 53.8 471
of 89 143.5 362
g 88 73.0 345
ot 124 15Le1 326
2 124 107.1 252
rot 171 198.2 242
} 171 123.8 207
ot 365 186.0 105
Q 365 170.6 102

Sure jit) Ale)?
Ratios: {9 1:0

Number of youngcells

Bw
SEAR ER eso eseeeees

aI eS FT hehe

SRE AAS RSA

ia
pan Ba

0 100 150 200 ne sane 350

Chart 4 Based on table 7. Showing the changes in the number of young cells
in the superior cervical sympathetic ganglion of the albino rat between sixty and
365 days of age. Males ——————. Females --------------- ;

300 CHI PING

toward the end of one year. A study of the rate in the decrease
of the young cells will serve as a means of measuring this change
at later ages. For this purpose counting was undertaken.
Because of their considerable number during the early prepubertal
stage, as well as their small size and irregular distribution, it is
almost impossible to obtain a satisfactory value by a single
count, so that a second and a third count were usually made.

The numbers recorded in table 7 represent averages of three.
counts of young cells in each ganglion. The cells, selected and
counted as young cells, have the following characters: They
are 5 to 10 uw in long diameter; more or less pyriform, and the
cytoplasm is little differentiated.

The graphs in chart 4 represent the numbers of the cells as
given in table 7, on age. The decrease in number at first shows
no tendency for one sex to outrun the other, but a difference
appears soon after puberty, and such a difference in decrease of
the young cells between the male and the female persists till the
end of one year. The young cells of the female rat in relation
to age are transformed more rapidly than those of the male;
that is, these cells grow faster in the female. This phenomenon
is in accord with what has been seen in the growth of large cells
in diameter, as shown in charts 1 and 2.

DISCUSSION

There is reason to think that at birth the full number of cells
in the superior cervical sympathetic of the albino rat has been
attained and that no more cells wander in and mitosis is finished.
These cells appear to persist throughout the span of life.

Postnatal development of these cells consists in the enlarge-
ment of all parts of the neuron accompanied by differentiation.
In the nucleus there is less change in size than in the cell body.
The increase in number of nucleoli has been frequently noted,
but it is not within the scope of the present paper to discuss this
point.

Bringing the observations together, we see that the male and
the female do not differ clearly from each other in the growth of
these nerve cells until the animal has become sexually mature.

SYMPATHETIC CELLS: ALBINO RAT 301

The rate of increase in the number of the large cells in the
superior cervical ganglion is, if anything, a little lower in the
females before twenty days and a little higher later. On the
other hand, the increase in size shows only chance variations
during the first seventy days. These variations are subject to the
influence of both age and body weight of the young animal, but
puberty is attained, sex begins to be significant in addition to
the other two factors.

If there is not too much difference between the ages and the
body weights of the male and of the female, then the difference
found in the quantitative development of the cytoplasm between
the two may be attributed to this influence of sex. In table 8

TABLE 8

Giving, according to sex, the average computed diameters of the cell and the nucleus
for three groups of body weights of albino rats. In the last column are given the
ratios between the cell and the nucleus diameters. Data condensed from table 2

DIAMETERS

SEX ie daaad eF BODY WEIGHT Ses ie DIAMETER OF

Cell Nucleus NUCLEUS TO CELL

See Wap Gams i) Ge. | eal

sh 4 5.6- 18.9 22.9 12.1 1:1.89

Q 5 6.3- 19.0 23.1 11.4 1:2.03

a 7 23.8-105.1 27.2 12.9 Ieper

2 9 25.5-107.0 27.7 13.1 ede iltt

of 5) 143. 5-230.0 30.6 13.6 1:2.25

2 4 123.8-170.6 31.6 13.5 1:2.34

is given a condensed statement of the cell measurements according
to sex, based on body weights as these appear in table 2. For
the cell body the values are in favor of the female for all three
groups.

After puberty the sympathetic neurons in the female tend to
have larger cell bodies and the small cells transform more rapidly.

At the moment it would not be wise to infer that similar rela-
tions would be found in other sympathetic ganglia or in other
strains of rats; nevertheless, as they stand, the results agree with
the suggestion of Dunn (’12) that the mass of the peripheral
nervous system in the female albino rat is greater in proportion
to the body weight than in the male.

302 CHI PING

In table 9 are given the amounts by which the cell diameters
of the females differ from those for the males at four ages after
eighty days. The mean excess for the females is about 6.9 per
cent, which represents approximately an excess of 20 per cent
in volume. When’ a corresponding comparison is made for the —
diameters of the nuclei in these four groups, the average difference
according to sex is feund to be zero.

TABLE 9

Showing, in four age groups, the absolute and percentage difference in the cell
diameters of the largest cells in the superior cervical sympathetic ganglion of the
female albino rat as compared with the male, based on the values in table 1.
Because of the great difference in the body weights, the data for the growp at 250
days are omitted

CELL DIAMETER IN THE FEMALE DIFFERS FROM THAT IN THE MALE BY
AGE
Absolute u Percentage

days

80 —0.8 — 3.0

124 210 + 9.6

171 +3.9 +14.4

365 +1.8 + 6.0

INS SIN ae Oe ae RES RR ee Oe Cone oe ee =e (OY

On the size of these cells in the inbred albino rat

. To determine whether the size relations according to sex which
have just been described for albino rats belonging to the so-
called ‘standard strain’ are generally found, a series of inbred
Albinos was examined, for comparison.

The specimens used in this study were furnished by Dr. Helen
D. King. - These rats had been closely inbred for thirty-four to
thirty-five generations. Seven pairs were used, ranging from
eighty-nine days to 154 days and each pair was from the same
litter. The preparation of the specimens was made in the same
manner as that for the series just described.

The records on sex, age, body weight and length and the
measurements of the cells and nuclei are given in the following
table 10.

SYMPATHETIC CELLS: ALBINO RAT

Using table 10, chart 5 was plotted on age.

303

The graphs show

a slight difference in the size of the cells of the male and female.
The male, as indicated by the graphs, seems to have a better
growth in the cytoplasm than the female of the same age, but
the difference is small and cannot be considered as primarily

TABLE 10

Data on the inbred albino rats from the colony of Doctor King.

Diameters of largest

cells in the superior cervical sympathetic ganglion—on age

SEX AGE reereeen nec
a 89 144 176
9 89 100 156
o 103 232 202
9 103 203 189
a 123 206 193
Q 123 176 187
a 116 140 177
) 116 110 172
o 131 310 220
9 131 205 192
J 136 179 188
9 136 148 182
ee 154 251 212
9 154 188 189

30 {Diameters of Cells and Nuclei in micra

SERRA cee see eee 6eNSe eee! ZI
aE Le L194 canes

10

80 90 100 110 120

DIAMETERS

RATIO OF
DIAMETER OF

Cell Nucleus NUCLEUS TO CELL

KB B
21.90 12.90 1:1.69
21.60 13.00 1:1.66
24.60 14.05 I hen PS)
24.20 13.25 1:1.82
24.70 13.12 1:1.88
24.21 13.14 1:1.84
23.21 12.60 1:1.84
22.81 12.50 1:1.82
28.80 14.00 1:2.07
26.00 13.50 1:1.92
24.80 12.79 1:1.94
25.80 13.79 1:1.87
27.21 15.21 ie
26.00 13.45 1:1.93

BERBER SSSeee

po ae a ST ae |e |

130

140

Age—days

150

160

Chart 5 Based on table 10. Showing the computed diameters of the cells

and their nuclei, according to sex, on age in days (inbred Albino).

ents |e

1

304 CHI PING

due to sex, because in each pair the male has a greater body
weight than the female.

In table 11 the data have been ee according to body
weight. Using these, chart 6 was plotted. In only one instance
is the value for the female below that for the male in the case of
the cells, while the female values for the nuclei are always above
those for the male.

TABLE 11
Giving the computed diameters of the cells and their nuclei arranged according to

body weight. The same data as in table 10. Inbred albino rats.

DIAMETERS

dae’ | aaa || mORe, | eee 1 a eee

Cell Nucleus

Me im
Q 89 100 156 21.60 13.00 eM
se) 116 110 172 22.81 12.50 INSt} 1
ot 116 140 177 23.21 12.60 5ia2
rol 89 144 176 21.90 12.90 1:3.9
2 136 148 182 25.80 13.79 12550
Q 123 176 087 24,21 13.14 1.522
ot 136 179 188 24.80 12.79 1:6.3
Q 154 188 189 26.00 13.45 Ave
9) 103 203 189 24.20 13.25 LBs dl
is) 131 205 192 26.00 13.50 Gye!
rou 123 206 193 24.70 13.12 S554
rot 103 232 202 24.60 14.05 1:4.0
Sie 154 351 212 27.21 15.21 UA 7
rot 131 310 220 28.80 14.00 WS That

100 200 300

Chart 6 Based on table 11 and giving the compareda diameters of the cells”
and their nuclei, according to sex, on body weight (inbred Albinos).
Male ——————. Female -------------=- 5

SYMPATHETIC CELLS: ALBINO RAT 305

As shown in table 10, the female is smaller in size than the
male at each age, the female at equal body weight must therefore
be older, consequently the cells might have a slightly larger
diameter as the result of age, but. the difference is small. It
would be fair to say, therefore, that the cells, as well as the nuclei,
as shown in chart 6, do indicate a sex difference although it is
slight.

This result supports in principle the earlier findings on the
standards rats.

TABLE 12

To illustrate the way in which the ‘inbred’ differ from the ‘standard’ albino rats in
respect of the diameters of the largest cervical sympathetic cells
and their nuclei. Data from tables 1 and 11

DIAMETERS

AGE BODY WEIGHT
Cells Nucleus
ue Me
89 122* DATE 12.95 Inbred albino rat
124 129 28.80 13.40 Stock albino rat
Terenoe PAIDSOUGCH. yo asa obtener —7.05 —0.45
e IBETCENUADEH ase oe eae —32.0 —4.0
123 i91 24.45 13.13 Inbred albino rat
i7al 161 28.90 13.00 Stock albino rat
OX saltities ts: samuel: —4.45 +0.13
Banranee | Percentage pert seer oe efas

*The values given are the average for the male and femaie in each instance.

In table 12 is given a comparison of the diameters of the cells
and nuclei of the inbred Albino with those of the stock albino
rat. The data for the latter have been selected from tables 2
and 11. The sexes are combined. |

According to table 12, the inbred has its largest cells in the
superior cervical sympathetic ganglion decidedly smaller than
those in the stock albino rat of approximately the same body
weight. The nucleus, however, shows only a little difference
between the two forms, though this difference is in the same
direction.

306 CHI PING

In her studies on inbreeding, King (’18) states that the closest
form of inbreeding, continued for many generations, has not
caused a diminution in the average body weight of the inbred
rat at any age, and that through the selection of the largest and
most vigorous animals for mating, inbred rats are superior in
body size to the stock animals reared under similar environmental
conditions. Nevertheless, our data as they stand indicate that
in the inbred rats the largest cells in this ganglion are clearly
smaller in size than in the standard strain. It seems best not to
comment on this relation until studies have been made on the
wild Norway, and these I hope soon to undertake.

SUMMARY
A. Based on the data for the ‘standard’ strain

1. Between birth and maturity the largest cells in the superior
cervical sympathetic ganglion increase about 55 per cent in
diameter, while the increase in the nuclei is less than half of this
amount.

2. The growth occurs in two phases: the first phase of rapid
growth ends at about twenty-five days and the second phase of
less rapid growth continues to the end of the record. The present
data do not show a marked alteration in rate at puberty.

3. The size of these cells is more closely related to the body
weight than to the age of the rat, but there is a marked tendency
after puberty for the females to have slightly larger cells than
the males of the same age.

4. The nucleus-plasma ratio increases from 1 to 4 at birth to
about 1 to 12 at maturity.

5. At maturity the large cells may be classified in three groups:
1) those with Nissl bodies accumulated at the periphery of the
cell; 2) those with large masses of Nissl bodies accumulated
around the nucleus; 3) those with larger Nissl bodies mingled
with small ones, and more or less evenly distributed within the
cell. Moreover, a few binuclear cells are found, and in the older
rats some pigmented cells are present.

SYMPATHETIC CELLS: ALBINO RAT 307

6. Taking the ganglion as a whole, the large or advanced cells
may be present, though in very small numbers, even at birth.
This number is slowly increased up to about the fifteenth day,
after which the increase is more rapid. Correlated with. this
is a decrease in the number of small cells which are transformed
into the large cells. This transformation continues during the
first year and probably throughout life. It appears to occur
slightly earlier in the female.

B. Based on the data for the ‘inbred’ strain

7. The inbred rats ranged from 89 to 154 days in age. When
the values for the diameters of the cells and of the nuclei were
plotted on age, these values were greater for the males. The
-males were also consistently greater in. body weight. When
the values were plotted on body weight, the values for the females
were in general above those for the males. In this case the
females were older than the males with which they were compared.
It seems probable that at like ages and like body weights, the
females would show slightly higher values, but this may be
merely an expression of precocity in this growth change in the
females.

8. When these cells in the inbred rats are compared with those
in the standard animals, table 12, it is seen that while the nuclei
differ but little in diameter, the cells in the standard Albinos
have a diameter some 25 per cent greater than that found for the
inbred cells. It is to be noted that this difference in diameter
would make the volume of these cells in the standard Albino
about twice that in the inbred, while the nuclei differ but slightly.
This difference in the cells is definite, but the significance of it is
not discussed here.

9. The ratios of the diameter of the nucleus to that of the
cell are in the inbred distinctly less than in the standard Albino,
within the same age limits. Compare data in table 1 with those
in table 10. 3

10. The nucleus plasma ratios in the inbred are only about
half as great as in the corresponding cells of the standard Albino.
Compare data in table 2 with those in table 11.

308 CHI PING

LITERATURE CITED

ApoLant, H. 1896 Ueber die sympathischen Ganglienzellen der Nager. Arch.
f. mikr. Anat., Bd. 47, 8. 461-471. ;

CARPENTER, F. W., AND ConzL, J. L. 1914 A study of ganglion cells in the

sympathetic nervous system, with special reference to intrinsic sensory

neurones. Jour. Comp. Neur., vol. 24, pp. 269-279.

Donaupson, H. H. 1915 The rat. Reference tables and data for the albino
rat (Mus norvegicus albinus) and the Norway rat (Mus norvegicus).
Memoirs of The Wistar Institute of Anatomy and Biology, no. 6.
1917 Growth changes in the mammalian nervous system. The
Harvey lectures, series 12, pp. 133-150.

Donaupson, H. H. anp Nagasaka, G. 1918 On the increase in the diameters of
nerve-cell bodies and of the fibers arising from them—during the later
phases of growth (albinorat). Jour. Comp. Neur., vol. 29, pp. 529-552.

Donn, EvizasetH H. 1912 The influence of age, sex, weight and relationship
upon the number of medullated nerve fibers and on the size of the
largest fibers in the ventral root of the second cervical nerve of the
albino rat. Jour. Comp. Neur., vol. 22, pp. 131-157.

GASKELL, WattTER H. 1920 The involuntary nervous system. Longmans,
Green & Co. New York

Kine, Heten D. 1910 The effects of various fixatives on the brain of the
albino rat, with an account of a method of preparing this material
for a study of the cells in the cortex. Anat. Rec., vol. 4, pp. 214-244.

1918 Studies oninbreeding. I. The effect of inbreeding on the growth
and variability in the body weight of the albino rat. Jour. Exp. Zool.,
vol. 26, pp. 1-54.

Kuntz, A. 1910 The development of the sympathetic nervous system in mam-
mals. Jour. Comp. Neur., vol. 20, pp. 212-259.

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PLATE 1

EXPLANATION OF FIGURES

Cells from the superior cervical sympathetic ganglion of the albino rat. Fig-
ures 1 to 7 magnified in plate by 2000.

1 An advanced cell and several young cells; male, one day old.

2 An advanced cell with Nissl bodies evenly distributed; male, five days old.

3 A cell with Nissl bodies accumulated at the periphery, common between
twenty days and sixty days; female, twenty days.

4 A cell with Nissl bodies accumulated around the nucleus, common between
twenty days and sixty days and also found at later ages; male, sixty days old.

5 A cell with larger Nissl bodies more or less evenly distributed common
after one hundred days; female, 124 days old.

6 A binuclear cell; female, one day old.

7 A binuclear cell; male, one year old.

PLATE 1

ALBIN

SYMPATHETIC CELLS

CHI PING

oll

*

ees

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THE JOURNAL OF COMPARATIVE NEUROLOGY,
VOL. 33, NO. 4, OCTOBER, 1921°

Resumen por el autor, Chi Ping.

Sobre el crecimiento de las mayores células nerviosas del ganglio
cervical superior del simpatico en la rata noruega.

Las mayores células de este ganglio crecen de un modo muy
semejante al observado en el caso de la rata albina. Estas
células en la rata noruega presentan nucleos con los mismos
didmetros que las de la rata albina,: pero los cuerpos celulares
son algo mds pequefios. La relacién nticleo-plasmatica es menor
que en la rata albina. Las células binucleares son menos nu-
merosas en la rata noruega, pero las células pigmentadas son
solamente un poco mds numerosas. Existen unas cudntas
células vacuoladas que parecen ser caracteristicas de las ratas
noruegas salvajes mds viejas. En las albinas “inbred,” com-
paradas con el tronco tipo, estas células poseen ntcleos del
mismo tamafio, pero el didimetro celular es mucho menor; por
esta causa sus relaciones ntcleo-plasmaticas son muy bayjas.
No puede decirse todavia si estas diferencias se deben a la domes-
ticacién o es simplemente una particularidad de este tronco.

Translation by José F. Nonidez
Cornel! Medical College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, SEPTEMBER 26

ON THE GROWTH OF THE LARGEST NERVE CELLS
IN THE SUPERIOR CERVICAL SYMPATHETIC
GANGLION OF THE NORWAY RAT

CHI PING
The Wistar Institute of Anatomy and Biology
FIVE CHARTS

INTRODUCTION

This study is a continuation of my first work ‘‘On the growth
of the largest nerve cells in the superior cervical sympathetic
ganglion of the albino rat from birth to maturity” (Ping, ’21).

In that paper the significance of the age, the size of the animal,
and of sex on the growth of the cells was examined, and an un-
expected difference in the size of these cells was found in the
‘inbred’ as contrasted with ‘standard’ strain of Albinos. It
was deemed important, therefore, to examine the Norway rat
in the same way in order to determine how the size and the
growth changes in these cells were related in the wild Norway
to those found in the two domesticated albino strains.

MATERIAL AND TECHNIQUE

The eighty-five specimens of the wild Norway used in this
study belong to two groups from different sources. One group,
comprising twenty-two individuals, was reared at The Wistar
Institute, and the ages of these animals range from one day to
134 days.

The other group of sixty-three animals was collected from
different localities in Philadelphia and its vicinity, and the ages
of these are unknown. Their body weights range from 37 to
402 grams, corresponding to ages from twenty days to three
years, as generally estimated.

313

314 CHI PING

The technique employed in preparing the specimens is the
same as that given in my former paper. The trapped rats were
carefully examined when dissected, and all the specimens used
were considered normal. As the ages of the trapped rats were
unknown, the determination of the percentage of water in the
brain was made in each case, since by this means the approximate
age of the animal may be estimated, as pointed out by Donald-
son (10), Donaldson and Hatai (’11, and ’16).

TABLE 1
Giving according to body weight the computed diameters of the largest cells and
nuclei in the superior cervical ganglion of the Norway rat. Sexes separated.
Data condensed. Thirteen groups .

FORTY-THREE MALES FORTY-TWO FEMALES
m
Number eis Computed diameter Computed diameter eae, Number
ge Nucleus Cell Cell Nucleus CGE SE

grams Le ML ML LB grams
1 6 9.9 17.2 16.5 9.6 6 1
3 15 12.9 22.8 22.9 12.5 14 3
5 35 °12.4 23.4 23.2 itr 28 2
3 76 13.0 2020 PAD) 13.0 31 1
3 104 1335 i 26.5 24.3 PASO 51 8
2 117 13.2 Hf 26.0 320 73 2
3 157 13.3 26731 24.9 13.0 103 4
3 186 13.4 27.9 28.3 13.3 157 4
5 220 13.6 29.7 20.0 13.0 179 6
4 244 14.0 31.3 26.6 12.8 192 3
3 276 13.5 33.3 27.8 7 214 3
5 SYA} 14.0 Bie) 31.6 a7 227 2
3 385 S37 Sled) Beso il 13.9 258 3

In measuring the cells and the nuclei of the superior cervical
ganglion I followed the same procedure as in my former study
of the Albino. The data represented by the computed diameters
of the cells and nuclei, and their ratios, together with the sex,
age, body length, and body weight, as well as the percentage
of water in the brain, were tabulated in the first instance for
each individual according to increasing body weight, but these
data have been condensed for the purpose of this paper—and
only the condensed tables will be used for discussion. The

SYMPATHETIC CELLS: NORWAY RAT aie)

individual data are on file in the archives of The Wistar Institute
and available there for reference.

The values for the diameters of the cells and their nuclei are
the averages of measurements on the twenty largest cells in each
ganglion. ;

GROWTH OF THE CELLS

A. In relation to body weight

The computed diameters of the cells and the nuclei of the
eighty-five cases have been condensed to thirteen groups for

se YSIS a a a ed hehe Sa fed SSE
Diameters of Cells and Nuclei in micra [7] |_| REET ESE eRe
pep al sles

oS an See ata ees EE RRPeReeaees

san beeeeereeeeeettneee= ca SSS Lai
30

| |

BERG SESaaR eee
Se eS ok z= SSE EEE ER
— ee

BERR eerer aan tie CL EE eee
Remnant non ooo rab [alot
ae i) fr ff ea lhe ae att et
BREE EEE EEE EERE pet
peice enlace fey oie PT LIE hdeted
0 50 100 150 200 250 300 350

Chart 1 Based on table 1 and giving the computed diameters of the cells
and their nuclei according to sex—on body weight in grams. Wild Norway rat.
Males Females ----------

each sex and arranged according to body weight as shown in
table 1. From these data chart 1 was plotted.

The graphs in chart 1 show both the cell bodies and the nuclei
as growing rapidly up to a body weight of 15 grams, but after
that period they grow more gradually. The graphs for the
males and females run close together at all body weights, and
there is no indication of the difference according to sex shown in
chart 2 for the Albino (Ping, ’21).

316 “| SCHI PING

B. In relation to body length

Following the procedure just used, the condensed data have
been arranged to show the relations of the cell diameters accord-
ing to increasing body length, and are given in table 2. There
are thirteen groups for the males and eleven for the females.
The corresponding graphs are plotted in chart 2. According

TABLE 2
Giving according to body length the computed diameters of the largest cells and nuclei

in the superior cervical ganglion of the Norway rat. Sexes separated. Data
condensed

MALE FEMALE

Minti joey. Computed diameter Computed diameter oa Nuamibe

eee VETERE Nucleus Cell Cell Nucleus NENA oF ORES

mm. Me iu Me be mm.

1 53 9.9 17.2 16.5 9.6 50 1
2 71 12.5 21.9 22.5 12.5 71 3
1 84 1358 24.4 23.9 12.1 103 3
2 104 12.6 23.6 23.4 12.8 114 1
3 112 12.3 23.3 24.6 12.8 123 5
3 144 12.9 25.5 24.7 12.7 130 3
5 167 13.1 26.8 PAR Fi 12.9 146 3
5 189 13.3 26.4 27.3 13.2 184 12
6 209 13.5 29.8 28.3 12.8 202 6
3 214 14.8 32.6 28.9 13.5 213 2
5 225 13.5 32.6 31.5 1350 222 3
4 233 13.8 32.4
3 251 113} 7/ 31.1

to the graphs, there seem to be two periods in which the diameter
of the cell is showing a rapid growth; one-period at a body length
of about 80 mm. and the other at a body length of about 200 mm.
Without entering into the details, it will be sufficient to point
out that both periods are those in which the body weight is
increasing rapidly in relation to the body length, and it is prob-
ably the influence of the body weight which appears in the
eraphs. At the same time there is no sex difference to be seen
in the diameters of either the cells or their nuclei.

SYMPATHETIC CELLS: NORWAY RAT 317

C. In relation to observed age

The rats with known ages form a separate series for the pres-
ent discussion. The computed diameters of the cells and the
nuclei are given in table 3 according to age. Examination of
the table reveals that the cells and nuclei grow comparatively
fast during the first twenty-five days of life. In order to show
the contrast between the early growth and that which follows,
data selected from table 3 have been arranged in table 4.

10

SORE eeReeee
Se Body length—mm.
50 100 150 200 250

Chart 2 Based on table 2 and giving the computed diameters of the cells
and their nuclei according to sex—on body length in millimeters. Wild Norway
rat. Males Females ----------

Table 4 shows that at the end of twenty-five days the cells
and the nuclei have increased in diameter 1.31 and 1.14 times,
respectively, for the male, and 1.48 and 1.25 times for the female;
while the increase from 25 days to 134 days—a period which
is more than five times as long—is 1.19 for the cells and 1.21
for the nuclei of the male and 1.04 for the cells and 1.06 for the
nuclei of the female.

The graphs in chart 3 illustrate the fact that during the first
twenty-five days there occurs a sudden and rather irregular
increase, which is followed by a slow advance. No clear indi-
cation of a difference according to sex is to be seen.

318

CHI PING

TABLE 3

Giving according to age the computed diameters of the cells and nuclei in the superior

cervical ganglion of the Norway rat.

AGE

COMPUTED DIAMETERS

Sexes separated. From detailed record

8 ATIOS OF DIAMETER
NUCLEUS TO

Cell iNadleus D IAMETER OF CELL
days fa ML
(of 1 vedo 9.85 Leas
£ 1 16.51 9.60 Li ede72
fot 5 22.20 12.41 ils a)
om 5 22.40 12.30 1182
of 10 21.66 12.50 Wee ie ee:
2 10 22.21 12.60 1:1.75
or 15 24.40 13.82 Le eG
¢ 15 22.82 12.60 peat col
of 19 26.00 13.86 ae SS,
Q 19 25.21 13.00 1:1.94
& 25 22.61 11.25 Ug Aston
2 25 23.80 12.06 121286
a 28 26.00 13.40 Lea 94
of 31 21.21 11.25 11.89
of 31 21.02 10.55 1 e309
2 60 27.22 14.20 ih eal bee:
of 65 25.00 13.04 es 92,
Q 65 26. 24 13.49 1:31.95
of 80 23.25 13.40 1 tis
g 80 23.41 12.80 oz 1382
of 134 27.00 13.70 De oF
2 134 24.70 12.85 TES ibatey!

SYMPATHETIC CELLS: NORWAY RAT 319

TABLE 4
Increase in diameters of cells and nuclei at three different ages

DIAMETERS

SEX AGE
Cell Nucleus
days

J 1 17.19 9.85

Q 1 16.61 9.60

of 25 22.61 11.25

Q 25 23.80 12.06

of 134 27.00 13.70

o} 134 24.70 12.85
: } of 1B US 1:1.14
Ratios between 1 day and 25 days......... 9 11.43 1:1.35
: 5 ot Is dle tks) Slee
Ratios between 25 and 194 days ........-..49 1:1.04 1: 1.06

iyaaee nada acaat

Zum FS ae
SHIRE Zakk eeeee BGagaeaee Bae
SEER EERE eee EE EEE
| | EAB E Ramer tL

ca aati ace sacs
a Baieigtemieret tar EEE EH Age days
0 20 50 100 200 200 300 400

Chart 3 Based on table 3 and giving the computed diameters of the cells and
their nuclei according to sex—on age in days. Wild Norway rat. The values
for one female at 450 days are given in the chart. This case is not entered in
table 3. At 200 days, as indicated by a break in the graph, the time unit is
changed—one division being made equal to twenty-five days instead of ten days as
heretofore. Males ————- Females ----------

320 CHI PING

To return to table 3, the ratios between the cell and nucleus
tend to increase up to the age of twenty-five days; from that
time on the ratio in every case is almost 1:1.9. It should be
noticed, moreover, that there is during this later period no in-
crease in the ratios as the animal increases in age or in size.
This fact will be discussed later on.

TABLE 5
Giving according to percentage of water in the brain the computed diameters of the
cells and nuclei in the superior cervical ganglion of the Norway rat. Sexes sep-
arated. Data condensed

MALE FEMALE
‘ P eS
Mean | Number Daan aes Computed Computed : ae Number |: Mean
pia | Soe ee |, aiemeter | attri | on | oe
age le average Nucleus Cell Cell Nucleus meee ced tics
iu Lu iD M
100 1 80.4 13.4 26.0 Qa40 ii 80.4 2 68
54 2 79.5 Ze 24.2 DRY Fi 12.6 79.6 5 46
153 5 COG See 28.4 \ 26.6 13.0 (820 4 129
172 5 78.3 13533 28.2 Pai 574 S351 78.5 5 164
258 i 78.1 14.1 32.4 29.5 3355 18.2 5 203
194 2, Holl 13.0 28.9 30.2 1320 Was ay 205
253 3 774 14.1 31.4 26.6 PA} 77.4 -2 175
3381 i 76.8 leo 30.8 28.2 12.8 77.0 5 203

D. In relation to the percentage of water in the brain as an
indication of age «

Accepting the conclusion (Donaldson, ’10) that the percentage
of water in the central nervous system is more closely correlated
with age than with body weight and brain weight, it was thought
worth while to study the growth of these cells in relation to the
percentage of water in the brain, in order to supplement what
has been presented in the preceding paragraph, based on ani-
mals of known ages. |

Table 5 is, therefore, to a certain extent, a continuation of
table 3. For each sex there are eight entries based on the con-
densed data, those cases with known ages being excluded. ‘The
corresponding graphs appear in chart 4. According to Don-

SYMPATHETIC CELLS: NORWAY RAT a2]

aldson (711) the percentage of water in the central nervous system
of the Albino and Norway rat at like ages is nearly the same,
so the ages of the Norway rats whose percentages of water are
known may be obtained from table 74 of ‘The Rat’ (Donaldson,
15) and the growth of the cells as shown in chart 4 can be trans-
lated into age. Using this procedure, the curves in chart 4
represent the gradual growth from twenty-five days of age
to maturity.

ae ees
BEE Eero
aan RENE ns es

20 PRBEECECEEEEE EEE ei
a Fa Oca Fo a BD
REPOS eeeh sae
SE ete EH
se canuasaunaseeeneedaas
Poo Snes ees eee eee eee ee

is spauece

81 80 19 78 it 16

Chart 4 Based on table 5 and giving the computed diameters of the cells
and their nuclei according to sex—on percentage of water in the brain. Wild
Norway rat. Males Females ----------

The examination of table 5 enables us to see that in general
the female has a slightly higher percentage of water in the brain
than the male, due probably to the smaller absolute size of the
brain (Donaldson, ’16), but the cells of the male exceed in diam-
eter those of the female in seven out of eight cases.

MORPHOLOGY OF THE LARGE CELLS

The general morphological changes in the large cells from
birth to maturity are similar to those in the cells of the Albino.
The large cells of the first few days are much alike in the two
forms. The distribution of Nissl granules and the tendency to
accumulate at different regions in the cell in the later ages are
also alike in the two forms. There is, however, some difference

Say CHI PING

between the Albino and the Norway rat in regard to the degree
of aggregation of the Nissl bodies in the cells. Thus, in the
cells of the Norway, these are not so densely crowded either at
the periphery or around the nucleus as in the cells of the Albino.

Furthermore, the space left at one region through the crowd-
ing of Nissl bodies toward another is not so clear. There is
therefore always found a gradual thinning out of the granules
toward the periphery or the nucleus whenever the accumulation
of them takes place in a reverse direction.

When the Norway rat reaches twenty days of age, we find .
in the cells of the superior cervical ganglion a tendency for the
Nissl bodies to aggregate at the periphery, though the region
around the nucleus is by no means devoid of them. Likewise,
when the Norway rat is about sixty or more days old, the Nissl
bodies tend to accumulate around the nucleus, while the periph-
ery. still has some of them thinly scattered. Unless carefully
examined, therefore, the distribution of the Nissl bodies in the
cells at twenty to sixty days and in those sixty days and later
would appear the same, i.e., as if they were evenly distributed.

Like the Albino, the distribution of the granules in the cells
of the older Norways is fairly even. On the whole, then, there
is no marked distinction between the two forms, so far as the
general morphology of these cells is concerned.

According to Gaskell (’18), both the motor and the inhibitory
cells are found in the sympathetic ganglion, and Cajal (11),
using the Golgi method, shows in the superior cervical ganglion
of a mouse several days old cells which appear to represent three
different types (fig. 550, B, C, and D). That the four types
of cells (Ping, ’21) found in the superior cervical ganglion of
both the Albino and Norway rat are correlated with the several
functions of the ganglion is‘by no means determined, but, gen-
erally speaking, we should expect that the morphological dis-
tinctions would have a functional significance.

SYMPATHETIC CELLS: NORWAY RAT BAS

Binuclear cells

The number of the binuclear cells in one ganglion in each
case is recorded in table 6. Generally speaking, we may say
that this type of cell is found through the whole span of life.

TABLE 6

Norway rat: Data for eighty individuals arranged in eighteen groups according
to the decreasing value of the percentage of water in the brain. This is indicative
of increasing age. Under the ‘number of cells’ the number of-any class of cells in
one ganglion of each individual in which such cells occur is given. Thus in the
first group one individuals showed three binuclear cells, but no pigmented or vacuo-
lated cells were found. In the fifteenth group binuclear cells were found in four
out of the five individuals, pigmented cells in four, and vacuolated cells in one

AVERAGE

pale OF; SOREENE: AVBRAGE NUMBER OF CELLS IN INDIVIDUAL CASES—ZEROS OMITTED
WATER IN WEIGHT a ae eo ee Sa he os hee Sash”
Me | Bs (otal (THs Binuclear Pigmented Yacue:
grams
iL Fe 33 88.33 11 3 — _
2 il 3 87.98 iil 1 _ —
2 1 3 84.42 21 3,3 =_ —
~ 3 i 81.45 29 eal — —
2 2 4 80.30 67 1 _ _
1) 3] 4| 79.83 40 indies) = =
3 1 4 79.43 46 Mpc Loa | — —-
3 2 5 78.87 119 11 Sy, toh 8 — -
2 3 5 78.69 120 Up Oy ap oh 2 —
1 4 5 78.47 183 5, 9, 2, 4 I =
3 2 5 78.41 123 By! _ _
4 i 5 78.29 201 2, 2, 4, 14 2:13 —
See Bula tSP) W314 220 5, 2, 10, 1 digs 3
3 2 5 78.01 236 Uo & ley! 3
4 1 5 77.60, 240 Pos Pas MO) 9, 3, 15, 4 1
1 4 5 Ula 189 5, 2, 4, 2 85, 3, 2, 6 —
2/3] 51 77.00 279 2,1,1,2,2 1, 2,5
5 1 6 76.66 307 9, 3, 3,4 Py Spl evils al 1

There are, however, exceptions, since, as the table shows, this
type of cell is present in all the individuals couiposinie a group
in two instances only.

As can be seen, binuclear cells appear eantty in very small
number and frequently only one is found in the entire ganglion,

324 CHI PING

and the maximum number of these cells is never above ten.
There seems to be a tendency for the binuclear cells to increase
in number during the middle age of the animal. ‘The diameters
of some of these cells and those of their two nuclei, as well as the
nucleus-plasma ratios derived from these diameters, are recorded
in table 7.

According to table 7, the nucleus-plasma relation is 1:4.6
in a very young rat and 1:4.1 in an old rat, as indicated by the
body weight, and throughout the series the values are fairly
constant, though with a slight tendency to diminish. This
agrees with what has been found in the Albino and shows that

TABLE 7
Diameters of some of the binuclear cells and of the two nuclei in each of them. Data
arranged according to body weight. The nucleus-plasma ratios are shown in the
last column. Norway rat

DIAMETER OF RATIO OF VOLUME OF
BODY WEIGHT : CYTOPLASM TO VOLUME OF
Cell Two nuclei ENAGIINITE EIDE

grams iM M

13.6 25.0 11.6 + 10.7 1 :4.6

33.0 29.9 13.4 + 13.4 1 :4.6

115.0 26.3 13.4+ 8.9 1:4.8
169.2 28). 1 13.4 + 12.9 12329
243.2 204 12.0 + 13.4 I 401
230.2 25-1 13.4 + 13.4 WB ioe

in the binuclear cells there is an increase in the nuclear mass
which is not accompanied by the same enlargement of the cyto-
plasm, as is found in mononuclear cells. In their nucleus-plasma
relation, therefore, these binuclear cells are like very young cells,
and in the older animals at least they certainly do not represent
two normal mononuclear cells pressed together.

Pigmented cells

It is somewhat surprising to find that the pigmented cells
in the superior cervical ganglion of the Norway are not very
numerous, as table 6:shows. In no case were they found in all
of the individuals of a group. They do not appear in the young

SYMPATHETIC CELLS: NORWAY RAT 325

animal, the first being found in my series at a body weight of
120 grams. In this case the percentage of water in the brain
was 78.69, which corresponds to the age of eighty-eight days—
this happens to be exactly the age at which they first appeared
in the Albino. The number of the pigmented cells tends to
increase as the animal grows older. There are two cases among
the older rats which show large numbers of pigmented cells,
but most ganglia have only a few, even at the later ages. There
is less increase in the number of these cells in the Norway, as
contrasted with the Albino, than we should have expected. The
pigment granules are black or greenish black in color. It was
a matter of some surprise to find that in the gray pigmented
rat the cells contained hardly more pigment than appeared in
the Albino.

Vacuolization of the cells

Incidentally vacuoles have been noted in a very few of the
cells in the superior cervical ganglion of the Norway, although
none were observed in the Albino. Out of eighty cases only
eight showed vacuoles in the cells and in an entire ganglion
only one to three vacuolated cells have been found (table 6).
Most of these cells have but one vacuole, which is oval in shape,
and which may lhe close either to the nucleus or to the periph-
ery, but in one cell two vacuoles were found. ‘The size of the
vacuole varies; generally it is smaller than the nucleus, rarely
larger. It resembles the nucleus in outline, but, owing to the
absence of any internal structure, can be recognized without
difficulty. All the vacuolated cells were found in older rats.

Increase in the number of the large cells

In counting the large cells found during the first twenty-five
days, I followed the procedure previously used for the Albino.
The cells of the Norway are small as compared with those of
the Albino, especially at birth, so I have extended the limiting
values, 19 to 25 », which were used for the diameters in counting
the large cells of the Albino, to 16 to 25 » for the Norway.

326 CHI PING

Table 8 gives the number of these large cells recorded in the
ganglion for each sex during the first twenty-five days.

There are a few advanced cells and a few comparatively large
cells at birth, or just after, but the number is strikingly small:
By the end of the fifth day there is a great increase in number—
about twenty times that of the preceding stage. Then the

TABLE 8
Increase in number of large and advanced cells, 16 to 25 w in diameter, during the
first twenty-five days of age. The ratios for the increase in the total number for
both sexes between one day and nineteen days stand at the foot of the last column.
Superior cervical sympathetic ganglion. Norway rat

SEX AGE BODY WEIGHT NUMBER OF LARGE CELIS
days grams

Si 1 5.9 22
g 1 5.6 * 24
oh 5 12.7 416
g 5 13.4 515
rot 10 13.6 417
a 10 13.5 530
of 15 1726 833
2 15 13.6 827
oe 19 31.1 3, 066
Q 19 31.1 3, 084
Q m4) 28.5 3, 074

of 1 :'139.3

Ratios between and 19" days. ot2. 2 os. .c so Se ees 3 1: 129.0

increase is slow and slight until the age of fifteen days. At this
time the cells again show a considerable increase in their number,
and this is still more marked at nineteen and twenty-five days.
Thus the greatest increase in number of the large cells is between
one and five days, and again between fifteen and nineteen as
in the case of the Albino (Ping, ’21, table 6).

SYMPATHETIC CELLS: NORWAY RAT BA

' The ratio of increase is a trifle higher for the male Norway,
as was found for the male Albino; indeed, the relations of the
ratios according to sex are strikingly similar in the two strains.

As will be seen by comparing the ratios (1 to 19 days) for the
Norway with those (1 to 20 days) for the Albino, the rate of
increase is apparently ten times as great in the Norway as in
the Albino. This will be discussed later.

The nucleus-plasma relation

In determining the increase in volume of the cytoplasm in
relation to that of the nucleus, the computed diameters of the

TABLE 9
Giving the average diameters of the cells and nuclet according to body weight. The
nucleus-plasma ratios based on these diameters are given in the last column.
Data condensed. Superior cervical sympathetic ganglion. Norway rat
|

MEAN ; DIAMETERS

NUMBER OF BODY WEIGHT Kany NUCLEUS-PLASMA
CASES RANGE Gin ait ees. RATIOS
grams Me [a
16 6- 38 22.2 22.3 12.1 3553
16 41-100 65.9 25.0 12.8 12624
8 104-150 121.4 26.8 13.0 IOI Le.
16 152-195 175.4 27.9 13.1 IS Bile
16 206-250 226.6 30.1 13.5 Teel O20
5 - 259-290 270.1 32.7 13.7 1:12.4
8 311-402 346.1 32.2 13.8 pestle 6

cell and of the nucleus have been condensed according to body
weight and arranged in table 9. By subtracting the volume
of the nucleus from that of the cell, the volume of the cytoplasm
is obtained, and the ratios between the latter and the volume
of the nucleus are given in the last column of table 9. This
table shows that the increase of the cytoplasm is progressive,
except in the last group.

In general it may be said that for each 50 grams of increase
in body weight, the increase in the ratio is one unit. For a
body weight of 22.2 grams (about twenty-five days of age)
the ratio is about one-half that for the oldest group with a body

328 CHI PING

weight of 346 grams. As compared with ratios for standard
Albinos of like body weights (table 2, Ping ’21), the ratios
for the Norway are clearly low.

DISCUSSION

In the foregoing paragraphs the growth of the cells in the
Norway rat has been treated in relation to the body weight
and length and in relation to age, either observed or inferred
from the percentage of water in the brain of the animal. In
each case the results show that the growth is comparatively
rapid at first and then becomes gradual. Moreover, as was to
be expected, the growth of the sympathetic nerve cells of the
Norway resembles in a general way that of the standard Albino.
There are, however, differences between these forms, worthy
of note, and to make the comparison as complete as possible,
the data for the inbreds will also be taken into consideration.

Although the form of these data and the numbers of cases
are not the same in the three series, they are yet sufficiently
similar to make several comparisons worth while.

Before attempting this, a word about the general relations
of the three strains here examined is in place. Both the Albino
strains contained animals which had been in captivity for many
generations, and were also domesticated in the sense that they
had lost the fear of man and were easy to handle. The Norway
strain, on the other hand, had two groups in it: (1) the rats
caught wild and of unknown age—represented in this series by
animals 37 grams or more in body weight-—and (2) a group
which were the F, or F; descendants of Norway parents caught
wild. Although this second group was composed of captive
individuals, they were by no means domesticated and for the
most part were still timid and excitable.

The differences between these strains may be tabulated as
follows:

Albinos Norways
Not pigmented Pigmented
Captive Wild or captive
Domesticated Not domesticated
(A) ‘Standard’ (not inbred) Not inbred

(B) ‘Inbred’

SYMPATHETIC CELLS: NORWAY RAT 329

By the aid of such a tabulation, there seems to be a chance
to consider the possible influence of albinism, captivity, domes-
tication, and inbreeding on the cells under discussion.

The characters which may be compared in the several strains
are:

A. The morphology of the largest cells.

B. Special cell forms (binuclear, pigmented, or vacuolated).

C. The increase in the diameter of the cells from birth to
maturity.

D. The absolute size of the cells at different ages.

E. The nucleus-plasma ratios.

F. The rate of the formation of large from small cells.

In making the comparison, the three strains will be briefly
designated as ‘standards,’ ‘inbreds,’ and ‘Norway,’ and for
convenience the values for the ‘standards’—which have been
most completely studied—will be those to which the values for
the other strains are referred.

A. The morphology of the largest cells

Figures 1 to 5 (Ping, ’21) show the morphology of the largest
cells in the standards. In the other strains these cells have in
general a similar appearance. However, it was: noted in the
Norways that the Nissl granules were less segregated than in
the standards. Whether this difference is correlated with albin-
ism or domestication cannot be determined at present, because
the Norways have not yet been domesticated.

B. Special cell forms

1. Binuclear cells. In the standards, binuclear cells were
found in every ganglion examined (table 5, Ping, ’21) and the
average number was 4.2 per ganglion. In the Norway they
were found in only 54 out of 80 ganglia studied (table 6). The
average number for the entire series of 80 ganglia was 2.2 per
ganglion, and for the 54 ganglia in which they occurred, 3.3. In
the Norways therefore, binuclear cells are less abundant. Again
this difference cannot be correlated with either albinism or
domestication.

330

Increase from birth to maturity.

CHI PING

TABLE 10

The data for the ‘standards’ are jrom table 1

(Ping, ’21) and for the Norways from table 1 of the present paper. In each in-
stance the values are the means for the male and female records at the corresponding

body weights

| DIAMETERS
BODY WEIGHT =
Cell Nucleus
| im a
Standards 5.9 19.7 10.8
178.0 30.5 1350
TU ARIOS 202 or siete se See oe Ca aw a Saree eae ras prneree 1.54 1225
Percentage dnGrCases.s ciec.<c 5 sclaicsscieyaie's = sjare afer 55 5 25
Norways 5.8 16.9 ih
182.0 27.8 1135,
Pat insee pedashik cars meshed Sci Wee ele De ets | 1.64 1.36
PeTrcentace MMCPeAse 55.5 pas «0.0 sige o sicctte eae ale 36

ganglion (Donaldson and Nagasaka, ’18)

TABLE 11
Showing, in the standard Albino and in the Norway rat at about 19 grams and 178
grams of body weight, the diameters in u of the largest cells and nuclei in the superior
cervical sympathetic ganglion, compared with those of the cells from the ventral
horn of the spinal cord (seventh cervical segment) and from the corresponding

LOCALITY

Superior cervical sympa-
thetic ganglion..........

Motor cells, spinal cord....

Spinal ganglion cells.......

Superior cervical sympa-
thetic ganglion. .........

STRAIN

Standard

Standard

Standard

Norways

aT 19 AT 178

GRAMS | GRAMS

Cells 24.2 | 30:5
Nuelei-<|\ 12.1) |. 1325
Cells 23.9 29.1
Nuclei 12.9.1) 15.1
Cells 21.6 | 34.2
Nuclei 10.6 | 16.4
AT 23 GRAMS meee
Cells | 23.0 | 27.8
Nuclei | 12.4 | 13.2

GAIN

Percent-
Absolute See

6.3 26
1.4 12
one 99
2.2 il
12.6 58
5.8 55
4.8 21
0.8 6

SYMPATHETIC CELLS: NORWAY RAT Soil

2. Pigmented cells. In the standards the pigmented cells do
not occur before puberty (table 5, Ping, ’21); the same is true
for the Norways (table 6). In the standards there were found
about 3.4 pigmented cells in each of the eleven ganglia in
which such cells occurred, while in the Norways there were
3.8 pigmented cells per ganglion, after these cells were first noted
at 120 grams of body weight.

However, in one ganglion of the Norways eighty-five, and in
another thirty-seven pigmented cells were found, and it is these
two cases which make the average for the Norways slightly
above that for the standards. This is a surprising result, for,
according to conmon teaching, we should have expected a
much greater number of pigmented cells in the Norways than
in the standards. Until thoroughly domesticated Norways
are available, these relations cannot be interpreted.

3. Vacuolated cells. These vacuolated cells were not found
in the standards, but do occur in the Norways after maturity.
Their number is small (table 6). As the data stand, the vacuo-
lated cells are characteristic for the mature wild Norways.

C. The increase in the diameters of the cells from birth to maturity

As between the standards and Norways, it is possible to make
this comparison between birth and maturity; table 10 gives
what has been found.

As the ratios and percentages show, the cells and nuclei have
increased a little more in diameter in the Norway than in the
standards, but this difference appears to depend mainly on
the small size of these cells at one day of age in the Norway.

If we choose the values at about 19 grams of body weight for
the initial data, it is possible to compare the changes in the
diameters of these cells, not only in the standards and Norways,
but also with those of the cells in the spinal ganglia and ventral
horn of the spinal cord, as observed in the standard rats (Don-
aldson and Nagasaka, 718). The relations appear in table 11.

This table 11 may be used for two purposes. First, it shows
that within the limits of body weight chosen the percentage

BOL CHI PING

gains in these cells are of the same order in the Norways as in
the standards—21 per cent and 26 per cent, respectively, for
the cell body. The two strains are therefore similar in this
character. Second, that this order of enlargement is, on the
one hand, similar to that found for the large motor cells in the
spinal cord, and, on the other, very different from that found for
the corresponding spinal ganglion cells. This later relation
indicates that the sympathetic ganglion cells, which are efferent
in function, behave like motor cord cells during their later growth.

TABLE 12
Comparing, according to body weight, the average diameters of the cells and nuclei
in the swperior cervical sympathetic ganglion of the Norway with that of the stand-
ards. The data for the Norway rat are condensed from table 2, and for the Albino
from table 2 of the former study (Ping, ’21). Sexes combined

ALBINO RAT : NORWAY RAT
Body weight Nucleus Cell Cell Nucleus Body weight
grams ye a Le ia grams
6 10.8 19.7 16.8 9.7 6
15 113351) 2557 22.6 127 14
31 12RD 25.0 23.9 12.4 BY
53 118357 Die, 24.3 ea 51
73 ils} 7 29.2 25.8 13.0 74
106 13.6 29.8 25.7 13.0 103
151 11832 28.9 Zie2 13.3 157
178 13H0 30.5 Dee? ifs}, i 189
Average .. 12.8 27.0 24.2 1245
Percentage difference in cell diameter = 11 per cent
Percentage difference in nucleus diameter = 3 per cent

D. Absolute size of cells at different ages

E. Nucleus-plasma_ ratios

In the first instance we shall take up cell size alone and limit
the comparison to that of the Norways to the standards.

In table 12 the mean diameters of the cells and nuclei of the
standards and Norways are arranged according to their respective
body weights, the former data being from table 2 of my previous
paper, the latter from the foregoing table 2 of the present paper.

SYMPATHETIC CELLS: NORWAY RAT BB

A study of table 12 gives a clear idea of the quantitative dif-
ference in growth between these two forms, and the same data
are represented by graphs in chart 5. From the chart it is
evident that the diameters of the cell body in the Biot ays are
consistently below those in the standards.

The data for these two strains have been compared also on body
length, on age, and on the percentage of water in the brain and
by all of these methods of comparison show relations essentially
like those just presented. Thus, no matter what the basis of

se es cok i ean ae redo aT es a
jameters of Cells and Nuclei in micra Pee eel tale oll eelali|
ie i ant aT Pe das

fest teal =n
Bas SSS==s5-s5 0”
Snecma aaecnaaa eases

SeesGhS ees ERR weas
ia || BEE Eee
PEEEEHTEE fr woat-ars|
0 25 50 15 -Gae 125 150 175° jek 200

Chart 5. Based on table 12 giving the diameters of the cells and their nuclei
on body weight in the Norway rat compared with the standard albino.
Norway 4—A Albino O—O

comparison, these cells in the Norways are smaller than those
in the standards.

In order to compare with the standards the inbreds as well
as the Norways, a series of data, selected from the previous re-
cords, have been assembled in table 13.

An examination of table 13 brings to light several interesting
relations. While in five out of the six instances the diameters
of the nuclei are similar, there is nevertheless a great difference
in the diameters of the cells according to strain. The differences
therefore appear mainly in the cytoplasm. As compared with
the standards, the inbred cells are small, while the cells of the
Norways, though also small, differ much less.

334 CHI PING

The smaller size of the cells in the inbreds and Norways is
not related to pigmentation, for the inbreds with the smaller
cells are not pigmented. It is not due to domestication, for
both the standards and inbreds are domesticated.

It would appear therefore to be a characteristic of this inbred
strain, but without further evidence it cannot be said to be due
to inbreeding.

Since the differences according to strain are mainly in the cell,
the nucleus-plasma ratios show the same relations as do the
cell diameters, and the most striking feature is the small value.

TABLE 13

Giving the diameters and nucleus-plasma ratios of the standard, inbred and Norway
rats according to body weight, based on previous tables. In each instance the
values given are for the two sexes combined

DIAMETERS
STRAIN BODY WEIGHT piesa:
Cells Nucleus
grams Le M
Stancarc seer ee 145 28.9 tee Ts (925
Imbnedie <.nkensoe 144 24.5 13.2 Pp 534
INOTwiiyeceee cece 146 28.0 1335 1b Sas
Standard. f\\.0 <0. 200 34.1 (14.5 esdiead
lhilomteclSkeaoe wooo 206 25.4 3.33 ts &8
INOGWaynecnteie acre 203 28.0 Teel iTepctn cones

for the ratios in the inbreds. The data for the heavier body
weights show that in this group these cells in the inbreds have
only half the volume of those in the standards.

Perhaps the point of most general interest which is thus brought
out is the plasticity or variability in the size of this group of
cells according to strain.

F. The rate of the formation of large from small cells

As table 6 (Ping, ’21) shows, in the standards the number of
large and advanced cells increases about thirteen-fold from one
to twenty days. On the other hand, table 8 of this paper indi-

SYMPATHETIC CELLS: NORWAY RAT 335

cates in the Norway an increase during this same interval, which
is about 135-fold, or nearly ten times as large. One circumstance
which contributes to this result is the lower standard which was
taken for the large cells in the Norway, in order to get a reason-
able number of such cells at birth. The use of this lower standard
gives of course a higher number at maturity in the Norway.
This is, however, a trifling matter, and the more important dif-
ference between the two strains lies in the relatively smallnumber
of large cells in the Norway at one day of age.

This is an expression of retardation in the early growth of
the Norway in captivity—a peculiarity which has been noted
by several observers and which has been demonstrated by Dr.
Helen D. King for the general body growth. .

Here again it is not possible to offer a precise explanation,
for this relative retardation may be a character of the Norway
in the wild state and hence difficult to study, or it may be a
response to captivity, as these observations were made on young
born in captivity from Norways not yet domesticated.

It is possible, however, to conclude that the retardation in
the early development of these cells is found in the wild as con-
.trasted with the domesticated standard strain.

To determine the general significance of these results, a brief
review of some earlier observations along similar lines may be
of value.

In the presentation of these the data for the wild Norway
will be taken as the standard, as this is the strain from which
the Albino has been derived.

As compared with wild Norways of like body weights, the
standard Albinos have lighter brains and lighter spinal cords
(16 and 12 per cent, respectively; Donaldson and Hatai, ’11).
Further, two portions of the brain, the olfactory bulbs and the
paraflocculi, are, relatively, still lighter in the Albino.

As between the sexes within the same strain, it was found that
while the weight of the spinal cord (on body weight) was similar
in the two sexes in the Norways, yet in the standard Albinos
the weight of the cord in the female was relatively heavier than
in the male (Donaldson and Hatai, ’11).

336 CHI PING

On comparing the diameters of the cells in the cerebral cortex
of the rat, Sugita (18) found that those in the lamina pyramidalis
and in the lamina ganglionaris of the Albinos were, respectively,
4 and 7 per cent less in diameter than the corresponding cells
in the Norway.

In the foregoing instances it appears that there is a reduction
in the relative size of the parts of the nervous system, as well
as in some of the cell elements, in the Albino as compared with
Norway.

Incidental observations on captive Norways suggest that
this reduction is the result of captivity or domestication. There
remains, however, the difference in the relative weights of the
spinal cord which occurs in the Albino, but is absent in the
Norway, and in this instance it is possible that albinism plays
a part. From these earlier observations we should have ex-
pected to find the cells in the superior cervical sympathetic
ganglion smaller in the Albinos.

Contrary to expectation, these cells in the standards are
larger than in the Norway, but also exhibit a sex difference,
being larger in the females. Nevertheless, in the case of the
inbreds, these cells are much smaller than in the Norways, and.
show only a slight difference according to sex. At the moment,
therefore, the size of these cells in the standards does not agree
with the expectation based on the results obtained from other
parts of the nervous system, but any simple interpretation is
at once precluded by the very small size which they have in the
inbreds.

SUMMARY

1. The growth of the largest cells in the superior cervical
sympathetic ganglion of the Norway has two phases: (1) a com-
paratively rapid growth from. birth to twenty-five days, (2)
followed by a slow and gradual growth continuing to the end
of the record.

2. The growth of these cells in relation to body weight, body
length, age, and the percentage of water in the brain does not
show differences due to sex after the animal has passed puberty.

SYMPATHETIC CELLS: NORWAY RAT 337

3. The morphological changes in the cells are in general similar
to those found in the Albino, but the stainable substance is
somewhat less aggregated. The binuclear cells and the cells
with pigment are present, as in the Albino, but the latter are
only slightly more numerous in the Norway. Besides, there
are in the Norway a few vacuolated cells.

4. The mode of growth of the cells, as in the Albino, shows
similarity to that of the motor cells of the spinal cord.

5. The nucleus-plasma ratio is 1 to 5 at birth and 1 to 12
after maturity.

6. The cells at birth are about 16 per cent less in diameter
than those of the Albino. The growth acquired after the
weaning period is about 10 per cent less than that of the
Albino, such difference being probably due to domestication.

7. Comparing with the standard Albino the data for the wild
Norways, it appears that in the Norways the diameters of the
nuclei are the same, but those of the cell bodies are slightly
less. As a consequence, the nucleus-plasma ratios are smaller
in the Norway. Moreover, there is no clear sex difference in
the size of these cells.

8. Comparing the inbreds with the standard albinos in re-
spect of these cells, it appears that in the inbreds the sex dif-
ference in size is less marked. The absolute diameters of the
nuclei are similar to those for the standards and Norways, but
the diameters of the cells are much less, and in consequence
the nucleus-plasma ratios are only about half those for the
standards.

9. The interpretation of these differences awaits the data
to be obtained from Norways after long-continued domestica-
tion and inbreeding.

338 CHI PING

LITERATURE CITED

Apotant, H. 1896 Uber die sympathischen Ganglienzellen der Nager. Arch.
f. mikr. Anat., Bd. 47, S. 461-471.

Donaupson, H. H. 1910 On the percentage of water in the brain and in the
spinal cord of the albino rat. Jour. Neur. and Psychol., vol. 20, pp.
119-144.

Donaupson, H. H., Aanp Hatar, 8. 1911 A comparison of the Norway rat with
the albino rat in respect to body length, brain weight, spinal cord
weight and the percentage of water in both the brain and the spinal
cord. Jour. Comp. Neur., vol. 21, pp. 417-458.

1915 The Rat. Reference tables and data for the albino rat (Mus
norvegicus albinus) and the Norway rat (Mus norvegicus). Memoirs
of The Wistar Institute of Anatomy and Biology, no. 6.

1916 A revision of the percentage of water in the brain and in the
spinal cord of the albino rat. Jour. Comp. Neur., vol. 27, pp. 77-115.

Donaupson, H. H., anp Nacasaka, G. 1918 On the increase in the diameters
of nerve cell bodies and of the fibers arising from them—during the
later phases of growth (albino rat). Jour. Comp. Neur., vol. 29,
pp. 529-552.

GasKELL, W. H. 1920 The involuntary nervous system. Longmans, Green
& Co., New York.

Hatar, S. 1907 On the zoological position of the albino rat. Biol. Bull.,
vol. 12, pp. 266-273.

1914 On the weight of some of the ductless glands of the Norway
and of the albino rat according to sex and variety. Anat. Rec., vol.
8, pp. 511-523.

Pine, Cut 1921 On the growth of the largest nerve cells in the superior cervical
sympathetic ganglion of the albino rat—from birth to maturity.
Jour. Comp. Neur., vol. 33, no. 3.

Ramon ¥y CasaAu, 8. 1911 Histologie du systéme nerveux, T. 2, pp. 891-942.

Sueira, N. 1918 Comparative studies on the growth of the cerebral cortex.
VI. Part I. On the increase in size and on the developmental changes
of some nerve cells in the cerebral cortex of the albino rat during the
growth of the brain. Part II. On the increase in size of some nerve
cells in the cerebral cortex of the Norway rat (Mus norvegicus) com-
pared with the corresponding changes in the albino rat. Jour. Comp.
Neur., vol. 29, pp. 119-162.

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Resumen por el autor, Howard Ayers.
Cefalogénesis de los Vertebrados.

V. Origen del aparato mandibular y del complejo del trigémino
en Amphioxus, Ammocoetes, Bdellostoma y Callorhynchus.

Las mandibulas de los Vertebrados presentan una larga his-
toria de transformaciones y adaptacion. Han comenzado con
‘el aparato mandibular del Amphioxus, que contiene solamente
el elemento mandibular de las formas superiores mas otras
estructuras que desaparecen en los peces cartilaginosos, en los
cuales aparecer bajo forma de cartilagos labiales. El ejemplo
mejor conservado de mandibula del tipo del Amphioxus entre
los gnatostémos se encuentra en Callorhynchus. Su mayor
desarrollo se aleanza en Bdellostoma. El mecanismo hioideo se
desarrolla en forma de una estructura de soporte a lo largo del
mecanismo mandibular, mucho antes de aparecer los elementos
maxilares, y experimenta una regresién al mismo tiempo que
los llamados cartilagos labiales y en este caso, de nuevo, su
seccion anterior se conserva mejor entre los gnatostémos en
Callorhynchus.

Mientras que la historia del elemento mandibular de la mandi-
bula de los gnatostémos es completa en general, el curso de la
evolucién del esqueleto del elemento maxilar necesita todavia
trazarse. El aparato mandibular ha precedido a las barras
branquiales, y por consiguiente, no esta genéticamente relac-
ionado con ellas. El nervio trigémino se origina en Amphioxus
como un grupo de cinco nervios segmentarios que inervan el
aparato mandibular y ha sufrido una condensacidn de sus partes
hasta que en los gnatostémos aparece como un solo tronco
nervioso complejo... En Bdellostoma los cinco nervios ances-
trales que le componen pueden todavia distinguirse.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED BY
THE BIBLIOGRAPHIC SERVICE, SEPTEMBER 28

VERTEBRATE CEPHALOGENESIS

V. ORIGIN OF JAW APPARATUS AND TRIGEMINUS COMPLEX—
AMPHIOXUS, AMMOCOETES, BDELLOSTOMA, CALLORHYNCHUS

HOWARD AYERS

THIRTY-SIX FIGURES

The current theory of the origin of the vertebrate jaws assumes
their derivation and descent from a pair of gills, with all the
structural implications involved in that assumption. The food
canal antedated gills, mere appendages of the wall of the food
canal. The anterior and posterior openings of the canal un-
doubtedly antedated not respiration, but the complicated bran-
chial respiration of vertebrates. Branchial respiration in these
forms is carried on by means of the richly vascular walls of holes
through the gut and body walls.

The phylogeny of these structures is insufficiently indicated
by their ontogeny in the lower vertebrates, and comparative
anatomy has not as yet revealed the story of their descent.
In any event, the armature of the anterior opening of the food
canal antedated the highly specialized gills of vertebrates as
we yet know them, and all the evidence we have on this subject
indicates that the jaws are in no way genetically related to
branchial structures of any kind.

»The testimony of Amphioxus, Ammocoetes, Bdellostoma,
and Callorhynchus, bearing on this subject, is here presented
for the purpose of solving a few of the host of questions which
are involved in the phylogeny of the vertebrate jaws. Two
conclusions that the testimony fully sustains are: 1) That the
jaw apparatus is in no way related to gills, but is genetically
related in the forms mentioned in the order named, and since
Callorhynchus is an accepted Gnathostome, it follows that the
Amphioxine jaw apparatus is the earliest known condition of

339

340 HOWARD AYERS

the gnathostome jaw. 2) That the jaw apparatus is the end-
organ of the trigeminus. These two conclusions are presented
before the evidence, in order that the reader may better weigh
the evidence given.

In the past the study of the jaw began with the examination
of the human jaw and worked down through the long series of
vertebrate forms, seeking evidence to homologize and explain
the origin and transformation of these parts in the vertebrate
stock. Embryological evidence was abundantly used to check
the results and to clarify the relationships. A theory of the
origin of the jaw was thus worked out, and it became generally
accepted among morphologists that the jaw was derived from
one of the anterior gills, presumably the first pair of the ancestral
vertebrate. However, indications of presumptive premandib- ~
ular structures were found which cast doubt on the accuracy
of this conclusion as regards the particular pair of gills involved.

The Myxinoids could not be brought into this arrangement
because they were held to have no jaws. They were set aside
in a special group—Cyclostomes—and contrasted with the
Gnathostomes. Amphioxus was considered to have nothing
even suggesting Jaws.

The jaw apparatus plays such an important réle in cephali-
zation of the vertebrate body that I have for quite some time
given special attention to the subject, particularly to what
Amphioxus, Ammocoetes, and Bdellostoma have to tell, and
I think they give a clear, straight story of how the jaw came to be.

The material used in this study consisted: 1) of adult and young
stages of Amphioxus from the Naples Station treated with dif-
ferent reagents. 2) Ammocoetes and Petromyzon from Cayuga
Lake, consisting of a series of stages covering the whole Ammo-
coetes period, stages illustrative of the metamorphosis and heads
of the adult Petromyzon. It is a pleasure to acknowledge my
indebtedness to Prof. Simon H. Gage, of Cornell University,
for this adequate material, which was of great value in my in-
vestigation. I am particularly grateful to Professor Gage for
his generosity in supplying me with Ammocoetes undergoing
metamorphic changes. 3) Bdellostoma from the Bay of Mon-

ORIGIN OF JAW APPARATUS 341

terey, partly collected by myself and partly by Miss Julia Worth-
ington, who generously placed her extensive collections including
adults, larvae, and embryos at my disposal. 4) Bdellostomas
from Cape of Good Hope, Myxine from Norway and Torres
Straits, Hexanchus from Bay of Monterey, various Elasmo-
branchs from Woods Hole and Trieste, Callorhynchus, Chimaera,
several species of Urodeles both larval and adult, and several
species of Anura both larval and adult.

A discussion of the literature is reserved for subsequent
publication.

AMPHIOXUS

Under the term jaw apparatus are included all the structures
constituting the cirri or tentacular apparatus of Amphioxus
as recorded in the literature of the lancelet—skeleton, muscles,
nerves, etc.

The jaw bars form the internal skeleton of the armature of
the buccal aperture of the oral hood. They are two elastic
chondral bars, one in the left lip, the other in the right lip of
the mouth. Each bar is composed of from seventeen to twenty
segments progressively smaller from the base behind to the
terminal segment in front. Each segment is prolonged into
a tapering filament or tentacle which projects from the distal
end of each segment.

The two jaw bars are bilaterally symmetrical and, seen from
above, curve outward from the median line in a gentle curve,
until about midway of their course, when they curve inward
so that the tips lie side by side in the anterior pocket of the
buccal cavity (fig. 1). Seen from the side, the bars curve dor-
sad from the base and then ventrad, then dorsad again to form
an open S-shaped figure. These curves have their part in the
functioning of the jaw apparatus in opening and closing the
‘mouth. Under varying pull of the muscles, the bars change
their shape.

Thus the bases of the bars lie in the ventral wall of the body
and the tips lie close under the notochord. The buccal aperture,
thus looks forward and downward; it is placed at an angle to

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 4

342

A, anastomosis
B, buccal cavity

b, attachment of jaw bar to hyoid

Br, brain

C, cornual cartilage

D, skin

d, dorsal spinal nerve

E, endostyle

Hi, eye

G, gills

H, hyoid

H’, hyoidean procartilage
Hs, hyoid arch

Hy, hypophysial canal

J, jaw bar

Jd, dentigerous jaw cartilage
Jl, jaw ligament

Jo, median ventral jaw bar
L, left side

l, left median tentacle

li, ligament

M, myotome

mc, circular velar muscle
mn, nasalis muscle

Me, endostyle muscle
Md, mandibular nerve
Mj, jaw muscle

Mn, ventral transverse muscle
mr, radial velar muscle
mv, velar muscle

Mzx, maxillary nerve

N, nasalis nerve

n, nerve

Na, nose

Ne, notochord

NI, nasal capsule

No, nasal canal

HOWARD AYERS

ABBREVIATIONS

Nov, velar nerve

O, oral hood

ol, ophthalmic lateralis nerve

on, ophthalmic nerve

opth, ophthalmic nerve

os, oral sphincter

P, palatine

Pc, palato-coronarius muscle

Pl, plexus of jaw apparatus

Q, quadrate

R, right side

r, right median tentacle

Rm, rectus muscle

S, subnasal bar

Se, septalis nerve

sm, myoseptum

sp, Occipitospinal nerves

T, tentacle

t, thyroid

Te, terminalis nerve

Tm, median tentacle of Ammocoetes

U, upper lip

Ul, lower lip

V, velum

v, ventral spinal nerve

ve, velar chamber

Vl, velar ligament

Vn, velar nerve

vg, veloquadratus muscle

W, wall of oral hood

I to V, jaw muscles of Amphioxus

VII, facial nerve

VIII, a and b, two acusticus nerves to
snout complex

IX, glossopharyngus nerve

X, vagus nerve

Figs. 1 to 13, except fig. 10, Amphioxus.

Figs. 14 to 21, Ammocoetes.

Figs. 22 to 36, except fig. 30, Bdellostoma.

Fig. 30, Chimaera.
Fig. 10, Callorhynchus.

ORIGIN OF JAW APPARATUS 343

the long axis of the body. ‘This angle is greater in both Ammo-
eoetes and Bdellostoma than it is in Amphioxus.

The jaw bars extend the entire length of the buccal aperture,
which is not quite as long as the buccal chamber, owing to the
pocket which lies behind the base of the jaw bars and ends in
front of the velum. The bases of the jaw bars are tied together
by tendons and muscle.

Fig. 1 Ventral view of the anterior end of Amphioxus to show relation of the
jaw apparatus to the body wall, myotomes, and velum.

The extension of the muscles forward to the tips of the bars
and out onto the tentacular projections ties all skeletal parts
into a unit structure for the work it has to do.

The chondral jaw bars which frame the buccal opening in
Amphioxus are the simplest expression among living forms of
the vertebrate mandible. But they include much more than
definitive mandible, e.g., of the cartilaginous fishes.

The jaw bars rotate 180° from their position when the buccal
cavity is wide open to their position when the buccal aperture

344 HOWARD AYERS

is tightly closed, and they carry the tentacles with them in
this semicircular sweep. When fully expanded the tentacles
form a widespread fringe encircling the buccal opening. When
fully contracted the tentacles form an interdigitated and usually
jumbled mass crowded into the buccal cavity and entirely fill-
ing it, and then the apparent buccal opening is formed by the
contracted right and left metapleural folds.

There is thus evidence of abundant and varied muscular
action, and the skeletal parts are so arranged as to give free and
extensive change of motion of the parts. The jaw apparatus
of Amphioxus has thus the motions of lateral biting Jaws, but
Amphioxus does not use them as such. It is important to note
that the jaws can be and are frequently withdrawn inside the
buccal cavity.

While the Amphioxus buccal skeleton is developed and ad-
justed to opening and closing the oral aperture, since the food
of Amphioxus consists of minute organisms suspended in the
respiratory current, no biting or comminuting function is
needed. It is apparent how readily adaptable this apparatus
is to such function, and when change of food habit occurred
the jaws could be readily modified to meet the requirements
of the new conditions.

Rising from the dorsal borders of the jaw bars, the right and
left walls of the oral hood arch upwards to fuse together in the
median line below the notochord; posteriorly they pass into
the velar curtain which is perforated by the velar mouth (figs.
PNB ANS attr)

The tentacular rods are generally single, but branched rods
occur in forked tentacles. The tentacles from the second quarter
of the jaw bar reach to the tip of the buccal cavity when retracted
and the short ones project outside (laterad) of them.

The tentacles are covered by extensions of the tissues of the
jaw bar, consisting of a thick layer of surface epithelium and
subepithelial connective tissue in which the nerves pass to their
endings and the vascular channels ramify. The epithelium is
produced at intervals into hillocks containing sensory cells.

ORIGIN OF JAW APPARATUS 345

The anatomy and histology of these structures is quite fully
described in the literature of Amphioxus.

The two mesial tentacles at the base of the jaw bars differ
from the others in being shorter and broader and they leave the

k ie
y
f
A.
pie MI

Fig. 2 Ventral view of the median part of the right and left jaw bars of Am-
phioxus. A shows the usual symmetrical arrangements of the tentacles on the
bar. B shows an irregular arrangement; r and J are the first tentacular rods
given off from the right and left jaw bars.

center of their basal plates, not at the anterior (distal) border
as is the case with all the others. They have the appearance
of palps.

The oral hood has the following attachments: 1) Directly
continuous with the metapleural folds. 2) Strong tendinous
attachment along the dorsal longitudinal lines of contact with

346 HOWARD AYERS

the mesial faces of the myotomes. 3) Bandlike tendons to the
ventral edges of the myotomes. Dorsomesad of the dorsal longi-
tudinal attachment to the myotomes there is a large lymph space
separating the oral hood from the surface of the myotomes and

yc

SISA - i My
PEO IIS Fe EG AD be.

Fig. 3 Sagittal section of the head of Amphioxus near the median line to
show the lower lip, jaw base, velum, and endostyle. The relations of the jaw
muscles to the parent transverse muscle, to the velum, and to the jaw base are
indicated. The jaw apparatus is in contraction.

the notochord across which connective-tissue threads connect
to the adjacent structures.

The muscles which move the jaw bars and their tentacular
projections appear to be derived from the large transverse ventral
muscle of the body wall (figs. 2, 3, 4, 5, 6, and 7). They consist

ORIGIN OF JAW APPARATUS 347

of bundles of fibers turned in varying degrees out of the trans-
verse direction of the fibers of the parent muscle bed. Thus
all the jaw muscles may be said to have their origin from the
ventral transverse muscle and to insert on the jaw bars and their

Fig. 4 A. Composite section to show relation“ of jaw base to velum of Amphi-
oxus when jaw apparatus is expanded. The first three consecutive divisions of
the gut are shown. 8B, buccal cavity; vc, velar chamber; G, pharyngeal canal.

B. Dorsal view of jaw base, velum, velar chamber, median part of gills, to-
gether with median muscle which arises from the transversus ventrad of the
velum.

tentacular projections. The jaw muscles are divisible into two
groups. The first group contains the muscles of attachment;
the second group the intrinsic jaw muscles, yet the latter are
in a way extensions of the former. In figure 3, the relations
of some of these muscles are shown as they appear in median

348 HOWARD AYERS

section through the sagittal plane of the body. It will be noted
how the ventral transverse muscle thickens as it approaches
the base of the jaw bars, also how bundles of muscle fibers take
on a longitudinal direction to insert on the jaw bars. Two large
muscles and one small one which surround the base of the jaw
follow the bars forward as the main intrinsic muscles of the
jaw bars. The largest and most caudad of the three is seen to
contain a large element of transverse fibers — the anterior bundles

Fig. 5 Dorsal view of dissection of jaw base and ventral floor of velar cham-
ber and endostyle of Amphioxus, showing the muscles attached to ventral sur-
face of anterior end of endostyle arising out of the transversus and the lateral
lines of attachment of endostyle to sheath of the transverse muscle.

of the transverse ventral muscle. One longitudinal bundle
is shown which originates from the base of the endostyle and
ventral border of the velum. In figure 2 is indicated the inter-
digitation and the crossing of these three bundles at the base
of the jaw bars. In figure 4B and figure 5 is shown the bundle
which arises from the base endostyle and velum.

In figure 6 the manner of origin of the left half of the jaw
muscles from the transverse muscle to the left of the median
line is indicated. Figure 7 shows a dissection of the jaw muscles,

ORIGIN OF JAW APPARATUS 349

their relation to the transverse ventral muscle, the jaw bars
and basal tentacular palps.

The group of intrinsic jaw muscles has been previously de-
scribed. It consists of a large fusiform bundle (outer muscle)
running the entire length of the jaw bar of either side and a
smaller muscle consisting of long bundles and short thin plates

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Fig. 6 Dorsal view of dissection of jaw muscles of the left side arising from
transversus. Amphioxus.

of muscle fibers extending from one tentacular bar to the next one
in front (inner muscle). These two groups we may call, one
the mandibular muscle and the other the tentacular muscle.
They are both complicated in their make-up.

_ The rectus-abdominis portions of myotomes 1 to 4, and some-
times myotome 5, are attached to the wall of the buccal cavity

350 HOWARD AYERS

along a horizontal line of insertion lying below the dotted outline
of the dorsal border of the oral hood, as shown in figure 12. When
the muscles of the jaw apparatus are fully contracted, the rectus
abdominis section of each myotome contracts in unison to assist
in withdrawing the jaw apparatus up into the boat-shaped cavity
formed by the myotomes on each side and the notochord in

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Fig. 7_ Dorsal view of dissection of jaw base and its muscles. Amphioxus.

the middle and at the same time the metapleural folds are folded
inward, the right over the left, so that the buccal opening is
entirely closed and covered over and protected by these folds
of the skin. The buccal cavity at such time is entirely filled by
the tentacular projections of the jaw bars, crowded together
like so many straws.

ORIGIN OF JAW APPARATUS 351

There are six pairs of muscles which move the jaw apparatus
of Amphioxus. Based on their action they may be listed as
follows:

I. Transversus abdominis, the parent muscle of them all.
Is attached by tendons to basal part of jaw and supplies the
bundles which compose muscles 2, 3, 4, 5. Assists in retracting
the jaw apparatus and contracting the buc¢al cavity.

II. Retractor of the jaw apparatus, arises from transverse
muscle and inserts into base of jaw. Draws the jaw apparatus
backward and, in connection with the transversus, Inward and
upward into the buccal cavity.

III. Adductor of the jaw bars, arises from transverse muscle
and inserts into basal part of jaw dorsolateral of the retractor.
Rotates the mandible inward through an angle of 180° and
adducts the jaw.

IV. Extensor of the jaw bars, arises from transverse muscle
and inserts into basal part of jaw ventrolaterad of retractor.
Rotates the mandible outward through an angle of 180° and
extends the jaw. .

V. Extensor of the jaw bars, arises from each segment of
jaw and inserts into the more distal segments. The powerful
extensor of the mandible.

VI. Adductor of the tentacles, arises from base of tentacles
and inserts into next anterior tentacle.

All of these muscles, which have become largely longitudinal
in direction, do not appear to have distinct sheaths; but from
the clear-cut motions of opening and closing the buccal opening,
retracting and in-folding the jaws and tentacles, it is certain that
they have sharply defined mechanical pulls and are subject to
an equally distinct nerve control. The whole jaw apparatus
is anchored by tendons which extend from the muscles to the
mesial surfaces of the body wall in front of the velum. Both
tendons and slips of muscle extend from the velum to the jaw
base.

The jaw apparatus as a whole is anchored by tendons to the
ventral transverse muscle and to the tough aponeurosis covering
the mesial surface of the myotomes. It functions in existing

aoe HOWARD AYERS

Amphioxus: a) to keep the mouth, at times, wide open; b) to
screen its entrance; c) to close it entirely; d) to serve on occasion
as a strainer for the food current; e) to direct the food current
to the-oral aperture.

The major motions of the jaw apparatus are three. First,
the motions of approximation and separation of the two arms
of the loop, with the resultant interdigitation and separation of
the tentacles. Second, the movement of elevation of the whole
jaw apparatus to the roof of the oral hood so that its long axis
lies nearly horizontal and parallel to the notochord, this move-
ment affecting the posterior end of the loop most, and that of
the depression of the jaw apparatus resulting in the increase of
the vertical diameter of the buccal cavity. Third, the retrac-
tion and extension of the jaw apparatus due to the pull of muscles
attached to the base of the loop and to the general contraction
of the walls of the oral hood and body wall. The buccal frame-
work can thus be withdrawn entirely within the oral hood or
fully protruded as a projecting structure beyond the limits of
the oral aperture. |

The jaw apparatus arises in the young Amphioxus in the lip
fold as two skeletal bars symmetrically placed on either side
of the sagittal plane. These bars begin at the middle of the
lower lip fold and grow outwards, then upwards, and forwards
into and through the edge of the lip fold. The bars send out
buds at intervals which later form the tentacles. These buds
of the jaw bars push the muscular layer, the connective-tissue
layers and the covering epithelium before them in such a manner
that they fit over the skeletal axis of each tentacle as the finger
of a glove covers a finger, except that the muscles extend only
part way up the tentacle.

Amphioxus—jaw nerves

The researches of Hatschek, Heymans and van der Stricht,
van Wijhi, Kutschin, Dogiel, and others, as well as my own,
on the nerves of the head of Amphioxus have one outstanding
result, viz., the oral hood is under the control of five pairs of

ORIGIN OF JAW APPARATUS aH3

nerves from the central nerve cord which lie between the second
nerve—the septalis—and the velum. These five nerves are the
nerves of the jaw apparatus and velum, supplying also the walls
of the buccal cavity and the other organs connected therewith,
besides giving branches to the skin and myotomes. My own
_ repeated studies of the head nerves of Amphioxus are in agree-
ment with the major facts which may be classed as well estab-
lished. There is great flexibility in the arrangement of the
nerves of the head. The septal nerve and the eighth may also
take part in the nerve supply of the jaw apparatus, but this is
not usual.

The frequent presence of unusual dorsal roots and the occa-
sional absence of one of the important visceral trunks are purely
individual variations, and do not affect the large facts of the
nerve supply of the jaw apparatus which are given here in con-
densed form for convenience of comparison with the nerve supply
of the jaw apparatus of Bdellostoma.

An accurate drawing of the five nerves of the left side control-
ling the jaw apparatus, together with a posterior ventral branch
of the septal nerve and the first postmandibular nerve or the eighth
nerve counting from the terminalis, is shown in figure 12. The
course of the visceral branches over the myotomes, and the
larger branches to the jaw apparatus are accurately drawn. There
is great variation in method of branching in different individuals,
but this figure is typical for Amphioxus lanceolatus. The velar
nerves are not drawn in, and only one of the deep branches turn-
ing mesad under the ventral edge of the body muscle is partly
shown. In general these nerves show only a slight tendency
to run cephalad, most noticeable where the ventral branches
emerge from behind the myotome. Their general course is nearly
due ventrad to their terminals, which in Amphioxus is the plexi-
form nerve net described by Fusari, Dogiel, and Kutchin as con-
sisting of three main sections, an inner and outer jaw bar plexus
and a more or less distinct tentacular plexus. From the plexiform
structures the final terminal nerves are given off. The plexi-
form structures are associated with a longitudinal nerve trunk
composed of a number of relatively large nerves running the

354 HOWARD AYERS

length of the jaw bar near the base of the tentacles made up
of the larger nerve branches given off from the five jaw nerves.
A satisfactory analysis of this longitudinal trunk, its components
and the relations of the plexiform structures to it has not been

Fig. 8 Dorsal view of velum, velar tentacles and chamber, and anterior end
of gill basket of Amphioxus. Six of the velar tentacles are protruded into the
buccal cavity. The attachment of the velar ligaments is shown on the left side.
The insertion of three myotomes is seen on the left side.

published. One or more branches of the longitudinal trunk
run out into each tentacle. Thus the ventral branches of five
pairs of segmental nerves of unmodified spinal type supply the
jaw apparatus and exercise control of the whole mechanism

ORIGIN OF JAW APPARATUS Shi,

through a complex nerve-net system of fiber distribution which
involves both sensory and motor terminals. The special asym-
metry of the nerve supply confined almost entirely to the
velum will be taken up later, but it may be said in passing

Fig. 9. Compound tendon connecting muscles of velum to body wall of Am-
phioxus. At the velar end, Mc, are shown a few cut ends of the circular muscles
of the velum while the tendons pass into the radial fibers. At the distal end the
tendon spreads out into a brush of fine tendonous threads which insert in the
connective tissue wall of the myotomes.

that it does not affect the result of the innervation or
the conclusions regarding the innervation of the jaw apparatus.
Kutchin’s observations that a branch of the third nerve of the
right side furnishes a branch to the inner mouth plexus, hereto-

356 HOWARD AYERS

fore held to be formed by branches from left nerves only, shows
that we have yet to learn the complete story of the innervation
of the jaw apparatus. Even though van Wijhi’s explanation
of the asymmetry of the formation of the inner buccal plexus
and the innervation of the velum is largely correct, the fact
remains that the right side can and does supply its bilateral
share to the buccal nerve supply of the inner plexus occasionally

to show jaw apparatus. The remnant of the distal part of the jaw bar is attached
to the hyoid as in Bdellostoma and runs forward and upward to the tentacular
cartilages of the nasal region, thus preserving the ancient amphioxine jaw bar in
a Gnathostome head.

if not frequently, and it would not require much more than has
actually been observed to establish the complete bilateral sym-
metry in the descendants of Amphioxus which we observe in the
higher forms, where the nerve supply of the right and left sides
is usually finely balanced.

Thus the five nerve trunks innervating the jaw apparatus
run ventrolaterad with a slight inclination cephalad. In the
jaw their main branches run longitudinally. They leave the
central nerve cord as separate and distinct dorsal roots. There

ORIGIN OF JAW APPARATUS aot

has thus been but slight displacement of the root portions of
these nerves caudad. The septal nerve, while largely devoted
to the sensory innervation of the snout, frequently sends one or
two branches to the anterior end of the jaw apparatus.

The innervation of the jaw apparatus is quite sharply limited
posteriorly. There is thus a section of the central nervous system

Fig. 11 Front view of velar disk of Amphioxus with the dorsal six of the velar
tentacles projecting into the buccal cavity. When the lower six are thrust for-
ward the lip fold disappears.

lying in front of the jaw apparatus whose peripheral nerves do
not typically invade the jaw territory and another section of
the central nervous system whose nerves supply territory nor-
mally behind the jaw apparatus; between these two lies a section
of the central nerve cord which, to judge by the five nerves it
sends out, is devoted exclusively to the control of the jaw appara-
tus. In the peripheral reaches of its nerves is built up a plexi-

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 4

358 HOWARD AYERS

form structure which does not appear to be duplicated in other
peripheral territories.

The velum is a part of the jaw apparatus; it is the organ about
which has centered all this structural activity. It is lodged
between two folds of the mucosa which fuse to form the velar
lip and the coverings of the tentacles, and is continuous with
the lining of the buccal cavity anteriorly and the foid which is
continuous with the lining of the branchial cavity posteriorly
(figs. 1, 3, 4, 8). The velum of Amphioxus is a muscular dia-

Fig. 12 Left side of jaw apparatus of Amphioxus. The dotted outline marks
the position of the dorsal wall of oral hood. The relations of the first seven
myotomes and the nerves 2 to 7 are accurately drawn. Nerves 1 to 6, the jaw
nerves. The heavy dashes mark the position of the ventral spinal roots.

phragm. It varies much in shape, depending on the degree of
contraction. When fully expanded it forms a thin circular
ring with a large central hole, from the anterior rim of which
twelve or more velar tentacles project forward into the cavity
of the oral hood. In this state the radial muscles are contracted
and the circular muscles are relaxed. This is the condition of
the velum when the oral hood is wide open and the jaw apparatus
fully expanded. When fully contracted the velum has the shape
of a thick doughnut minus the hole and the tentacles project
backward into the velar chamber or are folded across the posterior
face of the muscular lump. When Amphioxus is killed with

ORIGIN OF JAW APPARATUS 359

quick-acting reagents (e.g., osmic acid) it often happens that
the velar tentacles are not all withdrawn and some of them still
project forward (fig. 11), while others project backward. In
life the velar tentacles were long ago observed to occasionally
flip back and forth while the velum was fully expanded. Their

\

& >
a< 5

Fig. 13 Part of the brain and the terminal and septal nerves of Amphioxus.

Two branches of the latter supplying the anterior end of the jaw apparatus are
shown.

normal position while the food current is flowing in through the
velar aperture is that of full erection projecting forward into
the oral hood, forming a funnel-shaped fence about the velar
mouth. In the normal closure of the velum the tentacles are
folded toward the center and caudad due to the fact that the

360 HOWARD AYERS

radial muscles of the posterior face of the velum insert into the
base of the tentacles and draw the lip caudad, thereby flexing
the tentacles caudad and pulling the lip caudad of the muscular
diaphragm. In expanding, the motions are the reverse of these.
The velum is firmly tied to the surface of the myotomes on each
side as well as to the base of the jaw apparatus in front and the
gill basket behind. It thus forms a flexible curtain between
the buccal cavity and the gill chamber.

AMMOCOETES

The jaw apparatus of Ammocoetes is laid down after the same
plan as that of Amphioxus. It consists of two lateral jaw bars
of soft cartilage and procartilage, which are largest at their
union in the midventral line and taper to their anterior dorsal.
ends located on the ventral surface of the upper lip. They
form the boundary of the buccal aperture with their proximal
vertical portions and inclose between them a remnant of the
ancestral buccal aperture of Amphioxus in their horizontal
distal portions. They bear tentacles throughout their entire
length (figs. 14, 15, 18, 19, and 21).

In the midventral line they fuse, and from this junction is
given off dorsally a flask-shaped body which is crowned by the
median tentacle, and ventrally the jaw bars are continued cau-
dad in the form of a large spike of cartilage or median projec-
tion of the jaw bars (figs. 15, 18, 19, 21).

A reduction of the distal part of the jaw bars together with
an extension of their proximal portions has taken place since
the Amphioxus stage was passed, but the amphioxine charac-
ter of the jaw bars is plain to see. Viewed from in front the
jaw bars curve outward and upward nearly vertically from the
midventral line, then inward, but do not meet in the middorsal
line, thus forming an incomplete ring with the opening above.
Up to this point they are inclosed in the lips of the buccal aper-
ture. The ends of the open ring lie at the junction of the upper
lip with the head. Here the jaw bars bend forward nearly at
right angles and lie on the ventral surface of the upper lip, and
the position of the anterior part of the ancestral buccal chamber

ORIGIN OF JAW APPARATUS 361

is marked by the tentacle-covered surface which leads back into
the buccal opening between the vertical parts of the jaw bars.
Thus the posterior part only of the ancestral buccal aperture
is functional in Ammocoetes (figs. 14, 18).

Fig. 14 Upper lip of a 15-em. Ammocoetes seen from ventral surface. The
positions of jaw bars are in part outlined by two of the rows of tentacles and the
long muscle of the jaw between the rows of tentacles. In front of the functional
buccal aperture is included the remains of the anterior part of the buccal aperture
of the amphioxine ancestor. The number of rows of tentacles and consequently
the width of this space varies in different stages of growth. The jaw muscles
shown are those belonging to the jaw bars and tentacles and the muscles at the
base of the jaw apparatus. On the right side of the figure the forward extension
of the parietal muscles is shown. They attach to the upper and lower lips. In
front of the buccal aperture only the two main rows of tentacles are indicated.
In this 15-em. Ammocoetes there are four additional rows of smaller tentacles
on each side. The dotted outlines show the boundary of the tentacular region
in an 8-cm. Ammocoetes and the rows of dashes the position of rows of small
tentacles of this stage. The median line of dots marks the position of a median
line of small tentacles sometimes present.

362 HOWARD AYERS

When seen from the side (fig. 18), the proximal part of the
jaw bar stands perpendicular to the ventral bar. Where the
upper lip joins the head the jaw bars bend from the vertical to
a horizontal position and run forward in the ventral part of
the upper lip just below the skin, converging as they go forward.

Fig. 15 Dorsal view of the skeleton of the jaw apparatus of Ammocoetes,
showing the jaw bars, bases of part of the tentacles, the large median tentacle,
the extent of the hyoidean skeleton and outline of the lower lip.

The vertical portions of the jaw bars form the elastic skeletal
support of the buccal aperture. The basal median bar is a more
or less cylindrical piece with its largest diameter where it unites
with the jaw bars; here it gives off a short process which pro-
jects forward and upward, swelling into a bulb or flask-shaped
body with a slender neck crowned by a bush of tentacular branches

ORIGIN OF JAW APPARATUS 363

which are mainly directed forward and upward. When the
tentacle is fully erected the branches stand nearly vertical to
the long axis of the body. About this bush are grouped ten
other similar bushes, the crowns of ten lateral tentacles, five on
each side. These bushes are so set that their branches inter-
digitate, forming a screen which separates the functional part
of the buccal cavity from the outside (figs. 14, 18).

The posterior part of the median bar projects caudad in the
ventral wall of the body as far back as the branchial region,
where it tapers to a point in the connective-tissue framework
of the body wall. Below the base of the jaw bars and the median
bar is a sheet of procartilage which thins out laterally as it passes
upward in the body wall on each side and also as it runs caudad
below the median bar. This sheet of procartilage gives rise
to the hyoidean apparatus of Petromyzon (figs. 15, 21).

As in Amphioxus, the jaw bars bear tentacles their entire
length. Present as a single row about the functional part (buc-
cal aperture) they appear in a varying number of rows on the
surface of the lip, the rows being more numerous in younger than
in the older stages of growth. The tentacles vary from short
finger-shaped projections, such as are given off at the distal
extremity of the rows, through all gradations to the bushy struc -
tures given off from the base of the jaw bars (fig. 16). The
branching is dichotomous. As Prof. 8. H. Gage has explained,
this group of tentacles forms a strainer or sieve in front of the
buccal cavity. Along the horizontal part of the jaw bars the
tentacles project from the ventral concave face of the upper
lip, due to the spreading out of the skin cover of the jaw bars
as part of the lip covering. They decrease in size toward the
tip of the bars and also from the mesial rows laterad. They
form converging rows of fungoid outgrowths, coursing laterad,
the more posterior ones terminating in the border of the ten-
tacular area, the anterior (mesial) ones converging toward the
middle line near the front border of the lip (fig. 14). The median
tentacle mentioned above is the largest of all and surmounts a
bulbous structure which after the metamorphosis appears as
part of the skeleton of the mandibular mechanism of Petromy-

364 HOWARD AYERS

zon. Kaensche has described the part taken by the skeleton
of the median tentacle in forming the jaw of Petromyzon as
well as the origin of the hyoid from the sheet of procartilage
of the Ammocoetes which lies ventrad of the jaw bars. The
buceal screen is figured by Professor Gage on plate VI, figure
22, and, as he states, this tentacular sieve prevents the entrance

Fig. 16 A. One of the larger-branched tentacles from the jaw bar.
B. Four small tentacles to show method of branching.

of large particles into the buccal chamber and when tightly
closed shuts off the flow of water through the buccal aperture.

The mandibular bars are not so conspicuous superficially
as in Amphioxus, but they occupy the same relative position
in the head. Due to the shortening of the notochord, the for-
ward overgrowth of the brain, and the housing of the nasal
organs, the head does not project beyond the boundary of the
buccal chamber marked out by the proximal part of the jaw

ORIGIN OF JAW APPARATUS 365

bars. The head has also acquired a large muscular upper lip
which carries on its undersurface the anterior part of the terri-
tory of the oral hood, or correctly put, the distal part of the
oral hood has acquired a muscular roof which has suppressed
the cavity. The muscles of the mandibular bars consist of two
tapering fusiform bundles of fibers which accompany the bars,
flattening and thinning out on the horizontal part of the jaw bars.

Fig. 17 Ventral view of superficial muscles of 10-cm. Ammocoetes, to show
the transverse muscle and its derivations that lie in the lower lip.

At the base of the jaw bars they are thicker than the bars and
arise from the lateral borders of the median bar and the floor of
the buccal chamber. They pass foward, outward, and upward
along the outer border of each jaw bar. Where they originate they
interdigitate with the circular muscles of the head. They furnish
the tentacles with muscles, sending off shoots into the larger ones
(fig. 16), which attach to the walls. The jaw muscle is attached to
the skeletal tissue at the base of all tentacles, both large and small.
The motions of the jaw bars in Ammocoetes are not as varied

366 HOWARD AYERS

or extensive as in Amphioxus, owing to the fixation of the anterior
part of the jaw apparatus to the upper lip. The contraction
of the vertical portions of the jaw muscles closes the buccal
aperture and they are aided in this action by the circular muscles
of the head, which form a special sphincter surrounding the
buccal aperture outside the jaws. The expansion of the buccal
aperture seems to be largely due to the elasticity of the chondral
jaw bars aided by the ventrolateral retractors of the jaw bars.

Jo

Fig. 18 Composite picture of sagittal sections of head of Ammocoetes. The
position of the eye is indicated, as well as the circular muscle which begins in
front of the eye and runs ventrad to insert on the jaw apparatus. The left jaw
bar with its tentacles and the median tentacle are shown as though lying in the
plane of the section. The tentacles are outlined as though solid bodies.

The head muscles of the Ammocoetes consist of the parietal
or trunk muscle, the branchial, velar, buccal, jaw, and lip mus-
cles. The buccal and velar muscles are composed of two groups,
the ring and the longitudinal. ‘The ring or transverse muscles
consist of two layers of fibers separated by a layer of procartilage.
They are the continuation forward of the muscles of the branchial
basket. The group of buccal muscles attach to the base of the
cranial skeleton, both the cartilaginous and membranous cranium,
and to the nasal capsule. The general direction of the fibers is ven-
trad in the sidewalls of the head, to curve inward from right toleft
to the fusion of the muscles of the two sides along the median ven-

ORIGIN OF JAW APPARATUS 367

tral line; the only indication of a median raphé is the interdigita-
tion of some of the muscles, those not interdigitating have their
fibers continuous. In front of the nasal capsule the dorsal

B ZEN
= yy 3

Fig. 19 Dorsal view of the floor of the buccal cavity, velar chamber, and
anterior end of branchial cavity of Ammocoetes. The right and left arms of the
jaw bar are shown to the left and right of the median tentacle, also its median
tendon extending caudad in the ridge in which the opening of the thyroid gland
occurs. The velum with the ventral attachment of its muscle and the positipn
of the anterior gills are also shown.

layer runs into the upper lip and the bundles cross each other.
Below the capsule two heavy bundles arising from the lateral
surface of the ring muscle next the mucosa run into the upper
lip, crossing one another in front of the palatine loop. In the

368 HOWARD AYERS

ventral body wall, especially in the velar region, the fibers of the
ring muscle interlace across the middle line of the body.

The velar muscles arise from three territories: first, a strong
group from the skull behind the eye; second, a band of fibers,
not continuous, from the lateral part of the neighboring ring
muscle; and, third, a long but heavy bundle from near the mid-

WY
Y=
= = = _ Yy ————
SS Se
SSS

Fig. 20 Ventral view of the jaw and velar muscles, derivatives of the trans-
verse or circular muscles of the head. Ammocoetes.

ventral line of the branchial skeleton. This latter bundle is
long, and sweeps forward, upward, and outward into the velar
body to form the most important muscle of the organ. It is
the retractor veli. The velum is greatly assisisted in closing
by the elastic velar cartilage.

Besides the large jaw muscle already described, which lies
on the outer face of each jaw bar and spreads out on the ventral

ORIGIN OF JAW APPARATUS 369

face of the upper lip, other muscles belonging to the constrictor
group assist in the control of the jaw apparatus through their
insertion into the base of the upper lip. The upper lip muscles
as a whole form a concavoconvex muscular flap attached to the
anterior end of the head—indeed, the upper lip is the muscular
roof of the oral-hood territory and the jaw bars and tentacular
skeleton form its main support. Muscle bundles run into it
from the right and left sides from above and below and the
lower lip is a unit part as far as the muscles are concerned. The

Fig. 21 Sagittal section of the anterior end of the head of Ammocoetes, to
show the jaw bar in the median plane with the large tentacle, the position of the
velum and the dorsal body wall.

lips thus form a funnel with the lower third of its wall cut away
almost back to the jaw bars.

The jaw apparatus is under the control of the trigeminus and
facialis. I have not given these nerves of Ammocoetes the
necessary analytical dissection, and cannot therefore make com-
parison with the nerves of the jaw apparatus in Amphioxus.
The Marsipobranch innervation will be described as it occurs in
Bdellostoma. Here a thorough dissection shows the Marsipo-
beanch innervation of the jaw apparatus is fundamentally in
harmony with that of Amphioxus.

370 HOWARD AYERS

BDELLOSTOMA

Great as the structural differences between Amphioxus and
Ammocoetes undoubtedly are, they are not in some respects
greater than the structural differences between Ammocoetes
and Bdellostoma.

It is an accepted fact that Ammocoetes has progressed much
further toward the condition of anatomical organization found in
fishes higher in the scale of development than has Amphioxus.
It is also true that Bdellostoma has reached a much higher state
of organization than Ammocoetes and in general in the direction
toward fishes reckoned as more advanced in structure, e.g., the
Chimaeroids and sharks. It is generally held that the Myxi-
noids are not jaw-bearing fishes, although the evidence is accumu-
lating to prove that they are. Having presented evidence to
show that the buccal skeleton and some associated structures
in Amphioxus are homologous with the parts of the jaw apparatus
of Ammocoetes, if we can now show that the heretofore puzzling
jaw apparatus of Bdellostoma is built on the same plan and can
establish the essential homology of its parts with these of Am-
phioxus and Ammocoetes, we shall advance far in our effort
to solve the jaw problem.

The skeleton, mus¢éles, and nerves of Bdellostoma have been
subjected to thorough investigation, but have not by any means
yielded up all they have to tell. No complete description of
these structures will be given here, but only such details as seem
pertinent to the problem under consideration.

The skeleton of Bdellostoma is both cartilaginous and mem-
branous; many structures important for comparison with higher
forms are still in the membranous stage of their development.
The comparison of the parts which have become cartilaginous
with the same membranous territories in Ammocoetes and Ani-
phioxus is indeed instructive and the same may be said for carti-
laginous parts in the higher fishes which remain membranous in
Bdellostoma. In such comparison is a rich mine of morpho-
logical data.

For more detailed accounts of the anatomy of the skeleton,
muscles, and nerves the reader should consult the works of

ORIGIN OF JAW APPARATUS 371

J. Miller coveringa general survey of the anatomy of Bdellostoma,
Ayers and Jackson on the anatomy of the skeleton, Julia Worth-
ington on the head nerves, EK. P. Allis on the skeleton, nerves,
and muscles of the head, F. J. Cole on the skeleton and muscles
of Myxine, and P. Firbringer on the skeleton, nerves, and
muscles of Myxine.

The jaw apparatus of Bdellostoma consists of a pair of den-
tigerous jaw plates located in the buccal cavity and lying above
a large cartilaginous plate which forms a support for it, of a
pair of bars starting from near the sides of the jaw plates, but

Fig. 22 Front view of face of adult Bdellostoma, with jaws protruded, show-
ing external relation of tentacles to narial and buccal apertures. With the hyoid
as a base, the jaw bars pass upward giving off the tentacles and framing in the
buccal opening. The dotted line on the left side indicates the course of the
distal part of the jaw bars in front view.

attached to the large support plate which lies ventrad of the jaw,
and which run forward and upward in the lips of the buccal
cavity (figs. 22, 28, 24, 25, 26, 27, 29). They attach at their
distal ends to the lateral borders of the nasal tube near to its
anterior end. The first part is the basal or proximal part of
the Amphioxine jaw bar and the second part is the distal section.

The proximal part has been transformed into a tooth-bearing
protrusible jaw, while the distal part serves as the skeleton of
the lips of the buccal aperture and is the only part bearing
tentacles in Bdellostoma. The number of tentacles is reduced
to four, two of which (the first and third) are unit structures

3f2- HOWARD AYERS

with the cartilaginous jaw bar, while the other two (the second
and fourth) are connected only by tendons to the jaw bar. The
cartilages of the tentacle, two on either side, are fused together
where they meet in the middle line below the nasal tube, and
through this bar they are connected with the subnasal cartilage.
The cartilages of tentacle four lie in the heavy lip folds at the
sides of the ventral part of the mouth opening, and are so held
by tendons and muscles as to serve as a firm support for the pair
of thick flaps of skin which operate as a mouth guard and jaw
cleaner, as well as a palp. The other three tentacles are sensi-

a B Mj
es ee ee

Fig. 23 Composite figure from sagittal sections of head of adult Bdellostoma
showing outlines of important parts of jaw apparatus. The tentacular part
of the jaw bar, of course, lies in the side of the head. The tooth-bearing jaw
cartilage is also shown, but it also lies at one side of the median plane.

tive fingers used in connection with both nasal and buccal
apertures.

The important relation is that the jaw bars are the skeletal
framework of the buccal aperture just as they are in Amphioxus
and Ammocoetes. We found in Ammocoetes that a new skeletal
structure had been added ventrad of the base of the jaw bars and
that this sheet of procartilage of Ammocoetes developed into the
submandibular cartilage of Petromyzon. In Bdellostoma we
find this element larger and assuming proportions unusual, so
much so that Prof. G. B. Howes called it “the dominant mon-
ster of the Hag.’”’ It is however, admirably adapted to its
functions. It serves as a traveling floor for the jaws in their
extensive excursions out of and into the buccal cavity. Its

ORIGIN OF JAW APPARATUS aie

front edge also serves as the fulcrum over which the power-
ful pulls are exerted in the rasping and tearing action of the
teeth of the jaw. It further serves as the firm base of attach-
ment of a large group of muscles which take part in the work of
operating the jaw and in closing the buccal cavity as a whole.
The attachment of the distal ends of the jaw bars to the anterior
end of the base plate or hyoidean apparatus is well accounted
for by the close association of the two structures both in posi-
tion and in functioning and by the necessity for the separation
of the distal from the proximal division of the jaw bar in order
to secure freedom of motion of the latter. The position of the
distal part of the jaw bar varies with the degree of contrac-

Fig. 24 The adult cartilaginous skeleton of the jaw apparatus of the right
side, including the corneal cartilage, anterior end of jaw and hyoid to which the
jaw bar attaches. Bdellostoma.

- tion of the snout region of the fish, but, as figures 22, 23, 24,
25, 26, 29 show, it runs forward and upward at an angle that
recalls the slope of the jaw bars in Amphioxus.

The hyoidean apparatus has acquired cartilaginous connec-
tion with the skull above (figs. 25, 26, 27), and is also connected
thereto by strong tendinous bands. ‘The jaw itself has no attach-
ment to the cranial skeleton except the tendinous attachment to
the anterior end of the nasal capsule and a similar connection
through the corneal cartilage with the palatine and subnasal
bars. As the jaw works longitudinally while in the head, it
has no vertical stresses not already provided for by the hyoidean
supporting mechanism, and it is the latter which has developed
the cranial attachments mentioned above.

374. HOWARD AYERS

The muscles of Bdellostoma have been differentiated to such
a degree beyond the Ammocoete condition that they have not
yet been reduced to a grouping that permits comparison with
Ammocoetes. IJ had planned to publish an account of the muscles
of Bdellostoma in 1898; it would not have cleared many of the
problems seeking solution. That paper went to the waste-
basket two years later. Since then I have sought a basis for
comparison of the muscles of Bdellostoma with both lower and
higher forms. In 1896, F. J. Cole published a very full account

T Ae SSE Nn ee eee
f a og

Fig. 25 Dissection of the left side of skeleton of jaw apparatus of Bdellostoma,
to show the relation of its parts with dentigerous jaw protruded and retracted.
The hyoidean mechanism bears the tentacular portion of the jaw bars fastened
to its anterior end, thus leaving the dentigerous jaws free to move unhampered.

of the muscles of Myxine with frequent references to the muscles
of Bdellostoma, but he recognized that the comparative mor-
phology of his subject was not ripe for solution. What is here
offered is a grouping of the Myxinoid muscles based on the
comparative anatomy of the jaw apparatus as outlined within.

It is evident that we have to deal with two quite distinct
major groups of muscles associated with the jaw apparatus.
The first and oldest group is that of the intrinsic muscles of
the jaw apparatus which are developed out of the ventral plate
of transverse muscle fibers of the Amphioxine ancestor. Al-
ready in Amphioxus the group of intrinsic muscles is sharply

ORIGIN OF JAW APPARATUS 315

defined, and they only grade into the ventral transverse muscles
where the bases of the jaw bars attach to the parent muscle.
In Ammocoetes we have the system of constrictors quite com-
plete as such, forming a broad girdle of muscle bands encircling
the branchial pharyngeal and velar region, attaching to the
axial skeleton above and holding the base of the jaw bars in
their grasp. With many interruptions of the continuity of the
ring muscles in the branchial region these groups of muscular
girdles send out offshoots—to the velum, to the jaw apparatus,
and to the lip flaps. The phylogenetic history of these ring-

Fig. 26 Side view of dissection of head of 37-mm. Bdellostoma embryo, to
show relation of jaw apparatus at this stage. The dentigerous jaw is omitted.
The abrupt bend ventrad of the nasal region is shown, as also the folding of the
‘skin on the dorsum of head.

—s

muscles is not clear. We find in Ammocoetes the hyoidean
apparatus laid down in procartilage within the ring-muscle
system and extending from below the jaw bars backward. It
cares for stresses beyond the function of the muscles, but the
ring-muscle bundles still largely retain their continuity as rings.
In Bdellostoma, however, a larger and more active fish, the
ring-muscles have been broken into segments of rings by the
very great development of the hyoidean apparatus in both
cartilaginous and membranous form and the segments of the
ring muscle of the Ammocoetes stage now insert, by muscle ends

376 HOWARD AYERS

on the cartilages of the hyoidean apparatus and the cranial
skeleton which has also developed a number of new cartilages
and increased the relative size and strength of its tendon and
fibrous bands. The muscles of Bdellostoma may be grouped
according to derivation as follows:

Group 1. Muscles of the jaw apparatus. The most ancient muscles of the
vertebrate head, not considering the but little differentiated myotomes. The
muscles of this group are all derived from the ventral transverse muscle of Am-
phioxus. Some of them have already acquired a longitudinal direction in Am-
phioxus. This process is continued in Ammocoetes and Bdellostoma.

A. Muscles of the jaw bars

a. Of the dentigerous jaw:

Retractors
INGE ees tyes 1. Mandibularis (longitudinalis linguae)
11, (Cs Revel Ra a 2. Perpendicularis

Protractors

3. Hyo-copulo-glossus
4. Copulo-glossus superficialis
5. Copulo-glossus profundus
b. Of the distal part of the jaw bar and tentacles:

QipGh. atone ae 6. Nasalis

Opthi:aseseeece 7. Tentaculo-posterior
Onthieseceesc nas 8. Tentaculo-ethmoidalis

Opthos eendsioes 9. Transversus oris
Optlissse.ecase ke 10. Ethmoideo-nasalis

Optics ae 11. Palato-ethmoideo-superficialis
INES aterm eetne Meet 12. Palato-ethmoideo-profundus

B. Muscles of the hyoidean apparatus

U5 ote ae eae 13. Palato-coronarius

IM eaxetarr aster cay sare 14. Copulo-tentaculo-coronarius

IMeixatat spans ots oer 15. Coronarius

Tea sresek <2) acy cts es 16. Copulo-ethmoidalis

IW ilifears 5 keep 17. Copulo-copularis

Max (VII?)...... 18. Copulo-palatinus
EDT et ees 19. Hyo-copulo-palatinus
VIL......... 20. Copulo-quadratus superficialis
VIL......... 21. Copulo-quadratus profundus
VII......... 22. Cranio-hyoidens

WEES a ths 58 Medora ce 23. Quadrato-palatinus

C. Muscles of the velum

Md.............. 24. Velo-quadratus
Md?....3<......0. 25: ‘Velo-spimalis

ORIGIN OF JAW APPARATUS atl

D. Muscles of the pharynx and gills

IX............... 26. Constrictor pharyngis
1D Fae ee re ee 27. Constrictor branchiarum et cardiae

E. Muscles of the trunk

Bpeljs.sl-...:..2 28: | Parietalis
Spec doe wi eestes 29. Obliquus
2) Deed oo eS Sea 30. Rectus
F. Muscles of the cloacal region
SDMA he deste Seales 31. Sphincter cloacae
G] hoe GoORToe ene 32. Transversus caudalis
Bile cn arcs, = 33. Cordis caudalis

Fig. 27 Front view of the same dissection (37-mm. embryo), to show -rela-
tion of the external tentacles to mouth and the cartilaginous skeleton of the jaw
apparatus.

This may be subject to some changes as a result of further
study of the interrelations of the muscles and skeleton, partic-
ularly after a study of their embryology and a more complete
knowledge of their innervation which as published is both in-
accurate and incomplete.

The attachment of the mandibular mechanism to the base
of the skull and its association with the maxillary mechanism
is a remarkable work of codrdination and correlation which has
taken place in the vertebrate stock above the Myxinoids. An
interesting variety of experiments involving the use of numerous
mechanical devices is presented by the cartilaginous fishes,

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 4

378 HOWARD AYERS

not to mention other groups. The acquisition of biting jaws
was an important event in the evolution of vertebrates, for it
marked a great advance in the mechanics of food-gathering and
of defence. What advance, if any, Bdellostoma has made toward
the acquisition of a maxillary mechanism is not particularly
clear. Allis is the only investigator who has sought to identify
the maxillary element by comparative means, taking into con-
sideration skeleton, nerves, and muscles. His conclusion that
the cartilage of the fourth tentacle represents the cartilaginous

Fig. 28 Face view of 37-mm. embryo before dissection was made for figures
26 and 27. Note the small buccal aperture and the relations of tentacles to both
nasal and buccal openings.

precursor of the osseus maxilla of Ganoids and Teleosts and that
the lateral labial cartilage is in fact the cartilaginous palatine
element does not appear to me to be well founded. Iam not yet
willing to say that it is impossible, but it is very improbable in
my judgment. This much is certain, the fourth tentacle is used
by Bdellostoma as an antagonist to the jaw as already described,
in that it is at times pressed against the dental surface of the
dentigerous mandible. In the event the fourth tentacle proves
to be the nascent maxilla, we would have the whole mandibulo-
maxillary apparatus of vertebrates developed out of the Amphi-

ORIGIN OF JAW APPARATUS 379

oxine jaw bars, which would be a peculiarly fit use to make of
such building material.

Fig. 29 Three figures to show the basic-similarity in the arrangement of the
jaw apparatus in A. Amphioxus, B. Ammocoetes, C. Bdellostoma. 1 to 4 are
five nerves of jaw apparatus.

When the jaw is protruded the fourth tentacle lies above and
upon the exposed dental surface, and when the jaw is being re-
tracted the tentacular flap serves to clean the edge of the jaw
of foreign particles.

380 HOWARD AYERS

Amphioxus, Ammocoetes, and Bdellostoma, in the order named,
represent three stages in the phylogenetic development of ver-
tebrates. They are the only known living representatives of
the earliest vertebrate forms, as far as our knowledge reaches.

(4

Fig. 30 Left side of facial region of skull of Chimaera, to show amphioxine
jaw apparatus still in existence in a Gnathostome. / to 9 are parts of jaw bar
and tentacles.

In view of the great structural differences between Bdellostoma
and the Chimaeroids and sharks, the extensive changes involved
in the evolution of the Amphioxine type into the Myxinoid
type and our incomplete knowledge of both the anatomy and
embryology of the latter it is evident that additional studies are
needed if we expect to arrive at satisfactory solutions of the

ORIGIN OF JAW APPARATUS 381

problems concerning the phylogeny of the organs of the verte-
brate body.

The solution here offered of the Trigeminal complex of Bdello-
stoma is the outline of a new description of these nerves based
on my investigation of the head nerves of this fish. Out of
the several hundred dissections made in detail, to cover the
relations of each nerve as completely as possible, finally came the
realization that the trigeminal complex of Bdellostomais arranged
after the Amphioxine plan and apparently inherited directly
from Amphioxine ancestors. Although the relations of the
five segmental nerves which compose the nerve supply of the
Bdellostoma jaw apparatus are masked by being crowded together
and bound up in dense connective sheaths, they can be made
visible, and their homology then becomes evident. The figures
31 td 35 are drawn from dissections and are selected from a large
number made to illustrate the anatomy of these nerves. They
show some details of root origins and terminal distribution of
importance in establishing the structural similarity of the in-
nervation of the jaw apparatus in Amphioxus and Bdellostoma.
It is not intended to fully describe these nerves, but merely to
present certain features pertinent to their comparative anatomy
as Jaw nerves.

A detailed account of the head nerves of Bdellostoma will be
given in another contribution. Miss Worthington, Allis, and
Miiller have described the head nerves in Bdellostoma and
Fiirbringer and Bridge those of Myxine, to mention the more
important contributions. The ophthalmic nerve is uniformly
called the first or anterior nerve of the group.

In order to lay bare the medullary bundles as they issue to
make up the nerves, they must be freed from the connective
tissue so abundant about and between the roots where they
pass through the walls of the cranium and the ganglionic chamber.

In Bdellostoma five nerves supply the jaw apparatus. They
all leave the medulla cephalad of the ear capsule and, as the
snout region extends far in front of the brain, the course of most
of these nerves approaches the horizontal. From before, back-
ward, they are:

382 HOWARD AYERS

1. Nasalis

2. Ophthalmicus
3. Maxillaris

4. Mandibularis

5. Facial

Fig. 31 Ventral view of roots of V nerve of Bdellostoma, to show the relation |
of the root bundles of the nasalis, ophthalmic, maxillaris, and mandibularis
leaving the medulla.

The nasalis is the mesial nerve, the ophthalmic next, and so
on. The nasalis is therefore the most anterior nerve of the
group (figs. 29 and 36). It is the nasal nerve par excellence,
being largely devoted to the hollow nasal apparatus, both inside
and outside. The nerve supply to the structures of the outer
walls is largest, but perhaps not most important, for the nerves
to the structures placed inside the nasal capsule and tube

ORIGIN OF JAW APPARATUS 383

innervate not only the general mucosa, but a branch is supplied
to each olfactory plate which enters the olfactive membrane
along the dorsal aspect of the olfactory nerve bundle supplying

Fig. 32 View of the dorsal surfaces of the nasalis, ophthalmicus, and maxil-
laris roots of Bdellostoma seen from the ventral surface of brain. The roots
are dissected out and turned back, thus exposing their dorsal surfaces. The
root of the nasalis receives the mesial bundles from the medulla.

the plate. The nasal nerve sends an important branch to the
roof of the mouth, which palatine branch caused this nerve to
be named the palatine, and since it is not visible until it breaks

384 HOWARD AYERS

through the connective-tissue root sheath on the ventromesial
face of the larger maxiliomandibular nerve trunk, it has been
described as a branch of the latter.

A a=

J
>

cS

ZL
\) wi
}

VX
ay

Fig. 33 Ventral view of the left trigeminus of Bdellostoma, showing some of
the anastomoses of the root bundles and the manner of origin of the maxillaris
and mandibularis.

Most of the root bundles of the nasalis nerve arise (figs. 31
and 32) from the mesial part of the tip of the fork of the medulla,
cross ventrad of the root bundles of the ophthalmic and turn
sharply to enter the base of the nasalis root. Other bundles

ORIGIN OF JAW APPARATUS 385

issue direct from the medulla in line with the axis of the nasal
trunk, still another group of root bundles arise laterad of the
root and curve mesad to enter the root of the nasalis. The
nasalis is not a branch of any of the other roots, but a distinct
and important nerve certainly equal in morphological value to
the ophthalmic. Its position ventrad and sometimes laterad
of the ophthalmic is due to the crowding of the parts of the tri-
geminal complex into such narrow quarters that the fiber bundles
leaving both the mesial and lateral borders of the medullary
projection are both crowded ventrad on both edges, and curved
inward below the medullary lobe.

These root bundles unite into a cylindrical trunk which bends
at once mesad or toward the brain running cephalad along the
ventral border of the brain, nasal capsule, and nasal tube. It
gives off small twigs to the fat-body inclosing it, then a large
anastomosis to the ophthalmic near where the optic nerve passes
outward between the two. The optic nerve passes dorsad of
the nasalis, but ventrad of the ophthalmic, and the nasalis and
ophthalmic exchange fibers at the cephalic end of the ganglion,
and both give off a small number of fibers to the optic nerve
and its fat-body where it passes between them. It thus reaches
the nasal territory mesad of the ophthalmic and thus lies mesad
of the ophthalmic for the larger part of its course. The terminal
branches of the nasal division of the nerve in the anterior snout
region are mainly close to and upon the nasal tube. The pala-
tine division of the nerve ends in a brush of terminal branches
which supply the mucosa of the roof of the mouth, the median
tooth and from this region forward anastomose with the nerve
veil and network of the snout.

A bird’s-eye view of the nasalis nerve may be given as follows:
It is the mesial root and thus the most anterior nerve of the
jaw complex. It anastomoses with the ophthalmic nerve at
its root also where the optic nerve passes between them, both
giving off fibers which join the optic nerve trunk. It sends
branches: 1) to the fat-body covering the V root; 2) to the optic
nerve and fat-body; 3) to the posterior end of the nasal capsule;
4) to the lateral fat-body of the nasal capsule; 5) to the ventral

386 HOWARD AYERS

floor of nose, dorsal wall of the mouth, and to the hypophysial
canal; 6) to the fat-body on the opthalmic nerve; 7) to the sides,
dorsal and ventral faces of the nasal tube out to the tip of same;
8) to the roof of the mouth and nasal capsule and nasal tube;
9) five branches to connective tissue and mucosa of the roof
mouth and to the palatine tooth; 10) to the nerve net of the jaw
apparatus; 11) branches to each nasal fold, companions of the
branches of the olfactory nerve in the innervation of the sen-
sory olfactive epithelium.

The nasalis nerve is clearly the most anterior nerve of the
trigeminal complex for two reasons. In the first place, it retains
its own independent roots of origin from the central nervous
system which leave the medulla in part at least mesial to those
of the ophthalmic, and, in the second place, because the periph-
eral distribution of its fibers is to structures lying in general
mesad of those supplied by the ophthalmic nerve whose territory
lies laterad and consequently morphologically caudad to nasalis
territory. It carries motor fibers and innervates the nasalis
among other muscles. It is therefore a mixed nerve.

The terms nerve veil and nerve network refer to the nerve
structures of the distal portion of the jaw apparatus of Bdello-
stoma which represent the jaw plexuses of Amphioxus. The
characteristics of this plexiform structure will be given when we
come to the branchings of the jaw nerves in and about the snout,
buccal rim, and tentacles.

The term fat-body will be used to designate a variety of lip-
oid structures varying from the continous dermal sheet to large
and small, continuous and limited bodies about the nerves on
and between the muscles, to others attached to the connective
tissue sheets in the interspaces between the organs, about the
nasal capsule, the eye, and optic nerve. Usually they receive
branches from a passing nerve, with surprising frequency they
receive the whole of the terminal branches of relatively large
nerves, thus forming an end-organ. They are evidently often
more than accumulations of fat in the connective tissue. Am-
phioxus lacks such structures entirely, but they play an important
part in the organization of the nervous apparatus of Bdellostoma.

ORIGIN OF JAW APPARATUS 387

The roots of the ophthalmic nerve leave the brain, as shown
in figures 31, 32, laterad and dorsad of the mesial bundles of
the nasalis. They form the large nerve trunk which holds the
mesial position among the nerves Jeaving the tip of the medulla
seen from above, before the roots are dissected out. It soon
anastomoses with both the nasalis and the maxillary trunks
and runs forward diverging slightly laterad from the sagittal
plane. Near the level of the posterior end of the nasal capsule
it gives off a superficial branch, sometimes two branches, which
rises to the surface of the head just in front of the eye and enters
the subdermal lymph space and joins the complex of nerves
which ramify throughout the subdermal lymph space, in the
skin and over the muscles forming the floor of the lymph space.
Some of these ventral branches pass down between and also
through the muscles to join the coarse nerve net of the inter-
spaces and the loose connective tissue holding the muscle of
the mouth in place. Similar branches rise from the deeper
branches of the ophthalmic to join the complex and nerve net
in the subdermal space (figs. 34 and 35 B). On reaching terminal
territory the superficial branch breaks up into small nerves
which join the jaw plexus which covers the snout region hke
a fine-meshed veil (fig. 34).

The main trunk of the ophthalmic runs forward and branches
continuously to serve the muscles and other structures it passes,
and the branches to the skin and tentacles terminate in brushes
of fine fibers (fig. 835A). A large commissure of fibers passes
between the ophthalmic and mandibular nerves at A, figure
35. Altogether it is a remarkable bundle of fibers.

The maxillary nerve roots form a large nerve leaving the
dorsal lateral edge of the tip of the medulla as seen from above,
and its roots are drawn together from a wide territory in the
medulla (figs. 31 and 32). It is closely bound up with the man-
dibular nerve and the anastomoses between the two near their
origin from the medulla are extraordinarily extensive and com-
plex, as shown in figure 33. There is great variation in the
interchange of fiber bundles. I have selected the drawing shown
in figure 33 as on the whole typical of the relation of these

388 HOWARD AYERS

eA
fe AY

f
WA,
sie

ys

Fig. 34 Dorsal view of a dissection of the left ophthalmicus lateralis, VIIIa,
(buccalis) and VIIIb complex in the subdermal space and skin and the character
of plexus formation in the tentacular region of Bdellostoma. Only a part of this
complicated system is shown in the figure. Note the recurrent branches to the
aponeurosis and muscles at the anterior end of the nasal capsule.

ORIGIN OF JAW APPARATUS 389

ae

Fig. 35 A. Dorsal view of a dissection of some of the snout nerves of Bdellos-
toma. Forward extensions of the two trunks of the ophthalmic and the maxillary
nerves are shown. The branches of the latter to the second tentacle and the
skin between it and the first tentacle are drawn in more detail to show the tuft-
like terminals. At A is shown an anastomoses between ophthalmic and maxillary
nerves in the form of a large bundle of fibers. Near by is seen an exchange of
fibers in the form of a small bundle between the two branches of the ophthalmic.

B. Figure to illustrate the connection of the subdermal nerve net with deeper-
lying structures. Nerve branches descend through the muscle and rise from nerve
net and nerve trunks through the muscle to the surface. The arrangement re-
calls the association of parts found in the plexiform innervation of the jaw
apparatus of Amphioxus.

390 HOWARD AYERS

two nerves as they are being made ready for their peripheral
distribution.

As the drawing shows—and great care was taken to reproduce
accurately the number, relative size, and the course of the nerve
bundles present in this individual as seen from the ventral sur-
face—a large addition is made to the mandibular nerve bundles
from the fibers which leave the brain in the maxillary trunk.
Between x, which is a part of the maxillary trunk, and z, the
mandibular trunk, a thin band of fibers running apparently
from the latter to the former, is shown in the small insert at
the side. Two anastomoses with the ophthalmic trunk are
also shown. The method of branching to the dermal structures
is shown in figure 35. The maxillary nerve is also connected
up to the jaw plexus in the snout region. In figure 35, at D,
are shown four considerable branches which run from the trunk
direct to the skin. Besides innervating a number of muscles
of the jaw apparatus, it furnishes much material to the sensory
jaw plexus. It also innervates the velum by atleast two branches.
It is particularly the nerve of the distal portion of the jaw appara-
tus, both motor and sensory, sharing with the ophthalmic in
this control. It shares with the VII the control of the hyoidean
apparatus.

‘The mandibular nerve is not gathered into a single trunk in
any part of its course. The main part of it leaves the medulla
laterad of the maxilla, is closely applied to it and tied to it by
anastomoses.

Another part leaves the trunk of the maxillary nerve as a thin
band of fibers issuing from the dorsomesial surface near the
edge and wrapping around the ventral face of the trunk to run
laterad across it to join the main nerve, as shown in figure 35.
One division of the veloquadratus muscle is soldered to the
maxillary nerve at this place (fig. 35, v.q.). The mandibular
innervates the dentigerous jaw or basal portion of the ancestral
jaw bar, also the mandibular muscles, both retractors and pro-
tractors, and the muscles controlling the anterior part of the
hyoidean apparatus or the Jaw-supporting mechanism, as well
as supplying the skin structures of its territory. As figure

ORIGIN OF JAW APPARATUS 391

35 indicates, the fibers of the mandibular nerve are made up into
discrete bundles very soon after leaving the brain, which are
destined for distinct peripheral structures and run out to them
without further complications; 1.e., its main branchings occur
close to its origin.

From the foregoing sketch of the maxillary and mandibular
nerve trunks it is quite apparent that these nerves were present
long before the dentigerous jaws arose out of the Amphioxine
jaw apparatus and that they have been gradually transformed,
as the jaw apparatus evolved, into the characteristic nerves
of the maxillary and mandibular mechanisms of the higher verte-
brates, gradually assuming the condition and appearance of
nerves devoted almost exclusively to these important structures.
In Bdellostoma, as the mandibular mechanism has already been
established and the maxillary mechanism is still undifferenti-
ated, the nerves of the former show greater specialization for
the control of a motor mechanism than does the maxillary nerve.
Neither the maxillary nor the mandibular nerves are new
branches of old nerves, they are segmental trunks more ancient
than definitive gnathastome jaws.

The fifth member of the trigeminal complex is the so-called
seventh of vertebrate anatomy otherwise known as the N. faci-
alis. It connects with the lateral border of the medulla a short
distance behind the group-of four which leave the tip. After
viewing the crowded condition of the roots on the tip of the
medulla this interspace, bare of nerve trunks, is noticeable.
The manner of connection of the VII roots with the edge of the
medulla varies. Sometimes it appears as a single trunk; usually
it shows two roots, one motor root from the ventral edge of the
medulla and an acoustic root more dorsal. Occasionally a
third fine strand of fibers leaves the medulla cephalad of the
motor root and runs out some distance before it joins the nerve
trunk. However, these two or occasionally three external
roots do not indicate the internal complexity of its make-up.
Into its root enter fibers from six sources.

Its peripheral course is outward and backward around the
auditory capsule, as though. the latter by growing forward had

392 HOWARD AYERS

carried the nerve with it. After its curve around the anterior
end of the auditory capsule it runs ventrad and laterad, keeping
close for a time to the hyoid arch, and then it sweeps forward,
in a curve for a distance, on the surface of the muscles just above
the lateral border of the hyoidean plate to reach its final termina-
tion in the nerve veil about the fourth tentacle. In its course
it sends branches to several hyoidean muscles. As it crosses
the copulo-palatinus it sends a small branch dorsad into a small
muscle (fig. 36, VIZ m.) about an eighth of an inch long and thin-
ner than tissue-paper. This muscle lies closely applied to the
surface of the copulo-palatinus. Perhaps it is a vestigial muscle.
Its function is not apparent. I have found several structures
in Bdellostoma which are as non-conformist as this minute
muscle and will consider them in a later contribution.

This fifth segmental nerve, the most posterior of the group,
does not reach any part of the distal jaw apparatus other than
the fourth tentacle and its connections are mainly to the jaw
supporting apparatus. It is thus distinctively a hyoidean nerve.
It also does not innervate any of the muscles which move the
dentigerous jaw nor the mucosa in any part as far as my observa-
tions go. In this it differs from the homologue in fishes higher up.
It is postvelar and in Bdellostoma postmandibular. It has less
to do with the structures of the jaw bar than either the ophthal-
mic, maxillary, or mandibular nerves. Like the nasalis at the
anterior end of the series which has specialized in the service of
the olfactory apparatus, the facial at the posterior end of the
series has specialized in the service of the hyoidean apparatus.
It takes part in the formation of the nerve veil, sending its con-
tributions into the net near the fourth tentacle. The connection
with the ophthalmicus superficialis noted by P. Fiirbringer is
one of these net connections, and it is impossible, as I find it, to
yet determine what nerve elements the facial connects with,
as not only the ophthalmicus superficialis, but also the other
nerves of the jaw apparatus take part in the net building. The
nearest trunk to the facial in this locality is what Miss Worthing-
ton has called the nerve VIII a, and Allis the buccalis. This
nerve runs out below the eye, 1.e., posterior to it and conse-

ORIGIN OF JAW APPARATUS 393

quently ventrad of the ophthalmicus lateralis on the side of the
snout. It will take more study and different methods from any
yet applied to differentiate the elements entering the nerve veil of
the snout so that they may be followed back to the parent roots.

Having briefly described the origin and interconnections of
the roots of tle jaw nerves and their course to the periphery,
it is in order to explain how they intercommunicate and are tied
together at the periphery. This anastomosing of peripheral
branches of the main nerve pathways is apparently for the pur-
pose of shunting impulses by several paths toward the brain
-centers and permitting a diversified radiation of motor and other
impulses toward the periphery for one or several choices out of
the many possible combinations of peripheral reactions. This
veil is worthy of physiological study. Since time after time new
dissections have disclosed additional interconnections, I am far
from saying that the following account is complete. I present
these results of my dissections to serve as a basis of further study.
A general view of the peripheral relations may be given in few
words. The snout region of Bdellostoma in and under the
skin is veiled over by a wide-meshed network of anastomosing
branches of the nerves of the jaw apparatus (figs. 34, 35, 36).
This net is located in the skin and between the skin and outer
muscles of the head. It is most highly developed in the region
between the eye and the tentacles. It is connected with numerous
anastomoses, penetrating all parts of the snout, forming here
and there special net coverings, e.g., the network about the
anterior section of the velo-quadratus.

The nerves contributing branches to the network associated
closely with the eye and nose complex are:

. Nasalis

. Ophthalmicus profundus
. Ophthalmicus lateralis

. Acusticus a

. Acusticus b

. Mandibularis

. Maxillary group

. Facial

Noe

CoN Or Ww

394 HOWARD AYERS

What the distribution of these several elements through the
net is, has not been made out. It is notable that so many brain
centers have established connection with the sensory outposts
of the anterior end of the head, and that the muscles of the jaw
apparatus are closely tied up with this mechanism, as also are
the eye and nose. .

TO SUM UP THE NERVES

In Amphioxus the five nerves of the jaw apparatus arise seg-
mentally and evenly spaced from a long section of the central
nervous system and run to the walls of the buccal chamber, the.
jaw apparatus and its tentacles, the velum and the cheeks and
lips of the buccal region. Here they form nets or plexuses, the
most extensive of which surround the jaw bars and their muscles
and extend part way out on the tentacles. The nerves from the
right and left sides also anastomose freely. ‘Thus the parts of
the jaw apparatus and the walls of the buccal cavity are tied
together as a physiological or operating unit, and this extensive
peripheral intercommunication may be an expression of the lack
of a similar central mechanism and an effort to make good the
deficiency. ‘The jaw nerves are thus tied together peripherally
more intricately than we yet know them to be centrally. The
whole of the jaw apparatus is used in the work of food collec-
tion more or less passively.

In Bdellostoma the origins of the five nerves are crowded
together by the condensation of the long segment of the spinal
cord of Amphioxus into a short section of the spinal brain, the
medulla. They all leave a projecting lobe of this spinal brain
with their root bundles more or less intermingled, but still rec-
ognizable as separate nerves. They all run to the Jaw appara-
tus, which has been separated into two distinct parts, one of
which is put into active use in food gathering in a predacious
manner, and the rest of the skeleton of the jaw apparatus is
left for the passive work of a supporting skeleton for parts of
the mouth having little motion.

Arriving in the territory of the jaw apparatus, the nerves
intercommunicate by extensive anastomoses to build a plexi-

ORIGIN OF JAW APPARATUS 395

form veil and smaller nets with their connecting strands, en-
closing the structures in an Amphioxine plexiform nerve appara-
tus, the character of which leaves no doubt of its ancestral origin.

The labial cartilages of the cartilaginous fishes have attracted
the attention of anatomists from the time of Cuvier, who held
that they are related to the jaws and represent the maxillary
and intermaxillary, among other bones of the fish head. J.
Miiller held that they were aberrant structures and had no re-
lation to the jaw structures or any part of the mouth. Geg-
enbaur thought they were remnants of premandibular gill arches.
It has been an open field for guessing. The latter view seems
to be the one in favor, notwithstanding the fact that no traces
of gills or nerves that could belong to such body segments have
been. found.

Excepting Ammocoetes and the Myxinoids, the Chimaeroids
have preserved the Amphioxine jaw apparatus more completely
than any other of the cartilaginous fishes. Callorhynchus and
Chimaera serve to bridge over the gap between the Myxinoid
and what we are pleased to call the Gnathostomes. In figure
10 the relations of the parts of the skeleton of the jaw apparatus
of Callorhynchus are shown, and in figure 30 those of the related
Chimaera are displayed, both from the left side, for ready com-
parison with that of Bdellostoma (fig. 25). It will be noted that —
in Callorhynchus the mechanism is about as complete as in
Bdellostoma. It consists of two well-defined cartilaginous jaw
bars, marked 3, fastened to the hyoid below and just outside of
the mandible, 3b. They run forward and upward to the sides
of the nasal region, 4 and 8, where they are connected with
the tentacular cartilages grouped lateroventrad of the nasal
capsules. The tentacular cartilages of the third and fourth
tentacles of Bdellostoma, which are placed opposite the middle
and lower part of the buccal aperture (figs. 22 and 25), are here
located with the cartilages of the first and second tentacles
close about the nasal apertures, and they no longer extend out-
ward as slender tentacular rods but lie in flaps of the skin and the
large rostral snout. As in Bdellostoma, the distal part of the

396 HOWARD AYERS

fh

ay

Fig. 36 Composite picture to illustrate the amphioxine condition of the in-
nervation of the jaw apparatus of Bdellostoma. The nerves are sketched in over

a dotted outline of the major part of the head skeleton. These fundamental’

facts are illustrated: 1) The position of the buccal cavity (oral hood cavity)
between the buccal aperture and the velum. The innervation of the latter.
2) The persistence of an extensive amphioxine jaw apparatus composed of jaw
bars with the proximal dentigerous jaws disconnected for free action, and ten-
tacle bearing distal sections of the bars framing in the buccal aperture as in
Amphioxus. 3) The plexiform innervation of the tentacular portion of the jaw
apparatus forming the snout. 4) The enormously developed hyoidean jaw sup-

ORIGIN OF JAW APPARATUS 397

port. 5) The course of the five nerves controlling the jaw apparatus, forward
and ventrad, carrying motor and sensory fibers for the innervation of jaw ap-
paratus and its enclosed buccal cavity. 6) The branches of the nasalis innervat-
ing the olfactory mucosa together with other features of innervation acquired
since passing the Amphioxus stage, such as the lateralis nerves. 7) The caudad
displacement of the velar folds with the muscles which operate the velum re-
maining in their original territory near the mandibular nerve trunk.

The significance of the numbers is as follows: 1, terminal nasal cartilage;
2, olfactory lobe; 3, forebrain; 4, midbrain; 4, cerebellum; 6, medulla; 7, spinal
cord; 8, first spinal nerve; 9, eye; 10, olfactory branches of nasalis nerve.

398 HOWARD AYERS

jaw bar is separated from the mandible or basal part and has
the same attachment, viz., to the hyoid, not at the tip, but fur-
ther back, as shown at b, figure 10. This distal part is composed
of seven pieces of cartilage on each side of the head; the basal
piece is strongly attached by tendon to the dorsal edge of the
hyoid and bends upward and forward to connect midway to the
nose with the second segment, 4, the body of which is sickle-
shaped, curving forward and inward to its attachment to 5,
at the side and in front of the nasal capsule. Segments 3 and 4
span the distance between the ventral mandibular territory to
the nasal region and the maxillary territory. Segment 4 has
a prong which leaves the curve of the sickle about midway and
runs up, back and inwards, to attach at the side of the skull. Seg-
ments 6, 7, 8, and 9 are, respectively, the equivalents of the
first, second, third, and fourth tentacles of Bdellostoma. The
cornual and subnasal cartilages do not seem to be represented
as separate cartilages in Callorhynchus unless the cartilage of
the median nasal fold is the remains of the subnasal bar of Bdell-
ostoma. The long nasal tube of the latter is reduced to the
capsular parts, which in Callorhynchus are well developed as two
deep cartilaginous pockets set on either side of the median nasal
cartilage. Callorhynchus has in addition a large long median
dorsal snout cartilage which extends forward into the fleshy snout.
The two cartilages of the first and second tentacles extend into
the snout along either side of and somewhat ventrad of the
median snout cartilage.

The large hyoidean support cartilage lying below the mandible
in Callorhynchus was called by J. Miller a second lower jaw,
and he held it to be a peculiar cartilage not belonging in the
vertebrate plan of structure, in which category he also placed
the lower labials of Callorhynchus and other cartilaginous fishes.
G. B. Howes homologized the labials of Callorhynchus with
the mouth supports of Bdellostoma, but failed to see the relations
of these parts to the mandibular cartilages and did not under-
stand the morphological nature of this apparatus. His homol-
ogies are correct only in part.

The relation of the parts in Chimaera is shown in figure 30.
They are reduced in number in this species and as a whole fall

ORIGIN OF JAW APPARATUS 399

short of preserving the Myxinoid stage of theprimitive jaw appa-
ratus. They present us with a stage of reduction lying between
Callorhynchus and the Elasmobranchs, and the parts in Chimaera
are important for that reason.

The series of facts presented shows the genetic relationship
of the skeleton of the jaw apparatus step by step from Amphioxus
to Ammocoetes to Bdellostoma and to the Chimaeroids, which
are accepted Gnathostomes. It follows that the associated
structures, such as muscles, nerves and blood vessels are likewise
genetically related. As the nerves are dominantly associated
with the jaw apparatus, an examination of them should disclose
some indication of this genetic relationship, as after the skeleton
they are usually less changed during the course of phylogenetic
development than either the muscles or the vascular supply.
The results of such an examination of the nerves of Bdellostoma
have been presented in the foregoing pages. I believe that the
two conclusions given at the beginning of this contribution are
therefore fully supported by the evidence submitted.

It is also evident that the lower jaw or mandible is the first
part of the biting Jaws to be established. The maxillary struc-
tures arose in response to the stress and pressures developed
as the Bdellostomid mandible (which on first sight appears to
operate largely in a horizontal direction, but which develops its
main stresses while in a vertical position outside the mouth)
acquired relations such that these vertical stresses were developed
with the jaw inside the mouth. With the jaw in action, the
hyoidean apparatus of Bdellostoma takes up the vertical stresses
developed in the movements of the mandible, through its muscu-
lar and tendinous attachments to the skull above it, especially
the cartilaginous frame which includes the quadrate, palatine,
and other elements of the skulls of higher forms. But the hyoid-
ean stresses have not brought forth a definite maxillary anvil to
take the pressures of the mandibular hammer. They have,
however, brought forth a chondral condensation in the membranes
tying the hyoidean base plate to the skull. In the Chimaeroids
the skull has solidified into cartilage in a large way; only rem-
nants of the membranous skull remain, all the separate cranial

400 HOWARD AYERS

elements of Bdellostoma being fused together in a cartilaginous
unit.

A maxillary region is marked out by the presence of teeth and is,
however, limited behind by the articulation of themandible with the
base of the skull. The hyoidean base plate is still present in the
form of a large support plate below the mandible, and we are thus
supplied with an instructive intermediate stage between the
Myxinoids and the sharks in which the Amphioxine jaw appa-
ratus is still present in its modified Myxinoid form with a well-
defined mandible underlaid by the old time Myxinoid hyoidean
support. The parts of the distal section of the ancestral jaw
apparatus are not fused together nor are they fused with the
cranium. Butanew relation of parts has appeared—the mandible
is articulated to the base of the cranium, being tied to its ful-
crum by strong tendons. From this time on the distal section
of the jaw apparatus fades away, the hyoidean apparatus under-
goes reduction, the second mandible disappears and the newly
acquired maxillomandibular articulation becomes the seat of
stresses which call forth many different mechanical devices in
the effort to improve the biting jaws and to adapt them to special
conditions of food gathering and defense which are constantly
arising in the efforts of animals to adjust themselves to their
environment. The conditions of life become complicated and
survival more difficult and the differentiation of the biting Jaws
is an expression of this struggle. We thus see that Bdellostoma
has not lost definitive maxillary cartilages because they were
not developed in vertebrates until the Myxinoid stage was passed.
However, the fundamental cartilages of the head framework
with which the maxillary mechanism is associated and to which
they are more and more mechanically united as we ascend the
vertebrate scale are well developed. The pulls and stresses
set up in the skeletonous tissues by the movements of the head
parts have caused the appearance of cartilage and chondral
tissues in those localities most needing support and those which
form the foci of the mechanical stresses due to the pull of the
contractile tissues, until in Bdellostoma the basic cranial carti-
lages found in the vertebrate stock above the Myxinoids in great
and increasing variety and flux, have been built up into a head

ORIGIN OF JAW APPARATUS 401

skeleton of considerable extent; but as related to vertebrates
above the Myxinoids it represents only the early stages in the
ontogeny of their head skeleton. This is as one would expect.

Therefore the development of the craniofacial apparatus in
Bdellostoma is not an abnormal feature of vertebrate morphology
either from the viewpoint of: palingenesis or ontogenesis. It
is in harmony with the main current of cause and effect which
we call evolution of structure.

This is clearly shown by a comparison of the musculature,
nerve and vascular supply and the distribution of the connective-
tissue sheets and tendons, indicating the directions of the lines
of stress and the location of future structural changes, for where
stresses are developed new structural arrangements are evolved
and hard parts come into being to take up and transmit the
major stresses.

As the embryo of Bdellostoma is flattened between egg-shell
and food yolk, there is no possibility of development in full form
as occurs in the free embryo of Ammocoetes and larvae of Pet-
romyzon. ‘The jaw mechanism is flattened and vertically com-
pressed under the overgrowing brain, but the early larval stages
show that it grows cephalad and soon projects beyond the anterior
end of the brain. As the food yolk is absorbed and a place made
for the growing head, the parts expand dorsovertrally as well,
the snout region projecting into the space left free by the absorp-
tion of the yolk. The jaw mechanism passes through the Am-
phioxus and Ammocoetes stages, which are functional structures
in these animals in early embryonic life, by a series of rapid
transformations so that the structural characters of the ancestral
stages represented by Amphioxus and Ammocoetes, while marked
out, do not progress beyond embryonic tissues and by the time
definitive cartilage and muscle tissue appear the characteristic
structure of the Myxinoid jaw mechanism has been established.
There is apparently no remnant of a latent period, such as occurs
in the larval Petromyzon, even if such a larval state ever formed
a part of the life-history of the ancestral Myxinoids.

The jaw bars occupy the same relative position about the
mouth of the Bdellostoma embryo as in Amphioxus and Ammo-
coetes. From the basal part the jaw bars pass out and forward,

402 . HOWARD AYERS

tapering as they go, to meet in front of the mouth behind and
below the nasal aperture. The tentacles bud out from the bars.
They are four in number in nearly all stages after they appear
and it is uncertain if more are formed, although there are indica-
tions of two or three similar buds near the nasal end of the bars.
They do not develop far, even if-they are tentacular buds, and
soon disappear. The basal section of the jaw bar is broad and
the two attach to the hyoidean base plate in the earliest stages
I have dissected.

We thus find, even in early larvae, that the jaw apparatus
has the characteristics of the adult. It is desirable to carry
the study of the jaw structures back to their first appearance as
differentiated organs.

A few words about the brain.

That seetion of the spinal cord of Amphioxus from which the
five nerves of the jaw apparatus issue forms in Bdellostoma the
anterior part of the medulla. As a result of the growth of the
brain vesicle in the descendants of Amphioxus this medullary
territory has been formed out of the symmetrical spinal cord by
the pressure of the posterior part of the brain wall in such fashion
that the lateral walls of the spinal cord have been spread apart
with the extended and thinned dorsal wall stretched between their
edges. The ventral wall of the cord, being thick and resistant,
is not disrupted, remaining intact. The brain crowding back into
the V-shaped space thus formed has slid backward over the
ventral portion of the cord, leaving it unaffected. This process
can be observed in the ontogeny of many living forms. An
excellent series of figures illustrating the effects of the mechani-
cal stresses produced by the increase in volume of the forebrain
vesicle is given by Kerr for Lepidosiren. Of course, in the higher
vertebrates the process cannot be seen, or up to date has ‘not
been seen in full detail, as the nerves leaving the medullary
segments are already much crowded when recognizable.

Nevertheless, the medullary region is at first a cylinder and
is later split open on top to form the medullary V. In all cases
the trigeminal complex leaves the prongs of the V, which usually
form terminal lobes. At all times during this growth the brain
and cord are under tension, due to being inclosed in their en-

ORIGIN OF JAW APPARATUS 403

velopes and surrounded by the growing tissues of the body. The
spreading walls of the medulla meet with resistance to their
lateral travel, since they must displace other structures to make
room for themselves. Crowded back from in front by the growing
brain, they expand laterally against the resistance of the structures
laterad of them. Since in the growing organism, as elsewhere,
the forces of reaction are equal to the forces of action, the effect
of the lateral pressures will inevitably tend to compress the wall
of the medulla. We find evidence of this pressure in the relation
of the nerve roots to the tip of the medulla. If in the transforma-
tion of cord into medulla only the pressure of the brain in front
had been operative, the nerve roots would be found leaving the
edge of the arms of the Y on the same level and along a line curv-
ing about the rounded anterior end of each arm. However, we
find the arms have by lateral pressure been buckled upward and
their inner and outer edges, originally anterior and posterior
limits, bent ventrad, the inner edge curving laterad and the outer
edge curving mesad. The root of the nasalis (palatine) nerve
is thus forced below the inner edge of the arm and the ophthalmic
occupies the apical position on the edge of the tip of the arm. On
the outer edge of the arm we find the mandibular similarly curved
downward and appearing to arise from the ventral and lateral
surfaces of the arm. The maxillary trunk dominates the field on
the dorsal surface of the arm.

Up to the present only facts favorable to the view of the
genetic connection of the jaw apparatus of Amphioxus with that
of the Marsipobranch and through the latter with the true
Gnathostomes have been brought forward. Are there any
facts that do not harmonize? It would not bea problem awaiting
solution if there were none, and furthermore, when one can work
- out the solution of a problem in comparative anatomy and get
all the facts and leave nothing unexplained or unaccounted for,
the millennium will have arrived and comparative anatomy will
probably lack incentive.

Some investigators will raise the objection that the m. trans-
versus abdominis of Amphioxus is not homologous in its anterior
part with the circular muscles of the jaw and velar territory of
Ammocoetes, for example. I think they are, as I shall explain

404 HOWARD AYERS

in a subsequent contribution. But even though it could not
be shown that they are homologous, how much will that uncer-
tainty detract from the body of fact which shows that the more
or less longitudinal muscles of the jaw of Amphioxus arise out
of the transversus, that the transversus is innervated by the
nerves of the jaw apparatus, that in Ammocoetes these muscles,
although attached to the skull on both sides and continuous
across the midventral line, bear the same relations to the jaw bars,
are innervated in this territory by the nerves of the Jaw apparatus
(trigeminus), and further only differ from those of Amphioxus in
having developed two layers with skeletal tissue between them;
that in Bdellostoma they are split up into segments of circles
and have attachments to numerous cartilages, although they
still maintain the same fundamental relation to the jaw-bars
and continue to be innervated by the nerves of the Jaw apparatus?
In the face of such facts, what is a denial of their homology worth?

Take the old-time puzzle—the location of the nerves of Amphi-
oxus outside the myotomes and their submyotomic position in
all the other vertebrate forms. Even if we cannot here solve
this problem, does the fact that, at some time and somehow, the
muscle seems to have disappeared from under the proximal part
of the nerves of the jaw apparatus invalidate the evidence that
nerves supplying homologous structures in Amphioxus and
Bdellostoma are not themselves homologous? In other words,
does a non-essential fact destroy the value of an essential fact?

Other problems like these might be mentioned, but none seem
to me to constitute valid objections to the conclusions drawn from
the facts that have been presented, viz.: That the Amphioxine
jaw apparatus is the parent structure out of which has been
evolved the mandibular mechanism of the Gnathostome’ ver-
tebrates and its hyoidean companion mechanism. That to
branchial structures the jaw has no genetic relationship, while
to the trigeminus complex it bears the relation of end-organ.
The mystery of the labial cartilages dissolves when we recog-
nize their origin, follow their unfolding, obsolescence, and final
disappearance. .

Winding Way and Valley Road, Cincinnati, Ohio,
April 27, 1921

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THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 5
DECEMBER, 1921

Resumen por el autor, Halbert L. Dunn

El crecimiento del sistema nervioso central en el feto humano
expresado por medio del andlisis grafico y formulas
empiricas

El crecimiento del sistema nervioso central durante el periodo
fetal es semejante al aumento del peso del cuerpo durante esta
época. El crecimiento del sistema nervioso central demuestra,
mediante un andlisis, la existencia de tres subtipos diferentes de
crecimiento, a saber: 1) Crecimiento cerebral, el cual es lento y
continuo antes del sexto mes de la vida fetal (embrién de unos
30 em CH) y mas rapido desde dicha época hasta el nacimiento;
2) Crecimiento del tallo cerebral y de la médula espinal, mas
rapido desde el segundo hasta el final del quinto mes que en
épocas posteriores; y 3) Crecimiento del cerebelo, muy lento
desde el segundo hasta el sexto mes y sumamente rapido durante
los cuatro ultimos meses de la vida intrauterina.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, NOVEMBER 7

THE GROWTH OF THE CENTRAL NERVOUS SYSTEM IN
THE HUMAN FETUS AS EXPRESSED BY GRAPHIC
ANALYSIS AND EMPIRICAL FORMULAE

HALBERT L. DUNN

Department of Anatomy, University of Minnesota

THIRTY-EIGHT FIGURES

CONTENTS
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DU eHE ELON ee ers re MERI Seah AR i fed asage, Saja i Milde abe Gina Ree eK gars 407
Methodstomeollectimesdatanse: sss see osteo ae Pee et Sn ere ees Se 409
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Sala GLOW SCL Vb] ONS fae ek scm acct were PE yale, soy tree hy ies waked chsiccesh khan hv ene 417
Growth of the central nervous system as a whole.....................- 417
Crowcuromuecncepnalomeennns. here os yan ree tne hermes tale 419
Crowonombacwercorumisn tt): Sttleis Se Mes Pd: DR 423
Carnal ls THescebe me 25) sei Ss eh.)s cede os he AE pam beter Sletaere 2 hu 435
owas bie, ane ane: meduila ys soo k Shes cc a/c soo ke oe AAS 6G copied 442
GARONA An OLE AH) CxS Fogel aE Oe Sia gem aaa ate AE Rik ee mn Ne oie et eet et eee ea ys 447
Growmubrotethelspinalecords: ce aratritad Aas ey tee Male oe aa here 451
BP IGMSETOND e780 ce F de Sieh cis te Pa commons 8k ute iu) Fo 2 ban ced eke 456
Growth of the central nervous system as a whole...................... 458
SHETW DT TDE Weer My By RS a Ae 9 nay anne le DPN tear at Kis ke Bane a aaa can 488
INTRODUCTION

The growth of the human central nervous system has long been
a matter of interest and many studies on this subject have been
published, some of the earlier ones dating back almost a century.
These investigations have given us a general knowledge of the
growth of the brain as a whole from birth to maturity and have
furnished us some information regarding the modifications in
absolute and relative size of the various brain parts in early post-
natal life.

405

406 ‘ _HALBERT L. DUNN

Our knowledge of the growth of the central nervous system in
the prenatal period is much less complete. While the numerous
studies on the morphogenesis of the brain and spinal cord in the
first trimester of fetal life enable us to draw some general con-
clusions concerning the early prenatal growth of these structures,
there are practically no quantitative data available for this period.
Data on the later prenatal growth of the brain are more ex-
tensive, and a survey of the literature gives about 500 published
records of the weight of the brain between the third fetal month
and birth. The more important of these collections are those of
Rudinger (’77), Brandt (’86), Arnovljevicé (’84), Michaelis (’06),
Jackson (’09), and Valtorta (’09). A small amount of this ma- ~
terial has been presented in graphic form by Zangemeister (711).
The only collection of records of the weights or volumes of the
various parts of the brain in fetal life seems to be a short series
of figures on the cerebrum, cerebellum, and brain stem published
by Valtorta (’09).

The spinal cord has received far less attention than the brain,
doubtless because of its smaller size and the technical difficulties
of its removal. Our knowledge of the growth of the cord in post-
natal life is very incomplete, being limited to the data of Pfeister
(703) and Danielbekoff (’85). Information regarding the changes
in fetal life is apparently confined to a study by Giese (’98),
published as a St. Petersburg dissertation, which is available only
in abstract at the present time.

The following study was undertaken with the hope that a part
of this large gap in our knowledge of human growth might be
spanned by the systematic examination and measurement of the
brain and spinal cord in a large series of human fetuses ranging
from the second fetal month to birth, and by the treatment of
these data by some of the more modern methods of statistical and
graphic analysis. The work was done under the direction of
Dr. R. E. Scammon, to whom the writer is very much indebted
for constant advice and aid throughout the entire study.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 407

MATERIAL AND METHODS
Material

The material used in this study consisted of 156 human fetuses.
Fifty-four of these specimens were males, fifty-six were females,
and the sex of forty-six was unknown. The specimens ranged
from 3.1 to 53.6 em. in total or crown-heel length and were quite
evenly distributed between these extremes. At least one speci-
men was available for each centimeter interval of crown-heel
length and in many of the centimeter intervals below 30 cm. there
were three, four, or five specimens. All of the material was
selected from a much more extensive series of cases, with a view
to securing specimens in which the brains were well preserved;
however, a number of brains which were rather soft were used
for certain measurements which were not affected by this con-
dition. .

The great majority of the specimens were fixed in 10 per cent
formalin to which 1 per cent chromic acid had been added in some
instances, and all were afterwards preserved in 10 per cent forma-
lin. Five specimens of the entire series were fixed in Zenker’s or
Miiller’s fluid and were subsequently preserved in 70 per cent
alcohol. All of the material had been in the preservation for six
months or more.

The effect of formalin fixation upon the fetal brain must be
known in order to interpret the data correctly. It is unfortunate
that no comparison has been made between fresh and preserved
material. The fresh brain of the fetus is too friable for any deli-
cate, quantitative measurements to be taken upon it. Conse-
quently, fixation was prerequisite to the collection of the material
for this study.

The only checks upon the relationship of fixed and unfixed
material at our disposal consist of, 1) the determination of the
effects of formalin upon the size of the fetal body and, 2) the re-
lation of the curves of fresh brain weight to that of fixed brain
volume.

Several investigators have determined the effects of formalin
fixation on the fetus. Patten and Philpott (’21) record an

408 HALBERT L. DUNN

average increase in the crown-rump length of 4.8 per cent in a
series of twenty-two pig embryos which had been in 10 per cent
formalin for six months. Schultze (19) reports an average gain
in crown-rump length of 2.3 per cent in a group of eighteen human
fetuses which had been in 10 per cent formalin for nine months.
The average gain in head length and head breadth of the same
series was 1.1 per cent and 5.2 per cent, respectively. Calkins
(21) has found that the effect of fixation on the several diameters
of the head is very small and that formalin injection causes the
greatest part of this by a distention of the superficial tissues of
the scalp. The fetal cranium increases, therefore, very little in
size when it is fixed by formalin. The brain, on the other hand,
is supposed to gain greatly in volume. King (10) obtained a
percentage gain in weight of from 30 to 50 when the brains of
several white rats were subjected to formalin fixation. Such an
increase in brain weight or volume could take place in the fetus
only at the expense of the dural and subdural spaces, since the cir-
cumference and the diameters of the fetal head are but little
affected. However, these spaces were found to be quite large in
most of the preserved specimens in this series. The volume of
the brain, therefore, could not have been increased to any great
extent.

The relationship of the curve of fresh brain weight to that of
fixed brain volume can be compared readily by tables 2 and 3.
Table 2 shows the calculated volume of the central nervous system
for each 5 cm. C H interval. Table 3 sets forth the empirically
determined values for weights of fresh fetal brains collected from
the literature. In each respective C H interval the weight is seen
to be slightly greater than is the volume. This difference may
be caused in part by the formalin fixation, but the greater part
seems to be due to the fact that midsagittal sections of the brain
were made before volumes were determined and that intraven-
tricular fluid was lost by this procedure. On the whole, there-
fore, it is doubtful if there is any considerable change of the fetal
brain volume upon formalin fixation.

Five cases of the series were subjected to Zenker- or Miiller-
alcohol fixation instead of formalin. The effects of various

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 409

fixing fluids upon brain tissues have been determined by King
(10) and Hrdliéka (06). The work of King was done upon the
brain of the rat, and is analogous, in all probability, to the effects
of the fixing fluids upon the fetal brain. Calculated from her
findings, the absolute values of the five fetal brains subject to
Zenker- or Miiller-aleohol fixation would be 10 to 40 per cent
lower than the brains which were fixed by formalin. ‘Three of the
fetal brains in the present series which were fixed by the Zenker-
or Miiller-aleohol method lie in the C H interval of 5 to 10 em.
and two are in the C H interval of 10 to 15 cm. A glance at
figure 2 shows that these cases adhere closely to the central
tendency. Hrdlicka subjected the brains of twenty-seven mam-
mals and birds to 10 per cent formalin fixation. ‘Ten sheep brains
showed an average increase of 13 per cent after one month of
fixation; seven brains of birds an average gain of 20 per cent and
six brains of various mammals about 24 per cent. He left the
brains in 10 per cent formalin for eighteen months, and then
found that they had decreased in size to about 92 per cent of
their original weight.

From the results of these investigators as well as the data of
this series, it is obvious that 10 per cent formalin fixation has a
comparatively slight effect upon the volumetric determination of
the fetal brain.

Methods of collecting data

The magnitudes and proportions of the various parts of the
central nervous system were determined by several different
methods—by lineal measurements, by weighing and determining
the volume of each part, by making tracings of median sagittal
sections, and by photographing the lateral surfaces of the speci-
mens. The details of these methods and the descriptions of the
measurements taken will be considered separately.

Lineal measurements. All lineal measurements were made with
a steel vernier caliper which could be read accurately to 1 mm.
by the major and to 0.1 mm. by the minor scale. ‘They were
made with the brain and spinal cord in situ and always represented
the shortest distance between two given points.

410 HALBERT L. DUNN

Weight. The ponderal determinations were made on a scale
accurate to 0.01 gram. Before weighing in every instance the
brain part was stripped of its meningeal coverings and was placed
on a pad of dry gauze for 30 seconds to remove approximately the
same amount of surplus fluid in each case. Weights were always
taken upon preserved material.

Volumes. The volumetric determinations were made with a
special apparatus constructed according to a plan suggested by
Prof. L. W. Jones, formerly of the Department of Chemistry of
the University of Minnesota. A sketch of this device is shown in
figure 1. It consists of an iron chemical stand (A) to which a
clamp (B) is attached. This clamp holds a hard-rubber stopper
(C), to which is attached a slender wire (D) about 15 cm. inlength.
A small disk of mica about 1 cm. in diameter slides freely on this
wire. The wire is suspended in a jar or wide-mouthed bottle
(F) which has a spigot at its base and which is partially filled with
water. In using the apparatus the water in the bottle is drawn
off until the mica disk which floats on its surface touches the hair
line on the wire. The mass to be measured is then placed in the
bottle and the level of the fluid rises, carrying the disk with it.
The water is then drawn off into a beaker of known weight until
the mica disk has sunk again to the level of the hair line on the
wire. The beaker with the contained water is then weighed and
the known weight of the beaker is subtracted from the total, thus
giving the weight of the water displaced by the brain part.
After the proper temperature correction, this value can be con-
verted into a measure of volume. With practice and proper pre-
cautions, it is possible to determine small volumes quite accurately
with this apparatus. When working with small bodies, it is de-
sirable to use a bottle with a capacity of not over 30 to 40 ce.
A difference of approximately 0.1 cc. in volume can be determined
without trouble if a small container is used.

The lineal, volumetric, and ponderal determinations made were
as follows:

1. CH: Crown-heel length, or total body length; from the
vertex to the tip of the heel.

2. CR: Crown-rump length, or the sitting height; from the
vertex to the tip of the coccyx.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 411

3. FO: Fronto-occipital diameter; from the frontal pole to
the occipital pole of the cerebral hemisphere. The frontal pole
is defined as that point where the anterior, lateral, and inferior
surfaces of the frontal lobes meet. In large brains, in situ, this

Fig. 1 A figure representing the volumetric apparatus used in the collection
of data. A, iron standard; B, clamp; C, stopper; D, wire; E, movable isinglass
disk; F, bottle with spigot. The arrow indicates a hair line upon the wire.

point is adjacent to the crista galli. The occipital pole is defined
as that point where the posterior, lateral, and the interior sur-
faces of the occipital lobes meet.

4. TT: Temporal diameter; from the right temporal pole
to the left temporal pole of the cerebral hemispheres. The
temporal pole is defined as the most lateral point on the anterior

412 HALBERT L. DUNN

projection of the temporal lobe. This pomt becomes less defi-
nite in large fetuses, but can still be recognized with sufficient
accuracy.

5. FS: Frontospinal length; from the frontal pole of the
cerebral hemisphere to the point of origin of the first cervical
nerve. This measurement was taken to the middle of the body
of the first cervical vertebra, for it was sometimes difficult to
identify the first cervical nerve because of error in securing exact
midsagittal sections of the brain and the spinal cord.

6. SC: Spinal cord length; from the point of origin of the
first cervical nerve to the tip of the conus medullaris;.the meas-
urement was made with cord in situ.

7. BC: Brain and cord length; the sum of S C (6) and
FS (5).

8. PL: Pons length; the greatest inferior length of the pons.

9. CL: Colliculi length; from the most anterior point of the
superior colliculus to the most posterior point of the inferior
colliculus.

10. VL: Vermis length; the greatest length of the vermis
cerebelli parallel with the axis of the pons and medulla.

11. VH: Vermis height; the greatest height of the vermis
cerebelli perpendicular to the long axis of the pons and medulla.

12. VSC: Volume of the spinal cord; cord taken from the
first cervical nerve to the tip of the conus medullaris and freed
of meninges and nerve roots. |

13. VPM: Volume of the pons and medulla; pons and
medulla cut posteriorly at the first cervical nerve and anteriorly
just in front of the pons, meninges removed.

14. V MB: Volume of the midbrain; the midbrain was cut
just in front of the pons posteriorly and just in front of the
superior colliculi and through the pedunculi cerebri behind the
mammillary bodies anteriorly, meninges removed.

15. VC: Volume of the cerebellum; the velum medullaris was
severed anteriorly and posteriorly to free the cerebellum from the
pons and medulla, meninges removed.

16. VRH: Volume of the right hemisphere, meninges re-
moved.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 413

17. V LH: Volume of the left hemisphere, meninges re-
moved.

18. V BH: Volume of both hemispheres: computed by adding
V RH (16) and V LH (17).

19. V EB: Volume of the entire brain or encephalon computed
by adding 13, 14, 15, 16, and 17. This method was found to be |
more accurate in dealing with the soft brains than to take the
actual volumes of all the parts, because of the possible loss of
brain tissue in handling.

20. VCNS: Volume of the central nervous system; found
by adding the volume of the spinal cord (12) to the entire brain
or encephalon volume (19).

The ponderal determinations corresponding to the volumetric
measurements were taken and are included in table 41.

Methods of treating data

While the accuracy of data of the kind here presented depends
upon the nature of measures employed and the care with which
these measures are determined, their significance can only be
ascertained by the use of a variety of methods of graphic and
numerical analysis. A number of these methods have been used
in the present study. They may be arranged in order of pro-
cedure in five main groups, namely: 1) Construction of field
graphs with points of central tendency and establishment of pre-
liminary curves by inspection. 2) Reduction of these prelimi-
nary curves, as established by inspection, to empirical fomulae
based on total body length, and construction of primary data
tables. 3) Calculation of values at 5-cm. intervals of body
length, calculation of absolute and percentage increments for
each 5-cm. interval of body length, calculation of newborn ratios
for each 5-cm. interval of body length, and expression of these
values in the graphic form. 4) Conversion of the various values
as determined by empirical formulae from functions of the body
length to functions of the age in fetal months by interpolation on
the basis of Mall’s (712) tables of crown-heel length in fetal life.
Determination of the monthly percentage and absolute increment
on the basis of these conversions; graphic expression of these

414 HALBERT L. DUNN

values. 5) Determinations of the relative weights of the various
parts of the central nervous system as compared to that of the
encephalon. These several methods of analysis will now be con-
sidered in more detail.

1. Construction of field graphs and establishment of preluminary
curves by inspection. The data secured for each measurement of
the central nervous system were first plotted on a field graph in
which the length of the body (C H) was used for the abscissa and
the measurement in question for the ordinate. The material was
then divided into classes on the basis of 5-em. intervals of crown-
heel length and the arithmetic mean or average and the median
were determined for each of these classes. ‘These means were then
indicated in the graphs by special symbols, their exact position
being determined by weighting for the distribution of the cases
according to crown-heel length in the 5-cm. intervals. A pre-
liminary curve was then drawn by inspection on the basis of the
combined evidence furnished by the position of the arithmetic
mean, the median, and the general distribution of the cases. This
method has distinct advantages over the practice of establishing
curves of inspection on the basis of the arithmetic mean alone,
particularly in those instances where a curve is rising rapidly in
a single interval or where the cases are not regularly distributed.
The use of both the arithmetic mean and the median enables
one to correct for chance deviations in the mean alone with con-
siderable confidence, particularly when the cases are all spread
before one on the field graph.

2. Reduction of preliminary curves of inspection to numerical
expression by means of empirical formulae. The curve of each
value as determined by inspection was reduced to numerical ex-
pression in the form of an empirical formula on the crown-heel
or total length of the body expressed in em. This procedure in
no way increases the reliability of the curve as determined by
inspection nor does it increase the accuracy of the points upon
which it is based, but it has several important advantages, for it
permits an accurate and abbreviated expression of the curve,
facilitates exact interpolation and the conversion of values into
different scales and forms of expression, and, in some cases, aids
in classifying the curves as to form.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 415

The empirical formulae expressing the curves obtained from the
material are of several types. As certain of the lineal dimensions
of the brain are straight lines when plotted against crown-heel
length, they may be expressed by the formula:

Dei AL)
where Y is the value in question expressed in em., X is the crown-
heel length in cm., and a and 6 are empirically determined con-
stants.

The growth of other lineal dimensions of the central nervous
system is not so simply related to the lineal growth of the body
as a whole and the expression of this relation is more complex.
But all may be represented by formulae having the general form:

Vo (aXe ae
where Y is the value in question (in em.), X is the crown-heel
length of the body (in cm.), and a, b, and c are empirically de-
termined constants, b being a fractional exponent. Curves cor-
responding to formulae of this type may also be expressed fairly
satisfactorily in the logarithmic form:
Y =aX +blogX +c

which will be recognized as the formula which Hatai employed
with great success in the study of the growth of the various organs
of the albino rat and which is regarded by him as of fundamental
importance as exemplifying the application of the law of Mau-
pertuis to the process of growth. However, the exponential form
has been found the more convenient for application in the pres-
ent work.

The formulae for the expression of the inspected curves of
volumes of the various parts of the central nervous system have
as their simplest form:

ake
which is modified in certain instances to:
GX) Pear

and in others to:

Y =d[(aX)® +¢|
In these formulae Y is the value in question in ec., X is the crown-
heel length in cm., and a, b, c, and d are constants, b being always
an exponent with a value greater than 1.

416 HALBERT L. DUNN

Having obtained calculated values for the different measure-
ments under consideration, tables of each dimension were drawn
up in some detail. ‘These are tables 1, 3, 7, 9, 11, 13, 15, 17, 19,
21, 23, 25, 27, 29, 31, 33, 36, 38 of the present publication. In
each of these is given the crown-heel length in 5-cm. intervals,
the average crown-heel length, the maximum, minimum, and
arithmetic mean (average) of the considered measurement in the
interval, the calculated mean for the same measurement, and the
absolute difference between the calculated and the observed
means. Both the calculated and observed means are recorded
to the nearest millimeter for length and to the nearest 0.1 cc. for
volumes in these tables.

3. Calculation of secondary tables. Secondary determinations
made on the basis of the above calculations are included in tables
2, 4, 5, 6, 10, 12, 14, 16. These include the absolute value of
each measurement, as determined by calculation with the em-
pirical formula, for each 5 em. of body length from 5 to 55 em.,
and the percentage and the absolute increment of these values
for each 5-cm. interval between these points.! Included in these
tables is also the ratio of the calculated value of the measurement
at each 5 cm. of the total body length to the calculated value of
the new-born. It was assumed that the value at 50 cm. of crown-
heel length represented the new-born and the calculated values
for the 5-cm. points below 50 are expressed in per cents of this
one. The reduction of values to this common scale is of great
convenience, since it permits their direct comparison regardless
of their absolute magnitudes.

4. Conversion of the various values from functions of the crown-
heel length to functions of the age in fetal (lunar) months. This was
done by calculating the values by means of the empirical formulae

1The percentage increment was determined in the customary manner. The
absolute value at the beginning of an interval is subtracted from the absolute
value at the close of the same interval. The result is divided by the absolute
value at the beginning of the interval and the quotient thus obtained is mul-
tiplied by 100 in order to reduce it to a percentage basis. The percentage in-
crement is the only simple form of expression of relative growth now in use which
is at all satisfactory. However, it gives only a rough approximation of the actual
rate of growth in a given interval.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 417

from Mall’s tables of the relation of crown-heel length to age in
prenatal life (Keibel and Mall, Human Embryology, vol. 1, p.
199). The results obtained by this procedure are not to be
regarded as final, for the exact relation between body length and
age in fetal life is still open to question. They are included, how-
ever, because they give, in a rough way at least, some concept
of the relation of the growth of the central nervous system to
time in the prenatal period and because they aid in the inter-
pretation of its percentage increment. Since Mall’s table stops
at 270 days, it has been extended by graphic exterpolation to
ten full lunar months (280 days) and the length at this age (51.5
em.) was considered as the norm for the new-born in this type
of calculation. The monthly percentage increments of the prin-
cipal volumetric determinations were calculated secondarily from
the values obtained in this manner. They are shown in table 42.

5. Determination of the relative weights of the various parts of the
central nervous system in terms of per cents of the encephalon is an
obvious procedure which needs no particular description. The
results obtained are shown in table 40.

SUMMARY OF OBSERVATIONS
Growth of the central nervous system as a whole

The curve of the absolute volume of the central nervous system
(fig. 2), when based upon crown-heel or total body length, is con-
cave like all other curves of volume growth of the fetal organs.
It may be expressed by the empirical formula:

Central nervous system volume (cc.) = [0.114 (C H length
em.) ]*-34 + 2.0

The absolute volume of the central nervous system is about
7 ec. at 10 to 15 em. (CH). This increases steadily to 36 ce.
at 25 em. (C H), and is then deflected upward sharply to reach
3a/ cc. at 50 cm. (CH).

When calculated according to age in fetal months, the absolute
volume of the central nervous system (fig. 34, curve I) is found to
be 2 cc. at the beginning of the third month, rising to 36 ce. at the
beginning of the sixth month, and to about 340 ce. at birth.

418 HALBERT L. DUNN

The rate of growth of the volume of the central nervous system
as determined by the 5-cm. interval percentage increment
(page 413), (fig. 26, curve I) is approximately 120 per cent for the
10-to-15-cm. interval and descends gradually to about 40 per
cent for the 50-to-55-cm. interval.

TABLE 1
Volume of the central nervous system

Formula: Volume of the central nervous system (cc.) = [0.114 (crown-heel
length em.)]3-34 + 2.0

(146 cases)

ORSERVED VOLUME OF
CENTRAL NERVOUS SYSTEM

DIFFERENCE
BETWEEN
OBSERVED AND
CALCULATED

CROWN-HEEL LENGTH CALCULATED
VOLUME OF
CENTRAL NER-

VOUS SYSTEM

NUMBER
OF CASES

Range Mean saan ats Mean MEANS
cm. cm. cc. ce. cc. cc. cc.
Oto 5 3.9 0.7 0.1 0.3 4
5 to 10 ez 2.4 0.6 1.3 12
10 to 15 12.2 8.6 2.1 5.4 5.0 —0.4 21
15 to 20 16235) 2220 3.0 12.5 10.8 —1.7 16
20 to 25 22.6} 49.5 17.4 ole 25.6 —5.5 19
25 to 30 Patt Ak 65.4 | 24.7] 46.9 45 .3 —1.6 12
30 to 35 382.8 | 144.2 | 45.8 | 81.8 83.9 +2.1 18
35 to 40 37.2 | 222.8 | 102.2 | 141.4 126.7 —14.7 iff
40 to 45 42.2 | 256.9 | 145.3 | 197.7 187.6 —10.1 12
45 to 50 48.0 | 412.5 | 207.0 | 306.0 294.0 —12.0 7
50 to 55 52.2 | 414.3 | 332.3 | 367.5 388 .5 +21.0 8
TABLE 2

Calculated volume of the central nervous system at 5-cm. intervals of crown-heel length

VOLUME OF CENTRAL INCREMENT IN EACH

CROWN-HEEL LENGTH

NERVOUS SYSTEM

5-cM. INTERVAL

cc.

4.4
a
17.3
27.8
40.7
57.4
76.4
99.5
125.4

per cent

122.0
121.2
97.6
79.4
65 .0
50.4
47.5
41.9
37.2

RATIO TO NEW-BORN

per cent

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 419

The percentage increment, when calculated for time, is about
600 in the fourth fetal month. This drops to 185 in the fifth
month and to 22 per cent in the tenth month.

a SE ae ae OS Ge Oma”. OR Oa

Fig.2 Field graph and curve of the growth of the central nervous system in
fetal life. Abscissa: total body length in cm. Ordinate: central nervous system
weight and volume in grams (cc.). Individual cases are indicated by solid dots
(for weight) and by circles (for volume). Curve drawn to the formula: Y =
(0.114X)334 + 2.0. (Data from tables 1 and 2.)

Growth of the encephalon

The curve of the absolute volume of the entire brain (fig. 3)
when based on crown-heel length is a concave curve expressed by
the empirical formula:

Encephalon volume (cc.) = (0.125 (C Hlength cm.))#6 + 1.5

420

HALBERT L. DUNN

The absolute volume of the entire brain when based on crown-

heel length (fig. 3) is about 3.5 cc. at 10 em. (C H).

This in-

creases to 38 ec. at 25 em. (C H) and then more rapidly to 330 ce.
at 50 em. (C H).

Encephalon volume

TABLE 3

Formula: Encephalon volume (ce.) = (0.125 crown-heel length em.)*16 + 1.5

CROWN-HEEL LENGTH

(144 cases)

OBSERVED VOLUME

OF ENCEPHALON

Range Mean
cm. cm.
Oto 5 3.9
5 to 10 (POP
10 to 15 ES
15 to 20 16.9
20 to 25 Pass
25 to 30 20 1
30 to 35 32.2
35 to 40 37.2
40 to 45 42.8
45 to 50 48.0
50 to 55 52.2

Ne

Neeey
TE ARANORMOS §
mM CO WAT ORO ON

FON OnNNOO
DONWNOO De

ie
oOrPN

TABLE 4

- CALCULATED
VOLUME OF
ENCEPHALON

DIFFERENCE
BETWEEN
OBSERVED AND
CALCULATED

MEANS

cc.

NUMBER
OF CASES

Calculated volume of encephalon at 5-cm. intervals of crown-heel length

CROWN-HEEL LENGTH

VOLUME OF THE

ENCEPHALON

cc.

3.0
8.4
19.6
38.1
66.6
107.6
163.3
236.1
328.9
444.0

INCREMENT IN BACH 5-cM.
INTERVAL OF
CROWN-HEEL LENGTH

cm.

per cent

140.0
133.5
94.4
74.8
61.5
51.8
44.5
09.4
30.0

RATIO TO NEW-BORN

per cent

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM

When calculated to age in fetal months (fig. 34, curve I), the

volume of the entire brain is about 1 cc. at the beginning of the
third month. It increases to 14 ce. by the first of the fifth month,

and reaches the value of 330 cc. by birth.

The rate of growth of the encephalon when determined by
5-em. percentage increment (fig. 26, curve II) is approximately

Calculated weight of the encephalon (based on collected data) at 5-cm. intervals of

CROWN-HEEL LENGTH

CROW N-HEEL LENGTH

Range

cm,

Oxtoms
5 to 10
10 to 15
15 to 20
20 to 25
25 to 30
30 to 35
35 to 40
40 to 45
45 to 50
50 to 55

TABLE 5

crown-heel length

Tae re | eee ae | saara ne manor

grams grams per cent per cent

3.3 0.8

9.4 6.1 185.0 yal

PPA AL PAT 135.0 5.6

44.0 21.9 97.5 1 4

77.8 33.8 76.8 19.8

126.6 48.8 62.7 Bye

193.3 66.7 52.7 49.1

281.0 87.7 45 .4 71.4

392.9 111.9 42.4 100.0

pyAai 139.2 35.4 163565)

TABLE 6
Volume of the encephalon (data of Jackson, ’09)
(31 cases)
ENCEPHALON VOLUME
Mean Maximum Minimum Mean
cm. cc. cc. cc.

1.8 0.4 0.25 0.18
Ue! 3.6 2.0 2.83
ee 20.0 10.0 1340
WZ 40.0 18.0 22
22, A 60.0 30.6 45.9
27.4 129.0 HZ ea 85.4
S208 126.7 120.0 123.3%
38.5 140.0 140.0 140.0
44.2 BPA A es) 321.3
45.9 358 .0 278 .2 322.6
51.9 385 .0 363 .0 374.0

NUMBER
OF CASES

NONrRre NW Or OW W &

422 HALBERT L. DUNN

140 per cent for the 10-to-15-cm. interval. It descends rapidly
to 75 per cent of the 25-to-30-cm. interval, and then more
slowly to 40 per cent in the 50-to-55-cm. interval.

The percentage increment of the entire brain volume when
calculated against age in fetal months (fig. 28, curve VI) is about
580 per cent in the fourth month, descending to 22 per cent in
the tenth month.

Ort 5) 0p ee ON SIO 05S SAO Bam.

Fig. 3 Field graph and curve of the growth in weight and volume of the en-
cephalon in fetal life (the curve of growth taken from brain weights reported in
the literature). Abscissa: total body length in cm. Ordinate: entire brain
weight and volume in grams (cc.). Individual cases are indicated by solid
dots (for weight). J, curve of encephalon volume drawn to the formula:
Y = (0.125X)316 + 1.5.——== (Data from tables 3 and 4.) JJ, curve of
encephalon weight based upon data from the literature and drawn to the form-
Was Yi (ONS A Ee = RO n oe) ie eeeee (Data from tables 5 and 6.)

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM

Growth of the cerebrum

423

The growth of the cerebral hemispheres is portrayed by figures
3 to 9, inclusive, and figures 37 and 38, and is shown numerically

in tables 3 to 16, inclusive.

formula:

It may be expressed by the empirical

Volume of the cerebrum (ce.) = (0.12 (C H length cm.))*19

TABLE 7

Volume of the right hemisphere
Formula: Right hemisphere volume (cc.) = (0.1 crown-heel length (cm.))*1
(115 cases)

CROWN-HEEL LENGTH | RIGHT HEMISPHERE VOLUME

Range

cm.

Oto 5

5 to 10
10 to 15
15 to 20
20 to 25
25 to 30
30 to 35
35 to 40
40 to 45
45 to 50
50 to 55

Mean

cm.

12.5
16:9
22.6
27.1
32.6
37.2
42.5
48 .0
52.2

Maxi- Mini-
mum mum
cc. cc.

4.2 1.6
10.6 2.1
21.8 9.3
32.7) 12.4
eb 218

109.3 | 48.7
126.6 | 67.2
191.5 | 102.9
185.3 | 148.0

Mean

cc.

CALCULATED

RIGHT

HEMISPHERE

VOLUME

cc.

MEANS

cc.

DIFFERENCE
BETWEEN
OBSERVED AND
CALCULATED

NUMBER
OF CASES

Calculated volume of the right cerebral hemisphere at 5-cm. intervals of crown-heel

CROWN-HEEL LENGTH

VOLUME OF RIGHT
CEREBRAL HEMISPHERE

2.6 2.0 —0.6 12
6.0 5.2 —0.8 14
14.2 12.8 —0.4 19
21.8 22st +0.9 12
39 .2 40.4 ale 17
65.8 61.1 —4.7 16
94.6 92.6 —2.0 10
139.0 135.6 —3.4 a
166.0 176.4 +10.4 8
TABLE 8
length
ja eens ee RATIO TO NEW-BORN
cm. per cent per cent
0.6
2.6 260.0 2.3
5.2 144.0 Dds
8.8 100.0 11.4
13.5 76.6 20.2
19.4 62.3 32.7
25.9 51.2 49.5
34.4 45.0 71.8
43.4 39.2 100.0
53.0 34.7 134.5

424 HALBERT L. DUNN

The absolute volume of the right and of the left hemispheres
(figs. 4 and 5) shows no marked differences—the minor distinctions
between the two falling within the margin of experimental error.

aaa eemee
O 5 10 15 20 25 30 35 AO A5 50 55cm

Fig.4 Field graph and curve of the growth in weight and volume of the right
cerebral hemisphere in fetal life. Abscissa: total body lengthinem. Ordinate:
right hemisphere weight and volume in grams (cc.). Individual cases are in-
dicated by solid dots (for weight) and by circles (for volume). Curve drawn
to the formula: Y = (0.1X)*18, (Data from tables 7 and 8.)

The absolute volume curve of the hemispheres shows the same
concave form as the volume of the central nervous ststem (fig. 2).
At 15 em. (C H) the hemispheres are 1.8 ce. in volume, while in

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 425

fetuses of 25 to 30 em. (C H) they have reached an average volume
After this time the volume curve increases more
rapidly, attaining the average of about 300 cc. at 50 em. (C H).

When examined in relation to age (fig. 34, curve II) the ab-
solute volume of the right cerebral hemisphere is found to be 2 ee.

of about 32 ce.

CROWN-HEEL LENGTH

Range

cm.

Oto 5

5 to 10
10 to 15
15 to 20
20 to 25
25 to 30
30 to 35
35 to 40
40 to 45
45 to 50
50 to 55

TABLE 9

Volume of the left cerebral hemisphere

Formula: Volume of the left cerebral hemisphere (cc.) = (0.105 crown-heel
length (cm.))*%

OBSERVED VOLUME OF THE

LEFT HEMISPHERE

Maxi-

Mean mish
cm. ce.
12.5 4.4
16.9} 10.5
22.4 | 20.0
Phd | 3020
32.8 | 50.0
37.2 | 89.6
42.5 | 110.7 |
48.0 | 188.5 |
52.2 | 192.0

1.3
2.3
10.2
13.0
21.0
46.8
67.0
98.5
142.0

Mean

ce. ce

2.6 2.3

6.0 sac
14.8 13.5
21.7 24.0
35:2) 429
64.1 62.9
92.2 94.4
138.0 136.6
170.0 176.2
TABLE 10

CALCULATED
VOLUME OF
THE LEFT
HEMISPHERE

DIFFERENCE
BETWEEN
OBSERVED AND
CALCULATED

MEANS

cc.

—0.3
—0.3
—0.3
+2.3
+4.7
ee
+2.2
al:
+6.2

NUMBER
OF CASES

Calculated volume of left cerebral hemisphere at 5-cm. intervals of crown-heel length

CROWN-HEEL LENGTH

VOLUME OF LEFT
CEREBRAL HEMISPHERE

200.3

INCREMENT IN EACH
5-cM. INTERVAL

cc.

2.8
5.6
9.2
14.0
1957
25.8
32.9
43.7
45 .4

per cent

233 .0
140.0
96.0
74.5
60.0
49.1
42.0
39.2
29.3

RATIO TO NEW-BORN

OF CROWN-HEEL LENGTH

per cent

0.8
2.6
6.2
12.1
21.2
33.9
50.6

\

426 HALBERT L. DUNN

at the beginning of the third month, and this rises at first slowly
and then rapidly to about 150 cc. at birth.

aa 0 i) aes ies 05.40.40. 450. SO nee

Fig.5 Field graph and curve of the growth in weight and volume of the left
cerebral hemisphere in fetal life. Abscissa: total body length in cm. Ordi-
nate: left hemisphere weight and volume in grams (cc.). Individual cases
indicated by solid dots (for weight) and by circles (for volume). Curvedrawn
to the formula: Y = (0.105X)* 4. (Data from tables 9 and 10.)

The percentage increment of the hemispheres (fig. 26, curve VI)
approximates that of the entire brain. It is about 265 per cent
for the interval between 10 and 15 cm. (C H) length and steadily
declines from this value until it reaches the level of 35 per cent
for the 50-to-55-em. (C H) interval (birth).

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 427

The percentage increment of the hemispheres, when calculated
by fetal months (fig. 28, curve V), shows an approximate rate
of growth of 600 per cent in the fourth fetal month. This rate
descends rapidly at first, and then more slowly to about 22 per
cent in the tenth fetal month.

TABLE i1
Volume of both cerebral hemispheres

Formula: Volume of both cerebral hemispheres (cc.) = (0.12 crown-heel length
(em:))*2"
(115 cases)

OBSERVED VOLUME OF
BOTH HEMISPHERES

DIFFERENCE

CROWN-HEEL LENGTH
BETWEEN

CALCULATED

VOLUME) OF 1) soe an AD NUMBER
Range Mean ee Mint Mean HEMISPHERES ee ae Ween
cm. cm. cc cc. cc cc. cc
Oto 5
5 to 10
10 to 15 12.5 8.6 2.2 5.2 3.65 —1.55 12
15 to 20 16.9 | 20.5 AS5|l2ez 9.5 —2.7 14
20 to 25 22.4) 46.2 18.0 29.4 22.9 —6.5 16
25 to 30 Piel 62.5 25.4 | 46.6 43.0 —3.6 12
30 to 35 SZEON| Lode eoOs om Oe. 79.1 +3.0 18
35 to 40 37.2 | 210.8 95.5 | 131.4 118.2 —13.2 17
40 to 45 42.3 | 239.3 | 184.2 | 184.8 178.1 —6.7 il
45 to 50 48.0 | 380.4 | 201.2 | 277.0 266.5 —10.5 a
50 to 55 52.2 | 377.3 | 302.0 | 334.4 348.3 —13.9 8
TABLE 12

Calculated volume of both cerebral hemispheres at 5-cm. intervals of crown-heel length

VOLUME OF BOTH
CEREBRAL
HEMISPHERES

INCREMENT IN EACH

5-CM. INTERVAL RATIO OF NEW-BORN

CROWN-HEEL LENGTH

ce.

1.79
6.52
16.33
33.27
59.51
97.31
149.0
216.9
303.6
411.2

cc.

4.73
9.81
16.94
26.24
37.80
51.69
69.7
86.7
107.6

per cent

264.0
150.0
103.0
78.8
63.5
53.2
45.5
40.0
34.5

per cent

onoonne
me & OO Re SD

Hy Oo et et

A428 HALBERT L. DUNN

The per cent which the volume of both hemispheres forms of
the entire brain volume (fig. 7) reflects the relative growth of the
other brain parts. The cerebral hemispheres form only from

7A

JO

0 eet F - — - :
O 5 10 15 40) 05 SO oD AO AD5 50 55cm.

Fig. 6 Field graph and curve of the growth in weight and volume of both
cerebral hemispheres in fetal life. Abscissa: total body length in em. Ordi-
nate: hemispheres weight and volume in grams (ce.). Individual cases indicated
by solid dots (for weight) and by circles (for volume). Curve drawn to the
formula: Y = (0.12X)*19. (Data from tables 11 and 12.)

88 to 90 per cent of the entire brain volume in the third fetal
month. ‘This proportion increases rapidly to between 94 and 95
per cent during the sixth fetal month, but after this time it de-
scends to about 91.5 per cent at birth. This decline is due
mainly to the tremendous relative growth of the cerebellum.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 429

The two linear measurements of the cerebral hemispheres (the
fronto-occipital and the temporal diameters) also furnish some
information concerning the growth of this part of the brain. The

855 5 10 15 20 kD 2.8) 35 40 45 HOM MDI.

Fig. 7 Field graph and curve (connecting each 5-cm. crown-heel interval)
of the per cent which the volume of both cerebral hemispheres forms of the en-
cephalon volume. Abscissa: total body length inem. Ordinate: hemisphere
volume in per cent of the encephalon volume. Individual cases indicated by
solid dots. (Data from table 40.)

fronto-occipital diameter is a very constant measure, due perhaps
to its close conformance to the skull and to the fact that it is not
affected greatly by the torsion of the brain parts or of the brain
axis during growth. Its reliability has justified its use to some
extent as a basic measurement.

430 HALBERT L. DUNN

The absolute fronto-occipital diameter (fig. 8) when plotted to
the crown-heel length is a straight line ascending from 2 cm. at
10 cm. (C H) to 9 em. at 50 em. (C H). Its empirical formula is:

Fronto-occipital diameter (em.) = 0.175 CH body length (cm.)
+ 0.25

TABLE 13
Fronto-occipital diameter

Formula: Fronto-occipital diameter (em.) = .175 crown-heel length (em.) + .25
(151 cases)

cnowienmsniaans | Gmennyan rnow7e:, | cazconamen | PHERRENR |
ip, EET eee ee eee OCCIPITAL ee ae OF CASES
Range Mean naa secre Mean AES MEANS
cm. cm. cm. cm. cm. cm. cm.
Oto 5 4.0 ibaa ORG 0.9 0.9 0 3
5 to 10 Cell 2.0 1.2 1.6 os —0.1 10
10 to 15 12.3 Deel. 1.4 2.3 2.4 +0.1 21
15 to 20 16.7 She 38) ae) Oo. 0 15
20 to 25 22.6 4.9 3.6 4.3 4.2 —0.1 21
25 to 30 27.0 6.5 4.4 Sol 5.0 —0.1 16
30 to 35 32.6 GOO) Sone 6.0 6.0 0 19
35 to 40 BYfoll Oth || ea 6.8 6.7 —0.1 19
40 to 45 42.3 8.4 bo 7.8 7.6 —0.2 11
45 to 50 48.2 9.3 hold) 8.6 8.7 +0.1 8
50 to 55 52.2 10.0 8.0 9.2 9.4 +0.2 8
TABLE 14

Calculated fronto-occipital diameter at 5-cm. intervals of crown-heel length

CROWN-HEEL LENGTH | eT eee hi are ea RATIO TO NEW-BORN

cm. cm, cm. per cent per cent

5 1.125 12.5
10 2.000 0.875 78.0 222
15 2.875 0.875 43.7 31.9
20 3.750 0.875 30.3 41.6
25 4.625 0.875 23.3 51.4
30 5.500 0.875 18.9 61.1
35 6.375 0.875 15.9 70.8
40 7.250 0.875 13.6 80.5
45 8.125 0.875 12.1 90.3
50 9.000 0.875 10.8 100.0
55 9.875 0.875 9.7 109.8

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 431

The absolute fronto-occipital diameter when considered in re-
lation to age (fig. 32, curve I) is about 1.3 cm. in the middle of
the third fetal month and increases at first rapidly and then more
slowly to 9 em. at birth.

The percentage increment of the fronto-occipital length for
5-em. intervals of body length (fig. 25, curve I) gradually declines

O | |
0 5 10 15 £0 £5 30 3D AO 4d 50 55cm.

Fig. 8 Field graph and curve of the growth of the cerebral hemispheres in
fetal life as shown by the fronto-occipital diameter. Abscissa: total body length
in em. Ordinate: fronto-occipital diameter in em. Individual cases indicated by
solid dots. Curve drawn to the formula. Y = 0.175X + 0.25. (Data from
tables 13 and 14.)

from about 80 per cent in the 5-to-10-cm. interval to about 10
per cent in the 50-to-55-cm. interval.

The percentage increment of the fronto-occipital length, when
calculated for fetal months (fig. 27, curve I), shows an approxi-
mate rate of growth of 150 per cent in the third fetal month.
This descends rapidly to 31 per cent in the fifth month and gradu-
ally to 9.5 per cent in the last fetal month.

432 HALBERT L. DUNN

The linear measurement from the frontal to the occipital pole
indicates therefore a steady absolute cerebral growth in the
anteroposterior axis (when compared to crown-heel length) and
a constant diminution of the rate of growth (when compared to
time).

TABLE 15
Temporal diameter
Formula: Temporal diameter (em.) = 0.138 crown-heel length (cm.) + 0.3 -
(141 cases)

Borers pare | Oe
pe || SY OMPORAL J'OBEERVED AND On GAanS
Range Mean Maxi- Mini- Mean ear ¢ peas
mum mum
cm. cm. cm. cm. cm. cm. cm.
Oto 5 3.9 1.0 0.7 0.85 0.85 0 4
5 to 10 Se) 250 ibs 1.45 1.4 —0.03 7
10 to 15 12.4] 3.5 11'5 pal 2.0 —0.1 19
15 to 20 16.6 3.4 1.8 2.6 2.6 0 15
20 to 25 2246, | “422 2.6 3.3 3.4 +0.1 21
25 to 30 27.0 4.6 3.2 4.1 4.0 —0.1 15
30 to 35 32.6 | 5.0 3.8 4.5 4.8 +0.3 16
35 to 40 Ofelia) (628 4.6 5.5 5.4 —0.1 18
40 to 45 42.3 | 6.7 5.6 6.2 6.1 —0.1 11
45 to 50 48.2 7.2 6.6 Ged 6.95 +0.25 8
50 to 55 52.2 | 10.4 6.9 7.9 C5 —0.4 of
TABLE 16

Calculated temporal diameter at 5-cm. intervals of crown-heel length

INCREMENT IN EACH

CROWN-HEEL LENGTH TEMPORAL DIAMETER 5-CM. INTERVAL
VL

RATIO TO NEW-BORN

em, cm, cm. per cent per cent
5 0.99 13.7
10 1.68 0.69 69.7 23.4
15 2.37 0.69 41.1 32.9
20 3.06 0.69 29.1 42.6
25 3.75 0.69 22.5 52.1
30 4,44 0.69 18.4 61.6
30 5.13 0.69 15.5 71.3
40 5.82 0.69 13.4 81.0
45 6.51 0.69 11.8 90.4
50 7.20 0.69 10.6 100.0
50 7.89 0.69 YEG 109.5

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 483

The curve of absolute temporal diameter (fig. 9) is also a
straight line when based on crown-heel length. Starting at about
1 cm. at the 5 cm. (C Hy), it increases steadily to about 72 cm.
at 50 cm. (C H). Its empirical formula is:

Temporal diameter (cm.) = 0.1388 C H body length (cm.)
+0.3

When studied in relation to time (fig. 32, curve IT), this diam-
eter is found to be 1 cm. at the beginning of the third fetal

0 5 10 15 c0 05 30 35 40 £45 50 55cm

Fig. 9 Field graph and curve of the growth-of the cerebral hemispheres in
fetal life as shown by the temporal diameter. Abscissa: total body length
in em. Ordinate: temporal diameter in em. Individual cases indicated by
solid dots. Curve drawn to the formula: Y = 0.138X +0.3. (Datafrom tables
15 and 16.)

month and to rise at first fairly rapidly and then more slowly to
7.2 em. at birth.

The rate of growth of the temporal diameter (fig. 25, curve IT)
decreases steadily from 70 per cent for the 5-to-10-cm. interval
to about 10 per cent for the 50-to-55-cm. interval. : ;

The percentage increment of the temporal diameter, when cal-
culated for fetal months (fig. 27, curve II), is approximately
70 per cent in the fourth fetal month and falls to about 35 per

434 HALBERT L. DUNN

cent in the fifth month and to 9.2 per cent in the tenth month.
Or, in other words, the transverse axis of the cerebral hemispheres
shows a constant diminution in the rate of growth throughout
the fetal period.
TABLE 17
Volume of the cerebellum

Formula: Volume of cerebellum (cc.) = .01 (.095 crown-heel length
(em.))49 + 20.0
(114 cases)

hte Seals mic ted o eanMRaLeM ii CALCULATED potecaane
VOLUME OF | OBSERVED AND epee
Maxi. Mini- CEREBELLUM | CALCULATED
Range Mean arin aaa Mean MEANS
cm. cm cc ce. cc cc cc

Oto 5

5 to 10
10 to 15 12.8 0.5 0.1 0.3 0.5 +0.2 10
15 to 20 16.9 0.7 0.2 0.4 0.5 +0.1 14
20 to 25 22.6 2.0 0.5 0.9 0.6 —0.3 18
25 to 30 27.0 Piste 0.9 1.3 12 —0.1 13
30 to 35 aPAsTI 4.0 HO; 2.4 2.8 +0.4 16
35 to 40 37.2 9.3 3.4 5.4 Nal —0.3 Nef
40 to 45 42.3 15 N76 8.9 9.3 +0.4 11
45 to 50 48.0 22.0 14.0 19.0 ilvfeal —2.0 7
50 to 55 aoe 26.0 18.0 Dilisres PANU +4.4 8

TABLE 18

Calculated cerebellum volume at 5-cm. intervals of crown-heel length

INCREMENT IN EACH

CROWN-HEEL LENGTH | CEREBELLUM VOLUME 5-CM. INTERVAL

RATIO TO NEW-BORN

cm. ce. ce. per cent per cent

5
10 0.20 1.0
15 0.28 0.08 40.0 1.3
20 0.43 0.15 53.6 2.1
25 0.89 0.46 107.0 4.3
30 1.89 1.00 112.0 9.1
35 3.80 io 100.1 18.2
40 7.13 3.39 87.8 34.1
45 12.60 5.47 76.8 60.3
50 20.90 8.30 65.8 100.0

50 33.40 12.50 59.8 160.0

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 435

Growth of the cerebellum

The growth of the cerebellum is portrayed by the graphs in
figures 10 to 13, inclusive, and by the outline sketches in figure
38. The absolute volume of the cerebellum, when plotted against

0 5 10 15 20 25 30 551) “AO Ad 5O 55cm

Fig.10 Field graph and curve of the growth of the cerebellum in weight and
volume in fetal life. Abscissa: total body length in em. Ordinate: cerebellum
weight and volume in grams (cc.). Individual cases indicated by solid dots
(for weight) and by circles (for volume). Curve drawn to the formula: Y =
0.01 [(0.095X )4:9 + 20.0]. (Data from tables 17 and 18.)

crown-heel body length (fig. 10), presents a concave volume curve

expressed by the empirical formula:

Volume of the cerebellum (ec.) = 0.01 (0.095 C H body length
(em.))4-? +20.0

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 5

436 HALBERT L. DUNN

The cerebellum is not grossly visible before the fetus is about
10 em. (C H) long. At 20 cm. (C H) its volume is about 0.4 ec.,
and this increases slowly to about 2 cc. at 30 em. (C H). The

% 5 10 15 £0 5 50 35 40 45 50 55cm

Fig. 11 Field graph and curve (connecting the average points of each 5-cm.
crown-heel interval) of the per cent which the volume of the cerebellum forms of
the encephalon volume. Abscissa: total body length in cm. Ordinate: cere-
bellum volume in per cent of entire brain volume. Individual cases indicated
by solid dots. (Data from table 40.)

absolute volume curve then turns sharply upward and rises very
rapidly to 21 cc. at 50 cm. (C H).

On the basis of time (figs. 34 and 35), the volume of the cere-
bellum is about 0.4 ec. at four fetal months and approximately

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 437

2.3 cc. at six fetal months. At this time it begins to enlarge
rapidly, increasing almost ten-fold in the last four months of
intra-uterine life and reaching a volume of 21 cc. at birth. This
tremendous growth in volume in the last three or four months of
fetal life, at a time when all other brain parts are growing far less
rapidly, is a peculiar characteristic of the cerebellum.

The percentage increment of the cerebellum volume (fig. 26,
curve VII) reflects the peculiarities of the curve of absolute
volume. Starting at a comparatively low value, it rises to a
peak of 112 per cent for the 25-to-30-cm. interval and descends
to 60 per cent in the 50-to-55-cm. interval.

The percentage increment of the cerebellum volume, when
calculated for time (fig. 28, curve IV), shows an approximate rate
of growth of about 160 per cent in the fifth month, and then falls
rapidly to 54 per cent for the tenth month.

The relative growth of the cerebellim is also illustrated by the
per cent which it forms of the entire brain volume at various
periods in fetal life (fig. 11). This value is, with slight variations,
about 3 per cent until the middle of the sixth fetal month. At
this time the tremendous growth of the cerebellum becomes an
important factor in the total increase of the brain as a whole, and
by birth it forms about 6 per cent of the entire brain volume.

The absolute length of the vermis cerebelli, when plotted to
crown-heel body length (fig. 12), is expressed by the formula:

Vermis cerebelli length (cm.) = 0.01 [(C H length (cm.))'!.#
+0.15]

It also portrays the great growth of this part of the brain in the
last four fetal months. At 10 em. (C H) the vermis cerebelli
length is only about 0.4 em. It ascends in a very shallow curve
40 2.6 em. at 50 em. (C Hy).

The absolute length of the vermis cerebelli when calculated for
fetal months (fig. 31, curve I) rises in practically a straight line
from 0.4 cm. at the beginning of the fourth month to 2.64 cm.
at birth.

The rate of growth of the vermis cerebelli length (fig. 25, curve
IIT) shows a gradual decrease from 50 per cent in the 10-to-15-cm.
interval to 13 per cent at the 50-to-55-cm. interval. The entire

438 HALBERT L. DUNN

percentage increment curve, however, is much higher than any
other percentage increment based upon a straight-line measure-
ment (with the single exception of the vermis cerebelli height,
fig. 25, curve IV).

TABLE 19

Length of the vermis cerebelli

Formula: Vermis cerebellilength (em.) = .01 [(crown-heel length (em.))!4! + 0.15]
(124 cases)

—_ vermis [OBSERVED AND| Op Cases
Range Mean sees abyte Mean CeReeEEut MEANS
em. cm, cm. cm. cm. cm. cm.

Oto 5

5 to 10
LOOMS 27; 0.9 0.3 0.5 0.5 0.0 16
15 to 20 16.9 deal 0.4 0.6 0.7 +0.1 14
20 to 25 eRe 15) 14 0.6 0.9 1a +0.1 18
25 to 30 27.0 1.8 0.9 1.15 ie2 +0.05 14
30 to 35 32.8 az 1 eet J25 1.5) 0 16
35 to 40 Ove: 2.6 1.3 1.8 1.8 0 18
40 to 45 42.2 220) ed Pelt ral 0 i
45 to 50 48.2 27 Peo eo Bay 0 8
50 to 55 522 3.4 20 Bath 2.8 +0.1 8

TABLE 20

Calculated vermis cerebelli length at 5-cm. intervals of crown-heel length

CROWN-HEEL LENGTH Mak ieee aera Se ene RATIO TO NEW-BORN
cm, cm, cm. per cent per cent
5
10 0.41 15.5
15 0.61 0.20 49.0 23.1
20 0.83 0.22 36.1 31.5
25 1.08 0.25 30.2 40.9
30 1.36 0.28 25.9 51.6
35 1.65 0.29 21.3 62.6
40 1.96 0.31 18.8 74.3
45 2.29 0.33 16.8 86.8
50 2.64 0.35 15.3 100.0
55 2.99 0.35 13.3 118.2

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 439

The percentage increment of the vermis cerebelli length when
calculated for fetal months (fig. 27, curve VIII) is approximately
85 per cent in the fifth month, descending at first swiftly and then
more slowly to about 12.5 per cent in the tenth month.

oy J [
cm e
3.0
e 2
ie e ee
ee ee e |
2.0 . |
2. eo Ue
ry e e 1
oe °
eferece
se) © efse 4
e eo eo ¢«
10 comgna «
ee e se «
e . o
5 e om ane ae
e eS ee
fo) | | |

0 5 10 15 £0 £5 30 35 = 40 AD 50 55cm

Fig. 12 Field graph and curve of the growth of the cerebellum in fetal life
as shown by the vermis cerebelli length. Abscissa: total body length in em.
Ordinate: vermis cerebelli length in em. Individual cases indicated by solid
dots. Curvedrawn tothe formula: Y = 0.01 (X!.41+ 0.15). (Data from tables
19 and 20.)

440 HALBERT L. DUNN

The absolute height of the vermis cerebelli when plotted
against crown-heel body length (fig. 13) also forms a shallow
curve, expressed empirically by the formula:

Vermis cerebelli height (em.) = 0.01 (C H body length (em.))1-37

It is 0.23 em. at 10 em. (C H) and rises to about 2.1 em. at 50 em.
(COMAGE

TABLE 21
Height of the vermis cerebelli
Formula: Vermis cerebelli height (em.) = .01 (crown-heel length (em.))1!37

(122 cases)

caown-nmetiexons | ovmmevepsercunor | cstoouanen | REFERENCE
meron OF | oneenvay axo | XTMBEE

Range Mean aa eae Mean ORO EDEEE a |

Oto 5

5 to 10
10 to 15 128 |) 1026 0.3 0.4 0.3 -—0.1 14
15 to 20 16.9 ONT. 0.2 OS 0.5 0 14
20 to 25 22.6 |. 0.9 0.5 0.7 0.7 0 18
25 ‘to 30 27.0 1.0 0.6 0.8 0.9 +0.1 14
30 to 35 Byers) 1.4 0.8 ital 1.2 +0.1 16
35 to 40 =f 1.9 1.0 1.4 1.4 0 18
40 to 45 42.2 2.0 eal 1.6 Ue 76 +0.1 12
45 to 50 48.2 PaaS 1.6 220 2.0 0 8
50 to 55 Es 2.8 LG Pp 2; 0 8

TABLE 22

Calculated vermis cerebelli height at 5-cm. intervals of crown-heel length

CROWN-HEEL LENGTH | Coppnuiey REIGHT eS Rie eae RATIO TO NEW-BORN
cm. cm. cm. per cent per cent
5
10 0.23 10.8
15 0.41 0.18 78.1 19.2
20 0.61 0.20 48.8 26.6
25 0.82 0.21 34.4 38.5
30 1.06 0.24 29.2 49.8
35 1.31 0.25 23.6 61.5
40 1.57 0.26 19.8 73.7
45 1.84 0.27 17.2 86.3
50 2.13 0.29 15.8 100.0
55 2.42 0.29 12.0 113.7

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 441

The absolute height of the vermis cerebelli when calculated for
age in fetal months (fig. 31, curve IJ) rises in practically a straight
line from 0.23 em. at the beginning of the fourth month to 2.1 cm.
at birth.

30

cm.

ED

£0

O 5 10 15 £0 25 30 a}, AO A5 5O 55cm.

Fig. 13 Field graph and curve of the growth of the cerebellum in fetal life
as shown by the vermis cerebelli height. Abscissa: total body length in cm.
Ordinate: vermis cerebelli height in em. Individual cases indicated by solid
dots. Curve drawn to the formula: Y = 0.01 (X!-37). (Data from tables 21
and 22.)

The percentage increment of the vermis cerebelli height when
calculated for 5-em. intervals of crown-heel length (fig. 25, curve
IV) is similar to that of the vermis cerebelli length except that it
is somewhat greater at first and falls more rapidly.

The percentage increment of the vermis cerebelli height, when
calculated for fetal months (fig. 27, curve IX), shows a rate of

442 HALBERT L. DUNN

growth of about 126 per cent for the fourth month, falls to 58 per
cent for the fifth month, and then descends steadily to about 13
per cent for the tenth month.

The growth of the cerebellum during the latter part of fetal
life is also strikingly illustrated by the midsagittal sections
shown in figures 37 and 38. In the third fetal month (figure
38, C) the cerebellum grows posteriorly and parallel to the pons
and medulla; by the middle of the sixth fetal month (fig. 38, D, E,
F) it assumes its characteristic shape and position in relation to
the cerebral hemispheres. From the middle of the sixth fetal
month to birth (fig. 38, G to I, inclusive) each succeeding tracing
indicates a greater relative size in this portion of the brain.

It will be noted in this connection that the graphs of absolute
growth of the cerebellum in both of the linear dimensions resemble
volume curves much more closely than does the graphic expression
of any other lineal dimensions of the nervous system.

Growth of the pons and medulla

The growth of the pons and medulla (fig. 14) presents a typical
volume curve having the empirical formula:

Pons and medulla vohime (cc.) = 0.01 [(0.2 C H body length
(em.))2-67 + 20.0]

At 10 em. (C H) their volume is about 0.3 cc., by 30 em. (C H)
‘ it is 1.4 ec., and at birth (50 cm. C H) it is approximately 5 cc.
When calculated for age in fetal months (fig. 35, curve IJ), their
volume is 0.2 ce. at three months and 1.4 cc. at six months.
From this time it rises steadily to about 5 ce. at birth.

The percentage increment of the volume (fig. 26, curve VIII)
is approximately 50 per cent for the 10-to-15-em. (C H) interval
and rises to about 56 to 57 per cent for the interval between 15
to 20 em. It then descends steadily to about 27 per cent from
50 to 55 em. (C H).

The percentage increment of the pons and medulla volume as
calculated for fetal months (fig. 28, curve II) shows a rate of
growth of about 96 per cent for the fourth month, and then de-
scends steadily to about 27.5 per cent for the last month of pre-
natal life.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM

One of the most interesting facts concerning the prenatal
growth of the pons and medulla is the per cent of the entire brain
volume which they form during fetal life (fig. 15 and table 40).
In the middle of the fourth fetal month the pons and medulla
make up 5.5 per cent of the entire brain volume, at 5 months they

TABLE 23

Volume of the pons and medulla

Formula: Pons and medulla volume (ec.) = .01 [(.2 crown-heel length

(em.) )?-§7 + 20]

(112 cases)

CROW N-HEEL LENGTH

OBSERVED PONS

AND MEDULLA VOLUME

Range Mean Bae
cm. cm. ce.

Oto 5

5 to 10
10 to 15 1256 O25
15 to 20 16295 ORG
20 to 25 DOEo 1.0
25 to 30 tbe) eal War!
30 to 35 Sth | pal |
35 to 40 SY/ UME Bee
40 to 45 42.3 3.9
45 to 50 48.0} 5.5
50 to 55 8222 | 6:3

Mini-

mum

cc.

PWNEe OOOO
or SI OD O

Hm e bo
On

CALCULATED pea
PONS AND | op cERVED AND
| Mean “VOLUME ye ee |
ce. cc. aa
ORS 0.3 0
oe te +0.1
0.8 0.8 0
ni teh +0.2
< we +0.2
2.4 2.3 0.1
ae: He +0.1
ae Be +0.1
i ce 40.2
TABLE 24

NUMBER
OF CASES

Calculated pons and medulla volume at 5-cm. intervals of crown-heel length

CROW N-HEEL LENGTH

PONS AND

MEDULLA VOLUME

INCREMENT IN EACH

ce.

5-CM,. INTERVAL

per cenit

00.0
56.5
04.1
49.0
43.5
38.3
34.2
30.8
27.4

RATIO TO NEW-BORN

per cent

5.3
8.0
12.5
19.3
28.7
41.2
57.0
76.4
100.0
128.0

443

444 HALBERT L. DUNN

form only about 2.7 per cent, and from that time on to birth they
gradually decrease in relative volume, so that they form only
1.5 per cent of the entire brain volume in the new-born. This

0 5 10 15 £0 (ay 350 35 40 AD 50 55cm.

Fig.14 Field graph and curve of the growth of the pons and medulla in weight
and volume in fetal life. Abscissa: total body length in cm. Ordinate: pons
and medulla weight and volume in grams (cc.). Individual cases indicated by
solid dots (for weight) and by circles (for volume). Curve drawn to the formula:
Y = 0.01 [(0.2X)2-57 + 20.0]. (Data from tables 23 and 24.)

relative decline is clearly due to the great increase in the ab-
solute volume of the cerebral hemispheres and of the cerebellum
during the last half of fetal life.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 445

The pons length also shows some interesting changes in the
course of fetal life. The graph of absolute pons length plotted
against total body length (fig. 16) is practically a straight line
having the formula:

Length of pons (em.) = 0.0263 C H body length (em.) + 0.16

Fig. 15 Field graph and curve (connecting the average points of each 5-cm.
crown-heel interval) of the per cent which the volume of the pons and medulla
forms of the encephalon volume. Abscissa: total body lengthin cm. Ordinate:
pons and medulla volume in per cent of the entire brain volume. Individual
cases indicated by solid dots. (Data from table 40.)

The length increases steadily from 0.42 em. at 10 em. (C H) to
about 1.53 cm. at birth (50 em. C H).

When plotted against time, the pons length increases rather
rapidly from the third to the beginning of the sixth month, and
then grows more slowly at a uniform rate until birth.

446

HALBERT L. DUNN

The percentage increment of the pons length against intervals
of growth in body length (fig. 25, curve V) starts at 30 per cent
between 10 and 15 cm. (C H) and falls steadily but slowly to
9.8 per cent in the 50-to-55-cm. interval.

The percentage increment of the pons length as calculated for
fetal months (fig. 27, curve III) shows a rate of growth of about

TABLE 25
Pons length

Formula: Pons length (em.) = 0.0263 crown-heel length (cm.) + 0.16
(124 cases)

CROWN-HEEL LENGTH OBSERVED PONS LENGTH eae
— Cau EAE GHSEe RAND NUMBER
Range Mean Sa Nips ae te x cas air oe ie GIP eaters
cm, cm, cm. cm. cm, cm, cm.

Oto 5

5 to 10
10 to 15 UPA 0.6 0.3 0.5 0.5 0 18
15 to 20 16.7 0.8 On5 0.6 0.6 0 13
20 to 25 22.6 0.9 0.7 0.8 0.8 0 18
25 to 30 27.0 TA 0.6 0.9 0.9 0 13
30 to 35 32.8 its 0.9 1.0 10) 0 16
30 to 40 Sieik i) 1.0 lig? ipa =e 18
40 to 45 42.2 1.4 1.0 es 1,333 0 12
45 to 50 48.2 ie 11824 1fa8ts) 1.4 —0.05 8
50 to 55 52.2 176 eS 15) 125 0 8

TABLE 26

Calculated pons length at 5-cm. intervals of crown-heel length

CROWN-HEEL LENGTH

PONS LENGTH

INCREMENT IN EACH
5-CM. INTERVAL

ee

29
55
68
81
94
.08
21
34
47
60

p
~)

Peer OOOCOCo

cm.

plas
alec
PRE
.13-++
“BSS
si BSE
.13+-
sllsse
.13+
0.13+

SSeS} (See)

per cent

RATIO TO NEW-BORN

per cent

28.7
37.6
46.5
50.4
64.3
73.3
82.3
91.2
100.0
108.8

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 447

75 per cent in the third month and descends steadily to approxi-
mately 7 per cent in the tenth month.

There is no doubt that the pons and medulla form a large part
of the brain in early fetal life. Figures 38, a, b, ce, and d, sub-
stantiate the percentage curve (fig. 18). They show that the
pons and medulla form a very large portion of the entire brain
volume at this time. By the middle of the third fetal month
(fig. 38, b) fully one-third of the brain, as it is seen in mid-

20

cm

0 5 10 15 £0 e5 30 35 AO A5 50 55cm.

Fig. 16 Field graph and curve of the growth of the pons and medulla in fetal
life, as shown by the pons length. Abscissa: total body length in em. Ordi-
nate: pons length in cm. Individual cases indicated by solid dots. Curve
drawn to the formula: Y = 0.0263X + 0.16. (Data from tables 25 and 26.)

sagittal section, is formed by the pons and medulla. In the
fourth and fifth months (figs. 38, b, e, d, and e) the pons and
medulla become relatively smaller.

Growth of the midbrain

A consideration of the colliculi and the midbrain does not lead
to quite such clean-cut results as does the study of the other
brain parts. The midbrain, as it was prepared for the volume and
weight determinations, included not only the colliculi above the
iter, but also a portion of the brain stem below it.

448 HALBERT L. DUNN

The absolute volume of the midbrain when plotted against
body length (fig. 17) presents a shallow curve and has the em-
pirical formula:

Volume of midbrain (cc.) = 0.01 [(0.168 C H
length (em.))?-5§+12.0]

TABLE 27
Volume of the midbrain
Formula: Midbrain volume (cc.) = .01 [ (.168 crown-heel length (cm.))?*5§ + 12]
(109 cases)

Se ee ety ce See CALCULATED peta
BPE Se) Foe Lee ee B VOLUME OBSERVED AND ones
Mane ME OF MIDBRAIN CALCULATED
Range Mean aes Area Mean MEANS
cm. cm. ce. ce. cc. ce. cc.

Oto 5

5 to 10
10 to 15 12.9 0.3 0.1 0.2 0:2 0 9
15 to 20 16.9 0.4 0.2 0.3 0.3 0 14
20 to 25 22.0 0.8 0.3 0.5 0.4 —0.1 lg
25 to 30 27.0 0.9 0.4 0.6 0.6 0 13
80 to 35 ByA5 6 1.4 0.4 0.8 0.9 +0.1 16
35 to 40 Sino lente 0.8 1.4 1.2 —0.2 14
40 to 45 42.3 2.2 f2 16 1.6 0 il
45 to 50 48.0 3. 1 125 2.4 2.2 —0.2 Ui
50 to 55 5222 3.0 1.8 2.4 Papi +0.3 8

TABLE 28

Calculated midbrain volume at 5-cm. intervals of crown-heel length

INCREMENT IN EACH

CROWN-HEEL LENGTH MIDBRAIN VOLUME Boe eR RATIO TO NEW-BORN
cm. ce. ce. per cent per cent
5
10 0.16 6.6
15 0.23 0.07 43.7 9.4
20 0.34 0.11 47.8 13.9
25 0.51 0.17 50.0 20.9
30 0.75 0.24 47.0 30.7
35 1.05 0.30 40.0 43.0
40 1.48 0.38 36.2 58.5
45 1.89 0.46 32.0 77.4
50 2.44 0.55 29.1 100.0
55 3.12 0.68 27.8 128.0

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 449

At 13 em. (C H) the absolute midbrain volume is about 0.2 ec.,
and this steadily rises to 0.75 cc. at 30 cm. (C H). From this
point the absolute volume increases at a faster pace to about
2.5 cc. at 50 cm. (C H).

SOMES Sem:

Fig. 17 Field graph and curve of the growth of the midbrain in weight and
volume in fetal life. Abscissa: total body length in cm. Ordinate: midbrain
weight and volume in grams (cc.). Individual cases indicated by solid dots
(for weight) and by circles (for volume). Curve drawn to the formula: Y =
0.01 [(0.168X)2-5§ + 12.0]. (Data from tables 27 and 28.)

The volume of the midbrain, when calculated for age in fetal
months (fig. 35, curve ITI), is about 0.15 cc. at the third month
and increases to about 0.8 cc. at six months. Thereafter it
grows slowly to about 2.5 ce. at birth.

450 HALBERT L. DUNN

The percentage increment of the midbrain volume, when cal-
culated for 5-em. intervals of crown-heel length (fig. 26, curve IX),
is about 45 per cent for the 10-to-15-cm. interval. It rises
slightly at the 20-to-25-cm. interval and descends to 30 per cent
for the 50-to-55-cm. interval.

The percentage increment of the midbrain volume as calculated
for fetal months (fig. 28, curve IIT) is approximately 81 per cent

5
J
%

Ot

0 ] t
6) 5 s1) 15 £O 25 50 35 40 45 50 55am

Fig. 18 Field graph and curve (connecting the average points of each 5-cm.
crown-heel interval) of the per cent which the midbrain volume forms of the
encephalon volume. Abscissa: total body length in em. Ordinate: mid-brain
volume in per cent of the encephalon volume. Individual cases indicated by
solid dots. (Data from table 40.)

in the fourth month and descends to about 25 per cent in the last
fetal month. '

The slow growth of the midbrain is reflected in the per cent
which this brain part forms of the entire brain volume (figure 18
and table 40). Beginning at 2.9 per cent at three months, it
steadily declines to but 0.7 per cent of the entire brain volume
at birth.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 451

The length of the colliculi is not a true index of the mid-
brain growth in early fetal life, and its significance as a measure
of growth of that portion of the mesencephalon below the iter
is particularly questionable. In some fetuses of three to four
months a sharp line of demarcation between the superior col-
liculi and the thalamencephalon may be present, while in other
specimens of the same age no line of separation can be distin-
guished.

This conclusion is supported by evidence shown in figures 36,
a,b, ec. Figures 36, a, and 36, b, of specimens prior to the third
fetal month show no colliculi; figure 36, c, shows colliculi well
marked off and relatively larger than in a new-born specimen.

The curve of the absolute length of the colliculi, when plotted
against body length (fig. 19), isa straight line. It starts at 0.7 cm.
at 15 em. (C H) and rises to 1.22 cm. in length at 50 cm.
(C H). It may be expressed by the empirical formula:

Length of colliculi (em.) = 0.015 C H body length (em.)
+0.47

The growth of the colliculi in absolute length when plotted
against time (fig. 38, curve VI) approaches a straight line, al-
though the rate of growth is more rapid in the fourth and fifth
months than in the last four fetal months.

_ Growth of the spinal cord

The growth of the spinal cord resembles that of the brain and
particularly that of the pons and medulla. The absolute volume
of the spinal cord plotted against crown-heel length (fig. 21) is a
very shallow but typical volume curve, which may be expressed
by the empirical formula:

Volume of the spinal cord (ec.) = 0.01 [(0.17 C H
length (em.)) 2:57 + 11.0]
The absolute volume at 15 em. (C H) is approximately 0.22 ce.
The curve rises steadily until it reaches about 1 ec. at 35em. (CH).
From this point it proceeds upward at a sharper pitch, reaching
about 2.5 cc. at 50 em. (C H).

The growth in absolute volume against time (fig. 35, curve IV)

corresponds almost exactly with that of the midbrain and shows

452 HALBERT L. DUNN

a fairly rapid rate of absolute growth from the third to the be-
ginning of the fifth month and a slower and more regular growth
thereafter.

The data of Jackson (’09) (table 35) show a uniformly higher
average for the absolute spinal cord volumes. His small fetuses

TABLE 29
Length of the colliculi
Formula: Colliculi length (em.) = .015 crown-heel length (cm.) + .47.
(117 cases)

E D
CROWN-HEEL LENGTH CAEL NAGLE) DSU DIFFERENCE

eee) penne on || emer aire | SE
Rene Minie THE COLLICULI | CALCULATED
Range Mean main Tali Mean MEANS
cm. cm. cm. cm. cm. cm. cm.

Oto 5

5 to 10
10 to 15 | 13.0 0.9 0.5 0.66 0.66 0 11
15 to 20 | 16.9 0.9 0.5 0.74 0.73 —0.01 14
205025) 92255 1.0 0.7 0.80 0.81 +0.01 18
25 to 30 | 26.9 1.0 0.7 0.90 0.87 —0.03 12
30 to 35 | 32.8 1.7 0.8 0.99 0.96 —0.03 16
35 to 40 | 37.1 ileal 0.9 0.98 1.03 +0.05 18
40 to 45 | 42.2 1.2 1.0 1.10 1.10 0 12
45 to 50 | 48.3 1.4 ileal 1.23 20 —0.03 8
50 to 55 | 5252 1.3 Weal 1.20 1.25 +0.05 8

TABLE 30

Calculated length of the colliculi at 5-cm. intervals of crown-heel length

INCREMENT IN EACH

CROWN-HEEL LENGTH LENGTH OF COLLICULI 5-OM. INTERVAL

RATIO TO NEW-BORN

cm. cm. em. per cent per cent

5

10

15 0.695 57.0
20 0.775 0.75 10.8 63.1
25 0.845 0.75 9.8 69.3
30 0.920 0.75 8.9 75.4
35 0.995 0.75 8.2 81.5
40 1.070 0.75 7.6 87.8
45 1.145 0.75 7.0 93.9
50 1.220 0.75 6.6 100.0
55 1.295 0.75 6.2 106.2

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 453

(obtained by sectioning) show an absolute volume of 0.034 ce.
at approximately the close of the first month, while his two
largest fetuses (new-born) have a cord volume of about 3 ce.
The percentage increment of the spinal cord volume when de-
termined for 5-cm. intervals of crown-heel body length (fig. 26,

7d =

14

13

Le

ia

fo a vorleee
al . eee e

éL aicactatens

0) Lee tl cine ail, acl
OND: 10 15 £0 £5 30 355 40 45 5O) 755cmn:

Fig. 19 Field graph and curve of the growth of the midbrain in fetal life as
shown by the colliculi length. Abscissa: total body length in em. Ordinate:
colliculi length in em. Individual cases indicated by solid dots. Curve drawn
totheformula: Y = 0.015X + 0.47. (Data from tables 29 and 30.)

curve X) is 47 per cent for the 10-to-15-cm. interval. It rises to
about 55 per cent for the 15-to-20-em. interval, and then falls to
about 25 per cent for the 50-to-55-cm. interval.

The percentage increment of the spinal cord volume when
calculated for fetal months (fig. 28, curve I) shows a rate of growth
of approximately 94 per cent in the fourth month and then de-
scends steadily to 26 per cent in the last fetal month.

454

HALBERT L. DUNN

The relation of the spinal cord volume to the volume of the
entire brain is shown in figure 22 and in table 40. At the middle
of the second month the cord volume is equal to 4.4 per cent of

the total brain volume.

It then drops rapidly at first and

afterward more slowly to about 0.85 per cent of the total brain

volume at birth.

TABLE 31
Frontospinal length
Formula: Frontospinal length (em.) = (2 crown-heel length (em.))47 — 2.2
(145 cases)

embrace ieeancrs’ “ (ueleawaien eee eee
FRONTOSPINAL | OBSERVED AND Aten nae
Maxi- Mini- LENGTH CALCULATED
Range Mean eat tah Mean MEANS
cm. cm. cm. cm. Luts cm. cm.
Oto 5 AON AKO 0.7 0.9 0.5 —0.5 3
5 to 10 618 |. 1.7 1.0 1.4 1.2 =e 9
10 told 2. | 2.9 1.9 2.1 23 ae 20
15 to 20 | 16.6] 3.5 2.6 DG 3.0 eet 14
20.46: 251\ 22.6.) “4.5 3.0 3.8 3.8 0 20
25 to 30 | 27.0| 4.6 3.4 4.1 4.3 0), 2 15
30 to 35 | 32.8 | 5.3 4.3 4.8 4.9 +0.1 rely;
35 to 40 | 37.1 | 6.5 4.0 5.3 5.4 220.2 19
40 to 45 | 42.2| 6.8 5.1 5.9 5.8 Sail 12
45 to 50 | 48.2] 6.9 5.9 6.3 6.4 eet 8
50 to 55 | 52.2| 7.3 6.3 6.8 6.7 =n 8
TABLE 32

Calculated frontospinal length at 5-cm. intervals of crown-heel length

CROWN-HEEL LENGTH

CALCULATED FRONTO- |
SPINAL LENGTH |

INCREMENT IN EACH
5-CM. INTERV!

AL

cm.

0.75
1.89
2.75
3.46
4.09
4.65
Dold
5.64
6.09
6.51
6.91

cm. per cent
1.14 152.0
0.86 44.8
0.71 28.2
0.63 18.2
9.56 13.4
0.52 12.4
0.47 9.3
0.45 8.0
0.42 6.7
0.40 6.3

RATIO TO NEW-BORN

per cent

11.5
29.0
42.2
53.1
62.8
71.4
79.5
86.6
94.5
100.0
106.2

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 455

The absolute length of the spinal cord (fig. 23) ascends rather
sharply from 2.2 em. length at 5 em. (C H) to 9.2 cm. at 25 cm.
(CH). It then ascends more slowly to 14.2 cm. at birth (50 cm.
C H). Its empirical formula is:

Length of spinal cord (em.) = (10.0 C H body
length (cm.))-467 — 4.0
The curve of cord length, when plotted against time, is quite
similar to that plotted against body length, but the rise is a more
rapid one (fig. 32, curve IV).

& l |
cm ‘
~

O 5 10 i5 £0

05 30 Dpy 0) 45 50 55cm

Vig. 20 Field graph and curve of the growth of the brain stem in fetal life as
shown by the fronto-spinal length. Abscissa: total body length inem. Ordi-
nate: fronto-spinal length in em. Individual cases indicated by solid dots.
Curve drawn totheformula: Y = (2.0X)-47—2.2. (Data from tables 31 and 32.)

The percentage increment of the cord length as calculated for
5-em. intervals of crown-heel length (fig. 25, curve VIII) is about
109 per cent for the 5-to-10-cm. interval, falls rapidly to 23 per
cent for the 15-to-20-cm. interval, and then gradually to 6 per
cent for the 50-to-55-cm. interval.

The percentage increment of the spinal-cord length when cal-
culated for fetal months (fig. 27, curve V) shows a rate of growth
of approximately 400 per cent in the third month, descending
rapidly to 26 per cent in the sixth month, and then gradually to
nearly 6 per cent in the last month of the fetal period.

456 HALBERT L. DUNN

DISCUSSION

The discussion of the results of this work will be limited to,
1) a comparison of the growth of the central nervous system as a
whole with that of the various other viscera and parts of the body
in the fetal period; 2) the analysis of the type of growth of the

TABLE 33
Volume of the spinal cord
Formula: Spinal cord volume (ec.) = .01[(0.17 crown-heel length (em.) )?:57 + 11.]
(128 cases)

VOLUME OF | OBSERVED AND or CABES
Maxi- Mini- SPINAL CORD CALCULATED
Range Mean ain ae Mean MEANS
cm. cm. ce. Ge. (HP: ce. ce.

Oto 5

5 to 10 fea OXO5n| 02 05m 0205 0.13 0.08 3
10 to 15 1220" 7OF3 0.1 0.2 0.2 0 18
15 to 20 1GRS 5) O25 0.1 0.3 0.3 0 14
20 to 25 22.6 | 0.7 0.3 0.5 0.4 —0.1 19
25 to 30 26.9 iil 0.3 0.65 0.6 0 14
30 to 35 32.6 1.4 O25 0.9 0.9 0 17
35 to 40 SHC) | 24a 0.7 15d ila 0 17
40 to 45 41.9 2.3 52 ise led 0 11
45 to 50 48.0 3.1 1.4 2.4 2.3 —(): 1 7
50 to 55 52.2 3.8 1.8 2.5 2.8 +0.3 8

TABLE 34

Calculated spinal cord volume at 5-cm. intervals of crown-heel length

INCREMENT IN EACH

CROWN-HEEL LENGTH SPIN CORD
RO H PINAL CORD VOLUME 5-CM. INTERVAL

RATIO TO NEW-BORN

cm. ce. ce. per cent per cent

5

10 0.15 5.9
15 0.22 0.07 46.7 8.6
20 0.34 0.12 - 64.5 13.3
25 0.52 0.18 53.0 20.3
30 O47 0.25 48.1 30.1
30 1.09 0.32 41.5 42.5
40 1.49 0.40 36.7 58.2
45 1.98 0.49 32.9 77.4
50 2.56 0.58 29.2 100.0
55 3.24 0.68 24.5 126.5

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 4097

central nervous system into its component parts; 3) observations
upon the relation of volumetric and lineal measurements of the
nervous system, and, 4) a summary of the proportions which the

35 }
iS |
Oo
30 Be
°
5 e
%
e °
£5 e e
°
e
oe fe °
e e
= ° ° °
20 o ©
e
@ of, °
°
o /*
15 0° of .
e
e °
Seen. Ke
° fe ee 00
° ee/fo &
10 ° e°8 6 50
oad oo
oe ; ® 9
- % g 8 og
af he Pawn Bos
2) ° 9s Pf 2
e © a he -.
39 ED ee. °
ns eo fis}
(6) oh oe ©
2} 10 15 20 25 30 35 40 45 50 55cm.,

Fig. 21 Field graph and curve of the growth of the spinal cord in weight and
volume during fetal life. Abscissa: total body length in em. Ordinate: spinal
cord weight and volume in grams (cc.). Individual cases indicated by solid
dots (for weights) and by circles (for volume). Curve drawn to the formula:
Y = 0.01 [(0.17X)?-57 + 11.0]. (Data from tables 33, 34, 35.)

various brain parts form of the encephalon in prenatal life. The
consideration of the growth of the fetal brain and the spinal cord
in relation to the postnatal changes of these structures is reserved
for a later publication. It is hoped also that it will be possible

458 HALBERT L. DUNN

to present at a later time an outline of the prenatal growth of the
various internal parts of the brain for which data are now being
collected.

Growth of the central nervous system as a whole

The growth curve of the central nervous system (represented by
the volume of the central nervous system, fig. 2) is analogous
in both its character and its slope to the growth curves of nearly
all the fetal viscera. - The abscissa of this curve is the crown-

TABLE 35
Spinal cord volume and weight
(data of Jackson, ’09, and of Volpin, ’02)

SPINAL-CORD VOLUME (JACKSON, ’09) SPINAL-CORD WEIGHT (VOLPIN, ’02)
RANGE OF
CROWN-HEEL 5 Be, |
mmrans| | crowo-teel [gg Asatte g| Naber | crowncnect | ,Averaee, | umber
cm. cm. cc. cm. grams

Oto 5 sre 0.02225 3

5 to 10 CRC 0.115 3

10 to 15 1253 0.1671 3

15 to 20 hfs 1h 0.3447 5 16.0 0.75 1
20 to 25 22.4 0. 664 4 20.0 La | 1
25 to 30 27.4 0.908 5 28.0 2.0 1
30 to 35 ooaT 1.53 2 32.7 3.0 3
35 to 40 38.5 1.8 1 38.5 3.75 2
40 to 45 44,2 3.3 il 41.8 4.5 3
45 to 50 45.7 6.6 3
50 to 55 51.9 3.04 2 51.0 9.5 3

heel length and represents a simple lineal type of growth. The
value of the central nervous system is therefore a function of the
crown-heel length which has been raised to approximately the third
power. Obviously, this is a typical concave volumetric curve. It
is similar in type to all of the visceral volumetric and ponderal
curves of growth which have been worked out in this laboratory
and of which a summary has been published elsewhere (Scammon,
21). It is also analogous to the curves of fetal body weight
and of weights of body parts which have been expressed in func-
tions of the crown-heel length of the fetus. In order to compare

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 459

the growth of the central nervous system to the growth of other
parts of the body it has been considered as a unit. It is however
a complex of four distinct subtypes of growth, all of which are
dominated by the growth of the bulky cerebral hemispheres.?

Jo}

4

55cm

Fig. 22 Field graph and curve (connecting the average points of each 5-cm.
crown-heel interval) of the per cent which the spinal cord volume forms of the
encephalon volume. Abscissa: total body length in em. Ordinate: spinal
cord in per cent of the encephalon volume. Individual cases indicated by solid
dots. (Data from table 40.)

2In order to compare these four types of growth, ten methods were used: 1) the
various absolute length curves (fig. 29) were calculated against crown-heel body
length (em.) and reduced to a percentage basis; 2) the absolute volume curves
likewise (fig. 30) were taken to crown-heel length and reduced to a per cent basis;
3) the absolute length curves (fig. 33) were calculated for age in fetal months and
reduced to a common per cent; 4) the absolute volume curves also (fig. 36) were
taken to age in fetal months and reduced to a percentage basis; 5) percentage
increments of linear curves (fig. 25) were calculated for 5-em. C H intervals;

460 HALBERT L. DUNN

TABLE 36

Length of the spinal cord
Formula: Spinal cord length (em.) = 10 crown-heel length (em.).467 — 4

(144 cases)
ROWS HE ENGre EPINAL CORD LENGTH CALCULATED DEAE
SPINAL CORD | OBSERVED AND a pone
Maxi: | Mini- LENGTH CALCULATED
Range Mean main ant Mean MEANS
cm. cm. cm. cm. cm. cm. cm.
Oto 5 3.9 2.5 1.4 1.9 1.4 —0.5 4
5 to 10 Coll 4.0 253 3.0 3.6 +0.6 9
10 to 15 12.1 6:2 4.0 4.7 5.4 +0.7 20
15 to 20 16.7 9.7 5.3 7.0 6.9 Oat 14
20 to 25 Ded el Ole UD 8.7 8.6 —0.1 20
25 to 30 26.9 10.7 8.5 9.75 9.6 —0.1 14
30 to 35 32.6 isto3 9.0 10.6 10.9 +0.3 18
35 to 40 37.1 16.5 9.5 12.2 11.8 —0.4 17
40 to 45 42.2 14.0 11.0 12.4 12.8 +0.4 12
45 to 50 48.2 14.8 Pe 7 1325 13.9 +0.4 8
50 to 55 52.2 15.8 13.2 14.5 14.6 +0.1 8
TABLE 37

Calculated spinal cord length at 5-cm. intervals of crown-heel length

INCREMENT IN EACH

CROWN-HEEL LENGTH SPINAL CORD LENGTH 5-CM. INTERVAL

RATIO TO NEW-BORN

cm. cm. cm. per cent per cent

5 2.2 15.5
10 4.6 2.4 109.0 32.4
15 6.4 1.8 39.2 45.0
20 7.9 1.5 23.4 55.6
25 9.2 ive 16.4 64.8
30 10.4 12 13.0 73.3
35 11.4 1.0 9.6 80.3
40 12.4 1.0 8.2 87.3
45 13.3 0.9 7.5 94.0
50 14.2 0.9 6.5 100.0
55 15.0 0.8 5.9 105.6

6) percentage increments of volume curves (fig. 26) were determined for 5-cm.
intervals; 7) percentage increments of linear curves (fig. 27) were calculated for
age in fetal months; 8) percentage increments of linear curves (fig. 28) were
determined for age in fetal months; 9) the various linear formulae of the respective
absolute curves (table 43), and, 10) the volumetric formulae of the absolute
volume curves (table 43) were compared.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 461

The first type, cerebral growth, is characterized by a constant
growth in linear measurements from the second fetal month until
birth, and shows a steady and relatively slow increase in volume

O 5 10 15 LO £5 30 35), = 40 A5 50 55«

Fig. 23 Field graph and curve of the growth of the spinal cord in fetal life
as shown by the spinal cord length. Abscissa: total body length in em. Ordi-
nate: spinal cord length incm. Individual cases indicated by solid dots. Curve
drawn to the formula: Y = (10.0X).467 — 4.0. (Data from tables 36 and 37.)

before the sixth fetal month and a constant but more rapid in-
crease from that time until birth. Brain stem and cord growth, on
the other hand, advances much faster previous to the seventh
fetal month than it does in the last four months of fetal life.
Cerebellum growth, on the contrary, proceeds slowly until the

462 . HALBERT L. DUNN

seventh fetal month and from that time until birth it exceeds all
other growth activity in the central nervous system. The last
type, compound growth, represents merely the combined effect
of two or more of the above types predominated by the mass of
the cerebral hemispheres. ‘These varieties of growth will now be
considered in more detail.
TABLE 38
Combined brain-cord length
Formula: Brain-cord length (em.) = (10 crown-heel length (cm.))-*!4 —4

(137 cases)
BRAIN-CORD | OBSERVED AND On ORaEE
Mee Me LENGTH CALCULATED
Range Mean muni mum Mean MEANS
cm. cm. cm. cm. cm. cm. cm.

OMORS 4.0 4.9 Sea woe O Dall —=O(023 3

5 to 10 Tall On 4.0 4.4 4.9 +0.5 G
10 to 15 12:1 8.5 5.0 6.9 7.8 +0.9 20
15 to 20 16.6 Woh eo 9.8 9.8 0 14
20 to 25 22.6 14.7 LORZ Ze. 12.2 0), 19
25 to 30 26.9 Mayall 12.1 13.8 13.8 0 13
30 to 35 32.8 18.4 14.1 15.4 15.6 — (52 16
35 to 40 37.1 20.2 13.5 17.4 16.9 =(0).5) 17
40 to 45 42.2 ORs 16.1 18.1 18.4 —0.3 11
45 to 50 ASeD | 217 18.8 19.8 19.9 +0.1 9
50 to 55 92.2 23.1 20.4 | 21.3 20.9 —0.4 8

TABLE 39
Calculated combined brain-cord length at 5-cm. intervals of crown-heel length

INCREMENT IN EACH

= v y. D
CROW N-HEEL LENGTH BRAIN-CORD LENGTH 5-CM. INTERVAL

RATIO TO NEW-BORN

cm. cm. em. per cent per cent
5) 3.5 17.2
10 6.6 3.1 88.5 32.3
15 9.2 2.6 39.4 . 45.1
20 11.2 2.0 74 NY 6 54.9
25 13.1 1.9 17.0 64.2
30 14.8 Lid, 13.0 72.5
35 16.3 1.5 10.1 79.9
40 17.8 1.5 Or2 87.2
45 19S 1.3 7.3 93.6
50 20.4 1.3 6.4 100.0
55 21.6 ie” 5.9 106.0

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 463

Cerebral growth. The growth of the cerebrum is characterized
by a steady absolute increment in all of its lmear measurements
from the second fetal month until birth. This can be observed
by the inspection of the curves of the fronto-occipital length and
the temporal diameter (fig. 29, curves I and II, respectively).

4 = 3), "oP et al Pes Geel le T i T — |
cm. ; ‘ ‘ ,

O coe | eines | eee
O 5 {0 15 £O Eby 830 IDO) 45 50° 55cm.

Fig. 24 Field graph and curve of the growth of the spinal cord and brain stem
in fetal life as shown by the brain-cord length. Abscissa: total body length in
em. Ordinate: brain-cord length in em. Individual cases indicated by solid
dots. Curve drawn to the formula: Y = (10.0X)-*!4 —4.0. (Data from tables
38 and 39.)

These curves are both straight lines and lie intermediate to the
vermis cerebelli absolute curves below and to the curve of the
spinal cord above. When calculated against time in fetal months
(fig. 33, curves I and II), the fronto-occipital length and the

(Text continued on page 479)

464 HALBERT L. DUNN

TABLE 40

Ratios of the volumes of the various parts of the central nervous system to the entire
brain volume

RATIO OF:
CROWN-HEEL
ane ae bal BEE rr os eet pee Midbrain | Cerebellum hei hee
cm. ih ae ian | aK Ginnie bs.)
5 to 10 4.70
10 to 15 3.30 aia) 2.90 3.0 87.7
15 to 20 2.40 3.6 2.40 3.0 90.9
20 to 25 1.70 Ded 1.60 3), 1h 93.1
25 to 30 1.50 ee, 1.40 3.0 94.0
30 to 35 1.20 1.9 0.96 De teh 94.2
35 to 40 0.81 ire 0.90 3.8 94.1
40 to 45 0.90 1.6 0.87 4.4 93 .2
45 to 50 0.84 il 4) 0.70 One, 92.0
50 to 55 0.70 1.4 0.67 yas) 91.8
TABLe 41

Comparison or THC WeiGnTs AND VoLUMES
or THE Spina Corp, THE Exmrc Brain AND THe Various Parts or THE Brain
IN THE FetaL Periop

Averact WeicHt (Gm) anpVotume (cc)in Each Ser Intervat (CH)

Spinal Cord
128 cases)

Pons and Medulla

109 cases

Mid brain
(109 cases)

Cerebellum
(115 cases)

Righ! Cerebral Homspher
(115 cases)

|
Left Cerebral Hemisphere
(114 cases)

Dolire Brain
(I44 cases)

Central Nervous Syslem’
6 cases,

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM

TABLE 42.

465

Catcutatep ABSoLuTE AND PERCENTAGE INCREMENTS oF THE Srverat Parts or THE Brain
AND OF THE SPINAL Corp IN Each MontH or THe fetat Periop

CreREBRUM

CEREBELLUM

Pons aND Meputta| Mipprain} Spina Corp

Cerebellum
Volume

Both | [ronto la “ij
Hemispheres occipilal ses
Vine | Lebgin | Demeter

Vermis
Cerebelli
| Reight

Vermis

Cerebelli
Length

Spinal
ord
Length

Pons and
Medulla

Volume

Mid brain
Volume

Pons

Lenglh

a4

| %

1.56

20

C26) 141) 247 2.31
ie

3.03

TABLE 4)

EpintcAt Formutac Representinc Curves or Linetar AND VoLuMeTRIC GROWTH
or THe Central Nervous System nw otHe Terat Pentop

Seeciric FoRMuLAE

Messonevent (ExPonenTiaL)
Fronfo-occipital Lenglh Y= O.1T5X+025
Temporal-lemporal Diameter = Y: 0.138X+ 03
Pons Lenglh Y= 0.0£263X+ 0.16
Colliculi Length Y: 0.015X+04T

inal Cord Length Y: (10.0x) 4674.0
Ban Cord tat Y: (10.0X)?!4-4.0
Tronlo- spinal Length Y- (2.0K)47-2.2
Vermis Cerebelli Length Y= 0.01(X!4!+15.0)
Vermis Cerebelli Height ¥: 0.01(K!>")

Spinal Cord Volume Y= 0.01[(017X)2 7% 11.0]

Pons and Medulla Volume Y= 0.0{|(0.2X)* £7 +200

Y= 0.01{(0.168x)* +120]

Mid Brain Volume
Cerebellum Volume Y+ 0.01[(0.095X)*%+200]
Bight Hemisphere Velume ¥: (0.1X)29
t Hemisphere Volume Y= (0.105x)?
Both Hemispheres Volume Y: (0.12X)>'9
Bneephalon Volume Y: (O.125X)" ler 15
Y= (0.13X)7'9+1.0

Y= (O,)14X)°7442.0

Encephalon Weight
Central Rages Sysen Volume

Type or JORMULA

Y-aX+b

Y=0.01(X%+b)

Y- 0.01[(ax)+ ¢]

Y= (ax)>

Y- (oX)P+ c

Seeciric FORMULAE
(LoeaRritHMic)

Y= O.1T5X+025
Y= 0.138X +03

Y= 0.0263K+0.16

Y: 0.015X+ O47
Y=O.0TX+ 10 /ogX-65
Y= O.1TX +11 /ogX-GT
Y-O0.051K+4/0gX-2.85
Y- 0.01(0.073X+Z.85)>
Y=0.01(.07X+245)>

Y= 1.4(0.02X+031)-O0I3X+0.1
Y=094 (O03K+02)"+ Ol

Type oF FORMULA

Y=aX+b

Y= aX+b ogkK-c

Y=0.01(aX+b)?

Y=a(bX+c)°+dX+e

Y=1.5(0.02X+0:3!)+-O0Z7X+038
Y=0.01(0.020X+1.03)? Y= 0.01 (aX+b)?
Y= (01085X-0.09)* :

Y= (O.108X-0.05)° Yea(bX-c)”

Y=1.1(0.137X-0.07)=0.4X+5.0
se (0.14X+0.2)-X+10.0
|

Y=a(bXtc)-dX+e
Y=1.17(O.137X-0.03)*-1.12K+16.0

466 HALBERT L. DUNN

| ]
0-5 510 {0-15 1520 «62075 «6530 0) = 0055S 540) =4045> 4550 25055em.

Fig.25 <A series of curves illustrating the percentage increments of the various
linear dimensions of the brain, brain parts, and spinal cord. Abscissa: total
body length in em. Ordinates: percentage increments calculated from the
formulae of the respective absolute curves. (Data from tables 14, 16, 20, 22, 26,
30, 32, 37, and 39.) I, percentage increment of fronto-occipital diameter. II,
percentage increment of temporal diameter. III, percentage increment of vermis
cerebelli length. IV, percentage increment of vermis cerebelli height. V, per-
centage increment of pons length. VI, percentage increment of colliculi length.
VII, percentage increment of frontospinal length. VIII, percentage increment
of spinal cord length. IX, percentage increment of brain-cord length.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 467

fe)
05 S10 «(1015 §=61520 0 = 2025) 530 HSH 3H540 )=—40-45 «24550 BO55an.

Fig.26 A series of curves illustrating the percentage increments of the various
volumetric determinations of the brain, brain parts, and spinal cord. Abscissa:
total body length incm. Ordinates: percentage increments calculated from the
formulae of the respective absolute growth curves. (Data from tables 2, 4, 5, 8,
10, 12, 18, 24, 28, and 34.) I, percentage increment of central nervous system
volume. II, percentage increment of encephalon volume. III, percentage in-
crement of entire brain weight by literature. IV, percentage increment of right
hemisphere volume. V, percentage increment of left hemisphere volume. VI,
percentage increment of both hemisphere volume. VII, percentage increment
of cerebellum volume. VIII, percentage increment of pons and medulla volume.
IX, percentage increment of midbrain volume. X, percentage increment of
spinal cord volume.

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 5

468 HALBERT L. DUNN

80

TO

50

£0

0 | id 9) at 5 6 {( 6 9 10mos.

Fig.27 A series of curves illustrating the percentage increments of the various
linear dimensions of the brain, brain parts, and spinal cord when interpreted in
time in fetal months (as determined by Mall’s convention). Abscissa: total
time of fetal life inmonths. Ordinate: percentage increments calculated from
the formulae cf corresponding absolute curves and transposed into time. (Data
from table 41.) I, percentage increment of fronto-occipital diameter. II,
percentage increment of temporal diameter. III, percentage increment of pons
length. IV, percentage increment of colliculi length. V, percentage increment
of spinal cord length. VI, percentage increment of brain-cord length. VII,
percentage increment of frontospinal length. VIII, percentage increment of
vermis cerebelli length. IX, percentage increment of vermis cerebelli height.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 469

0 1 & E) 4 5 6 T 8 9 Omos.

Fig. 28 A series of curves illustrating the percentage increment of the various
volumetric determinations of the brain, brain parts, and spinalcord. Abscissa:
total time in fetal months as determined by Mall’s convention. Ordinates: per-
centage increments calculated from the formulae of respective absolute growth
curves. (Data from tables 2, 4, 5, 8, 10, 12, 18, 24, 28, and 34.) I, percentage
increment of spinal cord volume. II, percentage increment of pons and medulla
volume. III, percentage increment of midbrain volume. IV, percentage in-
crement of cerebellum volume. V, percentage increment of both hemisphere
volume. VI, percentage increment of the encephalon volume. VII, percentage
increment of the volume of the central nervous sytem. VIII, percentage in-
crement of the encephalon weight (cases collected from the literature).

470 HALBERT L. DUNN

0 5 10 15 £0 £5 £910) 35 AO 45 50cm.

Fig.29 A series of curves illustrating the growth (in linear dimensions) of the
brain, brain parts, and spinal cord in per cent of the respective new-born linear
measurements. Abscissa: total body length in em. Ordinates: linear dimen-
sions calculated in per cents of the respective new-born measurements. Data
from the formulae in table 438. I, frontal-occipital diameter. II, temporal diame-
ter. III, vermis cerebelli length. IV, vermis cerebelli height. V, pons length.
VI, colliculi length. VII, frontospinal length. VIII, spinal cord length. IX,
brain-cord length.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 471

100

80

TO

50

AO

30

£0

0 3} 10 15 20 e5 ro.) 35 AO 45 50cm

Fig. 30 A series of curves illustrating the growth in the volumes of the brain,
brain parts, and spinal cord in percent of the respective new-born volumetric
determinations. Abscissa: total body length in em. Ordinate: volumes eal-
culated in per cents of the respective new-born volumetric determinations.
(Data from the formulae in table 43.) I, volume of the central nervous system.
II, encephalon volume. III, encephalon weight by cases from the literature.
IV, right hemisphere volume. V, left hemisphere volume. VI, both hemi-
sphere volumes. VII, cerebellum volume. VIII, pons and medulla volume.
IX, midbrain volume. X, spinal-cord volume.

472 HALBERT L. DUNN

Fig. 31 A series of curves illustrating the growth of the pons, colliculi, and
vermis cerebelli by linear measurements. Abscissa: fetal months (crown-heel
reduced to time by Mall’s convention). Ordinates: lengths of pons, colliculi,
and vermis cerebelli in em. (Data from formulae in table 43.) I, vermis cere-
‘bellilength. II, vermis cerebelli height. III, ponslength. IV, colliculilength.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 473

0 ! ie J A 5 6 G 8 9 10m.

Fig. 32 A series of curves illustrating by linear measurements the growth of
the cerebrum, brain stem, and spinal cord. Abscissa: fetal months (crown-
heel reduced to time by Mall’s convention). Ordinates: lengths of the cere-
brum, brain stem, and spinal cord in em. (Data from formulae in table 43.)
I, fronto-occipital diameter. II, temporal diameter. III, frontospinal length.
IV, spinal cord length. V, brain-cord length.

474 HALBERT L. DUNN

O ! fé, 3 4 5 6 iG 8 9 1Omos.

Fig. 33 A series of curves illustrating the growth (in linear dimensions) of the
brain, brain parts, and spinal cord calculated in per cents of the respective new-
born linear measurements. Abscissa: fetal months (crown-heel length reduced
to time by Mall’s convention). Ordinates: linear dimensions calculated in
per cents of the respective new-born measurements. (Data from the formulae
in table 43.) I, fronto-occipital diameter. II, temporal diameter. III, ver-
mis cerebelli length. IV, vermis cerebelli height. V, pon length. VI, collic-
uli length. VII, frontospinal length. VIII, spinal-cord length. IX, brain-
cord length.

GROWTH

300 :
ce ey /
yf
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250 / /
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[fi
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/ a
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200 i rf} j
ip EL
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i 1/
150 / hi; ve
rif ;
ida wih
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ee
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Fig. 34 A series of curves illustrating the growth of the cerebrum and _ cere-

bellum as shown by volumetric determinations.
heel length reduced to time by Mall’s convention).
encephalon, right hemisphere, and cerebellum.
II, right hemisphere volume.

I, encephalon volume.
IV, central nervous sytem volume.
the literature).

OF THE FETAL CENTRAL NERVOUS SYSTEM

475

Abscissa: fetal months (crown-
Ordinates: volumes of the
(Data from formulae in table 43.)
III, cerebellum volume.

V, encephalon weight (data collected from

VI, both hemisphere volume.

476 HALBERT L. DUNN

on
|

(0) —-———
0 | g 3 4 5 6 iG 8 9 = [Omos.

Fig. 35 A series of curves illustrating the growth of the cerebellum, pons and
medulla, midbrain, and spinal cord. Abscissa: fetal months (crown-heel re-
duced to time by Mall’s convention). Ordinates: volumes of the cerebellum,
pons and medulla, midbrain, and spinal cord. (Data from the formulae in table
43.) I, cerebellum volume. II, pons and medulla volume, III, midbrain
volume. IV, spinal cord volume.

GROWTH ' OF THE FETAL CENTRAL NERVOUS SYSTEM 477

1Omos.

i a 4 5 6 16 8 2

Fig. 36 A series of curves illustrating growth of the brain, brain parts, and
spinal cord in per cent of the respective new-born volumetric determinations.
Abscissa: fetal months (crown-heel reduced to time by Mall’s convention).
Ordinates: per cents of volumes as compared with the respective new-born volu-
metric determinations. (Data from the formulae in table43.) I, encephalon
volume. II, cerebral hemispheres volume. III, cerebellum volume. IV, pons
and medulla volume. V, midbrain volume. VI, spinal cord volume.

478

HALBERT L. DUNN

MBL

G7 8

Fig. 37. A series of midsagittal tracings of the brain and brain stem. Each
figure represents an average brain, m each 5-cm. crown-heel interval. A, ca.
4em. CH length. B, ca. 7.5 em. CH length. C,ca.2.5 em.C Hlength. D, ca.

17.5 cm. C H length. E, ca. 22.5 cm. C H length. F, ca. 27.5 cm. C H length

G, ca. 32.5em.C Hlength. H, ca. 37.5 em.CH length. I, ca. 42.5 em. C H length

J, ca.47.5 em. C H length. K, ca. 52.5 em. C H length.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 479

temporal diameter are practically identical. They occupy the
same characteristic intermediate position between the cerebellum
and the brain stem and spinal cord absolute growth curves. The
rate of growth of the fronto-occipital diameter, when calculated
from 5-em. C H intervals (fig. 25, curves I and If) and also when
calculated for age in fetal months (fig. 27, curves I and IJ), por-
trays an intermediate percentage increment curve lying between
the cerebellum above and the brain stem below. The formulae
of the fronto-occipital length and the temporal diameter (table
43) likewise show a characteristic grouping of the hemispheres to
the general formula: Y = aX + b, in which Y is the fronto-
occipital length or the temporal diameter in cm., X is the crown-
heel length in em., and a and b are constants determined for the
two formulae.

Cerebral growth in volume shows a steady and relatively slow
increase before the sixth fetal month and then a constant but more
rapid increase from that time until birth. This is indicated by
the right hemisphere volume, left hemisphere volume, and both
hemisphere volume (figs. 4, 5, and 6, respectively). When cal-
culated against crown-heel length and reduced to percentage
basis (fig. 30), the right hemisphere volume, left hemisphere
volume, and the volume of both hemispheres (curves IV, V, and
VI, respectively), as well as the absolute curves of the central
nervous system and the total brain volume (curves I and II, re-
spectively), fall practically upon one another. The cerebellum
volume curve lies below and the brain stem and spinal cord vol-
ume curve above the curves dominated by the factor of cerebral
growth. Upon inspection of the curve of both hemisphere
volume when calculated against time in fetal months and reduced
to a percentage basis (fig. 36, curve II), a similar central grouping
between the cerebellum volume, the brain stem volume, and
spinal cord volume is observed. ‘Turning to the figures upon the
rate of growth of the cerebrum, which is determined for 5-cm.
intervals (fig. 26, curves IV, V, and V1) and for age in fetal months
(fig. 28, curve V), one observes the unswerving tendency of the
cerebral growth to proceed faster before the sixth month and
slower thereafter.

480 HALBERT L. DUNN

Fig. 38 A series of midsagittal tracings (of the specimens shown in figure 35)
drawn to a standard fronto-occipital length. A, ca. 4 em. C H length. B, ea.
7.5 em. C H length. C, ca. 12.5 em. C H length. D, ca. 17.5 em. C H length.

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 481

ia

ite

Uy

Px ose
G7»
y »

E, ca. 22.5 em. C H length. F, ca. 27.5 cm. CH length. G, ca. 32.5 em. C H
length. H, ca.37.5em.C H length. I, ca. 42.5 cm. CHlength. J, ca. 47.5 cm.
CH length. K, ca. 52.5 cm.C H length.

482 HALBERT L. DUNN

The volumetric formulae of the right hemisphere volume, left
hemisphere volume, and both hemisphere volume (table 43) are
all expressed by the simple formula:

Y = (aX)

in which Y is the value to be determined in cc.; X, the crown-
heel length in cm., and a and b are constants for the respective
volumes. |

Brain stem and spinal cord growth. The growth of the brain
stem and spinal cord, as shown by curves of absolute linear di-
mensions and absolute volumes, represents a second definite type
of growth in the central nervous system. It is characterized by
a relatively rapid absolute and relative increase previous to the ©
sixth fetal month and a slow increase during the last four months
of intra-uterine life.

The linear determinations showing this type of growth are the
frontospinal length, the spinal cord length, and the brain-cord
length. When plotted against crown-heel length in centimeters
and reduced to percentages of new-born values (fig. 29, curves
VII, VIII, and IX) these three curves approximate one another
and ascend more rapidly prior to the sixth fetal month than they
do in the last four months. They are distinctly separated from
the cerebral type of growth and, when calculated in fetal months
and reduced to a per cent of the new-born (fig. 33, curves VII,
VIII, and IX, respectively), they show even more clearly the
almost identical path which they all follow. When calculated to
time, they likewise show a rapid increase to the fifth fetal month
and a comparatively slow one thereafter. The rates of growth of
the brain stem and the spinal cord as shown by these three linear
determinations can be observed equally well when taken for
5 cm. intervals: (fig. 25, curves VII, VIII, and IX, respectively),
or when calculated for age in fetal months (fig. 27, curves VII, V,
and VI, respectively). The brain stem and spinal cord grow at
a slower rate both relatively and absolutely than do all other
brain parts during the last four months of fetal life.

The classification indicated by comparison of the absolute
linear curves is substantiated by a glance at their respective

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 483

formulae (table 43). The growths of most of the group are ex-
pressed by the general empirical formula:

Y = (ax)* —e

in which Y is the brain-part value in em., X is crown-heel length
in em., and a, 6, and c are constants determined separately in
each case for the frontal spinal length, spinal cord length, and
the brain-cord length.

Two other straight-line measurements, one of the colliculi
length and the other of the pons length, do not follow the typical
brain stem and spinal cord type of growth for some unknown
reason. ‘They are straight-line curves and evidently express the
growth of some specific factor rather than the growth of the
brain stem of which they are a part. ,

The brain stem and cord growth are portrayed also in the
curves of the spinal cord volume, the pons and medulla volume,
and the midbrain volume. When calculated against crown-heel
length (fig. 30, curves X, VIII, and XI, respectively), these
volume curves show a relatively rapid absolute growth previous
to the sixth month and a slow growth in the last four months of
fetal life. They also approximate each other and fall definitely
above the cerebral type of growth curve. They are practically
identical when calculated against fetal months and reduced to a
per cent of the new-born value (fig. 36, curves VI, IV and V, re-
spectively). When the percentage increments of the spinal cord
volume, the pons and medulla volume, and the midbrain volume
are compared (either when caleulated for 5-em. C H intervals
(fig. 26, curves X, VIII, and IX, respectively) or for age in fetal
months (fig. 28, curves I, IJ, and III, respectively)), the distinctive
types of the brain stem and spinal cord growthare observed. They
show a slow rate of relative growth in the third month, which in-
creases three-fold in rapidity in the fourth month. From this time
until birth the growth rate decreases in rapidity so that these por-
tions show a less rapid relative increase in the last three fetal
months than does any other brain part.

The classification is indicated also by volumetric formulae
(table 48) which can be expressed by the general equation:

Y = 0.01 [(aX)*+ ¢|

A484 HALBERT L. DUNN

in which Y is the value desired in cc., X is the crown-heel in cm.,
and a, b, and c, are constants determined for the spinal-cord
volume, the pons and medulla volume, and the midbrain volume.

Cerebellum growth. Cerebellum growth, as shown by linear
and volume absolute curves, proceeds slowly until the sixth fetal
month, and from that time until birth outstrips all other growth
activities in the central nervous system.

The lineal determinations expressing this type of growth are
the vermis cerebelli length and the vermis cerebelli height (fig.
29, curves III and IV). If plotted against crown-heel length
and reduced to per cents of their new-born values, they lie close
together and ascend slowly at first and then more rapidly to birth
in very shallow concave curves. They are decidedly different
from all other straight-line absolute curves of the nervous system,
resembling volumetric curves rather than lineal progressions.
They show a similar definite classification when calculated against
fetal months (fig. 33, curves III and IV). The rate of growth
of the cerebellum, as shown by the percentage increment curves
of the vermis cerebelli length and the vermis cerebelli height
either when calculated for 5-em. C H intervals (fig. 25, curves
III and IV) or for age in fetal months (fig. 27, curves VIII and
IX), reflect the cerebellum type of growth. During the last four
fetal months the percentage increment of the vermis cerebelli
length and the vermis cerebelli height is higher by 8 to 10 per
cent than all other linear percentage increment curves of the cen-
tral nervous system.

A consideration of the vermis cerebelli length and the vermis
cerebelli height formulae (table 43) shows the volumetric tendency
of these linear formulae. They can be expressed by the general
empirical formula:

Vi = Oi Xe)

in which Y is the value desired in em., X is crown-heel in cm.,
and a and 6 are constants determined for both the vermis cere-
belli length and the vermis cerebelli height.

The curve of the volume of the cerebellum is striking in char-
acter and demonstrates the slow growth of the structure prior

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM A85

to the sixth fetal month and the tremendously rapid increment
from that time until birth. When calculated against crown-heel
length (fig. 30, curve VII), and also when calculated against time
in fetal months (fig. 36, curve III), the distinct character of the
cerebellum volume is apparent. The formula of the growth
curve of the cerebellum (table 48) is also characteristic, although
it falls with the same general formula as the brain stem and spinal-
cord volume, viz.:

Y = 0.01 [(aX)e + ¢]

However, it will be noticed that the b factor is 4.9 in the case of
the cerebellum volume. This means that while the spinal cord
volume, midbrain volume, and the pons and medulla volume are
increasing practically as the cube and therefore like typical
volume curves, the cerebellum volume is growing at a rate
approaching the ninth power of the body length. The percentage
increment of the cerebellum volume both when calculated for
5-em. C H intervals (fig. 26, curve VII) or for age in fetal months
(fig. 28, curve IV), also demonstrates a specific type of growth
and shows the great increment of the part in the later fetal
months as indicated by other measurements of cerebellum
growth.

Compound growth. Compound growth indicates a summation
of two or all types of growth in the central nervous system. It
is typified by the volume and weight of the encephalon and the
central nervous system volume. It is dominated by the growth
of the cerebrum, since the cerebrum ranges between 85 and 95
per cent of the encephalon throughout fetal life. The rates of
growth of the compound growth curves, as shown by percentage
increments calculated for 5-cm. intervals (figs. 26, curves I, II,
and III) and for fetal months (fig. 28, curves VI, VII and VIII),
approximate the growth rate of the cerebrum. However, it
will be noticed that in the early fetal months the rate of growth
is not quite so high, due to the slow development of the other
brain parts at this period.

The volumetric formulae (of the type here presented) of the
central nervous system, the entire brain volume, and the en-

486 HALBERT L. DUNN

cephalon weight require a third constant in their expression, while
the volume of the cerebral hemispheres can be expressed mathe-
matically with two constants. The general empirical formula
of compound growth can be expressed:

y 2 axe

in which Y is the volume of the encephalon or entire central
nervous system in cc., X is the crown-heel length in cm., and a,
b, and ¢ are constants separately determined for the entire brain
volume and the central nervous system volume.

The significance of the relation between lineal and volumetric
dimensions in the growth of the parts of the brain. The classifica-
tion of the types of growth of the central nervous system is shown
by both the volume and linear curves. For instance, both the
volume curve of the cerebellum and the linear measurements of the
vermis cerebelli length and the vermis cerebelli height indicate
the characteristic cerebellum type of growth. It is logical that
there should be a definite relation between the linear and the
volume measurements of a brain part. It is also evident that
the length of a brain part when it is cubed should form a curve
identical in type and constant in relationship to the correspond-
ing volume curve, provided that no additional factor, other than
that which influences the growth in the linear dimensions, influ-
ences the growth in volume. Such a relationship actually exists
between the formulae of the linear cerebellum curves and the
formula of the cerebellum curve of volume. The formula for the
cerebellum length when cubed is practically identical to the
formula of the cerebellum volume. This relationship is seen
somewhat better when those values are based on the cube of the
body length as modified by three constants rather than by a
formula based on the body length raised to a fractional exponent.
The first formula is expressed below:

Cerebellum volume (cc.) = [0.01 (0.073 Vermis cerebelli
length (em.) + 2.85)3]§
Likewise, the formula of the cerebellum height when cubed and

multiplied by the constant 2.13 is practically identical with the
formula of the cerebellum volume:

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 487

Cerebellum volume (ce.) + [0.01 (0.07 Vermis cerebelli
height (em.) = 2.45)*]?

These indicate that the cerebellum volume is growing at relatively
the same rate as are its vertical and horizontal diameters. The
growth of the cerebellum must be controlled, therefore, by factors
which act alike upon the entire cerebellum.

The formulae of the fronto-occipital length and the temporal
diameter when cubed bear no relation to the cerebrum volume
which may be expressed by a simple constant, and the same
holds true of the spinal cord length when cubed. It is obvious,
therefore, that in all other brain parts except the cerebellum
there must be more than one factor influencing growth which act
differently on the volumetric and linear growth.

The per cent of the brain parts and the spinal cord of the encepha-
lon. The per cent which the various brain parts and the spinal
cord form of the encephalon at each fetal month can be observed
by a glance at table 40 and at figures 7, 11, 15, 18, 22, and 38.
Table 40 tabulates the per cent which a respective volume of a
brain part or of the spinal cord forms of the encephalon. Figures
7, 11, 15, 18, and 22 demonstrate graphically the per cent whichthe
pons and medulla, midbrain, and the spinal cord, respectively,
form of the encephalon volume. Figure 38 represents a series of
composite midsagittal sections which are based upon average
measurements and which represent therefore a typical fetal brain
for the 5-em. C. H. interval indicated. For purpose of compari-
son, a magnification of each individual figure was used. To
obtain this magnification, the ratio of the respective fronto-
occipital diameters to the fronto-occipital diameter of the new-
born was used. The detailed results of both figures and the table
have been presented in the body of this paper. Taken in toto,
the evidence shows conclusively: first, the high percentages which
the pons and medulla, midbrain, and the spinal cord volumes
form of the encephalon volume from the second to the fifth fetal
month; second, the increased percentages which the cerebellum
volume forms of the encephalon volume in the last three fetal
months, and, third, the predominating percentages which the

A488 HALBERT L. DUNN

volume of the cerebral hemispheres form of the encephalon
volume at all times, rising to its maximum per cent (94.2 per
cent) in the sixth fetal month.

SUMMARY

I. The growth of the central nervous system in the fetal period
is similar in general character to the growth of the other viscera
and of the major parts of the body in this period.

II. An analysis of data on the volume and dimensions of the
central nervous system in the fetal period shows four distinct
subtypes or varieties of growth. These are:

1. The cerebral subtype, which is characterized by, a) a steady
and relatively slow increase in volume from the second to the
beginning of the sixth fetal month and a constant and more rapid
increase from this time to birth, and, b) by a steady and constant
growth in linear dimensions from the second fetal month to
birth.

2. The brain stem and cord subtype, which shows a much more
rapid growth from the second to the end of the fifth fetal month
than it does in the last five months of fetal life.

3. The cerebellum subtype, which proceeds very slowly from
the second to the end of the fifth fetal month and then increases
tremendously from the sixth month to birth.

4. The compound subtype, which represents the combined
effect of two or three or all of the above varieties, predominated
by the cerebral subtype.

Ii. The factors which control the various subtypes of growth
in the central nervous system may influence the volumetric and
linear determinations of a respective brain part in two ways:
1) the factors may act differently upon the volumetric and linear
values as they do in the case of the cerebral growth, brain stem,
and cord growth, and in compound growth, or, ,2) they may
act in the same manner as they do in the case of the cerebellum
subtype of growth.

IV. An estimation of the percentages which the cerebrum,
cerebellum, and the brain stem form of the encephalon at the
various periods of fetal life offers a means of comparison of the

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 489

relative growth of these parts. The percentages which the brain
stem and the spinal cord form of the encephalon are relatively
high from the second to the fifth fetal month; the per cent which
the cerebellum form of the encephalon is at its height in the last
three fetal months, while the per cent which the hemispheres form
of the encephalon reaches its maximum in the sixth fetal month.
V. The four subtypes of growth of the central nervous system
may be clearly expressed by a classification of the empirical for-
mulae of the growth curves of the various parts as follows:
1. Cerebral growth:
a. General linear formula: Y = aX + 6.
b. Specific linear formulae:
a’. Fronto-occipital diameter: Y = 0.175 X + 0.25.
b’. Temporal diameter: Y = 0.188 X + 0.3
c. General volumetric formula: Y = (aX).
d. Specific volumetric formulae:
a’. Right hemisphere volume: Y = (0.1 X)*.
b’. Left hemisphere volume: Y = (0.105 X)*-%.
c’. Both hemisphere volume: Y = (0.12 X)*1°.
2. Brain stem and cord growth:
a. General linear formula: Y = (aX) — ¢.
b. Specific linear formulae:
a’. Fronto-spinal length: Y = (2.0 X)-47 — 2.2.
b’. Spinal cord length: Y = (10.0 X)-4*7 — 4.0.
c’. Brain cord length: Y = (10.0 X)-*4 — 4.0.
c. General volumetric formula: Y = 0.01 [(aX)® + ¢].
d. Specific volumetric formulae:

a’. Spinal cord. volume: Y = 0.01 [(0.17 -X)?-
+ 11.0].

b’. Pons and medulla volume: Y = 0.01 [(0.2 X)?-57
+ 20.0].

c’. Midbrain volume: Y = 0.01 (0. 168 X)2-55 + 12.0].
3. Cerebellum growth:
a. General linear formula: Y = 0.01 [X¢ + bj.
b. Specifie linear formulae:
a’. Vermis cerebelli length: Y = 0.01 [X!-* + 0.15].
b’. Vermis cerebelli height: Y = 0.01 [X'-37].

490 HALBERT L. DUNN

c. General volumetric formula: Y = 0.01 [(aX)® + cl.
d. Specific volumetric formula:
a’. Cerebellum volume: Y = 0.01 [(0.095 X)4:9
+ 20.0].
4. Compound growth:
a. General volumetric formula: Y = (aX) + ce.
b. Specific volumetric formulae:
a’. Encephalon volume: Y = (0.125 X)?-16 + 1.5.
b’. Brain and spinal cord volume:
Y = (0.114 X)3-4 + 2.0.
c’. Encephalon weight by literature:
Y=" (O13) Xe e ieeal0:

(In all of the above formulae, Y is the volume or length of the
part under consideration (in em. or eec.), X is the crown-heel or
total length of the body in em., and a, 6, and ¢ are empirically
determined constants.)

BIBLIOGRAPHY

ARNOVLJEVIC, S. 1884 Das Alter, die Gréssen und die Gewichtsbestimmungen
der Fétalorgane beim menschlichen Fétus. Diss., Miinchen.
Biscuorr, E. 1863 Hinige Gewichts und Trockenbestimmung der Organe dese

menschlichen Kérpers. Zeitschr. f. rationelle Med., III Reihe, Bd. 20.

Biscuorr, T. L.W. 1880 Das Hirngewicht des Menschen. Bonn.

Boyp, R. 1861 Tables of the weights of the human body and internal organs
in the sane and insane at various ages, arranged from 2614 post-mor-
tem examinations. Phil. Trans. Roy. Soc. London, vol. 151, pt. 1,
pp. 241-252.

Branpt, EH. 1886 Das Alter, die Gréssen und Gewichtsbestimmungen der
Foetalorgane beim menschlichen Foetus. Diss., Miinchen, 30 pp.

Bucustas. H. 1884 (Material on the question of the weight and volume of the
brain.) (Russian Diss. St. Petersburg.)

Cauxins, L.A. 1921 The growth of the external dimensions of the human body
in the fetal period and its expression by empirical formulae. Report
of meeting of Am. Assoc. of Anat., Mar. 24-26, 1921.

DANIELBEKOFF, A. 1885 Data on the weight and the volume of the brain
and the spinal cord in children of both sexes under one year of age.
(Russian.) Diss., St. Petersburg, 24 and II pp.

Dunn, H. L. 1921 The growth of the brain and the spinal cord in the human
fetus and its expression by empirical formulae. Anat. Rec., vol.
21, p. 55 (Abstract).

Gigse, 1898 Diss., St. Petersburg. (Quoted from Gundobin; Die Besonder-
heiten des Kindesalters, pp. 481 et seq.)

GROWTH OF THE FETAL CENTRAL NERVOUS SYSTEM 491

Hanpmann, E. 1906 Uber das Hirngewicht des Menschen auf Grund von
1414 im pathologischen Institute zu Leipzig vorgenommen Hirnwig-
ungen. Arch. f. Anat. u. Physiol., Anat. Abt.

Hatar, S. 1911 An interpretation of growth curves from a dynamical stand-
point. Anat. Rec., vol. 5. pp. 373-382.

HrouicKa 1906 The physical changes in human and other brains collected
under different conditions and preserved in various formalin prepara-
tions. Proc. U.S. Nat. Museum. vol. 30, pp. 245-320.

Huscuke, E. 1854 Schaedel, Hirn und Seele des Menschen und der Thiere
nach Alter, Geschlecht und Race. Dargestellt nach neuen Methoden
und Untersuchungen. Jena.

Jackson, C.M. 1909 On the prenatal growth of the human body and the rela-
tive growth of the various organs and parts. Am. Jour. Anat., vol. 9,
pp. 117-165.

Kine, H.D. 1910 The effects of various fixatives on the brain of the albino rat,
with an account of a method of preparing this material for a study of
the cells in the cortex. Anat. Rec., vol. 4, pp. 213-239.

MicuaeEuts, P. 1906 Das Hirngewicht des Kindes. Monatsschr. f. Kinder-
heilk., Bd. 6, 8. 9-26.

Prister, H. 1903 Zur Anthropologie des Riickenmarks. Neurologisches
Centralblatt. Bd. 22, 8S. 757-762; 819-824. ;

RupINGER 1877 Vorliufige Mittheilungen itiber die Unterschiede der Gross-
hirnwindung nach dem Geschlecht beim Foetus and Neugeborenen
mit Beriicksichtung der angeborenen Brachycephalie und Dolicho-
cephalie. Beitr. zur Anthropol. u. Urges. Bayerns, Bd. 1, S. 24.

RupotpH, O. 1914 Untersuchungen iiber Hirngewicht, Hirnvolume, und
Schidelkapazitit. Beitr. z. path. Anat., Bd. 58.

Scammon, R. E. 1921 On the growth in weight of the human body and its
various parts and organs in the fetal period, and its expression by
empirical formulae. Anat. Rec., vol. 21, p. 79 (abstract).

Scuuttz, A. H. 1919 Changes in fetuses due toformalin preservation. Am.
Jour. Phys. Anthrop., vol. 2, no. 1, pp. 35-41.

Vattorta, F. 1909 Richerche sullo svilippo dei visceri del feto. La indi-
vidualita nel neonato. Ann. Ostet. e Ginécol., I, XX XI, 673-718.

VouriIn, L. L. 1902 (Weight determinations on the growth of the brain in
children.) Diss., St. Petersburg, 110 pp. (Russian). (Rev: Jahresb.
f. Anat. u. Ent., 1903, Abth. 3, S. 640-645. )

We tcKER, H., uND Branpt, A. 1903 Gewichtswerte der Kérperorgane beidem
Menschen und den Tieren. Arch. f. Anthropol., Bd. 28.

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 5

Resumen por los autores 8. R. Detwiler y Henry Laurens
Estudios sobre la retina.
Histogénesis de las células visuales en Amblystoma

El estado inicial del desarrollo de la célula visual es la pro-
duccién de una yema protoplasmica y un glébulo acromatico
claro en las células de la capa nuclear externa. El glébulo, que
se transforma en el paraboloide del segmento interno, parece
ser de origen citoplasmico y no de origen nuclear, segin ha indi-
cado Cameron. En los estados tempranos del desarrollo de las
células visuales pueden percibirse masas de granulos fuertemente
tefiidos en las preparaciones tenidas por la hematoxilina férrica.
Estos grdinulos contribuyen principalmente a la formacién del
elipsoide del segmento interno y del material granular del externo.
En nuestras preparaciones no existe prueba alguna que indique
su origen como productos del pigmento ingerido de la capa
epitelial conforme supone Cameron. Pueden representar pro-
ductos de la transformacion de las mitocondrias.

En los estados tempranos del desarrollo las células visuales son
todas ellas cénicas, coincidiendo de este modo con las condiciones
caracteristicas de la retina de los anfibios en vias de desarrollo
(Cameron y Bernard). Los bastones y los conos se desarrollan
mis tarde a expensas de las células visuales primitivas no espe-
cializadas. Los bastones no aparecen en su forma definitiva
hasta un periodo relativamente tardio. Las dos clases de ele-
mentos visuales cénicos son consideradas como células visuales
de especializacién baja las cuales se desarrollan, respectivamente,
en los conos y bastones caracteristicos mediante diferenciacién
divergente. Esta interpretacién no apoya la hipotésis enun-
ciada por Cameron. Los dobles conos aparecen temprano
durante el desarrollo. La ausencia de figuras mitésicas durante
este periodo indica que nacen de la fusién de elementos sencillos
mas bien que por divisién de un solo elemento.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHORS’ ABSTRACT OF THIS PAPER ISSUED BY
THE BIBLIOGRAPHIC SERVICE, NOVEMBER 28

STUDIES ON THE RETINA

HISTOGENESIS OF THE VISUAL CELLS IN AMBLYSTOMA

S. R. DETWILER AND HENRY LAURENS
The Anatomical Laboratory, Peking Union Medical College, Peking, China, and
the Osborn Zoological Laboratory, Yale University

THIRTEEN FIGURES
INTRODUCTION

The anatomical features of the visual cells in larval Amblys-
toma have been described and figured by Laurens and Williams
(717). According to this account, the retina in larvae of 30 mm.
possesses characteristic rods and cones in the approximate pro-
portion of four to three. In their figure (p. 76) and in figure 11
in this paper the identity of these elements is clearly displayed.
The rods are quite large with typical cylindrical outer segments
and pear-shaped nuclei, the latter of which are considerably at-
tenuated on their inner pole. The cones, of which there are three
kinds, are much smaller elements characterized by the possession
of conical outer segments, long myoids, and more or less oval
nuclei. These nuclei are more deeply situated than are those of
the rods, which typically comprise the outer cell layer of the
external nuclear layer, and which project through the external
limiting membrane for variable distances.

In looking over some sections of the retinae of young larvae,
our attention was attracted to the striking similarity of the
visual cells, all of which possessed conical outer segments and oval
nuclei, characteristics typical of cones. The general impression
was received that in early stages of development the visual cells
are all of one type (cone-like), which later differentiate into the
two distinct categories of elements which characterize the fully
developed retina.

493

494 S. R. DETWILER AND HENRY LAURENS

From our observations several developmental possibilities were
- suggested: first, if all of these typical cone-like elements are to be
regarded as cones, then the rods, which do not appear in their
distinctive form until later, represent transformation products of
cones; secondly, that some of these elements functionally repre-
sent rods even though possessing anatomical features character-
istic of cones; and, thirdly, that all of these cone-shaped elements
represent indifferent visual cells and the characteristic rods and
cones come about by divergent differentiation of a visual cell of
low specialization.

In general, the rods are regarded as the more primitive type of
visual cell (Graham Kerr, 719, and Parsons, 715) and the cones
are regarded as specialized rods. If this be true, their appearance
in ontogenetic development might be expected to antedate that
of the cones—a condition just opposite to that which is suggested
in the developing retina under consideration. The idea that
cones represent specialized rods does not seem borne out by the
condition found in many reptiles in which only cones are pres-
ent (Detwiler, ’16). If they do represent specialized rods, then
the early developing visual cells in pure cone retinae should have
to be regarded as rods—a condition which seems very unlikely.

According to Leboueg (’09), Magitot (710) and Seefelder (’10),
the two kinds of visual cells in the human retina develop simul-
taneously, and are mainly distinguishable by the fact that the
axis of the diplosome (the centrosomes) is perpendicular to the
long axis of the cone and parallel to the long axis of the rod. This
is not entirely borne out in Seefelder’s illustration which is re-
produced in figure 12. The cones would also seem to be dis-
tinguishable by their much larger size. Cajal (’11) says that cones
and rods evolve in essentially the same manner and it is difficult
to distinguish them at the beginning (see also Fiirst, ’04).

Bernard (’03) and Cameron (’05 and ’11), who studied the de-
veloping retina in amphibia, conclude that cones represent early
stages in the formation of rods. Their conclusions are based on
the fact that in early stages of the developing retina only conical
elements are to be seen, while in later stages typically shaped
rods predominate,

VISUAL CELLS IN AMBLYSTOMA 495

The difficulty in accepting the conclusions of Cameron and
Bernard lies in the fact that the cone-like elements in the early
stages, which they call cones, may functionally represent rods
even though possessing the anatomical configuration characteris-
tic of cones. The variability in the shape of the visual cells has
made absolute rod and cone distinction difficult in some cases.
We have already referred to this difficulty in an earlier paper
(Laurens and Detwiler, ’21, p. 225).

From our first survey of the developing retina in Amblystoma
the conditions appeared to bear out the conclusions of Cameron
and Bernard. However, more critical examination has shown
that, although all the elements are cone-like in early stages of
development, they possess characteristic features by which they
ean be divided into two groups, one of which can be readily iden-
tified as the progenitors of the definitive rods, the other of the
cones. It seems hardly wise to regard the forerunners of the
rods as functional cones even though possessing certain cone-
like features.

METHODS

Our observations were made upon the retina of Amblystoma
punctatum. The larvae were preserved in corrosive sublimate-
acetic fixing fluid at successive twenty-four-hour intervals for a
period of twenty-six days following the tail-bud stage of develop-
ment. From this period on, specimens were fixed at successive
intervals of from three to five days. Sections were cut 8» in
thickness and stained in Ehrlich’s haematoxylin and erythrosin,
iron haematoxylin and eosin, and in Held’s molybdic acid haema-
toxylin.

OBSERVATIONS

The visual elements make their initial appearance in retinae
of approximately eleven days after the tail-bud stage of develop-
ment. They are first seen in the posterior portion of the optic
cup, just above the point of entrance of the optic nerve—a region
corresponding to the optical axis of the eye (fig. 1). Prior to this

141] references to ages are based upon days after the tail-bud stage of develop-
ment. ;

496 S. R. DETWILER AND HENRY LAURENS ‘*

period, the nuclei of the external nuclear layer show numerous
mitoses.

The first step in the formation of the visual cells is the simul-
taneous appearance of a protoplasmic bud and a small achromatic

PED Ve

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wees eS
SS.

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se STA ay } oo
Es R a
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Fig. 1 Transverse section of optic cup and lens in an embryo of 11 days after
the tail-bud stage of development. ON, optic nerve; PE, pigment epithelium;
R, retina; V.C, developing visual cells. XX 275.

globule on the pole of the nucleus applied to the external limiting
membrane (fig. 2, Aand B). In sections stained with iron haema-
toxylin, groups of deeply stained granules are usually seen just
distal to the globule. The protoplasmic buds appear to be pushed
through the external limiting membrane.

VISUAL CELLS IN AMBLYSTOMA 497

The conditions in retinae of forty-eight hours further develop-
ment are seen in figure 3. The protoplasmic buds have become
larger and assume a blunt conical shape. The globules occupy
the proximal portion of the protoplasmic mass, whereas distal to
them occur masses of deeply staining granules similar to the con-
ditions shown in figure 2.

As regards the origin of the clear achromatic globule, Cameron
claims that it is extruded from the nucleus. He supports his
contention by observations which both he and Bernard made in

Fig. 2 Developing visual cells from the retina in an embryo of 11 days after
the tail-bud stage of development. X 1040.

Fig. 3. Developing visual cells from the retina of an embryo of 13 days after
the tail-bud stage. XX 1040.

which they found a collapsed condition of the nucleus subsequent
to the supposed ‘sudden’ discharge of its achromatic material.
This view is contradicted by Leplat (713). Graham Kerr (’19)
states that the ‘fatty’ globule, which stains black with osmic
acid, has its origin in the protoplasmic bud from the primitive
visual cells.

Cameron attributes to this clear globule the power of ingest-
ing pigment from the hexagonal pigment cells, and the intensely
stained granules which he found on the summit of this globule
(figs. 2 and 3) he claims to be the pigment ingested from the pig-

498 S. R. DETWILER AND HENRY LAURENS

ment layer. Because of the fact that only the merest trace of
cytoplasm can be shown to envelop the nucleus of the primitive
visual cell, Cameron claims that the visual element cannot be of
cytoplasmic origin, and he thus believes that it is compositely
developed from the nucleus and from the ingested pigment of the
epithelial layer.

It seems to us that, even if pigment ingestion can be really
observed, the fact must not be overlooked that pigment inges-
tion could not take place unless there were cytoplasm present to
ingest it. Cameron holds that the clear globule does the ingest-
ing, but it so happens that the heavily stained granules which he
saw, as well as those which we have observed, do not lie within
the globule, but in the cytoplasm beyond it (fig. 3). Cameron
himself remarks, ““ . . . . and the product of the ingestion
of the pigment becomes deposited upon the summit of the glob-
ule in the form of a substance which stains intensely with iron-
alum-haematoxylin. In this manner, layer after layer becomes
superimposed until all the visual elements without exception now
assume the shape of cones, with the achromatic globules, much
reduced in size, situated in their bases. ”’

From our observations the origin of the globule cannot be
stated decisively, but whenever it is present, it is always sur-
rounded by cytoplasm and the two seem to appear simulta-
neously. In many initial developing cases, cytoplasmic buds can
be seen in which there is no globule and yet they contain the
heavy granules which are shown in figures 2 and 3. That these
granules should represent products of epithelial pigment seems
very doubtful from our observations. If they are ingested under
the chemotactic influence of the globule, as Cameron says, then
visual cell buds which lack the globule should be devoid of this
deeply staining substance. Moreover, the pigment needles of
the hexagonal cells are not stained nearly so heavily as are those
in the visual cell buds. The former are brownish and only appear
black when heavily massed. The granules in the visual cell bud
are jet black in our preparations, even when appearing singly.
Our observations suggest that the heavily stained granules are
elaborated in the cytoplasm of the visual cell and that the globule

VISUAL CELLS IN AMBLYSTOMA 499

comes about through partial vacuolization of the cytoplasm.
Occasionally, small refractive granules can be seen in the clear
globule. These may represent ingested pigment, but the heavily
stained ones situated distal to the globule and which comprise a
large part of the visual cell, certainly give no indication of origi-
nating from the pigment epithelium.

Leplat (713) has investigated the réle of mitochondria in con-
nection with the development of the rods and cones in the chick.
He also found deeply stained granules in the initial protoplasmic
buds. These granules, which he has called mitochondria, are to be
seen in figure 13A, which is a reproduction of one of his illustra-
tions. They are seen to bear a striking resemblance to those
which we have observed (figs. 2 and 3), yet our preparations were
fixed with the aid of 5 per cent acetic acid, which is a mito-
chondrial solvent. Leplat regards the transverse striation of the
external segment and its cleavage into dises (fig, 18 B), as the
expression of its mode of construction from mitochondria. In
our preparations a similar transverse arrangement of the material
is readily seen (fig. 5), though they cannot be regarded as mito-
chondrial granules for reasons previously stated. They may,
however, represent chemical transformation products of mito-
chondria with the retention of the plan of the mitochondrial
arrangement.

No evidence of rod and cone identity based on the posture of the
diplosome could be gathered from our preparations. In iron-
haematoxylin stained sections, the protoplasmic buds exhibited
only large groups of granules as seen in figures 2 and 3. If a
typical diplosome is present, such as is shown in Seefelder’s
illustration of the developing human retina (fig. 12) and Leboueq
(709) and First (’04), it is entirely masked by other granules.

Leboucq and Seefelder describe the cone bud as being much
stouter than that of the rod (fig. 12). This figure also shows that
the nuclei of the cones lie in a plane closer to the external limiting
membrane than the rod nuclei. In Amblystoma the initial pro-
toplasmic buds are approximately of the same size. ‘The visual
elements in a retina of several days’ further development show
marked differences in size (fig. 4). This relationship remains

500 S. R. DETWILER AND HENRY LAURENS

unchanged, and when the retina is fully differentiated the rod
nuclei are seen to constitute the outer row of the external nuclear
layer. The same relationship between the position of the rod
and cone nuclei was found in the eyes of the alligator (Laurens
and Detwiler, ’21). In the human retina the cone nuclei occupy
the more superficial position.

In retinae of fifteen days the visual cells typically show two
groups of granules (fig. 4), between which can be seen a clear re-

Fig. 4 Developing visual cells from the retina of a larva of 15 days following
the tail-bud stage of development. The larger elements are the progenitors of
the rods; the smaller, of the cones. XX 1040.

Fig. 5 Visual cells from the retina of a larva of 17 days following the tail-bud
stage of development. The three large conical-shaped elements are differen-
tiating rods. Their nuclei lie partly beyond the external limiting membrane.
The granular material of their outer segments shows a lamellar arrangement
characteristic of the fully differentiated rod (fig. 11). Arrow indicates double
cone. X 1040.

fractive disc marking the position of the oil drop. The original
clear globule is retained in the proximal portion of the visual
element and apparently develops the paraboloid. The proximal
group of granules develops into the ellipsoid portion of the inner
segment. The distal group of granules, as far as can be ascer-
tained, becomes transformed into the typical granular discs
which characterize the outer segment particularly in the rod.
How this group of granules develops is uncertain from our prep-
arations. That they may be separated off from the original

VISUAL CELLS IN AMBLYSTOMA 501

group by liquefaction is suggested by observations which show
that in the center of the single group of granules a clear area
(granule-free) can be seen as shown in figure 3. Whether or not
this is actually the case, cannot be definitely determined. At
any rate, the clear refractive oil drop or dise which separates these
two groups of granules gives no indication of coming from the
original globule, which persists in the proximal portion of the
inner segment as the paraboloid.

Double cones can be seen in early stages of development, as
illustrated in figure 5. These double elements give no indications
as having arisen by division of single elements. When they are
first observed, each member possesses its own nucleus. The
total absence of mitotic figures at this stage when double cones
first make their appearance suggests that they result from the
fusion of preexisting single elements rather than by division.

In the retina of seventeen days (fig. 5), the inner and outer
segments of the visual cells are readily distinguishable. The
inner group of granules has enlarged and is more deeply stained.
They constitute the ellipsoid portion of the inner segment. The
granular material of the outer segment of the large conical
visual cells has undergone cleavage into discs which are separated
by clear non-stainable bands. The lamellar arrangement of this
granular material is quite prominent in the outer segment of the
large visual cells, but less so in the smaller ones (fig. 5). The
difference in the structural arrangement of the material in the
outer segment which can be observed at this period serves as
another index that the individuality of the larger conical-shaped
elements is different from that of the smaller ones. Although
these large elements are conical in shape, the structural arrange-
ment of the material in the outer segment is quite similar to that
which characterizes the fully differentiated rod, and all the
evidence suggests that these large elements are not to be regarded
as cones.

In embryos of twenty-one days (fig. 6) the visual cells show
increased growth. The nuclei of the large conical-shaped ele-
ments (differentiating rods) begin to assume a pear-shape—a
condition which, added to their more external situation, readily

502 S. R. DETWILER AND HENRY LAURENS

distinguishes them from the cone nuclei. The latter are oval in
shape and occupy a deeper level.

A count of the two kinds of visual cells shows that the number
of larger elements slightly exceeds that of the smaller. The
ratio is approximately the same as the rod-cone ratio in the
fully differentiated retina, viz., four to three.

Successive stages in the development of the retinal visual cells
are shown in figures 7, 8, 9, 10, and 11. These figures show the
developmental conditions at twenty-six, thirty-one, thirty-six,

Fig. 6 Visual cells from the retina of an embryo of 21 days after the tail-bud
stage of development. The nuclei of the differentiating rods are being drawn
out on their inner pole, (cf. fig. 11). The cone nuclei are more deeply situated
and are oval in shape. X 1040.

forty, and fifty days, respectively. The difference in size be-
tween the two types of cells gradually increases as development
proceeds. The large conical element gradually becomes more
and more rod-shaped, and the final typical rod form which it
acquires is illustrated in figure 11. Throughout these successive
stages, the nucleus of this visual cell (rod) not only becomes
more and more drawn out on the inner pole, but it gradually
increases in its stainability, as is shown in figures 10 and 11.
These nuclei extend for variable distances beyond the external
limiting membrane, whereas the cone nuclei lie beneath it. The
outer segment of this element shows throughout a disc-like
arrangement of its granular material.

VISUAL CELLS IN AMBLYSTOMA 503

Fig. 7 Double cone, single cone, and four rods from the retina of a larva of
26 days following tail-bud stage. X 1040.
Fig. 8 Double cone, single cone, and three rods from the retina of a larva of

31 days after tail-bud stage. X 1040.

Fig. 9 Double cone, single cone, and two rods from the retina of a larva of
36 days following tail-bud stage of development. X 1040.

Fig. 10 Double cone, single cone and two rods from the retina of a larva of

40 days following tail-bud stage of development. X 1040.

504 S. R. DETWILER AND HENRY LAURENS

In the fully formed rod the myoid is exceedingly short. The
presence of a narrow refractive disc between the outer and inner
segments is demonstrable in most of the rods. In others it ap-
pears to be lacking.

Owing to the greater growth of the rods as development pro-
ceeds, the difference in size between the cones and the rods in
the fully differentiated retina (fig. 11) is greater than in earlier

Fig. 11 Four rods and three single cones from retina of larva of 50 days after
tail-bud stage. XX 1040.

stages of development (fig. 8). The final form of the cones is
seen in figure 11. Their shape and structure agree with the
description given by Laurens and Williams (717). The outer
segments are bluntly conical, and the granular material is not
so characteristically arranged in lamellae as in the rods. As re-
gards the inner segment, the size of the paraboloid is variable
and is present in single as well as in both members of the double
cones.

Fig. 12 Developing rod and cone visual cells, from the retina of a 345-mm.
(six months) human fetus. /, diplosome in a Miiller’s fiber at the level of the
external limiting membrane; C, diplosome in a cone cell; F, fiber growing out from
the cone diplosome; FR, diplosome in a rod visual cell. XX 1000. (AfterSeefelder.
From Jordan and Ferguson’s Text-book of Histology.)

Fig. 13 Two early stages in the development of the rod and cone visual cells
in the chick. A, from twelve-day embryo, showing the visual cell-buds con-
taining mitochondria. L.e., external limiting membrane; C. v., visual cell;
R.e., external reticular layer; C.p., bipolar cells. B, from one-day-old chick,
showing two complete rods and one cone, both elements containing a large lipoid
spherule (@) and mitochondria. The cone contains an ellipsoid (1); seg. int.,
internal segment; seg. ext., external segment. X 2000. (After Leplat. From
Jordan and Ferguson’s Text-book of Histology.)

505

506 S. R. DETWILER AND HENRY LAURENS

The character of the connections of the visual cells with the
internal nuclear layer could not be determined. In preparations
stained with Held’s molybdic acid haematoxylin, the internal
prolongation of the rod nuclei could be followed for a considerable
distance, but its final connection could not be seen.

With the establishment of the character of the centripetal con-
nections of the visual cells which remains to be brought out by
impregnation or other methods, in addition to observations on
the functional behavior prior to the period when typical rods
first appear, more light may be thrown upon the identity of the
developing visual cells in Amphibia.

SUMMARY

1. The fully differentiated retina of Amblystoma punctatum
possesses characteristic rods and cones (fig. 11).

2. The initial stage in the development of the visual cell is the
production of a protoplasmic bud and a clear achromatic globule
from the cells of the external nuclear layer (fig. 2). The globule,
which becomes the paraboloid of the inner segment, appears to
be of cytoplasmic origin, and not of nuclear, as asserted by
Cameron.

3. In early stages of the development of the visual cells (eleven
to thirteen days after tail-bud stage) masses of deeply staining
granules are seen in iron-haematoxylin preparations (figs. 2 and
3). These granules contribute mainly to the formation of the
ellipsoid of the inner segment and to the granular material of the
outer segment. There is no evidence from our preparations
which indicates that they are products of pigment ingested from
the epithelial layer. They may represent transformation pro-
ducts of mitochondria.

4. In these early stages the visual cells are all cone-like (figs.
2 and 3), agreeing in this respect with the conditions characteris-
tic of the developing amphibian retina (Cameron and Bernard).
Rod and cone identity based on the position of the diplosome
such as has been described by Fiirst, Leboucq, and Seefelder has
not been demonstrated. Rods and cones are later differentiated
from the primitive, non-specialized visual cells.

=

VISUAL CELLS IN AMBLYSTOMA 507

5. Rods do not appear in their definitive form until relatively
late in development. ‘Their progenitors consist of large conical-
shaped elements which possess, in early stages of development
(fifteen days), an individuality by which they can be distinguished
from the true cones. These elements are larger, their nuclei
occupy a more external position than that of the cones, and the
granular material of their outer segments becomes arranged in
lamellae characteristic of the fully differentiated rod (figs. 5 and
6). The ratio of these conical-shaped rod progenitors to that of
the cones is approximately the same as that of the rod-cone ratio
in the fully differentiated retina (viz., four to three).

6. The two kinds of conical-shaped visual elements as seen
in figure 4 (large and small) are regarded as visual cells of low
specialization, which develop, respectively, into characteristic
rods and cones by divergent differentiation (fig. 11). This inter-
pretation is not in support of the view advanced by Cameron,
who regards all of the conical-shaped visual cells in early stages of
amphibian development as cones and the later appearing rods
as transformed cones.

7. Double cones appear very early in development (fig. 5).
The absence of mitotic figures in the nuclei of the external nuclear
layer during the period when double cones first appear suggests
that they are formed from the fusion of single elements rather
than from division.

BIBLIOGRAPHY

BERNARD, H. M. 1903 Studies in the retina. Q. J. Micros. Sci., N. S8., vol.
43, pp. 23-48; vol. 44, pp. 443+468; vol. 46, pp. 25-75.

Cameron, J. 1905 The development of the retina in Amphibia. Jour. Anat.
and Physiol., vol. 39, pp. 135-153, 332-348, 471-488.
1911 Further researches on the rods and cones of vertebrate retinae.
Jour. Anat. and Physiol., vol. 46, pp. 45-53.

Detwiter, 8. R. 1916 The effects of light on the retina of the tortoise and the
lizard. Jour. Exp. Zodl., vol. 20, pp. 165-191.

First 1904 Zur Kenntnis der Histogenese und des Wachstums der Retina.
Lund’s Universitits Arsskrift, Bd. 40, adfeln 1, No. 1 (cited from
Cayjale les oo8)-

Kerr, J. GraAnAM 1919 Text-book of embryology, vol.2. London: Macmillan
& Co.

THE JOURNAL OF COMPARATIVE NEUROLOGY, VOL. 33, NO. 5

508 S. R. DETWILER AND HENRY LAURENS

Laurens, H., anp Detwiter, 8. R. 1921 Studies on the retina. The struc-
ture of the retina of Alligator mississippiensis and its photomechanical
changes. Jour. Exp. Zo6l., vol. 32, pp. 207-234.

Laurens, H., anp Witutams 1917 Photomechanical changes in the retina of
normal and transplanted eyes of Amblystoma punctatum. Jour.
Exp. Zoél., vol. 23, pp. 71-83.

Lesouce, G. 1909 Contribution 4 l’étude de l’histogenése de la rétine chez les
mammiféres. Arch. d. anat. micros., Paris, T. 10, pp. 555-606.

Lepiat, G. 1913 Les plastosomes des cellules visuelles et leur rdle dans la
différentiation des cOnes et des batonnets. Anat. Anz., Bd. 45, S.
215-221.

Maarror, A. 1910 Etude sur le développement de la rétine humaine. Ann.
d’oculistique, T. 143 (cited from Leplat).

Parsons, J. H. 1915 An introduction to the study of colour vision. Putnam’s
Sons.

Pirrer, A. 1912 Organologie des Auges. Leipzig: Wm. Engelman.

Ramo6n y Casa, 8. 1911 Histologie du systéme nerveux de l’homme et des
vertébrés. Paris: A. Maloine.

SEEFELDER, R. 1910 Beitriige zur Histogenese und Histologie der Netzhaut,
des Pigmentepithels und des Sehnerven. V. Graefe’s Ar. f. Ophthal-
mol., Bd. 73, S. 419-537.

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Resumen por los autores, O. Larsell y M. L. Mason

Degeneracién experimental del nervio vago y su relaci6én con las
terminaciones nerviosas del pulmén del conejo

La degeneracién experimental del nervio vago del conejo,
seguida de tefido intravital de los pulmones con el azul de meti-
leno, conduce a las siguientes conclusiones: Los nervios vagos
susministran las fibras pregangliénicas que terminan en las células
ganglionares intrapulmonares, diseminadas a lo largo de los
bronquios y sus ramas, como demuestra la desaparicién de la
mayor parte de las redes pericelulares alrededor de dichas células
después de dejar pasar suficiente tiempo para que degeneren las
fibras del vago del mismo lado. La mayor parte de estas fibras
pregangliénicas provienen del vago del lado homolateral, pero
algunas de las redes pericelulares persisten en el pulm6n después
de la degeneracion del vago del mismo lado. Estas redes derivan
parcialmente, con toda probabilidad, del lado contralateral
mediante entrecruzamiento de las fibras en el plexo pulmonar
posterior.

El ntimero de fibras de la musculatura bronquial que aparecen
teflidas varfa muy poco o nada al degenerar las fibras del nervio
vago. Despues de la degeneracién de dicho nervio persisten en
el pulm6n unas pocas terminaciones sensoriales, en el lado
operado. Estas terminaciones se cree provienen en parte del
vago del lado opuesto. El ntimero de fibras nerviosas que se
tifien en las paredes de los vasos sanguineos pulmonares no es
afectado por la degeneracién del nervio vago.

Translation by José F. Nonidez
Cornell Medical College, New York

AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, NOVEMBER 28

EXPERIMENTAL DEGENERATION OF THE VAGUS
NERVE AND ITS RELATION TO THE NERVE TERMI-
NATIONS IN THE LUNG OF THE RABBIT!

O. LARSELL AND M. L. MASON

FIVE FIGURES

In a recent article one of the authors (Larsell, ’21) has demon-
strated histologically the presence of nerve terminations in the
lung of the rabbit and their distribution in relation to the various
parts of the bronchial tree and of the pulmonary blood-vessels.
An adaptation of the methylene-blue technique was employed
which, with the normal lung, gave results that appeared suffi-
ciently constant to warrant an attempt to repeat the method
on experimental animals. The hope was entertained that it
might be possible to determine the elements which belong to the
vagus nerve as shown by their disappearance by degeneration,
following section of the vagus nerve in the neck region.

As in the previous work, rabbits were utilized. The vagus
was sectioned in the midcervical region and the cut ends were
allowed to retract. Ether anaesthesia was used during the
operation, and aseptic precautions were observed. ‘Twelve
animals were operated upon, but only four lived for the period
of twenty-eight days which was deemed necessary for complete
degeneration of the severed fibers.

It is worthy of remark that in these four rabbits the left
vagus had been sectioned, while in the animals which died,
apparently more or less as a result of the operation, either the
right vagus or both right and left nerves had been cut. All of
these animals died within seven days after the vagotomy, and
were not utilized. Except that two showed tuberculous lesions

1 Contribution from the Zoological Laboratory, Wm. A. Locy, Director, and
from the Anatomical Laboratory of Northwestern University.

509

510 O. LARSELL AND M. L. MASON

on autopsy, no other serious lung disturbance was noted, and
they apparently died from other causes. Two of the animals
which were subjected to the double vagotomy, however, died
within forty-eight hours from respiratory difficulties due to
closure of the glottis.

The lungs of the four animals that survived gave results which
were in such accord that they are believed to be typical. In
these four animals the lungs were stained and treated by the
procedure described in the article to which reference has already
been made. In each case both lungs from a given individual,
namely, the right normal lung and the left lung from the operated
side, were carried through the various steps of technique together,
in order to insure uniformity of treatment. It was believed
that in this way the right lung, with its nerve intact, would serve
as a check on the quality of staining of the nerve fibers in the left
lung, the vagus fibers to which had been severed.

A portion of the left vagus, including the neuroma at the level
of the lesion, was also removed. ‘This was fixed in 0.5 per cent
osmic acid, imbedded in paraffin by the usual process, and
sectioned. Transverse sections 10u thick were made of portions
from the proximal and distal ends, respectively, of the removed
portion of the nerve, and longitudinal sections, mounted in
series, were made of the remainder, including the neuroma.

The corresponding portion of the right vagus was also re-
moved from one animal, fixed and stained in osmic acid, im-
bedded and sectioned transversely. It showed no indication of
difference from the proximal end of the left vagus which was
taken from the same animal (ef. fig. 1). There appears, there-
fore, to have been no upward degeneration in the operated nerve.

All of these nerves which had been subjected to operative
procedure showed degeneration of the myelin sheaths below the
level of section (fig. 2). In one animal (R7) there appeared a
few rings in the distal part of the nerve (fig. 2, my.) which bore
the semblance of normal myelin sheaths, as observed in trans-
verse section, but these could not be followed consecutively
through the series of sections, and probably represent only
short stretches of myelin sheaths which had not yet completely

NERVE TERMINATIONS IN THE LUNG 511

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Fig. 1 Transverse section of left vagus nerve of rabbit (R7) which had been
vagotomized twenty-eight days prior to killing of the animal. The section
represents a portion of the nerve proximal to the level of the lesion, showing the
myelinated fibers of various sizes in apparently normal condition. bl.v., blood
vessel. Osmic acid stain. 10u. X 85.

Fig. 2 Transverse section of same nerve distal to level of lesion, showing
complete degeneration of the myelinated nerve fibers, with the possible exception
of those marked my. bl.v., blood vessel; my. dr., droplets of degenerated myelin.
Osmic acid stain. 104. XX 85.

52 O. LARSELL AND M. L. MASON

broken up into the characteristic myelin droplets (fig. 2, my. dr.)
of degenerated nerves. In the nerves from the other three
animals (R8, R9, and R14) degeneration of the distal portion of
the cut nerve was undoubtedly complete.

Since the preparations were stained by the methylene-blue
technique, the degeneration of fibers within the lungs themselves
could only be inferred from the fact that a relatively very small
number of fibers were stained in the main nerve trunks of the
left lung. The corresponding nerve bundles of the right (normal)
lung were very well stained in three of the animals, but very
poorly in the fourth. The left lung of this animal (R14) gave

Fig. 3 A cluster of four ganglion cells within the right lung of a rabbit (R7)
whose left vagus had been sectioned and degenerated. The pericellular net-
works about the cells have the normal appearance. Methylene-blue stain.
60u. X 600.

results comparable with those obtained from the corresponding
lungs of rabbits R7, R8, and R9. The right lung of R14 was
not well injected with the stain because of blocking of its pul-
monary artery, so that a sufficient quantity of the stain did not
have access to the proper regions of the lung to insure a good
stain of the nerve fibers and terminations.

Study of the intrapulmonary ganglia of the right lung in the
three well-stained cases revealed an apparently normal picture.
All or nearly all ‘of the cells were surrounded by more or less
complete pericellular networks (fig. 3) which represent the termi-
nations about these cells of preganglionic fibers from the vagus
nerve. An occasional cell appeared destitute of such a network,

NERVE TERMINATIONS IN THE LUNG ole

but this condition had previously been found to obtain also in
non-experimental rabbits when stained by the same method.

The left lung, however, revealed a strikingly different picture
with respect to the ganglia as well as to the intrapulmonary
nerve trunks. As shown in figures 4 and 5, nearly all of the
cells were characterized by an absence of the synaptic network.
As compared with the ganglionic cells of the right lung, the
majority of nerve cells in the left lung appeared also to be some-
what shrunken in size.

Fig. 4 Ganglion cells from left lung of same rabbit (R7) showing a single
pericellular network. Methylene-blue stain. 60. X 600.

An occasional pericellular network was found about a cell here
and there in the left lung, but these were few in number. They
evidently represent the terminations of preganglionic fibers
from other sources than the homolateral vagus. Schiff (50)
states that each lung receives fibers from both vagi, and MOll-
gaard (712) more recently has come to similar conclusions. It
appears probable, therefore, that preganglionic fibers from the
contralateral vagus cross through the posterior pulmonary
plexus, enter the lung at the hilum and terminate around some

514 O. LARSELL AND M. L. MASON

of the cells of the intrapulmonary ganglia. The possibility that
some of these fibers may come through the sympathetic from the
upper white rami is not excluded.

The nerve fibers in the smooth musculature of the bronchial
tree, so far as could be determined, were stained in apparently
equal numbers in both lungs and with equal intensity. This
would appear to indicate that these fibers are all postganglionic,
although it does not follow that all have their origin from intra-
pulmonary cells. SSome may be axonic processes from cells of the
ganglionic clusters in the posterior pulmonary plexus or else-
where. Chase and Ranson (’14) state that the bronchial rami

Fig. 5 Ganglion cells and nerve fibers from left lung of same animal (R7).
No pericellular networks are present in this cluster of cells, although fine nerve
fibers, believed to be postganglionic, are stained in the immediate vicinity of
the cells. A single large fiber (s.fi.), probably sensory, is also present. Methy-
lene-blue stain. 60u. X 600.

of the vagus contain large numbers of myelinated fibers and
take from the vagus nerve a considerable proportion of the
myelinated fibers which have continued down to this level.
Some of these are of large size and are undoubtedly sensory, but
the majority are small and probably represent the preganglionic
fibers to the lungs within the vagus nerve. The absence of any
obvious degeneration of the nerve fibers to the bronchial muscu-
lature speaks against Mollgaard’s statement that the bronchio-
dilatator fibers take their origin from cells of the vagus ganglia.
The number of stained nerve processes in the walls of the pul-
monary blood-vessels also appears to be undiminished on the

NERVE TERMINATIONS IN THE LUNG IS,

experimental side as compared with the intact side. The source
of these fibers, according to Méllgaard, is the second and third
thoracic ganglia of the sympathetic trunk, chiefly of the homo-
lateral side, but also from the ganglia of the opposite side. We
have nothing to add to this statement. The nerve processes of
the pulmonary vessels are probably all postganglionic fibers.

No exhaustive study was made of the sections from the ex-
perimental side with reference to sensory nerve terminations.
However, a few such endings were observed at some of the
stations which have been found to be characteristic (Larsell, ’21).
Also a few large nerve fibers (fig. 5, s. fi.) may be followed for
considerable distances along the bronchial tree in the lung of the
experimental side.

These sensory terminations and the fibers of large size which
probably lead to them are very likely derived, in large part,
from the vagus of the opposite side, although Mollgaard, in the
article previously cited, has brought forth evidence that the
lung receives sensory fibers also from the second and third
thoracic spinal gangla. The physiological experiments of
Barry (13) and Roger (717) clearly demonstrate that afferent
impulses from the lung travel through the sympathetic trunk
and white rami communicantes to the spinal cord. Accordingly,
some of the sensory fibers and their terminations which remain
in the lung after degeneration of the vagus branches leading to
it may have their origin in the second and third thoracic spinal
gangha. Both Mollgaard (?12) and Molhant (13) state that
the origin of the sensory fibers which pass to the lung by way of
the vagus nerve is in the body of the ganglion nodosum, well
above the level at which the vagotomy was performed. Hence
it is not likely that any aberrant cells from this ganglion were
responsible for the sensory terminations which remained in the
lung after degeneration of the lower part of the vagus leading to it.

516 O. LARSELL AND M. L. MASON

SUMMARY AND CONCLUSIONS

The vagus nerves supply the preganglionic fibers to the in-
trapulmonary ganglion cells which are scattered along the
bronchi and their branches, as is shown by the disappearance of
most of the pericellular networks about these cells following
experimental degeneration of the vagus nerve of the same side.

Most of these preganglionic fibers come from the vagus of the
homolateral side, but some pericellular networks remain in the
lung after degeneration of the vagus of the same side. These
are probably derived in part from the contralateral side by the
crossing over of fibers through the posterior pulmonary plexus.

The number of fibers in the bronchial musculature which
take the stain is affected very little, if at all, by the degeneration
of the fibers of the vagus nerve.

A few sensory terminations remain in the lung of the operated
side after degeneration of the vagus fibers leading to it. These
endings are believed to be derived, in part, from the vagus of the
opposite side.

The number of nerve fibers in the walls of the pulmonary
blood-vessels which take the stain is unaffected by degeneration
of the vagus nerve.

In conclusion we desire to express our thanks to Dr. 8. W.
Ranson for valuable suggestions in the course of the work on
which the present paper is based.

LITERATURE CITED

Barry, D. T. 1913 Afferent impressions from the respiratory mechanism.
Jour. of Physiol,; vol. 45, p. 473.

CuaseE, M. R., AND Ranson, 8. W. 1914 The structure of the roots, trunk and
branches of the vagus nerve. Jour. Comp. Neur., vol. 24, pp. 31-60.

LARSELL, O. 1921 Nerve terminations in the lung of the rabbit. Jour. Comp.
Neur., vol. 33, pp. 105-129.

Moxuant, M. 1913 Les ganglions périphériques du vague. Le Névraxe, T. 15,
pp. 525-579.

Moéuticaarp, H. 1912 Studien iiber das respirationische Nervensystem bei
den Wirbelthiere. Skandin. Arch. f. Physiol., Bd. 26, S. 315-383.

Roarr, H. 1917 Les réflexes pnéo-pnéique et pnéo-cardiaques.* Presse méd.
IPaie, I, A, [> 78s

Scutrr, Moritz 1850 Ueber den Einfluss der Vagusdurchschneidung auf des
Lungengewebe. Archiy fiir physiologische Heilkunde, Bd. 9, S. 625-
662.

SUBJECT AND AUTHOR INDEX

LBINO rat—from birth to maturity. On

the growth of the largest nerve cells in the

superior cervical sympathetic ganglion

Oil UL St nacsacoe Meret «ort aarp awn ee eee 28

rat. The vascularity of the cerebral
Contexso fit hese c mare e caters saci eae 2

Amblystoma. Studies on the retina. Histo-
genesis of the visual cells in..............

Amphibia. The connections of the vomerona-
sal nerve, accessory olfactory bulb and
Enpemay fo W EET ales Saree pig Dordt, eerie aaa te eee pe

Amphioxus. Ventral spinal nerves in........

Amygdala in amphibia. The connections of
the vomeronasal nerve, accessory olfactory
oxo o}:F2y 2c | RR ee s e: eee

Annelid nerve cord. Regeneration in the. .

Ayers, Howarp. Ventral spinal nerves in
Array DOR US aents Ete teresa cae rttes. Son ont

Ayers, Howarp. Vertebrate cephalogenesis.
V. Origin of jaw apparatus and trigemi-
nus complex—Amphioxus, Ammocoetes,
Bdellostoma, Callorhynchus

ic

reptile, Desmatosuchus spurensis, from
the upper Triassic of western Texas..

Cat, with special reference to the occur rence of
intrinsic commissural neurons. An experi-
mental study of the sacral sympathetic
trum OM bereits shoe ede oe kane

Cells in Amblystoma. Studies on the retina.
Histogenesis of the visual.................

Central nervous system in the human fetus as
expressed by graphic analysis and empir-
ical formulae. The growth of the........

Cephalogenesis. V. Origin of jaw apparatus
and trigeminus complex—A mphioxus, Am-
mocoetes, Bdellostoma, Callorhynchus.
Vertebrate Se ee Pere a er nor cer eee

Cerebral cortex of the albino rat. The vas-

Re ilantayviot Ghe see team mea ne ee mene

Cervical sympathetic ganglion of the albino
rat—from birth to maturity. On the
growth of the largest nerve cells in the
superior....

sympathetic ganglion of the Norway
rat. On the growth of the largest nerve
cellsyimithe superior. 22a; cscs ns tee

Commissural _ neurons. experimental
study of the sacral sympathetic trunk of
the cat, with special reference to the occur-
rence of intrinsic............-....--..-+--

Cook, Marcaret H., anp Neat, H. V. Are
the taste-buds of elasmobr anchs endo-
GeruraleinvOrigine eee ee ee

Cortex of the albino rat.
uescene braless hres com soc sesieo nest eee

Craicin, Epwarp Horne. The vascularity
of the cerebral cortex of the albino rates

EGENERATION of the vagus nerve and
its relation to the nerve terminations in
the lung of the rabbit. Experimental.

Detwiter, 8S. R., aNpD LAvuRENS, HENRY.

Studies on the retina. Histogenesis of the

visual cells in Amblystoma

SE, E.C. Onan endocranial cast froma

339

133

313

Dog. Effect of cutting the lingual nerve of
Dunn, Hatpert L. The growth of the cen-
tral nervous system in the human fetus as
expressed by graphic analysis and empiri-
callitonmul ae seen ro aan eee eee See,
LASMOBRANCHS endodermal in origin?
Arevthe itaste-pudsjiote oss) acce cen eae ee

Endocranial cast from a reptile, Desmato-
suchus spurensis, from the upper Triassic
ofiwesterm Pexas: “On ans. 20) assoc eee

ETUS as expressed by graphic analysisand

empirical formulae. The growth of the

central nervous system in the human... 405

ANGLION of the albino rat—from birth
to maturity. On the growth of the
largest nerve cells in the superior cervical
Sympavhetics perert ae ee eco ee cian 281
Ganglion of the Norway rat. On the growth
of the largest nerve cells in the superior cer-
WicHlisyvmipabhetich eet ys eee eeeeter ears
Growth of the central nervous system in the
human fetus as expressed by graphic
analysis and empirical formulae. The. .
of the largest nerve cells in the superior
cervical sympathetic ganglion of the
albino rat—from birth to maturity. On
Lo to Aare nau tie Se Ly, eA By Sen a 2
——— of the largest nerve cells in the superior
cervical sympathetic ganglion of the Nor-

way rat. On the
Hire Apa R. Regeneration in the anne-
Indinenvercondneneeecnteene cena eee Eee

Head of the urodeles. The fate of the neural
CLestainyuNere eee erence cer cee ae
Herrick, C.Jupson. The connections of the
vomeronasal nerve, accessory olfactory
bulb and amygdala in amphibia........ 2
Histogenesis of the visual cells in Amblys-
toma. Studies on the retina............
Human fetus as expressed by graphic analy-
sis and empirical formulae. The growth

of the central nervous system in the

AW apparatus and trigeminus complex—
Amphioxus, Ammocoetes, Bdellostoma,
Callorhynchus. Vertebrate cephalogen-
EsIsey VLOriPingaOtasc eect cee eee

JOHNSON, SYDNEY An experimental
study of the sacral sympathetic trunk of
the cat, with special reference to the occur-
rence of intrinsic commisural neurons.

JOHNSON, S. E., anD Mason, M.L. The first
thoracic white ramus communicans in

339

ANDACRE, F. L. The fate of the neural
crest in the head of the urodeles........

LARSELL, O. Nerve terminations in the lung
of the rabbit

Ld ed

518

LARSELL, O., AND Mason, M. L. Experimen-
tal degeneration of the vagus nerve and
its relation to the nerve terminations in the
Tung’ of. the rabbit @a 8--0-kp eee cee eee 509
Laurens, Henry, Detwiter, S. R., anv.
Studies on the retina. Histogenesis of the
visual cells in Amblystoma............... 493
Lingual nerve of the dog. Effect of cutting the. 149
Lung of the rabbit. Experimental degenera-
tion of the vagus nerve and its relation to

the nerve terminations in the.......... -- 509
of the rabbit. Nerve terminations in
tHe eh. Te is oe deen eee eee 05

AN. The first thoracic white ramus com-
Municans AM, 354 eee tee 7

Mason, M. L., Jounson, S. E., anp. The
first thoracic white ramus communicans
ph awscst:)s0 CQ, Sara ema, OR ciate «) "TS ae 77
Mason, M. L., Larsreitt, O., anp. Experi-
mental degeneration of the vagus nerve
and its relation to the nerve terminations
im the lung ofthe rabbit eee... ©... 26. 509

EAL, H. V. Nerve and plasmodesma.... 65

Neat, H. V., Cook, MARGARET H., AND.
Are the taste-buds of elasmobranchs endo-

Nerve, accessory olfactory bulb and amygdala
in amphibia. The connections of the
vomeronasal, Met. te. ae alee 213
and plasmodesma: 4: nae. eer ear aoe 65
——— cells in the superior cervical sympa-
thetic ganglion of the albino rat—from -
birth to maturity. On the growth of the
larrestey y 5.07 ean taney mice, coe an oe eet 281
cells in the superior cervical sympa-
thetic ganglion of the Norway rat. On
the growth of the largest:............... 313
—— cord. Regeneration in the annelid.... 163
of the dog. Effect of cutting the lingual. 149
terminations in the lung of the rabbit.. 105
Nervous system in the human fetus as
expressed by graphic analysis and empiri-
cal formulae. The growth of the central. 405
Neural crest in the head of the urodeles. The
fateiol the: ar. eee ee ook Se scicaee 1
Norway rat. On the growth of the largest
nerve cells in the superior cervical sympa-
thetic’ ganglion’ of thet.........05) .66ne oo. 313

CCURRENCE of intrinsic commisural
neurons.

special reference to the.................. 85

AECESSONY,.«).2.. .)-: > uy) PO eo le,
Oumstep, J. M.D. Effect of cutting the lin-

gual nerve of the dog.................... 149
Origin? Are the taste-buds of elasmobranchs

endodermal ina. tc. ee ee 45

of jaw apparatus and trigeminus com-
plex—Amphioxus, Ammocoetes, Bdellos-
toma, Callorhynchus. Vertebrate cepha-
logenesis. V...... Pale eMcRrslscicieeicitie eieerres 339

INDEX

ING, Cur. On the growth of the largest
nerve cells in the superior cervical sympa-
thetic ganglion of the albino rat—from

birth: tohmaiburity = <2 | ee eee 281

Pine Cut. On the growth of the largest nerve
cells in the superior cervical sympathetic

ganglion of the Norway rat............... 313

Plasmodesma. Nerve and................... 65

Re. Experimental degeneration of
the vagus nerve and its relation to the
nerve terminations in the lung of the.. 509
Nerve terminations in the lung of the 105
Ramus communicans.in man. The first
ChOrseie? White: fo. ices Ses oe fi
Rat—tfrom birth to maturity. On the growth
of the largest nerve cells in the superior
cervical sympathetic ganglion of the
albinos <eeeee ee oot tee oe i ee 281
On the growth of the largest nerve cells
in the superior cervical sympathetic gang-

lion! ofsrbe N omyaye os). Shee. Scie ene 313
——— The vascularity of the cerebral cortex

of the*albino 36.2 oe ese 193
Regeneration in the annelid nerve cord...... 163

Reptile, Desmatosuchus spurensis, from the

upper Triassic of western Texas. On an

endocranial cast from a.................. 133
Retina. Histogenesis of the visual cells in

Amblystoma. Studies on the............ 493

PINAL nerves in Amphioxus. Ventral 155
Sympathetic trunk of the cat, with special
reference to the occurrence of intrinsic
commissural neurons. An experimental

Shudyeot thessacral:....i:c.. aoc /o. meee 85
feat Same of elasmobranchs endodermal
INOnIGIN, Amen Me Fotis... ii. Sc: ee 45

Terminations in the lung of the rabbit. Experi-
mental degeneration of the vagus nerve

and its relation to the nerve............ 509
Thoracic white ramus communicans in man. oF
PRE ATSG Se thee ae ec ae eae 77

Trigeminus complex—Amphioxus, Ammocoe-
tes, Bdellostoma, Callorhynchus. Verte-
brate cephalogenesis. V. Origin of jaw
ApPPATabus-and) =... eee mre ees + eens 339

RODELES. The fate of the neural crest
imthe) HeaGu.of the:sec.smsiietc.e ouceaee on 1

AGUS nerve and its relation to the nerve
¥: terminations in the lung of the rabbit.
Experimental degeneration ofthe...... 509
Vascularity of the cerebral cortex of the albino
nish meee Dal: ne cholo ae oe Be Oh obo Sapo 193
Ventral spinal nerves in Amphioxus..... pears tits)
Vertebrate cephalogenesis. V. Origin of jaw
apparatus and trigeminus complex—
Amphioxus, Ammocoetes, Bdellostoma,
Callorhynehus.; comet. a2 2~ Bee Sra 339
Visual cells in Amblystoma. Studies on the
retina. Histogenesis of the.............. 493
Vomeronasal nerve, accessory olfactory bulb
and amygdala in amphibia. The connec-
FOMS "Ol Chews Ae omgen ecins eos 7 kero 213

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