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THE AMERICAN JOURNAL
OF
ANATOMY
EDITORIAL BOARD
CHARLES R. BARDEEN
University of Wisconsin
Henry H. DoNaALpson
G. Cart HUBER J. Puayrain McMourrica
University of Michigan
University of Toronto
GerorGE S. HUNTINGTON GEORGE A. PIERSOL
The Wistar Institute Columbia University University of Pennsylvania
Simon H. GaaGe
Henry McE. KnNower, SECRETARY
Cornell University
University of Cincinnati
VOLUME 25
PAIN UA YY
1919
THE WISTAR INSTITUTE OF ANATOMY AND BIOLOGY
PHILADELPHIA, PA.
CONTENTS
No. 1. JANUARY
GerorGE L. STREETER. Factors involved in the formation of the filum terminale. Three
ENGST OUI CS Sarge Panta te ENED Pree mente treed cB 2 Hat scch adie plea otis RTE ME OED Sree ft
J. A. BapertscHer. The ultimobranchial bodies in postnatal pigs (Sus scrofa). Four
HOU CSeeee eee BO Greboiatg a hia ke Ut Gas Gord OER CE eae ORE oie ecto eae ic Orne C.
FRANK haees edema “The origin of the phagocytic mononuclear cells of the
peripheral blood. Eleven figures. . Ba 27
H. D. Sentor. The development of the sieibes a tite ipa apes eaeuien "Bleven
THEW INS Sh oe ctor rg Etna EN get oo Se PURGE CR i Se a Do ME 19)
No. 2. MARCH
C. H. DanrortH. The ane relations of brachydactly in the domestic fowl.
Five figures. oe o We
Dexa Dries. eraciee on inet ovary Os “fhe pperamonie (Gomaneniite eielioe eudecont
lineatus) with special reference to the corpus luteum. Twenty-nine figures........ 117
Royat Norton CHapMan. A study of the correlation of the pelvic structure and the
habits of certain burrowing mammals. Five plates (twenty-six figures)............ 185
No. 3. MAY
C.M. Jackson. The postnatal development of the suprarenal gland and the effects of
inanition upon its growth and structure in the albino rat. Ten figures. sae PPall
Joun C. Donatpson. The relative volumes of the cortex and medulla of ang eee
gland in the albino rat. Four charts.. Ae. Me eee. 20
FRANKLIN PARADISE JOHNSON. The eooleamiont of the jopule an ie pig’s silva ne
eight figures. . : .. 299
Lucite WITTE. Pie eoeeneais a ae fees mmmeele ae the pig in reiaiee fa the poueinemnes
and development of the intercalated discs. Two plates (eighteen figures).......... 333
NozA JULY
Epwarp Puecps Auuis, Jr. The homologies of the maxillary and vomer bones of Polyp-
terus. Three hee (eighteen figures)... pee . 349
J.A.Mynrs. Studieson themammary Hemel, “iy, The fieealoey a hed mammary ation in
male and female albino rats from birth to ten weeks of age. Nineteen figures (eleven
text figures and two plates).. ; . 395
H. E. Jorpan. The histology of fhe blood one fis réd bone -marrow oe ihe aed frc og,
Rana pipiens. Two plates (seventy-three figures) >...........:..0ccccecccsccccvsecs 437
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AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, JANUARY 6
FACTORS INVOLVED IN THE FORMATION OF THE
FILUM TERMINALE
GEORGE L. STREETER
From the Department of Embryology, Carnegie Institution of Washington,
Baltimore, Maryland
THREE TEXT FIGURES
In a study recently published by the writer! on the develop-
ment of the cartilaginous capsule of the ear in human embryos
it was pointed out that the changes in size and form which the
capsule undergoes during its development are accomplished not
only by a progressive differentiation, but also in part by a retro-
gressive differentiation of its constituent tissues. The margins
of the cartilaginous cavities are in a continual state of change;
they exhibit an unstable equilibrium between two opposing ten-
dencies: on one hand, toward the deposit of new cartilage, and
on the other, toward the excavation of the old. The margins
thereby are always advancing or receding, and as a result of
this there is provided a suitable suite of chambers for the
contained membranous labyrinth in all stages of its development.
It is the feature of retrogressive differentiation or dedifferen-
tiation that I wish particularly to recall here. The fact that
certain areas of cartilaginous tissue revert to an earlier em-
bryonic type and are subsequently redifferentiated into a tissue
of a widely different histological character, is very clearly shown
in the case of the otic capsule, and is a factor of great embryo-
logical significance. Such a process of retrogressive change,
combined with redifferentiation of the same tissue, greatly in-
creases the facilities for and the range of certain structural
1 Streeter, G. L., 1917. The factors involved in the excavation of the cavities
in the cartilaginous capsule of the ear in the human embryo. Amer. Jour. Anat.,
vol. 22.
1
ys GEORGE L. STREETER
adjustments that occur in many regions in the development of
the human embryo.
Another instance of dedifferentiation has recently been pointed
out by Kunitomo.? This writer has published the results of a
careful study of the tail region in alargenumber of human embryos,
representing the period of greatest development of the caudal
appendage, and also the later period of its gradual reduction.
He shows that in very young specimens the spinal cord reaches
the extreme tip of the tail and throughout its length is quite
uniform in structure. Somewhat later (11 to 15-mm. stage) it
can be divided at about the level of the thirty-second vertebra
into two parts—a cranial or main part, having a wide central canal
and thick walls in which can be recognized well-developed mantle
and marginal zones, and a caudal slender part, having a narrow
canal with walls consisting only of an ependymal zone. Kunitomo
shows that it is this caudal atrophic portion that eventually
forms the filum terminale. The main part lying cranial to the
thirty-second vertebra undergoes uninterrupted and progressive
differentiation, whereas the portion caudal to this undergoes re-
gressive changes and, with the exception of the extreme tip,
finally becomes converted into a fibrous strand, the tip forming
the coccygeal medullary vestige. This, therefore, is another
instance in which an absorptive adjustment is brought about by
the reversion of the tissue to an earlier embryonic type with a
certain amount of subsequent redifferentiation.
Kunitomo further calls attention to the fact that in the forma-
tion of the filum terminale, in addition to the dedifferentiation
of the caudal end of the medullary tube, there is also the
mechanical disproportion between the growth of the medullary
tube and that of the vertebral column. How much of one and
how much of the other of these two factors is responsible for the
further development of the filum terminale was not determined
by him. It has occurred to the writer that this question could
be answered by the determination of the elongation of the nerve
2 Kunitomo, K., 1918. The development and reduction of the tail and of the
caudal.end of the spinal cord in the human embryo. Contributions to Embry-
ology, vol. 8, Publication No. 271, Carnegie Inst. of Wash.
FORMATION OF THE FILUM TERMINALE 3
roots. In the younger stages the spinal cord and the vertebral
column lie alongside of each other in a metameric manner, cor-
responding in position segment for segment. Owing to their
disproportion in growth, there occurs a relative displacement of
their segment levels, so that, for instance, the thirtieth segment
of the cord comes to lie opposite the twentieth segment of the
vertebral column. The segment levels of the vertebral column
are, of course, evident; in the spinal cord they are just as plainly
marked by the attachment of the nerve roots, for these become
attached to the cord before the displacement begins, and thus
permanently mark the various segmental levels. In the case of
each segment of the spinal cord there are two fixed topographical
points: the spinal ganglion, which is held in the intervertebral
foramen and registers the original position of the segment relative
to the vertebral column, and the place at which the dorsal root
is attached to the cord and which moves as the cord moves. By
locating those points for the different stages one can determine
the exact elongation of the nerve roots, and this in turn is the
index of the relative displacement of the spinal cord as regards
the vertebral column. Conversely, it will be seen that the
alteration not explained by mechanical displacement must be
attributed to the retrogressive changes referred to above. The
determination of the amount of displacement was made by
comparison of selected stages by means of profile reconstructions
of the smaller specimens and actual dissection of the older ones.
I was assisted in this by Mr. James F. Didusch, of the Carnegie
Embryological Laboratory, who made careful dissections of these
structures in several older fetuses, two of which will be used for
illustration. The results of this determination are given in the
following note as a matter of interest to those who have read
the paper by Kunitomo, and also because it offers an opportunity
to emphasize the significance of dedifferentiation of tissues in
the processes of development in the human embryo.
The part played by dedifferentiation in the caudal region of
the spinal cord is more apparent in the younger stages of develop-
ment, as pointed out by Kunitomo. The so-called ‘absorption’
of the tail is completed before the embryo reaches a length of
4 GEORGE L. STREETER
30 mm. It is also well known that the remodeling which takes
place in the gill region completes the obliteration of the gill bars
before the embryo is 20 mm. long. One well might expect these
processes of dedifferentiation and redifferentiation to be more
active in the earlier stages. They are not confined, however, to
- this period, for in the case of the ear capsule they were found to
be very active throughout fetal life. In the case of the spinal
cord dedifferentiation is well demonstrated in the period repre-
sented by embryos between 11 and 30 mm. long. A comparison
of these two stages can be made in figure 1. It will be noted in
the first place that the spinal ganglia show a regression varying
from arrest in development to complete disappearance. All but
two of the coceygeal ganglia have disappeared in the 30-mm.
specimen, and the remaining two are of about the same size as —
the same two ganglia in the 11.5-mm. specimen.’
As for the cord itself, the changes are equally marked. In the
younger stage (embryo 11.5 mm. long) the extreme caudal end
of the spinal cord, the part belonging to the non-vertebrated tail,
shows little differentiation, consisting only of indifferent cells
resembling embryonic ependyma. In the coccygeal region, how-
ever, the development is more advanced. Opposite the five
coccygeal ganglia the wall of the cord is differentiated into dis-
tinct ependymal, mantle, and marginal zones, with well-developed
rootlets entering into it from the first two ganglia. Sections
through it show nothing to indicate that this region is not going
on to complete its differentiation into the adult condition. When,
for comparison, one examines the very same region in the older
specimens (fig. 1, embryo 30 mm. long) it is found that its con+
dition, relative to the remainder of the cord, has undergone a
marked change. While the precoccygeal cord has continued to
increase in the thickness of its walls and in the elaboration of the
mantle and marginal zones, the coccygeal region is less advanced
3 Throughout this paper the twenty-fifth to the twenty-ninth segments have
been uniformly regarded as sacral. The slight variation which is known to exist
in this respect is too small to be taken into account in our genéral conclusions,
and for convenience the regional terms, lumbar, sacral, and coccygeal, will be
used, upon the assumption that the specimen concerned has the usual regional
distribution of its segments.
FORMATION OF THE FILUM TERMINALE 5
in its development than it was in the younger stage. Whereas in
the 11-mm. embryo there existed a distinct elaboration- into
ependymal, mantle and marginal zones, the mantle zone is com-
pletely missing in the 30-mm. embryo, and we find thin walls
consisting only of ependymal cells covered by a thin marginal
zone. The coccygeal spinal cord in the 30-mm. embryo is in an
earlier embryonic state than that of the 11.5-mm. embryo; that
0080U)
ve.
11.5 mm (x25)
30mm (x 12.5)
Fig. 1 Profile reconstructions showing the spinal ganglia and their dorsal
roots in the tail region of the human embryo. The last two lumbar ganglia are
shown in white, the sacral ganglia are stippled, and the coccygeal ganglia are
solid black. It will be noted that in the period included between these two
stages marked regressive changes have affected the entire coccygeal region of the
spinal cord, with complete disappearance of the last three coccygeal ganglia, in
sharp contrast to the sacral region of the cord, which undergoes uninterrupted
development. The reconstructions are taken from embryos No. 544, 11.5 mm.
long, and No. 75, 30 mm. long, belonging to the Carnegie Collection.
is, it has undergone dedifferentiation. In later stages the proc-
ess goes still farther and, as has been pointed out by Kunitomo,
this ependymal tube eventually becomes converted or redif-
ferentiated into a fibrous strand.
How much of the spinal cord is involved in this retrograde
process can be seen by comparing the two stages shown in
figure 1. In the region of the attachment of the fifth sacral
nerve the wall of the cord remains thick and develops a well-
6 GEORGE L. STREETER
differentiated mantle zone. About opposite the first coccygeal
nerve in the 30-mm. embryo the mantle zone abruptly disappears,
and there is a corresponding enlargement of the lumen of the ~
cord, thereby producing the thin-walled ventriculus terminalis.
There is some variation in different embryos as to the segmental
level caudal to which the mantle zone has dedifferentiated and
also in the manner of transition from the well-developed sacral
cord into the atrophic coccygeal cord, including sometimes the
doubling or partial obliteration of the central canal. The tran-
sition is quite abrupt, involving only one segment. In the
30-mm. embryo in figure 1 the cord at the level of the first
coccygeal nerve shows some decrease in the size of its mantle-
zone area. Opposite the second coccygeal nerve the mantle
zone is entirely gone. The second coccygeal ganglion present
in this case would probably soon have disappeared.
The ventriculus terminalis at this stage tapers caudally and
may be said to extend to the third coccygeal segment. Caudal to
this the differentiation of the cord is more complete and results
in the gradual obliteration of the lumen and the replacement of
the ependymal substance by a fibrous strand, embedded in
which can be found isolated groups of persistent ependymal
cells. At its extreme tip there is often found a more or less
detached group of such cells which undergoes cystic enlargement
and constitutes the coccygeal medullary vestige. The interval
of cord lying between this and the ventriculus terminalis con-
stitutes what is later known as the filum terminale. Thus far
its formation is based upon the process of dedifferentiation; its
subsequent growth and elongation is accomplished by an inter-
stitial increase of its constituent fibres, and not by the further
invasion of the process of dedifferentiation into the sacral region
of the cord. -This will become evident on examination of
figure 2
It has been pointed out that in embryos 30 mm. long a ven-
triculus terminalis is formed opposite the second and third
coccygeal vertebrae, owing to a retrogressive thinning out of
the walls of the spinal cord, with a consequent irregular enlarge-
ment of the central canal. In fetuses with a crown-rump
FORMATION OF THE FILUM TERMINALE
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length of 100 to 200 mm. the ventriculus terminalis can be
recognized in gross dissection with the naked eye, as a piriform,
translucent area at the tip of the conus medullaris. Apparently
in the natural condition it presents smooth outlines, but in pre-
pared sections its thin walls are thrown into what are evidently
shrinkage folds. This ventricle bears a permanent relation to
the rootlets of the fifth sacral nerve at their entrance into the
cord, as is shown in figure 2. In the four stages there represented
it lies just caudal to the entrance of the fifth sacral nerve. The
rootlets of the first coccygeal nerve in these specimens were so
delicate that they could not be traced with certainty and were
therefore omitted. The roots of the five sacral nerves, however,
could be very accurately followed, and are all indicated in the
figures. Their entrance into the substance of the cord con-
stitutes in each case a firm attachment and remains a fixed topo-
graphical point. By comparing the four stages from 30 mm. to
221 mm. it will be seen that the ventriculus terminalis and the
points of attachment of the sacral roots maintain the same
relative positions, there being no further encroachment of the
former into the territory of the more cephalic lying spinal cord.
In other words, there is no further dedifferentiation of the sacral
region of the cord after the embryo has attained a length of 30
mm. The cephalic migration that is subsequently experienced
by the ventriculus terminalis and points of attachment of the
sacral nerve roots, relative to the bodies of the vertebrae, is
clearly a result of the fact that the vertebral column gradually
extends farther caudalward than the spinal cord, and since the
nerve roots and the filum terminale are attached at both ends
they are correspondingly elongated. The latter process is not a
simple stretching, for, as these structures lengthen they actually
become thicker. In other words, there is a compensatory inter-
stitial growth. This increase in thickness is not apparent in
figure 2, as the older stages are shown at a progressively de-
creasing scale of enlargement.
The rapidity and extent of the caudal thrust of the vertebral
column—that is, its caudal displacement in relation to the
terminal ventricle—can be seen in figure 2. This covers a little
FORMATION OF THE FILUM TERMINALE 9
over the first half of fetal life (twenty-five weeks). In the
adult the corresponding points fall at the interval between the
bodies of the first and second lumbar vertebrae. Thus in the
first twenty-five weeks there is an ascent of the ventriculus
terminalis from the level of the second coccygeal to the third
lumbar vertebra, or a distance of nine segments, and there
remain but two segments before the adult position is reached.
One may say that the principal part of the migration is accom-
plished during the first half of fetal life.
The dura mater and its relations can be plainly recognized in
the 67-mm. fetus, where it can be seen to reach and adhere to
the filum terminale at the lower border of the fourth sacral
vertebra, thereby sealing off the lower end of the subdural
space. It is of interest to note that it undergoes very little
change from its position here and that which it occupies in the
adult. In the 111-mm. fetus it extends to about the same level
and ends in the same manner. In the adult it terminates about
two segments higher up. Thus the dural sac conforms more to
its bony environment than does the spinal cord and shows very
little of the migratory adjustment of position that is noted in
the latter. We therefore find the ventriculus terminalis
gradually receding cranialward from the caudal end of the sac.
In figure 2 the specimens are enlarged upon a decreasing
scale of magnification according to age, so that the segments of
the different stages are brought to about the same size. This
has been done in order to facilitate the comparison of segment
levels. The actual elongation of the spinal root of a given nerve
is greater, therefore, than would appear from the figure.
Measurements of the dorsal root of the first sacral nerve from
the margin of the ganglion to the point of entrance into the
cord yield the following figures: 30-mm. fetus, 0.65 mm. long;
67-mm. fetus, 4.75 mm. long; 111-mm. fetus, 12.25 mm. long;
221-mm. fetus, 32 mm. long.
The actual elongation of the first sacral root is indicated for
the first three of these stages in figure 3, in which the topography
of the spinal cord and the vertebral column is drawn on the same
scale of enlargement. The dorsal root of the first sacral nerve
10 GEORGE L. STREETER
Fig. 3 Topography of the spinal cord and the dorsal root of the first sacral
nerve in three fetal stages. 'These are taken from three of the same specimens
shown in figure 2, but here they are shown on one scale of enlargement in order
to indicate the actual changes in size. The 30-mm. specimen is shown both in
figure 2 and figure 1. y
FORMATION OF THE FILUM TERMINALE Lt
is indicated by a heavy black line; the first thoracic, first
lumbar, first sacral, and first coccygeal vertebrae are marked
by small circles. Comparison of the stages, as shown in this
figure, gives perhaps a better representation of the actual topo-
graphical changes that occur in this apparent ascent than does
figure 2.
From these results we may conclude that in the human
embryo the greater part of the coccygeal and post-coccygeal
cord—that is, the part caudal to the thirtieth segment—under-
goes dedifferentiation, the more cephalic part of it persisting as
the ventriculus terminalis and the more caudal part redif-
ferentiating into a fibrous strand—the fi!um terminale, with the
coccygeal medullary vestige at the tip. The first twenty-nine -
segments of the spinal cord are not affected by this process of
dedifferentiation, but continue in a progressive development.
When the embryo reaches 30 mm. in length there begins a dis-
‘proportion in the rate of growth as between the vertebral column
and the spinal cord, the former elongating more rapidly than
the latter. This results in a relative displacement of the two,
the ventriculus terminalis in the 221-mm. fetus (twenty-five
weeks) lying nine segments higher than it did originally, and by
the time the adult form is attained two more segments have been
added to this disproportion. We may say, therefore, that the
filum terminale represents that portion of the spinal cord caudal
to the second coccygeal segment (thirty-first segment), which
has undergone dedifferentiation and has finally become con-
verted into a fibrous strand. This strand, like the sacral nerve
roots, elongates by interstitial growth in adaptation to the
ascending displacement of the spinal cord. The caudal tip of
the dural sac maintains its relation to the vertebrae rather than
to the spinal cord and remains attached to the filum terminale
in the sacral region at a more or less fixed point.
Resumido por el autor, J. A. Badertscher.
Los cuerpos tiltimobranquiales después del nacimiento en el
cerdo (Sus scrofa).
Una porcién de los cuerpos tltimobranquiales puede persistir
durante largo tiempo en la glindula tiroides del cerdo, después
del nacimiento. En cerdos hasta de 56 dias de edad, estan
formados por areas de cordones sincitiales nucleados y masas en
las cuales est’ comenzando a formarse materia coloide, o por
dreas en las cuales los foliculos son pequenos pero con la estruc-
tura tipica de la glindula tirofdes. La porcién central de
algunas de las dreas sincitiales esta desprovista de coloide. Un
' drea alargada de pequefios foliculos que contienen coloide, la
cual representa probablemente un cuerpo tltimobranquial, fué
encontrada por el autor en dos entre tres glindulas tiroides de
cerdos subadultos. Los cuerpos tltimobranquiales estan colo-
cados generalmente en la mitad posterior de la glandula tiroides,
cerca de su borde dorsal o dorso-lateral. Es imposible deter-
minar exactamente la proporcién relativa en que los cuerpos
ultimobranquiales y el esbozo tiroideo medio contribuyen a la
formacion de la glandula tiroides. A causa del comportamiento
variable durante el desarrollo, por parte de los cuerpos tltimo-
branquiales, la proporcién relativa en que dichos cuerpos con-
tribuyen a la constitucién de la glindula tiroidea es indudable-
mente variable en diferentes cerdos. Es sin enibargo evidente
que solo una pequefia porcién de la glandula deriva de los cuerpos
ultimobranquiales. Folfeulos excepcionalmente grandes (cis-
toideos) abundan en algunas de las tiroides examinadas. De un
modo general puede decirse que la situacién de estos foliculos
esta limitada a la porcién de la tirofdes en la cual se encuentran
generalmente los cuerpos ultimobranquiales, es decir, en la mitad
posterior de la glandula.
Translation by Dr. José F. Nonidez,
Columbia University
AUTHOR’S ABSTRACT OF THIS PAPER ISSUED BY
THE BIBLIOGRAPHIC SERVICE, DECEMBER 23
|
THE ULTIMOBRANCHIAL BODIES IN POSTNATAL
PIGS (SUS SCROFA)
J. A. BADERTSCHER
From the Department of Anatomy, Indiana University, Bloomington, Indiana
FOUR FIGURES
In a recent study (’18) of the ultimobranchial bodies in a wide
range of successively older developmental stages of pig embryos
(before their fusion with the median thyroid anlage to full
term), the writer was convinced that these bodies contribute to
the structural elements of the thyroid gland. The time at
which they are completely transformed into typical thyroid
structures, that is, when they can no longer be recognized
structurally from the median thyroid anlage, varies greatly.
Even in a full-term embryo a portion of the ultimobranchial
bodies may be free from colloid. It thus became evident that in
order to follow out a more complete developmental history of these
structures, the thyroid gland of postnatal pigs must be examined.
The material used for this investigation was obtained from a
litter of pigs and from three young adult hogs (age unknown).
The pigs were killed at the following ages: one a few hours
after birth, one 7.5 days old, one 15 days old, one 28 days old,
one 42 days old, and one 56 days old. The thyroid and a portion
of the trachea were removed from the pigs, while only the thyroid
was removed from the adults. All the material was fixed in
Zenker’s fluid and imbedded in paraffin. The thyroid glands of
the pigs were cut transversely into sections 15y in thickness, all
the sections were mounted in serial order, and stained with
eosin and Unna’s alkaline methylene-blue solution. The thyroid
gland of the adults were cut transversely into sections 20y
thick, only every fifth section was mounted, and stained with
hematoxylin and eosin.
13
14 J. A. BADERTSCHER
In the description of the following stages special attention will
be given: 1) to the structure of the ultimobranchial bodies and
to their location in the thyroid gland, and 2) to the location and
extent of areas of unusually large (cystoid) follicles. Since in
the embryonic material it was found that cystoid follicles may
develop in the ultimobranchial bodies, the latter consideration
is of importance.
Pig at birth. The thyroid gland is 9.3 mm. long and its
greatest width is 4.8 mm. Caudally it terminates in a rather
blunt point, while the anterior portion is drawn out into a slender
streamer. The greater portion of the bulk of the gland is thus
located in its posterior half. The more bulky part of the gland
is crescent in shape in transverse sections. In the middle third
of the left lateral half of the thyroid gland the ultimobranchial
body is represented by three small areas which are composed of
tortuous syncytial cords and masses quite closely packed to-
gether. These areas are embedded beneath the dorsal surface
of the gland lateral to its medial plane, a position usually
occupied by the ultimobranchial bodies in the later embryonic
- stages. Anterocaudally, they extend through eight, six, and
twelve consecutive sections, respectively. The most anteriorly
located of these areas is free from colloid and lies in a field of
follicles that are on an average smaller than the average size of
the majority of follicles present in the thyroid gland. In the
central and caudal areas the colloid is just beginning to form.
On account of the absence of colloid in one and its scanty amount in
the other two of these areas, they stand out sharply from the thy-
roid follicles immediately surrounding them. No difference could
be observed between the structure of the nuclei in these areas and
the nuclei in the cells composing the follicles. In the right lateral ,
half of the thyroid gland the ultimobranchial body is absent.
1 As this work is practically a continuation of a previous investigation (18)
by the author of the ultimobranchial bodies in pig embryos, it was deemed
unnecessary to repeat an historical sketch of this subject in this article. Also
the bibliography includes only those references to articles in which may be’
found more orless definite statements concerning the fate of the ultimobranchial
bodies. If an extensive bibliography on this subject is desired, reference should
be made to the, works of Verdun (’98) and Grosser (12).
ULTIMOBRANCHIAL BODIES IN POSTNATAL PIGS 15
Unusually large follicles are present in both lateral halves of the
caudal fourth of the thyroid gland. These are located mainly
near the dorsal and dorsolateral surface of the gland. A few
very large follicles are located near the most caudally located
area of the ultimobranchial body. Also an area of very large
follicles (extending through a series of thirty-four sections) is
present in the caudal portion of the middle third of the right
lateral half of the thyroid gland just below its dorsal surface.
Pig 7.5 days old (fig. 1). The thyroid gland is 15 mm. long
and its greatest width is 6.8 mm. The greater portion of the
anterior half of the thyroid gland is in form a slender band, so
that by far the greater portion of its bulk les in the posterior
half of the gland. The more bulky portion of the gland is
crescent in shape in transverse section. Traces of both ultimo-
branchial bodies are present in the caudal fourth of the thyroid
gland. The left one is represented by three oblong areas com-
posed of closely packed syncytial cords and masses which are
almost free from colloid. The nuclei in the syncytium have a
structure identical to the structure of the nuclei in the cells
composing the follicles. Anterocaudally, these areas extend
through nineteen, seven, and thirty consecutive sections, re-
spectively. The most anteriorly (fig. 1, U.) and caudally lo-
cated of these areas are partially exposed to the free surface
on the dorsal border of the thyroid gland, while the centrally
located one is embedded only a short distance below the dorsal
surface of the gland. The ultimobranchial body on the right
side has a structure similar to the left one. It extends through
a series of ten sections and is entirely embedded below the
dorsal surface of the thyroid gland.
In the left lateral half of the thyroid gland and just anterior
to the ultimobranchial body represented in figure 1 is the caudal
termination of an elongated area containing many large (cys-
toid) follicles. This area of large follicles is located just below
the dorsal border of the thyroid ‘gland and extends through
seventy consecutive sections. Very large follicles are also found
in the immediate neighborhood of the centrally located area of
the ultimobranchial body. Two rather large follicles lie near
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, NO. 1
16 J. A. BADERTSCHER
the right ultimobranchial body. A few large follicles are found
near the caudal end of the thyroid gland.
Pig 15 days old (fig. 2). The thyroid gland is 10.5 mm. long
and its greatest width is 6.3 mm. It tapers to a blunt point at
each end. Only the right ultimobranchial body is present. It
is embedded deeply below the dorsal surface in the posterior
half of the thyroid gland. It is represented by two areas which
are composed of syncytial cords and masses in which follicles
containing colloid are quite numerous, but on an average much
smaller than the majority of follicles in the immediate neigh-
borhood of these areas. The more anteriorly located and larger
of these areas (fig. 2, U.) extends through a series of eighty-five
sections, while the more caudally located one extends through
sixteen consecutive sections. Thirty-six sections intervene be-
tween the two areas.. The structure of the nuclei in these areas
is identical to that of the nuclei in the cells composing the
follicles. In the caudal fourth of the thyroid gland there are
many very large follicles which are located chiefly in the dorso-
lateral margin of the gland.
Pig 28 days old. The thyroid gland is 11.1 mm. long and its
greatest width is 7.7 mm. It tapers to a blunt point at each
end and its more bulky portion is crescent in shape in cross-
section. Both ultimobranchial bodies are present. They are
located just below the dorsal surface in the caudal portion of the
middle third of the thyroid gland, lateral to its medial plane.
The right one is similar in structure to the ultimobranchial body
in the thyroid gland of the 15-day-old pig. It extends through
a series of sixteen sections. The left ultimobranchial body is
composed largely of an attenuated syncytial mass in which are
found a few small follicles. It extends through a series of
thirty-eight sections.
In the immediate neighborhood of the left ultimobranchial
body are found a few very large follicles. Many very large
follicles are present in the taudal fourth of the thyroid gland.
These are located chiefly near the dorsal and dorsolateral border
of the gland, excepting near its caudal end where they are
scattered throughout the entire thickness of the gland.
ULTIMOBRANCHIAL BODIES IN POSTNATAL PIGS 17
Pig 42 days old (fig. 3). The thyroid gland is 14.6 mm. long
and its greatest width is 7.3 mm. It tapers to a point at each
end and, excepting near its ends, is crescent in shape in trans-
verse section. The ultimobranchial body on the right side is
located midway between the two ends of the thyroid gland and is
embedded just below the dorsal surface of the gland lateral to
its medial plane. It extends through a series of twenty-six
sections and is composed of syncytial cords and masses. In
both ends and in the peripheral portion of this structure the
follicles are quite numerous but comparatively small, while in
places its center is free from colloid. The ultimobranchial body
on the left side is located in the anterior portion of the caudal
fourth of the thyroid gland and is embedded just below the dorsal
surface of the gland lateral to its medial plane. It extends
through fifty-four consecutive sections and has a structure
(fig. 3, U.) similar to the right one. In place it is almost
separated from the rest of the thyroid gland by connective
tissue.
An area of large follicles in the right lateral half of the thyroid
gland extends anteriorly from the ultimobranchial body. This
area of large follicles is located chiefly just below the dorsal
border of the gland, but in places it extends to its dorsalateral
margin. In the dorsolateral portion of the middle third of the
left lateral half of the thyroid gland is an area, variable in width,
of very large (cystoid) follicles. Near the caudal portion of the
anterior third of the thyroid gland these areas of large follicles
become continuous just below its dorsal surface and gradually
extend more deeply into the gland so that large follicles are
found throughout its extreme anterior portion. No follicles of
an unusually large size are found in the extreme caudal portion
of the thyroid gland.
Pig 56 days old. The thyroid gland is 13.5 mm. fae and its
greatest width is 10.2 mm. It tapers to a blunt point at both
ends. The left lateral half of the gland is considerably more
bulky than the right lateral half. The only traces of an ulti-
mobranchial body is an area of very small follicles near the
dorsolateral margin in the right lateral half of the thyroid gland.
18 J. A. BADERTSCHER
This area extends through a series of ten consecutive sections
and is located nearly midway between the two ends of the
gland. A feature very noticeable in the thyroid of this pig is
the presence of only a small number of large follicles. These
are located chiefly near the dorsal border in the caudal fourth of
the gland.
Young adult hog no. 1 (fig. 4). The thyroid gland is 26 mm.
long and its greatest width is 18.5 mm. Its anterior end termi-
nates in a single blunt point, while its posterior end terminates
in two blunt processes each about 3.5 mm. long.. The only
structural feature representing a possible derivative of an ulti-
mobranchial body is an elongated area of small follicles (fig. 4,
U.) which extends through 125 consecutive sections. This area
is located along the dorsal border in the posterior fourth of the
right lateral half of the thyroid gland and extends for a short
distance into its right terminal process. More interfollicular con-
nective tissue is present in this area than in other parts of the
gland. The anterior two-thirds of the thyroid is free from
unusually large follicles. Many are found in the posterior third of
the gland. Two large follicles (C.F.) are represented in figure 4.
Young adult hog no. 2. The thyroid gland is 29 mm. long
and its greatest width is 20 mm. The anterior end terminates
in two blunt processes each 6.5 mm. long, while the posterior end
terminates in a single blunt point. The only structural feature
representing a possible remnant of an ultimobranchial body is
an area of small follicles in the posterior fourth of the left lateral
half of the thyroid gland. This area extends through a series
of forty sections and has a structure similar to that of the
ultimobranchial body represented in figure 4.
Two areas of large follicles are present. In the interior and
middle thirds of the thyroid gland these areas extend from the
tip end of the two anterior processes along the dorsolateral-
margin of the thyroid. In the caudal third of the gland these
areas gradually become larger, so that at the extreme caudal
end they are found throughout the entire thickness of the gland.
Young adult hog no. 3. The thyroid gland is 26.5 mm. long
and its greatest width is 20mm. At each end it tapers to a blunt
ULTIMOBRANCHIAL BODIES IN POSTNATAL PIGS 19
point. No remnants of the ultimobranchial bodies are present.
Only a few follicles of an unusually large size are present in the
extreme anterior and posterior ends of the gland.
SUMMARY
In pig embryos? it was found that the peripheral portion of
the ultimobranchial bodies generally develops into typical thyroid
structures before its more central portion. It thus becomes
evident that the ultimobranchial bodies which can be recognized
structurally as such in the thyroid gland of postnatal pigs are
mere remnants of these structures that have not fully developed
into typical thyroid structures. Also, the structure of the
ultimobranchial bodies in the late developmental stages of pig
embryos and in the postnatal pigs are similar, namely, areas
composed of nucleated syncytial cords and masses the central
portion of which may be free frum colloids. Areas of small
follicles (developmentally young) are not so marked in the
immediate neighborhood of the ultimobranchial bodies in post-
nata pigs as in the immediate vicinity of these structures in
most of the late embryonic developmental stages.
One of the ultimobranchial bodies in the thyroid gland of
three pigs is not a continuous structure, but is broken up into
segments. For example, the one on the left side in the 7.5-day-
old pig is composed of three oblong areas which vary in length.
It seems that a satisfactory explanation for this condition is a
more rapid transformation into typical thyroid structures of
some parts of the more central portion or core than of other
parts, thus dividing it into segments which are separated from
each other by follicles. The length of the portion of an ultimo-
branchial body broken up into segments is obtained by measur-
ing the distance between the anterior and posterior parts of the
most anteriorly and most posteriorly located segments, re-
spectively. Thus the left ultimobranchial body in the 7.5-day-
old pig extends through a series of seventy-five sections. The
2 In the summary it will be necessary to refer quite frequently to the writer’s
previous work on the ultimobranchial bodies in pig embryos. This will be done
without calling attention to the bibliographic reference of that article.
20 J. A. BADERTSCHER
right ultimobranchial body in the same pig is represented by a
single area which extends through a series of ten sections. Judg-
ing from the so variable developmental behavior of these
structures in pig embryos, this single area (and similar single
areas in the thyroid gland of other pigs) undoubtedly does not
represent the greatest length of the central core of this ultimo-
branchial body, but only what is left of it at the time the pig
was killed. It underwent a more rapid transformation than the
left one.
A considerable lapse of time intervenes between the age of the
pigs and the young adult hogs, thus producing an undesirable
break in the continuity of the postnatal developmental history
of the ultimobranchial bodies. Although the conclusion in re-
gard to these structures in the young adult hogs is thus rendered
somewhat uncertain, I have a strong feeling that the areas of
small follicles in the thyroid of the young adult hogs nos. 1 and 2
represent the ultimobranchial bodies in an advanced stage of
development. Their structure, location in the thyroid gland,
and their proximity to unusually large follicles strengthen this
- interpretation.
In some of the later developmental stages of pig embryos it
was found that unusually large (cystoid) follicles develop in
connect’on with the ultimobranchial bodies. The extent and
location of areas of this type of follicles in the thyroid gland of
postnatal pigs need therefore to be considered.
The existence of a possible interrelationship between the large
follicles and the ultimobranchial bodies is exemplified in a single
thyroid gland in scme of the postnatal pigs. Thus, in the pig
at birth the ultimobranchial body that is present is located in
the middle third of the left lateral half of the thyroid gland,
while an area of large follicles occupies an almost corresponding
position in the right lateral half of the gland. Also a few very
large follicles are found near the most caudally located segment
of the ultimobranchial body. In the thyroid gland in pig 7.5
days old large follicles are found in the immediate neighborhood
of the right ultimobranchial body and near the central segment
of the left one. Also cephalad to the most anteriorly located
ULTIMOBRANCHIAL BODIES IN POSTNATAL PIGS 21
area of the left ultimobranchial body is an elongated area, of
large follicles that extends into the middle third of the thyroid
gland. r9
a (ah EO
LMASSOPUST 78.
a
PLATE 3
EXPLANATION OF FIGURES
Oil-immersion objective and no. 4 ocular (Leitz) except fig. 8 which was made
with the high-dry lens (no. 7, Leitz) and fig. 10 which was made with the low-
power objective and no. 2 ocular (Leitz).
8 Carbon-containing endothelial cells lining a dilated lymph-vessel in the
subcutaneous tissue ten days after first subcutaneous injection (dog 152,
table 2).
9 Carbon-containing phagocytic mononuclear leucocyte and one containing
a polymorphonuclear leucocyte in the peripheral sinus of a retroperitoneal
lymph node one day after subcutaneous injection (dog 177, table 2).
10 Lymph nodule from a retroperitoneal lymph node ten days after subcu-
taneous injection (dog 152, table 2). Most of the carbon is in phagocytic mono-
nuclear leucoctyes in and near the sinuses.
11 Blood-vessel at the periphery of a germinal center (retroperitoneal lymph
node, dog 151, table 1). The endothelial lining shows carbon. 20 diams.
Fig. 6 Reconstruction showing the arteries of the right side of the pelvis and
right lower extremity in a human embryo of 18 mm. (C.E.1.C., 409) xX 20
diams.
69
70 H. D. SENIOR
as the a. interossea. The intervening part will be called the a.
poplitea profunda.
At this stage the a. poplitea profunda has two branches which
course longitudinally through the posterior crural region. One
of these arteries resembles, in a general way, the a. tibialis
posterior of the adult, it may be called the a. tibialis posterior
superficialis. The other slightly more distal in its place of
origin is the a. peronaea posterior superficialis (of Hyrtl).7 The
relations borne by these arteries to the n. tibialis remain constant
throughout a prolonged period of development.
The a. tibialis posterior superficialis passes at first backward
upon the medial side of the n. tibialis to reach its posterior
aspect. Running upon the posterior aspect of the n. tibialis in
the leg, it passes into the sole upon the inferior aspect of the n.
plantaris medialis. The part of the artery which enters the
sole becomes the a. plantaris medialis of the adult.
The a. peronaea posterior superficialis passes from the medial
to the lateral aspect of the n. tibialis by skirting around the
proximal and lateral sides of a large muscular ramus of the
nerve.’ It follows the anterior and lateral aspect of the n.
tibialis as far as the place of origin of the nn. plantares. In this
situation the artery bifurcates into a lateral and a medial branch.
The branches are both short and end blindly.
Two branches now spring from the lateral side of the a.
femoralis. One of these is situated upon the proximal side of
the r. saphenus and represents the r. musculo-articularis of the
adult a. genu suprema. The other is the a. circumflexa femoris
lateralis of the adult a. profunda femoris.
The branches arising from the medial side of the a. femoralis
which will be discussed under stages E and F.
The a. glutaea superior is present and takes origin from the
root of the a. ischiadica.
7 According to the system of nomenclature in use at the present time, the
adjective ‘posterior’ as used by Hyrtl is redundant. In view of the essentially
fugitive nature of anatomical terminology it has seemed unnecessary to modify
Hyrtl’s original term, which adequately meets the requirements of the case.
8 This is resolved later into the rami musculares for the mm. popliteus,
tibialis posterior, flexor digitorum longus, and flexor longus hallucis.
ARTERIES OF HUMAN LOWER EXTREMITY 71
e. Stage of 17.8 mm. H. E. C., no. 839, figs. 5 and 9 E
During the preceding stages of development the arterial retia
of the foot have received their blood supply exclusively through
the a. interossea; the reta plantare directly, and the rete dorsale
by means of the r. perforans tarsi. The mesenchymal skeleton
of the foot is now definitely circumscribed, and it is plain that
the latter vessel reaches the dorsum by way of the tarsal sinus.
The arterial retia of the foot lie in close contact with the tarsus
and metatarsus and represent the following vessels of the adult
circulation—the arcus plantaris and its articular branches, the
aa. tarseae mediales and laterales, the a. arcuata, the aa. meta-
tarseae and digitales (both dorsal and plantar), and the rami
perforantes (including the ramus plantaris profundus).
The present stage is characterized by the presence of four
arteries which traverse the leg for the supply of the sole. The
a. interossea is still present, the aa. tibialis posterior superficialis
and peronaea posterior superficialis have formed their definitive
connections, and a new vessel, the a. tibialis anterior, pars
distalis, has arisen.
The a. tibialis posterior superficialis and the a. peronaea
posterior superficialis preserve in the leg the same relations to
the n. tibialis as obtained in the preceding stage. At the ankle
the medial terminal branch of the a. peronaea posterior super-
ficialis has traversed the fork formed by the diverging plantar
nerves to join the a. tibialis posterior superficialis. The lateral
terminal branch of the same artery has reached the lateral side
of the sole to join the plantar arterial rete. The connection thus
formed between the a. tibialis posterior superficialis and the
rete plantare becomes the a. plantaris lateralis of the adult foot.
At the present stage of development it receives blood from two
sources.
° The greatest total length of this embryo, measured in formalin, was 17.8 mm.
In 80 per cent alcohol it measured 13.6 mm. It has been described by Thyng as a
17.8 mm. embryo (Am. Jour. Anat., vol. 17, p. 31), and referred to by Thyng
(Am. Jour. Anat., vol. 7, p. 489) and by Thyng and Lewis (Am. Jour. Anat., vol. 7,
p. 505) as an embryo of 13.6 mm. The distribution of the arteries of the lower
extremity resembles that of other embryos of about 15 or 16 mm.
v2 H. D. SENIOR
The part of the a. tibialis posterior superficialis distal to the
newly formed origin of the a. plantaris lateralis now becomes
the a. plantaris medialis. The portion of the remainder of that
vessel which is not concerned in the formation of the a. poplitea.
becomes the a. tibialis posterior.
The branches of the medial plantar artery anastomose, upon
the lateral side of the foot, with the plantar rete, thus consti-
tuting a (transitory) superficial plantar arch.
The a. tibialis anterior, pars distalis, extends from the part of
r. perforans cruris which lies in the extensor region of the leg to.
the rete dorsale. The portion of the r. perforans which now lies
beyond the proximal end of the pars distalis corresponds to the
adult a. recurrens tibialis anterior. The portion which extends.
from the a. poplitea profunda to the pars distalis enters into.
the composition of the adult a. tibialis anterior. The a. recurrens.
tibialis anterior is connected by means of a plexiform anastomosis,
with the r. saphenus of the a. femoralis.
Upon the medial side of the a. femoralis there are at this.
stage three branches exclusive of the r. saphenus. Of these the
most distal, which has a longer individual course than the others,
takes a recurrent direction. The three branches break up to.
form an extensive plexus (not indicated in the figures) which
ramifies throughout the thigh and is particularly rich around.
the perichondrium. It is probable that this plexus is eventually
taken over by the adult a. profunda femoris and its branches.
Two branches arise from the lateral side of the a. femoralis as.
in the preceding stage.
The a. glutaea superior now takes direct origin from the a. hy-
pogastrica upon the proximal side of the origin of the a. ischiadica.
f. Stage of 18 mm. C. 1. E. C., no. 409, figs. 6 and 9 F
From the time of the junction between the r, communicans
superius and the axial artery the caliber of the a. femoralis has.
gradually exceeded that of the a. ischiadica, which has now
become exceedingly slender. After having traversed the lower
portion of the sacropudendal plexus, the a. ischiadica passes.
ARTERIES OF HUMAN LOWER EXTREMITY io
to the region of the hiatus tendineus upon the lateral side of the
n. tibialis. The n. peronaeus lies immediately upon its lateral
side.
In a general way the aa. poplitea profunda and interossea
and their branches have the same arrangement as in the pre-
ceding stage. The exact course taken by these vessels can now
be clearly recognized, since it is possible to identify the individual
muscles of the limb.
The a. poplitea profunda runs between the m. popliteus and
the tibia. At the proximal border of the muscle the artery
gives off the a. genu media and a short trunk which is being
formed by the progressive union of the proximal ends of the aa.
tibialis posterior superficialis and the a. peronaea posterior super-
ficialis. This short trunk, which lies upon the posterior surface
of the m. popliteus, may now be called the a. poplitea super-
ficialis. It forms the distal part of the a. poplitea of the adult.
A short distance below the origin of the a. poplitea superficialis
there arises the a. genu inferior medialis.
The a. interossea passes down the leg between the m. tibialis
posterior and the interosseous membrane. Just above the
medial malleolus it gives off a branch which passes around the
posterior to the medial side of the tibia, the ramus coronarius
of the medial malleolus (Hyrtl).1° At the malleolus the a.
10 In this vessel lies, according to Hyrtl, the key to the deep supramalleolar
anastomosis (i.e., the r. communicans) between the a. peronaea and the a.
tibialis posterior. The r. coronarius is regarded by Hyrtl as primarily passing
from the a. peronaea around the medial side of the tibia to unite with the a.
tibialis anterior. By means of a secondary connection occurring between the r.
coronarius and the a. tibialis posterior, the former vessel eventually becomes
converted into an anastomosing chain in which the aa. malleolaris anterior
medialis, malleolaris posterior medialis and r. communicans are the named
components. At thestage of 18 mm. of this series the r. coronarius arises from the
part of the a. interossea which persists to take part in the formation of the adult
a.peronaea. It runs, at this stage, as at the stage of 22 mm., toward the a. tibialis
anterior as described by Hyrtl, and would, no doubt, eventually join it. Since
in both these stages the a. tibialis posterior is situated upon a plane superficial
to that occupied by the r. coronarius, no junction of the vessels in question has
occurred. There can be little doubt, however, that Hyrtl has interpreted the
nature of the r. coronarius correctly. His views upon this subject, which were
derived from an extensive study of the normal and abnormal conditions found
in adult legs, have been of the greatest assistance in the interpretation of the
conditions obtaining during development.
74 H. D. SENIOR
interossea lies upon the lateral side of the tendon of the m.
tibialis posterior (i.e., between tendon and malleolus) and then
passes across the plantar end of the sinus tarsi to join the plantar
rete.
The a. peronaea posterior superficialis is placed anteriorly and
laterally to the n. tibialis and les upon the posterior surface of
the m. flexor hallucis longus. At the distal border of the muscle
the artery lies near the a. interossea and interosseous membrane.
In this situation it gives off a very large r. calcaneus lateralis
which runs upon the lateral side of the tuber calcanei. This
branch eventually forms the termination of the a. peronaea.
The a. tibialis posterior superficialis follows the posterior
surface of the n. tibialis until the latter bifurcates in the neigh-
borhood of the medial malleolus. Here the artery is placed
between the n. tibialis and the tendon of the flexor hallucis
longus. The a. tibialis anterior, pars distalis, has relations
identical with those of the adult.
The relations existing between the various structures of the
leg were ascertained by making a reconstruction which, from
the fact that it yields but little information when entirely built
up, is unsuitable for reproduction. The data gained from an
examination of its separate parts are embodied in two diagrams
(fig 7, A and B) which may serve as a rough guide to the inter-
pretation of anomalies. These diagrams show, approximately,
the course taken by the embryonic aa. poplitea profunda,
interossea, and peronaea posterior superficialis.
The order in which the various structures passing from the
leg to foot are arranged in the hollow between the medial mal-
leolus and tuber calcanei is as follows: m. tibialis anterior and
a. interosseus together; m. flexor digitorum longus; a. peronaea
posterior superficialis; n. tibialis, a. tibialis posterior super-
ficialis, and m. flexor hallucis longus.
Embryos of about this age are instructive mainly by reason
of the fortunate circumstance that the individual muscles are
differentiated before the continuity of the axial artery has
been broken. Only one important change has occurred at this
stage of development; this consists in the appearance of a com-
ARTERIES OF HUMAN LOWER EXTREMITY 15
municating branch which foreshadows the development of the
future a. peronaea. :
- The communicating branch in question leaves the a. peronaea
posterior superficialis at the proximal border of the m. flexor
hallucis longus. It passes distally between the mm. flexor
hallucis longus and tibialis posterior to join the a. interossea,
t.p., Mm. tp, m.
a |||, Fe)
5
Fig. 7 Indicates the course of three arteries of the embryonic lower extremity,
represented diagrammatically as they would appear if persisting in the adult.
The vessels which normally persist are indicated by shading. A, A. poplitea
profunda and a. interossea. B. A. peronaea posterior superficialis. d. 1. m.
flexor digitorum longus; h. l., m. flexor hallucis longus; t. p., m. tibialis posterior.
as that artery lies upon the interosseous membrane. It may be
called the r. communicans inferius.
The entire r. communicans inferius persists in the adult as
the portion of the a. peronaea which lies between the mm. flexor
hallucis longus and tibialis posterior. The portions of the aa.
peronaea posterior superficialis and interossea with which the
76 H. D. SENIOR
proximal and distal ends of the r. communicans inferior are
respectively connected also take part in the formation of the
a. peronaea. ‘
Of the branches of the a. femoralis, those arising from the
lateral side are identical with the branches of the preceding
stage. There are three branches upon the medial side. The
recurrent branch (R in figures) seems to be identical with the
similar branch of the preceding stage. It is doubtful whether
the two other medial branches correspond to those observed in
the preceding stage or not.
g. Stage of 22mm C.I.C no. 1, figs. 8 and 9 G
At this stage of development the continuity of the a. ischiadica
has been interrupted and the a. femoralis alone conveys blood
to the region beyond the knee.
The femoral artery pursues a more direct course than before
and gives origin to most of its important branches. In addition
to the a. circumflexa lateralis which appeared at the stage of
14 mm., it gives origin to the a. pudenda externa and the a.
profunda. The r. musculo-articularis has migrated to the em-
bryonic r. saphenus. The root of the latter branch, therefore,
now appears as the a. genu suprema and gives origin to the r.
saphenus and the rr. muscularis and articulares of the adult.
The a. profunda arises from the a. femoralis quite close to the
origin of the a. circumflexa lateralis. It seems probable that
the embryonic a. profunda sometimes ar:ses from the root of
the latter artery and sometimes directly from the femoral. In
the former case the adult a. circumflexa lateralis would
appear to arise from the a. profunda and in the latter from the
femoral. The great variation in the site of the origin of the
vessel is well known.
The a. profunda femoris gives origin to one perforating artery
which, since it pierces the adductores brevis and magus, repre-
sents the first or second of the adult series. The single a. per-
forans is connected, by means of an extensive plexus, with the
more distal section of the now interrupted a. ischiadica.
ARTERIES OF HUMAN LOWER EXTREMITY 77
_ The a. ischiadica passes through the sacropudendal plexus as
before and reaches the lateral side of the a. tibialis, which is now
contained in the same sheath as the n. peronaeus. After its
exit from the pelvis the artery gives origin to a few gluteral
branches and leaves the n. tibialis with the n. cutaneus femoris
posterior. In company with the latter nerve and the v. ischiadica,
it passes to the posterior surface of the thigh and ends by
dividing into several cutaneous branches. This part of the a.
ischiadica persists as the a. glutaea inferior.
The more distal portion of the a. ischiadica is traceable as a
definite path through a plexus upon the posterior surface of the
m. adductor magnus, but has lost its continuity with the a.
poplitea. The other vessel participating in the formation of
the plexus is the perforating branch of the a. profunda femoris.
This plexus evidently represents the terminal anastomosis
between the perforating arteries of the adult. It probably takes
a large share in the formation of the perforating arteries
themselves.
The distal part of the anterior tibial artery has received blood
since the time of its first appearance by way of the a. poplitea
profunda and r. perforans cruris. An alternative path has now
been provided by the appearance of a vessel which, passing
around the distal border of the m. popliteus, connects the
developing a. peronaea with a more distal part of the a. poplitea
profunda. It is the r.communicans medius which will later become
the definitive proximal end of the a. tibialis anterior. The
significance of this vessel in relation to the formation of the
adult aa. profunda and tibialis anterior will be dealt with in the
succeeding section.
The principal changes which have occurred in the embryonic
arteries of the posterior crural region since the preceding stage
of development involve the vessels which participate in the
formation of the adult a. peronaea. These changes cons st in
the disappearance of considerable parts of the aa. interossea and
peronaea posterior superficialis.
The portion of the a. interossea which formerly extended from
the r. perforans cruris.to the distal end of the r. communicans
78 H. D. SENIOR
a. iliaca. com.
a. iliaca. ext.
q. hypogast.
a. epigast. inf.
@CiiGal| 3 p,0.
a. pudend. ext. F
gd. circ. fem. lat.—_«g
a. prof. femor. \
dg. genu. suprem.
rf muse. et artic:
‘
q. perforans.
a. ischiad.
a.genu med.
a. popl. prof.
a. popl. superf.
r. com. med.
a. peron. p. superf.
lL 6. peronsea:
a. interos.
Sik ele peron. p. superf.
___ — ram. calean. lat.
_@. plant. lat.
arcus plant.
r. saph sete
a. genu. inf. med.
a. tb. p. superf.
GICOLONnGi: aaa We
r. pert. ag ae se
rete calc.
a. plant. med. (cut)
f.. profund.
FEPRiOn
Fig. 8 Reconstruction showing the arteries of the right side of the pelvis
and right lower extremity in a human embryo of 22 mm. (C.E.I.C., 1.). Medial
aspect. X 20 diams.
ARTERIES OF HUMAN LOWER EXTREMITY 79
inferius has practically disappeared. ‘The portion immediately
beyond the distal end of the latter vessel lies upon the posterior
surface of the interosseus membrane and gives origin to two
branches. The remainder of the vessel is involved in the meshes
of the rete calcaneum.
The portion of the a. peronaea posterior superficialis which
extends from the present termination of the a. poplitea super-
ficialis to the proximal end of the r. communicans inferius per-
sists to become a part of the adult a. peronaea. A large part of
the remainder of the artery has. been lost, but the distal portion
of the vessel can still be recognized as forming a projection upon
the proximal aspect of the rete caleaneum. This portion of the
artery can be traced through the meshes of the rete as far as
the a. plantaris lateralis.
The rete calcaneum is very extensive at this stage of develop-
ment and occupies the concavity upon the medial side of the
caleaneum and surrounds the deep flexor tendons in the
malleolar region of the leg.
The component parts of the a. peronaea are now recognizable.
The proximal part is derived from the a. peronaea posterior super-
ficialis. The part between the aa. flexor hallucis longus and
tibialis posterior represents the entire r. communicans inferius.
The immediately succeeding part, which rests upon the mem-
brana interossea, is derived from the a. interossea. The terminal
part of the artery consists mainly of the r. calcaneus lateralis of
the a. peronaea posterior superficialis. The connection between
the a. interossea and the part of the latter artery which gives
origin to the caleanean branch is SugerE through the agency of
the plexus caleaneum.
The part of the a. interossea which takes part in the formation
of the a. peronaea has been noted as giving rise to a branch, the
r. coronarius, at the stage of 18 mm. At the present stage
another branch arises from it, namely, the r. perforans. Both
of them become branches of the adult artery. At the stage of
22 mm. the r. coronarius can be traced around the posterior
and medial aspects of the tibia almost as far as the a. tibialis
anterior. The views expressed by Hyrtl upon the conversion
!
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, No. 1
n. gen. fem. a.iliac.ext.
oa. mes. inf. ; g n. fem. eS
a. umbilic. g.hypog SS
{red ventr n. per. SS
radix dors. a
L3 plex.
a axis. dors
a. pudend. int i q. Iliac. com.
plex. pelv. SNa. pudenel. int.
-LS. gd. OXIS.
q. glutaea. sup.
a. glut. inf.
a. circ. il. prof.
a. epigast. inf.
a. circ. fem. lat.
a. prof. femor.
n.cut. fem. post.
a. glut. inf.
a. perforans.
gd. genu. suprem.
a. popl. superf.
a.tib.psupert .
_-@. peron. p. supert.
rete calcan.
my =-q, plant. med.
im
RET}.
MM
qd. peron. p. superf. :
a. peronaed. / a. plant.lat. + refund.
q.dorsal. ped. F EP
9
Fig. 9 Reconstructions showing the arteries of the right lower extremity in
the seven human embryos shown in the preceding figures. The cephalic (em-
bryonic pre-axial, adult medial) aspect is shown in all cases. A, X 40, the others
X 20 diams. The skeleton is‘shown, in part, in E, F and G. Parts of the tibia
80
G. 22 mm.
qa. fernor
a. epigast. inf.
r. saphen.
g.iliaoco ext.
@ epigast. inf.
oa. circ. fem. lat.
n. saphen. n. femor. ?
Ff. Com. sup. Sr
mob
a. axis Q a
> r muse.
gee , artic.
rete plantar.
r.saphen,/ £ Q
g. pudend. int. nN. peron: E ~ a. tib p. supert.
n. tibial.
a. ilioc. ext.
a. epigast int.
a circ. fem. lat. pw AE
a. genu. inf. med.
a. recur. t.@. / «=m
a. tib. ant. /
past
\e pudend. inf.
. ischiad.
. tih. p. super.
a. peron. p. supert
r. calc. lat.
r. com. int
Pak
“a. plant. lat.
a. peron.p. supert.
a. pop!. prot.
a.recur.t.a.
r pert crur.
@.interos. |
r pert. tarsi.
rete. plantar.
14 mm.
rete.
a. epigast. inf.
a. circ.fern, lq
r. musc.-artic.
. ischiad.
= \% . popl. prof.
rper B AGE —«. tib. p-superf.
. peron. p. superf
go.
Zs
. @.tib. ant . plant. med.
r pert. tarsi. :
rete plantar p. distal. . plant. lat.
. perf. tarsi.
and tarsus are omitted in E, the tibia in F and the tibia and medial femoral
condyle in G. The fibula is omitted in all. The medial plantar artery has
been almost entirely removed from E and F. In C, a. femor. should read a.
iliaca ext.
81
82 H. D. SENIOR
of the r. coronarius into the communicating branch and medial
malleolar arteries of the adult have been noted above (footnote
WO; th G3):
A small portion of the a. peronaea posterior superficialis
appears at the present stage as a branch of the a. peronaea. It
runs upon the posterior surface of the m. flexor hallucis longus.
The disintegrating a. interossea has left a similar vestige of its
proximal end upon the posterior surface of its interosseous
membrane. This vestigial branch occasionally persists as a
branch of the adult a. tibialis anterior.11. The terminal part of
the main trunk of the a. peronaea posterior superficialis is now
involved in the rete caleaneum. It usually leaves its mark in
the adult as a recurrent element among the leash of branches —
known as the r. calcaneus medialis of the a. plantaris lateralis.
The aa. iliolumbalis and sacralis lateralis now arise from the
a. glutaea superior. The root of the latter artery has conse-
quently become the posterior division of the adult a. hypo-
gastrica. All the visceral arteries of the pelvis, with the excep-
tion of the a. vesicalis superior, have assumed their adult
relations.
h. Changes occurring in the principal arieries subsequent to the
stage of 22 mm.
A. Formation of the adult a. poplitea and a. tibialis anterior.
Fig. 10, AS Co The.) 22: B,C EXCrAl Cr Cr ls tne eae
The r. communicans medius, the presence of which was noted
at the stage of 22 mm., seems to arise at about that stage of
development. In C. E. C., no 1, it is larger upon the right side
than upon the left and in C. E. C, no. 6, which also measures
22 mm., it is absent altogether.
11 Mr. Kimbrig has kindly dissected six adult legs for evidence of persistence
of this vessel. In two instances he found a branch accompanying the n. interos-
seus cruris for a short distance. In both cases the branch arose from the root of
the ramus fibularis.
It is very questionable whether the B. N. A. is correct in classifying the r.
fibularis as belonging to the a. tibialis posterior. It has usually been regarded,
in English-speaking countries at all events, as a branch of the a. tibialis anterior.
ARTERIES OF HUMAN LOWER EXTREMITY 83
At the stage of 20 mm. (fig. 10, A) blood traverses the pop-
liteal region to reach the r. perforans cruris through the original
channel, namely the a. poplitea profunda. The a. poplitea
superficialis is longer than at the stage of 18 mm., having in-
creased in length at the expense of the aa. tibialis posterior
superficialis and peronaea posterior superficialis.
At the stage of 22 mm. (fig. 10, B), a junction between the
developing a. peronaea and the distal part of the a. poplitea
Fig. 10 Three reconstructions, each showing a sagittal segment (0.25 mm.
thick) from the left leg of an embryo of the length indicated. Lateral aspect.
xX 19. A, 20 mm.; B, 22 mm.; C, 24.8mm. The tibia appears in all cases. P.
indicates the position of the m. popliteus. c.m., a. communicans media; g. 7. m.,
a. genu inferior medialis; g. m., a. genu media; 7., a. interossea; 7.m., interosseous
‘membrane; p., a. poplitea; p.c., a. perforans cruris; p. d., a. tibialis anterior, pars
distalis; p. p., a. poplitea profunda; p. s., a. poplitea superficialis; per., a.
peronaea; 7. a., a. recurrens tibialis anterior; r. p., a. recurrens tibialis posterior;
t. a., a. tibialis anterior; ¢. p. s., a. tibialis posterior superficialis.
profunda has been effected by the development of the r. com-
municans medius. Blood may now pass from the proximal part
of the a. poplitea profunda to the a. tibialis anterior, pars dis-
talis by two routes: 1) anterior to the m. popliteus, by way of
the distal part of the a. poplitea profunda and r. perforans cruris,
or 2) posterior to the m. popliteus, by way of the a. poplitea
superficialis, developing a. peronaea, r. communicans medius, a.
poplitea profunda, and r. perforans cruris.
84 H. D. SENIOR
At the stage of 24.5 mm. (fig. 10, C) the direct arterial route
upon the anterior surface of the m. popliteus has ceased to
exist. The part of the a. poplitea profunda not incorporated in
the definitive a. poplitea or tibialis anterior is represented:
1) by the root and a small branch of the a. genu inferior medialis
of the former and 2) by the a. recurrens tibialis posterior of the
latter. The main arterial channel through the popliteal region
has thus been transferred from the anterior to the posterior
surface of the popliteus muscle. Both the a. poplitea and the
a. tibialis anterior have assumed their adult arrangement.
The formation of the adult a. poplitea results from the com-
bination of two embryonic elements. The part of the vessel
which extends from the hiatus tendineus to the origin of the a.
genu inferior medialis is derived from the embryonic a. poplitea
profunda. The remainder of the artery corresponds to the
embryonic a poplitea superficialis.
The embryonic components of the a. tibialis anterior consist
of the r. communicans medius, a short section of the distal end
of the a. poplitea profunda, the proximal part of the r. perforans
cruris, and the entire tibialis anterior, pars distalis.
The a. poplitea superficialis reaches completion at the stage
of 24.5 mm. by extending as far as the r. communicans medius
which then becomes one of its terminal branches. By this
means the a. peronaea relinquishes its temporary participation
in the formation of the a. tibialis anterior.
B. The conversion of the embryonic a. tibialis posterior super-
ficialis into the adult a. tibialis posterior.
The a. tibialis posterior superficialis is originally a branch of
the a. poplitea profunda and extends into the sole. The distal
portion of the artery, marked off from the remainder of the
vessel upon the development of the a. plantaris lateralis, becomes
the a. plantaris medialis. The proximal portion of the a. tibialis
posterior superficialis has blended with the corresponding section
of the a. peronaea posterior superficialis at the stage of 24.5 mm.
as far as the termination of the popliteal artery. The further
blending between these two arteries which is to occur during
the later stages of development will cause the migration of the
ARTERIES OF HUMAN LOWER EXTREMITY '85
origin of the a. peronaea from the termination of the a. poplitea
to a point upon the a. tibialis posterior. :
After the formation of the a. tibialis posterior has been com-
pleted by the occurrence of these changes the vessel still retains
its original relation to the n. tibialis. The relations between
the a. poplitea and the n. tibialis also differ from those of the
adult.
The entire a. poplitea lies upon the medial side of the n.
tibialis. The a. tibialis posterior crosses the medial side of the
nerve to gain its posterior aspect upon which it runs until it
ends behind the medial malleolus. The relations of the aa.
poplitea and tibialis posterior to the n. tibialis have been
thoroughly studied up to the stage of 22 mm. They seem to
remain unaltered in the oldest serially cut embryo which has
been examined in this regard. It is C. I. E. C., no. 1134, which
measures 33.5 mm. .
It is not difficult to see how an agency capable of moving
the termination of the a. poplitea across the anterior aspect of
the n. tibialis from the medial to the lateral side would produce
a condition differing little from that normally encountered in
the adult. Since the knee is flexed throughout intrauterine life
it would seem that the modification of the relative positions of
the arteries and nerve might depend upon the straightening of
the n. tibialis which occurs at birth. In order to put the matter
to the test, a foetus of 7.9 cm. was dissected.!2 The rela‘ions,
however, were found to be identical with those of the adult.
Apart from the study of a few special points, this investigation
has not extended beyond the stage of 22 mm. At that period
all the vessels of the adult limb are present with the exception
of two of the three aa. perforantes, which usually arise from the
adult a. profunda femoris, the a. circumflexa femoris medialis,
and the a. obturatoria. At the stage of 22 mm. two obturator
veins are present in C. E. C., no. 1, one of these is tributary
to the v. hypogastrica, the other to the v. femoralis.
The leading features in the process of the arterial development
of the limb are indicated in figure 11.
122 For this dissection and for many others made upon various mammals, I
wish to express my indebtedness to Mr. B. Spector.
86 H. D. SENIOR
III. BRIEF DEVELOPMENTAL HISTORY OF THE INDIVIDUAL
ARTERIES
A. Axis. This artery is a branch of the dorsal root of the a.
umbilicalis, its presence has been noted by Tandler as early as
the stage of 5mm. ’03). At the stage of 6 mm. it terminates by
giving origin to the r. perforans tarsi and to the rete plantare.
The axial artery has three rami communicantes, and gives
origin to three rami perforantes as follows:
The r. communicans superius, from the a. femoralis, joins it
near the hiatus tendineus shortly before the stage of 14 mm.
The r. communicans medius, from its own branch the a.
peronaea posterior superficialis, has joined it near the distal
border of the m. popliteus at the stage of 22 mm.
The r. communicans inferius, from the a. peronaea posterior
superficialis, joins it, near the distal border of the m. tibialis
posterior, between the stages of 17 and 18 mm.
The r. perforans cruris arises between the stages of 13 and
14 mm.
The r. perforans artertae peronaea arises between the stages
of 18 and 22 mm.
The r. perforans tarsio is present at the earliest stages of de-
velopment at which the axial artery has been observed.
Two points have been marked upon the axial artery at the
stage of 14 mm. which may be used for the convenient sub-
division of the axial artery into three parts. These points
correspond to the termination of the r. communicans superius
and to the origin of the r. perforans cruris, respectively.
The respective parts of the artery are defined and named as
follows: The part upon the proximal side of the termination of
the r. communicans superius is the a. ischiadica, while that
upon the distal side, of the origin of the r. perforans cruris is
the a. interossea. That part intervening between the other two
is the a. poplitea profunda. Further information regarding the
axial artery may be found under the headings devoted to the
description of its respective parts.
A. dorsalis pedis. ‘This artery is a channel through the em-
bryonic rete dorsale. Its importance dates from the appearance
a. iliac. ext.
a. epig. inf inf" _,
‘
i
&
&
a. prof. fem.
ones
g.femor.
is saphenu
a. genu Sup.
a. popl. prof.
a. recur. tib. post
F. perf.cruris:
i}
" Ste
Bee)
SA
vs
\:
ETA
, WY
Bose
a.tib.ant.,pars dista/_—+
a. interosseq——
r. perf. PUTER 4 a a
rete plant. te
a. dors. ped. pe
arcus plant. a,
Fig. 11 Diagram to illustrate the general development of the arteries of the
human lower extremity. Adult arteries are stippled and their names underlined.
The chief embryonic channels are outlined in black. The black line is
continuous only in the case of the axial artery, otherwise it is broken.
letter P indicates the position of the m. popliteus; 7, that of the m. tibialis
posterior, and H that of the m. flexor hallucis
87
a. iliac. com. dext.
a. hypog.
a. glut. sup.
a. glut. inf.
a. pudend. int.
a. ischiadica.
r. commun. sup.
r. commun, med.
a.tib. a.tib. poster
<—a. tib. post. superf
-r. commun. int.
+#—a. peron. p. super.
@. peronaea.
a. plant. med.
longus.
ie ad
88 H. D. SENIOR
of the distal part of the a. tibialis anterior which is developed
between the stages of 15 and 16 mm.
Branches. One of the adult rr. tarseae laterales represents
the dorsal end of the r. perforans tarsi, all other branches are
derived from the rete dorsale.
A. femoralis. This artery arises from the a. iliaca externa
between the stages of 11 and 12 mm. From the time of the
first appearance of the femoral artery its growing end is bifur-
cated into the r. communicans superius and the r. saphenus.
The r. communicans superius traverses the hiatus tendineus and
joins the a. axis between the stages of 12 and 14 mm., while the
r. saphenus terminates below the region of the knee-joint.
Branches. The A. profunda femoris seems to be derived from
an arterial plexus which envelops the femur as early as the stage
of 14mm. The proximal end of the artery is distinguishable at
the stage of 22 mm. The a. circumflexa lateralis arises from the
femoral trunk shortly before the stage of 14mm. Althoughthis
artery is commonly regarded as a branch of the a. profunda, its
original connection with the a. femoralis is frequently retained
in the adult. The a. circumflexa medialis is not present at the
stage of 22 mm. One of the a. perforantes, the first or second,
is present at the stage of 22 mm., and is involved at that period
of development in an extensive arterial plexus connected with
the remainder of the distal part of the a. ischiadica. The other
perforating arteries are not present. at the stage of 22mm. The
a. pudenda externa is the only superficial inguinal branch present
at the stage of 22 mm. The a. genu suprema corresponds to
the part of the r. saphenus upon the proximal side of the origin
of the rr. musculares and articulares. These branches arise
directly from the a. femoralis, but have migrated to the em-
bryonic r. saphenus by the stage of 22mm. The distal portion
of the latter vessel corresponds to the r. saphenus of the adult.
A. hypogastrica. The main stem of this artery represents the
part of the dorsal root of the a. umbilicalis which lies beyond
the place of origin of the a. iliaca externa.
Branches. The anterior division of the a. hypogastrica is de-
rived from the proximal part of the a. axis, which becomes the
ARTERIES OF HUMAN LOWER EXTREMITY 89
a. glutaea inferior, from the root of the a. pudenda interna, and
from the part of the a. umbilicalis beyond the distal end of its
dorsal root. The portion of the original a. umbilicalis which
contributes to the formation of the a. hypogastrica is the source,
in all probability, of all the visceral branches of the artery. The
a. obtoratoria is not present at the stage of 22 mm. In conse-
quence of the transference of the origins of the aa. iliolumbalis
and sacralis lateralis to the a. glutaea superior, the proximal
part of that artery becomes the posterior division of the a.
hypogastrica.
A. iliaca communis. This artery is derived from the portion
of the dorsal root of the a. umbilicalis which lies upon. the
proximal side of the origin of the a. iliaca externa.
A. iliaca externa. This artery already springs from the con-
vexity of the a. iliaca communis at the stage of 8.5 mm. Its
termination is unbranched until a period shortly before the stage
of 12 mm., when it divides into the a. epigastrica inferior and
the a. femoralis. The a. circumflexa ileum profunda arise
from the a. iliaca externa between the stages of 18 and 22 mm.
A. interossea. The course of this part of the axial artery is
described on page 73.
The a. interossea is joined before the stage of 18 mm. by the
distal end of the r. communicans inferius. The junction occurs
at a point immediately beyond the inferior margin of the m.
tibialis posterior. This artery has given origin to the r. coro-
narius at the stage of 18 mm. and to the r. perforans and to
the branch of communication with the a. peronaea posterior
superficialis at the stage of 22 mm.
The part of the a. interossea extending from the termination
of the a. poplitea profunda to the distal end of the r. communicans
inferius disappears between the stages of 18 and 22 mm. A
vestige of the proximal end of this part of the artery sometimes
persists in the form of a small branch of the a. tibialis anterior
or of the r. fibularis.
The small portion of the a. interossea which extends from the
distal end of the r. communicans inferius to that of the membrana
interossea persists as the third part of the a. peronaea (p. 79).
90 H. D. SENIOR
The branches which arise from this portion of the a. interossea
are transferred to the a. peronaea. They are the r. coronarius
malleolaris, part of which becomes the adult r. communicans,
and the r. perforans. The remainder of the a. interossea has.
lost its identity at the stage of 22 mm. The r. perforans tarsi
disappears at about the same period.
A. ischiadica. The course taken by this part of the axial
artery is described on page 73.
The continuity of the a. ischiadica has been broken in the
region of the glutaeal fold, at the stage of 22 mm. The proximal
part of the vessel, which has now become the a. glutaea inferior,
follows the v. ischiadica and the n. cutaneus femoris to the
surface of the limb where it ends by dividing into cutaneous
branches. The more distal portion of the artery is involyed,
with the single perforating artery present at the stage of 22
mm., in the formation of an extensive plexus upon the posterior
surface of the m. adductor magnus. _
A. peronaea. This artery is clearly recognizable at the stage
of 22 mm. (p. 79). It may be divided into four parts:
The first part of the a. peronaea, which extends from the
origin of the artery to the proximal margin of the m. flexor
hallucis longus, is a persisting portion of the a. peronaea posterior
superficialis (p. 74).
The second part, which lies between the m. tibialis posterior
and the m. flexor hallucis longus, represents the entire embryonic
r. communicans inferius (p. 76).
The third part, which is short and rests upon the interosseus
membrane near the distal border of the m. tibialis posterior, is
a persisting portion of the a. interossea (p. 73).
The fourth part, which ramifies upon the fibula and caleaneum
in the neighborhood of the lateral malleolus, represents a sur-
vival of the short portion of the a. peronaea posterior super-
ficialis and of the entire r. calcaneus lateralis of that vessel
(p. 74).
Branches. The r. perforans and r. communicans, belong
orig nally to the a. interossea. The rr. calcanei laterales are.
branches of the r. calcaneus lateralis of the a. peronaea posterior
superficialis.
ARTERIES OF HUMAN LOWER EXTREMITY 91
A. peronaea posterior superficialis. The course of this em-
bryonic artery is described on pages 70 and 74. The main stem
of the vessel is present at the stage of 14 mm., its terminal
branches form their definitive connections at a stage of 15 or 16
mm.
The proximal part of the a. peronaea posterior superficialis
unites with the a. tibialis posterior superficialis to form the .a.
poplitea superficialis and the part of the a. tibialis posterior
above the root of the a. peronaea. A more distal portion persists
as the first part of the a. peronaea, and a still more distal portion
enters slightly into the formation of the fourth part of that
vessel.
The terminal branches of the a. peronaea posterior super-
ficialis form the adult a. plantaris lateralis. The r. calcaneus
medialis of the latter artery represents the end of the stem of
the parent vessel. The r. calcaneus lateralis, which arises from
the a. peronaea posterior superficialis shortly before the stage
of 18 mm., persists to form the major portion of the fourth part
of the a. peronaea (p. 79).
A. plantaris lateralis. The proximal part of this artery
represents the medial terminal branch of the a. peronaea pos-
terior superficialis (p. 71), which unites with the a. tibialis
posterior superficialis. The distal part represents the lateral
terminal part of the same vessel.
Branches. The r. caleaneus medialis represents the terminal
portion of the main trunk of the embryonic a. peronaea posterior
superficialis. The arcus plantaris and all its branches are de-
rived from the embryonic rete plantare.
A. plantaris medialis. This artery is the distal portion of the
embryonic a. tibialis posterior superficialis. It is marked off
- from the remainder of that vessel by the medial terminal branch
of the embryonic a. peronaea posterior superficialis.
A. poplitea. The part of this artery above the origin of the a.
genu inferior medialis represents a surviving portion of the
embryonic a. poplitea profunda (p. 73). The part lying upon
the posterior surface of that muscle represents the embryonic a.
poplitea superficialis. The proximal portion of the a. genu
inferior medialis is derived from the a. poplitea profunda.
92 H. D. SENIOR
Branches. The a. genu media is present at the stage of 18
mm. The other branches appear at a later stage of the
development.
A. poplitea profunda. The course of this vessel is described
on page 73. The part of the a. poplitea profunda which extends
from the hiatus tendineus to the origin of the a. genu inferior
medialis becomes the proximal part of the adult a. poplitea.
The remainder of the artery is represented in part by the root
of the a. genu inferior medialis. It is represented also by the a.
recurrens tibialis posterior and by the second part of the a.
tibialis anterior (p. 84).
A. poplitea superficialis. The formation of this artery results
from the gradual union which takes place between the proximal
part of the embryonic a. tibialis posterior superficialis and that
of the a. peronaea posterior superficialis. The vessel lies upon
the posterior surface of the m. popliteus and has nearly reached
completion at the stage of 22 mm. The a. poplitea superficialis
persists as the distal portion of the adult a.. poplitea (p. 84).
A. tibialis anterior. This vessel may be divided into four
parts which correspond to the four embryonic components of
the adult artery.
The first part extends from the origin of the vessel to the root
of the a. recurrens tibialis posterior. It corresponds to the
whole of the r. communicans medius (p. 84).
The second part occupies the immediate neighborhood of the
origin of the a. recurrens tibialis posterior. It represents, like
that artery, a persisting portion of the embryonic a. poplitea
profunda (p. 84).
The third part extends from the a. re¢currens tibialis posterior
to a point immediately beyond the root of the a. recurrens
tibialis anterior. It is derived from the proximal portion of
embryonic r. perforans cruris (p. 68).
The fourth part of the artery extends from the termination of
the third part to the proximal end of the a. dorsalis pedis. It
represents the entire a. tibialis anterior, pars distalis, of the
embryo (p. 72).
ARTERIES OF HUMAN LOWER EXTREMITY 93
Branches. The a. recurrens tibialis anterior represents NG
terminal portion of the a. perforans cruris (p. 68).
The a. malleolaris anterior medialis is derived from the r.
coronarius of the a. interossea which arises shortly before the
stage of 18 mm.
A. tibialis aniertor, pars distalis. This embryonic artery
arises at about the stage of 15 or 16 mm. Its course is identical
with that of the fourth part of the a. tibialis anterior of the
adult.
A. tibialis posterior. The part of this artery proximal to the
origin of the a. peronaea is the product of the union between the
a. tibialis posterior superficialis and the a. peronaea posterior
superficialis. The remainder of the vessel is a survival of the
part of the stem of the a. tibialis posterior superficialis proximal
to the point at which it is joined by the medial terminal branch
of the a. peronaea posterior superficialis.
Branches. The a. peronaea is described on page 90. The
a. malleolaris posterior medialis and the r. communicans are
derived from the embryonic r. coronarius malleolaris medialis of
the a. interossea (p. 73).
A. tibialis posterior superficialis. The course of this artery,
which arises shortly before the stage of 14 mm., is described on
page 70.
The whole of the a. tibialis posterior superficialis persists in
the adult life. In combination with the a. peronaea posterior
superficialis, the more proximal part of the artery forms the
part of the a. poplitea in contact with the m. popliteus and the
part of the a. tibialis posterior proximal to the origin of the a.
peronaea.- The more’ distal part of the artery persists as the
portion of the a. tibialis posterior beyond the origin of the a.
peronaea and as the a. plantaria medialis.
The relation of the part of the a. tibialis posterior super-
ficialis which forms the a. tibialis posterior of the adult to the
n. tibialis is discussed on page 84.
Rete dorsale. The dorsal rete of the embryonic arises from
the r. perforans tarsi. It forms all the arteries which are
distributed upon the dorsum of the adult foot.
94 H. D. SENIOR
Rete plantare. This embryonic plexus arises from the terminal
branches of the a. interossea. It furnishes the arcus plantaris
and all the other arteries of the adult sole excepting the aa.
plantares.
BIBLIOGRAPHY
CaILLARD 1832 Proportions de Méd. et dela Chir. Thése inaug., Paris.
CRUVEILHIER, J. 1843 Traité d’Anotomie descriptive. Paris.
DeVrigEsE, BertHa 1902 Récherches sur I’évolution des vaisseux sanguins
chez ’homme. Archiv. de Biologie, T. 18, p. 665.
DusrveEIL, J. M. 1847 Des anomalies artérielles, Paris.
Exits, G. V. 1853 An account of an instance of remarkable deformity of the
lower limbs. Medico-Chir. Trans., vol. 36, p. 489.
Faace, C. H. 1864 Case of aneurism nenied on an abnormal main artery ie
the lower limb. Guy’s Hosp. Rep., vol. 10, p. 15.
Froriep, L. Fr. V. 1832. Notizen aus dem Gebiete der Natur u. Heilkunde,
B. 34, S. 45.
GREEN, P. H. 1832 Ona new variety of the femoral artery, with observations.
Lancet for 1831, vol. 1, p. 731.
Grosser, O. 1901 Zur Anat. u. Entwichlungsges. des Gefiss-systems der
Chiropteren. Anat. Hefte, B. 17, H. 2, S. 203.
Hocustetrrer, F. 1890 Uber die urspriingliche hauptschlagader der hinterén
Gliedmasse des Menschen, u.s.w. Morph. Jahrb., B. 16, 8. 300.
Hyrrt, J. 1864 Uber normal u. abnorme Verhiltnisse der Schlagadern des
Unterschenkels. Wien.
Lespoucg 1893 Verhandl. d. Anatom. Gesellsch., auf. d. siebenten Versamml.
Anat. Anz., Erginzungsh. zum. B. 8.
Levy, G. 1902 Morphologia delle arteriae iliache, Parte 2. Archiv. Ital. di
Anat. e di Embriol., V. 295.
McMoraricn, J. P. 1904 The development of the human body, 2nd ed., Phila-
delphia.
PorowskI, J. 1893 Uberbleibsel der Arteria saphena beim Menschen. Anat.
Anz., B. 8, S. 580.
1894 Das Arteriensystem der unteren Extremititen bei den Primaten.
Anat. Anz., B. 10, 8S. 55 u. 99.
QuaINn, R. 1844 The anatomy of the arteries of the human body. Lenore
Rue, C. 1863 Anomalie der Arteria cruralis. Wiirzb. mediz. Zeits., B. 4,
S. 344.
Ruce, G. 1894 Varietiiten im Gebiete der Arteria femoralis des Menschen.
Morph. Jahrb., B. 22, 8S. 161.
Satyr 1898 Arteria dorsalis pedis. Pisa.
Senior, H. D. 1917 The development of the external iliac artery in man.
Ann. N. Y. Acad. Sci., vol. 27.
Srrepa, H. 1893 Verhandl d. Anatom. Gesellsch., auf d. siebenten Versamml.
Anat., Anz., Erginzungsh. zum B. 8.
ARTERIES OF HUMAN LOWER EXTREMITY 95
Suressporr, M. 1895 Lehrbuch der vergleichenden Anatomie der Haustiere.
Stiittgart. i
TANDLER, J. 1903 Zur Entwickelungsges. der menschlichen Darmarterien.
Anat. Hefte, B: 23, Hi, S: 187.
VELPEAU 1839 Eléments de Médicine Opératoire.
ZAGORSKI 1809 Mém. de l’Académ. des Sci. de Petersb.
ZUCKERKANDL 1894 Zur Anat. u. Entwickelungsges. der Arterien des Vor-
derarms (1. theil). Anat. Hefte, B. 4, H. 1, 8S. 1.
1895 Zur Anat. u. Entwickelungsges. der Arterien des Unterschenkels
u. des Fusses. Anat. Hefte, B. 5, H. 2, S. 207.
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AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, FEBRUARY 24
THE DEVELOPMENTAL RELATIONS OF BRACHY-
DACTYLY IN THE DOMESTIC FOWL
C. H. DANFORTH
Department of Anatomy, Washington University School of Medicine
FIVE [FIGURES
The factors which exercise determining influences in the
ontogeny of animals have proved difficult of recognition and
evaluation. To what extent any given structure is the direct
expression of some more or less specific ‘determiner’ in the
germ cell and to what extent it represents the product. of
reactions to the influence of other parts of the developing
organism cannot often be estimated in any reliable manner.
Nevertheless, some knowledge of the relative weight of the
two kinds of factors involved is essential to a satisfactory con-
ception of the processes .of embryology and morphology. The
observations recorded in the present paper are reported because
they seem to throw some light on this problem with reference
to a rather special case; namely, the correlation between skeletal
peculiarities of the fourth toe and the presence of feathers on
the tarsi in the common fowl.
The data to be presented were collected as the result of the
following observations: In July, 1915, an egg incubated in the
laboratory hatched out a chick (no. 12) which showed an almost
complete absence of the fourth toe on each foot. This particular
chick was helped from the shell and the condition of its toes
noted at that time, so there is no possibility of the malformation
having been due to injury received after hatching. Subsequent
dissection showed that the three terminal phalanges and the nail
were lacking on both sides (fig. 1, B). During growth the
lateral rudimentary toes turned under the balls of the feet
causing the bird some inconvenience, especially in perching.
97
98 Cc. H. DANFORTH
The specimen, a male, had moderately booted tarsi and was
also strongly polydactyl. It met an accidental death at about
nine months of age before any offspring had been obtained.
The parents of this chick were full brother and sister (no. 8,
#, and no. 7, 2). Both were polydactyl and both had booted
tarsi. On closer inspection it was found that their fourth toes
were somewhat shorter than normal. It was at first thought
possible that the brachydactyly and polydactyly might be cor-
rq
A B
Fig. 1. Bones of the left foot from (A) normal and (B) brachydactyl cockerel.
related, that the excessive toe development on the medial side
of the foot might be more or less at the expense of the tissues
on the lateral side. The following data show that such was not
the case.
From the mating of nos. 7 and 8 only a few chicks were ob-
tained. These included specimens with four and five digits,
bare and feathered tarsi, brachydactyl and normal fourth toes.
Some additional specimens secured by crossing no. 8 with a
barred Plymouth Rock hen gave essentially the same distri-
BRACHYDACTYLY IN THE FOWL 99
bution of characteristics. The association of characters in these
chicks was such as to suggest that the brachydactyly might be
correlated with booting but the numbers were not adequate to
warrant definite conclusions.
Through the courtesy of the officials in charge, it was possible
to make a superficial examination of the feet in a considerable
series of booted fowls exhibited at poultry shows in St. Louis.
Such examinations sufficed to show that brachydactyly occurs in
several races with booted tarsi. One highly valued buff Cochin
fowl was found to present the characteristic in as pronounced a
form as did the specimen which first attracted our attention to
the peculiarity. Owners and judges are inclined to attribute
such cases to accident, but the fact that the standard of require-
ments for some of these breeds specifically calls for well formed
toes suggests that the tendency to brachydactyly has been
vaguely recognized by breeders. No instance was found in
which a short lateral toe occurred on a smooth-shanked bird.
These observations .were sufficient to indicate that brachy-
dactyly is of rather common occurrence in the domestic fowl
and seemed to justify a more careful investigation of the
peculiarity. Consequently a further breeding experiment was
undertaken in order to furnish material for study of the heredity
and embryology of the condition. -
MATERIAL AND METHODS
To obtain this material male no. 8 was mated to a flock of 13
white Leghorn hens. All eggs laid between February 15 and
April 18, 1917, were incubated, except when, as occasionally
happened, the number of eggs remained for several days in
excess of the capacity of the incubator. At such times the
oldest eggs were discarded. It is not probable that this method
could have resulted in any differential selection. A total of 300
eggs were used. Of these, 108 yielded living embryos and 86
were allowed to hatch. Two of the remaining eggs were infected
with bacteria, 34 were non-fertile or developed only slightly,
and 70 contained embryos that died before hatching.
100 Cc. H. DANFORTH
The living embryos were fixed in Zenker’s fluid or in 10 per
cent formalin. They were subsequently examined under a
binocular microscope and parts of many of them were either
sectioned or stained by the van Wijhe method and studied in toto.
The contents of eggs found to contain dead chicks were put
in formalin and after a short period of hardening the embryos
were examined, their age estimated and characteristics recorded.
All the toes on both feet of each of the 86 living chicks were
measured, the grade of polydactyly, the grade of booting and
also the form of the comb were noted. These data likewise
were recorded so far as possible for the older embryos. For
each of the living chicks an index of brachydactyly was obtained
by dividing the sum of the lengths of the two lateral toes (digits
IV) by the sum of the lengths of the two medial toes (digits II).
This gives a value varying above and below 1. The value
obtained is multiplied by 100 in each case to eliminate fractions.
The index is thought to be fairly reliable, since careful measure-
ments have shown that the fourth toe is the one chiefly affected
while the third is only slightly so and the second possibly not at
all. The toe measurements represent the distance, obtained by
the use of dividers, from the base of the nail to the metatarso-
phalangeal articulation.
DESCRIPTION OF STOCK USED FOR BREEDING
The females employed in the breeding experiment were all pure
bred white Leghorns of a well established strain. Their shanks
were free from feathers and there was no indication of poly-
dactyly or brachydactyly. The fourth toe index ranged from
106 to 121 with an average of 112. All were pure white with
large single combs. Eight of these and a male were obtained
in 1916 from a poultryman who devotes himself exclusively to
this breed. The other five were raised at the laboratory from
this original flock. There can be little doubt as to the purity
of the stock especially in regard to the characteristics under
investigation. At the time of the experiment five of the hens
were less than ten months old, the rest were two years or more.
BRACHYDACTYLY IN THE FOWL . 101
The male (no. 8) differed from these hens in a considerable
number of characteristics, for most of which he seems to have
been heterozygous. His color was partly that of a barred
Plymouth Rock, although hackle, saddle feathers, and_ tail
contained much white and the breast was somewhat spotted.
The comb was broad and long with irregular elevations and a
flattened, truncated and slightly trifid posterior prolongation.
It did not seem to be either typically ‘rose’ or ‘walnut’ although
perhaps more closely approaching the latter. The tarsi were
moderately booted (grade 2 in an arbitrary scheme adopted for
purposes of this paper). Digit I on the left foot was partially
doubled (grade 2) while on the right foot it was normal. Digits
IV were short, giving an index of 89. At the time of the
experiment this bird was in the last half of his third year of life-
HEREDITY OF CHARACTERISTICS
The heredity of the five outstanding characteristics, color.
comb form, polydactyly, booting and brachydactyly may be
briefly summarized. The first four of these have been subjected
to careful study by Davenport (’06, ’09) and others.
A. Color. All. chicks that hatched and all embryos that
reached the stage for developing down were white, or white with
occasional small dark spots. White was to have been expected
in the offspring since the white of the Leghorn is a well known
dominant character.
B. Comb. According to the prevailing view as to the unit
factors involved (Bateson, ’09) four types of comb should have
occurred in equal numbers: walnut, rose, pea and single. These
four types did occur, but since in the embryo and chick it is
often difficult to differentiate with certainty between rose and
walnut (of the type met in this experiment), pea and single,
the four categories were grouped into two; broad combs and
narrow combs. Of 172 chicks and late embryos, 88 had broad
combs and 84 narrow, the expectation being 86 of each. This
is very close to the Mendelian ratio.
2 -. Cc. H. DANFORTH
C. Polydactyly. For the sake of convenience three grades of
polydactyly, corresponding essentially to those designated by
Anthony (’99), were employed. Grade 1 includes cases in which
only the distal phalanx is involved; grade 2 includes cases
where two phalanges are involved but with the toes usually
more or less united and equal; grade 3 includes all cases of a
more pronounced form. In grade 3 the metatarsal is also
affected and the accessory toe generally has three phalanges.
Only one instance of six toes occurred. Occasional specimens in
which there were really only four toes but in which the first
digit showed three phalanges instead of two were classed as
polydactyl, grade 1. Such individuals were included because
the evidence seems to the writer to indicate that polydactyly
in the fowl is not so much a matter of added elements as of
readjustments following early hypertrophy of a single anlage.
In possibly a third of the cases the two sides were not strictly
symmetrical. Of the 28 polydactyly chicks that hatched, 8
were of grade 1 and 10 each of grades 2 and 3. The father
‘was of grade 2 (on the left side only).
As shown by Kufmann-W olf (’08), some cases of polyadactyly
can be recognized as early as the fifth day of incubation. Prob-
ably all individuals which would develop info grade-3 speci- .
mens can be recognized as polydactyl at this age. But for
purposes of the present record no individual under seven
days is included. Of 220 chicks and embryos of 7 days and
over, 147 were normal, 73, or 33 per cent, were polydactyl.
If polydactyly behaved as a strictly dominant character the
expectation in this case would be 50 per cent. The failure of
polydactyly in poultry to give typical Mendelian ratios has
been discussed by several authors, especially Davenport (’09).
D. Booting. In chicks that are to be booted, feather (down)
rudiments generally become apparent on the tarsi during the
eleventh day of incubation (fig. 2) or considerably later than on
other parts of the body. After the twelfth day, booting is
easily recognized. Thirty-six living chicks which were booted
were classified into three grades, based upon the amount of down
on the tarsi. Grade 1 represents individuals with from one
BRACHYDACTYLY IN. THE FOWL 103
to several feathers on the shank. Grade 3 stands for speci-
mens with about twelve feathers on the shank, three or four
on the lateral toe, and occasionally one or two on the middle
toe. Grade 2 includes specimens intermediate between 1 and
3. The distribution among the three grades was as follows:
grade 1, 14; grade 2, 10; grade 3, 12. The father was grade 2.
Out of a total of 129 chicks and embryos of thirteen days and
over, 80 were normal and 49 booted,—38 per cent. Booting,
EXIT.
FLEX.
Fig. 2 Sketches of cross-sections of comparable regions of the right shank
from (A) nine-day and (B) eleven-day embryos, both of which were strongly.
brachydactyl. Dr., down rudiment; Fzt., extensor group of tendons; Flez.,
flexor group of tendons; Mz., metatarsal cartilages.
like polydactyly, does not give strictly Mendelian proportions
(Hurst, 705, Davenport, ’09).
E. Brachydactyly. In chicks with normal feet the lateral toe
is about 10 per cent longer than the medial; in brachydactyly
chicks it is of equal length or shorter. While the differences are
only slight and in embryos difficult to measure a certain number
of obvious cases can be recognized in nine-day specimens. From
the tenth day on they are clearly distinguishable, probably in
nearly all cases. It is possible, however, that a small number
104 Cc. H. DANFORTH
of brachydactyl specimens, having a relatively high index, may
be overlooked even among late embryos. Among 90 embryos
of ten days and over, 57 had long fourth toes; 33, short. Of 86
young chicks, 55 had an index of more than 100 while 31 showed
an index of 100 or less (brachydactyl). The totals for embryos
and chicks are: out of 186 individuals, 112 normal, 64 brachy-
dactyl. This is 36.4 per cent, which is intermediate between
the percentages for booting and polydactyly. These character-
istics behave in heredity quite differently from color and comb
form which give results corresponding closely to the Mendelian
expectation.
EMBRYOLOGY OF BRACHYDACTYLY
Since brachydactyly in poultry has not seemed to attract
much attention (Anthony, ’99, states that he has seen specimens
which showed four and six phalanges in their fourth toes), a
fuller account of the condition may be entered upon at this
point. Inasmuch as the shortening of the other toes is relatively
slight and difficult to measure the fourth only will be considered.
The skeleton of a normal foot is shown in figure 1, B. Digit IV
has five phalanges, the terminal one bearing a nail. Among
brachydactyl specimens, the fourth toe varies from a condition
in which nail and all five phalanges are present with a total
length equalling that of the second toe (but not exceeding it as
in normal feet) to a condition showing complete absence of the
nail bearing phalanx and only two remaining phalanges. Inter-
mediate conditions are represented by toes with three or four
well formed phalanges and by toes with the full number of
joints but of considerably reduced length. The brachydactyl
toes also varied from the normal in other characteristics such
for example as the number of rows of scales. The two sides of
the same individual were found to be very closely correlated.
Among the 31 brachydactyl chicks that hatched, the index varies
from 70 to 100, with an average of 89, which is identical with the
index of the father. All of the specimens had the nail reduced
and seven of them had only four bones in the toe as shown
either by dissection or surface configuration (fig. 3).
BRACHYDACTYLY IN THE FOWL 105
Tracing the condition back in the development of the chick,
we find, as already stated, that brachydactyl can be recognized,
at least in many cases, as early as. the ninth day, when the
fourth toe in certain individuals can be seen to be conspicuously
shortened. It has not been possible to determine whether or
not the shortening is actually present at an earlier date. The
cartilage of the third phalanx of the fourth toe is laid down
during the 8th day. The fourth and sometimes the fifth car-
tilage are formed on the 9th day. By the end of the 10th day
all of the phalanges to develop are present in cartilages. Definite
reduction in the number of cartilages cannot be recognized with
certainty earlier than the tenth day.
e @
Fig. 3 Foot-prints of (A) brachydactyl and (B) normal left feet. In A the
absence of one phalanx in the fourth toe is revealed.
The fact that brachydactyly is apparent at the time when
cartilages are formed, if not actually before then, would seem to
indicate that the reduction of the skeleton takes place in re-
sponse to conditions already manifest in the toe, and not as a
result of any factor acting specifically on cartilage or bone forma-
tion. In other words the cartilage anlagen form while subjected to
a kind of compression resulting from the shortening of the toe.
The consequent reduction in the cartilages, and the correlated
changes in ligaments, tendons, etc., would therefore seem to be
106 Cc. H. DANFORTH
due not to any inherent peculiarity of the structures themselves,
but rather to the conditions under which they develop. The
shortening of the soft parts, seemingly secondary is more
probably the primary factor in causing brachydactyly. In the
adult foot, however, the bony framework provides an accurate
index of toe length.
As already indicated, the reduction in the skeletal elements
is both qualitative and quantitative. It may be that this
reduction does not take place in the same way in all short-toed
specimens, but a series of grades may be described which
seem to represent successive steps in the process. 7
The first degree of shortening represented by the least pro-
nounced type is characteristic of the majority of brachydactyl
specimens. Here all of the phalanges are present, but the
fourth, or sometimes the third, is more or less shortened. The
amount of shortening in these phalanges varies greatly and can
often be estimated in the living chick. It has not been possible
to determine accurately whether or not the other phalanges are
also shortened, although it is probable that they are, especially —
- the fifth since the nail is almost always abnormally small. The
fact that the fourth, or occasionally the third, phalanx is the
one first affected suggest that the reducing factor is most effective
at the time these phalanges are being laid down, viz: on the
ninth, or occasionally the eighth day.
The second degree, found in slightly over a fifth of the cases
studied, shows the first numerical reduction in the phalanges.
Here the third and fourth segments are replaced by a single
element. The resulting cartilage is commonly a slender rod
slightly longer than a normal fourth which it otherwise re-
sembles. This cartilage also varies in length so that there is
some fluctuation within this grade. It has not been possible to’
determine with certainty whether the coalescence of the two
elements is due to one cartilage forming from the substance
that normally gives rise to two, or whether both elements form
separately and then fuse. The specimen shown in figure 4 seems to
favor the latter alternative, but where the adult organ presents
an unlimited series of gradations it is impossible to say in the
BRACHYDACTYLY IN THE FOWL 107
case of any individual embryo whether the observed condition
is transient or definitive.
The third degree is represented by one twenty day embryo
and possibly one or two younger embryos. In this case there
seems to be a general reduction in which the terminal phalanx
suffers most. The third and fourth phalanges are fused. The
claw is represented only by a small flat scale and the fifth bone
Fig. 4 Feet of a brachydactyl embryo on the tenth day of incubation. In
the fourth toe of the right foot, at B, is shown the completely fused third and
fourth phalanges. In the left foot, at A, these two phalanges are seen to be
only partially fused.
by a little nodule into which the terminal slip of the flexor
profundus tendon is inserted. Since the flexor profundus is
also inserted into the plantar surface of the normal fourth
phalanx or the fused third and fourth phalanges, a stage showing
a complete absence of the fifth segment would differ very slightly
from the case here described. It is interesting to note in this
connection, however, that the basal and terminal phalanges are
108 Cc. H. DANFORTH
the most stable, the intermediate elements being the ones first
affected.
The fourth degree was found in no. 12 (half brother of these
chicks), referred to at the beginning of the paper, and in several
chicks obtained later. Here there are only two bones present
in the fourth toe (fig. 1, B). There is no nail. The tendon of
the flexor profundus is inserted on the plantar surface of the
terminal bone and the superficial flexér into the lateral aspects
of both segments. In view of this arrangement, the conforma-
tion of the two bones, and the condition observed in other
specimens it would seem probable that the proximal element
represent the normal basal phalanx while the distal one repre-
sents a fusion (in effect, if not in reality) of phalanges II, III
and IV; phalanx V being entirely absent.
Owing to the brief period between the first indication of
clearly demonstrable brachydactyly and the appearance of
phalangeal cartilages, the possibility that the precartilage cells
are the ones first affected by the shortening factor can not be
entirely excluded. If such be the case, it is the skeletal elements
that from the earliest period determine the form and proportions
of the toe; but as already indicated, the evidence seems to point
quite as strongly to the alternative possibility, namely, that the
form of the skeletal elements as first laid down is the result of a
more or less passive response of cartilage forming cells to the
influence of surrounding conditions. The relation of brachy-
dactyly to booting (to be discussed in a later paragraph) adds
considerable support to the latter interpretation. If this inter-
pretation is correct, it follows that the early shortening of the
toe as a whole results in a reduction of the skeletal elements,
the degree of reduction being directly correlated with the degree
of shortening, and not improbably passing successively through
the above described stage till the definitive condition is reached
on the tenth day or shortly thereafter.
INTERRELATION OF CHARACTERISTICS
The relations of brachydactyly and the other hereditary
characteristics observed in this experiment are indicated in the
BRACHYDACTYLY IN THE FOWL 109
TABLE 1
A summary of the data arranged to show the relation between brachydactyly
polydactyly, booting and comb form in the eighty-six
living chicks
A. BRACHYDACTYLY AND D. BRACHYDACTYLY AND
COMB FORM POLYDACTYLY
Broad comb Narrow comb Five toes Four toes
Obs. Exp. Obs. Exp. Obs. Exp. Obs. Exp.
Shontmounth toe... 2-54. 15 15e5 16 15e5 10 10 21 21
Monestourth tev... +... .4.4)) 28 DAL ® Dal 1 ies) 18 17.9 37 347/01
B. POLYDACTYLY AND E. POLYDACTYLY AND
COMB FORM BOOTING
Broad comb Narrow comb | Booted shank | Smooth shank
Obs. | Exp. | Obs. | Exp. | Obs. | Exp. | Obs. | Exp.
TRINVEATE Ss Oo ete See eee 16 14 12 14 13 pes 15 | 16.2
HOMERGOCS Hearne ao. teas Pal 29 31 29 23 24.2 35 | 33.8
Cc. BOOTING AND COMB F. BOOTING AND BRACHY-
FORM DACTYLISM
Broad comb Narrow comb |Short fourth toe|Long fourth toe
Obs. | Exp. | Obs. | Exp. | Obs. | Exp. | Obs. Exp.
DOoLed Shakes a .2c)225...| AT 18 19 18 dl 12.9 iy | aye
Smooth shank.............| 26 25 24 25 Osieis 50 | 32
accompanying table (table 1, A—F), which is based on the 86
chicks hatched. Since the male used was, in Mendelian terms,
heterozygous for all four of the peculiarities studied, the de-
terminers for each (with the partial exception of comb form)
having been supplied by his paternal gamete alone, and since
the females were homozygous for the absence of all these
characters, an especially favorable opportunity was afforded for
testing linkage relations. If each character behaved in a
strictly Mendelian fashion and segregated independently, the
expected value for each combination would be 21.5, and the
sum of each pair added either vertically or horizontally would
be 43. While, as already indicated comb form agrees exactly
with the Mendelian expectation (43: 43 for those that hatched)
110 Cc. H. DANFORTH
the other three characters do not, so that linking can only be
tested by comparing the proportional distribution of one pair of
contrasted characters with reference to other pairs. The table
shows the observed values for all possible combinations and the
expected values for each of these based on the behavior of the
several characters considered separately.
It will at once be apparent that the observed and expected
values agree very closely in the first five sets of combinations.
The slight apparent deviations in favor of a correlation between
polydactyly and broad comb and between polydactyly and
booting are too small to be of signficance. So far as this part
of the data goes it is in full accord with Davenport’s (06) con-
clusion that correlation of characteristics in poultry is very rare.
But, on the other hand, between brachydactyly and booting
there is found to be a close correlation. Here the values are
clearly significant. Of thirty-one brachydactyl chicks, all had
booted tarsi; and of thirty-six chicks with booted tarsi, thirty-
one were brachydactyl. The distribution is shown graphically in
figure 5 where it also appears that the length of the fourth toe is
rather more variable in specimens with booted shanks than in
those with smooth shanks.
There is, then, a distinct relation, which becomes apparent in
embryonic stages, between brachydactyly and booting, but
there is no evidence of a significant connection between either
of these characters and comb form, color or polydactyly.
SIGNIFICANCE OF THE CORRELATION BETWEEN BRACHY-
DACTYLY AND BOOTING
The explanation for the relation between short toes and
feathered tarsi is not obvious. Several possibilities suggest
themselves.
In the first place, it might be that we have here a case of true
linkage, such, for example, as Morgan (715) and others have
successfully demonstrated in fruit flies. Morgan believes that
in such cases the determiners for both of the linked characters
are located in the same chromosome. If this were the true
explanation, ‘crossing over’ might possibly be expected to occur
BRACHYDACTYLY IN THE FOWL 112
in a certain percentage of cases, the frequency depending on the
distances of the two determiners from each other. There “is
some evidence that could be regarded as favoring this inter-
pretation. Five chicks with more or less booted tarsi (grades 1,
1, 1, 1, 2) were not brachydactyl. If these represented true
cases of ‘crossing over’ there should have been an equal number
of brachydactyl chicks. which were not booted. One specimen
was obtained. To fully substantiate this hypothesis it would
Fig. 5 Curve showing the relation between brachydactyly and booting in
86 living chicks. Each square represents an individual, the shaded ones being
booted. The indices of brachydactyly are indicated below.
be necessary by further breeding experiments to derive from the
same hybrid stock one strain with feathered feet and long toes
and another strain with unfeathered feet and short toes.
In this connection it was thought desirable to investigate
the relation of these characters in other species. The writer is
greatly indebted to Mr. Louis Agassiz Fuertes for carefully
examining for him a large number of booted grouse, ptarmigans,
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, NO. 2
ae Cc. H. DANFORTH
and owls. Mr. Fuertes was unable to find any indication
whatever of brachydactyly in any of these birds.
In the case of the domestic pigeons on the other hand, the
writer has found somewhat the same condition that occurs in
hens. ‘Swallow’ and ‘pouters’ both of which are booted, often
show short lateral toes. Among ‘muffed tumblers’ also heavily
booted, several specimens were found to have the third and
fourth toes fully webbed, and I have been informed that the
same condition sometimes obtains in ‘pouters.’ Davenport (’09,
pp. 29 to 36) describes a similar condition in a strain of fowl.
It is not clear whether all of his syndactylous specimens were
also booted but the fact that the first cases arose in breeds that
are normally booted, is probably significant. In another flock
of fowl, related to those here described and having so far as
known no syndactyly in the ancestry, a pair of brachydactyl
birds produced two syndactylous chicks of the type described by
Davenport. These cases suggest that syndactyly like brachy-
dactyly may represent an arrest of development, occurring in
the former case a little earlier than in the latter, and that the
two conditions may be dependent upon a single factor.
While the suggestion of linked determiners cannot be entirely
ignored as a possible explanation of the results obtained,
there is still further evidence against such an interpretation.
This rests chiefly on the fact that there is a correlation between
the degree of booting and the index of brachydactyly. Despite
wide fluctuations, this fact is brought out by averaging the
indices for the different grades. After excluding the two ex-
treme cases with indices of 70 and 120 the average index for
each group is as follows, grade 1, 93; grade 2, 91; grade 3, 89.
If the two extremes are included, they tend to raise the value
of grade 1 and lower the value of grade 3, thus accentuating
the differences. Such a result is not easily explained on the
assumption of two independent determiners but, on the con-
trary, strongly suggest that the association is due to some-
thing more than a chance proximity of the determiners in a
chromosome.
BRACHYDACTYLY IN THE FOWL fale
If this is the case the question presents itself as to whether the
relation is causal, one condition producing the other or whether
the two conditions are both dependent on some one determining
factor. It was at first thought probable that the abnormal
development of feathers or down on the feet overtaxes the
nutritional supply of the limb with the result that the fourth
toe becomes somewhat stunted in its development. The em-
bryology of the condition, however, does not support this view.
On the contrary, in light of the sequence in which the character-
istics make their appearance in ontogeny, brachydactyly would
seem to be the cause of feathered tarsi, an interpretation which,
a priori, does not seem plausible.
Since, on the whole, the evidence does not favor the idea
that either condition is really the cuase of the other the only
alternative that would seem to be left is that they are both
caused by the same factor. The close association of the two
characteristics and the fact that they tend to fluctuate together,
points strongly toward this conclusion. What the factor may
be or precisely how it acts cannot at present be answered. Nor
can the number of characteristics which it affects be stated
since it is quite possible that some of them have been overlooked
in the present study.
If the connection between brachydactyly and booting had
been a little less obvious the former might have been described
as an independent unit character and a separate determiner in
the germ plasm postulated for it, as has already been done for
booting by the several authors who have studied that character.
In general the relations here described would seem to be such
as tend to support the contention of Morgan (loc. cit.) that the
determiners in the germ plasm are not strictly specific, but tend
to influence the character of the organism as a whole.
In the particular case under consideration it is quite possible
that the determining factor may actually be effective for only
a short time, possibly a few days. After two cartilages have
once fused it is doubtful if the removal of the exciting cause
would in itself induce them to separate. It is also possible that
a feather germ once formed would need no further stimulus than
114 Cc. H. DANFORTH
proper nutritional conditions to continue its development to the
end. If these suppositions are correct it is quite conceivable
that the whole complex is due to a slight irregularity (delay or
acceleration) in the beginning functioning of some one or other
of the endocrine glands. The character of the gland would of
course be determined by other factors which would have an
ultimate basis in the nature of the germ plasm itself. An ex-
planation of this sort, both for these characters and for poly-
dactyly would much more readily account for the aberrant
nature of the ratios obtained in breeding experiments.
SUMMARY
The observation reported in this paper establish the fact
that, at least in certain strains of poultry (and probably in
pigeons) there is a close correlation between brachydactyly
involving the size and number of bones in the feet, and the
presence of feathers on the tarsi. The embryology of both
conditions has been studied, the evidence from this source
indicating that while the size and number of skeletal elements
are determined by the length of the embryonic toe, there is no
causal relation between toe length and feathering. The data
cited seem to indicate, on the contrary, that brachydactyly,
feathering of the tarsi, and probably syndactyly are all de-
pendent on one and the same factor. No attempt is made to
postulate the nature of this factor but it is suggested that a
study of the early functioning of the endocrine glands in normal
and abnormal embryos might throw some light on the question.
No. correlation could be detected between either of these
characters and polydactyly or comb form.
BRACHYDACTYLY IN THE FOWL its
LITERATURE CITED
Antuony, R. 1899 Etude sur la Polydactylie chez les Gallinacés. Jour. de
l’Ant. et de la Physol., T. 35, pp. 711-750.
Bateson, W. 1909 Mendel’s principles of heredity. Cambridge University
Press.
Bateson, W., AND PunNETT, R.C. 1905 Poultry. Jn Report to the Evolution
Committee of the Royal Society. Report II, pp. 99-131.
Bepparp, Frank E. 1898 The structure and classification of birds. Long-
mans, Green & Co., London, New York, and Bombay.
Davenrort, C. B. 1906. Inheritance in poultry. Publications of the Carnegie
Institution of Washington, no. 52.
1909 Inheritance of characteristics in domestic fowl. Carnegie In-
stitution of Washington, Publication no. 121.
Horst, C. C. 1905 Experiments with poultry. In Report to the Evolution
Committee of the Royal Society. Report II, pp. 131-154.
KAvuFMANN-Wo Lr, Marie 1908 Embryologische und anatomische Beitrige zur
Hyperdactylie (Houdanhuhn). Morph. Jahrb., Bd. 38, 8S. 471-531.
Lituin, Frank R. 1908 The deyelopment of the chick. Henry Holt & Co.,
New York. ;
Moraan, T. H., Sturtevant, A. H., er AL 1915 The mechanism of Mendelian
heredity. Henry Holt & Co., New York.
Resumido por la autora, Della Drips.
Estudios sobre el ovario del esperméfilo, con especial menci6én del
cuerpo amarillo.
En el presente trabajo se considera el ciclo de cambios que
ocurren anualmente en los ovarios del espermO6filo, dando detal-
ladas descripciones histol6gicas del cuerpo amarillo en cada uno
de los estados de su desarrollo. Se han empleado coloraciones
especificas para poner de manifiesto los caracteres nucleares y
protopldsmicos de las células amarillas. En el ciclo vital del cuerpo
amarillo se reconocen tres fases: Primera, una fase que se carac-
teriza por la presencia de un gran nimero de granulos rojos, in-
dudablemente grénulos de secreccién, en el protoplasma de las
células amarillas. Esta fase comprende, practicamente, todo el
periodo de la prefiez. Segunda, la fase lipoide, asi, llamada por
la abundancia de gotitas de substancia lipoide en el protoplasma
de dichas células. Esta fase comienza algin tiempo antes del
parto y dura pr6éximamente unas seis semanas después de este,
que es también pr6éximamente el tiempo que requiere para com-
pletarse la involucién normal del utero. Tercera, la fase de re-
eresion. Se dan a conocer también ciertos estudios experimen-
‘tales, tales como los efectos de la ovariotomia sencilla y doble,
practicada sobre animales prefiados y no prefiados. La ovario-
tomia sencilla produce resultados negativos. La ovariotomia
doble en animales no prefiados causa una atrofia funcional del
utero, que se manifiesta muy gradualmente. En las hembras
prefiadas esta operacién da lugar a abortos, excepto cuando se
practica cuando la prefiez est’ muy avanzada. Como resultado
de los estudios histol6gicos y experimentales, la autora llega a la
conclusién de que los cuerpos amarillos producen dos secrecciones
internas que presiden sobre los cambios que tienen lugar en el
utero a consecuencia de la prefiez. La primera secreccién pro-
duce la implantaci6n normal y desarrollo del embrién, y la se-
creccién lipoide ulterior ayuda a la involucién normal del utero.
Translation by Dr. José F. Nonidez
Columbia University
AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, FEBRUARY 24
STUDIES ON THE OVARY OF THE SPERMOPHILE
(SPERMOPHILUS CITELLUS TRIDECEMLINEATUS)
WITH SPECIAL REFERENCE TO THE CORPUS
LUTEUM!
DELLA DRIPS
Mayo Foundation, Rochester, Minnesota
Division of Experimental Surgery
TWENTY-NINE FIGURES
CONTENTS
LLNS IRS s RSS os SEG AN Sy les ao ke. al gala ce oe lane te te ge eae ae Se Pe 117
The ovarian cycle. A. Material and methods. B. Histologic descriptions
of ovaries at consecutive intervals of the annual cycle. 1. The ovary of
the early fall. 2. The ovary of the late fall. 3. The ovary of the early
spring. 4. The ovary of early pregnancy with corpora lutea of the red
granule stage. 5. The ovary of the middle of pregnancy with its char-
acteristic corpora lutea. 6. The ovary at parturition with its char-
acteristic corpora lutea. 7. The ovary of July with corpora lutea of
the lipoid stage. 8. The ovary of the late summer with corpora lutea
TEE, RETEST 011 | LN pala eG + Coe 2 ie oe a SOY a Oe eer eS 126
Experimental studies. A. Effects of the removal of the uterus on the
ovaries. 1. In non-pregnant spermophiles. 2. In pregnant sper-
mophiles. B. Effects of the removal of both ovaries on the uterus. 1,
In non-pregnant spermophiles. 2. In pregnant spermophiles. C,
Effects of the removal of one ovary. 1. In non-pregnant spermophiles.
2. In pregnant spermophiles. D. Effects on the remaining ovary of the
removal of one ovary and the uterus in pregnant spermophiles. E.
Studies on the normal bursting of follicles and the formation of corpora.
lutea. F. Studies on ovulation, fertilization and corpora lutea forma-
Summary and Discussion. A. Summary of the results obtained from the
histological and experimental studies. B. Conclusions as to the func-
tions of the corpora lutea in the ovaries of spermophiles............... 160
LITERATURE
The first papers written on the corpus luteum of the ovary had
to do largely with its formation, particularly with the origin of
the luteal cells. Were these cells connective-tissue elements
1 Thesis for degree of Master of Science, University of Minnesota (Mayo
Foundation, 1917).
1ly/
118 DELLA DRIPS
from the internal theca of the follicle according to the hypoth-
’ esis of von Baer, or were they epithelial in nature and derived
from the membrana granulosa of the follicle as advocated by
Bischoff? Marshall, in his book, ‘‘The Physiology of Repro-
duction” (’10), carefully reviews this early literature.
It was not until Sobotta (96) published the first of his series
of papers on the corpus luteum that the discussion over the origin
of the luteal cells began to come to an end. Sobotta’s study on
the corpus luteum of the mouse was the first systematic record
of the transformation of the follicle into a corpus luteum and
the latter’s subsequent development. He describes, first, the
follicle about to burst; then, one just after bursting; one, one-half
hour afterward, and others at succeeding short intervals up to
seventy-two hours after bursting. All the descriptions are very
detailed, every change in the structure as it proceeds in its de-
velopment being noted. Sobotta traces the origin of the luteal
cells to the epithelial cells of the granulosa of the follicle. The
internal theca cells, he says, expend themselves utterly in the
formation of connective tissue and blood-vessels. The external
theca remains as it was.
‘Sobotta believed the function of the corpora lutea was to
maintain a constant equal tension in the ovaries, which shows
how little thought he gave to this phase of the problem. He was
all absorbed in the formation of the luteal structure. This paper
of Sobotta’s was followed shortly by a very similar one on the
rabbit, in which he confirmed all of his findings in the mouse.
These papers started a real investigation of the corpus luteum,
and for the following ten or fifteen years there were any number
of papers written regarding it. A few writers continued the
discussion as to the origin of the luteal cells. Some criticised
Sobotta, trying to disprove his statements. Among these was
Clarke (’98), who contributed an account of the formation of the
corpus luteum in thesow. His studies were made on serial sections
of pig’s corpora lutea and follicles in different but not subsequent
stages of development. The sections in each set were treated
alternately to a process of tryptic digestion and a picro-fuchsin
stain. From a study of these sections he concludes that the
THE OVARY OF THE SPERMOPHILE 119
luteal cells are of a connective-tissue origin and that the function
of the corpus luteum is to preserve the ovarian circulation.
Sobotta (99) published his third paper on the corpus luteum
of mammals. Honore (’00) was one of the first to confirm
Sobotta’s findings. He wrote concerning the corpus luteum in
the rabbit. Marshall (01) published a paper on the corpus
luteum in the sheep, coming to practically the same conclusions
as Sobotta. Cohn (’03) further confirmed Sobotta’s work on
the rabbit. Jankowski (’04) published a paper in which he
came to entirely different conclusions. They are as follows:
‘““He (Sobotta) simply lets the internal theca vanish. If one
layer must vanish, it would be the one for whom the conditions
after the follicle bursting are very unfavorable and that is the
case with the epithelium of the follicle. The corpus luteum is
not an epithelial but a connective tissue structure.” Sobotta
(06) published a fourth paper on the formation of the corpus
luteum in the guinea-pig, again confirming his former work.
Such an amount of histologic investigation over the formation
of the corpus luteum could not very well go on without arousing
much interest in regard to the physiologic function of the struc-
ture. The men who upheld the connective-tissue origin held
many curious ideas as to its function, all of which tended toward
making its action more or less mechanical. Several thought the
only function of the luteal structures was to prevent ovulation.
Clarke thought its function was to preserve the ovarian
circulation. —,
On the the other hand, those who accepted Sobotta’s con-
clusions began looking for a much more important function for the
corpus luteum. If the luteal cells were epithelial and each was so
intimately in contact with the blood stream, and the whole organ
had so much the appearance of a gland, why couldn’t it be a
gland of internal secretion? Several histologists began studying
the cells for evidences of a secretory product. Regaud and
Policard in 1901 were the first to publish any results. They
stained sections of ovaries of the dog, that had been fixed in
acetic potassium bichromate, with a copper-hematoxylin method
of Weigert. They described some black secretion droplets in
120 DELLA DRIPS
the luteal cells. In Cohn’s paper previously mentioned, he
describes some granules which he thinks may be the same as
those described by Regaud and Policard. He also gives his
reasons for believing that the fatty osmic-stained droplets,
seen in the luteal cells in greatest abundance when the cells
reach their maximum hypertrophy, are real secretion droplets
and not evidences of fatty degeneration as had been previously
contended.
Along with the attempt to discover by a special histologic
technic, evidences of secretory products in the luteal cells, a
number of men sought to discover, by animal experimentation,
proofs of the glandular action of the corpus luteum. The uterus
and the mammary glands, in the minds of all, were the most
closely associated of all the organs of the body, with the ovaries.
Ovulation had been observed to be closely related to menstrua-
tion in the human and to the heat periods in animals. Complete
double ovariectomy was known to stop. menstruation, cause an
atrophy of the uterus, and, in young women, to bring on meno-
pause symptoms. In the very young castration had prevented
the development of the uterus and mammary glands. Double
ovariectomy in pregnant women, especially in the first months,
was known to be followed by abortion.
Various attempts had been made to overcome the bad effects
of ovariectomy by the transplantation of ovaries and by ovarian
medication. These were reported to have given good results,
at least partially.
All these facts seemed to speak for the production of an
internal secretion in the ovaries which affected the uterus and
mammary glands.
Fraenkel (’06) was the first to attempt to prove by a series of
experimental studies on rabbits and cats that the corpora lutea
in the ovaries were responsible for the various effects produced
on these organs.
He begins his papers as follows: ‘‘The corpus luteum must,
from its structure and development, be a gland of internal
secretion, made to insure the implanting and development of
the fertilized egg in the uterus. The corpus luteum maintains
THE OVARY OF THE SPERMOPHILE a
the state of nutrition of the uterus during the years of sexual
activity.”” He thought it ruled over the phenomena of rut as
well as pregnancy.
About this same time Marshall and Jolly published a series of
experiments very similar to Fraenkel’s. They used dogs and
rabbits, and their conclusions were much like those of Fraenkel.
Daels (08) published a paper in which he gives his several
objections to Fraenkel’s theories and records a series of experi-
ments on guinea-pigs. In his first series of experiments he tried
to determine the influence of bilateral ovariectomy on the
pregnant animal, concluding that this operation in the pregnant
animal always interrupts pregnancy during more than the first
half of its duration. He also tried giving Fraenkel’s lutein
tablets, with no results.. He had better results from a product
of the whole ovary.
Ancel and Bouin (’08-’09) contributed several papers to the
literature on the corpus luteum. They believed with Fraenkel
that the corpus luteum produced rut and the other changes
incident to pregnancy. They performed a series of experiments
on rabbits in which they produced an unfertile coitus either
between a normal female and a male in which a part of the vas
deferens had been resected, and a female in which a part of the
uterus had been resected and a normal male. They wished by
these experiments to eliminate any action of the egg and the
placenta on any changes taking place in the uterus incident to
ovulation. They describe structural changes in the uterus and
mammary glands for a period equal to the period of activity
of the yellow body.
In this same year (’09) two other Frenchmen, Regaud and
Dubreuil, published several articles. ‘They were particularly in-
terested in the cause of rut and ovulation. They made asystem-
atic study of a large number of uteri and ovaries in different
phases of the genital cycle and concluded that rut is independent
of the corpora lutea, and that it is improbable that the corpora
lutea plays a réle in originating the pregestative changes in the
uterus, for the graphic curve of their development is much later
chronologically than the curve of its changes. They claim that
122 - DELLA DRIPS
coitus only will bring about ovulation. The congestive phenom-
ena which they notice in the ovaries during rut will not produce
the rupture of a single follicle without coitus.
Niskoubina, the same year (’09), published a series of studies
which confirmed the observations of Ancel and Bouin. He first
made a histologic study of a series of ovaries removed at varying
intervals after coitus and then did some experiments similar to
Fraenkel’s to determine the period during which the ovaries
seem to exert an influence on the pregnant uterus. From his
experimental studies, he concludes: ‘‘The corpus luteum exer-
cises an obvious action on the physiology of pregancy. It puts
the uterus in a condition necessary to assume the development
of the fertilized egg. This action lasts during the first half of
pregnancy, after which it ceases to act.”
Loeb (’08) published a paper in which he states that deciduo-
mata can be produced experimentally in the uterine mucosa of
guinea-pigs by making a number of transverse and longitudinal
cuts so as to break the continuity of the tissue. He states that
this can happen only during a certain definite period after
copulation, between the fourth and the eighth days. This is
the time when freshly formed corpora lutea are present in the
ovaries. These changes were not excited by the presence of
ova, since they took place when the uterus was ligated and the
passage of ova prevented. If the ovaries were removed, de-
ciduomata could not be produced. He concluded, then, that
the ovaries at certain periods after ovulation elaborate a pre-
disposing substance, in the presence of which indifferent stimuli
may produce deciduomata.
Parhon, Dumitresco and Nissipesco (’09) published a paper
on the lipoids of the ovary. From various staining reactions
on sections of ovaries and from chemical reactions of a powder
made from ovaries, they conclude that in the interstitial cells
of the ovary and in those of the yellow body fats are found which
differ in many characteristics from the fat of the adipose tissue.
Mulon (’10) reported that the fat in the corpus luteum, which
stains only faintly with osmic acid, was similar to other fats
found in the adrenal and other organs of the body which form a
THE OVARY OF THE SPERMOPHILE 123
class of fats different from the ordinary body fat. He thought
these so-called glandular fats had to do with the neutralization
of the glandular excretions or the ordinary poisons formed in
the cellular activity. The specific action of the fat of the corpus
luteum was to neutralize the poisonous products formed by the
developing embryo.
Miller (’10, ’14) published some studies on human corpora
lutea. He tried out many fat stains on fresh corpora lutea and
claimed they contain no fat. He says the negative result of
the fat reaction on fresh corpora lutea makes it possible to tell
the difference easily between these and other ovarian structures.
When the involution of the yellow body begins, the neutral fat
reaction begins. The peripheral parts show the fat reaction
first. In the corpus luteum of pregnancy the reaction to neutral
fat remains negative to the end of pregnancy. He says that in
a corpus luteum of five days the fat reaction was negative, in
one of six to eight days there was a little, in one of eleven days
more, and in one of twelve to sixteen days the cells were rich in
fat (not neutral fat).
Meyer (’11, 713) wrote on the human corpus luteum. In his
first paper he described the development and regression of the
human corpus luteum of pregnancy, which confirms Sobotta’s
and Cohn’s work.
Van der Stricht (’12) published the results of his studies on the
corpora lutea and the interstitial cells of the ovary of the bat.
This is one of the most valuable studies ever published on the
corpus luteum, because the ovary of the bat, with its contained
structures, is one of the simplest ever studied. No confusion
arises from old corpora lutea; these are gone before the new ones
are formed, as the periods of ovulation are so far apart, occurring
each spring only. The author describes two secretions in the
luteal cells. The first is a serous secretion which is very like the
liquor of the follicle. This is secreted by the cells from the time
of bursting until about the time the egg enters the uterine horns.
The second is a lipoid secretion. Beginning some time after the
bursting, there is a slow elaboration of fatty granules from the
depth of the cytoplasm of the luteal cells, the amount of which
increases as the cells increase in size.
124 DELLA DRIPS
He says: ‘‘Far be it from our idea of admitting two abso-
lutely distinct phases for serous secretion and lipoid secretion.
On the contrary, at a moment in the development of the corpus
luteum these two processes coexist, but the first is especially
marked at the beginning of gestation and the second exists alone
during the following period.’”’ He concludes that: ‘‘the serous
secretion exercises its influence on the transformations of the
uterine mucosa of the first phase of gestation during the dis-
placement of the egg and that the lipoid secretion intervenes
principally to provoke the arrest and fixation of the blastocyst
and the formation of the placenta.” Van der Stricht has not
been able to demonstrate this serous secretion in the luteal cells,
but because it is present in the central cavity of the young corpus
luteum and in the lymphatics when they are first formed, he
thinks the cells must be secreting it. Both the serous secretion
and the lipoid secretion are carried away, he says, by the
lymphatics.
Corner (’16) published a paper on the corpus luteum of preg-
nancy in swine. He claims to have found in the corpora lutea
of pregnancy, beside the cells which are descendants of the
granulosa cells and the cells which are descendants of the theca
cells, two more types of cells which can be found at all stages of
pregnancy.
Livon (’09) published the results of the effect of injecting
luteal extract into guinea-pigs. He writes as follows: “We have
employed an extract of the corpora lutea of the sow and the
cow, a product that I have today called the Product A. In-
jected into the peritoneal cavity of guinea-pigs, we find a toxic
action varying with the rapidity of the absorption and with the
individual. The toxic.dose obtained generally is 20 to 30 centi-
grams per kilogram of animal. The animals die presenting
general tremors,dyspnea, convulsions and uttering weak cries.”’
Champy and Gley (11) are said to have been the first to show
that the corpus luteum from pregnant cows was exceedingly
active, whereas that from non-pregnant animals possessed little
physiologic action.
Hare (712) reported very favorably on the clinical use of
corpus luteum extract.
THE OVARY OF THE SPERMOPHILE 125
Frank and Rosenbloom (’15) published the results of some
experimental work done on rabbits with extracts of the placenta
and the corpus luteum. They claim to have gotten better results
than former investigators because they used a more concentrated
and an alcoholic (fat solvent) solution of the active substance
of the luteal extract. This active substance, they say, “‘is not a
lipoid but is carried along the lipoids.’”’ They state that the
only corpus luteum substance extracted, which was found to be
active, was derived from pregnant animals.
Dannreuther (’14) reported his results obtained clinically from
the use of corpus luteum extract. He calls attention to the
necessity of using the extract of pregnant animals only.
Up to date, the most noteworthy publication concerning the
clinical value of the extract of the corpus luteum is that of
Culbertson (’16), entitled, ‘‘A study of the menopause.” He
regards the climacteric as a’ ‘‘functional disarrangement on the
part of the endocrine glands, the ovarian secretion having
ceased.” His theory concerning the value of luteal extract in
the treatment of menopause disorders in as follows:
“Thus the chief characteristic stamping the vasomotor dis-
turbances of the climacteric seems to be a disarrangement of the
systolic-diastolic relation producing elevation in the pulse
pressure. In blood-pressure estimations, then, we find a fairly
reliable measure of the vasomotor disturbances of the menopause,
as will be shown, a satisfactory method of treatment.
“Tf we accept the propositions thus far laid down, that the
cessation of ovarian activity leads to a functional over-efficiency
on the part of the pituitary and adrenal glands and that this, in
turn, produces an arterial hypertension, the corollary is that by
the administration of corpus luteum extract, the pressor sub-
stances will be neutralized and the tension will decrease.”’
In summing up, it may be stated that up to 1906 practically
all the literature on corpus luteum had to do with the histologic
origin and consequent structure at various succeeding periods in
its life cycle. The writings of Sobotta and Cohn practically
established the epithelial’ nature of the luteal cells and the
glandular character of the structure as a whole. From 1906 to
126 DELLA DRIPS
1912 most of the papers written were reports of experimental
studies undertaken to prove that the corpus luteum is the gland
of internal secretion in the ovaries, and that, through this
secretion, the luteal structure produces specific effects on other
organs, particularly the uterus. In spite of the many criticisms .
directed against his work, Fraenkel still stands preéminent
among the experimental workers who established beyond a doubt
the foregoing hypothesis.
Since 1910, efforts have been put forth by investigators actually
to demonstrate this secretion in the corpus luteum. Van der
Stricht comes nearer the goal than any others. The greater
part of recent literature, however, concerns the extract of the
corpus luteum, its chemical constituency, its physiologic action,
and its clinical value. .
THE OVARIAN CYCLE
In the summer of 1914, while studying microscopic sections of
the various tissues and organs of the spermophile, the relatively
immense size of the ovaries, compared with those observed the
previous spring, strikingly presented itself. On further compari-
son, it was very evident that this great increase in size had been
brought about by a growth in the corpora lutea only. One
ovary contained eleven of these bodies; there remained only
a framework of ovarian stroma with a few atretic follicles.
With the particular: stain the luteal cells bore a marked re-
semblance to the cells of the cortex of the adrenal. The former
- were much larger, but the shape of both, their arrangement in
columns, the position and appearance of the nuclei, and the
presence of lipoid droplets in the protoplasm accounted for the
likenesses. In fact, the luteal cells resemble secreting cells.
From observations that had been going on, it was known that
these spermophiles had given birth to young about a month
before. According to most writers, degeneration of the luteal
cells begins not later than birth. Here were what looked like
actively secreting cells a month after birth. ‘Thereupon it was
decided to try out some differential stains on these luteal cells
at every stage in their life history, and to study the complete
THE OVARY OF THE SPERMOPHILE Wea
ovarian cycle in the spermophile with a view to gaining some
accurate information of the origin, development, life history,
and function of the corpora lutea of the ovary.
In the spring of i915, numbers of spermophiles were captured.
But not having realized how very soon the animals become im-
pregnated after coming out of hibernation, no strenuous efforts
were made to obtain them until they were quite numerous in
the fields, and consequently they were found to be either in
advanced stages of pregnancy or lactating. However, all the
animals that could be gotten were used for a study of the ovarian
cycle through the summer and fall, until hibernation began.
Several animals were sacrificed each week. They were killed
quickly with ether and bleeding. The ovaries were immediately
placed in one of several fixatives, 10 per cent formalin, Zenker’s
fluid with acetic acid, Bensley’s formalin Zenker, and Bensley’s
acetic acid bichromate. Many stains were experimented with.
After considerable study of the fixed and stained sections, it was
decided that for the problem in hand, two fixatives seemed
best, Bensley’s formalin Zenker and Bensley’s acetic osmic
bichromate—the former particularly for the nuclear structures
and the latter for the elements in the protoplasm. In all the
work of the past spring (’16), one ovary of each animal sacrificed
was routinely placed in formalin Zenker and the other in acetic
osmic bichromate.
Of the sections fixed in zenker, the best results were obtained
with a modified Weigert stain (copper-chrome-hematoxylin),
Ehrlich’s hematoxylin and eosin, Mallory’s connective-tissue
stain, and Bensley’s acid fuchsin and methyl green. A few
sections of each series were prepared with these stains.
Of the sections fixed in acetic osmic bichromate, one of each
series was stained with the Weigart stain and several with the
acid fuchsin and methyl ‘green of Bensley. Complete paraffin
serial sections were made of all the ovaries studied.
The spermophiles went into hibernation about the middle of
October although many of them became partially torpid earlier
than this. The next spring it was determined to get them early
enough. The frost was not out of the ground until about
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, Nu. 2
128 DELLA DRIPS
April 15. As a result of strenuous efforts, the females were
obtained while in rut and every day through the period of
pregnancy, which was found to be twenty-eight days. Two or
more were sacrificed each day.
The ovarian cycle was now completed. Ovaries were at hand
for every week of the year except during the hibernation period
when only a sufficient number of animals was sacrificed to make
sure there were no changes taking place in these organs. For
the period of pregnancy, a time which is especially related to
ovarian activity, there were ovaries for even fractions of a day.
The period of rut evidently follows immediately on the awaken-
ing of the spermophiles in the spring. Ovulation follows on
coitus. Most of the females are impregnated in a very short
time. The period of pregnancy follows. From the time of
ovulation until about September 1 the ovaries contain corpora
lutea. These approach their greatest size about July 1. From
September 1 to 15 the large corpora lutea have disappeared.
With a disappearance of the corpora lutea, there is a very notice-
able rapid growth of the follicles together with a noticeable
decrease in size. The, ovaries which in July consisted almost
entirely of large corpora lutea with a small amount of ovarian
stroma containing a few atretic follicles, by September 15 con-
tained no corpora lutea, but instead, many medium and good-sized
growing follicles containing very little liquor folliculi, but filled
with mitotic figures.
In this paper descriptions will be presented, gross and micro-
scopic, of the ovaries which, as the year goes around, show the
characteristic changes of their cycle.
A typical ovary of the early fall will be presented first; second,
one of the late fall just before hibernation, with its inactivity,
and, third, one of the early spring, showing the characteristics
of the rutting season. For the fourth period, the period of
pregnancy, several ovaries will be described, marking the suc-
cessive changes occurring in the corpora lutea principally.
Finally, will be presented descriptions of a number of ovaries
of the summer months, which show the final stages in the life
history of the corpora lutea and the corresponding notable
differences in the rest of the ovary.
THE OVARY OF THE SPERMOPHILE 129
Experiment 379-15 (spermophile 200). Captured during the spring
of 1915. The ovaries were removed surgically September 14, 1915.
Weight, 150 grams.
Microscopic observations of an ovary. Fixative, formalin zenker.
Stain, hematoxylin and eosin (fig. 10). There are nine good-sized
follicles, the larger of which measure about 0.4 x 0.4 mm. There are
twice as many follicles half as large. There is a goodly number of
small hyalinized follicles. Around the periphery of the ovary are
numbers of primordial ova. The larger follicles are approaching
maturity, but are still growing. They contain many mitotic figures
and very little liquor folliculi. They are located through the cortex
of the ovary, only three being near the surface. None of these larger
follicles appear to be atretic. Through the medullary portion of the
ovary are conspicuous clumps of interstitial cells.
The spermophiles begin to become torpid about September 15, but
they are active by spells for some time after this, depending on weather
conditions. In the laboratory, some are active until November 15.
Experiment 503-15 (spermophile 254). Captured during the spring
of 1915. Sacrificed November 15, 1915. The animal had been hiber-
nating six days.
Microscopic observations of an ovary. Fixative, formalin zenker.
Stain, hematoxylin and eosin. This ovary appears very similar to
that of spermophile 200. The larger follicles are about the same in
number, size, and location. There are about the same number of
smaller and hyalinized follicles as well as primordial ova. The size
of the ovaries as a whole, however, has decreased. This is probably
due to a marked decrease in the size of the blood-vessels and sinuses.
The clumps of interstitial cells are much less conspicuous. The
ovaries appear to have prepared themselves for their long period of
functional inactivity.
Experiment 246-16 (spermophile 291). Captured April 25, 1916.
Was injured in being caught, so was sacrificed immediately. Weight,
170 grams.
Gross observations of the uterus. The animal is in rut. The
rutting season evidently lasts about two weeks. During this time
practically all the females become impregnated. It may be stated
here that the laboratory animals come out of hibernation much earlier.
Those killed from the 1st to the 15th of March appeared to be in rut
and one killed April 11 was found to be pregnant, showing that weather
conditions set the time of the commencement of their sexual activities.
The large size of the uterus is immediately noted (fig. 7). There has
been an increase in length and breadth. It is twice as large as the
inactive uterus. The cervix shows the greatest increase in size and it
is filled with a thick mucoid substance. The walls of the vagina are
swollen. It is very evident that the great increase in the size of the
whole organ is due to a marked increase in the fluid content, which
gives it a pale appearance. (For measurements see table.)
130 DELLA DRIPS
Gross observations of the ovaries. The ovaries of rut show several
changes from those of the fall. They are larger and seem to be slightly
congested. On the surface of each ovary may be seen several slightly
raised, tiny, colorless, cyst-like bodies which are the mature follicles.
Microscopic observations of the ovaries. Fixative, formalin zenker.
Stain, hematoxylin and eosin. Serial sections show ten larger follicles,
all of which lie at the surface of the ovary and several of which are
projecting slightly from the surface. One of these follicles measures
0.5 mm. x 0.4 mm., and another 0.5 mm. x 0.6 mm. None of them
shows atretic changes and all are apparently mature. There is con-
siderable liquor folliculi, and few mitotic figures among the granulosa
cells (fig. 12).
The internal and external theca of these large follicles are distinct
and comparatively thick layers. There is a well-marked membrane
between the stratum granulosum and the internal theca. Acetic osmic
bichromate sections stained with acid fuchsin and methyl green show
a red secretion along this membrane and between the cells of the gran-
ulosum near it. The cells of the granulosum seem to have an increased
amount of protoplasm which makes them larger.
Of the smaller follicles, a few are in good condition, but the greater
number have become hyalinized. The hyalinization of so many of
the smaller follicles gives a characteristic appearance to the ovaries of
the spring (fig. 11). Indeed, it would seem that a few mature follicles
have been produced at the expense of many. The primordial ova are
few in number. ‘The interstitial cells are not at all conspicuous.
Toward the end of the rutting season many uteri present a different
picture. They are smaller and instead of appearing edematous appear
congested.
Experiment 256-16 (spermophile 298). Date of capture April 27,
1916. Sacrificed April 28, 1916. Weight, 105 grams. (For measure-
ments of uterus see table.)
Gross observations of the uterus. The blood-vessels to the uterus
are all much congested and the organ itself shows some congestion
throughout. There is one especially congested area in each horn
about 1 em. from their point of union. There is another specially con-
gested area in the body near the point of union of the horns.
As there are no corpora lutea in the ovaries of this spermophile, the
uterus is still one of rut, not pregnancy. What brings about this
change in the uterus is not evident. The congested condition is, how-
ever, without doubt preparatory to the reception of the fertilized ova.
This brings us in the life cycle to the ovaries of pregnancy. It
has been shown that the ovaries of the fall, winter, and early
spring contain no corpora lutea. Occasionally some remains of
these bodies of the previous year may be found, but this is very
unusual. Thus when coitus takes place during rut and the
THE OVARY OF THE SPERMOPHILE 131
follicles burst and become transformed into corpora lutea, these
new bodies are the only corpora lutea in the ovary. They are
all produced simultaneously and they also develop simul-
taneously if they are not abnormal in some way.
The picture of the ovary changes, then, when fertilization
brings on pregnancy. From this time on until the period of the
growing follicles is reached in the late summer, the ovaries con-
tain corpora lutea. These are the predominating structures in
the ovaries of the sprmg and summer. The changes which take
place in these organs during the period have to do with the
corpora lutea principally. The following descriptions of ovaries
will be attempts at picturing them with corpora lutea of various
ages. As this study is very largely concerned with these luteal
structures, they will be described in considerable detail. Prob-
ably the ovaries which contained the very youngest corpora
lutea seen were those in which ovulation took place in the
laboratory, the animals being sacrificed very shortly afterward.
Experiment 268-13 (spermophile 303). Captured May 3, 1916:
Sacrificed the same day. Weight, 112 grams.
Gross observations. There are no visible signs of pregnancy in the
uterus except congestion. The ovaries contain several slightly raised,
small spherical bodies which resemble mature follicles except that they
are red or pink instead of colorless.
Microscopie observations of the left ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. This ovary con-
tains five luteal bodies, three of which are normal and two of which
are not. Beside these, there are several growing medium-sized fol-
licles, no large ones, quite a number of small atretic follicles and a few
primordial ova. The interstitial cells are not as conspicuous in this
ovary as they were in some of the ovaries of early pregnancy. All the
blood-vessels and sinuses in the medullary portion of the ovary are
very much dilated. Most of the ovary is made up of the five corpora
lutea. Serial sections of the three normal ones show them to be of
different sizes from 0.7 mm. x 0.8 mm. in diameter to 1.3 mm. x
0.1 mm., depending on the amount of blood which they contain, for
practically all the young corpora lutea contain blood in their centers
(figs. 13 and 17). A hemorrhage from a blood-vessel in the wall of the
follicle must occur as the follicle bursts. The exact point of this burst-
ing cannot be made sure of in any of these structures. Each luteal
body is surrounded by a very thin connective-tissue capsule, no doubt
the same theca externa which surrounded the follicle. From this thin
capsule to the central core of blood are massed the luteal cells. They
1382 . DELLA DRIPS
seem to have no definite arrangement. Among them may be seen
numerous fibroblasts and endothelial cells. These are most numerous
about the periphery of the central mass of blood. Some are already
making their way intoit. The cells all seem to have their axes directed
radially as if they were approaching the central mass of blood from the
periphery of the luteal body. The luteal cells are of various sizes and
shapes. Some are spindle-form and some polygonal, but the majority
are spherical or oval. There is one specific characteristic of all young
luteal cells and that is the existence of spherical granules in their pro- —-
toplasm. In the sections fixed in Bensley’s acetic osmic bichromate
and stained with acid fuchsin and methyl green, these granules are
strikingly brought out (fig. 26). They are colored a brilliant red.
Their sizes vary somewhat, but they are all spherical. The protoplasm
of some cells is so full of these granules that it resembles a homogeneous
red secretion, but on examination with very high power, the separate
granules may be seen. In many cells where the granules do not fill
the protoplasm, they are grouped about the nucleus, leaving a narrow
clear zone about the periphery of the cell. The nuclei take the green
stain and are strikingly brought out against the red granules. Each
nucleus contains one or two good-sized bright, red staining nucleoli.
The chromatin threads stain green and do not show very well with
this stain. For the nuclear characteristics, another ovary stained
differently will be described.
The two abnormal luteal structures of this ovary are very interesting.
One of them is a luteal cyst (fig. 23). There is only a single layer of
luteal cells next to the thin capsule. No fibroblasts or endothelial
cells are present. The whole body is filled with a transparent greenish-
colored fluid which has every appearance of being of the same compo-
sition as the liquor folliculi.
The other structure (fig. 21) is much more normal. The center con-
tains what resembles the above green-tinged fluid containing numer-
ous red blood corpuscles. Masses of normal luteal cells are found most
of the way around the body. But on one side is a mass of follicle or
granulosa cells persisting untransformed. In several places, as in
figure 22, some granulosa cells are found among the luteal cells. This
peculiar luteal structure appears to give striking evidence to the theory
that luteal cells are simply transformed granulosa cells. The differ-
ences between them are well brought out in the picture. |
It may be well to state here that other fixatives and stains bring
out these same specific luteal-cell characteristics, particularly the
granules in the protoplasm. With a formalin zenker fixative and the
acid fuchsin and methyl green stain, the granules appear the same in
every way except in color. They are pink instead of red. With this
same fixative and a copper-chrome hematoxylin stain, the granules
appear brownish-black. With an acetic osmic bichromate fixative and
the copper-chrome hematoxylin stain, the granules appear bluish-black.
The nuclear characteristics of the early luteal cells are best brought
out with a formalin zenker fixative and a hematoxylin and eosin stain.
THE OVARY OF THE SPERMOPHILE 133
Experiment 275-16 (spermophile 310). Captured May 1, 1916.
One ovary was removed and the uterus ligated May 4, 1916. -
Gross observations. There are no recognizable signs of pregnancy
in the uterus, but the ovaries contain what resemble young corpora
lutea.
Microscopic observations of thé ovary removed. Fixative, formalin
zenker. Stain, hematoxylin and eosin (fig. 13).
The nuclei of these early luteal cells are strikingly like those of
the follicle cells. The nuclei of the latter have several small nucleoli
with quite conspicuous chromatin strands. The nuclei of most of
these early luteal cells also have several small nucleoli with numerous
chromatin strands. But in some cells, the nucleus is much larger and
contains only one or two larger and darker nucleoli, while the chro-
matin strands are finer and fewer.
A very few of the luteal cells in the early luteal structures show
mitotic figures. There were none at all in those of spermophile 303.
Of all the early corpora lutea studied, mitotic figures were found in
these structures in the ovaries of only three spermophiles. These
were apparently the earliest luteal bodies found. If mitosis occurs, as
a rule it occurs just after the bursting of the follicle. It may be that
the luteal cells which show mitotic figures are the transformed follicle
cells undergoing mitosis as the bursting occurred.
There is one abnormal early luteal structure in the ovary (spermo-
phile 310) which deserves mention, as it seems quite common and
furnishes further proof that the luteal cells are simply transformed
granulosa cells (fig. 24). The structure with this staining appears at
first glance like a mature follicle. The liquor folliculi is present and
the ovum lies over at one side of the central cavity against the sur-
rounding cells, which resemble the granulosa cells of the follicle.
On closer observation, however, it will be seen that the cells which
were thought to be the epithelial cells of the follicle are larger, richer
in protoplasm, and more irregular in shape and size. Scattered through
them are numerous fibroblasts and endothelial cells. These fibroblasts
are quite numerous about the ovum, as if they were attempting to wall
it off. The ovum has been stripped of its own rim of granulosa cells
and appears to be undergoing degeneration. The internal theca is
missing. In fact, what we have here is a corpus luteum formed in a
follicle which, if it burst, did not throw out enough of its contents to
get rid of the ovum. Practically all-of the epithelial cells must then
have been retained. Where are they if they are not the luteal cells?
The only missing cells are those of the internal theca and the only
new cells are the fibroblasts and endothelial cells. Does it not appear
reasonable that the internal theca cells which are of the same origin
as the connective-tissue cells expend themselves in the formation of
the new fibroblasts and endothelial cells? It is hard to account for
these abnormal structures. Several of them showed blood in the
central cavity, as if the normal hemorrhage had occurred into them.
Possibly they are formed in the mature follicles that are ready to burst,
134 DELLA DRIPS
and undergo the same changes incident to this phenomenon as the
others except that, on account of not occupying a position close enough
to the surface of the ovary, they are not able to discharge their contents.
Experiment 293-16 (spermophile 328). Captured May 4, 1916.
Both ovaries were removed May 6, 1916. Weight, 119 grams.
Gross observations. The fetuses-in the uterus measure about 2
mm. in length, which makes the luteal bodies in the ovaries older than
those previously described.
Microscopic observations of the right ovary. Fixative, formalin
zenker. Stain, acid fuchsin and methyl green. This ovary contained
six corpora lutea. These luteal bodies appear differently, due prin-
cipally to the rapid growth which has been going on among the fibro-
blasts and endothelial cells. These ovaries suffered some congestion
through the manipulation of removal, and this helps to show the great
numbers of capillaries and blood-vessels that have been formed in a
_short time. The central mass of blood is undergoing rapid organiza-
tion. No doubt the presence of this blood with its serum and fibrin is
the great attractive force which aids in the complete formation of the
luteal body. Fibroblasts and endothelial cells are always attracted by
serum and fibrin. As soon as the hemorrhage occurs in the follicle,
they start in to organize it. This is evident from the radial direction
which the axis of the fibroblasts all take very early. As they go into
the center, the transformed epithelial cells of the follicle are carried in
by them. Endothelial cells grow in, and so very early there is formed
in the corpus luteum a complex system of blood-vessels and capillaries,
as is seen in sections of this ovary.
In one of the corpora lutea in this ovary, the hemorrhage was so
extensive that instead of trying to organize it, the fibroblasts have.
formed a wall around it. Since the fibroblasts have not penetrated
very far, there is only a narrow rim of luteal cells. This structure is
what is ordinarily called a hemorrhagic luteal cyst (fig. 25). Beside
the corpora lutea in a section of this ovary, one notices readily the large
clumps of interstitial cells through the medullary portion. These are,
as a rule, conspicuous in the ovary of early pregnancy.
Microscopie observations of the left ovary. Fixative, formalin
zenker. Stain, hematoxylin and eosin. This ovary contains only two
corpora lutea. There are present in it several large, apparently mature
follicles. The number of such follicles in an ovary evidently depends
on the number of corpora lutea. Where there are a good many of the
latter, the follicles evidently cannot grow. When there are only a few
corpora lutea in an ovary, one or two follicles may reach the size of
0.5 mm. x 0.5 mm., or 0.56 mm. x 0.7 mm. These will, of course,
degenerate as ovulation takes place only once a year, during the rut-
ting season which has just gone by. And as there are practically no
large follicles ever seen in the ovaries of July which contain the largest
luteal bodies, they must degenerate before this time. Perhaps the
pressure of even one large, growing corpus luteum is enough to bring
this about.
THE OVARY OF THE SPERMOPHILE 135
Experiment 264-16 (spermophile 302). Captured May 1, 1916, and
sacrificed the same day.
Gross observations. This animal is definitely pregnant, the fetuses
measuring 7 mm. in length. The blood-vessels going to the uterus
and the ovary are very much congested. They stand out sharply,
showing plainly the blood supply to the two organs. The blood supply
to the ovary is practically separate from that to the uterus, there being
only one small anastomosing branch close to the ovary. There are
what look like corpora lutea in the ovaries, but they cannot be counted
with any certainty. They resemble little reddish-pink cysts sticking
out from the surface of the ovary. Naturally, from the size of the
fetuses, the corpora lutea in these ovaries are older than those of sper-
mophile 328.
Microscopic observations of the left ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. There are four
luteal structures in this ovary. They show some changes over the
younger ones previously described. They are slightly larger, measur-
ing about 0.7 mm. x0.8mm. The luteal cells have become larger and
more regular in shape. Many more have assumed an oval form and
all seem to be approaching this. They seem to be tending toward a
radial arrangement also. This is being effected evidently by the ar-
rangement of the connective-tissue strands. The latter are running
from the capsule to the central core of blood, which is almost organ-
ized. The system of capillaries and blood-vessels is even more com-
plex than that of the luteal body of spermophile 328. In the luteal
cells themselves the red granules have increased in number and vary
slightly in size. They occupy the same position in the cell. The
nuclei of all the cells are larger and contain one or two large bright
nucleoli with numbers of very fine chromatin strands.
Experiment 296-16 (spermophile 331). Captured and _ sacrificed
May 6, 1916. Weight, 146 grams.
Gross observations. The fetuses in the uterus are 1 ecm. in length,
which lead us to expect to find changes in the corpora lutea of the
ovaries.
Microscopie observations of the left ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. The size ofthe
luteal structures has increased. There are two in this ovary, one
measuring 0.9 mm. x 0.9 mm., and the other, 1 em. x 0.8 mm. The
most noticeable feature of this later luteal body is the absence of any
blood in the center. Instead, there is a core of connective tissue. The
size of this connective-tissue core depends evidently on the amount of
hemorrhagic material there is to organize. In some bodies the core is
much larger than in others. Very perceptible strands of connective
tissue run from this central mass to the capsule, enclosing columns of
luteal cells. Gross strands have developed also so that connective-
tissue strands seem to be enveloping each cell. There is a complete
capillary network following the arrangement of the strands of connec-
tive tissue. Good-sized blood-vessels are located about the periphery
136 DELLA DRIPS
of the structure. Several smaller ones are present in the central con-
nective-tissue core. There are several sinuses about the periphery
lined with endothelium which appear to contain lymph. The lutea
cells themselves have increased in size. Many more have taken on an
oval shape and are lying with their long axes perpendicular to the
capsule. The same red granules are still present in the protoplasm.
These do not seem to be quite as numerous in the cells, which fact is
in part due, no doubt, to the increased size of the latter. The granules
are now found scattered throughout the protoplasm, the clear zone
about the periphery of the cell having disappeared. The granules
show more difference in size than formerly, but they are still all spher-
ical. The nuclei have not changed. In a few cells there is a clear
space in the protoplasm on one side of the nucleus.
Microscopic observations of the right ovary. Fixative, formalin
zenker. Stain, hematoxylin and eosin. There are seven corpora lutea
which make this ovary larger than the other one. Beside these bodies,
this ovary contains approximately five good-sized, growing follicles
about 0.4mm.x 0.4mm. Four follicles nearly the same size are under-
going atretic changes and there are about twenty small follicles, some
of which are degenerating. Around the edge of the ovary are a few
primordial ova. Through the medullary portion are some small
groups of interstitial cells. They are not nearly as conspicuous as
they were earlier in pregnancy (fig. 14).
Experiment 355-16 (spermophile 375). Captured May 20, 1916.
Both ovaries were removed May 22, 1916. Weight, 128 grams.
Gross observations. The fetuses in the uterus are 1.5 cm. in length.
Microscopic observations of the right ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. There are six
corpora lutea in the right ovary, measuring about 0.8 mm. x 0.9 mm. in
diameter. Something is.noted in the luteal structures in this ovary which
has not been seen before, namely, that there are present in the cells close
to the periphery some osmic-stained droplets. The size of the corpus
luteum and the size of the individual cells are about the same as that
of spermophile 331. There is a slight increase in the number* of red
granules in the cells. The clear space next to the nucleus is present
in many more cells. The osmic-stained droplets are located at the
periphery of the cell. They are very large compared with the red
granules. They vary somewhat in size, but not in shape; all are
spherical. Of course, in sections fixed with formalin zenker and stained
with the various stains which were used, these lipoid droplets appeared
as vacuoles. But they could be easily recognized by their correspond-
ing size and location in the cells. For convenience, these droplets
will be called lipoid droplets, because they certainly are a lipoid prod-
uct. They do not appear in the luteal cells before the fetus is about
1.5 cm. in length or about fourteen days old, that is, until the period
of pregnancy is half over. These droplets make up the ‘lutein’ of the
corpus luteum. which has been described for many years and which
has given the corpus luteum its name.
THE OVARY OF THE SPERMOPHILE © 137
When the lipoid droplets appear in the cells, the period of the red
granules is waning. The latter seem to reach their crisis of abundance
when the fetus measures about 8 mm. in length. But the granules are
still very abundant in the cells until the lipoid droplets begin to appear. .
From this time on, the former grow fewer and fewer and the latter
increase in number, as will be shown, until they, too, reach a crisis of
abundance and then decline.
Experiment 363-16 Garenonhale 383). Captured and _ sacrificed
May 24, 1916. Weight 140 grams.
Gross observations. The animal was in labor when killed. The
uterus still contains two live fetuses, four having already been born.
The crown-rump measurement of a fetus is from 4 to 5 em.
Microscopic observations of the left ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. The luteal struc-
tures in the ovaries of this animal show some changes over those pre-
viously described (fig. 18). There are four of them. They have
increased slightly in size, measuring 0.9 mm. x 0.9 mm. and the indi-
vidual cells have increased correspondingly. Still more noticeable
than their increase in size is the regularity of their oval form and the
uniformity with which all their axes point in a radial direction. This
seems to be due to an increased amount of connective-tissue frame-
work, which, from the first, has seemed to govern the position and
shape of the cells. The increase of connective-tissue framework has
been accompanied by an increase in the size of the blood-vessels and
capillaries. In the luteal cells (fig. 27) the red granules have decreased
still more than in the cells last described, and this is very general
throughout the structure. The lipoid droplets are much more numer-
ous in all the cells. In fact, it seems hard to tell which is the predom-
inant product of the cells, the red granules or the black droplets. The
nuclei of these cells are slightly larger than the ones of the preceding
,description. Otherwise, they are the same. Most writers have agreed
“that degeneration of the corpus luteum begins about the time of birth.
None is in evidence here. Several writers have stated that the prin-
cipal reason for their belief was the entrance into the cells of osmic
staining droplets which they considered to be evidences of fatty degen-
eration in the cells. It would not seem from the nuclear character-
istics of the cell nor from the color, shape, and the regular size of the
droplets, that they could be fatty degeneration products, especially
when compared with the true fatty degeneration which occurs much
later in the life history of the corpus luteum and which will be described
accordingly.
Microscopic observations of the right ovary. Fixative, formalin
zenker. Stain, hematoxylin and eosin. There are two corpora lutea
in this ovary. A striking feature is the number of good-sized atretic
follicles. There are at least twelve. This seems to be a noticeable
feature of other ovaries about this same time. In fact, the follicles
which were growing when ovulation took place, evidently go on and
develop if there are not too many corpora lutea in the ovary, but as
138 DELLA DRIPS
the latter structures begin to increase in size quite rapidly about the
time of parturition, there seems to be a degeneration of all the mature
follicles even where there are only one or two luteal structures present.
There are five medium-sized and about twenty small growing normal
follicles, only a few of which show atretic changes. The primordial
ova are very few in number, and the interstitial cells can scarcely be
distinguished from the connective-tissue cells of the stroma.
Experiment 368-16 (spermophile 387). Captured May 24, 1916.
Sacrificed May 25, 1916, twelve hours after having given birth to
young. Weight, 115 grams.
Microscopie observations of the left ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. The three luteal
structures in the left ovary of the animal measure either 0.9 mm. x
1.1 cm. or 0.9 mm. x 1 em. The appearance of the luteal structure
as a whole and of the individual cells is very similar to that of sper-
mophile 383.
Experiment 369-16 (spermophile 388). Captured May 24, 1916, and
sacrificed May 26, 1916, twenty-four hours after having given birth
to young. Weight, 115 grams.
Microscopic observations of the left ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. The corpora lutea
of the ovaries of this animal measure 0.8 mm. x 0.9 mm. There is
scarcely any new noticeable difference unless it is a slight increase in
the number of lipoid droplets.
Experiment 370-16 (spermophile 389). Captured May 24, 1916.
It gave birth to young on May 25, 1916. It was with the young until
May 26, 1916, when it killed them. The next day, May 27, 1916, the
animal was sacrificed. Weight, 135 grams.
Microscopic observations of the left ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. The corpora
lutea in this ovary show a marked increase in the amount of lipoid in
the luteal cells. The red granules have correspondingly decreased in
number. There are no other differences except a slight increase in
size of the luteal structure and the cells.
Experiment 372-16 (spermophile 391). Captured May 18, 1916,and
gave birth to normal young May 22, 1916. The young were destroyed
May 26, and the animal was sacrificed May 31, 1916.
Microscopic observations of the left ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. The corpora lutea
in the ovaries show the lipoid content to be still more increased in
amount and the red granules to be very scarce. There are no other
differences. It might be stated here that there is some variation in
the time when this lipoid change comes on. For instance, the corpora
lutea of the ovaries of spermophile 390, ten days after parturition, do
not contain as much lipoid as those of spermophile 391, nine days after
parturition. But the majority of luteal bodies for any given time
before or after parturition appear very similar.
oe
THE OVARY OF THE SPERMOPHILE 139
Experiment 351-16 (spermophile 371). Captured May 20, 1916.
The left ovary was removed on May 22, 1916. Weight, 135.5 grams.
Gross observations. The fetuses in the uterus are 2 em. in length.
This animal gave birth to young May 31, 1916, and was sacrificed
June 17, 1916.
Microscopic observations of the right ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. The individual
luteal cells are of about the same size and shape. The red granules
are still quite abundant, but there seems to be a marked difference in
the cells as to their individual content of red granules. This appears
to vary with the number of lipoid droplets in the cell. In a few cells
where the latter are very abundant, the red granules are found only
in a rim about the periphery. Where the lipoid droplets are still few
the cell protoplasm still contains many red granules with these few
droplets scattered among them or occupying a peripheral position.
Many cells have not changed at all; they have no lipoid droplets.
Some cells have a peculiar appearance. Their protoplasm appears
honeycombed. Since none of the cells had this appearance in the
unstained sections, it was concluded that the cells must have been filled
with lipoid which was dissolved out in the staining process and the
result was this honey-combed appearance to the protoplasm. This
was later proved to be the case. The lipoid droplets, when they aré
numerous, show much less variance in size than when they are few in
number in the cell. The nuclei of these cells appear very similar to
those of earlier cells. In some cells there is a slight change in their
position. Instead of being directly in the center, in the cells filled
with lipoid, the nucleus lies a little to one side of the center. The Ist
of July, about thirty-five days after parturition, the corpora lutea in
the ovaries were larger than at any other time.
Experiment 412-16 (spermophile 415). Captured and sacrificed
July 1, 1916. Weight, 125 grams.
Gross observations. The ovaries are the largest yet seen, owing
to the comparatively immense size of the corpora lutea. These are
now of a yellowish-cream color and stand out prominently so that the
ovary looks as if it were made up of several spherical bodies 1.5 mm. in
diameter. Any other ovarian tissue except that holding the spherical
bodies together can scarcely be distinguished. There are three corpora
lutea in the right ovary and four in the left, which numbers correspond
to the tiny white spots marking the former placental sites in the uterus.
Experiment 413-16 (spermophile 416). Captured and sacrificed on
July 3, 1916. Weight, 157.3 grams.
Gross observations. The right ovary appeared grossly just like that
of spermophile 415 (fig. 15).
Microscopic observations of the left ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. The corpora
lutea are much larger than those previously described (fig. 19). They
measure 1.1 mm. x 1.5 mm. in diameter (1.7 mm. x 1.3 mm., grossly
some shrinkage). The luteal cells are correspondingly larger, and their
140 DELLA DRIPS
protoplasm is absolutely full of a mass of lipoid droplets. This is seen
to advantage in the unstained sections. In the stained sections the
cells appear more or less honeycombed, according to the amount of
lipoid which has been dissolved out. These lipoid droplets are very
uniform in size and are dark brown, quite a different color from the
black fat droplets of fatty degeneration. The red granules are gone.
Where there is any protoplasm visible, it appears granular and pinkish-
gray in color (fig. 28). The nuclei of the cells appear slightly smaller
than formerly, but this apparent decrease in size is evidently due to
the increase in the size of the cells, for the nuclei are no smaller by
measurement. The location of the nucleus in the cell is the same as
described under spermophile 371, either in the center or to one side of
the center. The nucleolus stands out large and bright and the chro-
matin strands appear as they did. There are no apparent degenerative
changes. Beside the three corpora lutea in this ovary, there are six
or seven medium-sized normal growing and three atretic follicles.
There are no large follicles or even any of good size. A few primordial
ova are present, fewer than in any of the ovaries described thus far,
and no interstitial cells can be distinguished as such (fig. 15).
Experiment 439-16 (spermophile 436). Captured June 23, 1916.
Sacrificed July 15, 1916. Weight, 194.2 grams.
Gross observations. The uterus still shows several tiny white spots
marking the placental sites. It is otherwise normal. The ovaries
appear to contain corpora lutea, but these latter are certainly much
reduced in size over those of spermophile 416. They appear congested
or of a reddish-yellow color.
Microscopie observations of the left ovary. Fixative, acetic osmic
bichromate. Stain, acid fuchsin and methyl green. The corpora
lutea are much smaller (fig. 20). They measure 0.7 mm. x 0.8 mm.
and 0.9 mm. x 0.9mm. The cells are smaller. The protoplasm of
the cells contains no red granules and very few lipoid droplets or any
honeycombing suggestive of these. It has a grayish, granular appear-
ance. Something is present, however, which has not been seen before,
and that is fat. Scattered here and there thoughout the luteal struc-
ture, fat globules, characteristic of fatty degeneration, are present in
the protoplasm of the cells. They are of various sizes and take on a
characteristic black color with the osmic acid in the acetic osmic bichro-
mate fixative. The nuclei of the cells show degenerating changes.
The nucleolus has disappeared in some cells and in others appears
pale and fringed. The chromatin strands are fewer and appear clumped
in some cells. One striking new feature in the luteal structure is the
great increase in the size of the blood-vessels and capillaries. The con-
gestion of blood is not common to the whole ovary, but is only in the
corpus luteum. The vascular change is apparently one factor in the
disappearance of the luteal body.
Experiment 515-16 (spermophile 458). Captured August 18,1916.
Sacrificed August 21, 1916. Weight, 190 grams.
Gross observations. There are no evidences of placental sites in the
uterus. There are no signs of corpora lutea in the ovaries.
THE OVARY OF THE SPERMOPHILE 141
Microscopic observations of the left ovary. Fixative, acetic osmic ®
bichromate. Stain, acid fuchsin and methyl green. The left ovary
shows three luteal stuctures (fig. 16). The largest measures 0.8 mm.
x 0.8mm. No red granules or lipoid droplets are discernible. The
greenish-gray granular protoplasm is everywhere filled with various
sized fat droplets. In some cells these fat droplets are so large that
they occupy nearly the whole cell, squeezing the degenerated nucleus
out to one side of the cell (fig. 29). The nuclei are so degenerated,
no. chromatin strands or nucleolus are recognizable as such. What
is left of the nucleus takes the acid fuchsin rather than the methyl
green stain—so it is red. All the blood-vessels and capillaries are
markedly dilated. There is a marked increase of connective tissue
throughout the body. The thin capsule of connective tissue which
before surrounded the luteal structure seems to have disappeared in
places, making it appear as if the connective tissue of the body were
continuous with that of the ovary around it. This connective-tissue
invasion is evidently another factor in the disappearance of the corpus
luteum. Three factors, then, are associated with the disappearance of
the corpora lutea in the ovaries, cellular degeneration, vascular dilata-
tion, and connective-tissue Invasion.
A word may be added concerning the changes in the ovary
outside the luteal bodies. There were in this organ six or seven
good-sized, normal, growing follicles and four and five atretic
ones. The most noticeable feature is the number of small
hyalinized follicles. This is a noticeable feature of all the
ovaries of this date which contain old corpora lutea. It would
appear that as long as there are still luteal bodies in the ovary,
there is very little growth in the follicles. There are a few
primordial ova. The interstitial cells are not recognizable as
such. By September 1 the corpora have disappeared from the
ovaries and the organs again have assumed the appearance
described for September 15.
There are occasional exceptions to the normal cycle. An
ovary of spermophile 462, sacrificed October 30, 1916, was
found with a little structure in it which appeared in every way
to be a young corpus luteum. The cells contained red granules.
Perhaps impregnation had occurred in the fall. This instance is
mentioned because one function of the corpus luteum in spermo-
philes has been suggested by it. What keeps the follicles from
becoming mature and ovulation from taking place at other
times in the year from the spring? Evidently, the corpora
142 DELLA DRIPS
v lutea help in regulating the periods between oestrus. No new
follicles become mature in the fall as long as there are luteal
bodies in the ovary and by the time the latter have disappeared
it is too late for the follicles to become mature before the period
of hibernation comes on.
A word concerning the interstitial cells may be added. These
appear very prominent at two periods of the cycle, during the
period of early pregnancy when the corpora lutea are young
and apparently very active, and in the early fall when the fol-
licles are developing and growing rapidly. Their significance is
not evident. Several writers have claimed that the cells of the
corpus luteum become the interstitial cells of the ovary. There
is absolutely no basis for such a supposition as far as the spermo-
philes are concerned. |
EXPERIMENTAL STUDIES
These experiments were made with a view to determining
whether or not the results of certain experimental studies on the
spermophile would substantiate the same work done on other
animals.
The spermophile stand experimental surgery well. They are
very satisfactory to work on as they are not nearly as susceptible
to infectious diseases and are more resistant to local infections
than most small laboratory animals. They are easily anes-
thetized. The surgical technic must be aseptic, and the animals
must be placed in separate cages after the operation or they will
chew one another’s wounds open. They recover from the effects
of the operation quickly, as a rule, and the wounds usually heal
by first intention.
To all workers who have been interested in the functional
relationship of the ovaries and the uterus and especially to those
trying to isolate a particular function or functions for the corpora
lutea, two problems have seemed of vital importance: First,
what are the effects on the uterus of the removal of one or both
of the ovaries? And, second, what are the effects on the
ovaries of the extirpation of the uterus? ‘These two problems
THE OVARY OF THE SPERMOPHILE 143
have a special bearing on the function of the corpora rhe
when they dea! with pregnant animals.
Another much studied problem closely related to the {unction
of the luteal bodies attempts to expla'n the means by wh ch
ovulation s brought about and the results on the development
of the corpora lutea i fertilization is prevented.
Following are protocols and results of experiments performed
in connection with these various problems. Only those experi-
ments which proved operative successes and in which reliable
data were obtained wil! be included.
Series 1. Effects of the removal of the uterus on the ovaries of non-
pregnant spermophiles
Experiment 230-15 (spermophile 146). Captured in the spring of
1915. Weight, 128 grams. Operated on June 11, 1915. Complete
removal of the uterus.
Gross observations. There are good-sized corpora lutea in the
ovaries.
Sacrificed July 20, 1915.
Gross observations. The ovaries appeared very much smaller.
Microscopic observations of the ovaries. Fixative, formalin zenker.
Stain, hematoxylin and eosin. The ovaries show no degenerative
changes except in the corpora lutea. These are undergoing regression.
Connective tissue has heavily invaded every part of the luteal struc-
ture. The blood-vessels are numerous and good sized. There are
many fat vacuoles present. The decrease in size of these ovaries is
no doubt due to the decrease in the size of the corpora lutea.
Experiment 443-15 (spermophile 247). Captured in the spring of
1915. Weight, 207 grams. Operated on October 1, 1915. Complete
removal of the uterus. '
Died October 8, 1915.
Gross observations. Death was due to peritonitis.
Microscopic observations of the ovaries. Fixative, formalin zenker.
Stain, hematoxylin and eosin. There is a marked congestion through-
out the ovary. Many of the larger follicles are undergoing degenera-
tion. Practically all the smaller follicles are markedly degenerated.
This, no doubt, is a pathologie condition.
Experiment 444-15 (spermophile 248). Captured in the spring of
- 1915. Weight, 196 grams. Operated on October 1, 1915.
Died January 13, 1916.
Microscopic observations of the ovaries. Fixative, formalin zenker.
Stain, hematoxylin and eosin. The great number of small hyalinized
follicles are immediately apparent. They are so numerous that the
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, No. 2
144 DELLA DRIPS
ovary has a lacy appearance. There are a few good-sized, apparently
normal follicles and on the periphery are numbers of primordial ova.
Experiment 445-15 (spermophile 249). Captured in the spring of
1915. Weight, 110 grams. Operated on October 4, 1915. Complete
removal of the uterus.
Sacrificed April 22, 1916.
Gross observations. The blood supply to the ovaries is intact.
The ovaries are very small; the left so small as to leave doubt as to its
identity. The right ovary appears to contain several tiny cysts.
Microscopic observations of the ovaries. Fixative, formalin zenker.
Stain, hematoxylin and eosin. Tissue removed for left ovary proves
to be a bit of granulation tissue. The right ovary appears to be nor-
mal. The several cysts noted grossly are large mature follicles which
normally occur on the surfaces of the ovaries at this time of the year.
No effects from the removal of the uterus are noted.
Discussionand summary of results. There seemsto be no marked
effect on the ovaries from the removal of the uterus They seem
able to repeat their life cycle as far as the follicles are concerned.
It was thought that the great number of hyalinized follicles noted
in the ovaries of spermophile 248 was a sign of degeneration due
to removal of the uterus. However, on studying the ovaries of
normal spermophiles killed about the same time for controls,
there were found in the ovaries of two a great number of these
hyalinized follicles (fig. 11). Evidently the hyalin degeneration
occurs normally at this time of the year. It is, no doubt, part
of the attempt to produce a few large mature follicles at the
expense 0 many smaller ones.
Series 2. Effects of removal of the uterus on ovariés in pregnant
animals
Experiment 270-16 (spermophile 305). Captured May 1, 1916.
Weight, 98 grams. Operated on May 4, 1916. The uterus was
‘removed to the cervix.
Gross observations. There are no signs of pregnancy in the uterus,
but the right ovary contains what appears to be a hemorrhagic cyst.
Died May 9, 1916.
Gross observations. There was no apparent cause for death.
Microscopic observations of one ovary. Fixative, formalin zenker. -
Stain, hematoxylin and eosin. The ovary contains several corpora
lutea of an early stage. One is markedly hemorrhagic. There is some
degeneration apparent in the luteal bodies. Some cells are undergoing -
chromatolysis. There is an increased amount of connective tissue.
The rest of the ovary appears to be in a very normal condition.
THE OVARY OF THE SPERMOPHILE 145
Experiment 294-16 (spermophile 329). Captured May 4, 1916.
Weight, 117 grams. Operated on May 6, 1916. The uterus was
removed to the cervix.
Gross observations. The animal appears to be in early pregnancy.
Sacrificed May 20, 1916. Weight, 130 grams.
Gross observations. There are no adhesions about the ovaries;
they appear to contain several white, transparent corpora lutea.
Microscopic observations of the right ovary. Fixative, formalin
zenker. Stains, hematoxylin and eosin, and acid fuchsin and methyl
green. There is no apparent pathologic condition of this ovary. Even
the corpora lutea appear normal. They have proceeded in their
development without the uterus and now have the same appearance
as others of the same age. The only detectable difference might be a
smaller number of red granules in the luteal cells.
Experiment 251-16 (spermophile 348). Captured May 6, 1916.
Weight, 108 grams. Operated on May 9, 1916. The uterus was
removed to the cervix.
Gross observations. The animal is pregnant. There are five
placental swellings in each horn, measuring 6.5 mm. in length.
Sacrificed May 19, 1916. Weight, 102 grams.
Gross observations. Everything in the abdominal cavity is in
excellent condition.
Microscopic observations of the ovaries. Right ovary: fixative
acetic osmic bichromate; stain, acid fuchsin and methyl green. Left
ovary: fixative, formalin zenker; stain, Weigert’s copper-chrome
hematoxylin. The ovaries are apparently normal. The corpora lutea
appear like those of the controls except that the luteal cells contain a
larger number of lipoid droplets and fewer red granules. There are no
signs of degeneration in the nuclei of the cells.
Experiment 314-16 (spermophile 349). Captured May 6, 1916.
Weight, 122 grams. Operated on May 9, 1916. The uterus was
removed to within 3 mm. of the cervix.
Gross observations. The animal is pregnant. There are four
placental swellings in the right horn and five in the left about 4 mm.
in length.
Sacrificed June 5, 1916. Weight, 135 grams.
Gross observations. Everything in the abdominal cavity is in
excellent condition. Both ovaries were readily found and appear to
contain large corpora lutea.
Microscopic observations of the ovaries. Right ovary: Fixative,
formalin zenker; stain, Weigert’s copper-chrome hematoxylin. Left
ovary: fixative, acetic osmic bichromate; stain, acid fuchsin and
methyl green and Weigert’s copper-chrome hematoxylin. There is no
apparent abnormality in the ovaries. The corpora lutea have devel-
oped normally. They are still in the red-granule stage. The cells
contain some lipoid droplets, but are full of red granules. The lipoid
droplets in these ovaries are not as numerous as would be expected,
but normal ovaries vary somewhat as to the time when the lipoid
droplets begin to appear in the cells.
146 DELLA DRIPS
Experiment 317-16 (spermophile 353). Captured April 25, 1916.
Weight, 116 grams. Operated on May 9, 1916. The uterus was
removed to the cervix.
Gross observations. The animal is pregnant and, having been kept
by itself since capture, the period of pregnancy must be at least fourteen
days. The placental swellings measure 13 mm. in length and 10 mm.
in width.
Sacrificed May 20, 1916. Weight, 130 grams.
Microscopic observations of the ovaries. Right ovary: fixative,
formalin zenker; stains, hematoxylin and eosin, and Weigert’s copper-
chrome hematoxylin. Left ovary: fixative, acetic osmic bichromate;
stain, acid fuchsin and methyl green. There is no pathologic condi-
tion apparent in the ovaries. The corpora lutea have gone on in their
normal development. They appear very ‘similar to those of spermo-
philes whose fetuses are nearing parturition.
Experiment 318-16 (spermophile 353). Captured April 28, 1916.
Weight, 127 grams. Operated on May 9, 1916. The uterus was
removed to the cervix.
Gross observations. The animal is pregnant and, since it has been
kept by itself since capture, it must have been pregnant at least eleven
days. The placental swellings measure 7 mm. in length.
Sacrificed May 31, 1916. Weight, 120 grams.
Gross observations. The ovaries appear very small. The blood
supply is intact and there are no adhesions. There is a mass of fat
around each ovary.
Microscopic observations of the ovaries. The right ovary was
carefully studied. The other was lost. Fixative, formalin zenker.
Stains, hematoxylin and eosin, and acid fuchsin and methyl green.
Many degeneration changes are present all through the ovary. Prac-
tically all the follicles are degenerating. The corpora lutea show many
degeneration changes also. There are practically no red granules in
the cells and many fatty degeneration vacuoles are present. The
nuclei of the cells show degenerative changes. Their margins are
indented and their nucleoli are faded and fringed. The strands of
chromatin are clumped. The capillary sinuses seem dilated and
filled with blood. Evidently the ovaries have been injured by the
operation.
Experiment 331-16 (spermophile 361). Captured May 4, 1916.
Weight, 141 grams. Operated on May 12, 1916. The uterus was
removed to the cervix.
Gross observations. The animal is pregnant, the placental swell-
ings being 9 mm. in length.
Sacrificed May 20, 1916. Weight, 120 grams.
Microscopic observations of the ovaries. Right ovary: fixative,
formalin zenker; stains, acid fuchsin and methyl green, and hema-
toxylin and eosin. The left ovary was lost. The right ovary shows no
abnormality. The corpora lutea have gone on in their development.
They are just entering the lipoid stage.
THE OVARY OF THE SPERMOPHILE 147
Experiment 61-16 (spermophile 381) Captured May 20, 1916.
Weight, 145.5 grams. Operated on May 22, 1916. The uterus was
removed to the cervix.
Gross observations. The animal has given birth to young, probably
a few days previously, as the uterus appears to be in an early stage of
involution. There are good-sized corpora lutea in the ovaries.
Sacrificed October 20, 1916. Weight, 170 grams.
Gross observations. The ovaries are so small they are hard to find.
Microscopic observations of the ovaries. Right ovary: fixative,
formalin zenker; stain, hematoxylin and eosin. Left ovary; fixative,
acetic osmic bichromate; stain, acid fuchsin and methyl green. There
is nothing abnormal about the ovaries. They are very similar to other
ovaries removed at this time. The corpora lutea have disappeared
normally.
Discussion and summary of results. Removal of the uterus
has no apparent pathologic effect on the ovaries conta ning the
corpora lutea of pregnancy. The latter pass through their
norma’ cycle of development and regression. There seems to
be a slight irregularity about the time at which the various
changes in the life cycle come on; for instance, the corpora lutea
in the ovaries of spermophile 348 seem to lose the red granules
and take on the lipoid droplets sooner than normally. Then, in
the ovaries of spermophile 349, the corpora lutea have retained
their red granules longer than usual. However, this may not
be significant since there is some irregularity about the cycle
normally.
Series 3. Effects of the removal of both ovaries on the uterus of
non-pregnant animals
Experiment 200-15 (spermophile 189). Captured in the spring of
1915. Weight, 130 grams. Operated on May 28, 1915. Both ovaries
were removed.
Gross observations. The uterus is in a stage of early involution.
Sacrificed April 22, 1916.
Gross and microscopic observations of the uterus. Fixative, for-
malin zenker. Stain, hematoxylin and eosin. The uterus is very
small. It shows no signs of rut of having been in rut, either grossly
or microscopically. This is very abnormal for this time of the year.
A comparison of this uterus with a normal uterus in rut at this time of
the year brings out the differences. (Compare figs. 1 and 3; also
2and 4.) A comparison of the dimensions of the two uteri brings out
the marked differences in size (table).
148 DELLA DRIPS
Microscopic observations of the uterus. Fixative, formalin zenker.
Stain, hematoxylin and eosin. Cross-sections through the horns and
body of the uterus show much fibrosis of all the layers. There is
much less muscular and glandular tissue than in a normal inactive
uterus (compare figs. 3 and 5, 4 and 6). One striking feature is the
number of obliterated blood-vessels with hyalinized walls. The lumen
of the uterus is closed and no mucous secretion is present. The cervix
shows these same changes to an even greater degree than the rest of
the organ.
Experiment 201-15 (spermophile 140). Captured in the spring of
1915. Weight, 101 grams. Operated on May 28, 1915. The ovaries
were completely removed.
Gross observations. The uterus is in the condition of early involu-
tion.
Postoperative notes. This animal became very fat and went into
a torpid condition at times. On July 24 it was changed to a cage with
two other animals who killed it. The body was badly traumatized
and specimens were not saved.
Experiment 202-15 (spermophile 141). Captured in the spring of
1915. Weight, 108 grams. Operated on May 28, 1915. Both ovaries
were removed.
Gross observations. The uterus is undergoing involution.
Died September 21, 1915.
Gross and microscopic observations of the uterus. Fixative, for-
malin zenker. Stain, hematoxylin and eosin. This uterus appears
very similar to the uterus of a normal animal killed the same day.
Experiment 218-15 (spermophile 142). Captured in the spring of
1915. Weight, 135 grams. Operated on June 3, 1915. Both ovaries
were removed.
Gross observations. The uterus is undergoing involution.
Sacrificed November 10, 1915.
Gross and microscopic observations. Fixative, formalin zenker.
Stain, hematoxylin and eosin. The uterus is very similar to the uteri
of the controls.
Experiment 220-15 (spermophile 151). Captured in the spring of
1915. Operated on June 28, 1915. Both ovaries were removed.
Gross observations. The uterus has become completely involuted.
The uterus was removed September 28, 1915.
Gross and microscopic observations. Fixative, formalin zenker.
Stain, hematoxylin and eosin. Compared with a normal control, this
uterus shows no abnormalities. .
Experiment 423-15 (spermophile 231). Captured in the spring of
1915. Weight, 200 grams. Operated on September 22, 1915. Both
ovaries were removed.
Sacrificed March 18, 1916. Weight, 100 grams.
Gross and microscopic observations of the uterus. Fixative, for-
malin zenker. Stain, hematoxylin and eosin. This uterus shows the
‘same changes as that of spermophile 139.
THE OVARY OF THE SPERMOPHILE 149
Experiment 424-15 (spermophile 232).. Captured in the spring of
1915. Weight, 205 grams. Operated on September 22, 1915. Both
ovaries were removed.
Gross observations. The uterus is normal.
Sacrificed May 8, 1916. Weight, 117 grams.
Gross and microscopic observations. Fixative, formalin zenker.
Stain, hematoxylin and eosin. This uterus shows the same changes as
that of spermophile 139. (Photograph of this uterus with a normal
uterus of rut removed the same day (fig. 7). Table for measurements
of these uteri).
Experiment 435-15 (spermophile 242.) Captured in the spring of
1915. Weight, 210 grams. Operated on September 28, 1916. Both
ovaries were removed.
Sacrificed May 9, 1916. Weight, 155 grams. The same day a
control animal, spermophile 233, which was captured the same time as
spermophile 242, and had lived in the laboratory under the same con-
ditions, was sacrificed also and the uteri of these two animals were
photographed together (fig. 8).
Gross and microscopic observations. Fixative, formalin zenker,
Stain, hematoxylin and eosin. The uterus of spermophile 242 shows
all the changes noted in that of spermophile 139. (For dimensions of
uterus see table.) The control uterus (table) does not show as marked
enlargement as the uteri of animals brought in from the fields in the
condition of rut, but aside from this variation in size, the uterus ap-
pears in every way like a typical one of rut.
Experiment 436-15 (spermophile 246). Captured in the spring of
1914. Weight, 95 grams. Operated on July 10, 1914. Both ovaries
were completely removed.
Sacrificed September 29, 1915.
Gross and microscopic observations. Fixatve, formalin zenker.
Stain, hematoxylin and eosin. Compared with the uteri of the con-
trols, this uterus shows atrophic changes of a similar nature to those of
the uterus of spermophile 139, except that the atrophy must be even
more marked to be noticed in a comparison with the controls of this
time of the year.
Experiment 354-16 (spermophile 874). Captured May 20, 1916.
Weight 128 grams. Operated on May 22, 1916. Both ovaries were
removed.
Gross observations. The uterus is undergoing involution.
Sacrificed October 30,1916. Weight, 120 grams. This uterus was
photographed with that of a control animal and that of an animal
Co eens and sacrificed on the same date—spermophile 378
fig. 9).
Experiment 358-16 (spermophile 378). Captured May 18, 1916.
Weight, 115 grams. Operated on May 22, 1916. Both ovaries were
removed.
Sacrificed October 30, 1916. Weight, 135 grams. This ae was
photographed with that of spermophile 374 and a control.
150 DELLA DRIPS
Gross and microscopic observations. Some slight differences can be
noted between the uteri of spermophiles 374 and 378 and their control.
The uteri of the doubly ovariectomized spermophiles are both smaller
than their control. This decrease in size is more noticeable in the
cervix than in the rest of the uterus. The cervices of these two uteri
are much firmer also and contain very little mucus. It is true, the
differences are only slight and are scarcely recognizable microscop-
ically, especially after fixation. The main microscopic difference is a
decrease in size of the blood-vessels in the uteri of the ovariectomized
spermophiles. Perhaps this accounts for the fact that the uterus of
the control has a healthier appearance grossly.
Discussion and summary of results. The uteri of animals
doubly ovariectomized in the spring of the year show some
changes over their controls by the fall of the same year. These
are slight and are all of the nature of a functional atrophy. The
cervix is affected the most. This atrophy increases so as to be
quite noticeable by the fall of the next year. The very striking
effect of double ovariectomy is the discontinuation of the changes
in the uterus incident to the phenomena of rut.
Series 4. Effects of removal of both ovaries on the pregnant uterus
Experiment 293-16 (spermophile 328). Captured May 4, 1916.
Weight, 119 grams. Operated on May 6, 1916. Both ovaries were
removed.
Gross observations. The animal is in an early stage of pregnancy,
the placental swellings being just visible grossly in the uterus.
Sacrificed May 20, 1916. Weight, 115 grams.
Gross observations. There are no signs of placental sites in the
uterus. To be sure the animal had been pregnant, the ovaries were
studied carefully. They contained early corpora lutea. Evidently
the regression changes began in. the uterus immediately after the re-
moval of the ovaries.
Experiment 300-16 (spermophile 335). Captured in the spring of
1916. Weight,101 grams. Operated on May 8,1916. Both ovaries
were removed.
Gross observations. The animal was in an early stage of pregnancy,
the placental swellings being just large enough to be recognizable.
Died May 12, 1916.
Gross observations. The external wound is in bad condition, per-
haps the fault of too much iodin. The inside of the abdominal cavity
appears normal. The uterus is in good condition save on the ends
where the blood supply has been injured. The congestion is much
reduced. There are placental swellings, hard and dark red; five in
the right horn and three in the left, very hard to see. The placentas
and fetuses are apparently undergoing degeneration.
THE OVARY OF THE SPERMOPHILE Lt
Experiment 302-16 (spermophile 337). Captured in the spring of
1916. Weight, 120 grams. Operated on May 8, 1916. Both ovaries
were removed. ;
Gross observations. The animal is pregnant, the placental swellings
being 6 mm. in diameter.
Sacrificed May 18, 1916. Weight, 142 grams.
Gross observations. The uterus is in a very unnatural condition.
It appears dark red in color all over. (The color is not due to conges-
tion as can be seen on microscopic section. Instead, it must be due to
the presence of old clotted blood in the lumen.) There is one hard
swelling still palpable. The pregnancy was interrupted and the
placentas with the fetuses have been undergoing degenerative changes.
Experiment 303-16 (spermophile 338). Captured April 23, 1916.
Weight, 221.5 grams. Operated on May 8, 1916. Both ovaries were
removed.
Gross observations. The animal is pregnant, the placental swellings
being of good size.
Sacrificed May 19, 1916. Weight, 140 grams.
Gross observations. The uterus is very dark-red colored, dead-
looking and contains several dark-colored swellings. The blood supply
to the uterus is intact as tested by Dr. Mann. It appears as though
degeneration of the placentas with the fetuses has been going on.
Experiments 305-16 (spermophile 340). Captured May 6, 1916.
Weight, 124 grams. Operated on May 8, 1916. Both ovaries were
removed.
Gross observations. The animal is in an advanced condition of
pregnancy, the placental swellings measuring 2.2 cm. in length and
1.3 em. in breadth.
Died May 11, 1916.
Gross observations. The cause of death could not be determined.
The uterus is very unhealthy appearing, dark red in color. The
remaining placental swellings are of various sizes. They are very dark
red and are hard. Apparently degeneration changes have been going
on.
Experiment 307-16 (spermophile 342). Captured May 6, 1916.
Weight, 142 grams. Operated on May 8, 1916. Both ovaries were
removed.
Gross observations. The animal.is pregnant, the placental swell-
ings being 6.5 mm. in length. There are nine swellings in the left horn
and three in the right.
Sacrificed May 20,1916. Weight, 152 grams.
Gross observations. The uterus is very dark in color. There are
some adhesions on the left end. In the left horn are four swellings,
the first and fourth measuring 4 mm. in length and breadth, the second
4mm. by 5 mm., and the third, 2 mm. x 2 mm. In the right horn
are three swellings—the first measuring 4 mm. x 4 mm., the second
L52Z DELLA DRIPS
5mm.x5mm., and the third, 3mm.x2mm. Each swelling con-
sists of a light-colored band surrounding a hard dark red mass. The
same condition was found in all the other cases. Some of the pla-
cental swellings with the fetuses have disappeared entirely, the others
are degenerating.
Experiment 333-16 (spermophile 362). Captured April 28, 1916.
Weight, 168 grams. Operated on May 12, 1916. Both ovaries were
removed. ,
Gross observations. The animal is in advanced pregnancy. The
operation was difficult as there were so many large fetuses. No at-
tempt was made to count the number for fear of disturbing them.
Sacrificed May 20, 1916. Weight, 125 grams.
Gross observations. The uterus appears about normal in color.
There are three swellings in the left horn, 4mm. by 5 mm., and ten in
the right horn, same size, dark red and hard.
Experiment 355-16 (spermophile 375). Captured May 20, 1916.
Weight, 143 grams. Operated on May 22, 1916. . Both ovaries were
removed.
Gross observations. The animal is in advanced pregnancy. The
operation was performed with very little trauma, the fetuses not being
removed from the abdominal cavity. The placental swellings were
1.5 em. or more in length.
Sacrificed May 26, 1916. Weight, 130 grams.
Gross observations. The animal had aborted some time previously,
perhaps twenty-four hours. There are seven placental sites in the
right horn and two in the left, all 5 mm. x 6 mm. The uterus is
normal in color. The placental swellings appear congested but not
hard. They look very different from those previously described.
Experiment 357-16 (spermophile 377) Captured May 18, 1916.
Weight, 116 grams. Operated on May 22, 1916. Both ovaries were
removed.
Gross observations. The animal is in advanced pregnancy, the
placental swellings measuring 2 cm. in length. The operation was
performed with very little trauma, the fetuses not being removed from
the abdominal cavity.
Sacrificed May 25, 1916. Weight, 110 grams.
Gross observations. The animal must have aborted shortly after
the operation. There are three placental sites in the right horn and
four in the left, 5 mm. by 4 mm.
Experiment 309-16 (spermophile 344). Captured May 6, 1916.
Weight, 120 grams. Operated on May 8, 1916. Instead of removing
both ovaries, the left ovary was removed and the right tube ligated on
the opposite side.
Gross observations. The animal is pregnant, the placental swellings
measuring 1.4 cm. in length. There are many of the latter in the left
horn and only one in the right.
Sacrificed May 20, 1916. Weight, 120 grams.
THE OVARY OF THE SPERMOPHILE 153
Gross observations. The uterus appears healthy, a normal pale
pink color, but it is very much swollen (table). There are three small
palpable swellings i in the left horn LS only from the increased
size of the uterus at these points. Placental sites in the right horn
not definitely established. Evidently the animal had aborted. The
cause of the swollen condition of the uterus is not evident.
Microscopic observations of the ovaries. Left ovary: fixative,
formalin zenker; stain, hematoxylin and eosin. Rfght ovary: fixative,
formalin zenker; stain, hematoxylin and eosin. The left ovary re-
moved at operation May 8, 1916, is normal. It contains eight corpora
lutea, 0.8 mm. x 0.9 mm. in size. The right ovary obtained at au-
topsy May 20, 1916, is very interesting. It contains only one corpus
luteum, 0.4 mm. x 0.5 mm. in size. This is in the final stage of its
life cycle, the degenerative stage described in the first part of this
work. The rest of the ovary is normal. Through its cortex are eight
large follicles, apparently mature, not showing any atretic changes.
One of these measures 0.8 mm. x 0.6 mm. There are many smaller
growing follicles also, and quite a number of primordial ova. Almost
in the middle of the ovary, just inside the cortex, is the one degenerated
corpus luteum. It is full of large fatty degeneratiye vacuoles and
enlarged blood-vessels and capillaries. The connective tissue has made
great inroads. What made this corpus luteum degenerate is not evi-
dent. It will be shown later that after the removal of one ovary the
other showsno pathologic effects. Andevenafter removal of one ovary
and a uterus containing fetuses, the other ovary with its corpora lutea
shows no pathology. Evidently the ligation of the tube caused the
corpus luteum in the right ovary to degenerate and abortion occurred,
the same as on removal of both ovaries. This would make it appear as
if the corpus luteum was the part of the ovary necessary to the develop-
ment of the fetuses.
Discussion and summary of results. Ten animals were operated
on to get results from the removal of both ovaries containing
corpora lutea on the uterus containing fetuses. One of the
animals died from too much ether. The others lived, but none
of them came to term. Those operated on early in pregnancy
did not abort. The placentas and fetuses degenerated. Those
operated on when the pregnancy was well advanced, aborted.
That this was not the effect of operative trauma will be shown by
another series of experiments on the removal of one ovary in
pregnant spermophiles.
154 DELLA DRIPS
Series 5. Effects of the removal of one ovary on the one remaining
in non-pregnant spermophiles
Experiment 219-15 (spermophile 143). Captured in the spring of
1915. Operated on June 3, 1915. The left ovary was removed.
Gross observations. There are spots marking placental sites in the
uterus. ‘
Right ovary removed September 20, 1915.
Microscopic observations of the ovaries. Both ovaries: fixative,
formalin zenker; stain, hematoxylin and eosin. The left ovary contains
several large corpora lutea of the late lipoid stage. The right ovary
contains no corpora lutea. It is a typical ovary of the fall of the year.
There are many medium-sized growing follicles and many primordial
ova. There is no demonstrable pathology. The corpora lutea must
have disappeared normally.
Experiment 221-15 (spermophile 159). Captured in the spring of
1915. Operated on June 29, 1915. The left ovary was removed.
Gross observations. The uterus is normal and inactive.
The right ovary was removed August 5, 1915.
Microscopic observations of the ovaries. Both ovaries; fixative,
formalin zenker; stain, hematoxylin and eosin. The left ovary con-
tains six very large corpora lutea of the lipoid stage. The right ovary
contains two small corpora lutea in the final stage of their life cycle,
evidenced by their size, the large amount of fatty degeneration, the
numerous large blood-vessels, and the great invasion of connective
tissue. This ovary contains many medium-sized, ripening follicles,
also many small follicles and primordial ova. No pathology is evident.
It is a typical ovary of August 5. The results of these two successful
experiments were so evidently negative that it seemed unnecessary to
repeat them the next year.
Series 6. Effects of the removal of one ovary on the other ovary and
the uterus in pregnant spermophiles
These experiments were controls for the experiments under
series 4.
Experiment 352-16 (spermophile 372). Captured May 20, 1916.
Weight, 148 grams. Operated on May 22, 1916. The left ovary was
removed.
Gross observations. The animal is pregnant, the fetuses measuring
1.5 em. in length. There are not many fetuses.
Sacrificed May 25, 1916. Weight, 135 grams.
Gross observations. All the fetuses are alive and no abnormalities
are apparent. There are five fetuses in the right horn and two in the
left. They measure 2.7 cm. by 1.5 cm.
THE OVARY OF THE SPERMOPHILE | 155
Microscopic observations of the ovaries. Right ovary: fixative,
formalin zenker; stains, hematoxylin and eosin, and Weigert’s copper-
chrome-hematoxylin. The left ovary was lost. The right ovary
shows no abnormality. It contains five corpora lutea. They measure
0.8 mm. x 0.9 mm. in diameter and are in the early lipoid stage, the
condition which exists about the time of the birth of the fetuses. They
still contain numbers of red granules, but the lipoid droplets are very
conspicuous in the cells. In fact, some cells are so full of the lipoid
that the red granules are in evidence only at the periphery of the cell.
Experiment 351-16 (spermophile 371). Captured May 20, 1916.
Weight, 135.5 grams. Operated on May 22, 1916. The left ovary
was removed.
Gross observations. The animal is pregnant, the fetuses measuring
2 cm. in length. The animal gave birth to young, May 31, 1916.
Sacrificed June 17, 1916. Weight, 130.2 grams.
Gross observations. The remaining ovary and uterus appear nor-
mal. There are three spots marking the placental site in the right
horn and three in the left.
Microscopie observations of ovaries. Left ovary: fixative, formalin
zenker, stains hematoxylin and eosin, and Weigert’s copper-chrome-
hematoxylin. Right: ovary: fixative, acetic osmic bichromate; stain,
acid fuchsin and methyl green. The left ovary contains three
corpora lutea. These are in the late red-granule stage. The red
granules are still quite numerous, but some cells contain lipoid drop-
lets about their periphery. The right ovary contains three corpora
lutea of the lipoid stage. It appears very similar to other ovaries
containing luteal bodies of this stage. There is no demonstrable
pathology in the ovary.
Experiment 359-16 (spermophile 379). Captured May 20, 1916.
Weight, 154 grams. Operated on May 22, 1916. The left ovary was
removed.
Gross observations. The animal isin advanced pregnancy. There
was more trauma experienced in this operation than in the one per-
formed on spermophile 377, in which both ovaries were removed. The
animal gave birth to young on May 28, 1916, and kept them until
June 8, 1916.
Sacrificed June 24, 1916. Weight, 136.5 grams.
Gross observations. The placental sites in the uterus are scarcely
visible.
Microscopic observations of the ovaries. Both ovaries: fixative,
formalin zenker; stain, hematoxylin and eosin. The left ovary con-
tains six corpora lutea of the same stage as the left ovary of spermo-
phile 371, the late red granule stage. There are a few lipoid droplets
located through the cells. The right ovary contains four corpora
lutea of the lipoid stage. There is nothing abnormal about it.
156. - DELLA DRIPS
Discussion and summary of results. Three spermophiles were
operated on in this series. The results were positive. The
removal of one ovary during the second half of pregnancy does
not affect in any way the normal development of the fetuses.
They go on to term. There is no effect produced on the re-
maining ovary and the development of rts corpora lutea.
rv
Series 7. Effects on the remaining ovary of the removal of one
ovary and the uterus in pregnant animals
Experiment 273-16 (spermophile 308). Captured May 1, 1916.
Operated on May 4, 1916. The right ovary and the uterus were
removed.
Gross observations. Three are four placental swellings in the right
horn of the uterus and three in the left about 4.5 mm. in length. Both
ovaries show hemorrhagic areas which look like tiny hemorrhagic
cysts. They are more noticeable in the right ovary.
Died May 11, 1916.
Gross observations. Absolutely no cause for death is apparent in
the abdomen. The blood-vessels to the remaining ovary are very
much congested.
Microscopic observations of the ovaries. Both ovaries: fixative,
formalin zenker; stains, hematoxylin and eosin, acid fuchsin and
methyl green, and Weigert’s copper-chrome-hematoxylin. The right
ovary contains five corpora lutea of a very early stage, measuring 0.7
mm.x 0.9mm. The left ovary contains four corpora lutea, measur-
ing 0.8mm.x0.5mm. This organ shows no pathology except in the
corpora lutea. These bodies show many degenerative changes. The
cells have lost their red granules and their nuclei are undergoing chro-
matolysis. The connective tissue has increased greatly.
Experiment 290-16 (spermophile. 325). Captured May 4, 1916.
Weight, 111 grams. Operated on May 6, 1916. The right ovary and
uterus were removed.
Gross observations. The animal is pregnant, the fetuses being just
recognizable.
Sacrificed June 8, 1916.
Microscopic observations of the ovaries. Fixative, formalin zenker.
Stain, acid fuchsin and methyl green. The right ovary contains five
corpora lutea of a very early stage, about 0.5 mm. x 0.6 mm. in diam-
eter. The luteal cells are filled with the red granules. There is a mass
of blood in the center and practically no organization of connective
tissue or blood-vessels. The left ovary contains six corpora lutea of
the late red-granule stage. They have increased in size to 0.9 mm. x
1mm. The normal development of the corpora lutea has apparently
gone on in this ovary, as it compares very favorably with other ovaries
of this date. There is no demonstrable pathology.
THE OVARY OF THE SPERMOPHILE 157
Experiment 279-16 (spermophile 314). Captured May 1, 1916.
Weight, 100 grams. Operated on May 5, 1916. The left ovary and
the uterus were removed.
Gross observations. It is impossible to recognize any fetuses in the
uterus.
Sacrificed May 18, 1916. Weight, 112 grams.
Gross observations. The right ovary appears to contain corpora
lutea. Everything in the abdomen is in excellent condition. There
are no adhesions.
Microscopic observations of the ovaries. Right ovary: fixative,
formalin zenker; stains, acid fuchsin and methyl green, Weigert’s cop-
per-chrome-hematoxylin, and hematoxylin and eosin. The left ovary
was lost. The right ovary shows no pathology. It contains two
corpora lutea which measure about 0.8 mm. x 0.8 mm. They seem
to have reached their normal development for eighteen days. The
number of red granules might have been slightly less than normal, but
they are much more abundant still than the lipoid droplets. There is
a well-developed connective-tissue framework and network of blood-
vessels throughout the luteal body. As there are only two corpora
lutea in this ovary, several follicles have had room to mature. Two of
these show atretic changes.
Experiment 292-16 (spermophile 327). Captured May 4, 1916.
Weight, 130 grams. Operated on May 6, 1916. The right ovary and
uterus were removed.
Gross observations. There are six swellings in the right horn of the
uterus and four in the left, 8.5 mm. in length.
Sacrificed May 31, 1916. Weight, 170 grams.
Gross observations. The left ovary is found embedded in a mass of
fat, but the blood supply to the ovary seems intact.
Microscopic observations of the ovaries. Right ovary; fixative,
formalin zenker; stains, hematoxylin and eosin, Weigert’s copper-
chrome-hematoxylin. Left ovary: fixative, acetic osmic bichromate;
stain, acid fuchsin and methyl green. The right ovary contains six
corpora lutea, measuring 0.6 mm. x 0.7 mm. They are of the red-
granule stage. There is no lipoid. The mass of blood in the center is
fairly well organized. The connective-tissue framework and network
of capillaries are fairly well developed. The left ovary contains six
corpora lutea which measure 0.7 mm. x 1.1 mm. in diameter. They
have developed very normally, although not quite as rapidly as their
controls. The red granules are still very numerous and there is scarcely
any lipoid present. The connective-tissue framework and the blood-
vessel network are very well developed.
Experiment 295-16 (spermophile 330). Captured May 4, 1916.
Weight, 116.5 grams. Operated’‘on May 6, 1916. The right ovary
and uterus were removed.
Gross observations. There are five swellings in each horn, all about
5.5 mm. in length.
Sacrificed May 26, 1916. Weight, 130 grams.
158 DELLA DRIPS
Microscopic observations of the ovaries.. Left ovary: fixative, acetic
osmic bichromate; stain, acid fuchsin and methyl green. The right
ovary was lost. The left ovary contains five corpora lutea, measuring
0.7 mm. x 0.8 mm., and showing many degeneration changes. These
are most apparent in the corpora lutea and the clumps of interstitial
cells of the stroma. The latter cells appear swollen and their nuclei
are undergoing chromatolysis. The cells of the corpora lutea are most
degenerated in the center of the structure. Some cells seem to have
disappeared entirely here, leaving holes. Many are undergoing fatty
changes. Toward the outside of the structure there are cells which are
still in fairly good condition, still containing red granules, though their
nuclei have not taken the stains well, have indented margins and
clumped chromatin strands.
Experiment 308-16 (spermophile 343). Captured May 6, 1916.
Weight, 110 grams. Operated on May 8, 1916. The ovary and the
uterus were removed.
Gross observations: The animal is pregnant. There are eight pla-
cental swellings in the left horn and one in the right, all 6.5 mm. in
length.
Sacrificed May 31, 1916. Weight, 153 grams.
Microscopic observations of the ovaries. Right ovary: fixative,
formalin zenker; stains, hematoxylin and eosin, Weigert’s copper-
chrome-hematoxylin, Mallory’s connective-tissue stain. Left ovary:
fixative, formalin zenker; stains, hematoxylin and eosin, Weigert’s
copper-chrome-hematoxylin. The right ovary contains one corpus
luteum, measuring 0.8 mm. x 0.8 mm., and is of the red-granule stage.
The left ovary shows no apparent pathology. The corpora lutea meas-
ure 1 cm. x 0.9 mm. and are of the early lipoid stage. They have
developed normally.
Discussion and summary of results. In four of these animals
the remaining ovary with its corpora lutea was not at all affected
by the operation. In two of the spermophiles operated on, the
second ovary showed changes. In spermophile 330 the whole
ovary was affected, and in spermophile 308, only the corpora
lutea. There was evidently some injury to the blood supply.
Series 8. Studies on the producion of the-normal bursting of follicles
and the formation of corpora lutea
Experiment 311-16 (spermophile 346). Captured in the spring of
1916. Weight 88 grams. This animal appeared to be in rut on May
7, 1916. She was placed with a male at 10:10 a.m. on this date and
removed at 3 p.m. on May 8, 1916.
Sacrificed May 8, 1916.
~
THE OVARY OF THE SPERMOPHILE 159
Gross observations. The uterus shows no signs of pregnancy.
The ovaries show very small hemorrhagic areas, the smallest noted so
far. In all probability, the animal has ovulated during the time if
was with the male.
Microscopic observations of the ovaries. Fixative, formalin zenker.
Stain, hematoxylin and eosin. There are three corpora lutea in the
right ovary and five in the left. They are in a very early stage of
development, appearing just like other very early ones. There were a
few mitotic figures, one very sure proof of their very recent develop-
ment.
Experiment 323-16 (spermophile 357). Captured April 27, 1916.
Weight, 125 grams. This animal was kept separate from the time
of her capture until May 8, 1916. At 3 p.m. on May 8 a male was
placed with her and kept there until 10:45 a.m. on May 10, 1916.
Sacrificed May 10, 1916, 10:45 a.m.
Gross observations. Gross evidences did not show that the animal
had ovulated.
Microscopie observations of the ovaries. Fixative, formalin zenker.
Stain, hematoxylin and eosin. Both ovaries showed beautiful early
corpora lutea, three in the right ovary and four in the left. These
could not have been more than twenty hours old.
Discussion and summary of results. The results obtained in
these two experiments prove that ovulation follows on coitus
during rut.
Series 9. To determine if ovulation is followed by the formation of
corpora lutea when fertilization is prevented by the resection of
1 cm. of the uterus. One ovary was removed, also, to determine
absolutely whether the animal was pregnant or not.”
Experiment 280-16 (spermophile 315). Captured May 3, 1916.
Weight, 100 grams. Operated on May 5, 1916. The right ovary
was removed and 1 cm. of the uterus just above the body was ligated
and resected.
Gross observations. The animal is not pregnant. She was placed
with a male on May 20, 1916, and remained with him until sacrificed.
Sacrificed June 5, 1916. Weight, 120 grams.
Gross observations. The remaining ovary appears to contain
corpora lutea. There is a good deal of pus about the ligature on the
uterus, with adhesions to the intestines.
Microscopic observations of the ovaries. Fixative, formalin zenker.
Stains, hematoxylin and eosin and Weigert’s copper-chrome-hema-
toxylin. There are no corpora lutea in the right ovary. Several
* This series of experiments is at present being continued. No conclusions
can be drawn from the results obtained so far.
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, NO. 2
160
large mature follicles are present.
in the left ovary.
mm. x 0.7 mm. or 0.7 mm. x 0.7 mm.
DELLA DRIPS
There are six corpora lutea present
They are of a very early stage, and measure 0.6
The cells are full of granules.
There is a core of blood in the center which is undergoing organization.
It certainly appears as if ovulation had taken place shortly before the
animal was sacrificed.
Comparative sizes of uteri
LENGTH | LENGTH SIZE OF WIDTH WIDTH
EXPERI- | SPERMO- DATE OF OF BODY SIZE OF OF OF
MENT PHILE RIGHT LEFT OF CERVIX RIGHT LEFT
HORN HORN UTERUS HORN HORN
cm. cm. mm. mm. mm. mm.
20(-15 | 139 | April 22, 1916 AS 4.5. | 92a |) 2206") ama eaten
42’-15 | 232 | May 8, 1916 5.6 | 5.6 | 4.0 | 1.8 4) O:75)\euee
42°-15 | 233 | May 9, 1916 7.0 | 6.5. | 8.0 9) 430) |) e0/a ieee
437-15 | 242 | May 9, 1916 6.3: | 6.3.) BION e250) soca anon
246-16 | 291 | April 25, 1916 12.4 || 10:31, 2895 1 Sy 0.) . Aa Oana
2°€-16 | 298 | April 29, 1916 7.2 VO 704) 900. |) sO. |) oe0nr aoe
354-16 | 374 | October 20,1916| 6.8 | 6.0 | 35 | 2.0 | 1.0) a0
35-16 | 378 | October 30, 1916 | 6.3 | 6.3 | 45 | 26 | 1.0 eee
657-16 | 462 | October 30,1916] 6.7 | 6.5 | 5.5 | 2.0 | 1.5 | 1.5
30-16 | 344 | May 20, 1916 6.8.9) 7.2 WeSs0). 18:0) 7) eos ae
SUMMARY AND DISCUSSION
The results obtained from the histologic and experimental
investigations carried on may be summarized as follows:
1. In the spermophiles, ovulation occurs only once a year,
during the rutting season in the early spring. Ovulation is
dependent on the stimulus of coitus, for no corpora lutea were
found in the ovaries of animals which were kept from the males.
2. The corpora lutea cannot be responsible for the phenomena
of rut for they are not present in the ovaries at this time.
3. The corpora lutea develop and pass through their normal
cycle after ovulation whether fertilization follows or not (foot-
note 2, p. 159).
4. While the corpora lutea are present in the ovaries, especially
during the two months following parturition, the process of
developing and ripening the follicles is at a standstill.
5. If the uterus is removed after conception, the corpora lutea
do not begin to degenerate, but pass through their normal cycle.
No effects are noted in the ovaries.
THE OVARY OF THE SPERMOPHILE 161
6. Removal of the uterus at any time does not produce
noticeable effects on the ovaries even after a year’s time. ;
7. Double ovariectomy performed at any time during the
period of pregnancy interrupts gestation. If the operation is
performed after a little more than the first half of pregnancy,
the placentas with the fetuses simply degenerate. The invo-
lution of the uterus in these cases is very abnormal. If both
ovaries are removed late in pregnancy, the animal aborts and
the uterus undergoes a much more normal involution. The
removal of only one ovary does not interrupt the pregnancy.
8. Double ovariectomy at any time prevents the recurrence
of the cyclic changes in the uterus and produces an atrophy of
the organ scarcely noticeable within a year.
9. The corpora lutea apparently do not influence the develop-
ment of the mammary gland. When the uterus was removed
very soon after conception, before any signs of pregnancy could
be noted grossly in the uterus, and before any development of
the mammary glands could be noted grossly, the developing
corpora lutea in the ovaries produced no development in the
mammary glands. This would seem to substantiate the work
of Lane-Claypon and Starling who attribute to the fertilized egg
the stimulus for the development of the mammary glands.
10. The corpus luteum of the spermophile derives its elements
from the follicle just as Sobotta states occurs in the mouse. The
luteal cells are the transformed granulosa cells of the follicle.
The connective tissue and vascular network are derived from
the cells of the internal theca which spends itself entirely in
their formation. The capsule of connective tissue surrounding
the luteal structure is the same external theca which surrounded
the follicle. ‘The microscopic pictures of the corpora lutea in
the succeeding stages of their development correspond also to
Sobotta’s descriptions. It may be well to emphasize the com-
plexity of the vascular network throughout the luteal structure
which, when it is completed, brings every luteal cell in intimate
contact with the blood stream.
11. The life cycle of the corpus luteum is made up of three
distinct phases. First, the phase characterized by the presence
162 DELLA DRIPS
of great numbers of red granules in the protoplasm of the luteal
cells. This phase embraces a period dating from the bursting
of the follicle and covering the entire period of pregnancy. From
a point of time very shortly after the bursting, the protoplasm
of the luteal cells shows these red granules which become more
and more abundant until they seem to reach a crisis of abundance
when the organization of the luteal structure is about perfected,
which is not until the placental swellings have reached a length
of about 8.5 cm. From this time on the granules seem very
gradually to decrease in number in the cells until parturition,
when there is a sudden considerable reduction in their number.
Some are found in the cells, however, even as late as the fourth
week after parturition. Second, the phase characterized by the
presence of many lipoid droplets in the protoplasm of the luteal
cells. This phase begins sometime before parturition and lasts
for about six weeks afterward. About the fourteenth day of
pregnancy when the placental swellings in the uterus measure
1.5 em. to 2 em. in length, the lipoid droplets usually begin to
make their appearance at the periphery of the luteal cells next
to the capsule of the corpus luteum. ‘They increase in number
until at the time of parturition, they are quite noticeable in the
cells, being found scattered all through the protoplasm among
the red granules. After parturition, there seems to be a more
rapid increase in the number of lipoid droplets, which coincides
with the sudden decrease in the number of red granules pre-
viously noted. With this increase in lipoid content the cells
which, from the beginning, have been growing constantly larger,
seem to begin to hypertrophy more rapidly. The luteal cells
are largest and contain the greatest amount of lipoid about six
weeks after parturition. In two more weeks practically all the
lipoid has disappeared from the cells and they are beginning
to show evidences of degeneration. Third, the phase of regres-
sion. This period begins about eight weeks after parturition and
lasts for four weeks. By the last of August the corpora lutea
have disappeared from the ovaries. This phase is characterized
by a fatty degeneration of the luteal cells by an increased
vascularization and a connective-tissue invasion.
THE OVARY OF THE SPERMOPHILE 163
From these results, the following conclusions were drawn as to
the functions of the corpora lutea in the ovaries of spermophiles:
The corpora lutea fix the period of estrus by preventing the
development and the ripening of follicles until the time for the
next rutting season is at hand.
The corpus luteum is a gland with two internal secretions,
both of which have specific effects on the uterus, one bringing
about the changes incident to pregnancy and the other effecting
the normal involution of the organ. The first internal secretion
is represented in the luteal cells during the period of pregnancy
by granules which are very similar in their location and staining
reactions to the granules in the A cells of the islands of Langer-
hans, the glands of internal secretion of the pancreas, described
by Bensley. The granules of the luteal cells, however, are
much larger than those of the A cells, being very easily seen
with high powers of the microscope. No mitochondrial granules
or filaments could be observed, perhaps because of the abun-
dance of the granulations in the protoplasm. These luteal cell
eranules are very much like other secretion granules described
by various writers as occurring in the secreting serous cells of
several glands of the body.
The majority of writers have agreed that there is no fatty
product demonstrable in the corpus luteum of several species of
animals and man in the very early stages. They all seem to
have been of the same opinion that the activity of the ovarian
gland of internal secretion begins with the appearance of the
lipoid droplets in the cells. These lipoid droplets were con-
sidered by them to be the evidence of the secretory activity of
the corpus luteum. Its period of activity would then begin
when these droplets begin to appear in the cells, which time
varies with different species, but in all seems to be about the
time of the fixation of the blastocyst. This activity lasts, they
consider, for varying periods in different species. In the rabbit,
Cohn, Fraenkel, and Niskoubina consider that it lasts for nine
or ten days, when regression sets in about the fifteenth day.
Van der Stricht says that in the bat the lipoid droplets are in
much greater abundance during the second half of the period
164 DELLA DRIPS
of pregnancy and that regressive changes do not begin until the
period of pregnancy is over. Miller says there is no neutral fat
in the human corpus luteum until regression sets in at birth.
Because the first-mentioned group of men found that double
ovariectomy did not cause abortion in rabbits after the fifteenth
day, and did so earlier than this, they considered this lipoid
secretion related to changes in the uterus occurring between the
fourth and fifteenth days after coitus, or between the time of
the fixation of the blastocyst and the middle of the period of
pregnancy.
Van der Stricht seems to have been the first to conceive of the
presence of a secretion in the luteal cells prior to the appearance
of the lipoid droplets which coexists with them for some time
after their appearance. He judges of the presence of this
secretion in the cells from its presence in the near-by inter-
cellular spaces and lymphatics. The latter, according to van
der Stricht, are the avenues of excretion of both the serous and
the lipoid secretions.
In spermophiles, the lipoid product does not begin to appear
in the luteal cells until the period of pregnancy is half over and
is not very abundant until after birth. As far as these animals
are concerned, then, the lipoid product is not the active sub-
stance of the corpus luteum which has specific effects on the
uterus during pregnancy. This active substance is rather a
secretion represented in the cells by the secretory granules
previously noted, which are of a very different nature from the
lipoid droplets.
The second internal secretion which is Rea eee in the
luteal cells by lipoid droplets and which formerly has been
considered the secretion which is responsible for the changes
occurring in the uterus incident to pregnancy, must be con-
sidered, as far as the spermophiles are concerned at least, as
having another function. There seems to be some relationship
in these animals between the period of greatest abundance of
the lipoid product in the cells and the period of regression and
atrophy in the uterus. The uterus of the spermophile atrophies
very slowly, much more so than in animals that bear several
THE OVARY OF THE SPERMOPHILE 165
litters of young every year. The atrophy is not completed until
six or seven weeks after parturition, about the time when the
lipoid product reaches its crisis of abundance in the cells and
begins to disappear. Another result which substantiates the
theory that the lipoid secretion brings about the normal invo-
lution of the uterus in the very abnormal, even pathologic process
which goes on in the uterus following the removal of both ovaries
during all but the more advanced stages of pregnancy. During
the first half of the period of pregnancy there is no lipoid in the
corpora lutea, which, according to this theory, would account
for the pathology in the uterus following double ovariectomy.
If the ovariectomy is performed late in pregnancy after the
lipoid droplets have become quite abundant in the luteal cells,
the animal aborts and the uterus undergoes an involution more
nearly like the normal, due to the specific effect of the lipoid
secretion which is already present in the circulation. Mulon
thought the lipoid of the corpus luteum had an antitoxic action
toward the poisons elaborated in the development of the fetuses.
It would seem more reasonable to suppose that it neutralizes
the toxic products produced in normal involution, which would
be only a part of its function as a specific agent in effecting
this normal involution of the uterus.
It may be added, in closing, that the two luteal secretions are
undoubtedly emptied into the blood stream in these animals.
An observation of the elaborate capillary network of these
structures could lead to no other conclusion. Lymphatic sinuses
are demonstrable in the corpora lutea, but they are found only
near the capsule in the proximity of the larger blood-vessels.
There is no anatomical evidence for concluding that the secre-
tions are carried away by the lymphatics.
The writer wishes to express her appreciation for the valuable
aid given her by Dr. Frank C. Mann, head of the Division of
Experimental Surgery and Pathology of the Mayo Clinic, under
whose direction this work was done.
166 DELLA DRIPS
LITERATURE CITED
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1908 Sur le follicule de De Graaf mur et la formation du corps jaune
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1909 Sur les homologies et la signification des glandes a secretion
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1916 The normal mode of secretion in the thyroid gland. Am. Jour.
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Bonp, C. J. 1906 An inquiry into some points in uterine and ovarian phys-
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CaRMICHEL, E. 8., anp Marsuatu, F. H. 1907 The correlation of the ovarian
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CHALFANT, S. A. 1915 Subcutaneous transplantation of ovarian tissue, report
of thirty-two cases with special reference to its effects on the meno-
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CHAUFFARD, A., LARocHE, G., AND Gricaut, ‘A. 1912 Function cholesterini-
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Cuark, J.G. 1898-99 The origin, growth and fate of the corpus luteum. John
Hopkins Hospital Reports, vol. 7, pp. 181-221.
Coun, F. 1903 Zur Histologie und Histogenese des Corpus luteum und des
interstitiellen Ovarialgewebes. Arch.f. mikr. Anat., Bd. 62, 8, 745-
Tie.
Corner, G. W. 1915 The corpus luteum of pregnancy in swine. Carnegie
Institution, Washington, Publication no. 222, pp. 69-94.
CuLBRETSON, C. 1916 A study of the menopause. Surg., Gynec, and Obst.,
vol. 23, pp. 667-685.
Darts, F. 1908 On the relations between the ovaries and the uterus. Surg.,
Gynec. and Obst., vol. 6, pp. 153-160.
DannreEvTHER, W. T. 1914 Corpus luteum organotherapy in clinical practice.
Jour. Am. Med. Assn., vol. 62, pp. 359-362.
THE OVARY OF THE SPERMOPHILE 167
De Lez, J.B. 1916 Auto-transplantation of the corpusluteum. Surg., Gynec.
and Obst., vol. 22, pp. 228-231.
FRAENKEL, L. 1903 Die Function des Corpus luteum. Arch. f. Gynak., Bd.
68, S. 438-545.
Frank, R. T., Aanp RosenBiLooM, J. 1915 Physiologically active substances
contained in the placenta and in the corpus luteum. Surg., Gynec.
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Graves, W. P. 1913 Influence of the ovary as an organ of internal secretion.
Am. Jour. Obst., vol. 67, pp. 649-665, 779-785.
Hare, H. A. 1912 Therapeutic application of ductless glands. Am. Jour.
Obst., vol. 64, pp. 514-518.
Harrower, H.R. 1916 Relation of the internal secretions to neurasthenia in
women. Am. Jour. Obst., vol. 73, pp. 630-637.
Hearst, W. 1906 The source of the stimulus which causes the development of
the mammary gland and the secretion of milk. Am. Jour. Physiol.,
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Hirst, J.C. 1916 The control of disagreeable symptoms of the surgical meno-
pause by the hypodermic intramuscular administration of corpus
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Iscovesco, H. 1912 Les lipoides de l’ovaire. Compt. rend. de la Soc. biol.,
T. 73, pp. 16-18, 189-191.
JANKowskKI, J. 1904 Beitrag zur Entstehung des Corpus luteum der Siuge-
tiere. Arch. f. mikr. Anat., Bd. 64, S. 361-389.
Lane-Criaypon, J. E., AND StaruinG, E. H. 1906 An experimental inquiry
into the factors which determine the growth and activity of the mam-
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Lane-Crayron, J. E. 1906 On the origin and life history of the interstitial
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vol. 77, pp. 32-58.
LetcutTon, A. P. 1915 The use of corpus luteum extract in the treatment of
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Lewis, M. R., ann WarreN, H. 1914-15 Mitochondria in tissue cultures.
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168 DELLA DRIPS
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Fig. 4 Portion of a section of the suprarenal gland of an adult albino rat
(F 3.2), after nine days of acute inanition. Formalin fixation; frozen section
stained with Herxheimer’s scarlet red. For explanations, see figure 3. No
apparent decrease in the liposomes, which appear more uniformly distributed
throughout the cortex. The light band at the transition between outer and
middle zones is obscured. X 90.
Fig. 5 Portion of a section of the suprarenal gland of an adult albino rat
(F 4.2) after twelve days of acute inanition. Formalin fixation; frozen section
stained with Herxheimer’s scarlet red. The liposomes persist in the outer zone
(O), but have nearly all disappeared elsewhere. A few are barely visible in the
outer half of the middle zone (Mo), and in the inner zone (J). X 90.
Fig. 6 Portion of a section of the suprarenal gland of a normal albino rat
(J 1.1) three weeks old. Zenker fixation, paraffin section stained with hema-
toxylin-eosin. Most of the middle cortical zone is omitted. F, fibrous capsule;
O, outer zone (glomerulosa), one cell in mitosis; 7’, transition band, nearly lipoid-
free; Mo, outer part, and M7, inner part of middle zone (fasciculata); 7, inner
zone (reticularis); M, medulla. X 300.
Fig. 7 Portion of a section of the suprarenal gland of an albino rat (S 7.31)
held at constant body weight by underfeeding from three to ten weeks of age.
For explanations, see figure 6. Progressive differentiation of lipoidal vacuoles
in the outer zone (O) and of pigment (P) in the inner zone (J). Some cell
atrophy, with hyperemia and increased degeneration in the inner zone. X 300.
Fig. 8 Portion of a section of the suprarenal gland of a normal albino rat
(St 7.45) ten weeks of age. For explanations, see figure 6. This represents the
normal adult structure. Lipoidal vacuoles well marked in the outer zone (OQ)
and outer part of the middle zone (Mo), but rare in the transition band (7).
One pigment mass (P) visible. > 300.
Fig. 9 Portion of a section of the suprarenal gland of an adult albino rat
(J 1.5) subjected to chronic inanition for five weeks. For explanations, see
figure 6. The lipoidal vacuoles have disappeared, except in the outer zone
(O). Marked atrophy with nuclear and cytoplasmic degeneration in the inner
cortical zone (J). Three pigment masses (P) visible. X 300.
Fig. 10 Longitudinal section of the suprarenal gland in an adult albino rat
(F 9.3) after ten days of acute inanition. Fixation in Miiller’s fluid; frozen
section (unstained). Normal chromaffin reaction in the medulla, which in this
case extends to the surface at the hilus. X 80.
253
254 Cc. M. JACKSON
In a rat at seventeen days (F 7.1) the fresh frozen sections
of the formalin-hardened gland show the characteristic cor-
tical opacity due to the liposomes. The clear line between the
outer and middle zones is distinct. The opacity decreases
greatly in the inner half of the middle zone and in the inner
zone, the medulla being perfectly clear.
This irregular distribution of the liposomes is still more ap-
parent when the sections are stained with scarlet red or osmic
acid. The liposomes are most abundant in the outer zone and
the outer half of the middle zone, where the largest droplets
now approach nuclear size. Toward the inner cortical zone
they become fewer and finer. Some of the inner zone cells are
entirely free from liposomes, others occasionally appear well
filled. Aside from a few scattered cortical cell islands, no lipo-
somes appear in the medulla.
The other supararenal gland from this rat was hardened in
Miiller’s fluid. The chromaffin reaction of the medulla is more
definite than in the earlier stages, though not so intense as later.
All of the medulla parenchyma cells are stained light brown,
the nucleus darker than the cytoplasm. The vacuoles of va-
rious size in the medulla cells are unstained, not giving the chro-
maffin reaction. The blood in the vascular spaces of the medulla
occasionally presents a reddish-brown color to a variable extent
(probably from absorbed epinephrin).
At three weeks. The paraffin sections were stained as usual.
The outer cortical zone (fig. 6, O) is narrow, usually 6 to 8 (rarely
12) cells deep. The cells are in irregular masses or columns,
separated by blood capillaries. Cell boundaries are ill defined.
The cytoplasm is scanty, contains fine eosinophile granules
and more numerous lipoidal vacuoles. The nuclei are hyper-
chromatic, some almost pyenotic. The deepest cells (fig. 6, T)
are transitional to the middle zone, and form a narrow zone
nearly free from lipoidal vacuoles.
The middle cortical zone is broadest. The cells (fig. 6, Mo,
M71) are arranged in very distinct cell columns, radially arranged,
and usually but one cell wide. The radial cell columns are
separated by blood capillaries, which become wider toward the
SUPRARENAL GLAND—EFFECTS OF INANITION 255
center of the gland. Cell boundaries are more distinct than
heretofore. The relatively abundant cyoplasm is filled with
characteristic eosinophile granules and a variable number of
lipoidal vacuoles. In some cases these vacuoles appear no
more numerous than in the second week. In others they are
more abundant especially in the outer half of the middle zone,
whereby these cells become much larger than the cells of the
inner half of the middle zone. The nuclei are typical in struc-
ture, spherical, and moderately rich in chromatin. Atrophic
cells occasionally occur in various stages of degeneration.
The inner zone (zona reticularis, fig. 6, 7) is rather narrow,
but of variable width, representing the area of irregular cell
columns next to the medulla. The irregularity of structure is
probably associated with the process of absorption accompany-
ing the expansion of the medulla, although only occasionally
are the border cells flattened as though atrophic from pressure.
Most of the cells in the inner zone are similar to those of the
adjacent middle zone, the cytoplasm containing eosinophile
granules and a few small lipoidal vacuoles. Some scattered
cells show various stages of degeneration and disintegration. Oc-
casional islets of such cortial cells occur also in the adjacent
medulla, but rarely deeper, toward the center of the medulla,
as in the earlier stages.
The medulla in stained sections (fig. 6, M) appears very light,
in’ strong contrast with the darker cortex. (The converse is
true if the suprarenal gland has been fixed in Zenker-formol
instead of Zenker’s fluid, the sections being stained with hema-
toxylin.) The parenchyma forms irregular cell masses, sepa-
rated by delicate fibrous stroma (with elongated nuclei) enclos- ‘
ing wide, sinusoidal blood-vessels. The cytoplasm of the
parenchyma cells is abundant, containing pale violet (chromaffin?)
granules and numerous non-lipoidal vacuoles, variable in size
and number. The nuclei are typically vesicular in form and
only moderately chromatic. A few are smaller and more deeply
staining, sometimes pycnotic. Degenerative cells with karyo-
lytic nuclei are rarely seen. Occasional large spherical sym-
pathetic ganglion cells and bundles of non-medullated fibers
appear.
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, No. 3
256 Cc. M. JACKSON
At eight weeks. The appearance of hematoxylin-stained sec-
tions under low power is similar to that at three weeks. The
cell structure in the outer and middle zones is also similar, with
some increase in the size and number of lipoidal vacuoles, es-
pecially in the outer half of the middle zone. The inner zone
shows in general a more atrophic appearance, and occasional
pigment cells occur for the first time. These are parenchyma
cells containing one (sometimes two) irregularly spheroidal,
light yellowish-brown, coarsely granular masses. As will be
shown later, this pigment is probably a lipochrome. The
nuclei of these pigment cells are irregular, sometimes central,
sometimes crowded to the side of the cell and flattened. The
inner zone cells bordering on the medulla are usually more or
less flattened and atrophic in appearance. The medulla ap-
pears similar to that at three weeks.
At ten weeks to adult. At ten weeks, the histological struc-
ture of the suprarenal (fig. 8) has reached practically the adult
condition. In fact, although the gland increases in size, but
few changes in structure are noticeable after the eight weeks’
stage previously described. In the usual hematoxylin-stained
sections the structure appears as follows (fig. 8):
The outer cortical zone (fig. 8. O) remains narrow, 6 to 12
cells deep. It is somewhat variable in structure, with irregular
cell cords separated by blood capillaries. The cells remain
relatively small, with nuclei of variable form and structure.
The cytoplasm is relatively scanty and somewhat granular. It
contains a variable amount of lipoidal vacuoles, usually giving
the cytoplasm a honey-comb appearance. The deepest cells,
on the border between the outer and middle zones (fig. 8, 7),
are relatively free from lipoidal vacuoles as found in the earlier
stages.
In the middle zone the only noteworthy change is in the
amount of lipoids, as shown by the vacuoles. These increase,
but to a variable extent. They are larger and more numerous
in the outer half of the zone (fig. 8, Mo), but smaller and fewer
in the inner half. In the larger cells of the outer part, the cyto-
plasm presents a reticulated (honey-comb) appearance, like
SUPRARENAL GLAND—EFFECTS OF INANITION 257
that of sebaceous gland cells. In cells with fewer lipoidal vac-
uoles, the eosinophile granules are more conspicuous. Occa-
sionally basophilic granules also occur. These are especially
evident in preparations stained with iron-hematoxylin, and prob-
ably in part correspond to the ‘corps sidérophiles’ or the mito-
chondria described by various French investigators. The nuclei
are usually central in position, spherical, and typical in structure.
A few degenerative cells occur as heretofore.
The inner zone (fig. 8, J) appears in general similar to that at
eight weeks, the outer cells of the zone being transitional to the
middle zone and the inner cells (next to the medulla) being typi-
cally more or less atrophic in structure. Atrophic or degenerative
changes have likewise been described in the inner cortical zone
of various animals (including the rat) by Gottschau (’83), Pfaund-
ler (92) Soulié (’03) and others. According to Kolmer (12 a,
12 b) these degenerative changes are increased by pregnancy
(guinea-pig).
Pigment cells in the inner cortical zone of the rat at ten w pole
occur more frequently than at eight weeks, but in variable de-
gree. They are somewhat variable in number, often numerous,
especially in the older rats, and may even extend somewhat into
the middle zone. The nuclei of the pigment cells may be central
in position, surrounded by the light yellowish-brown or greenish
yellow granular masses. In this case the nucleus is irregular in
form, but it is often pushed aside and flattened. The pigment
is still visible, though inconspicuous, in stained paraffin sections.
In thin sections the pigmented substance frequently appears
vacuolated (fig. 8, P). In fresh frozen sections the pigment cells
are clearly visible, and they are deeply stained by osmic acid or
scarlet red. This would indicate that the pigment is a lipochrome,
probably derived by a transformation of lipoids or other fatty
substances. The vacuoles probably represent untransformed
lipoids, observed by Ewald (’02) and Da Costa (’13).
The cortical fats (lipoids) in general are best studied in frozen
sections of suprarenals, either fresh or hardened a few hours in
formalin. In the unstained sections the distribution of the lipo-
somes is clearly evident on account of the varying degrees of
258 Cc. M. JACKSON
opacity caused by them. The extracapsular fat droplets, on the
other hand, appear clear and light. The liposomes present the
same reactions mentioned for the new-born.
The amount and distribution of the liposomes are most élearly
shown in the frozen sections stained with scarlet red (fig. 4). In
the narrow outer zone, they are usually very abundant and closely
packed, the largest droplets somewhat exceeding the average
nuclear size. The subjacent clear line or narrow band is rela-
tively (but not absolutely) free from liposomes. They are most
abundant and closely packed in the outer half of the middle zone,
where the largest may reach double the average nuclear diameter.
In the inner half of the middle zone they become more or less
reduced in amount, and are usually scanty in the inner cortical
zone (aside from those associated with the pigment cells). No
liposomes are present in the medulla, aside from occasional groups
which appear in the cortical islands near the boundary.
. The distribution of liposomes in sections stained with osmic
acid corresponds closely with that in sections stained with scarlet
red. Thestriking contrast in the staining reactions of the cortical
lipoids and the extracapsular ordinary fat was mentioned in the
new-born suprarenal, and is constant. The pigment cells (lipo-
chrome) of the inner cortical zone frequently react more like the
ordinary fat, however, staining a lighter reddish than the lipo-
somes with the scarlet red and darker than the liposomes in prep-
arations stained with osmic acid.
The amount and distribution of lipoids, as is well known, vary
much according to age and species. In general, the formation of
lipoids begins during the prenatal period (Poll, ’05; Starkel and
Weegrzynowski, 710), and increases during postnatal life, though a
decrease in the human suprarenal fat after childhood is noted by
Napp (’05). As to species the lipoidal content appears greatest
in man, carnivora, and rodents, less in ruminants and pachy-
derms (Ciaccio, ’10, and others). The lipoids are usually, as in
the rat, most abundant in the middle zone, with less in the outer
and inner zones. In some cases, however, a greater abundance
of lipoids of the outer zone has been observed (Hultgren and An-
derson, 99; Ewald ’02; Napp, ’05; Bonnamour, ’05 b; Starkel and
SUPRARENAL GLAND—EFFECTS OF INANITION 259
Weerzynowski,’10) ; and in the sheep, according to Mayer, André,
Mulon and Schaeffer (’12), the sparse lipoidal granules are lo-
cated exclusively in the outer zone. The characteristic fat-free
band between the zona glomerulosa and the fasciculata was
noted in the rat by Bonnamour (’05 b). Babes and Joneseo (’08)
likewise noted a scarcity in fat between these zones in the dog,
but Mulon (712), on the contrary, finds larger droplets at this
level. The composition of the suprarenal liposomes is generally
considered to be chiefly lecithin or cholesterin with a variable
admixture of ordinary fats (compare Ciaccio, 710, and Kawa-
mura, 711).
Pigment has often been described in the inner cortical zone of
the guinea-pig (even in the albino, according to Mulon) and man,
but less frequently in other forms. It was noted by Gottschau
(83) and Pfaundler (92) in most mammals, Baroncini and
Beretta (’01) in bats, and by Elliott and Tuckett (’06) sparsely
in the duckbill and pigeon. Bonnamour (’05 a) found it variable
in the dog and horse, and absent in the cat, rabbit, and marmot.
He found it rarely inthe rat, with no difference in pigment be-
tween white and black rats. DaCosta (713) also noted a few
pigment cells in the zona reticularis of Mus decumanus. Dewit-
ky (12) likewise noted brownish pigment in the cortex of the
rat at seven weeks. - Delamere (’03) and others have noted that
the amount of pigment usually increases with age, being rare or
absent in the young. Mulon (’02, ’03 a), Ciaccio (’05), Kolmer
(712 b), and others believe that the pigment formation may rep-
resent a secretion with physiological significance, and an in-
crease during pregnancy (guinea-pig) is claimed by Kolmer (12 a).
As to its composition, this pigment has usually been considered
as related to fat (lipochrome), although Starkel and Wegrzynow-
ski (710) and Thomas (’11) consider that the pigment appearing
in the degenerating inner cortical zone of the suprarenal in the
human new-born is of hemal origin and different from the pig-
ment in the adult gland.
The suprarenal medulla in the rat at ten weeks and older may
extend to the surface at the hilus (fig. 10), as found by Poll
(99). It is similar in structure to that described in earlier stages.
260 Cc. M. JACKSON
The stroma (also that in the inner cortical zone) stains indistinctly
bluish with Mallory’s anilin-blue connective-tissue stain. The
parenchyma cells (fig. 8, M) are large. With Zenker’s fixation
and hematoxylin-eosin stain, the cytoplasm contains the usual
faintly basophilic (chromaffin?) granules and occasionally a few
spherical eosinophile bodies of variable size. The cell periphery
frequently contains irregular vacuoles, non-lipoidal and of un-
known significance. They are variable in size and are somewhat
more prominent than in earlier stages. According to Ciaccio
(05), they are acidophile in reaction, like true nucleoli, and
are comparable to ‘plasmosomes.’ The structures described by
Ciaccio probably correspond to the eosinophile bodies referred
to above, and not to the characteristic vacuoles. The nuclei, as
heretofore, are typically vesicular, though a few of them are
small and hyperchromatic (sometimes pycnotic). The blood sin-
uses, sympathetic ganglion cells, and occasional atrophic cortical
islands occur as heretofore.
In preparations fixed in Miiller’s fluid, the chromaffin reaction
of the medulla is always well marked (as in fig. 10). The paren-
chyma cells appear as brownish masses separated by the un-
stained vascular areas. There is some variation in the intensity
of the reaction in different individuals, although the variation is
not great in sections of the same thickness. It also usually ap-
pears fairly uniform throughout the medulla, all of the paren-
chyma cells being somewhat similar in their reaction. Some
masses or clumps of cells may stain more deeply, however. The
brownish color appears in the cytoplasm, which may appear
homogeneous or granular (the granules being most distinct in
very thin sections). The cytoplasmic vacuoles of various size
remain perfectly clear and unstained. Whatever their nature,
they evidently do not contain epinephrin. The nucleus gives the
chromaffin reaction, being stained slightly darker than the cyto-
plasm. This was noted by Dostoiewsky (’86) confirming Henle
(versus v. Brunn). According to Ciaccio (’05), Diamare claims
that the chromaffin substance fills the whole cell, while Grynfeld,
localizes it in the cytoplasmic granules. The reaction is weak-
ened or lost a few hours post mortem (Dostoiewsky, ’86; Ciaccio.
SUPRARENAL GLAND—EFFECTS OF INANITION 261
705, and others), and is also affected by anesthetics (Schur and
Wiesel, 08; Hornowski, 09). The chromaffin reaction sometimes
appears also to a variable extent within the blood-vessels of the
medulla of the rat, as has often been noted in other animals by
various observers.
2. Changes in young rats stunted by underfeeding
Underfed from birth. In arat (St 80.9) underfed from birth to
twelve days and reaching a body weight of 8.9 grams, the supra-
renal gland has increased in weight to 0.0026 gram, which is about
normal for seven days of age. The normal differentiation of
the suprarenal cortex and medulla has occurred, and the struc-
ture (in sections stained with hematoxylin) is essentially similar
to that of the normal rat at seven days. Thus the histological
differentiation has continued, as in a norma! gland of correspond-
ing weight, although( as previously shown) the rate of mitosis has
been greatly diminished.
In a rat (St 247.5) underfed from birth to seven days, there is
apparently no increase in the weight of the suprarenal. Frozen
sections stained with scarlet red and osmic acid show the cortical
liposomes somewhat similar in appearance to those in the new-
born, but progressive absorption in the cortical cell strands of the
medulla is evident. The chromaffin reaction is normal.
In another rat (St 228.4) underfed from birth to fifty-eight
days, the suprarenal gland has increased markedly in weight and
shows a progressive differentiation of liposomes similar to those
in a normal gland of similar weight (F 7.1). The chromaffin re-
action in the medulla is variable. Some cell masses show a very
definite reaction. In others it is entirely absent, giving the
medulla a spotted appearance. This irregularity of the chro-
maffin reaction is probably due to the inanition, as it is not ob-
served in the normal animals.
Rats at maintenance from three weeks to eight, ten, or twelve weeks
of age. In these underfed young rats the body weight and supra-
renal weight (table 1) are but little above the initial weight at
three weeks. In general, the stained sections (Zenker’s fixation,
262 Cc. M. JACKSON
hematoxylin-eosin stain) of the suprarenal appear similar to
those normal at three weeks, though certain changes are clearly
evident (fig. 7).
The vacuoles in the outer-zone cells usually indicate a progres-
sive lipoidal differentiation (fig. 7, O). While in a few cases the
lipoidal vacuoles appear scanty (as normally at three weeks), in
others the outer lipoid zone was as distinct and clearly differen-
tiated as normally at ten weeks. In most cases the lipoidal
content appears somewhere between these two extremes. The
eosinophile cytoplasmic granules are somewhat indistinct.
The lipoidal vacuoles of the middle zone were also found in-
creased in size and number in most cases, though more variably
and not so definitely as in the outer zone. As in normal differen-
tiation, the lipoids usually accumulate to a greater extent in the
outer half of the middle zone. Otherwise there is no constant
change to be noted, excepting apparently a somewhat greater
number of degenerating cells than occurs normally.
The inner zone (fig. 7, 7), however, when compared with the
normal shows the most conspicuous changes. This zone is con-
stantly hyperemic and very markedly atrophic. The cells in
general appear greatly decreased in size, and many show various
stages of nuclear and cytoplasmic degeneration. More cells are
flattened at the medullary border than occurs normally. The
nuclei are frequently pycnotic or karyolytic. Pigment masses
(not present at three weeks) appear (fig. 7, P) and are even more
abundant than in the normal gland at ten weeks.
The medulla (fig. 7, 1) shows relatively less change than any
other part of the suprarenal. In most cases it resembles closely
the normal at three weeks, though in some cases there is an evi-
dent increase in the proportion of smaller, hyperchromatic (oc-
casionally pycnotic or karyolytic) nuclei. The cytoplasmic gran-
ules are usually indistinct. No chromaffin tests were made in
these cases.
Maintenance from three to fifteen or twenty weeks of age. In
these two young rats (St 33.1 and St 38.8) held at maintenance
for unusually long periods, the changes in the suprarenal are
more pronounced. The glands were fixed in Zenker’s fluid, sec-
SUPRARENAL GLAND—EFFECTS OF INANITION 263
tioned and stained as usual. In general, the nuclei appear more
hyperchromatic, though not much more than in the normal
gland at three weeks. Although the fat in the fibrous capsule
around the gland has apparently almost entirely disappeared,
the lipoidal vacuoles of the outer zone of the cortex are abundant.
In the middle zone, however, they are apparently not more num-
erous than in the normal at three weeks. In the inner part of
the middle zone many cells are degenerating some in various
stages of disintegration. The atrophic degeneration and pig-
ment formation in the inner cortical zone is very marked. The
greenish-yellow, vacuolated pigment masses in many cases have
entirely filled the cells and are quite numerous. The medulla
cells show increased vacuolization, sparser granulation, and hy-
perchromatic (frequently pyenotic, some karyolytic) nuclei. In
general, however, the structure is less changed than in the cortex.
3. Changes in young rats refed after stunting by underfeeding
In the rats refed fully after maintenance from three to twelve
weeks of age, the suprarenal gland rapidly returns to approxi-
mately normal structure. Even after one week of refeeding the
cortex appears distinctly lighter in stained sections, due to a de-
crease in the chromatin content of the nuclei. The lipoids ac-
cumulate more abundantly (especially in the outer half of the
middle zone) and the inner cortical zone becomes somewhat less
atrophic in appearance. By the end of two weeks of refeeding,
the gland has reached nearly normal structure, although some
areas of degeneration may persist for longer periods. Even those
held at maintenance up to twenty weeks of age (S 33.118 and $
33.120), with permanently stunted body weight, show practi-
cally normal structure in the suprarenals after being refed fully up
to about one year of age.
4. Changes in adult rats after acute or chronic inanition
Adult acute inanition. Adult rats were given water only for
seven to twelve days, with loss in body weight of 29 to 45 per
cent (table 1 D). The suprarenal glands, as previously shown
264. Cc. M. JACKSON
(Jackson, 715 a), lose but little in absolute weight during adult
inanition. The cells (especially those of the middle cortical zone
and the medulla) during acute inanition apparently diminish in
size, however, as shown in table 4. The decrease in cell size is
counterbalanced by hyperemia of the cortex, though apparently
the reverse (decrease in proportion of vascular stroma) occurs in
the medulla.
Sections of the suprarenal, stained with hematoxylin-eosin,
show changes in histological structure. The cytoplasm is re-
duced in amount and the eosinophile granules usually indistinct.
The nuclei are variable, frequently hypochromatic. More fre-
quently, however, they appear hyperchromatic, though less so in
the outer half of the middle zone. Pycnosis and deformity of
the nuclei are frequent, as described by Bonnamour (’05b) in
the outer cortical zone of the starved rat. The lipoidal vacuoles
are variable. Usually, however, they are absent or considerably
reduced in number and size in the middle zone, though persisting
nearly unchanged in the outer zone. The eosinophile granules
of the cortical cells, though sometimes indistinct, are often well
preserved and become more evident with the reduction in the
amount of lipoids present.
The inner cortical zone shows the hyperemia and atrophic con-
dition usually more pronounced than in the normal animal.
Pigment masses are frequent, but it is somewhat doubtful whether
they are increased in number. Degenerating cells with pycnotic
or karyolytic nuclei in various stages of disintegration are
numerous.
Scarlet red or osmic staining of fresh frozen sections reveals
_the cortical liposomes much more abundant than would be sus-
pected from the ordinary stained paraffin sections. In some
cases the usual fat-free boundary line between the outer and mid-
dle zones is nearly obliterated by an increased development of
liposomes, and they may appear more uniformly . scattered
through the middle zone, and even the inner zone (fig. 4).
This tendency toward a more uniform distribution of the lipo-
somes throughout the cortex was noted in three of the eight cases
of adult acute inanition stained especially for lipoids, though the
SUPRARENAL GLAND—EFFECTS OF INANITION 265
total amount of the lipoids appears somewhat reduced in one of
them (F 1.2). A similar ‘Verbreitung der lipoidhaltigen Schich-
ten’ was observed by Landau (713 b) in fasting guinea-pigs.
In five of the eight rats in my series, however, the change con-
sists In a very marked reduction in the liposomes of the middle
and inner zones. Under the low power of the microscope only a
few liposomes may remain visible in the outer half of the middle
zone, the remainder of the middle and inner zones being appar-
ently free from them (fig. 5). Under higher power, however,
numerous extremely fine granules are often still visible in all these
cortical cells. The effect in such cases is therefore apparently
merely to reduce the liposomes very muchinsize. In other cases,
the liposomes have entirely disappeared. The pigment cells
near the corticomedullary zone apparently persist nearly un-
changed, contrary to Rondoni and Montagnani (’15), who found
a decrease in fasting guinea-pigs.
In striking contrast with the remainder of the cortex is the
narrow outer zone, in which the liposomes persist with great
tenacity (fig. 5). With the scarlet stain, this zone therefore ap-
pears nearly unchanged as a deep reddish band, while the re-
mainder of the cortex appears pale, nearly colorless, or with a
pale, diffuse reddish color suggesting some fat-like substance in
solution. Osmic preparations give corresponding pictures. In
only one case (F 9.3) the outer zone in places appears somewhat
broken and irregular. The tenacity of the outer lipoidal zone is
not due to any visible difference in the structure or composition
of the liposomes in this zone. Landau (13 b), however, finds
that in the fasting guinea-pig, although there is no decrease in
the total lipoid content, the cholesterin decreases in all but the
outer cortical zone. It may be dependent upon the vascular
arrangement, as all of the blood to the cortex passes first through
the capillaries of this zone (Flint, ’00).
The amount of decrease in the liposomes of the suprarenal cor-
tex is usually, but not always, somewhat proportional to the
length of the fasting period or to the loss in body weight. In
general, the lipoids appear much more resistant to inanition than
does the ordinary fat just outside the suprarenal capsule.
266 Cc. M. JACKSON
The suprarenal medulla is often considerably affected during
acute inanition, though usually less so than is the cortex. The
‘eytoplasm frequently becomes more vacuolated, although the
characteristic (chromaffin?) granules persist in the parenchyma
cells. The nuclei may become more hyperchromatic, and a
larger proportion are pycnotic. In many cases the nuclei are
hypochromatic, undergoing chromatolysis. Some areas of
marked cellular degeneration occur.
The chromaffin reaction is apparently but slightly Gf at all)
affected by the inanition experiments. The slight variations in
the intensity of the brown color are apparently no greater than
appear in the normal controls. Even in the rat starved twelve
days with loss of 45 per cent in body weight (F 5.2) the chromaffin
reaction of the medulla appears fully as intense as in any of the
controls. This rat was still active when killed, but the reaction
persists in another (F 9.3) which was killed while very weak and
near death with a loss of 33 per cent in body weight after ten
days of acute inanition (fig. 10). .
In only one case (F 6.2) is there a marked decrease, only traces
of the chromaffin reaction being present. This was probably a
postmortem change, as the rat was found dead. It had lost only
34 per cent in body weight after seven days of inanition. It
therefore appears that in adult rats acute inanition produces no
appreciable decrease in the chromaffin reaction of the suprarenal
medulla, even (aside from postmortem changes) in those starved
to death.
Adult chronic inanition. In six adult rats underfed thirty to
thirty-five days with gradual loss in body weight amounting to
33 to 38 per cent, the histological changes (in the ordinary prepa-
rations) were found in general very similar to those after acute
inanition (fig. 9). These include a general atrophy of cells and
nuclei with a variable reduction in the amount of lipoids in the
middle (but not in the outer) zone (fig. 9, O). There is similarly
a pronounced cellular atrophy in the inner zone, and in one case
(J 1.5, fig. 9, J) the vacuolated cytoplasm in a few places had ap-
parently disintegrated to form extensive intercellular spaces, sim-
ilar to the condition described in the human suprarenal by Meyer
CZ):
SUPRARENAL GLAND—EFFECTS OF INANITION 267
The cells usually undergo simple atrophy, however. Many of
the nuclei appear chromatolytic, some pyecnotic. There is ap-
parently no change in the number and appearance of the pigment
cells, as a rule, although sometimes they appear increased in
number (fig. 9, P). The changes in the medulla are similar to
those noted under acute inanition. The special lipoid and
chromaffin stains were not employed in the chronic-inanition
series.
From the foregoing it appears that during inanition in the al-
bino rat the behavior of the suprarenal lipoids is somewhat vari-
able according to ircumstances. In young rats severely stunted
by underfeeding there is no apparent loss, the liposomes continu-
ing to differentiate as in the normal gland of corresponding
weight. Likewise in older rats there is sometimes no evident
decrease, but instead a tendency to more uniform distribution of
the liposomes throughout the cortex. This appears characteris-
tic where the inanition has not been carried to extremes. No de-
crease in suprarenal fat (lipoids) during inanition has also been
observed by Frederici (’03) in the guinea-pig, Traina (’04) in
man and rabbit, Napp (’05) in man, Kawamura (’11) in man,
Landau (713 b) in man, cat and guinea-pig. Bonnamour (’05 b)
even finds an increase in the starved rat, cat, rabbit, and guinea-
pig. Ciaccio (according to Landau, 713 b) during inanition found
first an increase, with subsequent decrease, in the suprarenal fat
content.
In most cases of acute and chronic inanition in the rat, espe-
cially in those carried to extremes, there is awell-marked decrease
in the suprarenal lipoids, although they are retained in the outer
cortical zone with remarkable tenacity in all cases. A decrease
in the fat (lipoids) of the suprarenal during inanition has been
observed by Orth (’93) in man, Beneke in man and animals,
Ewald (02) in the rabbit, Herman (’05) in man, Ponomarew
(14) in the mouse, and Rondoni and Montagnani (715) in the
guinea-pig. In the latter two investigations, more or less reten-
tion of fat in the zona glomerulosa (of mice and guinea-pigs) is
mentioned, which would be in agreement with my observations
on the rat. A variable decrease in suprarenal fat (lipoids) as a
268 Cc. M. JACKSON
result of various toxic conditions has been noted by various in-
vestigators in man and lower animals. The apparently contra-
dictory results of different observers as to the effect of inanition
upon the suprarenal fat (lipoids) is doubtless due to variations in.
the age, species, and individual animals, as well as to variations
in the extent and character of the inanition.
In hibernating gophers, Mann (716) found no change in the
lipoid content of the suprarenal. Frederici (’03) found no de-
crease in hibernating bats, but a progressive decrease is noted by
Baroncini and Beretta (’01) and Ciaccio (10). Bonnamour
(05 b) and Ciaccio (710) found a decrease in hibernating hedge-
hogs.
As to the effect of inanition upon the chromaffin reaction of
the suprarenal medulla, my results for the rat are more uniform.
They indicate that (with rare exceptions in extreme cases, espe-
cially in the younger stunted rats) there is no appreciable decrease
in the reaction, except as a result of postmortem changes. This
is in agreement with the results of Luksch (’05, ’11) and Kuri-
yama (718), who found no decrease in the epinephrin content of
starved rabbits, but contrary to Venulet and Dmitrowsky (710)
in the rabbit, Borberg (’12) in the cat and guinea-pig, Rondoni
and Montagnani (’15) in the guinea-pig, and Pellegrini (716) in
the later stages of fasting. In most cases, these positive results
were obtained on animals starved to death, and are therefore
probably explainable as due to postmortem changes. Effect of
anesthetics and acid in the fixative (for example, in the formalin
used for Wiesel’s mixture) are also possible sources of error, as I
have learned by experience.
MORPHOGENESIS OF THE SUPRARENAL GLAND
Some general features in the process of morphogenesis of the
suprarenal gland may now be discussed. As is well known (for
details in various species compare Poll, (’05), the medulla arises
in the embryo in connection with the sympathetic system, and
later migrates, usually in the form of multiple, small sympatho-
chromaffin cords or masses, which (in mammals) pass through
SUPRARENAL GLAND—EFFECTS OF INANITION 269
the cortical anlage and finally collect in the center of the gland.
Here they later become confluent, a few cells (‘sympathoblasts’)
forming the sympathetic ganglion cells, the majority ‘phiochro-
moblasts’ or ‘chromaffinoblasts’) ripening into the characteristic
parenchyma of the medulla.
The time at which this confluence of the medulla is finally
completed varies in different mammals, but is usually during the
late fetal period. In Echidna, however, the immigration of the
sympatho-chromaffin anlage is greatly delayed, and the supra-
renal medulla is not formed until long after birth (Keibel, ’04).
Also in the new-born mouse (Inaba, ’91), cat, dog, and guinea-pig
(Soulié, 703) and occasionally even in the human new-born
(Starkel and Wegrzynowski, ’10; Zuckerlandl, ’12), the process is
still incomplete and admixture of cortex and medulla persists to
a variable extent after birth. In the mouse, Inaba (’91) found
the corticomedullary boundary line still indefinite at ten days,
but distinct at thirty days.
Soulié (’03) described the confluence of the medulla in the
suprarenal of the rat (Mus decumanus) as appearing in the 25-mm.
fetus. Dewitzky (’12), however, found the suprarenal medulla in
the rat very ill defined at birth, becoming distinct at three days
and thereafter. This is in general agreement with my observa-
tions, although I find the confluence of the medulla in the albino
rat to be a gradual process and subject to some individual varia-
tion. Itis usually completed by the end of the first week. Even
in the adult, however, short cords of cortical tissue may occa-
sionally extend into the medulla, as observed by Bonnamour
(05 a) in the rat and rabbit, and small cortical islands occur
near the margin of the medulla.
As to the exact manner in which the confluence occurs, but
few definite statements appear in the literature. Flint (00)
mentions.appearances of pressure atrophy in the cortical strands
in the medulla of the suprarenal in the fetal pig, and Soulié (’03)
states that the cells of the medulla anlage ‘‘etouffent peu a peu
les cordons corticaux emprisonnés au stade de pénétration.”’ It
seems to be generally assumed, however, as is stated definitely
by Inaba (’91) for the mouse, that these cortical cell strands and
270 Cc. M. JACKSON
masses intermingled with the medulla are finally displaced and
- squeezed out of the medullary mass. In the rat, at least, there is
no evidence that such a displacement occurs. It appears rather
that the medulla becomes confluent through degeneration and
absorption of the intermingled cortical remnants. This process
is difficult to observe in preparations fixed and stained in the
usual manner, on which account it is easily overlooked, but it is
clearly evident in frozen sections stained with Herxheimer’s scar-
let red. In such preparations, the stained liposomes reveal the
atrophic cortical cells undergoing gradual atrophy and absorp-
tion. A careful study of similar preparations would probably
show that in other forms the primitive cortical strands in the
medulla likewise undergo degeneration and absorption.
The process of absorption of the cortical tissue continues during
the postnatal growth and development of the suprarenal gland,
associated with the expansion of the medulla. The continued
postnatal growth of the medulla has been noted in the rat, cat,
rabbit, and,guinea-pig by Elliott and Tuckett (06) and in man
by Scheel (’08), Starkel and Wegrzynowski (10), Thomas (’11).
Kern (711), and others. The continued postnatal increase in the
absolute volume of the medulla in the rat is confirmed by the
extensive data in the present study.
This expansion of the medulla necessarily involves an encroach-
ment upon the space formerly occupied by cortex. A priori, this
might happen in three ways: 1. There might be a corresponding
interstitial growth and expansion of the adjacent cortex. This,
however, would require continued multiplication and growth of
the cells in the inner cortical zone, and numerous observers (as
previously shown) agree that postnatal cell division in the inner
cortical zone during postnatal growth rarely or never occurs.
2. In the absence of cell division with interstitial growth, the
inner cortical zone might remain passive and be mechanically
displaced by the expansion of the medulla. Such a displacement,
however, would inevitably result in a very marked flattening of
the cortical cells on the adjacent surface of the expanding medulla
It cannot be denied that occasionally such a flattening does ap-
pear, but it is irregular and inconstant. Indeed the characteristic
SUPRARENAL GLAND—EFFECTS OF INANITION 271
irregularity of arrangement of the cell cords in the zona reticularis
is perhaps in part due to the pressure of the expanding medulla.
In the rat, however, and apparently in other forms, the histo-
logical structure of the inner (reticular) zone at the cortico-
medullary border does not, in general, support the idea of a
mechanical displacement by pressure.
3. The remaining possibility is that there is an actual absorp-
tion and removal of the cortex at the corticomedullary border.
This theory is strongly supported by the available evidence. Hy-
peremia of the inner cortical zone, absence of cell division and a
more or less well-marked cell atrophy with degeneration and pig-
ment formation are (as has been shown) characteristic not only
for the rat, but for mammals in general. In most cases, this proc-
ess of absorption is comparatively slow and inconspicuous, as in
the rat, but in the human infant it is more prominent. The ex-
tensive degenerative atrophy of the inner cortical zone of the
suprarenal in the human new-born, as described by Starkel and
Wegrzynowski (’10), Thomas C11), Kern (’11), Elliott and Ar-
mour (’11), Landau (’13 a) and Lewis and Pappenheimer (’16), is
therefore not a unique phenomenon, as heretofore supposed. It
appears to be merely an exaggeration of the same fundamental
process found in the development of the suprarenal in other
mammals. It is thus incorrect to claim that nothing similar
occurs in the lower animals (Kern, ’11 ; Dewitzky, ’12; Landau,
13a). The erosion of the inner cortical zone is evidently a con-
tinuation of the same process of degeneration and absorption of
the cortex which removes the cortical strands at the time of the
original confluence of the medulla. Minot (97), on the other
hand, believed that the cells of the primitive medulla anlage dis-
appear in the fetus, and agreed with the view of Gottschau (’83)
that the permanent medulla is derived by transformation of the
suprarenal cortex.
The process of cortical erosion by the medulla may be com-
pared with that of the absorption of the cartilage by the osteo-
genic tissue in the zone of enchondral ossification. In both
cases, small islands of the invaded tissue may persist for variable
periods. It is significant that such cortical islands in the medulla
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, No. 3
272 Cc. M. JACKSON
of the suprarenal in animals after the earlier postnatal stages
usually occur only in the immediate neighborhood of the cortico-
medullary border. Those originally in the central portion of the
medulla have usually undergone complete absorption. Flint
(00), Pellegrino (04), and others have described cortical islands
in the medulla of the adult suprarenal, and they occur also in the
rat; but these probably represent the results of later cortical ero-
sion, rather than persistent embryonic remnants.
The exact nature of this absorptive process is uncertain. It
does not appear to be a mere pressure atrophy, due to the expan-
sion of the cortex, although this may be a factor of subordinate
importance in the process. . It would appear to be rather a chem-
ical process of obscure nature, possibly a result of the contact
between the essentially alien cortical and medullary tissues, not
yet fully adapted to each other in their comparatively recent
phylogenetic association. The observation by Lewis and Pap-
penheimer (10) of similar involutional changes in accessory Su-
prarenals composed of cortical tissue only would seem to invali-
date this theory, but this point needs further investigation. It
does not appear probable that the absorption of the cortex at the
corticomedullary border is of functional significance, as claimed
by Gottschau (793) and Mulon (02, ’03. a, 03 b, ’05 a, 705 b,
1D):
If the inner zone of the cortex is subject to continued erosion
and absorption during the morphogenesis of the suprarenal, it is
evident that the zone must be constantly regenerated. As has
been previously shown, the abundant evidence in various animals
clearly establishes the fact that although during embryonic and
fetal periods cell division occurs throughout the cortex, during
postnatal development it becomes progressively restricted to the
outer region of the cortex in general, and to the zona glomerulosa
in particular. There is therefore during the postnatal growth
period a continued renewal of the suprarenal cortex, the cyto-
morphosis of the cells including an origin in or near the outer
zone (glomerulosa), a descent through the middle zone (fascicu-
lata), and a final atrophy, absorption and removal in the inner
zone (reticularis) at the corticomedullary border. The process
SUPRARENAL GLAND—EFFECTS OF INANITION Zhe
may be compared with the cytomorphosis of the cells of dhe epi-
dermis, where, however, the process is centroperipheral instead
of peripherocentral in direction.
The weight or volume of the suprarenal gland as a whole, to-
gether with the relative volumes of the cortical and medullary
constituents, will therefore vary according to the rate of expan-
sion of the medulla, the rate of erosion of the cortex at the inner
zone, and the rate of regeneration from the outer zone. Thus
the postnatal involution of the inner zone in the human suprare-
nal is so rapid that it is not fully compensated by regeneration
from the outer zone, or by expansion of the medulla; so the gland
during the first year actually decreases in weight, as shown by
the data of Scheel (’08) Starkel and Wegrzynowski (710), and
others. In the rat the retardarion in the growth of the gland
during the first week is perhaps explainable upon the same basis.
Subsequent changes in the absolute and relative volume of the
cortex in the rat and other forms are evidently subject to much
variation in different individuals and species. As a rule, how-
ever, as in the rat, the medulla appears relatively small in vol-
ume in the earlier prenatal stages (during immigration), expands
rapidly to a maximum relative size following its confluence (early
postnatal stages), and thereafter decreases relatively correspond-
ing to the later more vigorous growth of the cortex (Canalis, ’87;
Hultgren and Anderson, ’99; Soulié, ’03; Elliott and Tuckett, ’06;
Scheel, ’08; Starkel and Wegrzynowski, 710; Thomas, ’11, and
others).
SUMMARY
1. In the new-born rat, the suprarenal cortex and medulla are
not yet distinctly separated, the cortical cell strands in the
medulla being absorbed during the confluence of the medulla
in the first, week. During the second and third weeks after
birth, the cortex increases from 75 or 80 per cent to about 90 per
cent of the entire gland, by volume. It apparently continues to
increase relatively to about 93 per cent at ten weeks of age, de-
creasing slightly in the adult. The medulla increases more
slowly in absolute volume, thereby decreasing in relative volume
274 Cc. M. JACKSON
from 20 or 25 per cent of the gland at one week to 15 per cent
at two weeks, 10 per cent at three weeks, and 7 per cent at ten
weeks, increasing again slightly in the adult.
2. The relative volumes of cortex and medulla are subject to
considerable individual variation. Their ratio shows no distinct
difference according to sex and is not materially changed in
young rats stunted by underfeeding or in adults subjected to
acute or chronic inanition.
3. The vascular stroma (in comparison with parenchyma) nor-
mally shows considerable individual variabilty in relative volume,
due chiefly to the varying distention of the blood-vessels. In
general, however, there is evidently an increased vascularity upon
passing from the outer zone of the cortex (average 13 per cent)
toward the medulla (average 28 per cent), with no constant
change according to age.
4. The changes in the volume of the vascular stroma during
inanition are variable. In general, the stunted young rats show
a marked hyperemia in the inner cortical zone, with relative
anemia in the middle and outer ones; medulla unchanged. In
stunted rats refed one to two weeks, the relative volume of the
stroma in general returns toward normal, though the outer zone
remains anemic. In adult rats with acute or chronic inanition
the inner and middle cortical zones appear hyperemic, the medulla
relatively anemic.
5. With the exception of the first week, there is in general an
increase in the size of the suprarenal parenchyma cells from birth
to ten weeks of age, with little or no increase thereafter. The
average change in cell diameter for the various zones between
birth and maturity is as follows: outer zone, 7 » to 9 »; middle
zone (outer portion), 9 u to 15 w; middle zone (inner portion), 9 u
to 12 w; inner zone remains about 9 »; medulla, 8 » to 16 un.
6. The nuclei increase slightly in diameter during postnatal
life, excepting those of the inner cortical zone. The cytoplasmic
increase is much greater, however, so the nuclei in general lag
behind in relative size (nucleus-plasma ratio). Thus the relative
nuclear volume decreases, in the outer cortical zone, from about
44 per cent of the cell volume in the new-born to 23 per cent in
SUPRARENAL GLAND—EFFECTS OF INANITION 200
the adult; in the middle zone (outer part) from 33 per cent to
10 per cent; in the middle zone (inner part) from 28 per cent to
16 per cent; in the inner cortical zone from 28 per cent to 21
per cent; in the medulla from 46 per cent to 10 per cent.
7. In the underfed young rats stunted three to ten weeks or
more, the suprarenal cells may increase in size (outer part of
middle zone), or decrease (inner cortical zone and inner part of
middle zone), or remain nearly unchanged. The nuclei are
similarly variable, with slight changes in the relative volume.
In the stunted rats refed one or two weeks, the cells and nuclei
in general approach, but have not yet reached, their normal size.
8. In adult acute inanition there is but little change in the size
of the cells and nuclei in the outer and inner cortical zones. In
the middle zone and medulla, however, there is a marked loss in
size of the cells, though but slight loss in the nuclei (with corre-
sponding increase in relative nuclear volume). In chronic adult
inanition there is a greater decrease in the size of the outer- and
middle-zone cells, but about the same as during acute inanition
in the inner zone and medulla. The nuclear loss is somewhat
greater than during acute inanition, with relative nuclear volume
not very different from normal, except in the middle zone (where
it is high).
9, Amitosis in the suprarenal of the rat is infrequent and of
doubtful significance. Mitoses are frequent in the new-born (av-
erage about 20 per section), but fewer at the end of the first week
(10 per section). They increase to a maximum rate during the
second and third weeks, slowly decreasing in number thereafter,
although occasionally found even in the adult. Mlitoses are
most frequent in the outer zone and outermost part of the middle
zone (these forming the ‘germinative zone,) less frequent in the
medulla, and rare in the inner cortical zone.
10. In young rats stunted by underfeeding, mitosis is more or
less completely suppressed in the suprarenal. On refeeding one
week, mitosis begins again, and in two weeks the normal rate is
reestablished. Cell division in the suprarenal is therefore con-
trolled by the amount of nutrition.
276 Cc. M. JACKSON
11. The three cortical zones of the suprarenal are distinguish-
able from birth and well differentiated at three weeks. Lipo-
somes (lipoidal granules or droplets) are present in considerable
amount at birth, being rather uniformly distributed through
the cortex and the cortical strands through the medulla. They
increase slowly rather and the distribution changes. They be-
come more abundant in the outer zone and outer half of the
middle zone, decreasing in amount toward the inner zone, with
none in the medulla. The inner cortical zone is atrophic in char-
acter, with cells in various stages of degeneration and absorption.
Pigment (lipochrome) appears in these cells after eight weeks of
age. The degeneration and absorption of this zone is associated
with the expansion of the medulla in the morphogenesis of the
suprarenal gland. The absorption of the inner cortical zone dur-
ing the growth of the suprarenal is characteristic for mammals,
though greatly exaggerated in the human infant. The chromaf-
fin reaction of the medulla is weak at birth, becoming well marked
in the rat after the age of three weeks.
12. In the young rats stunted by underfeeding certain cell
changes occur in the suprarenal. The cortex tends to undergo
the normal differentiation of liposomes, but their amount is
variable. In some cases many of the cells appear atrophic and
degenerative, especially toward the inner cortical zone, where
the process is extreme. Pigment appears in unusual abundance.
The medulla is usually less affected, although pyenotic nuclei
frequently occur. Only in extreme cases is there any appreciable
decrease in the chromaffin reaction. In stunted rats refed two
weeks the structure is already gaining a normal appearance,
though some areas of degeneration persist.
13. In adult rats subjected to acute or chronic inanition the
suprarenal cells present a simple atrophy, together with a vari-
able amount of degeneration. The nuclei may be either hypo-
chromatic or hyperchromatic, with frequent pyenosis or karyoly-
sis. The liopsomes are retained tenaciously in the outer zone,
though usually decreased to a considerable extent in the rest of
the cortex, especially in extreme inanition. The pigment re-
mains unchanged. The medulla cells show degenerative changes,
SUPRARENAL GLAND—EFFECTS OF INANITION 277
though usually less marked than those of the cortex. The chro-
maffin reaction is apparently retained undiminished up to the
point of death, though occurring as a postmortem change.
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SUPRARENAL GLAND—EFFECTS OF INANITION 279
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280 Cc. M. JACKSON
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TABLE 1
Individual number, age and condition, sex, body length, body weight and weight of
the suprarenal glands tn the albino rats used for histological study
NOSE- BODY WEIGHT bee SEE
RAT NO. AGE, ETC. SEX ANUS GROSS ~— SUPRA- NOTE
LENGTH (OR NET) seas
A. Normal rats
mm. grams grams
F 10.1 New-born m. 46 4.2 0.0014 ss
F 10.2 1 day m. 50 4.8 | 0.0014 =
J1.7a New-born m. 4.9
J 1.7b New-born m. 4.9
F 8.1 New-born m. 50 il 0.0018 =
ype Ube 3 days m. 57 6.5 0.0020
Ret. 2 5 days tie 60 8.3 0.0020 *
halles 8 days m. 64 9.6 0.0018 cd
St 72.5 7 days ie 66 10.8 (10.1) | 0.0024
St 72.2 7 days m. 66 10.8 (10.0) | 0.0024
V 18.1 10 days m. 12.0 0.0030
St 80.5 12 days rs aD 15.5 0.0040
Vals 14 days m. 75 15.4
Be 17 days 1; 86 26.3 0.0054 =
V1.4 21 days m. 87 20.4
§ 5.2 21 days i 95 22.9 (21.1) | 0.0100
pils® 21 days ie 100 28.2 0.0078
J1.1 21 days m. 100 29.0 0.0078
St 47.2 21 days m. 102 34.3 0.0134
St 5.1 56 days ie 63.0 0.0116
St 5.2 56 days m. 77.0 0.0182
St 228.2 58 days m. 156 96.0 0.0183 a
St 47.6 67 days it 169 124.0 0.0364
St 47.5 67 days m. 191 196.0 0.0326
H 70.3 : 70 days m. 194 208 .0 0.0357
H 68.11 72 days m. 192 171.0 (164) 0.0228
H 68.8 74 days ie 184 142.0 (137) 0.0339
S 5.4 74 days fe 168 126.0 (118) 0.0319
S$ 5.3 74 days m. 180 172 0 (167) 0.0289
M 1.2 74 days m. 190 181.0 (173) 0.0264
piles 94 days m. 183 177.0 0.0249
H 64.3 101 days m. 191 185.0 (179) 0.0264
H 60.7 103 days te 203 (182) 0.0302
Leas 105 days m. 188 192.0 0.0234 .
H 58.3 106 days m. 225 (258) 0.0404
Apilire 112 days li 185 161.0 0.0365 | (1)
10) oil 116 days m. 176 149.0 0.0268 -
Mo. 9 120 days fe 180 157.0
H 36.3 138 days m. 202 (202) 0.0406
H 50.3 141 days 0 ee ee (222) | 0.0406
i)
90
bo
TABLE 1—Continued
WEIGHT
NOSE- BODY WEIGHT OF SEE
4 RAT NO. AGE, ETC. SEX ee See (on yan) SUFRA- NOTE
. mn grams grams
| aa | 141 days m. 195 200.0 0.0234
F 9.1 170 days m. 185 173.0 0.0246
F 6.1 150 days m. 188 178.0 0.0290
$14 (adult) m. 205 252.0 (247) 0.0414
21.3 202 days m. 216 (232) 0.0304
H 34.3 224 days i 194 (173) 0.0453
H 34.6 225 days E; 205 (188) 0.0604
L 3.7 234 days m. 190 241.0 (238) 0.0310
H 27.3 253 days ie 195 (166) 0.0475
H 27.6 254 days m. 214 (222) 0.0298
S 33.116 340 days Fi: 195 194.0 (188) 0.0414
S 33.117 346 days m
5 228 302.0 (294) 0.0330
B. Stunted young rats
!
53 4.5 0.0016 | *(2)
S 11.65 Maint. 21-73 days 100 23.8 (22.5) | 0.0118 | (2)
St 247.5 Maint. 1-7 days £:
St 80.9 Underfed 1-12 days 1 64 8.9 0.0026 (2)
St 228.4 Underfed 1-58 days m. 74 10.9 0.0054 | *(2)
St 9.36 Maint. 21-51 days m. 113 30.5 0.0124 | (2)
S 12.69 Maint. 21-66 days f. 100 24.5 (22.7) | 0.0090 | (2)
St 47.4 Maint. 21-66 days m. 113 32.3 0.0140 | (2)
St 47.3 Maint. 21-66 days m. 120 34.0 0.0136 | (2)
he oe ar Maint. 21-67 days m. 95 23.3 (21.2) | 0.0086 | (2)
S 7.34 Maint. 21-70 days m. P25 30.5 (29.0) | 0.0116 | (2)
8 7.31 Maint. 21-70 days m. 120 34.8 (31.4) | 0.0126 | (2)
S 7.32 Maint. 21-71 days m, 120 30.0 (28.8) | 0.0100 | (2)
9) 7.00 Maint. 21-71 days i 117 35.3 (31.0) | 0.0120 | (2)
m.
St 12.50 | Maint. 21-82 days m. | 123 45.0 (41.2) (2)
Sig Gayl Maint. 22-104 days it 89 19.1 (18.2) | 0.0086 (2)
St 38.8 Maint. 21-139 days m. 118 30.0 0.0161 | (2)
C. Refed after inanition
St 12.48 | 88 days Refed 7 days
St 11.42 | 88 days Refed 7 days
St.11.45 | 88 days Refed 7 days
St 10.27 | 89 days Refed 7 days
St 12.51 | 95 days Refed 14 days
St 11.43 | 95 days Refed 14 days
St 11.40 | 95 days Refed 14 days
St 10.26 | 96 days Refed 14 days
F 2.3 122 days Refed 6 days
S 33.120 | 339 days Refed 189 days
S 33.118 | 346 days Refed 206 days
125 50.2 (46.5) | 0.0105 | (3)
125 55.0 (50.3) | 0.0160 | (3)
130 67.6 (57.7) | 0.0156 | (3)
127 55.0 (51.4) | 0.0115 | (3)
137 77.2 (70.7) | 0.0150 | (3)
143 79.0 (74.7) | 0.0196 | (3)
142 84.5 (76.5) | 0.0158 | (3)
150 91.0 (86.3) | 0.0180] (3)
176 144.0 0.0350 | -*(3)
181 162.0 (156) 0.0396 | (3)
204 229.0 (219) 0.0348 | (3)
Bear Side) meee eee wee
TABLE 1—Concluded
NOSE- BODY WEIGHT | yaa | SEE
RAT NO. AGE, ETC. SEX ANUS GROSS SUPRA- NOTE
LENGTH (OR NET) Fenn e
D. Adult acute inanition
| mm. grams grams
22 116 days (loss 29%) m. 173 104.0 | 0.0316 | *(4)
ie 147 days (loss 30%) m. 188 162.0 0.0374 | *(4)
S 27 (loss 30%) m. 215 223.0 (219) 0.0424 | (4)
M 2 (loss 338%) m. 185 (?)| 170.0 (167) 0.0338 (4)
F 9.3 170 days (loss 338%) m. 179 129.0 0.0334 | *(4)
Rubee 151 days (loss 34%) m. 183 114.0 0.0422 | *(4)
Beane 130 days (loss 34%) m. 171 115.0 0.0280 | *(4)
J 1.4 94 days (loss 35% m. 180 107.0 0.0302 | (4)
S 25 (loss 35%) m. 205 202.0 (198) 0.0458 | (4)
F 9.2 170 days (loss 36%) m. 191 127.0 0.0374 | *(4)
S 26 (loss 39%) m. 205 174.0 (171) 0.0233 (4)
F 4.2 153 days (loss 43%) m. ie 105.0 0.0332 | *(4)
Heda 105 days (loss 45%) m. 185 110.0 - 0.0318 | *(4)
E. Adult chronic inanition
Jie5 117 days (loss 34%) m. 175 97.0 0.0240 (5)
M 3 (loss 34%) m. 175 125.0 (122) 0.0252 | (5)
M 12 (loss 36%) m. 173 128.0 (125) 0.0306 (5)
M5 : (loss 37%) m. 190 129.0 (127) 0.0320 (5)
M 6 (loss 37%) m. 175 138.0 (134) 0.0270 (5)
M 11 (loss 38% m. 190 163.0 (159) 0.0322 (5)
* The ‘F’ series, and a few others as indicated, were cut by frozen sections
for study in the fresh condition, or for lipoids, chromaffin reaction, ete. All
of the remaining glands were embedded and cut in paraffin sections.
1Rat J 1.7 had just given birth to a (first) litter. The remaining females
were virgins.
2 Of the stunted rats (table 1 B), St 80.9 and St 228.4 had practically doubled
their initial weight during the period of underfeeding. The others were held
nearly at maintenance (constant body weight) during the period indicated.
3 Of the rats refed after inanition (table 1 C), all excepting the last three
were refed after having been held nearly at maintenance from three to about twelve
weeks of age. S 33.118 and S 33.120 had been held with but slight increase in
body weight from 21 to 140 and 150 days of age, respectively, and were apparently
permanently stunted. Rat F 2.3 decreased from 151 grams to 104 grams (loss
of 31 per cent) during seven days of acute inanition, and was then refed as
indicated.
4 The rats subjected to acute inanition (table 1 D) had been given water only
for periods varying from seven to twelve days, with loss in body weight as indi-
cated. In four cases the age was unknown. The final body weights are given
in the table.
5 The rats with chronic inanition (table 1 E) had been fed gradually decreasing
amounts of food during a period of about five weeks, with losses in body weight
as indicated. The final body weights are given. The age was unknown in all
but the first rat.
284
‘
TABLE 2
Postnatal growth of volume in suprarenal cortex and medulla in the iho rat
SUPRA- |pERCENTAGE FORMED BY THE
eee supRa- | RENALS
2 1 : PER CENT
cata s | “AND SEX be WEIGHT none Cor tex! Medulla, average
ace |“VERAGE) \ picur Be (and range)
AVERAGE| ~~
A. Normal
I
grams grams | per cent oe per cent
20:
NOMA aY Ss... oo y= > oth 2 3 m | 13.7 |(0.0031)|(0.019)| 80.9) 19.1 (16. 8-22.3)
TIDE WIG NG ke (ee eae Df 15.5 | 0.0040 | 0.026 | 84.3] 15.7 (15.3-16.4)
UGE: Se cee ge ae 5 m | 27.9 | 0.0106 | 0.033 | 90.6} 9.4 (8.5-10.8)
ZAG Ey (stoi Gear ari See 2 £ 28.2 | 0.0078 | 0.028 | 89.5] 10.5 (10.0—11.0)
GR OLY GEER ora «ic svoles * 2 m | 77.0 | 0.0182 | 0.024 | 92.3) 7.7 (7.48.1)
Cia. G RNS. eee ee one 1 63.0 | 0.0116 | 0.018 | 82.7) 17.3
GYSVERO ES a eoaey hep ae 3 m |180.3 | 0.0218 | 0.012 | 92.8} 7.2 (5.8-8.0)
BTA IOMV Ss 5 neue f Sep30 3 f |132.6 | 0.0414 | 0.032 | 93.5} 6.5 (5.3-7.2)
94-346 days............- 8 m |220.1 | 0.0322 | 0.015 | 89.9) 10.1 (6-16)
112-253 pee ar eke: Sineeeie ee tenia wee A pte ee eA 4 f£ |161.1 |.0.0426 | 0.027 | 90.9} 9-1 (7.0-11.9)
B. det sk ee he be ee A Oe ie One Bebe 2 ee ae ee experiments
79.3} 20.6 (18.6-22. ert lis ed. | 2 2) foes heonasl| otnay| gala 206 cis.6-22.7)
90.2) 9.8
89.8) 10. 2 (8.7-12.5)
88.2} 11.8 (11.4-12.2)
89.3} 10.7
Underfed 1-12 days..... 8.9 | 0.0026 | 0.029
30.5 | 0.0124 | 0.040
f
Maintenance 21-51 days. m
a) 34.1 | 0.0131 | 0.041
m
m
2
1
Maintenance 3-10 weeks.| 7
1
a
1
Maintenance 21-82 days.
Maintenance 21-139 days 30.0 | 0.0161 | 0.054
mptiy Pires Ges Se wd. Tt; 1 m | 10.8 | 0.0024 | 0.025 | 74.5] 25.5 .
7 days Pep 10.8 | 0.0024 Oe ees 20.4 (18.6-21.7)
ated bweeks 1H) ps 6 £ | 57.6 | 0.0125 | 0.024 | 89.4) 10.6 (9.6-11.6)
2 s
See as as : oh 82.9 | 0.0171 | 0.622 | 90.4) 9.6 (8.8-11.1)
Refed 189 days..........| 2 £ |162.0 | 0.0396 | 0.024 | 85.7] 14.3 (13.4-15.3)
Refed 206 days.......... 2 m |229.0 | 0.0348 | 0.016 | 90.8} 9.2 (9.0-9.4)
Adult acute inanition...| 6 m |177.3 | 0.0393 | 0.024 | 90.1) 9.9 (5.8-12.8)
Adult chronic inanition.| 9 m|123.4 | 0.0278 }\0.023 | 88.7) 11.3 (8.1-13.1)
RES er ee
1'The ‘No.’ in the second column refers to number of glands, not animals.
The suprarenal weights were missing in one of the three normal males at ten
to fourteen days.
285
Cc. M. JACKSON
286
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SUPRARENAL GLAND—EFFECTS OF INANITION 287-
TABLE 5
Mitoses in the suprarenal gland of the albino rat
NORE AVERAGE NUMBER OF MITOSES PER SECTION IN
AGE AND CONDITION BER OF
GLANDS| Outer
zone
Inner | Me- | Total average
zone dulla |; (and range)
Middle
zone
A. In normal rats
ISG SL OVET E.G ge le i ee a 3 11 2 1 6 | 20 (12-29)
ODEON pena mee ep oat es ae ameemetr on 3 5 2 4 3 | 10 (7-18)
LU Sa a Se Be 6 5 3 4- | 16 (14-18)
LES ee An eed 1 18 17 0 5 | 40
ives. (OEE S FS che. hehe ie a 2 10 10 3 6 | 27 (22-381)
=D) ee a ae 9 9 12 z 2 | 24 (10-41)
SPOS 72 dg ae 1 3 5 0 S | 11
i 94 dances aie n Sh see. sort es. 4 2 2 a 0 4 (2-5)
CAE ICr hs 2 1 1 0 0 2 (1-3)
CUE ee, fe a tee Seat 3 oe as a el 1 1 0 0 0 1
B. In test rats
Maintenance birth to 12 days...... 1 1 1 0 mS) 14.
Maintenance 3-10 weeks...........| 10! 3 4 0 0 1 (0-8)
Maintenance 3-12 to 20 weeks..... 3 0 0 0 0 0
Refed 1 week........... Bs 23 =, 2 SS: 3 3 1 0 3 2 (1-3)
ELCONAtWEEKS este ot ccc en 2 3 5 0 2 LO“ (712)
PEI suG MUVGHT Ree ceteris Ss 3 553 1 0 0 0 0 0
1 Six of the ten held at maintenance from three to ten weeks of age showed no
mitoses in the sections examined.
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, No. 3
JACKSON
M.
C.
288
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289
EFFECTS OF INANITION
SUPRARENAL GLAND
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Resumido por el autor, John C. Donaldson. |
El volumen relativo de la corteza y médula en la glandula adrenal
de la rata albina.
Las medidas que han servido de base al presente estudio han
sido tomadas en las glindulas adrenales de diez y siete ratas
albinas, fijadas en solucién de Bouin y después reconstruidas por
medio de cortes seriados. La adrenal izquierda es, en general,
mas grande que la del lado derecho. El volumen relativo que
ocupa el tejido medular decrece desde préximamente el 12 por
ciento en los machos y el 10 por ciento en las hembras, ambos
recién nacidos, hasta llegar a ser el 7.5 por ciento y 6.5 por ciento,
respectivamente, hacia el periodo de la pubertad. Desde este
momento en adelante hay, relativamente, un cambio muy pe-
quenho. Cuando se comparan las glandulas de la hembra con las
del macho, en ejemplares del mismo peso, las de la primera con-
tienen relativamente menos médula.
Translation by José F. Nonidez
Columbia University
AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, MARCH 17
THE RELATIVE VOLUMES OF THE CORTEX AND
MEDULLA OF THE ADRENAL GLAND IN
THE ALBINO RAT
JOHN C. DONALDSON
Department of Anatomy, School of Medicine, University of Cincinnati
FOUR CHARTS
This study was undertaken at the suggestion of Dr. H. H.
Donaldson to determine the relations of the cortical and medul-
lary portions of the adrenal gland in the albino rat, M. nor-
vegicus albinus, and to demonstrate the changes in their volume
which take place with age.
In the albino rat at birth there is a fairly well-marked cortex
and medulla in the adrenals, though there are numerous groups
of medullary cells still scattered through the cortex. Within a
few days most of these have disappeared and the cortex and
medulla are sharply marked off from one another. There are
present in the albino rat, in addition to the tissues in the adrenal,
microscopic masses of chromaffin cells in the retroperitoneal
tissue, Fulk and Macleod (16), and a small but constant mass
of cortical cells in the epididymis of the testis, Swale Vincent
(12). No attempt was made to include these extra-adrenal
masses in this study.
The materials used were the adrenal glands from seventeen
rats, nine males and eight females, of Wistar Institute stock.
The glands were fixed in Bouin’s solution. After having been
stored in cedar oil, they were passed through xylol, embedded
in paraffin, and cut into serial sections 10 » thick. The sections
were stained with haematoxylin and eosin. This gives a very
sharp contrast between the cortex and the medulla, the latter
showing as a blue mass in the surrounding pink-stained cortex.
Every tenth section was projected with an enlargement of fifty
291
292 JOHN C. DONALDSON
diameters, and the outlines of the -entire section and of the
medulla were traced. The area of each of these figures was
measured with a planimeter. From these areas and the known
thickness of the section the actual volume of each section
traced and of its medulla was calculated. The nine sections
between each of those traced were considered as each having
the same volume as the nearest traced section. By adding
the volumes determined together the total volumes of the
gland and of the medulla were found. Blood sinuses and
blood-vessels were considered as a part of the tissues in which
they were found. In other words, no attempt was made to
subtract their volume from that of the gland. The thin capsule
was included in the area of most of the sections. In those
toward the end of the gland, where the capsule appeared thick
owing to the plane of section passing tangentially to the surface
of the gland, the capsule was omitted from the areas.
Somewhat similar methods were used by Elliot and Tuckett
(06) in their study of the adrenals of a number of mammals.
They examined the cat, the guinea-pig, and the rabbit, and found
in these animals that the relative volume of the medulla, com-
pared with the volume of the whole gland, decreased with age.
There were marked differences in the relative amount of med-
ullary tissue in the gland, the maximum being 14 per cent of
the entire gland, in a young guinea-pig, and the minimum 1 per
cent in an old one. The values for the cat and the rabbit fell
between these extremes. These authors give the value of 9 per
cent for the amount of medullary tissue in the gland of a male
rat of 120 grams, and 6 per cent for that in one of 190 grams.
They do not state the variety of rat. They admit that the method
they used would not give accurate results with so small a gland
as that of the rat. In his study of the adrenal of the rabbit,
Bertel Bager (’17) finds that the medulla forms 20 per cent of
the total volume of the gland at birth. At twelve months it
makes up a little less than 2 per cent. The ratio rises to 3 per
cent as the animal gets older. The medulla is relatively smaller
in the females.
VOLUMES, CORTEX AND MEDULLA, ADRENAL GLAND 293
TABLE 1
AGE |VOLUME OF BOTH GLANDS|THE RATIO OF THE PEPE RENCE OX
ESTIMATED VOLUME or |YOLUME BETWEEN
104.0 71 =: {17.60 | 1.00 |16.60
192.0 112. _|17.90 | 1.50 |16.40
OR etaiax: Bobs THOT DE eR THM MED ULNA ros Pee oe
BER WEIGHT eee LEFT ADRENAL.
Leyes Total dulla Cortex THE a ee H eae
en RRNA OO za60 (gn || pe ominellh’ percent Ont eter tent
1 M 4.6 1 0.825} 0.095} 0.730 33 4.0
2 F 5.9 2 1.042} 0.114} 0.928 10.2 20.0
3 M 17.0 14 2.010} 0.260} 1.750 13.0 4.0
4 F a0 16 2.130) 0.200) 1.930 9.1 11.0
5 M 26.0 29 ~ | 5.370} 0.580} 4.840 9.8 12.0
6 F 28.0 29 5.49 | 0.39 | 5.10 G2, 20.0
1h F BY (al) 35 7.35 |0.53' | 6.82 iad 15.0
8 F 39.0 35 7.55 0.61 | 6.94 8.0 8.0
9 M 39.0 38 7.46 | 0.60 | 6.86 8.0 —0.2
10 M 50.0 45 7.06 | 0.50 | 6.56 7.0 2.0
11 M 7A \0) 55 10.68 | 0.87 | 9.81 14.0
12 F 79.0 64 16.70 | 1.20 |15.50 11.0
13 M
F
M
8.2
6.9
103.0 69 {11.51 | 0.81 {10.70 7.0 —17.0
6.0
8.2
16 | Damaged in prepa-
ration
AT El e20SeOhN, 202) 1830) |) 2.2: | 20:8 6.6 12
18 Tumor of the adrenal
19 M | 300.0 | 2SO ye 2b 2 | S2e 2° O20 10.2 18.0
The values for the volume of the cortex were obtained by subtracting the
volume of the medulla from that of the total gland.
The results of my observations are shown in the accom-
panying table 1 and the four charts. All figures are for the sum
of the two adrenals of each rat. In this connection it is worth
noting that in fifteen out of the seventeen rats the left adrenal
was larger than the right; exceeding it in volume by an average
of about 10 per cent and in one case by as much as 20 percent.
Chart 1 shows the change in the volume of the cortical and
medullary material in the gland of the rat from birth to nine
months—5 to 300 grams. The values were gotten by finding
out what fraction of the whole gland was composed of medulla
and reducing that fraction to its percentage value. The values
294 JOHN C. DONALDSON
for the males and females have been plotted separately. It
will be seen that the curves for the two sexes are essentially
parallel, but that the one for the females, marked by crosses,
indicates a smaller relative value for the medulla, and at the end
of the female curve the relative volume becomes stationary.
Both curves show a rapid decrease in the relative volume of the
medulla until about the age of puberty, then in the males an in-
50 100 150 260 250 300
Chart 1 The change in volume of the medulla expressed as a percentage of
the whole adrenal gland. Dots, e, males, individual values. Crosses, +, females
individual values.
crease and in the females no further loss. Dr. Jackson, in some
recently completed work (Jackson, ’18), has found essentially
the same relative decrease in the chromaffin tissue in the rat. In
his rats, however, he did not find any differences between the
sexes. His results, as he has kindly told me, were not arranged.
to bring out any such differences which might exist.
The recent work of Elliot and Armour (11), and Lewis and
Pappenheimer (716) shows that in the human adrenal just after
birth there is a rapid degeneration and absorption of the inner
VOLUMES, CORTEX AND MEDULLA, ADRENAL GLAND 295
portion of the cortex. The resulting rapid reduction in the
relative size of the enormous cortex is much greater than any-
thing shown in chart 1, and is in the direction of a relative in-
crease of medullary tissue. This degeneration has never been
noted except in man. There is nothing resembling it in my
specimens.
VOLUME
CUBIC M.M.
BODY WEIGHT
GRAMS
30 !06 150 200 250 300
Chart 2 The growth in volume of the male adrenal cortex and medulla. The
lowest solid line represents the medulla. The broken line marked by crosses
represents the vakues for the medulla multiplied by ten, so that its growth can
more conveniently be compared with that of the cortex.
Charts 2 and 3 show, in solid lines, the growth curve, in volume,
for the cortex and for the medulla, chart 2 for the males and
chart 3 for the females. The broken line represents the values
for the medulla multiplied by ten; so that the relative growths
of the two parts can be more conveniently compared. The
values for both the cortex and medulla are expressed in cubic
millimeters.
Chart 4 shows the values for the whole of the adrenal in cubic
millimeters. When compared with the values for the weight of
296 JOHN C. DONALDSON
the adrenal found in table 71 of the rat (Donaldson,’15) and
shown in chart 20 of the same, it will be seen that the curves are
similar in form and relation, but that the values in chart 4 are
about one-half what they should be to satisfy the weights given
for the respective ages in table 71. Jackson (717) found a similar
VOLUME
CUBIC M.M
50 100 150 200 250 300
Chart 3 The growth of the female adrenal cortex and medulla. The lines
have the same significance as those in chart 2.
discrepancy between the volumes and the weights, when recon-
structing the hypophysis from serial sections. He suggests
that shrinkage by reagents and the weighing of a certain amount
of tissue which is not included when reconstructing the organ
explain this. Recent work at The Wistar Institute mdicates
VOLUMES, CORTEX AND MEDULLA, ADRENAL GLAND 297
that when specimens are embedded in paraffin directly from
cedar oil they may show a loss of as much as 41 per cent of their
fresh volume.
VOLUME
CUBIC M.M.
BODY WEIGHT
GRAMS
50 100 150 200 250 300
Chart 4 The growth of the whole gland expressed in cubic millimeters. The
upper line gives the values for the females, the lower line that for the males.
Dots, *, males, individual values. Crosses, +, females, individual values.
To sum up: The left adrenal in the albino rat is usually dis-
tinctly heavier than the right. The relative volume occupied
by the medullary tissue decreases rapidly from birth to puberty,
50 to 100 grams, and then remains stationary or increases a little
298 JOHN C. DONALDSON
with age. When compared with those of the male, the glands
of the female, body weight for body weight, contain relatively
less medulla. The difference, however, is slight, less than 2
per cent (chart 1).
Results found by the reconstruction method are comparable
with results of growth expressed by weight (chart 4).
LITERATURE CITED
Bacer, Bertret. 1917 Bidrag till binjurarnas 4ldersanatomi hos kaninen
(The anatomy of the adrenal in the rabbit as modified by age). Up-
sala Likareférenings Férhandlingar. Ny foljd. Bd. 23. H. I-2.
CrowrE AND Wistockr 1914 Experimental observations on the suprarenal
gland with special reference to the functions of their interrenal por-
tions. The Johns Hopkins Bull., no. 284, pp. 287-304
Donatpson H. H. 1915. The rat. Reference tables and data. Memoirs of
The Wistar Institute, no. 6, Philadelphia.
Exuiot, T. R., anp Armour, R. G. 1911 The development of the cortex in
the human suprarenal gland and its condition in hemicephaly. J.
Pathology and Bacteriology, vol 15, pp. 481-488.
Exuiot, T. R. anp Tuckett, I. 1906 The cortex and medulla of the adrenal.
J. Physiology, vol. 34, pp. 333-369.
Fuux, M. E., anp Macteop, J. J. 1916 Evidence that the active principle
of the retroperitoneal chromaphil tissue has the same physiological
action as the active principle of the suprarenal gland. Amer. J. of
Physiology, vol. 40, no. 1, pp. 21-29.
Jackson, C. M. 1917 Effects of inanition and refeeding upon the growth
and structure of the hypophysis in the albino rat. Amer. J. Anatomy,
vol» 21, mo. 2.
1918 The postnatal development of the suprarenal gland and the
effects of inanition upon its growth and structure in the albino rat.
(In the course of publication.)
Lewis, R. W., AND Parpenneimer, A. M. 1916 A study of the involutional
changes which occur in the adrenal cortex during infancy. J. of Med.
Research, vol. 34, pp. 81-93.
VINCENT, SwALE 1912 Internal secretion and fie ductless glands. Edward
Arnold. London.
1917 Experimental and clinical evidence as to the influence exerted
by the adrenal bodies upon the genital system. J. Surgery, Gynae-
cology, and Obstetrics, vol. 25, pp. 294-299.
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Resumido por el autor, Franklin P. Johnson.
El desarrollo de los l6bulos del higado en el cerdo.
En el presente trabajo el autor se ocupa principalmente del
desarrollo tardio del higado del cerdo, considerando especial-
mente la formacién de los tabiques interlobulares formados por
tejido conectivo, el modo de formacién de nuevos lébulos, el
crecimiento de las venas hepatica y porta y el desarrollo del
higado en conjunto. Los resultados obtenidos pueden resumirse
del siguiente modo: los tabiques de tejido conectivo se distin-
guen ya en el animal recién nacido, pero no se hacen bien patentes -
hasta que el animal tiene dos meses de edad. Se originan como
extensiones del tejido conectivo que rodea a la vena porta. La
excisiOn de los lébulos origina otros nuevos. Durante este pro-
ceso la vena central del l6bulo o bien se bifurca o produce una
nueva rama en uno de sus lados, y siguiendo su trayecto aparece
un nuevo tabique intralobular de tejido conectivo que se extiende
formando angulo recto con la superficie del l6bulo, entre las dos
nuevas venas centrales. Cuando se completa la excisién resultan
dos nuevos lébulos provisto cada uno de ellos de una vena cen-
tral, mientras que el tabique de tejido conectivo y la antigua vena
central adoptan posicién interlobular. Los tabiques de tejido
conectivo desempefan tan solo un papel pasivo en la formacién
de nuevos lébulos. El crecimiento del higado durante todos los
estados de su desarrollo tiene lugar: 1) por el aumento del nit-
mero de lébulos; 2) por el aumento de tamajio de estos ultimos.
En general todas las partes del higado crecen simultaneamente y
de una manera semejante. Hay un cambio constante en la posi-
cién de los lébulos en la periferia, pero este cambio se lleva a
cabo de tal modo que las relaciones de un l6bulo con otro sufren
una alteracién minima.
Translation by José F. Nonidez
Columbia University
AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, APRIL 7
THE DEVELOPMENT OF THE LOBULE OF THE PIG’S
LIVER
FRANKLIN PARADISE JOHNSON
Department of Anatomy, University of Missouri
TWENTY-EIGHT FIGURES
CONTENTS
LOSS TL Pe ghe B a SRS Sane een A ee ON Ree, rere ETO Tee a eee eee 299
Development of the connective tissue septa...................0 2 eee eee ee 301
CHEESES VES el See get a ae Neat nse dnd « bh DAEs Seay iat Ape ae at 2 lied 308
HOLM GION OMmneWPLODULES Hs cine o.cteoeee aie ook oe eee elo ae ACs cae Sete 308
Rilevof the/ connective tissue septa: :.:... 224 anh elas d. sie. ate 2 oda rinen 318
PmeeaRMLRMRE TEI UOT Sikes oh ete sade Ova a Was dei nines DEA Ro alngn are 5 cue Sty ae 319
UD TL SVE Cis ree ii inde as fe ec eee mete Mon op 1 bee onl cae), eda 325
INTRODUCTION
The development of the liver lobules offers a difficult problem
owing to the fact that in most animals the lobules have no defi-
nite boundaries, one running directly into the other without
demarcation. In section, therefore, the livers appear to con-
sist of solid masses of parenchyma, pierced at more or less reg-
ular and alternating intervals by various-sized branches of the
portal and hepatic veins. This lack of lobule definition is so
great that one readily appreciates why Weber, in 1842, denied
the presence of true lobules in the human liver, and one may
himself doubt the appropriateness of the term ‘lobule’ for either
the portal or hepatic units.
Early recognizing this difficulty, I chose for material the
liver of the pig, for I hoped that by using an animal in which
the liver lobules are definitely marked out, this difficulty would
be greatly overcome. I soon found that the pig’s liver does
not show indications of dividing septa until about birth, and
that the connective tissue septa are not definite until about
the second month of postnatal life. Contrary to the state-
299
300 FRANKLIN PARADISE JOHNSON
ment of Mall (06), lobule formation is not complete at this
time, for I find with Lewis (712) that ‘‘the multiplication of
lobules continues long after birth.”’ The few late stages of the
pig which I have been able to procure, consequently, have
been of great value in furnishing evidence concerning the de-
velopment of the hepatic lobules.
Because of the importance of the connective tissue septa,
I have found it advantageous to divide the development of the
pig’s liver into two definite periods—one before and one after the
septa are indicated. I shall speak of the former simply as the
‘early stages,’ of the latter as the ‘late stages.’ The early stages
include those up to but exclusive of an embryo of 254-mm.
in length; the later stages include the 254-mm. embryo and
extend to the adult.
In attempting to determine the manner in which the units
of the liver multiply, I first gathered together a series of se-
lected stages of developing livers. But so far as lobule forma-
tion is concerned, this was unnecessary. I agree with Mall,
(06) when he says, ‘‘the great difficulty is to recognize the
same thing from step to step,’”’ but I find the greatest difficulty
is to recognize the limits of a lobule in its three dimensions in
any single early stage. In the later stages, however, because
of the presence of the connective tissue septa, the lobules are
definitely bounded. Since in any developing liver the lobules
present numerous instances of every stage of development,
it is possible by a little study to arrange the various stages in
their proper sequences. In this manner the development of
the liver lobules may be easily and most satisfactorily deter-
mined from any single late stage.
Inasmuch as I have worked out the development of the liver
lobules from stages in which the connective tissue septa are
present and continually growing and delimiting new lobules, I
found it necessary to study first the origin of the septa. This
was the more essential, for an understanding of the different de-
velopmental stages of the septa makes it possible not only to
recognize dividing lobules, but to distinguish between the newly
formed and the old lobules.
DEVELOPMENT OF LOBULE OF PIG’S LIVER 301
DEVELOPMENT OF THE CONNECTIVE TISSUE SEPTA
In stages until nearly birth the liver of the pig shows no indica-
tions of connective tissue septa. The parenchyma is made up of
cells not greatly different from those of the adult. The sinu-
solids, which appear proportionately large, are lined with en-
dothelial cells. The ‘Gitterfasern’ or ‘reticulum of Mall’ (’96)
is demonstratable following Bielschowsky’s silver-impregnation
method, and is also slightly discernible after staining with
Mallory’s triple connective tissue stain. The branches of the
hepatic and portal veins interdigitate with one another and
furnish a means by which hepatic and portal lobules may in
certain areas be roughly outlined. The large branches of the
portal veins are readily distinguished from the hepatic, for they
are accompanied, as in the adult, by branches of the hepatic
artery and of the bile duct; some of the smaller veins are deter-
minable only with the aid of serial sections.
In figure 1 is shown a longitudinal section through a branch
of the portal vein, taken from the liver of a pig 229-mm. in
~length. The interdigitation of its branches with those of the
hepatic vein is clearly shown. Surrounding the larger branches
of the portal veins, bile ducts, and hepatic arteries is connective
tissue. Staining with Mallory’s triple connective tissue stain
clearly demostrates in it the presence of collagen fibrils. Where
the collagen fibrils are in contact with the parenchyma they
often send short fibrils into its reticulum. Surrounding the
hepatic veins there is but a thin layer of connective tissue,
which, moreover, does not extend out to as small branches as
does that surrounding the portal veins. The hepatic cells
are polyhedral in shape, with slightly granular protoplasm and
distinct chromatic nuclei. They are grouped in strands, but,
as stated by Theopold, a radial arrangement about the central
vein is not yet to be found. Here and there are to be seen small
clusters of nucleated round cells. These cells, which we know
to be developing blood-cells, were first described by Ko6lliker
C79), but were thought by Toldt and Zuckerkandl (’75) to be
young hepatic cells. That Kolliker’s interpretation is correct,
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, NO. 3
Do
°
a
FRANKLIN PARADISE JOHNSON
302
Mallory’s triple connective-
. in length.
ig 229 mm
ion of liver of p
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DEVELOPMENT OF LOBULE OF PIG’S LIVER 303
however, has been abundantly proved by the researches of Van
der Stricht (’91); Kostanecki (92), Engel (99), Lobenhoffer
(08), and others. Large mononuclear giant-cells, also de-
scribed by Kdlliker, are to be found sparsely scattered through
sections of the liver. In the later fetal stages both the group
of embryonic blood-cells and the giant-cells become fewer and
fewer, as is agreed by all investigators. I find with Theopold,
however, that they have not all disappeared at birth and that
a few may be found for several weeks after birth.
The first indication of a segmentation of the liver paren-
chyma into hepatic lobules which I have found has been in an
embryo of 254-mm. in length. According to Engel (’99), this
pig would be one at about full term, since he states that a pig
at birth measures 25 em. Theopold (10), however, shows that
pigs at full term vary considerably in length; that they even
attain a length of 35 em. before birth. In the 254-mm. speci-
men, the liver cells appear more vesicular, their protoplasm
stains but faintly. The boundaries of the hepatic lobules as
seen in figure 2 are in many places definitely marked. A close
analysis of the dividing medium, however, shows that it con-
tains no collagen fibrils, but rather its appearance is due to a
change in the parenchyma. A higher-power drawing of this
stage is shown in figure 3. To the right and left of the draw-
ing are shown terminal hepatic on central veins; above and
below are terminal portal veins. Extending from portal vein
to portal vein is shown the first evidence of an interlobular
septum. It consists of hepatic cells which are more coarsely
granular and stain more deeply than the remaining cells; similar
cells are found surrounding the portal veins. However, the
thing which makes the septum most apparent is the arrange-
ment of the cells. As seen in sections, they form two more
or less regular parallel rows, an arrangement which becomes
greatly accentuated in slightly older stages. Another factor
which in places intensifies the distinctness of the septa is the
presence of small branches of the portal veins. Nucleated
blood-cells found in the sinusoids along the septa also aid in
making it more distinct. I have studied carefully the retic-
304 FRANKLIN PARADISE JOHNSON
800.
Fig. 10 Drawn from a section through the terminal branches of a collateral
duct of a female albino rat of nine weeks. Zinker’s fixation; Weigert’s iron-
hematoxylin stain. Drawn with the aid of a camera lucida. c.t., connective
tissue. The cross-sections through the six terminal parts show in general a
single layer of epithelium. An occasional flattened nucleus represents the outer
or myo-epithelial layer. > 800.
422 J. A. MYERS
three parts: first, the short excretory duct which extends from
the milk-pore to a noticeable enlargement; second, the enlarge-
ment or milk-sinus, and, third, the milk-duct or canal. He
found the excretory duct lined with a stratified pavement epi-
thelium, while the sinus and milk-ducts possess two epithelial
layers. Benda (’94) found the epithelium near the attached
parts composed of two layers of cells, while in the deeper and
free parts of the ducts the epithelium was described as consisting
of a single layer. Raubitschek (’04) found in a human female
of seven days that the epithelium and the ducts is composed of
two layers. Brouha (’05) described this epithelium as consisting
of two layers throughout the entire system of ducts in virgin
Vespertilio murinus, the rabbit, and the domestic cat. Schil
(12) observed two distinct layers of cells around the ducts before
the stage of puberty. O’Donoghue (’12) confirms Brouha’s
finding in the domestic cat and writes concerning Dasyurus as
follows: ‘‘As in the foetus, so in the adult Dasyurus, all the
branching tubules of the mammary gland are lined by a double
layer of cells, at any rate in the resting animal, and for some
time after ovulation.”’
The fact that a few ducts were observed in the early postnatal
stages of the albino rat with small areas covered with a single
layer of epithelium probably has no significance as in later stages
all ducts observed were lined with a double layer of cells. The
outer layer of cells doubtless represents the so-called basket cells
or myo-epithelial cells of the active mammary gland, while the
inner layer represents the true glandular cells.
True alveoli lined with a single early of cells have been re-
ported in the glands of various animal species before puberty
by several writers. Such alveoli or acini have probably been
observed, as there is no reason to doubt that through individual
variation a few acini might develop just as a mature Graafian
follicle sometimes appears in the ovary of the human fetus or
new-born. However, Schil (’12) concluded that during this
period the mammary gland in the rabbit ordinarily presents
no glandular acini. O’Donoghue (’12), after studying the glands
of Dasyurus, made the following statement: ‘‘The true secre-
STUDIES ON THE MAMMARY GLAND 423
tory alveoli of the gland with their simple epithelial lining do
not make their appearance until the last few days of pregnancy
or until some time after ovulation when this has not been fol-
lowed by fertilization.” The work of these investigators, the
present work, and my previous work (Myers, 716) prove that in
certain animals true alveoli do not usually appear before puberty.
It has been proved quite conclusively that true secretory
alveoli do appear after the first ovulation. In fact, Ancel and
Bouin (711) and Schil (12) showed that in the rabbit the mam-
Fig. 11 Internal view of a wax model reconstructed from two terminal duct
and end-buds of the right first thoracic gland of a female albino rat of seven weeks.
e.b., end-bud; ep.w., epithelial wall; 7., lumen. X 100.
3
mary gland develops after each ovulation not followed by preg-
nancy to the same extent as it develops during the first half of
pregnancy. O’Donoghue (712) and Hartman (’18) reported
somewhat similar changes in the mammary glands of Dasyurus
and the opossum. Most of the investigators believe this develop-
ment is due to the corpus luteum. The enlarged terminations of
the ducts which resemble alveoli in some ten-week rats were
perhaps developed during the first period of heat or some time
after the first ovulation.
424 J. A. MYERS
Secretion in the new-born
It will. be recalled that twelve hours after birth a slight secre-
tion appears in the lumina of the milk-ducts. This secretion is
more abundant at the fourth and fifth days and remains present
in approximately the same amount during the first week. At
no time does it completely fill the lumina of the ducts. After
the second week the secretion gradually appears present in
smaller amounts; however, at the ninth and tenth weeks there
is still some secretion to be found in the lumina. There is only
a very occasional free epithelial cell present in the secretion.
During the first few days of postnatal life such cells manifest
no signs of degeneration, however, later a few were observed
undergoing degeneration. A few red blood corpuscles were ob-
served in the lumina of the ducts at twelve hours after birth,
but none appeared in any of the later stages. An occasional
leucocyte may be seen in the lumina at any stage from birth to
ten weeks. In none of the stages studied could fat be demon-
strated in the lumina or the epithelial cells lining the ducts.
The secretion in the new-born or witches’ milk has been ob-
served in human by a large number of investigators. According
to Brouha (’05), Aristotle called attention to it. The writers
of the second half of the nineteenth and the early part of the
twentieth centuries presented different theories concerning the
origin of this secretion. For example, De Sinety (’75), Rein
(82), Barfurth (82), Czerny (’90), Unger (’98), Schalachta (’04),
Brouha (’05), Berka (’11), Schil (12), and others regarded it
in the human new-born as a true milk secretion, while K6lliker
(54), Milne Edwards (’70), and Raubitschek (’04) believed it
is the product of necrobiotic changes which result in the for-
mation of the lumina. Still another view is that of Keiffer (02),
who described the formation of milk in the new-born as a patho-
logical process.
Several theories have been offered to account for the stimulus
which excites the secretion in the new-born human. The fact
that Brouha concluded that the new-born secretion in well-
developed human may completely precede the birth of the child
STUDIES ON THE MAMMARY GLAND 425
and that Schil observed true milk secretion in the ducts of a
human fetus at the beginning of the eighth month, proves fairly
conclusively that a secretion capable of stimulating the glands
to action is formed in the placenta or ovary of the mother and
transmitted to the blood of both mother and offspring, thus
bringing about the formation of milk in both maternal and fetal
mammary glands.
Berka (711) believes the contents of the lumina of the milk-
ducts through the virgin stage is a part of the witches’ milk
which has been retained in the ducts. On the other hand, Schil
(12) thinks this results from a constant slight secretory activity
of the epithelial cells. O’Donoghue (’12), after studying Dasyu-
rus agrees with Schil in the following:
A trace of a secretion somewhat resembling colostrum is always to be
found as a coagulum in the lumen of the tubules and ducts until it is
removed by the more active secretion of colostrum or milk. It would
appear, then, that the gland, quite apart from the proper milk flow, is
the seat of slow secretory activity, although this secretion is quite dif-
ferent in microscopic appearance from true milk.
Aristotle is said to have observed witches’ milk in the goat.
Creighton (’78) reported it in the guinea-pig in the following
manner:
The fluid expressed from the nipple of the new-born guinea-pig had
the appearance of a watery kind of milk; on microscopic examination,
the milk-globules of ordinary milk were not found, but a more uniform
fluid mass irregularly broken up under the cover-slip into large or small
drops, and without any mixture of cellular elements.
Barfurth noticed such a secretion in the guinea-pig, bitch, and
rabbit and regarded it as a coagulum resulting from simple
transudation. After studying it in Vespertilio murinus, rabbit,
and the domestic cat, Brouha (’05) came to the following con-
clusions in regard to these forms; “la sécrétion natale existe
également, mais elle n’aboutit pas, au moins dans les trois pre-
miéres semaines de le vie, 4 une véritable lactation.’ Schil
(12) observed secretion in the ducts of rabbit embryos as well
as postnatal stages. No secretion has been observed in the
ducts of rat fetuses by me. Brouha’s work and the present
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, NO. 4.
426 J. A. MYERS
study lead to the conclusion that in the rabbit, cat, bat, and
white rat the so-called new-born secretion does not contain fat
and hence is not a true lactation as is found in human. This
failure to produce true milk in some of the lower forms may be
due to the feeble development of the mammary gland in the
new-born. The lumina are not completely formed at birth in
the rat, cat, and rabbit, therefore the cells may be unable to
produce a true milk secretion when the stimulus is received.
On the other hand, Brouha found the lumina quite well repre-
sented in Vespertilio murinus at the time of birth. The question
then arises as to whether the stimulus which produces a true
lactation in the human new-born is actually present in the lower
forms of animals. Further experimental evidence is necessary
to decide this question.
Gland stroma
Attention was called to the fact that the stroma in the fetus
is divisible into the mantle layer closely surrounding the ducts
and the true stroma between the ducts. This arrangement of
the stroma persists through the ten weeks’ stage. The mantle
layer increases somewhat in thickness as age advances. ‘This
layer diminishes in thickness as one passes from the primary to
the terminal ducts. About the third week of postnatal life a few
elastic fibres appear in the mantle layer surrounding the primary
and secondary ducts. In later stages elastic fibers are more
numerous and extend further toward the free end of a system of
ducts. The true stroma is at first composed of loose connective
tissue, but soon after birth there appears such a tremendous
invasion of fat that the mantle layer of the ducts is almost en-
tirely surrounded by fat cells.
Free red corpuscles were observed in the stroma twelve hours
after birth, but were not seen in any of the later stages. White
blood-cells were observed in the true stroma in all stages studied,
but in no stage did they appear in greater numbers than they
are usually found in the neighboring tissues.
Berka (’11) called attention to the presence of elastic fibers
in the mantle layer of the virgin human ducts. Schil (12)
STUDIES ON THE MAMMARY GLAND 427
observed elastic fibers in rabbits six months of age and at the
age of puberty.
Red blood corpuscles were observed in the true stroma of the
glands of premature and mature infants by Schlachta (’04).
Brouha (’05) found capillary hemorrhage exists in the stroma of
all infants that died from abrupt asphyxia. He noticed, how-
ever, that in infants dying from other causes there may be a
considerable extravasation of red blood corpuscles. Both of
these authors concluded that the red corpuscles probably reached
the stroma through the process of diapedesis. Owing to the
presence of red blood-cells in the mammary-gland stroma of the
albino rat in comparatively small numbers, it is possible that
they reach the stroma through the process of diapedesis. How-
ever, it must be borne in mind that during parturition and the
removal of the skin there is considerable danger of trauma.
Several authors have reported the presence of an abnormally
large number of leucocytes in the true stroma of the mammary
gland at or soon after birth. Czerny (’90) believed they play
an important réle by entering the lumina of the ducts and carry-
ing the secretion into the lymphatics. Schlachta (04) was un-
able to confirm Czerny’s finding. Brouha (’05), however, found
the immigration of leucocytes in the gland stroma to be less
marked during the activity of the new-born gland than before
or after such activity. The fact that the secretion in the albino
rat is very slight and never becomes a true milk secretion proba-
bly accounts for the absence of an infiltration of leucocytes.
SUMMARY
The results of the present study on the mammary gland of
the male and female albino rat from birth to ten weeks of age
may be summarized briefly as follows:
1. Owing to the thickening of the epidermis over the develop-
ing nipple of the new-born female albino rat, the mammary-
gland area appears lighter than the surrounding epidermis. In
the male no epidermal thickening is present, hence the mammary-
gland area is not visible.
428 J. A. MYERS
The nipple elevation is oblong in shape and very slightly
elevated at the fourth or fifth day. At the end of the second
week it is still oblong, but much more elevated. After the second
week its growth is a gradual one until the time of puberty, when
it takes on the size and shape of the adult virgin nipple.
2. The sulcus around the nipple in the new-born female rat
is apparently a remnant of the prenatal mammary pit. During
the early days of postnatal life this sulcus disappears, but on
the fourth or fifth day an apparently new depression makes its
appearance. This depression deepens so that by the tenth week
there may be developed a definite pocket in which the basal part
of the nipple is located.
3. The epithelial ingrowth (hood) at birth presents a smooth
and quite regular outline. Two or three days after birth, how-
ever, the stratum germinativum becomes somewhat thickened
in some places, thus forming short processes. Such processes
appear on both the outer and inner surfaces of the hood through-
out the stages studied. The central portion of the proximal part
of the hood apparently degenerates, thus deepening the sulcus
around the base of the nipple.
4. Hair follicles are numerous in the integument peripheral’
to the hood and sulcus, but have not been observed nearer the
nipple.
5. Near the highest part of the nipple there appears, a few
days after birth, a slight excavation—the developing milk-pore,
communicating with the intra-epidermal part of the primary
duct. At two weeks there is a slight connection between this
cavity and the lumen of the primary duct. About the sixth
week a complete connection is established with the primary
duct. When the nipple becomes cone-shaped, the milk-pore
appears near the apex.
6. The lumina of the milk ducts are apparently formed by the
process of rearrangement of the cells. The lumen of the primary
duct is not completely formed until after the second week. The
other ducts possess fairly well-developed lumina at the time of
birth.
STUDIES ON THE MAMMARY GLAND 429
7. The walls of the milk-ducts are for the most part lined with
a two-layered epithelium. The inner or glandular layer being
composed of cuboidal to low columnar cells, while the outer (or
so-called myo-epithelial layer) is composed of cells irregular in
size, shape, and arrangement. At nine or ten weeks some of
the terminal processes show indications of developing into alveoli.
The walls of the intra-epidermal part of the primary duct are
lined with stratified epithelium similar to that covering the
surface of the nipple.
8. Masses of subcutaneous fat develop soon after birth. Such
masses Increase in size as age advances, and it is in this fat that
many of the milk-ducts ramify.
9. The stroma is divisible into its usual parts. The mantle
layer forms a thin sheath immediately surrounding the milk-
ducts. At the third week and thereafter elastic tissue fibers
were visible in this layer surrounding the larger ducts. The
true stroma is formed of a loose connective-tissue network in
the early stages, but later shows a very marked infiltration of
fat. Extending through the adipose tissue may be seen an
occasional fair-sized connective-tissue lamina.
10. In no stage was an infiltration of leucocytes observed in
the stroma.
11. A slight secretion appears in the lumina of the milk-ducts
soon after birth. In no case has such secretion appeared in
sufficient quantities to cause distention of the ducts. A trace
of secretion may be seen in the milk-ducts through the ten weeks’
stage. Specific fat stains fail to reveal fat in the secretion of
the milk-ducts or in the epithelial walls, notwithstanding the
fact that much fat was seen in the true stroma outside the walls
of the ducts.
BIBLIOGRAPHY
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430 J. A. MYERS |
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STUDIES ON THE MAMMARY GLAND 431
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quiry into the factors which determine the growth and activity of
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tribution of the milk-ducts and the development of the nipple in the
albino rat from birth to ten weeks of age. Am. Jour. Anat., vol. 19.
(Abstract also published in the Anat. Rec., 1916, vol. 10, p. 230.)
1917 a Studies on the mammary gland. II. The fetal development
of the mammary gland in the female albino rat. Am. Jour. Anat.,
vol. 22. (Abstract also published in the Anat. Rec., 1917, vol. 11,
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1917 b Studies on the mammary gland. III. A comparison of the
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late fetal stages to ten weeks of age. Anat. Rec., vol. 13. (Abstract
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1918 The histology of the mammary gland in the male and female
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Mamma des Neugeborenen. Arch. f. mikros. Anat. u. Entwick.
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SrernHAus, Juttus 1892 Die Morphologie der Milchabsonderung. Arch. f.
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Heft.
vonEpner 1902 Handbuch der Geweblehren des Menschen von Kélliker, Bd.3.
PLATE 1
EXPLANATION OF FIGURES
12 External view of wax model reconstructed from the right third thoracic
gland of a female albino rat of five days. Reconstruction built by A. H. Nerad.
ep., epidermis; m., nipple eminence; s., sulcus. X 37}.
13 External view of a wax model reconstructed from the right second in-
guinal gland of a female albino rat of two weeks. Reconstruction built by A. F.
Branton. ep., epidermis; m.po., milk-pore; n., nipple; s., sulcus. X 37%.
14. Internal view of a wax model reconstructed from the right second inguinal
gland of a female albino rat of two weeks. Reconstruction built by A. F. Bran-
ton. ep., epidermis; ep.in., epithelial ingrowth; p.d., primary duct. X 373.
15 External view of a wax model reconstructed from the right second in-
guinal gland of a female albino rat of six weeks. Reconstruction built by Nellie
Pederson and Frieda Radusch. ep., epidermis; m.po., milk-pore n., nipple; s.,
sulcus. X 373.
16 External view of a wax model reconstructed from the right abdominal
gland of a male albino rat of six weeks. ep., epidermis; m.po., milk-pore. X
373.
17 External view of a wax model reconstructed from the right first inguinal
gland of a female albino rat of nine weeks. Reconstruction by M. H. Litman
and A. F. Branton. ep., epidermis; m.po., milk-pore; n., nipple; s., sulcus. X
OTs.
432
STUDIES ON THE MAMMARY GLAND PLATE 1
J. A. MYERS
433
PLATE 2
EXPLANATION OF FIGURES
18 Drawn from a section through the right first inguinal gland of a female
albino rat at birth. Zenker’s fixation; Mallory’s connective-tissue stain. ¢.,
corium; d.f., developing fat cells; ep., epidermis; h., hair follicles; m.b., muscle
bundle; m.d., milk-ducts; t.s., tela subcutanea. X 83}.
19 Drawn from a section through the right first inguinal gland of a female
albino rat of nine weeks. Zenker’s fixation; Mallory’s connective-tissue stain.
b., blood vessel; c., corium; c.t., connective-tissue lamina; ep., epidermis; f., fat
cells; h., hair follicles; m.l., mantle layer; m.d., milk-ducts. X 833.
434
STUDIES ON THE MAMMARY GLAND PLATE 2
J. A, MYERS
435
Resumen por el autor, Harvey Ernest Jordan.
Histologia de la sangre y médula 6sea roja en la rana leopardo,
Rana pipiens. .
El autor susministra pruebas de que el homélogo del megalo-
eariocito de los mamfferos no es la célula fusfforme de los an-
fibios sino una célula gigante mononucleada que se deriva, como
el megalocariocito de los mamfferos, de un hemoblasto. Todos
los tipos de leucocitos, incluso las células gigantes, pueden formar
pseudépodos, los cuales pueden segmentarse para formar cuerpos
semejantes a plaquetas, granulares o hialinos. Los que tienen
erdnulos metacromiaticos son idénticos a las plaquetas de los
mamiferos. Tales corpusculos se producen también por un
proceso de fragmentacién del protoplasma de un leucocito o un
trombocito. La formacién de plaquetas es, aparentemente, una
produccién secundaria de la actividad normal de los leuco-
citos, la cual se expresa en la formacién y segmentacién de pseu-
dépodos y la degeneracién de estas células. La hematopoiesis
tiene lugar segun el modo monofilético. Los leucocitos neutré-
filos de nucleo polimorfo contienen un sistema astral bien patente,
cuyo centrosoma adopta diversas formas y puede ser sencillo,
doble o multiple; pero estas células no se dividen por mitosis en
su condicién de nucleares polimorfas. El autor discute la im-
portancia de estos hechos sobre la significacién de la amitosis.
Translation by José F. Nonidez
Columbia University
AUTHOR’S ABSTRACT OF THIS PAPER ISSUED
BY THE BIBLIOGRAPHIC SERVICE, JUNE 2
THE HISTOLOGY OF THE BLOOD AND THE RED
BONE-MARROW OF THE LEOPARD FROG,
RANA PIPIENS
H. E. JORDAN
Laboratory of Histology and Embryology, University of Virginia
TWO PLATES (SEVENTY-THREE FIGURES)
CONTENTS
esa MOND oe tips 3 (sone eckcnuiin boos for hore ie a anor bale ashes Ge Be Rene Es eae aes ae 437
Cer ee TGLOOOR Nest We AEs lacie oa aodlecs Rue RE Be A 4 ete tectee 438
PRRs aN ROTM ae pac cLace's urs ole ace Seca eee eke ee TR Rte CL an 439
ai Lhe histology ‘of the circulating blood)! )..0)0...2.. 00/0850. Rts 439
Pee ELE EM URTOP VES. 444). tare) Sunfss ys dyatowe oa Pacheco aos Bete 439
PAL VATU CV LES 8 \02/5 tae by Abbe e. q.es.s de Ss cape Be oe 439
ae eae POsinopnilic: leucacytes.% zc ss<. =. + 445,50 «8.0 WU ROS hs ee 441
4- ihe. basophilic leucocytes. //2 6. 6o7 oA Le ie 442
cpne MCULFOpHilic IEUCOGYLES... 1. 5. « Leis. satan unten ed dase eon e. 442
Gop tiie. ChmOMmOCVLER sn. poeknte i auars'sy 5s eae rene SSO Hetrecih kee 443
beeitne histology of the red bone-marrow... ..... 2 ..,02.ccsa cade deen une de. 445
1, The development of the erythrocytes......................4 Petes oes 446
2: The development of the lymphocytes. :... 2. 29.0.0. 2h Jcci aonb wee 447
3. The development of the eosiniplfilic leucocytes.................0. 448
4, The development of the basophilic leucocytes.................... 449
5. The development of the neutrophilic leucocytes................... 450
®,, he development of the thrombocytes..;. 0.2... 2 seat. sss tee 451
7. The development of the plasma-cells and giant-cells............... 451
Pireminuiand Conclusions 3a <2 sqia ceo pili ee a ere als Ae eae oe 452
SSUNLETIAOTAS AAR Ve ee eM SEER Par RAR dete cent eae eee eRe S Hicecrs och ORs Aone ee ee 471
INTRODUCTION
This investigation was begun with the study of the bone-
marrow. Special interest in this tissue was first aroused through
a previous investigation” of the giant-cells of the red bone-
marrow of certain mammals. The primary object of this work
concerns the identification of the amphibian homologue of the
hemogenic giant-cell of mammals, the so-called megakaryocyte.
437
438 H. E. JORDAN
Since the latter cell, under certain conditions, becomes the source
of the blood-platelets, interest centers also on the amphibian
homologue of the mammalian platelet.
It became apparent early in this investigation that a confi-
dent interpretation of the developmental stages of the hemal
elements of the marrow demanded a precise knowledge of the
types of cells of the circulating blood. Accordingly, blood and
marrow were: studied coincidentally and with the same tech-
nics. As regards the blood, the cells of special importance in
this connection are the polymorphonuclear neutrophilic leuco-
cytes and the spindle cells, both containing similar metachro-
matic (azurophil) cytoplasmic granules.
In the red marrow the origin and development of these same
elements claim first attention. Incidentally must be con-
sidered also the developmental history of the eosinophilic and
basophilic leucocytes, the origin of the lymphocytes and of the
erythrocytes, the genetic relationships among the several series
of blood-cells and the bearing of these data on the prevalent
monophyletic theory of blood-cell origin.
MATERIAL AND METHODS
The species of frog used in this work is Rana pipiens. Smear
preparations of blood were stained according to Wright’s technic.
The frogs employed had been left over from the early fall, kept
in cages until the middle of January. Blood smears were made
in October, in January, and in April. The bone-marrow was
taken from the femurs of frogs killed in January, in a certain
number of which the shaft was well filled with red marrow.
The marrow was fixed in a mixture of 100 parts of a saturated
normal-salt solution of corrosive sublimate and 10 parts of forma-
lin. Paraffin sections were cut at 54 and stained according to
Wright’s® technic. This technic gave very excellent results.
BLOOD AND BONE-MARROW OF FROG 439
DESCRIPTION
a. The histology of the circulating blood
Though the study was begun in the reverse order, it seems
preferable to pass from a description of the blood to that of the
marrow. The cellular elements include the following types
which will be described in the order enumerated: erythrocytes,
lymphocytes, eosinophilic leucocytes, basophilic leucocytes,
neutrophilic leucocytes, and thrombocytes.
1. The erythrocytes. 'The typical elliposidal, centrally nucle-
ated, discoid erythrocytes of amphibia are well known and need
no special description for this species of frog. However, certain
atypical forms, comparable to those previously specified for cer-
tain turtle bloods, should again be noted, namely, larger, smaller,
and senile types and their transition forms. The largest type,
comprising a relatively small number of cells, is approximately
one and a quarter times the average size; the smallest type, like-
wise including only a few cells, may be less than half the average
size; these are generally stoutly oval with a spheroidal nucleus
situated nearer one pole; the senile type (fig. 3) is relatively
abundant in the frogs kept under laboratory conditions over
winter; this cell has a large, spheroidal, lilac-colored nucleus and
an expansive shell of very faintly-staining cytoplasm. The
typical erythrocyte (figs. 1 and 2) has a much smaller oval nu-
cleus which stains deeply blue, and a greenish-yellow cytoplasm.
Account must of course be taken of the fact that in smear prep-
arations the cells are spread out under varying pressures and
tensions which operate to produce definitive size variations;
nevertheless, the above-enumerated atypical forms occur in
some degree. ‘This classification agrees in general with that of
Werzberg” except that he makes no mention of the senile forms.
2. The lymphocytes. Under this head may be listed a relatively
large group of mononuclear leucocytes, varying greatly in size,
but with the same nuclear and cytoplasmic characteristics. The
smallest are but slightly larger than the nucleus of a thrombo-
cyte, with a scarcely perceptible shell of basophilic cytoplasm
(fig. 4); the largest (fig. 9) have a diameter approximately twice
44() H. E. JORDAN
that of the smallest, with a larger nucleus and a more expansive
shell of cytoplasm. All the various sizes of lymphocytes have a
common type of nucleus, which stains deeply as a whole, due to
a coarse close-meshed reticulum with many net-knots. It
assumes a deep lilac color. Grouped according to. size, the
lymphocytes may be classified as large and small, but abun-
dant transition forms occur (figs. 6 to 10). Their resemblance to
mammalian lymphocytes is striking.
The most characteristic features of these lymphocytes are
their apparently granular cytoplasmic content and their numer-
ous pseudopods (figs. 5 and 7). Various distortions, nuclear
as well as cytoplasmic, demonstrate that these cells are extra-
ordinarily delicate. The cytoplasm is extremely viscid. Some
of the ‘pseudopods’ are most probably artifacts. But that
at least many should not be so interpreted is sufficiently indi-
cated by the fact that these same cells, or their ancestors, in the
sections of bone-marrow likewise are covered with abundant
pseudopods. For the same reason certain isolated portions of
such pseudopods (fig. 7) must be regarded as actual constrictions
or segmentation products.
In the bone-marrow the lymphocytes have a homogeneous
basophilic cytoplasm (figs. 34 to 40); in the blood smears they
appear to have a coarsely granular basophilic cytoplasm; the
‘oranules’ are very irregular in form and size and frequently
appear massed into clumps (fig. 9). It seems possible that the
granules in question are in fact fixation artifacts, that is, proteid
coagula of the basophilic cytoplasm. However, certain of the
lymphocytes in the blood contain also metachromatic granules
scattered throughout the basophilic substratum. These would
seem to correspond with the ‘leucocytoid lymphocytes’ of Werz-
berg, which this author regards as a distinct class. The evi-
dence seems to indicate that the potentiality to form metachro-
matic granules is a common property of the lymphocytes. There
is no good reason, in my opinion, for classifying lymphocytes on
the basis of the presence of ‘azurophil’ granules. ‘Lymphocytes’
comprise a group of cells characterized by common nuclear and
fundamental cytoplasmic (basophilic) ¢haracteristics; they vary
BLOOD AND BONE-MARROW OF FROG 44]
greatly with respect of size, and the presence or abundance of
metachromatic granules. Moreover, certain of the pseudopods,
especially isolated portions, contain masses of this basophilic and
metachromatic granulation. Such bodies resemble very closely
the blood-platelets of mammals.
There is abundant evidence, which will be detailed below, to
show that the various blood-cells undergo further growth and
development in passing from the blood spaces of the marrow to
the general circulation; in view of which it may be inferred that
the non-granular lymphocytes of the marrow elaborate meta-
chromatic granules after entering the blood stream.
The nucleus of the lymphocyte likewise suffers a considerable
change during the passage of the cell from the marrow to the
circulation. ‘The nucleus of the medullary lymphocyte is vesic-
ular; it contains a distinct plasmosome, several deeply chromatic
karyosomes, many smaller chromatic granules ranged mainly
along the nuclear membrane, and a very delicate chromatic
reticulum (figs. 35 and 37). In general appearance it is much
clearer and lighter staining than the nucleus of the circulatory
lymphocyte.
The attraction sphere of the lymphocytes is usually masked
by the basophilic granulation. Occasionally it appears very
conspicuously as a spheroidal clear area containing a centrosome
or diplosome (fig. 10).
While the evidence at hand does not permit of a final conclu-
sion regarding the genetic relationship between lymphocyte and
platelet, the striking resemblance of the lymphocyte pseudopods
and their constriction products to mammalian blood-platelets is
unmistakable.
3. The eosinophilic leucocytes. This type of cell will be fully
discussed below in connection with the description of its myelo-
cyte ancestors. It will suffice here to state that in its adult con-
dition it contains a polymorph or, in rare instances, a multiple
nucleus, generally located at one pole of the spheroidal cell. Oc-
casionally a mononucleated form may occur. Figure 11 shows a
binucleated form. The spheroidal eosinophilic granules are of
approximately uniform size. They appear hollow, or annular,
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, No. 4
442 H. E. JORDAN
due probably to difference in density between center and pe-
riphery. The granules are imbedded in a lightly basophilic
substratum. Eosinophils constitute only a very small propor-
tion of the leucocytic content of the frog’s blood. In the winter
specimens the eosinophils are rarer than the basophils; in the fall
specimens they are relatively more numerous.
4. The basophilic leucocytes. This type of cell is somewhat
smaller than the eosinophil. It is approximately the size of the .
medium-sized lymphocyte (fig. 12). Its nucleus is centrally
located and vesicular. The nucleus contains a very delicate
reticulum. The cytoplasm is closely packed with spherical baso-
philic, lilac-colored granules, larger on the average than the
eosinophilic granules, but varying somewhat in size and shape,
certain granules having an oval form. In the marrow these
cells have a deeply staining nucleus (figs. 51 to 53) and the cyto-
plasmic granules take a deep blue color with Wright’s stain.
5. The neutrophilic leucocytes. This is by far the most abun-
dant type of leucocyte (figs. 13 to 26). Its nucleus is generally
polymorphous. Mono- and binucleated forms also occur. The
cytoplasm consists of a homogeneous basophilic (blue-staining)
substratum, throughout which are scattered fine metachromatic
(neutrophilic) granules. The granules are always very small,
but vary in size from a scarcely perceptible dust to very definite
spheroidal granules of light lilac color. The resemblance to the
polymorphonuclear neutrophilic leucocytes of mammals (com-
pare figs. 22 and 23) is striking. In areas where the neutro-
philic granules are sparse, the basophilic (blue) substratum also
appears granular.
The most conspicuous feature of these cells, aside from their
characteristic polymorphous nucleus, is the attraction sphere.
‘This is generally situated within the concavity of the lobulated
nucleus (figs. 13, 16, 18, and 19). The neutrophilic granules are
arranged in lines radiating from the sphere (fig. 17). The attrac-
tion sphere may consist of a spherical mass of minute granules
imbedded in a denser protoplasm (figs. 16, 21, 23, and 25) or it
may occur as a Clear area (centrosphere) containing centrally -a
deep-staining granule or centrosome (centriole) (figs. 13, 17, 18,
BLOOD AND BONE-MARROW OF FROG 443
and 19). The latter may be simple, bilobed, or double (diplo-
some). These cells were never seen in mitosis in the circulating
blood. The persistence of such a definite and conspicuous at-
traction sphere in a cell whose nucleus undergoes successive
constrictions in the formation of a polylobular condition (figs. 13
to 21) is of much theoretical interest, a point which will be dis-
cussed below in connection with its myelocytic history.
The relationship between the two types of attraction spheres
is not perfectly clear. However, they correspond closely with
similar types occurring in other cells, e.g., maturing eggs of
Cummingia,? where the clear sphere with its central granule
-(centrosome) of earlier stages becomes converted into a granular
darker sphere through division of the centrosome. In these
leucocytes of the frog the monosome and diplosome occur more
commonly in those cells with the less complex nucleus, the finely
granular spheres in those with the more complex polymorphous
nucleus (compare fig’. 13 and 19 with fig. 25). Though figures
16 and 24 show that this relationship is not invariable, it may be
said that no cells with nuclei like that of figure 13 were seen with
the granular type of sphere. The inference seems justified that
here also the granular type of sphere arises from the clear type
with diplosome, by repeated division of the centrosome. The
granular type would thus represent a disintegration or partition
product. The bearing of this conclusion will be discussed in
connection with the description of these same features of the
medullary neutrophilic leucocytes, where also these same two
types of attraction spheres occur.
6. The thrombocytes. These cells typically are stout fusiform
elements with central, deeply staining (violet), dense, oval
nucleus (figs. 28 and 30). The cytoplasm stains only very faintly
(pink) and contains metachromatic spherical granules of uni-
form size. These granules appear identical with the larger
granules of the neutrophilic leucocytes. The granules are gen-
erally aggregated more or less closely into small groups. The
cytoplasm is apparently extremely viscid and is drawn out at
numerous points into pseudopod-like projections (figs. 27 and
31). These contain the majority of the granules. Certain of
444 H. E. JORDAN
these pseudopods appear to constrict off spheroidal . granular
fragments, resembling blood-platelets of mammals (figs. 30 and
31.) The extreme viscosity of the thrombocyte cytoplasm
would seem to account fully for the fact that these cells generally
occur in larger and smaller closely adherent groups. One line
of evidence casts doubt upon the interpretation of the projec-
tions of the thrombocytes as pseudopods and the isolated bodies
as segmentation products: The cytoplasm is unmistakably very
delicate and very viscid; in contrast with all of the leucocytes of
the marrow, the spindle cells of the marrow do not show pseudo-
pods. This would seem to indicate that the ‘pseudopods’ of the
thrombocytes are artifacts, formed through the operation of
mechanical factors in the process of making the smears. On the
other hand, numerous naked and disintegrating nuclei of throm-
bocytes occur in the blood (fig. 32); these recall the naked nuclei
of the megakaryocytes of mammalian marrows, and in a meas-
ure support Wright’s suggestion that the megakaryocytes of
mammals and the spindle cells of amphibia are homologous
elements.
The resemblance between the granular cytoplasm of the mega-
karyocytes of mammals and that of the thrombocytes of am-
phibia is very close; but it appears no more close than between
this cytoplasm and that of the neutrophilic leucocytes. More-
over, the constriction products of the pseudopods of thrombo-
cytes, neutrophilic leucocytes and certain lymphocytes of the
frog, and those of the pseudopods of the megakaryocytes of
rabbit and guinea-pig are very similar. These corpuscles consist
in common of a spheroidal viscid mass of faintly basophilic
cytoplasm containing a central group of fine metachromatic
spherical granules. The question of homology will be further
discussed below.
At this place must be noted the character of the nucleus. In
the smear preparations this appears dense and chromatic (deep
lilac) with numerous irregular vacuoles. In the marrow this
nucleus is more oval, it contains a chromatic reticulum with
numerous karyosomes, it stains much less deeply and it takes a
blue color (figs. 42 and 43). Moreover, it shows several deep,
BLOOD AND BONE-MARROW OF FROG 445
approximately longitudinal grooves; certain of these are so deep
as to appear to completely divide the nucleus. Similar condi-
tions were previously reported also in blood smears of certain
turtles.1! Downey* questions the accuracy of this interpretation
and inclines to regard all these phenomena as simply deep fur-
rows. One could hardly base final conclusions regarding this
point on smear preparations; the process of spreading the blood
on the slide might very readily divide the already deeply con-
stricted (grooved) nucleus. But similar appearances occur also
in the marrow sections. However, here it could always be
argued that in such cases the plane of section passed above the
bottom of the groove, thus giving the deceptive appearance of a
division of the nucleus. Against the interpretation that certain
of the thrombocyte nuclei are actually split must be cited the
fact that these cells in the circulating blood of the frog contain
generally a more compact, more nearly spherical and ungrooved
nucleus (figs. 27 to 32). If a certain groove of the nucleus of
the immature thrombocyte of the marrow is conceived to lead to
a direct division of the nucleus, then certain mature thrombo-
cytes should be binucleated, which is apparently never actually
the case in the frog. On the contrary, the mature nucleus seems
to have lost the grooves of its immature condition, apparently
through a process of dilatation of the nuclear vesicle, involving a
change from oval to spherical form and an obliteration of the
grooves by reason of a filling up of the nucleus.
b. The histology of the red bone-marrow
This description is based almost exclusively on the bone-mar-
row of the shaft of the femurs of an adult specimen killed the
middle of January. This particular specimen had an especially
abundant red marrow in its femurs. Practically the entire shaft
was packed with red marrow. Other specimens examined at
the same time and during the following month, as also specimens
from a new shipment in April, showed only small patches of red
marrow in the shaft of the femur. Sections were made of this
essentially yellow marrow and also stained according to Wright’s”®
446 H. E. JORDAN
technic. This tissue proved helpful for comparisons in the
course of this study, but the hemopoietic sequences were worked
out on the one specimen with the abundant red marrow. In
the frog the process of blood-corpuscle formation is said to occur
only in the early summer, at which time only the bones contain
red marrow (Starling). The specimen under consideration had
presumably simply anticipated his fellows in this process of con-
verting the yellow into red marrow.
The reticular stroma of the red marrow is packed centrally
with fat cells, blood-vessels, and hemogenic vascular spaces
(angiocysts), and covered peripherally by a layer of differentiat-
ing leucocytes. Erythrocytes are seen only centrally within the
vascular spaces.
The development of fat cells from the mesenchymal stroma
can be traced through a complete series of stages. The process,
however, presents nothing new except that a certain number of
practically adult fat cells contain two nuclei. Endothelium,
erythrocytes, lymphocytes, and leucocytes can likewise be traced
through a complete series of developmental stages back to the
marrow mesenchyma. Hemopoiesis is an essentially similar
process, involving the formation of ‘blood-islands,’ in yolk-sae
and red marrow. The spindle cells arise only intravascularly
as differentiation products from small lymphocytes and from
endothelium. Endothelium may also produce hemoblasts sec-
ondarily.
It seems preferable to describe the developmental history of
the blood-cells in the marrow in the following order: erythro-
cytes, lymphocytes, eosinophilic leucocytes, basophilic leuco-
cytes, neutrophilic leucocytes, and thrombocytes.
1. The development of the erythrocytes. This history can be
read in the smaller blood spaces of the central portions of the
red marrow. The larger spaces and blood-vessels contain only
more mature erythrocytes, mingled with the granular leucocytes
of extravascular origin. The erythrocytogenic process passes
from a stage of solid strands of hemoblasts (marrow ‘blood-
islands’) to one of forming vessels in which the central cells are
free mature erythrocytes and the peripheral cells represent
BLOOD AND BONE-MARROW OF FROG 447
various stages of the differentiation from original hemoblasts.
A certain number of the latter are in intimate connection with
the superficially differentiating endothelium. Endothelial cells
and blood-cells have differentiated from the same primitive cell
mass, and subsequently for a time young endothelium may give
origin to hemoblasts, including thromboblasts.
The original progenitor of the erythrocyte is a cell with a rela-
tively large vesicular nucleus and a narrow shell of slightly baso-
philic eytoplasm. The nucleus is granular in character (figs. 37
and 38). At first irregular in shape, i.e., polyhedral or even
fusiform, the hemoblast soon assumes a spheroidal form. These
cells proliferate mitotically. In becoming an erythrocyte the
hemoblast (erythroblast) again changes into a stoutly oval form
(fig. 41), the cytoplasm becomes oxyphilic as it elaborates hemo-
globin, and the nucleus becomes more compact and more
chromatic.
The great difference in size between the red cells of the marrow
and those of the circulating blood is surprising (compare figs. L
and 41). Those of the smear preparations are approximately
twice the size of those in the sections of the marrow. A certain
large amount of this difference may be accounted for on the basis
of a spreading out in the process of making the smear, but the
residue can only be explained in terms of continued growth after
leaving the marrow. The same phenomenon of postmedullary
growth is evident in some degree in the case of all of the other
types of blood-cells except the basophilic leucocytes (compare
figs. 12 and 452).
2. The development of the lymphocytes. As in the circulating
blood, so in the marrow two main types of lymphocytes can be
distinguished, differing in no essential nuclear or cytoplasmic
features, but only in size (figs. 34 to 41). Transition forms
occur abundantly. These intravascular lymphocytes are struc-
turally indistinguishable from similar extravascular cells from
which the granulocytes develop (compare figs. 88 and 41). Fur-
thermore, the smaller varieties correspond with the hemoblasts
from which erythrocytes, and in part thrombocytes, develop
intravascularly. Intravascularly, the original hemoblast is more
448 H. E. JORDAN
generally of the smaller lymphocyte type with granular vesicular,
nucleus; this may grow into the larger type. Extravascularly,
the original hemoblast is more generally of the large lymphocyte
type (figs. 34 and 35); this may divide mitotically to form small
lymphocytes. These marrow types correspond with the large
and small lymphocytes of the blood smears, except that in the
latter the originally basophilic homogeneous cytoplasm of the
marrow cells becomes granular‘and may in addition elaborate
metachromatic granules. Similarly, the small lymphocytes form
metachromatic granules in becoming thrombocytes within the
blood spaces of the red marrow.
3. The development of the eosinophilic leucocytes. ‘The eosino-
philic myelocytes arise in the extravascular connective tissue.
Their progenitor is indistinguishable from the large and medium-
sized medullary lymphocytes. The mature eosinophils second-
arily enter the vascular spaces, and continue their development
during their passage into the peripheral circulation (compare
fig. 49 with fig. 11). The first indication of the beginning of
differentiation of a ‘lymphocyte’ into an eosinophilic myelo-
cyte is the appearance of a few, very minute, purplish-red gran-
ules (figs. 36 and 44). The first granules are most commonly
aggregated in a certain restricted region, generally in the vicinity
of the centrosome, and only gradually appear in all portions of
the cytoplasm. They increase gradually in number and in size,
always maintain a fairly uniform size for any particular stage
(figs. 45 and 46), and during their later phases resemble those of
the circulating eosinophils in that their centers are less dense,
giving them the appearance of rings (figs. 47 to 50). The gran-
ules apparently grow by a process of swelling involving, beside
increase of diameter of the granules, a rarefaction especially of
_the core, and a slight general decrease in staining capacity.
The original eosinophilic granule has a purplish-red color, the
definitive form a slightly orange-red color. This material gives
no indication of the primitive basophilic granules described by
Downey® for the eosinophilic myelocytes of the guinea-pig.
Nor does either the developing or the definitive eosinophilic
leucocyte contain intranuclear eosinophilic granules, as claimed
BLOOD AND BONE-MARROW OF FROG 449
by Niegolewski!® for Rana esculenta. We are in like disagree-
ment with Niegolewski regarding the basophilic granules of the
mast-cells. Nor is there any evidence that the original eosino-
philic granules of the myelocytes have a nuclear origin, nor any
that they have an extracellular origin, as claimed by certain
investigators, e.g., Weidenreich.2t The eosinophilic granules
arise gradually within the cytoplasmic area, apparently as a
result of some specific activity of this protoplasm.
Coincident with the above-described changes in the granules,
the nucleus also passes through a series of structural and slight
tinctorial alterations. At first the nucleus is approximately
spherical, centrally located, contains a distinct plasmosome, and
is vesicular in character (figs. 44 to 48). It gradually moves
excentrically, meanwhile showing a coarser network and more
numerous karyosomes, and a modification in shape leading
through a reniform to a polylobular, and eventually, in some
cases a multiple, character. Only the younger forms with
spherical or stoutly reniform nucleus are seen in miototic divi-
sion. In common with all myelocytes, including the lympho-
cytes, these cells also form pseudopods which may constrict and
fragment to form free ‘hyaline bodies’ (fig. 36) or, very rarely,
globules with eosinophilic granules. Pseudopod formation and
segmentation seem to be a common property of leucocytes, both
circulatory and medullary.
4. The development of the basophilic leucocytes. 'The basophilic
leucocytes or mast-cells likewise take origin from the common
lymphocyte progenitor, of medium size, in the extravascular
tissue (figs. 38, 39 and 51). The nucleus is characteristically
deep-staining, apparently homogeneous except for a few, barely
visible, large nucleoli (fig. 52). The granules are at first small,
but always larger than the eosinophilic granules, and stain very
deeply blue (fig. 53). The granules become coarser, meanwhile
maintaining their deep coloration. These same cells in the
blood smears have a lighter-staining, centrally located nucleus,
and their basophilic granules show a violet or lilac tinge (fig. 12).
The nucleus of this cell in the marrow, due to its deep-staining
and apparently homogeneous character, suggests degeneration.
450 H. E. JORDAN
However, if this nucleus were properly interpreted thus, it could
scarcely attain a more normal appearance in the circulating
blood. It appears that these cells undergo a further differen-
tiation in passing from marrow to the circulation. They are the
rarest type of leucocyte, but are most probably to be inter-
preted as normal and specific blood elements, as maintained by
Maximow.'® There is no evidence that the granules are nuclear
extrusions, nor that they result from a mucoid degeneration of
the cytoplasm. As in the case of the eosinophilic granules, they
appear to represent the result of some metabolic activity of the
cytoplasm.
5. The development of the neutrophilic leucocytes. These cells
are in certain respects the most interesting among the blood
elements of the frog. It is only in the light of their origin in the
marrow that their true significance can be determined. Such
study shows that they correspond much more closely to the
polymorph neutrophils of mammals than to the leucocytes with
special eosinophilic granules of sauropsida, rabbit, and guinea-
pig. They are in fact the amphibian homologues of the neutro-
philic leucocytes of certain mammals (compare figs. 21 and 22).
These cells alse originate from the common lymphocyte an-
cestor (figs. 34, 35, and 54). The first indication of their differ-
entiation is the appearance of an oxyphilic halo about the
centrosome in the otherwise basophilic cytoplasm. This halo
becomes finely granular and spreads in radiating fashion toward
the periphery of the cell (fig. 54). This disposition of the neu-
‘trophilic granules in radii is, maintained in the definitive forms
(figs. 17 to 26). The granules vary somewhat in size and in the
degree of their lilac coloration, but are always smaller than the
eosinophilic granules. The basophilic substratum remains vis-
ible, and is variably conspicuous in different regions. Coin-
cident with the differentiation of the metachromatic granules,
the nucleus undergoes great morphologic changes, passing ulti-
mately into a polylobular (figs. 69 and 70), and occasionally a
polynuclear, condition (figs. 66 to 68). Only the earliest types,
in which the nucleus is still spheroidal or reniform (fig. 54),
divide by mitosis. The same statement can be made for the
BLOOD AND BONE-MARROW OF FROG 451
eosinophils; basophils were not seen in division. A conspicuous
and significant feature of these cells is the abundance and length
of their granular pseudopods, certain of which may be seen seg-
menting or fragmenting into faintly basophilic globules with
granular centers, simulating thus very closely the blood-platelets
of mammals in their structure and in their origin from mega-
karyocytes (figs. 62 and 68). However, no naked nuclei could
be found. The cells form common lyin the extra vascular tissue
(fig. 69), but a few may possibly arise also from hemoblasts
within the developing blood spaces of the marrow (fig. 41).
6. The development of the thrombocytes. ‘These cells only arise
intravascularly, from small lymphocytes, and in small part
directly from endothelium. As they take on their definitive
oval or fusiform shape they develop metachromatic granules.
In their passage into the circulation they undergo further nuclear
and slight dimensional changes as described above. They are
never seen to arise extravascularly, nor do they undergo divi-
sion; and the nucleus never assumes the crescentic lobulated
condition characteristic of certain leucocytes. In the marrow
they occur singly, in the blood smears generally in groups. The
blood smears show thrombocytes with pseudopods, certain of
which apparently constrict to produce platelets, as first described
by Wright?§ for Amblystoma, leaving eventually disintegrating
naked nuclei (figs. 31 and 32).
7. The development of the plasma-cells and giant-cells. Certain
large lymphocytes undergo a type of differentiation leading to
typical plasma-cells. These plasma-cells are characterized by
the coarse chromatic reticulum of their deep-staining nucleus,
their irregular shape, and their very faintly basophilic, exten-
sively vacuolated cytoplasm (fig. 71). This observation agrees
with Downey’s® conclusion regarding the chief source of origin of
the plasma-cells from lymphoid cells in the mesentery of Rana.
_ A small number of lymphocytes undergo also a hypertrophy
leading to mononuclear giant-cells. These cells contain a rela-
tively enormous nucleus very like that of the younger lympho-
cytes, and a variable shell of basophilic cytoplasm containing
many fine metachromatic (lilac) granules (figs. 72 and 73). Con-
452 H. E. JORDAN
stricting pseudopods of such cells produce bodies comparable
with the blood-platelets of megakaryocyte origin in mammalian
marrows.
DISCUSSION AND CONCLUSIONS
This search for the amphibian homologue of the mammalian
giant-cell from which the blood-platelets take origin has revealed
two types of cells which in a measure fulfill requirements, namely,
the polymorphonucleated neutrophilic leucocytes and the throm-
bocytes. That the so-called megakaryocytes of red bone-
marrow of mammals (e.g., of rabbit and guinea-pig) do actually
at certain stages liberate blood-platelets can be abundantly
demonstrated by the Wright” technic. These mammalian cells
are commonly polymorphonucleated, occasionally polynucleated.
Their faintly basophilic cytoplasm contains an abundance of fairly
uniform metachromatic granules. These cells are not phagocytic,
their occasional content of a leucocyte or two, generally eosinoph-
ilic, is probably to be interpreted as an invasion following early
stages of degeneration. Under certain conditions these cells are
erythrocytopoietic. ‘The mammalian megakaryocyte also traces
its origin to the common lymphocyte (hemoblast) ancestor of
the blood-cells, both in the yolk-sac and in the red marrow.
In both locations the nuclear characteristics are very similar,
and the apparently identical granular cytoplasm produces com-
parable blood-platelets in an identical manner (Jordan). More-
over, in the yolk-sac the smaller, usually binucleated, type of
these giant-cells may differentiate into erythrocytes (Jordan).”
Wright?’ has suggested that the thrombocytes of Amblystoma
represent the megakaryocytes of mammalian red marrow. But
the thrombocyte resembles the megakaryocyte only in respect of
its metachromatic granules scattered throughout a lightly baso-
philic cytoplasm, and its elimination within the blood of gran-
ulated globules similar to platelets, leaving eventually a naked
nucleus. Within the bone-marrow it apparently does not lib-
erate ‘platelets,’ in which respect it contrasts sharply with the
mammalian giant-cells. Moreover, these cells differ greatly in
regard to nuclear characteristics. The thrombocytes resemble
BLOOD AND BONE-MARROW OF FROG 453
platelets in respect of a very adhesive protoplasm, in consequence
of which they frequently become grouped into larger and smaller
masses. But this property of adhesiveness is characteristic also
especially of the lymphocytes, certain of which also have a
variable amount of metachromatic granules.
The polymorphonucleated neutrophilic leucocytes of the frog
seem at first to bear a much closer resemblance to mammalian
megakaryocytes. They are of course smaller than the mega-
karyocytes. While not of the extreme ‘basket’ form, the nu-
cleus is frequently extensively lobulated. The centrosome is
always conspicuous and frequently multiple, as described by
Heidenhain for megakaryocytes. These cells contain, moreover,
a very similar metachromatic granulation, which is likewise
scattered through a basophilic substratum, the latter forming
an exoplasmic layer of variable width. The granules of the
amphibian neutrophilic leucocytes stain less deeply than those of
the mammalian megakaryocytes, but otherwise they are very
similar. Moreover, these leucocytes protrude pseudopods, which
fragment to form platelet-like bodies, like those of megakaryo-
cytes (figs. 61 to 68). These pseudopods even project into
blood-vessels, as do those of the megakaryocytes (fig. 63). The
presence of these alleged giant-cell homologues in the amphibian
blood, as compared with the restriction of the megakaryocytes
of mammals to the bone-marrow, might be due simply to the
fact of the great size of the latter prohibiting entrance into the
capillary circulation. It would seem on the basis of the histo-
logic evidence that the neutrophilic leucocyte of amphibia meets
more nearly the requirements of a megakaryocyte homologue
than does the thrombocyte.
However, the fact that no naked nuclei of these neutrophilic
leucocytes occur, while such are numerous of thrombocyte origin,
contravenes in a measure the assumption of homology between
the polymorphonuclear neutrophilic leucocytes of the frog and
the megakaryocytes of the marrow of the rabbit. In addition to
this objection, there is the further contradiction that certain
mammals have a red marrow containing both typical megakary-
ocytes and typical polymorphonuclear neutrophilic leucocytes
(e.g., cat, dog).
454 H. E. JORDAN
On the other hand, the common property of pseudopod for-
mation and constriction possessed by all the types of leucocytes
within the marrow casts doubt upon the specific nature of the
process as restricted to megakaryocytes of mammalian marrow.
In the frog marrow, primitive lymphocytes, eosinophilic leu-
cocytes and hemoblasts produce ‘hyaline’ bodies by this
method of pseudopod ‘segmentation.’ The neutrophils produce
granulated bodies, resembling platelets. Basophilic leucocytes,
and to some extent eosinophiles, likewise produce granulated
globules. The evidence seems to indicate that this property is
common to both lymphocytes and granulocytes, and that plate-
let-like bodies are formed only incidentally. A thorough study
of the giant-cells of rabbit and guinea-pig also leads to the con-
clusion that these cells produce platelets to some extent by a
constriction of pseudopods, but chiefly by a process of fragmen-
tation of large cytoplasmic areas of degenerating giant cells.
The degeneration is indicated chiefly by the irregular and pyc-
notice character of the nucleus. Similarly, in the case of throm-
bocytes and leucocytes with metachromatic granules, degenera-
tion involves a fragmentation of the cytoplasm and the incidental
formation of platelet-like bodies.
In this connection must be considered the question whether
the cells above described as polymorphonuclear neutrophils are
actually such or only types of non-granular leucocytes, as main-
tained by Werzberg.2> Werzberg’s failure to interpret these
cells as granulocytes must be ascribed to his disregard of the
Wright or a similarly favorable technic. Studied with the
Wright technic, the metachromatic granular content is con-
spicuous and indubitable. This conclusion agrees with the
earlier one of Niegolewski!® and the later one of Downey.*°
Downey® made a special study of the polymorphonucleated
leucocytes of the amphibian Amblystoma. He employed only
smear preparations; these were stained with Wright’s blood
stain. He describes their granules as ‘azurophil,’ which he
regards as close to the ‘special’ granules of the higher animals,
i.e., the neutrophilic granules of the polymorphs of certain mam-
mals. He agrees with Werzberg®* that the lymphocytes of most
BLOOD AND BONE-MARROW OF FROG 455
other amphibia, including the frog, lose their azurophil (i.e.,
neutrophilic, metachromatic) granules when they differentiate
into polymorphonuclears, and maintains that the cytoplasm of
' the definitive polymorphs of these forms is oxyphilic. Neu-
mann!’ likewise denies the presence of granules in the poly-
morphonuclear leucocytes of the frog. However, my prepara-
tions of frog marrow very clearly show that the neutrophilic
myelocytes retain and increase their ‘azurophil’ granules as they
differentiate into the definitive forms of the circulating blood.
Downey* states further that all possible intermediate stages
between larger lymphocytes (with neutrophilic granules) and
neutrophilic polymorphs occur in the circulating blood of Am-
blystoma. This finding is at variance with that of Maximow"™
in the case of Axolotl.
In view of the fact that the complete developmental history
of the polymorphonucleated neutrophilic leucocytes from non-
granular basophilic primitive lymphocytes can be traced in the
sections of the red marrow, Downey’s interpretation of transition
forms between ‘definitive’ lymphocytes and neutrophilic granu-
locytes in the circulation of Amblystoma at first seems quite
improbable. The nucleus of the circulatory lymphocytes is
very different from that of the medullary lymphocytes (compare
figs. 4 dnd 37), a change which indicates progressive differentia-
tion. However, a careful study of the different types of circula-
tory lymphocytes in the frog forces the conclusion that these
lymphocytes do actually metamorphose into the neutrophilic
granulocytes, as urged by Downey* for Amblystoma, but denied
for the frog. Moreover, the nuclei of the medullary and circu-
latory neutrophils and of the circulatory lymphocytes are prac-
tically identical in their fundamental features. It would seem
that a lymphocyte with an already considerably differentiated
nucleus may develop neutrophilic granules abundantly and so
pass over into a neutrophilic granulocyte whose nucleus may
subsequently undergo lobulation. Such a developmental proc-
ess is illustrated in figures 8, 13, 16, and 19.
Figure 8 is a typical lymphocyte with a reniform nucleus.
The only perceptible cytoplasmic difference between it and a
456 H. E. JORDAN
lymphocyte like that illustrated in figure 6 pertains to the
relative proportion of neutrophilic and basophilic granules. The
designation ‘basophilic granules’ in connection with lymphocytes
is always used here with the reservation that the ‘granules’ may
be actually a coagulation phenomenon in an essentially homoge-
neous cytoplasm. Certain circulatory lymphocytes contain only
a few neutrophilic granules; these types lead through transition
forms to neutrophilic granulocytes which show many granules and
interspersed small areas of only basophilic granular material.
The conclusion seems inescapable that lymphocytes may, and
continually do, differentiate into polymorphonucleated neutro-
philic granulocytes within the circulation. This would seem to
dispose of Werzberg’s” classification of the lymphocytes of frog
as nongranular large and small lymphocytes, ‘leucocytoid lym-
phocytes with azurophil granulation,’ and ‘lympholeucocytes.’
In the frog the lymphocytes differ in their cytoplasmic features
only in respect of the relative abundance of neutrophilic (azuro-
phil) granules. This conclusion has an important bearing upon
the discussion regarding the validity of the monophyletic the-
ory of hemopoiesis. The circulatory lymphocyte, the slightly
modified persistent medullary lymphocyte which functions as
the common progenitor (hemoblast) of all types of blood-cells,
still maintains its capacity to differentiate at least into a neutro-
philic granulocyte.
The question then arises whether the Amphibian neutrophils
are actually the homologues of the mammalian megakaryocytes
or of the mammalian polymorph neutrophils. If these cells are
regarded as representing polymorph neutrophils of higher mam-
mals, then amphibian blood, except for thrombocytes, would
seem much closer to mammalian than to sauropsid bloods. This
would conflict with the accepted phylogenetic seriation. How-
ever, neither neutrophilic leucocytes nor recognized hemogenic
giant-cells occur in sauropsid marrows, while leucocytes with
‘special’ (eosinophilic, ellipsoidal) granules are abundant, facts
which favor Downey’s® interpretation. The evidence seems to
point to the homology of the amphibian and mammalian poly-
morph neutrophils. Whether both are the homologues of the
BLOOD AND BONE-MARROW OF FROG 457
‘special leucocytes’ with minute or ellipsoidal eosinophilic granules
characteristic of those forms which lack neutrophils, namely,
certain mammals (e.g., rabbit, guinea-pig) and sauropsids, respec-
tively, is a question of a different order, and one that need not
here be further discussed.
Having thus disposed of both these alleged amphibian homo-
logues of the mammalian megakaryocytes, we may analyze the
possibilities of still another type of cell. This cell occurs only
sparsely. It contains a relatively enormous nucleus, with one or
several plasmosomes, numerous irregular karyosomes, and a
delicately reticulated vesicular nucleoplasm (fig. 73). The
nucleus is enveloped with a variable shell of lightly basophilic
cytoplasm containing metachromatic, lilac-colored granules. It
may be assumed that this cell forms pseudopods which may con-
strict and become free corpuscles. This amphibian giant-cell
develops from a primitive lymphocyte and represents a hyper-
trophied hemoblast. In all these respects, then, it corresponds
with the mononucleated giant-cells of the red marrow of the
femur of the rabbit. It has been shown that the polymorpho-
nucleated giant-cells, the so-called megakaryocytes, develop
from mononucleated giant-cells through nuclear modifications
(Jordan). It seems that in the frog the marrow giant-cells
develop only to the mononucleated stage. Possibly at certain
periods this marrow also would show later polymorphonucleated
phases. The usual statement that giant-cells are not found in
the red marrow of forms below the mammals must therefore be
revised. Careful study of the marrow of sauropsida may pos-
sibly also reveal giant-cell homologues of even closer corre-
spondence.
Another important body of evidence supplied by this material
concerns the monophyletic theory of blood-cell origin. Maxi-
mow? has published a brief preliminary report on hemopoiesis
in Rana temporaria, and interprets his evidence in accord with
this theory. His material includes larvae and the adult bone-
marrow. As regards the process in the latter, his statements
are especially brief. However, the chief points are touched.
My results agree completely with his earlier findings except in
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, NO. 4
458 H. E. JORDAN
one important respect. Maximow claims that in Rana tempo-
raria lymphocytes arise only in the blood-vessels. He notes
that this is a significant departure from what occurs in the case °
of selachians, birds, and mammals, where the lymphocytes form
both extra and intravascularly, and he suggests that this differ-
ence may inhere in ontogenetic differences associated with holo-
blastic and meroblastic cleavage. My preparations of the
marrow of Rana pipiens show, however, that in this species
lymphocytes arise in both locations. If a difference actually
exists in the case of Rana temporaria it must be a specific dif-
ference or a chance variation in Maximow’s specimens and not
one characteristic of amphibia as a group. Maximow” further
calls attention to the fact that in amphibia the lymphoid and
myeloid tissues are not sharply separated topographically, there
being here no sharply defined lymphoid organs with the specific
function of lymphocyte production, like the lymph nodes of mam-
mals, and another tissue with specific granulopoietic function.
This mingling of lymphoid and myeloid tissues and functions in
the bone-marrow gives additional support to the monophyletic
theory.
The erythrocyte, lymphocyte, and leucocyte series in common,
can be traced through gradual steps of development from a type
of cell which is indistinguishable in the several series, namely, a
lymphocyte-like cell, the ‘hemoblast.’ The determining differ-
entiation factor seems to be exclusively environmental. ‘Lym-
phocytes’ that become enclosed by endothelium differentiate
into erythrocytes, or in small part they may proliferate or per-
sist as lymphocytes which undergo slight differentiation in pass-
ing into the blood stream, or some may differentiate into
thrombocytes. Furthermore, the already slightly differentiated
endothelium may to some extent during its younger stages further
differentiate into erythroblasts and into thromboblasts. Con-
sideration of the common origin of the endothelium and the
hemoblasts, from the original medullary ‘blood-islands,’ makes
such differentiation intelligible. Neutrophilic leucocytes may
also to some extent differentiate from intravascular hemoblasts
and from circulatory lymphocytes. But extravascular condi-
BLOOD AND BONE-MARROW OF FROG 459
tions seem more favorable for the neutrophilic differentiation in
the case of the majority of the lymphocytes. Eosinophilic and
basophilic granulocytes, apparenthky differentiate only extra-
vascularly. These enter the blood-channels secondarily through
ameboid activity.
Danchakoff? calls attention to an apparent contradiction in
the argument of the supporters of the monophyletic theory of
blood-cell origin: they note that morphologically identical mes-
enchymal cells and hemoblasts in the same limited regions
develop into both erythrocytes and granulocytes, and infer
from this fact the equipotentiality of these blood-cell ancestors.
Danchakoff argues that if these ‘stem-cells’ develop into differ-
ent products in the same region, then they must have had dis-
similar potentialities and were in fact originally distinct, as
claimed by the adherents of the polyphyletic theory. But this
argument must postulate an identity of the environmental
factors playing upon the hemoblasts in these restricted areas;
this involves an assumption which cannot be supported by tan-
- gible data. It is not at all inconceivable that two adjacent,
identically endowed, cells are nevertheless under environments
sufficiently dissimilar to determine erythropoiesis in one case
and granulopoiesis in the other case. It is no doubt generally
true, as abundant recorded observations show, that hemoblasts
enclosed by endothelium develop into erythrocytes, while ex-
travascular hemoblasts differentiate into granulocytes. But
Danchakoff‘ herself observed in regions in the allantois of the
chick embryo upon which had been grafted particles of adult
spleen, that extravascular hemoblasts could develop into eryth-
rocytes. She, however, interprets her sections to mean that
the extravascular hemoblasts in these instances had received an
erythropoietic bias while previously confined by endothelium,
which could not be reversed under the influence of the new
environment, or that the extravascular spaces containing these
displaced hemoblasts are actually in continuity with the orig-
inal lumen of the blood-vessel through breaks in the endothelial
wall. However, there are other instances in which such explana-
tions will not suffice. For example, in the area vasculosa of the
460 H. E. JORDAN
young turtle embryo I have seen an eosinophilic granulocyte
within an early blood-island, otherwise differentiating largely
into erythrocytes. Until it-can be actually demonstrated that
the environment is identical in such instances, these facts do not
contradict, but sustain the monophyletic view.
Unless we assume an identity of developmental potencies in
the case of the hemoblastic mesenchymal cells, one is forced to
the position that the mesenchymal ancestors of fat cells, pig-
ment cells, endothelium and smooth muscle cells also differ in
respect of specific developmental potencies. But such unquali-
fied position must ignore a large group of histogenetic data.
Mesenchyme cells are apparently of identical nature and en-
dowed with multiple potencies, that is, ‘equipotential’ and
‘polyvalent.’ The expression of any one type of mesenchymal
differentiation—whether as endothelium, erythrocytes, granu-
locytes, or fat—seems accordingly dependent upon extrinsic
factors of a type or degree not yet detectable or measurable by
our means of investigation. Indeed, Danchakoff describes endo-
thelial cells of the spleen graft separating from the wall of small
blood-vessels both centrally, where they pass into the lumen as
hemoblasts and differentiate into erythrocytes, and peripherally
where they become reincorporated with the mesenchyma and
may redifferentiate into granuloblasts. I haye observed com-
parable phenomena in the bone marrow and the body mesen-
chyma of the turtle embryo. These observations show that the
same cell, an endothelial hemoblast, may become either an
erythrocyte or a granulocyte according as the environmental
stimuli change ever so slightly. They furnish, moreover, the
very strongest support to the monophyletic theory. But they
show also that adjacent identically differentiated cells, namely,
young endothelial cells, suffer divergent further differentiation
in an apparently identical environment, namely, inclusion in an
endothelial wall with luminal and mesenchymal surfaces. But
since the cells are identical, i.e., mesenchymal cells slightly
differentiated into functional young endothelal cells, then the
environment of extrinsic stimuli determining the direction of
migration and the initial steps in the further differentiation into
BLOOD AND BONE-MARROW OF FROG 461
different varieties of blood corpuscles, must indeed have differed
to some extent, however slightly.
An interesting feature of this marrow concerns further the
fact of a more or less sharp grouping of the several types of cells
at the various phases of development, very much as is the case
in the red marrow of the femur of the pigeon and of the rabbit
during stages of intense hemopoiesis, that is, a certain group of
cells comprises predominantly small lymphocytes, another
large lymphocytes, another neutrophilic leucocytes, and another
eosinophilic leucocytes. Intravascularly, developing erythro-
blasts predominate; among these are intermingled lymphoblasts,
thromboblasts, and neutrophilic leucoblasts.
Here again it might be argued in favor of the polyphyletic
theory that since the general environment is apparently the
same, the progenitors of the several groups were cells with dif-
ferent and specific developmental potencies, in spite of their
apparent morphologic identity. But in view of the fact that
relatively slight environmental differences apparently determine
developmental differences in early ontogeny, as, for example, in
the developing gut of the mammalian embryo where smooth
muscle, connective tissue, and blood develop in the same re-
stricted regions, it seems more in accord with histogenetic data
to conclude that the several regions filled with different segre-
gated groups of cells were under the influence of different meta-
bolic (fundamentally perhaps only relational, both temporal
and spatial) factors, which determined the specific type of
development. -
This brings us to the question of the development of the
eosinophilic leucocytes. The red marrow of the frog offers an
especially favorable material for the study of the developmental
history of the eosinophilic granulocytes. Unequivocal histologic
evidence accrues with respect to the debated questions regarding
the origin of the cells, whether heteroplastic or homoplastic;
source of the eosinophilic granules, whether endogenous or exog-
enous; the alleged basophilic ‘unripe’ eosinophilic granules, and
the genetic relationship between mast-cells and eosinophilic
leucocytes. An enormous literature of conflicting opinion and
462 H. E. JORDAN
conclusions exists covering these disputed points; this need not
be here again reviewed; it is fully covered in a number of recent
works (e.g., Downey,’ Ringoen?!).
The superior feature of the frog material pertains primarily to
the absence of special eosinophilic leucocytes. The presence of
such types—with fine eosinophilic granules, as in guinea-pig and
rabbit, and ellipsoidal eosinophilic granules, as in sauropsida and
certain fishes—greatly confuses the picture in these forms. In
the marrow of the frog the neutrophilic ‘special’ granulocytes
and the eosinophils are clearly distinguishable from their earliest
stages of development from the common lymphocyte ancestor.
The neutrophilic granulocytes show at first an oxyphilic halo
about the centrosome, which early becomes granular, the granu-
lation spreading throughout the entire cell, the granules always
being of a lighter or darker lilac color. The eosinophils, on the
other hand, begin their differentiation from the parent lympho-
cyte by the elaboration of a few minute deep-staining (purple)
eosinophilic granules. These gradually increase in number and
size, and assume a more brilliantly red (or orange-red) color.
No basophilic granules (i.e., blue-staining) were seen among the
eosinophilic granules at any stage of the development of this
type of granulocytes. The earliest granules are, it is true, of
purple-red color and apparently less oxyphilic than the definitive
granules, but there is no evidence of a differentiation of definitely
basophil granules into eosinophil granules. The definitive gran-
ules, moreover, have a lighter-staining (less dense) center, giving
the appearance of a ring-shaped structure. This appearance is
due to a rarefaction of the center of the granules coincident with
its growth. The growth of the granules, and their change into
more intensively oxyphilic granules is apparently due largely to
a dilution of their substance, specially pronounced in the center.
Downey® comes to the conclusion, on the basis of his study of
the marrow of the guinea-pig, that the eosinophilic granules
develop from basophilic predecessors. His illustrations show
deep blue-staining granules among eosinophilic granules in the
eosinophilic myelocytes. Such blue-staining granules never
appear in the eosinophilic myelocytes of frog in my material;
BLOOD AND BONE-MARROW OF FROG 463
they are present only in the mast-cells. However, I find abun-
dant examples of just such cells as depicted by Downey in sec-
tions of young turtle embryos in the periaortic connective tissue.
It must be emphasized that the fixing and staining fluids in
these instances. (turtle embryo, Jordan; guinea-pig, Downey)
were the same, namely, Helly’s fluid followed by the Giemsa
stain (Downey employed also other similar stains but not
Wright’s combination). In the case of the frog’s marrow the
fixation was secured with a corrosive-sublimate-formalin mixture
and the staining was done with Wright’s stain. It seems prob-
able that a difference of appearance, as concerns presence or
absence. of blue-staining granules among the red granules in
eosinophilic myelocytes depends upon the type of fixing fluid and
staining combinations employed. In the frog material under
consideration, it can only be said that less oxyphilic granules
precede more oxyphilic granules, and that the former are always
smaller than the latter.
All the evidence in this material, moreover, clearly points to a
heteroplastic origin of these cells from lymphocyte ancestors,
and the endogenous origin of the eosinophilic granules. Only
the younger types of myelocyte, i.e., with non-polymorphous
nucleus, are capable of mitotic division and thus of forming new
eosinophilic myelocytes. The granules are in no case ingestion
products of hemoglobin-containing fragments of disintegrating
erythrocytes, but result from the specific activity of the myelo-
cyte protoplasm. Moreover, the nucleus is never invaded by
these cytoplasmic granules, as maintained by Niegolewski,'* nor
is there the slightest evidence in support of Weidenreich’s*
theory that eosinophilic granules are the ingested fragmenta-
tion products of erythrocytes. In this material the erythrocytes
develop only intravascularly, where they also fragment; the
eosinophils, on the other hand, develop only extravascularly.
All the evidence inclines towards the interpretation of these
granules as the product of a specific cytoplasmic activity.
The chief question regarding the basophilic leucocytes (mast-
cells) is whether they represent a specific type of normal blood-
cell or whether they represent degenerations of lymphocytes or
464 H. E. JORDAN
other leucocytes. This study does not touch the disputed ques-
tion of the genetic relationship between the so-called histogenous
and hemotogenous types of basophilic leucocytes. The litera-
ture on this subject has been very fully reviewed in the recent
papers of Maximow,'® Downey,’ and Ringoen.?®* This study is
concerned only with the hemotogeneous type of mast-cells. My
results are in accord with those of the above-mentioned authors,
who conclude that in the mammals investigated (including
guinea-pig and rabbit) the mast leucocytes represent a specific
‘and independent line of granulocytes, of heteroplastic origin,
with basophilic granules resulting from a specific activity of the
protoplasm.
In the marrow of the frog the first indication of mast-cells
‘appears in the nuclear modification of the lymphocyte parent,
namely, a relatively greater contraction and condensation of the
central spheroidal nucleus, giving a deep-blue coloration and a
more homogeneous character to this body. Coincident with
these early nuclear alterations, minute basophilic (deep blue-
‘staining) granules appear; these granules are, however, from the
beginning larger than the ancestors of the eosinophilic granules.
“They grow in size, exceeding that of the definitive eosinophilic
granules, and maintain their original deep blue coloration.
These cells also liberate granulated globules through segmenta-
tion of pseudopods. As found in the blood, these cells show a
centrally located, more vesicular nucleus, a somewhat larger
size, and cytoplasmic granules of deep lilac color. There is not
the slightest indication of a transformation of these basophilic
‘granules into eosinophils at any stage, nor of any degeneration
process connected with the formation of the granules. None of
these cells were seen by me in mitosis at any stage. The cells
apparently originate only heteroplastically from parent lym-
phocytes, and form their granules by an endogenous process.
These observations agree with those previously reported for the
basophilic granulocytes in the turtle embryo.!!' Not only can
both eosinophils and basophils be traced to the same type of
lymphocyte, but the smaller granules of the youngest basophils
are always considerably larger than the smallest eosinophilic
BLOOD AND BONE-MARROW OF FROG 465
granules, and the nucleus of the mast-cells is at all stages after
the earliest quite different from that of the eosinophils. The
presence of these cells in the smear preparations in considerable
numbers renders inadmissable their interpretation in terms of a
disintegration of other types of cells. The deep blue-staining
granules of the basophilic myelocytes change to a violet color as
seen in blood smears. The nucleus also changes from a deep-
staining homogeneous body to a vesicular body with a delicate
reticulum. It maintains an approximately central position in
this cell throughout its history.
The most perplexing matter regarding the thrombocytes con-
cerns their origin. In a study of hemopoiesis in the turtle,
Jordan and Flippin" have described their origin in part from the
endothelium of. the original vascular spaces of yolk-sac and red
marrow and in part from small lymphocytes. Danchakoff?
claims that they arise exclusively from lymphocytes (throm-
boblasts), while Werzberg?> maintains that they are a type sui
generis, having origin neither from endothelium nor lympho-
cytes. The evidence from this study of the red marrow of the
frog confirms our former conclusion regarding their origin, as
described in a study of turtle embryos. The thrombocytes
arise only intravascularly. They can be traced directly to
small lymphocyte-like cells. Secondarily, both lymphocytes
and thrombocytes may arise from endothelium. The close
relationship of small lymphocytes and thrombocytes appears
throughout their entire history. This is especially emphasized,
aside from the close similarity in structure of the nucleus, by the
presence in certain of the small lymphocytes of a variable quan-
tity of metachromatic granules.
The mode of multiplication of the constituent cells of this
marrow centers interest upon amitosis and throws additional
light on the question of this variety of cell division. The orig-
inal blood-cell progenitor (lymphocyte; hemoblast) arises from a
mesenchymal cell. Relatively little proliferative activity occurs
among these mesenchymal cells; the multiplication of hemoblasts
results mainly from a division of already differentiated mesen-
chymal cells. This differentiation process consists essentially in
466 H. E. JORDAN
the rounding up of an irregular, generally stellate cell (fig. 33),
and a condensation and increase in basophily of the cytoplasm
(fig. 34). The nuclei of the typical mesenchymal cells and the
derived hemoblasts are practically identical. These nuclei are
of a vesicular lightly-staining character, with a pale nucleolus,
several deeply chromatic karyosomes, and many very minute
chromatic granules scattered among a very delicate chromatic
reticulum. This description fits also the smaller types of hemo-
_blasts, both intra- and extravascular. In general, further dif-
ferentiation into the various types of blood-cells involves first an
increase in the number of larger, more regular karyosomes, so that
the nucleus has a coarsely granular appearance, many granules
lying peripherally upon the more robust, chromatic, nuclear
membrane. ‘The plasmosome meanwhile persists. Later stages
are characterized mainly by increase in the general chromaticity
of the nuclear sap, giving the entire nucleus, now with a coarser
reticulum, a deep blue coloration. Only the larger lympho-
cytes divide mitotically; the smaller lymphocytes do not divide
as such; they may grow to larger size and then divide mitotically.
Excessive growth of the large lymphocytes leads to giant-cells
(fig. 73). Both neutrophilic and eosinophilic leucocytes also
divide mitotically during their earlier stages, while the nucleus
remains of spheroidal shape. Similarly, young spheroidal eryth-
roblasts may divide mitotically within the blood-vessels.
Neither basophilic leucocytes nor thrombocytes were seen in
either mitosis or amitosis.
Besides proliferation by mitosis, large lymphocytes, young
erythroblasts, and young neutrophilic leucocytes also divide
amitotically. What determines whether a lymphocyte or eryth-
roblast shall divide directly or indirectly remains obscure;
possibly the two modes of division are determined by divergent
metabolic conditions as expressed in the nucleo-cytoplasmic
relationship. However, the amitotic division of the neutro-
philic leucocytes, of which the extreme lobulation of the later
nucleus must probably be reckoned a phase, is the more surpris-
ing, since here an astral system is maintained intact and is at all
stages conspicuous. Possibly here the mitotic incapacity of the
BLOOD AND BONE-MARROW OF FROG 467
centrosomes is related to the high stage of differentiation as
expressed fundamentally in the neutrophilic granules. This
suggestion is in harmony with the facts that the later eosino-
philic myelocytes, the basophilic myelocytes, the thrombocytes,
and the giant-cells also do not divide mitotically. All of these
cells express in their peculiar cytoplasmic condition a high degree
of differentiation. The reason for the failure of basophilic
myelocytes, thrombocytes, and giant-cells fot the most part,
to proliferate even amitotically is under this view not clear.
However, if we assume that the lobulated nucleus of the neutro-
philic myelocytes represents an unfinished amitosis, then pos-
sibly the bilobed nucleus of the basophilic myelocytes, the deeply
grooved condition of the thrombocyte nucleus, and the poly-
morphous character of the giant-cell nucleus (in mammals) may
legitimately be similarly interpreted.
The nuclear amitosis of the neutrophilic myelocytes and the
amitosis of the hemoblasts (large lymphocytes and erythroblasts)
are apparently similar phenomena resulting from quite different
causes— cytoplasmic specificity or high differentiation and intense
proliferative demands, respectively. These dissimilar causative
-factors may, however, be brought under a common head
as regards their effect on the potency and integrity of the
kinetic center, the centrosome. Both conditions may be con-
ceived to reduce relative nutritive conditions below the possi-
ble minimum for centrosomal activity. In essence, amitotic
proliferation, in contrast with mitotic proliferation, results
presumably when the metabolic demands of the protoplasmic
mechanism are such as to deprive the astral system of its
minimum nutritive requirements. This assumption can at
least harmonize the apparently contradictory facts that amitosis
occurs in the cells of rapidly growing tissues as well as in highly
specialized and degenerating cells, an idea first suggested by
Child.!
In this connection attention must again be directed to the two
main types of astral systems in the neutrophilic granulocytes:
one clear with a central monosome or diplosome, the other con-
sisting of a larger or smaller granular sphere. As described
468 H. E. JORDAN
above, these types correspond with those found in the earlier
and later stages respectively, of the first maturation spindle of
the Cummingia tellinoides egg, where the pluricorpuscular is
derived from the unicorpuscular sphere, and represents a dis-
integration or partition product.* This conclusion respecting
the significance of the pluricorpuscular centrosphere is sup-
ported by the facts that the second maturation spindle of Cum-
mingia has almost invariably this type of sphere and that the
segmentation spindles again show spheres of both types. It
would seem a legitimate inference that the two types of centro-
spheres of the neutrophilic leucocytes of frog bear to each other
the same genetic relationship, and that the pluricorpuscular
variety also signifies disintegration. The medullary neutrophils
support this inference; certain examples with lobulated nuclei
contain a granular sphere (fig. 56), but here the sphere is smaller,
the granules less numerous and more conspicuous. It seems
very probable that the mitotic incapacity on the part of these
cells results from an untoward influence upon the centrosome
due to a relative lack of nutritive materials following the main-
tenance of the high degree of specialization involved in the elab-
oration of metachromatic granules.
In a paper dealing with amitosis in the cells of the ioonee
efferentes of the testis of the mouse, 4° suggested that the ami-
totic division of these cells was a consequence of the loss of the
integrity of the centrosome through partition into the basal
granules which give rise to the cilia. While confirming the
observation that ciliated cells of vertebrates do not multiply by
mitosis, but may divide amitotically, Saguchi” claims to be
able to demonstrate the presence and integrity of the centrosome
in ciliated cells of vertebrates, and the origin of the basal gran-
ules and the cilia from mitochondria. Apart from the fact that
his illustrations are far from convincing, both as regards identi-
fication of the centrosome and the genetic relationship between
mitochondria and cilia, the divergence of such a process from the
known functional behavior of mitochondria in general renders
his claims dubious. The recent investigations on mitochondria
have demonstrated that these cytoplasmic elements have no
BLOOD AND BONE-MARROW OF FROG 469
direct genetic relationship to such structures as nerve, muscle
or connective-tissue fibrils, but are intimate cytoplasmic con-
stituents most probably subserving general cell metabolism, not
specific differentiations. On the other hand, we have the very
suggestive fact that the axial filament of the flagellum of the
sperm (comparable to a coarse cilium) does grow out from one
of the partition products of the centrosome of the spermatid.
The fact that a centrosome can still be detected in a ciliated cell
is not disproof that some of the original partition products served
as basal bodies for the development of the ciliary apparatus.
Not all of the partition products need have been thus employed;
several might have remained as discernible granules near the
central portion of the cell. Nevertheless, Saguchi’s” general
conclusion that the ‘‘occurrence of amitosis in ciliated cells is
not owing to the lack of centrosome,” but is ‘due essentially to
the degree of differentiation of the cell-plasm”’ (p. 262) is not in
contradiction with my earlier suggestion that the primary com-
mon cause of amitosis is some deleterious influence of variable
type upon the centrosome. This might be narcotization, rela-
tive lack of sufficient materials to meet metabolic demands as in
conditions of very rapid growth or differentiation, lack of suff-
cient supply of oxygen, or the presence of toxic substances as in
degenerating or pathologic tissues.
The theory that can in my opinion best harmonize the appar-
ently contradictory observations that amitosis occurs in rapidly
growing tissues, degenerating tissues, highly specialized cells
(e.g., secretory, ciliated, granulocytes) and in tissues grown
under experimental conditions (e.g., root tips grown in water
with ether) is one expressed in terms of primary influence upon
the centrosome, effecting either a loss of morphological integrity
as by partition in ciliated cells and certain leucocytes, or loss of
specific physiologic capacity as in narcotized, degenerate, or se-
eretory cells. The effective factor may in the latter case still be
fundamentally a disturbance of the optimum nucleo-cytoplasmic
relationship (producing a nutritive want, subsequently affecting
the centrosome), as recently suggested by Nakahara!’ for ‘se-
cretory or reserve-forming cells,’ where he concludes that amitosis
470 H. E. JORDAN
in adipose cells of insects ‘‘may be for the purpose of securing an
increase of the nuclear surface to meet the physiological neces-
sity due to the active metabolic interchange between the nucleus
and cytoplasm” (p. 509). Even in cases where the original
centrosome has fragmented, presumably under the influence of
cytoplasmic specialization, the effective factor in determining
amitotie division may still in part be the metabolic condition
following a certain degree of nucleo-cytoplasmic balance. That
the fragmentation of itself is not in all cases effective seems
proved by the case of the mitotically dividing blastomeres of
the Cummingia embryo in which pluricorpuscular centrosomes
abound. In the ciliated cells of vertebrates a partitioned cen-
trosome, under the metabolic conditions underlying the type of
differentiation characteristic of a cell developing cilia, is gen-
erally incapable of supporting mitotic division; in the more
vigorous blastomeric cells of Cummingia, characterized by pre-
sumably different metabolic conditions, such a centrosome may
still be able to function in indirect division.
Probably the most important result of this search for the am-
phibian homologue of the mammalian giant-cell turns out to be
the light shed upon the significance of the hemogenie giant-cells,
especially the so-called megakaryocytes of mammals, and upon
the morphologit and genetic significance of the blood-platelets.
Though both thrombocytes and polymorphonucleated neutro-
philic leucocytes show certain characteristics in common with
the mammalian megakaryocytes, the genuine amphibian homo-
logue is a very large mononucleated cell, comparable with a
similar giant-cell of mammalian marrow from which the poly-
morphonucleated megakaryocytes develop. These cells are the
homologues also of the mono- and polymorpho- and polynucle-
ated hemogenic giant-cells of the mammalian yolk-sac during its
hemopoietic phase.!2 These cells in all of these locations develop
from the primitive lymphocyte or hemoblast. This study of
frog’s marrow has shown that pseudopod formation and constric-
tion is a characteristic common to leucocytes at all stages of their
history. Moreover, it is well recognized that cytoplasmic frag-
mentation is a concomitant of degeneration. The processes by
BLOOD AND BONE-MARROW OF FROG 471
which blood-platelets arise from megakaryocytes in mammalian
marrow thus appear to be coincidences of these two cytoplasmic
properties of active and degenerating leucocytes respectively.
This conclusion is further strengthened by the fact that the hemo-
genic giant-cells of the yolk-sac (e.g., 12-mm. pig embryo) like-
wise produce typical platelets in abundance."
SUMMARY
1. The blood of the leopard frog contains the following cellular
elements; erythrocytes, thrombocytes, large and small lympho-
cytes, and neutrophilic, eosinophilic, and basophilic granulocytes.
2. Thrombocytes, neutrophilic granulocytes, and lympho-
cytes contain a variable quantity of metachromatic granules.
Both thrombocytes and lymphocytes show conspicuous granular
pseudopods which may constrict to form platelet-like bodies.
3. The polymorphonuclear neutrophilic leucocyte of the frog
resembles the cell of this designation in certain mammalian bloods.
It is characterized especially by its conspicuous astral system,
its extremely lobulated nucleus, and the presence of metachro-
matic granules arranged in lines radiating from the centrosphere.
4. The red bone-marrow contains the following types of
myelocytes; erythroblasts and thromboblasts, only intravas-
cularly: lymphoblasts, both intravascularly and extravascularly ;
granular myelocytes, including neutrophilic, eosinophilic, and
basophilic cells. All of the leucocytic series show pseudopods,
which may constrict to form ‘hyaline’ bodies and granular
platelet-like corpuscles. Certain lymphocytes differentiate also
into plasma-cells and giant-cells extravascularly.
5. All the types of myelocytes can be traced back to a similar,
apparently identical, progenitor, a lymphocyte-like cell or hemo-
blast arising from the mesenchyma. The evidence from this
material is wholly in accord with the monophyletic theory of
blood-cell origin. Environmental conditions are apparently the
chief factors which determine the line of differentiation a certain
hemoblast shall take. Cords of hemoblasts enclosed by endo-
thelium produce erythroblasts and thromboblasts, or persist in
472 H. E. JORDAN
part as lymphoblasts. The extravascular hemoblasts develop
into lymphocytes and granulocytes, which may enter the vas-
cular spaces secondarily. The lymphocytes of the circulating
blood are apparently only slightly modified marrow lympho-
blasts or hemoblasts, which occur both intra- and extravas-
cularly. A certain number of neutrophilic leucocytes also
originate intravascularly. Circulatory lymphocytes also may
differentiate further into neutrophilic leucocytes.
6. Pseudopod constriction and cytoplasmic fragmentation of
leucocytes are two fundamentally distinct processes leading to
similar results, namely, the production of free cytoplasmic
globules. Pseudopod formation and constriction is a common
property of leucocytes; fragmentation is a degeneration phe-
nomenon associated with nuclear pycnosis and subsequent dis-
integration. Lymphocytes and eosinophilic leucocytes produce
the hyaline bodies; neutrophilic granulocytes, thrombocytes,
and certain lymphocytes with metachromatic granules produce
platelet-like bodies. Globules with basophilic granules arise
from pseudopods of mast-cells. Platelet formation from mega-
karyocytes in mammalian red bone-marrow is apparently a by-
product of this common property of leucocytes and their deriva-
tives, and especially of the disintegration of senile types of these
cells.
7. The amphibian homologue of the mammalian hemogenic
giant-cell is a large mononucleated cell with a relatively large
nucleus, comparable to the similar mononucleated giant-cell of
mammalian marrow from which develop the polymorphonu-
cleated (‘megakaryocyte’ with ‘basket nucleus’) and multinu-
cleated older types. Both cells differentiate from a hyper-
trophied primitive lymphocyte or hemoblast.
8. The polymorphonucleated neutrophilic leucocytes contain
a conspicuous centrosphere which may include a simple, a bi-
lobed, a double, or a multiple centrosome. These cells do not
divide mitotically. It is suggested that mitotic incapacity on
the part of these cells is the result of a relative nutritive want in
consequence of the high degree of specialization involved in the
elaboration of the metachromatic granules. The underlying
BLOOD AND BONE-MARROW OF FROG 473
metabolic demands are conceived to effect an untoward influ-
ence upon the kinetic center, a morphologic aspect of which is
expressed in the pluricorpuscular variety of centrosome.
10
11
13
14
15
16
LITERATURE CITED
Cuitp, C. M. 1907 Studies on the relation between amitosis and mitosis
Biol. Bull., vol. 12, nos. 2, 3 and 4.
Dancuakorr, VERA 1910 Uber die Entwicklung der embryonalen Blut-
bildung bei Reptilien. Bd. 37 (Erginzungsheft).
1918. Cell potentialities and differential factors considered in relation
to erythropoiesis. Am. Jour. Anat., vol. 24, p. 1.
1918. Equivalence of different hematopoietic anlages (by method of
stimulation of their stem-cells). II. (Grafts of adult spleen on the
allantois and response of the allantoic tissues. Am. Jour. Anat., vol.
24, p. 127.
Downey, Hat 1911 The origin and structure of the plasma cells of normal
vertebrates, especially of the cold blooded vertebrates, and the eosin-
ophils of the lung of Amblystoma. Folia Haematologica, Bd. 11,
S. 25-
1913a The granules of the polymorphonuclear leucocytes of Ambly-
stoma, with a few notes on the spindle cells and erythrocytes of this
animal. Anat. Anz., Bd. 44, 8S. 309.
1913 b The development of histogenous mast cells of adult guinea-pig
and cat, and the structure of the histogenous mast cells of man,
Folia Haematologica, Bd. 16, S. 49.
1915 The origin and development of eosinophil leucocytes and of
haematogenous mast cells in the bone marrow of adult guinea-pig.
Folia haematologica, Bd. 19, 8. 148.
Jorpan, H. E. 1910 A cytological study of the egg of Cummingia with
special reference to the history of the chromosomes and the centro-
somes. Arch. f. Zellfors., Bd. 4, S. 248.
1913 Amitosis in the epididymis of the mouse. Anat. Anz., Bd. 43,
S. 589.
JorpAN, H. E., anp Furippin, J. C. 1913 Haematopoiesis in Chelonia.
Folia Haematologica, Bd. 15, 8. 1.
JorpAN, H. E. 1918 A contribution to the problems concerning the origin,
structure, genetic relationship and function of the giant-cells of
hemopoietic and osteolytic foci. Am. Jour. Anat., vol. 24, p. 225.
1918 The histogenesis of blood-platelets in the yolk-sac of the pig
embryo. Anat. Rec., vol. 15, p. 391.
Maximow, A. 1906 Uber entziindliche Bindegewebsneubildung beim Axo-
lotl. Zeigler’s Beitr. z. path. Anat. u. z. Allgem. Path., Bd. 39.
1910 Uber embryonale Entwicklung der Blutzellen bei Selachiern
und Amphibien. Anat. Anz., Bd. 37 (Ergiinzungsheft), 8. 64.
1913 Untersuchungen iiber Blut und Bindegewebe. VI. Uber Blut-
mastzellen. Arch. f. mikr. Anat., bd. 83, Abt. 1.
THE AMERICAN JOURNAL OF ANATOMY, VOL. 25, No. 4
474 H. E. JORDAN
17
18
19
20
NakaHara, W. 1918 Studies of amitosis: its physiological relations in
the adipose cells of insects and its probable significance. Jour. Morph.,
vol. 30, p. 483.
NeuMANN, E. 1903 Haematologische Studien. Virchow’s Archiv., Bd.
174.
NieGoutewskt, F. 1894 Die Ehrlische Granulation der weisen Blutkér-
perchen bei einigen Tierspezies. Inaug. Dissert., Miinchen (cited
from Werzberg).
Ringorn, A. R. 1915 a Observations on the origin of the mast leucocytes
of the adult rabbit. Anat. Rec., vol. 9, p. 233.
1915 b Observations on the differentiation of the granules in the
eosinophilic leucocytes of the bone-marrow of the adult rabbit. Anat.
Rec., vol. 9, p. 683.
Sacucu1, S. 1917 Studies on ciliated cells. Jour. Morph., vol. 29, p. 217.
Staruinc, E. H. 1915 Principles of human physiology, p. 833. Lea &
Febiger, Phila.
WerpenreicH, F. 19095 Zur Frage nach der Entstehung der eosinophilen
Leukozyten. Folia Haematologica, Bd. 2.
Werzsera, A. 1911 Studien zur vergleichenden Haemozytologie einiger
poikilothermer Vertebraten. Folia Haematologica, Bd. 11, 8. 17.
Wriacut, J. H. 1910 The histogenesis of the blood platelets. Jour. Morph.,
vol. 21, p. 263.
EXPLANATION OF PLATES
All figures were made by aid of ;'; Leitz oil-immersion lens and a drawing
camera at an original magnification of 2000 diameters, which was reduced one-
third in reproduction. Figures 1 to 32 are from smear preparations of blood,
stained with Wright’s stain; figures 33 to 73 are of sections of the red marrow of
the femur, fixed in a corrosive-sublimate-formalin solution and stained according
to Wright’s technic for demonstrating the origin of blood-platelets from mega-
karyocytes.
PLATE 1
EXPLANATION OF FIGURES
Blood
1 to3 Normal, older, and senile types, respectively, of erythrocytes. The
color of the nucleus changes from violet to lilac, the cytoplasm from bluish green
to yellow, in passing from 1 to 3.
4 to 6 Small, ameboid, and medium-sized lymphocytes, respectively. The
nucleus stains a deep lilac or violet color; the cytoplasm contains finer and
coarser irregular and clumped basophilic (blue) granules, among which may
appear a variable number of small spheroidal metachromatic (lilac-colored)
granules.
7 Similar medium-sized lymphocyte with numerous pseudopods, some of
which have liberated spheroidal granulated globules by a process of constriction.
Except for the blue color of their granules, these globules are apparently identical
with blood-platelets of mammals.
8 Large lymphocyte with reniform nucleus. The cytoplasm contains a
small amount of a metachromatic granulation scattered among the predomin-
ating basophilic (blue) granules.
9 Large lymphocyte, with nuclear and cytoplasmic characteristics like those
of small lymphocytes.
10 Similar large lymphocyte, with reniform nucleus and a conspicuous
diplosome opposite the nuclear concavity.
11 Binucleated eosinophilic leucocyte (mononucleated, polymorphonuclear,
and polynuclear types also occur). The nucleus has a deep lilac color. The
eosinophilic granules are imbedded in a homogeneous basophilic (blue) sub-
stratum. The granules maintain a fairly uniform size; they appear ring-shaped,
indicating a difference in condensation between center and periphery. The
nucleus almost invariably takes a polar position.
12 Basophilic granulocyte (mast-cell). The nucleus is located centrally; it
is relatively large and vesicular, staining a light blue color, and showing a delicate
reticulum. The granules have a deep lilac color, are fairly uniform in size, and
in general slightly larger than the granules of the eosinophilic leucocytes.
13 to 15 Mononucleated (young) types of neutrophilic granulocytes. The
nucleus has a lilac color. The fine neutrophilic granules are imbedded in a
lightly basophilic (blue) cytoplasm. Figure 13 shows a conspicuous dipiosome
about which the granules are disposed in radiating lines. Figure 15 shows two
pseudopods.
16 Neutrophilic granulocyte with bilobed (dividing) nucleus, and conspicuous,
finely granulated centrosphere. |
17 Binucleated neutrophilic leucocyte.
18 to21 Various types of polymorphonucleated neutrophilic leucocytes.
22 Polymorphonucleated neutrophilic leucocyte of blood of dog.
23 to 26 Various, more complex types of polymorphonucleated neutrophilic
granulocytes. Figure 23 shows a dense granular centrosphere; figure 24, a clear
sphere with a dumb-bell-shaped centrosome (centriole).
27 to 29 Types of thrombocytes. The nucleus is a dense granular body,
staining a violet color. The cytoplasm forms a meagre shell of viscid homo-
geneous character. This contains a variable amount of fine uniform, meta-
chromatic (lilac) granules. The general shape of the thrombocytes is fusiform,
but spheroidal and oval forms also occur (fig. 29).
30 and 31 Thrombocytes with pseudopods, some of which liberate granulated
stellate globules. The latter simulate very closely the blood-platelets of
mammalian bloods.
32 Naked disintegrating nucleus of a degenerating thrombocyte.
476
BLOOD AND BONE-MARROW OF FROG PLATE 1
H, E. JORDAN
477
PLATE 2
EXPLANATION OF FIGURES
Bone-marrow
33 A mesenchymal marrow cell, source from which hemoblasts develop.
The cytoplasm is only very slightly basophilic, staining very faintly blue, and
of homogeneous character. The nuclues is relatively large. It invariably con-
tains a faintly staining plasmosome, occasionally several, and a number of larger
karyosomes and innumerable minute chromioles scattered over a very delicate
chromatic reticulum. The nucleus is vesicular in character and takes only a
faintly blue stain.
34 Primitive large lymphocyte (hemoblast). The nucleus is practically
identical with that of its mesenchyme ancestor. It stains only slightly darker.
The cytoplasm likewise seems more condensed and slightly more basophilic, of
light blue color.
35 Slightly older extravascular lymphocyte, in ameboid activity. The
pseudopods may constrict off hyaline globules, similar to non-granular ‘platelets.’
36 Eosinophilic myelocyte; a slightly differentiated lymphocyte, with pseu-
dopods forming hyaline ‘platelets.’
37 Medium-sized primitive lymphocyte (hemoblast); a slightly differentiated
mesenchyme cell. Note the mesenchymal character of the nucleus. This cell
may likewise develop into a granulocyte.
38 Small extravascular primitive lymphocyte.
39 Slightly later stage in the development of an extravascular small lympho-
cyte, leading to a large lymphocyte or a granulocyte.
40 Similar small lymphocyte with numerous pseudopods, which may form
hyaline ‘platelets.’
41 Group of myelocytes from the periphery of a developing, small marrow
blood-space. The cells include in order from above an erythrocyte, a poly-
nuclear neutrophilic leucocyte, a large lymphocyte, an erythroblast, and a small
lymphocyte. From the latter type of cell develop intravascularly both erythro-
cytes and thrombocytes; from large lymphocytes develop intravascularly only
neutrophilic granulocytes. From these large and small intravascular lympho-
cytes develop also the various definitive lymphocytes of the circulating blood.
42 A medullary erythrocyte (above) and a thrombocyte. (Compare with
figs. 1 and 28 for demonstration of growth in passage from narrow to peripheral
blood stream.) The cytoplasm of the thrombocyte varies in color from a very
faint blue to a very light pink. It contains minute metachromatic (lilac) gran-
ules of uniform size, grouped at the poles, and ranged apparently in single file
along the lateral border. Thrombocytes apparently do not protrude pseudopods
within the marrow. Their nucleus has in general the features of a small or
medium-sized primitively mphocyte, from which cell the thrombocyte develops,
but it is characteristically furrowed by deep oblique and longitudinal grooves.
43 Similar medullary thrombocyte.
44 to 50 Successive steps in the development of an eosiniphilic myelocyte
from a primitive lymphocyte or hemoblast (compare figs. 50 and 11).
(Continued on page 480)
478
BLOOD AND BONE-MARROW OF FROG PLATE 2
H. E, JORDAN
(Continued from page 478)
51 and 52 Basophilic myelocytes (medullary mast-cells). (Compare with
ancestral cell, fig. 39, and circulatory mast-cell, fig. 12.)
53 Earlier stage of medullary mast-cell with finer basophilic (deep blue)
granules and a long pseudopod in process of constricting off a ‘platelet’ with
basophilic granules.
54 Young neutrophilic granulocyte. (Compare with the ancestral large
lymphocytes, figs. 34 and 35.) The centrosome is conspicuous in the concavity
of the reniform nucleus. The neutrophilic granules have not yet spread through-
out the entire cytoplasmic mass; a relatively wide non-granular hyaline area
appears at the left.
55 to 59 Suecessively later stages in the development of the neutrophilic
myelocytes. The polylobular nucleus may constrict to form a polynuclear cell
(fig. 57); this amitotic division of the nucleus may in some cases be followed by a
fission of the cytoplasm (fig. 59). These cells contain astralsy stems like those
of the circulatory cells.
60 Small polymorphonuclear neutrophilic myelocyte. (Compare with fig. 22,
a polymorphonuclear neutrophilic leucocyte of blood of dog.)
61 and 62 Young neutrophilic myelocytes with pseudopods.
63 A neutrophilic myelocyte which has protruded four pseudopods into a
capillary space of the marrow, a phenomenon duplicating that by which meg-
akaryocytes of mammals pass platelets directly into the blood stream.
64 to 68 Various forms of neutrophilic myelocytes with pseudopods which
constrict to form typical platelets (fig. 65).
69 Complex polymorphonuclear neutrophilic myelocyte arising directly from
mesenchyme among a group of fat cells, two of which are indicated at the right.
70 Similar neutrophilic myelocyte from the fatty mesenchyma. (Compare
with fig. 24.)
71 Plasma-cell from the medullary mesenchyma, a derivative from a large
lymphocyte. (Compare with fig. 35.)
72 and 73 Mononucleated giant-cells from the marrow mesenchyme. These
are strictly comparable to the mononucleated hemogenic giant-cells of mammals,
from which the polymorphonuclear giant-cells (so-called megakaryocytes), the
source of blood-platelets, develop. These amphibian homologues of mammalian
hhemogenic giant cells likewise possess a basophilic cytoplasm with metachro-
matic (lilac colored) granules, and arise from hypertrophying hemoblasts.
Segmenting pseudopods of such cells produce typical platelets.
480
SUBJECT AND AUTHOR INDEX
DRENAL gland in the albino rat. The
relative volumes of the cortex and me-
Gillaton thes sean eee reece 291
Albino rats from birth to ten weeks of age.
Studiesonthe mammary gland. IV. The
histology of the mammary gland in male
Cy aolliGicit Geeteeten SRR RMEE San ooae soDedce
Albino rat. The postnatal development of
the suprarenal gland and the effects of in-
pation upon its growth and structure in
the
Albino rat. The relative volumes of the cor-
i and medulla of the adrenal gland in
TLE epee, et Oak TRS DIRE ae Cir cer
Axuis, EpwarpD PHEtps, Jr. The homologies
of the maxillary and vomer bones of Po-
INO Sie gue secon OEE patos sn Ud nubeconer
Arteries of the human lower extremity. The
developmention ther....9-:t. ssn =
ADERTSCHER, J. A. The ultimobran-
chial bodies in postnatal pigs (Sus scrofa)
Blood and the red bone-marrow of the leopard
frog, Rana pipiens. The histology of the
Blood. The origin of the phagocytic mono-
nuclear cells of the peripheral............
Bone-marrow of the leopard frog, Rana pip-
aay The histology of the blood and the
EC eee ete Serine Ae cere cm eee oye a
Bones of Polypterus. The homologies of the
Maxi lanysanG wOMer s,s cee ee oe
Brachydactyly in the domestic fowl. The de-
velopmental relations of................-.
Burrowing mammals. Astudy of thecorrela-
tion of the pelvic structure and the habits
of certain
ELLS of the peripheral blood. The origin
of the phagocytic mononuclear..........
CHAPMAN, Royat Norton. A study of the
correlation of the pelvic structure and the
habits of certain burrowing mammals....
Corpus luteum. Studies on the ovary of the
spermophile (Spermophilus citellus tride-
cemlineatus) with special reference to the
Correlation of the pelvic structure and the
habits of certain burrowing mammals. A
SEUGVAOL tHE a5. hase ee ee OE ei dente
Cortex and medulla of the adrenal gland in
pe albino rat. The relative volumes of
WIG ond anon soctucdccdudoOocoucccononucuraL
ANFORTH, C. H. The developmental
relations of brachydactyly in the domes-
97
Development of the arteries of the human
KOWemextreniityn yer atte ieee ees 55
Development of the intercalated discs. His-
togenesis of the heart muscle of the pig in
relation to the appearance and............ 333
Development of the lobule of the pig’s liver.
21) TCU Eee aM en sent ee EOE EOE 299
Development of the suprarenal gland and the
effects of inanition upon its growth and
structure in the albinorat. The post natal 221
481
Discs. Histogenesis of the heart muscle of
the pig in relation to the appearance and
development of the intercalated..........
Domestic fowl. The developmental relations
of brachydactyly in the..................
DonaLpson, JOHN C. The relative volumes
of the cortex and medulla of the adrenal
gland in the albino rat
Drips, Detia. Studies on the ovary of the
sphermophile (Spermophilus citellus tri-
decemlineatus) with special reference to
the corpus luteum
XTREMITY. The development of
arteries of the human lower . . ....
333
the
ILUM terminale. Factors involved in
fie kormanomol theses ores esterase ne
Fowl. The developmental relations of brachy-
dactyly in the domestic...............--
Frog, Rana pipiens. The bistology of the
blood and the red bone-marrow of the
Jeopand ie sea. c as oss ase sees ce cee
LAND and the effects of inanition upon
its growth and structure in the albino
rat. The postnatal development of the
Suprarenalls. com. sc ach detest tems
Gland in the albino rat. The relative vol-
umes of the cortex and medulla of the ad-
renal
Gland. IV. The histology of the mammary
gland in male and female albino rats
from birth to ten weeks of age. Studies
One MamManys ose ee eee ice
Growth and structure in the albino rat. The
postnatal development of the suprarenal
gland and the effects of inanition upon its
395
221
ABITS of certain burrowing mammals.
A study of the correlation of the pel-
vic structure and the.............-..--
Heart muscle of the pig in relation to the ap-
pearance and development of the inter-
calated discs. Histogenesis of the........ 3
Histogenesis of the heart muscle of the pig in
relation to the appearance and develop-
ment of the intercalated dises............ 333
Histology of the blood and the red bone-mar-
row of the leopard frog, Rana pipiens.
MB aYS Yu rs Bh crc ca SOC TODO eee aicrtcicl
Histology of the mammary gland in maieand
female albino rats from birth to ten weeks
of age. Studies on the mammary gland.
TIW/, MB GH one): Seca ee ns HOME DOS eon ae
Homologies of the maxillary and vomer bones
of Polyptenuss he. 20 e622 ee oi - 4
Human lower extremity. The development
oltheartenies!Of thes:4<. <<< + sere
| Ey
development of the suprarenal gland and
tHeehectsOle s,s c-ee== Mess aes ee
Intercalated discs. Histogenesis of the heart
muscle of the pig in relation to the ap-
pearance and development of the
437
ANITION upon its growth and struc-
ture in the albino rat. The postnatal
482
ACKSON, C.M. The postnatal develop-
ment of the suprarenal gland and the ef-
fects of inanition upon its Bryn and
struture in the albino rat. : 221
JOHNSON, FRANKLIN PARADISE. ‘The ‘de-
velopment of the lobule of the pig’s liver 299
Jorpan, H. BE. The histology of the blood
andthe red bone-marrow of the leopard
frog. Ranaypiplenseessdsratemicer ce eeccl: 437
EOPARD frog, Ranapipiens. The histol-
ogy of the blood and the red bone-
marrow of the.. 437
Liv a ne development ‘of the lobule of the
Tebale of the pig’s liver. The dev clopment _
Gh the este aclacrrras panes wees eae ee 299
Lower extremity. The development of the
arteries of the human.. 55
Luteum. Studies on the ovary ‘of the spermo-
phile (Spermophilus citellus tridecemline-
atus) with special reference to the corpus 117
cJUNKIN, Frank Apam. Theorigin of
M the phagocyte mononuctlear cells of
the peripheral blood: e.s-- eee 27
Mammals. A study of the correlation of the
pelvic structure and the habits of certain
PULFO WINE, evs heen earl sees creer ees 185
Mammary gland. IV. The histology of the
mammary gland in male and female al-
bino rats from birth to ten weeks of age.
Studiesion GHel icc cs cen sleustestoe ee rors eichate 395
Maxillary and vomer bones of Polypterus.
Thevhomologiesiof the: ....5--+-s-2e-- <- 349
Medulla of thea ‘drenal gland in the albino rat.
The relative volumes of the cortex and... 291
Mononuclear cells of the peripheral blood.
The origin of the phagocytic............. 27
Muscle of the pig in relation to the appearance
and development of the intercalated discs.
Histogenesis of the heart................. 333
Myers, J. A. Studies on the mammary
gland. IV. The histology of the mam-
mary gland in male and female albino
rats from birth to ten weeks of age........ 395
VARY of thespermophile (Spermophilus
citellus tridecemlineatus) with special
reference tothe corpus luteum. Studies
OMMENEN eas gi mete cichetes see etee wabeereoanerase 117
ELVIC structure and the habits of certain
burrowing mammals. A study of the
COLLElAtIOMIOL thes eriemce eres cic ties eee 185
Peripheral blood. The origin of the phagocy-
tic mononuclear cellsof the............... 27
Phagocytic mononuclear cells of the periph-
eral blood. The origin of the............ 27
Pigs in relation to the appearance and devel-
opment of the intercalated discs. Histo-
genesis of the heart muscle of the......... 333
Pig’s liver. The development of the lobule
Gisbheres ast Pet eeehtce enema 299
an
INDEX
Pigs (Sus scrofa). The ultimobranchial bod-
Jes)in postnatal kes... =...ceoe eee eee 13
Pipiens. The histology of the blood and the
red bone-marrow of the leopard frog,
1 St hol: HAE a 5 eR eR eee aoac to tas Seinioac 437
Polypterus. The homologies of the maxillary
andtvomen bones ols teeeserece oe eee cae 349
Postnatal development of the suprarenal
gland and the effects of inanition upon its
growth and structure in the albino rat.
TCs. che a Ne eee OO eee oe God Clare 221
Postnatal pigs (Sus scrofa). The ultimo-
branchialibodiestineneeescst cee. s sees 13
R? ANA pipiens. The histology of the blood
eae the red bone-marrow of the leopard
Rats ae birth to ten weeks of age. * Studies
on the mammary gland. IV. The his-
tology of the mammary gland in male
anditemaleialbinosecacuese aoe ee een 395
Rat. The postnatal development of the su-
prarenal gland and the effects of inanition
upon its growth and structure in thealbino 221
Rat. The relative volumes of the cortex and
medulla of the adrenal gland in the albino 291
Red bone-marrow of the leopard frog, Rana
pipiens, The histology of the blood and res
Ne Bae ectnorecons UnUoR oo OQOope Codes aos 70°
ENIOR, H. D. The development of the
arteries of the human lower extremity... 55
Spermophile (Spermophilus citellus iiideoen
lineatus) with special reference to the cor-
pusluteum. Studies on the ovary of the. 117
SrrEETER, GeorGE L. Factors involved in
the formation of the filun terminale...... 1
Structure and the habits of certain burrowing
mammals.