THE
INTERNAL SHC
OF THE OVy
A. S. PARKES
m
MONOGRAPHS ON PHYSIOLOGY
EDITED BY
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THE INTERNAL SECRETIONS
OF THE OVARY
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THE
INTERNAL SECRETIONS
OF THE OVARY
BY
A. S. PARKES
M.A. (Caxtab.)» Ph.D. (Manch.), D.Sc. (London)
BEIT MEMORIAL RESEARCH FELLOW
DEPARTMENT OF PHYSIOLOGY AND BIOCHEMISTRY
UNIVERSITY COLLEGE, LONDON
WITH ILL USTRA TIONS
LONGMANS, GREEN AND CO.
LONDON I NEW YORK I TORONTO
1929
Made in Great Britain
AUTHOR'S PREFACE
This book has been written with the object of bringing together
the more important facts bearing upon the internal secretions
of the ovary. This subject now forms a vigorous growing-point
of physiology, and its innate complexity, combined with the
present activity of research workers, makes the presentation of
a coherent picture difficult. Nevertheless, while many aspects
must as yet be dealt with tentatively, my aim has been to
bring forward in their natural relevance and in due proportion,
the observational and experimental data which comprise our
knowledge of the ovary as an endocrine organ.
The more general sections of the book, notably the chapter on
the morphology of the female reproductive organs, are merely
introductory. No useful purpose could be served by reproducing
in detail what is found in numerous anatomical and histological
text-books. The morphological aspect of the oestrous cycle is of
necessity dealt with fairly fully, but only those species are
considered which have been studied in some detail. The re-
maining chapters on the endocrine control of the female
reproductive organs are intended, however, to be tolerably
complete, both as regards fact and hypothesis. The biblio-
graphy has been designed to include as much as possible of the
literature up to the end of 1928.
My obligation to workers in this and similar fields is un-
bounded. The development of my own research has largely
been directed by the critical interest of Dr. F. H. A. Marshall,
F.R.S., whose writings form the groundwork of the subject, and
who has most kindly read the proofs and allowed me to use
certain illustrations from his Physiology of Reproduction. I am
deeply conscious, also, of my debt to Prof. C. Lovatt Evans,
F.R.S., and Prof. J. P. Hill, F.R.S., for their continued advice
and encouragement. The work on X-ray sterilization has been
viii AUTHOR'S PREFACE
made possible by the kindness of Prof. G. Elliot Smith, F.R.S.,
in allowing me facilities in his Department, and by the interest
of Dr. H. A. Harris.
Mr. J. Hammond, in addition to allowing me to reproduce
Figs. 10 and 17, has also given me the benefit of his wide
experience. To Dr. J. H. Burn I am indebted for permission
to reproduce Figs. 68 and 69, while Mr. W. Shaw has kindly
supplied me with the human material shown in Figs. 4, 5 and 35.
Table 3 and Figs. 14, 18 and 63 respectively have been
included by the courtesy of Prof. H. M. Evans, Prof. Carl
Hartman, Prof. G. W. Corner, and Prof. B. Zondek, while I am
particularly grateful to Dr. P. E. Smith and Dr. E. T. Engle for
the illustrations in Figs. 59, 60 and 61. To all of these I would
offer my best thanks.
Finally, I would take this opportunity of thanking most
sincerely my friends and collaborators, Dr. F. W. R. Brambell,
Mr. G. F. Marrian, Mr. C. W. Bellerby, Miss U. Fielding, Dr.
A. R. Fee, and Mr. S. Zuckermann, who have taken a large
part in the research upon which this monograph is based, and
who have assisted me with the preparation of the book.
I have been fortunate in obtaining permission from the Council
of the Royal Society, from the Cambridge University Press,
and from the Editors of the Quarterly Journal of Experimental
Physiology and of the Lancet, to reproduce various illustrations
and figures.
The new illustrations are the work of Mr. F. J. Pittock.
A. S. Parkes.
University College,
London,
1929.
CONTENTS
CHAPTER
I. The Differentiation of the Sexes .
II. The Female Reproductive Organs .
(a) The ovary ......
(b) The accessory reproductive organs
III. Sexual Periodicity in the Female Mammal
(a) Puberty and the menopause
{b) The breeding season ....
(c) Essential features of the oestrous cycle
IV. Types of OEstrous Cycle
(a) Marsupials .
{b) Dog . . .
(c) Guinea-pig and cow
(d) Horse, sheep and pig
(e) Mouse and rat .
(/) Rabbit and ferret
(g) Primates .
(h) Summary table .
V. The Ovary as an Organ of Internal Secretion
(a) Effects of ovariectomy ....
(b) Ovarian transplantation ....
(c) Intersexuality ......
{d) Administration of ovarian preparations
(e) The internal secretion complex of the ovary
PAGE
I
4
4
12
i8
i8
22
23
28
28
31
34
39
43
53
6i
68
70
70
75
8o
8o
8i
33739
CONTENTS
VI. The (Estrus-Producing Hormone
(a) History of preparation
{b) Chemical properties
(c) Administration
(d) Test-objects
(e) Standardization
(/) Distribution
(g) Site of origin
(h) Pharmacological properties
Vn. The Function of ffisTRiN
{a) Action on test animals
{b) Action on normal animals .
(c) Limits of action
{d) Attainment of puberty
(e) QEstrin and the changes of the post-ovulation
phase
(/) Significance of distribution
Vin. The Periodicity of CEstrus
{a) Role of the cyclic structures of the ovary
(b) Relation between the Graafian follicle and the
production of oestrus ....
(c) Occurrence of oestrus after follicular ablation
(d) Occurrence of follicular maturation without
oestrus .
(e) Reasons for supposing ovarian regulation to be
external
83
83
88
91
93
lOI
106
113
113
115
115
115
121
123
125
131
134
134
134
137
148
149
IX. The Relation between the Ovary and the
Anterior Pituitary Body .... 152
(a) Introduction ....... 152
(b) Luteinization of the Graafian follicle . . . 153
(c) The production of ovulation .... 158
CONTENTS
XI
{d) Are two anterior pituitary substances concerned
in the regulation of the ovary ?
(e) Assay of anterior pituitary extracts .
(/) Preparation and properties
(g) Distribution of the anterior pituitary hormone
(h) Action on the normal animal
(/) The mechanism of ovarian regulation
X, The Internal Secretion of the Corpus Luteum
(a) Introduction ......
(b) Inhibition of ovulation and oestrus .
{c) Sensitization of the uterus ....
(d) Development of the mammary glands
[e) Maintenance of pregnancy ....
(/) The functional relation between the corpus
luteum and the interstitial tissue
XI. Parturition
(a) Correlation with ovarian cycle
{b) Direct action of the ovary on spontaneous
uterine contraction
(c) Role of oxytocin
(d) Relation of parturition to effects
during pregnancy .
Bibliography
Index of Authors
Index of Subjects
of ovariectomy
163
165
166
166
168
170
173
173
176
183
188
196
198
200
200
201
203
206
207
235
239
LIST OF ILLUSTRATIONS
FIG.
1. Ovary of mouse just before puberty
2. Ovary of dog ......
3. Part of corpus luteum of pregnancy of mouse
4. Human corpus luteum
5. Human ovary .
6. Ovary of rabbit
7. Fallopian tube of mouse
8. Clitoris of mouse
9. Lobule of mammary gland of cow .
10. Udder of cow ......
11. Ovary of mouse at three weeks old .
12. Uterus of mouse at three weeks old
13. Ovary of senile mouse ....
14. Diagram of oestrous cycle in the opossum
15. Prooestrous uterine mucosa of dog .
16. Uterine mucosa of dog at the end of pseudo-pregnancy
17. Diagram of ovarian cycle in non-pregnant cow
18. Diagram of ovarian cycle in non-pregnant sow
19. Ovary of mouse (Pasini stain) showing two sets of corpora
lutea ....
20. Uterus of mouse
21. Uterus of mouse during dioestrus
22. Uterus of mouse during oestrus
23. \^agina of mouse duri.ng dioestrus
24. \^agina of mouse during oestrus
25. Vaginal smears of the mouse during the oestrous cycle
26. Uterus of rabbit in oestrus
5
7
8
9
10
II
13
15
16
17
19
19
21
29
32
33
38
42
45
46
47
48
49
50
51
53
Xlll
XIV
LIST OF ILLUSTRATIONS
27. Uterus of pseudo-pregnant rabbit ..... 54
28. Photograph of mammary gland of pre-pubertal rabbit , 55
29. Photograph of mammary gland of rabbit in first oestrus . 56
30. Photograph of mammary gland of rabbit 12 days pseudo-
pregnant 57
31. Photograph of mammary gland of rabbit after pseudo-
pregnancy ........ 58
32 . Photograph of mammary gland of rabbit 23 days pregnant 59
33. Section of lobule of mammary gland of rabbit during
pseudo-pregnancy ....... 60
34. Section of mammary gland of 29 days pregnant rabbit . 61
35. Endometrium of human uterus on the first day of men-
struation ......... 62
36. Normal uterus of rat ....... 71
37. Uterus of rat after ovariectomy . . . . . y^
38. Uterus of mouse after pre-pubertal ovariectomy . . 74
39. Vagina of mouse after pre-pubertal ovariectomy . . 75
40. Ovary of rat after transplantation to the peritoneum . yy
41. Uterus of rat after transplantation of the ovary . . 79
42. Uterus and vagina of ovariectomized mouse injected
with oestrin ........ 94
43. Ovarian tissue re-formed after double ovariectomy . . 100
44. Diagram showing the growth curves and oestrous cycle
histories of rats injected with oestrin during vitamin
B deficiency an oestrus . . . . . .117
45. Ovary of mouse sterilized by exposure to X-rays when
three weeks old . . . . . . .139
46. Ovary of mouse sterilized when three weeks old . . 140
47. Ovary of mouse sterilized by exposure to X-rays when adult 141
48. Ovary of mouse sterilized at birth ..... 142
49. Uterus of mouse sterilized at weaning time . . . 143
50. Frequency polygons for length of oestrous cycle before
and after X-ray sterilization . . . . .144
51. Ovary of adult mouse injected with sodium hydroxide
extract of anterior pituitary body . . . .154
LIST OF ILLUSTRATIONS xv
PACE
52. Early stage in luteinization of the Graafian follicle of
mouse by sodium hydroxide extract of anterior
pituitary body . . . . . . -155
53. Later stage of luteinization ...... 155
54. Atretic corpus luteum in untreated mouse . . . 156
55. Ovary of mouse injected with sodium hydroxide extract
of anterior pituitary at three weeks old . . 156
56. Ovary of rabbit injected with sodium hydroxide extract
of anterior pituitary ...... 157
57. Luteal tissue of ovary of Fig. 56 . . . . .158
58. Luteal tissue produced in sterilized ovary of mouse by
anterior pituitary preparations . . . .159
59. Effect of anterior pituitary implants on ovary of im-
mature rat ........ 160
60. Effect of anterior pituitar}^ implants on ovary of im-
mature rat ........ 161
61. Group of tubal ova following super-ovulation . . 162
62. Effect of oestrin and anterior pituitary extracts on uterus
and ovary of young mouse . . . . .165
63. Diagram of amount of oestrin and anterior pituitary
substance in the urine of pregnancy . . .167
64. Uterus of mouse (after sterile copulation)with deciduoma 185
65. Uterus of mouse with deciduoma ..... 187
66. Photograph of mammary gland of rabbit during pseudo-
pregnancy prolonged by anterior pituitary extracts 194
67. Section of gland shown in Fig. 66 . . . . . 195
68. Effect of oestrin on the contraction of an isolated
guinea-pig uterus ....... 203
69. Action of oestrin in sensitizing an isolated uterus to
oxytocin ......... 205
CHAPTER I
THE DIFFERENTIATION OF THE SEXES
The sexes are distinguished primarily by the presence of ovary
or testis, i.e. by the power to produce ova or spermatozoa. This
production of germ cells is the essential function of the gonads,
and in lower animals, where the reproductive processes are
of the simplest, their only function. During the course of evolu-
tion, however, the tendency has been to limit the number of ova
produced, and to provide each embryo with a greater degree of
parental care. This tendency has culminated in the prolonged
period of internal gestation found in mammals, and has
resulted in the appearance of many organs and characters
designed to facilitate reproduction. Coincidently, a second
function of the gonad has appeared, the control of the develop-
ment of these accessory structures. These organs have reached
a higher degree of elaboration in the female, upon whom the
care of the young largely falls in most species.
In addition to the accessory organs, there are found secondary
sexual characters, which, though of no direct use in the repro-
ductive processes, are nevertheless valuable or necessary in
dioecious propagation.
It is thus possible to distinguish three types of sexual
differentiation :
(a) The gonad — ovary or testis.
[h) The accessory reproductive organs.
[c) The secondary characters.
The accessory reproductive organs in the male are all designed
to convey the spermatozoa in a suitable medium to the exterior
and thence into the female genitalia. The accessory organs of
the female mammal (individually described in Chapter II) are
adapted for reception of the spermatozoa, gestation of the
P.S.O. A
2 INTERNAL SECRETIONS OF THE OVARY
fertilized ovum, evacuation of the foetus, and subsequent
suckling of the young.
The secondary sexual characters in mammals are extra-
ordinarily diverse. The characters usually consist in the
appearance or accentuation of some attribute which serves to
attract the opposite sex, or combat others of the same sex.
Since the male is usually the active partner in mammalian
reproduction, this sex has the more definite secondary
sexual characters. Thus the possession of fighting weapons and
a stronger skeleton are typical of the male mammal. Sex
differences in the voice and in the amount and distribution of
hair are also found.
The differences in external appearance which result from the
possession of secondary sexual characters make it possible in
many animals to distinguish male from female without reference
to their genitalia. The degree to which these characters are
present, however, is subject to much specific variation, and
from the point of view of the experimental physiologist, their
distribution is disappointing. The common laboratory rodents,
for instance, have practically no secondary sexual characters,
and long experience is required to distinguish male from female
in such animals as mice, rats, guinea-pigs, and rabbits without
examination of the external reproductive organs.
The mechanism of sexual differentiation. The factors which
cause a fertilized ovum to develop a testis or an ovary,
namely, to become male or female, are at present inadequately
known, but there can be little doubt that the development of the
indifferent embryo into one sex or the other is normally
dependent upon its chromosome constitution. This aspect of
the problem has been dealt with in full by many writers (8i,
147, 149, 168, 248).
The nature of the subsequent sexual differentiation is better
understood, and it is clear that once the gonads have
developed, differentiation proceeds as the result of stimuli
from these organs. Gonadectomy experiments have shown
decisively that the development of the accessory organs and
secondary sexual characters is entirely dependent upon the
presence of the gonad, as are the skeletal and other structural
details typical of the two sexes.
THE DIFFERENTIATION OF THE SEXES 3
Moreover, grafting experiments have shown that the removal
of the gonad from its normal site in the body to an alien
situation does not affect the development of the associated
organs. Experiments of this nature soon suggested that the
gonads exert their control by means of some internal secretion,
and later work has substantiated this view. The extraction of
the active principles from the gonads has, however, lagged
behind work on the other internal secretory organs. This is
particularly true of the testis, from which no substance appears
to have been prepared capable of replacing its endocrine action
in the castrated male. As regards the mammalian ovary, at
least one definitely active substance has been prepared, and it
is reasonable to suppose that both ovary and testis control
the development of their accessory organs by means of
internal secretions. In the female mammal the study of the
endocrine activity of the gonad is comphcated by the cyclic
changes which take place in the reproductive organs during
the breeding season, and which have no analogue in the male.
CHAPTER II
THE FEMALE REPRODUCTIVE ORGANS
The morphology of the female reproductive organs is described
in a great number of text-books and monographs (some of the
chief of which are cited in the bibliography : 8i, 265, 429) and it
is not proposed to give a more detailed account here than is
strictly necessary as a basis for the rest of the book. Mor-
phological variation in the reproductive organs from one species
to another may make one animal useless and another ideal for
some particular experiment, and it is with this aspect of specific
variation, therefore, that the present account is largely con-
cerned.
[a) THE OVARY
The ovary is a bean-shaped organ attached to the broad
ligament by the hilum. The outer covering is the germinal
epithelium, which is continuous with the peritoneal epithelium,
and from which the definitive ova are proliferated early in life
and possibly after puberty (7, 275, 484) . The body of the ovary
consists of a stroma of connective tissue in which the follicles are
embedded, together with the products of follicular degeneration
or maturation. In addition, the ovaries of many animals
(notably the rabbit) contain blocks of epithelial cells, forming
the so-called interstitial tissue. This tissue appears to be
completely absent in certain animals, such as the mouse.
The Graafian follicle. The follicle, containing the ovum,
consists of two peripheral layers, the theca externa and the
theca interna, surrounding the follicular epithelium (membrana
granulosa and discus proligerus) which carries the ovum. In
the more mature follicles (except in the Monotremata) an
antrum appears which is filled with a viscous fluid, the liquor
folliculi.
THE FEMALE REPRODUCTIVE ORGANS 5
The theca externa cells are fibroblastic, and continue un-
changed during the whole life of the follicle. After ovulation
the theca externa produces trabeculae which carry blood and
lymph capillaries into the developing corpus luteum.
The accounts of the life history of the theca interna cells vary
greatly. In some species, such as the pig (Corner, I2i, 122),
\^^'rV
m.g.
Fig. I. — Ovary of Mouse just before Puberty.
Large follicles but no corpora lutea are present.
c. connective tissue; d.p. discus proligerus; g.e. germinal epithelium;
/./. liquor folliculi; m.g. membrana granulosa; o. ovum; t. theca.
and Ornithorhynchus (Hill and Gatenby, 299), they are de-
scribed as being small and flattened until just before ovulation
and then swelling up to a glandular type. Other authors,
however, describe them as undergoing degeneration on follicular
maturation or after ovulation {Dasyuriis : O'Donoghue, 477, 479,
480 ; Dasyurus and Didelphys : Hill and Gatenby, 299).
The granulosa is made up of polygonal cells arranged irre-
gularly, except round the periphery and round the ovum, where
the arrangement is columnar. Most recent authors have
followed Pfliiger (517), de Winiwarter (639), and Waldeyer (628)
6 INTERNAL SECRETIONS OF THE OVARY
in supposing that both ovum and fohicular epithehum are de-
rived direct from the germinal epithehum. The ovum itself con-
sists of an external zona pellucida which encloses a cytoplasmic
mass containing the nucleus and certain cytoplasmic inclusions.
Ovulation, which takes place spontaneously at oestrus in most
species, is preceded by certain changes in the follicle. The
distension due to the accumulation of liquor folliculi causes the
follicle to approach the periphery of the ovary, so that discharge
of the ovum becomes possible. The discus proligerus, containing
the ovum, breaks away from the membrana granulosa, and at
the same time the cells become arranged radially round the
ovum, from which they tend to withdraw, forming the corona
radiata or cumulus oophorus. Finally, the first polar body is
given off, and ovulation takes place. The nature of the final
stimulus required for ovulation is not known. If fertilization
takes place, the second polar body is given off, and the ovum
proceeds down the Fallopian tube. Otherwise, degenerative
fragmentation occurs, which has occasionally been mistaken
for parthenogenesis.
Although ovulation is the predestined end of the Graafian
follicle, the majority fail to complete the full life history
owing to their excessive number, and undergo atresia at some
stage of their growth. The degenerative processes usually
begin in the granulosa, from which odd cells are shed into
the antrum in a state of pycnosis. The degeneration of the
ovum, accompanied by spurious maturation divisions, usually
takes place later in atresia. Finally, the follicle is either entirely
absorbed or metamorphosed into interstitial tissue or a corpus
luteum atreticum. Cyclic bursts of atresia seem to take place
in the ovary corresponding with the stages of the oestrous
cycle. Atresia of small follicles tends to be most common
during pregnancy and in the vicinity of large corpora lutea.
Atresia of large follicles is very evident in the rabbit (in the
absence of the sexual stimulus required for ovulation) and in
the guinea-pig where only two or three of each batch of mature
fohicles ovulate. For the extensive literature on fohicular
atresia the bibliography given by Salazar (541) may be
consulted.
Abnormalities of the ovum and follicle, such as multinucleate
THE FEMALE REPRODUCTIVE ORGANS
7
ova, and poly- or anovular follicles, are not uncommon, but they
are of no particular physiological interest (see Hartman, 274,
for full bibliography).
TJie corpus liiteum. After ovulation the shell of the ruptured
follicle shrinks, and, owing to the previous rupture of capillaries,
becomes filled with a greater or lesser amount of blood. From
^
'^C-<^ "
f^.0*
4%
y-g'^A
>, »
•iU'
. A'
y\^ -^ -
' ^/''
a.f.
•i»
5*5
^
Fig. 2. — Ovary of Dog.
a.f. atretic follicle; c.l. corpus luteum; y.g.f. young Graafian
follicle.
the follicular remains, the corpus luteum, composed of large
glandular cells containing the so-called lutein granules, develops
with remarkable rapidity. In some species [e.g. cow, man) the
corpora lutea are coloured yellow, orange or reddish, by the
presence of carotene.
Much controversy centres round the exact manner of forma-
tion of the corpus luteum. The probability is that it develops
from the remains of the follicle by enlargement of the individual
cells and not by cell division, though Loeb (379) has described
8 INTERNAL SECRETIONS OF THE OVARY
mitotic proliferation. Doubt still exists as to the degree to
which the various follicular elements are concerned in the
transformation.
Von Baer (53) originally described the corpus luteum as
originating solely from the theca interna, and one or two
comparatively recent writers have adopted this view. It is
now generally admitted, however, that the remains of the
ii.—
Fig. 3. — Part of Corpus Luteum of Pregnancy
OF Mouse.
l.g. lutein granule; n. nucleus.
follicular epithelium play the major part in the formation of the
true lutein cells, and the real problem relates to the part played
by the theca interna. According to Sobotta for the mouse
(583-6) and guinea-pig (587), Van der Stricht for the bat (601-3),
and Marshall for the sheep (440, 442), the lutein cells are derived
exclusively from the follicular epithelium, the theca interna
merely assisting in supplying the vascular connective tissue
framework for the corpus luteum.
In other species, however, the theca interna has been described
as undergoing a burst of growth at the time of ovulation, and
THE FEMALE REPRODUCTRT ORGANS 9
as contributing clumps of cells to the corpus luteum. These
cells, the theca-lutein cells of Corner and para-lutein cells of
Gatenby, while clearly distinguishable from the true lutein cells,
are of the same general type and appear to be definitely of a
secretory nature. This type of corpus luteum has been described
in the sow by Corner (122), in Platypus by Hill and Gatenby
(299) and in the human by Gatenby (237) and by Shaw (560).
t.m.g : — - "^ •'
Fig. 4. — Human Corpus Luteum.
t.m.g. tissue derived from membrana granulosa; t.t.i. tissue
derived from theca interna (after Shaw).
In such circumstances, therefore, there is some histological
basis for supposing the corpus luteum to have a dual secretion.
In certain animals, notably the cow, many of the corpora
lutea are hollow and contain a viscous fluid. Some workers have
considered them to be cystic. Allen in the mouse (6) and
Hammond in the cow (265), however, found that the secretion
of liquor folliculi goes on temporarily after ovulation, and if not
completely reabsorbed by the developing corpus luteum, the
fluid remains in a central cavity. This inclusion of follicular
secretion is important from the point of view of extraction
of the corpus luteum (see p. 107).
10
INTERNAL SECRETIONS OF THE OVARY
In many species, notably polyoestrous ones, the corpus luteum
survives as a histologically intact body long after it must have
ceased to function; in the unmated mouse three, four or even
five sets of corpora lutea, representing successive ovulations,
may be present in the ovary at one time. The name corpus
luteum spurium has been given to these corpora lutea of the
infertile cycle, but the distinction seems unnecessary since the
difference between them and the corpora lutea of true pregnancy
M
n.c.J.
n.c.l.
Fig. 5. — Human Ovary.
n.c.l. new corpus luteum; o.c.a. old corpus albicans (after Shaw).
is merely one of degree. In Dasy virus such a difference has
not appeared, the duration of life and histological appearance
of the corpora lutea being uninfluenced by the fate of the ova.
The ferret appears to be similar in this respect.
The corpora lutea atretica, which are sometimes formed with-
out the intermediate stage of ovulation by atretic follicles or by
follicles in which ovulation is for some reason inhibited, probably
do not function; here again, however, the difference from
normal luteal tissue appears to be only one of degree. Corpora
lutea capable of performing all their known functions can be
produced experimentally without discharge of the ovum.
THE FEiMALE REPRODUCTIVE ORGANS
II
After a certain time, which varies according to the species and
the occurrence of lactation and pregnancy, the corpus luteum
retrogresses and finally is either reabsorbed entirely or dwindles
to a small corpus albicans.
The interstitial tissue of the ovary. The whole question of the
presence, formation and function of the so-called interstitial
tissue of the ovary is controversial. Clumps of large epithelial
^ v.o.
Fig. 6. — Ovary of Rabbit.
i.t. interstitial tissue; y.o. young oocjrtes.
cells, presenting the appearance of secretory tissue, are very
obvious in the ovaries of such mammals as the rabbit, but appear
to be absent in others such as the mouse. Schaeffer (545), Aime
(3), Fraenkel (210), and O'Donoghue (480) give lists of species
showing the presence or absence of interstitial cells. According
to Bouin and Ancel (77) they are not found in the ovaries of
animals which ovulate spontaneously. The confusion as to the
distribution of this tissue is accentuated by the lack of any
unanimous definition. Some workers describe only obviously
extrafollicular tissue as interstitial, while others, maintaining
that interstitial cells are of follicular origin, apply the term to the
12 INTERNAL SECRETIONS OF THE OVARY
products of follicular atresia and luteal degeneration. Their
origin is variously ascribed to three different sources :
(a) Direct derivation from the germinal epithelium; Paladino
(483), Lane-Claypon (340) and others.
(b) Derivation from the follicle, either from old corpora
lutea, atretic follicles or theca interna; Schron (550)
and Rabl (522).
(c) Derivation from transformed connective tissue cells ;
Sainmont (540), Regaud and Policard (527) and
Athias (47).
Others again, as Van der Stricht (603), consider a combination
of one or more of these sources to be probable. Criticisms of all
these views are obvious and further discussion is unwarranted
here.
The interstitial tissue has been said to show cyclic changes
during the oestrous cycle (O'Donoghue, 480, Lane-Claypon, 340),
but owing to the present doubt as to whether it is an essential
constituent of the mammalian ovary, and to the uncertainty of
its origin, it is difficult at the moment to assign to it any definite
physiological role. According to Steinach (590), Lipschiitz
(366) and others, it is the main endocrine tissue of the ovary and
constitutes the ' puberty gland ' of these authors.
Accessory ovarian tissue. Ovarian tissue apart from the two
main ovaries may occur in rare instances, according to Waldeyer
(628), Beigel (55), Wilhams (638), Hartman (276) and other
workers. In the dissection of some thousands of mice, however,
the writer has found only two with accessory ovaries, and in
each case the abnormality consisted of a supplementary body
within the same capsule as the normal ovary. The problem
of third or accessory ovaries is of physiological importance as
it may complicate the operation of complete ovariectomy
(see p. 98).
{h) THE ACCESSORY REPRODUCTIVE ORGANS
In mammals the accessory female reproductive organs are
designed for the fertilization of the ovum, the gestation of the
embryo, and the subsequent suckling of the young. To this end
THE FEMALE REPRODUCTIVE ORGANS 13
the more primitive oviduct of oviparous animals has undergone
great elaboration, while certain skin glands have undergone
alteration to produce the mammary tissue.
The Fallopian tube. After ovulation the ova are caught by the
funnel-shaped end of the Fallopian tube (the infundibulum] and
are passed down the tube by the action of the ciliated epithelium
with which it is lined, aided by the secretion of mucus which
takes place at the time of ovulation. The Fallopian tube may
Fig. y. — Fallopian Tube of Mouse.
e.l. epithelial lining; ))i.l. muscle layer; p.c. peritoneal covering.
be entirely independent of the ovary except for its common
anchorage in the broad ligament (as in man, the cow, sheep,
etc.) or the expanded end may open into an ovarian capsule
formed from a fold of peritoneum. A closed circuit of this
nature, which makes it impossible for the ova to fall into the
body cavity, is found in the bitch, mouse, and rat. The
Fallopian tube itself may be short and coiled as in the mouse
and rat, or long and comparatively straight as in the rabbit,
ewe and man. The tube is lined by mucous membrane and
covered by a serous layer from the peritoneum. Circular
and longitudinal muscle layers are found under the serous
layer.
14 INTERNAL SECRETIONS OF THE OVARY
The uterus. The Fallopian tubes open into the uterus, which
consists essentially of the same three layers, but the internal
mucous membrane is much thickened to form the uterine
mucosa (or endometrium) which consists of a glandular stroma
lined by epithelium. The shape of the uterus shows great
specific variation. At one extreme is the type having two
distinct cornua fusing only at the junction with the vagina, and
even there retaining two distinct cervical canals. This type of
uterus is characteristic of the rat and the mouse. At the other
extreme is the type where the cornua are entirely fused to form
one large (usually pear-shaped) uterine sack, into the top angles
of which open the two Fallopian tubes. This type is
characteristic of the human. Between these extremes every
gradation is found, from the rabbit, where the cornua are fused
to the extent of having a common cervical canal, to the goat
where fusion is complete except for the tops of the cornua,
which form two horns projecting from the main body of the
uterus.
The vagina. The vagina, which connects the uterus with
the exterior, possesses the two muscular layers found in the
other sections of the genital tract, and is lined internally with
epithelium, the nature of which varies greatly in different
animals and in different stages of the cestrous cycle. In man
erectile tissue is present, but this appears to be absent in the
lower species, in which the necessary facilitation to copulation
is obtained by copious secretion (cow), intense hyperaemia
(ferret) or cornification (mouse). The vagina opens to the
exterior at the vulva, the anatomy of which varies greatly
in different species, being a simple orifice in the case of most
lower mammals, and complicated in the human by inner
and outer labia.
In the rat and mouse the vaginal lumen is not complete during
pre-pubertal life. In the foetus the cord of cells destined to
form the vagina first shows a lumen at the anterior end. This
lumen extends until it is separated from the exterior only by a
thin wall of cells, which remains in the rat and mouse until
the first oestrous period, when it is ruptured by the enlargement
of the vagina. The closure of the immature rodent vagina may
be analogous to the partial closure of the human vagina effected
THE fe:\iale reproductive organs
15
by the hymen. In the guinea-pig a remarkable mechanism
exists whereby the vaginal closure membrane is regenerated
after each oestrous period.
The clitoris. The clitoris, situated anteriorly to the vaginal
orifice, is a vestigial homologue of the penis. It is composed
of connective tissue, surrounded by more or less cornified
epithelium. Its homology with the penis is emphasized by the
presence (in the human] of erectile tissue; in such lower
Fig. 8. — Clitoris of Mouse,
b.e. band of epithelium running down from distal cleft; b.v. blood
vessels; u. urethra.
mammals as the rat and mouse, the urethra, instead of opening
to the exterior at the vulva, as is usually the case, traverses the
clitoris and emerges at its distal cleft. In certain mammals the
clitoris plays a subsidiary part in copulation, but otherwise its
value in the reproductive processes appears to be negligible.
The mammary glands. The mamnicc are usually bilaterally
paired organs, consisting of secretory alveoli from which the
milk is carried to the exterior by ducts. The ducts are gathered
together on the surface into the nipple to facilitate suckling.
The exact nature of the secretion which takes place in the
mammae is still a subject of controversy, but, whether or not
i6 INTERNAL SECRETIONS OF THE OVARY
actual cell destruction is involved, it seems probable that the
secretion is the manifestation of a katabolic phase resulting
from withdrawal of a growth stimulus.
The number of mammae and their distribution varies
widely in different species. The smallest normal number
appears to be one pair, while in the larger polytocous animals
(pig), as many as six to nine pairs may be found. The rat,
rabbit and mouse have five to six pairs. Where there are only
a.
•>!•. ^
■-^'Vi
'^-i '
vrti »»•►"» -•> '
T>
^«.-:~^
.•»!- .,^
■ a.i.
c.t.
Fig. 9. — Lobule of Mammary Gland of Cow.
a. alveolus; a.i. adipose tissue; c.t. connective tissue.
one or two pairs they may be either thoracic (Primates, elephant)
or abdominal (guinea-pig, cow, etc.). The presence of a large
number of mammae involves their distribution over both thorax
and abdomen. In certain animals the mammae have milk
reservoirs, which involve alterations in the shape of the gland,
resulting in the udder typical of ruminants. The rat, mouse,
rabbit, and ferret, on the other hand, have no receptacles for
the milk beyond the ducts, which may become greatly
distended; the glands in these animals are flat strips of tissue
beneath the skin. In the rabbit the gland can readily be
stripped off from both skin and body wall and can be prepared
THE FEMALE REPRODUCTIVE ORGANS
17
entire for examination with great ease, a fact which makes this
animal invaluable for experimental work. The mammary
Fig. 10. — Udder of Cow; stained to show Mammary Tissue.
(From Hammond).
gland differs from the other accessory organs in being differen-
tiated at a much later stage of development and also in being
rudimentary when puberty is reached.
P.S.O.
CHAPTER III
SEXUAL PERIODICITY IN THE FEMALE MAMMAL
Sexual periodicity in the mammalian female may be said to
consist of three cycles. The first — the attainment of puberty,
sexual maturity, and the decline of sexual function at the
menopause — is passed through but once. The second — the
periodic occurrence of the breeding season — appears one or
more times each year (or in extreme cases every two years).
Finally, the third consists of the cyclic periods of oestrus, at
which the actual mating takes place, and of which one or more
occur during each breeding season.
{a) PUBERTY AND THE MENOPAUSE
The development of the ovary before puberty tends to be
sporadic, and may include one or more waves of growth followed
by retrogressive changes. In the mouse, for instance, the growth
of the follicles is quite advanced at three weeks old, but de-
generation subsequently sets in, accompanied by a decrease in
the size of the ovary (80). The growth of the accessory organs,
on the contrary, appears to be gradual and continuous from the
time when they are first differentiated. The first abrupt change
in the accessory organs occurs at the first oestrous period, the
definite sign of the onset of puberty. The attainment of puberty
may thus be said to consist of two phases:
(a) The gradual pre-pubertal development of the accessory
organs.
(b) The abrupt appearance of the first oestrus and ovulation.
The first cestrous period, however, is the same as any other,
except for minor changes such as the appearance of the vaginal
orifice for the first time in the mouse and rat; the stimulus
SEXUAL PERIODICITY IN THE FEMALE MAMMAL 19
J!^|l#^^;¥^-.
■^.
ft%>S^
Fig. II. — Ovary of Mouse at three weeks old, showing
LARGE NUMBER OF SmaLL FOLLICLES.
Fig. 12. — Uterus of Mouse at three weeks old, showing
general lack of development.
20 INTERNAL SECRETIONS OF THE OVARY
causing the first oestrous period must be supposed to be the same
as that responsible for the later ones. The problem of the
causation of puberty thus resolves itself into two parts:
(a) What stimuli cause the gradual pre-pubertal development
of the accessory organs?
(b) By what means is the oestrus-producing stimulus first set
in motion?
These questions are discussed later (see p. 123).
Functional puberty is attained in many animals (notably in
man) before body growth is completed. This is an anomaly,
since animals becoming pregnant before growth is completed
rarely reach maximum size. In the rat, according to Long and
Evans (425), the vaginal orifice usually appears a little before the
first ovulation. In the mouse and the rat it has been observed
that the intervals between the first few oestrous periods tend to
be somewhat longer than normal (425, 491).
Long and Evans give the following data for the attainment of
puberty in the rat:
Table i. — Attainment of Puberty in the Rat
(after Long and Evans).
Age at opening of vagina
Age at first ovulation -
Length of ist cycle
Length of 2nd cycle
Length of 3rd cycle
Length of 4th cycle
Average.
72nd day of life
77th day of life
10 days
9 days
8-5 days
7-5 days
On the whole, however, the remarkable thing about the onset of
puberty is that the normal periodic activity is attained so
abruptly.
The decline of sexual function at the menopause is character-
ized by retrogressive changes in the ovaries and by the gradual
atrophy of the entire accessory sexual apparatus — Fallopian
tubes, uterus, vagina, vulva, and mammary glands. These
changes, which finally are similar to those occurring after
SEXUAL PERIODICITY IX THE FEMALE MAMMAL 21
ovariectomy, have been described, especially for the human, b}^
many writers.
The fading out of the sexual cycle at the menopause is far less
abrupt and regular than is its appearance at puberty. In the
human, the menopause extends over two or three years,
menstruation gradually becoming less frequent and less regular.
According to Magian (433) the cessation of menstruation does
'.C.I.
- \
►.:.--^>.
■^i-X'*.' ■''■'''''•
Fig. 13. — Ovary of Senile Mouse 15 months old, showing
LACK of Follicles and New Corpora Lutea {cf. fig. i).
O.C.I, old corpus luteum; s.a.f. small anoxular follicle.
not necessarily involve the complete disappearance of fertility,
and the ovarian cycle may therefore go on longer than the
uterine.
Data relating to the menopause in lower mammals are very
difficult to obtain. Domesticated animals are usually slaugh-
tered when the breeding function declines, and in laboratory
animals the decreased resistance to disease which occurs in old
age usually results in death about the time when menopause
symptoms appear. Nevertheless some data are available.
22 INTERNAL SECRETIONS OF THE OVARY
In the mouse the dedine of the sexual function usually
proceeds by the following stages;
(a) Litter size gradually decreases.
(b) Coitus becomes infertile owing to ovulation having ceased.
(c) CEstrous symptoms in the accessory organs become
infrequent and irregular.
{d) Complete menopause anoestrus appears.
By the last stage the ovary has become entirely devoid of
follicles as well as of corpora lutea vera and resembles to a great
extent the type produced by exposure to X-rays (see p. 138).
(6) THE BREEDING SEASON
The term ' breeding season ' was originally proposed by Heape
(287) to cover the time when activity occurs in the reproductive
organs. This definition, however, includes the period of preg-
nancy and lactation, and since some mammals may spend all
their reproductive life in one or other of these states, the term
loses some of its force. In discussing here the time of the year
at which breeding takes place, the term ' breeding season ' will
be used to denote the time at which a species comes into
oestrus, namely, in the sense that ' sexual season ' was used by
Heape.
In captive and domesticated mammals, as well as in man,
living under conditions of fairly constant food supply (and often
of temperature), the occurrence of a restricted breeding season
has become rare. The lower mammals in a state of nature,
however, have a definite season of the year at which mating
takes place, and, in general, this season is so placed that the
young are produced at an auspicious time. Copulation takes
place, even during the breeding season, only at certain definite
periods of oestrus (or ' heat '). Qistrus may occur only once in
a breeding season (monoestrous animal) or, in the absence of
pregnancy, a regular series of periods may occur (polyoestrous
animal). During the non-breeding season (except in pregnant
animals) the reproductive organs are in a state of quiescence.
This period is known as anoestrus in contrast to oestrus.
The breeding season is well shown in wild rodents, where a
SEXUAL PERIODICITY IN THE FEMALE MAMMAL 23
series of cestrous periods, probably separated by pregnancy,
occur during the spring and summer, while during the winter
months the reproductive organs enter into a prolonged state of
rest. In animals with a long period of gestation, what would
be the anoestrous period may be occupied by pregnancy.
Mares, for instance, having a period of gestation of 11 months,
foal about the beginning of the next breeding season.
The wild prototypes of the cow, pig, sheep and goat all
probably exhibit a restricted mating season, which in the case of
the last two appears to be in the autumn, but domestication has
resulted in the gradual expansion of the season until the
domestic strains of these animals will breed at almost any
period of the year. The greatest readiness to breed, however,
is still found at a time which probably corresponds to the
primitive breeding season, especially in the less highly domes-
ticated varieties.
The dog has two breeding seasons a year, in spring and
autumn, and only one cestrous period occurs in each season.
Whatever the primitive state of affairs may have been, the
monoestrous cycle, with its limited opportunities for the animal
to become pregnant, is the rarer condition at the present time in
such animals as have been studied. The cause of the onset of
the breeding season of mammals is obscure, but, in so far as
it occurs in spring and summer, it is probably a combined effect
of raised temperature and increased food supply (359, 509).
Civilized man retains practically no vestiges of a breeding
season beyond a slight seasonal variation in the birth rate
(especially the illegitimate birth rate), and therefore in the
conception rate. Many primitive tribes, however, show a
marked lumping of the births in one season of the year (444), and
it is possible that man originally had a definite, if not entirely
restricted, breeding season.
[c) ESSENTIAL FEATURES OF THE CESTROUS CYCLE
The essential feature of ovarian activity is the maturation
of the Graafian follicle and the discharge of the ovum. This
periodic occurrence, together with the intervening growth of
the corpora lutea, constitutes the ovarian cycle. While these
24 INTERNAL SECRETIONS OF THE OVARY
ovarian changes are in progress cyclic events are proceeding
in the accessory organs of reproduction — uterus, vagina, and
mammary glands. These events are collectively known as the
' oestrous cycle,' from oestrus, the central point of the cycle,
when ovulation and copulation take place.
After the anoestrous or pre-pubertal period of comparative
quiescence the cycle starts with the preliminary phase of
prooestrus, during which the follicles ripen and growth changes
take place in the accessory organs. This is followed by the
period of oestrus proper, during which ovulation takes place (in
some species only after copulation), accompanied by further
changes, usually of a retrogressive nature, in the accessory
organs. It is at this period only, in the lower mammals, that the
female will receive the male (but see p. 55). (Estrus is usually
followed by a short recuperative period — the metoestrus; sub-
sequent events depend primarily on the fate of the ova produced
at oestrus and on the species of animal.
In the absence of pregnancy, the monoestrous animal with
only one oestrus per breeding season, returns to anoestrus,
usually with an intervening period of development in the ovary
and accessory organs. The polyoestrous animal, on the other
hand, with a series of cycles in a breeding season, enters upon a
very transitory period of development, the dioestrous interval.
At the end of this short phase prooestrus supervenes and the
cycle starts again. The occurrence of successive oestrous
periods with no real interval of rest constitutes a dioestrous
cycle, of which the essential feature is that growing follicles or
corpora lutea are present the whole time in the ovary.
After ovulation, conditions in the non-pregnant female are
determined by the behaviour of the corpus luteum. This may
show only the most transitory development, as in the rat and
mouse, or, as in Marsupials and the ferret, it ma}^ undergo
development equal or nearly equal to that found during preg-
nancy, with very striking growth effects upon the accessory
organs. In the latter case the term ' pseudo-pregnancy ' ^ is
1 This term, coined by Matthews Duncan (172) to indicate a
psychological condition in the human female, was applied to the post-
ovulation phase in Dasynrus by Hill and O'Donoghue (300), and later
to the same phase in the bitch by IVIarshall and Hainan (449).
SEXUAL PERIODICITY IX THE FEMALE MAMMAL 25
given to this period of post-ovulative activity. The two chief
tvpes of cvcle may therefore be illustrated as follows:
i
Procestrus
(Estrus
Pregnancy
I
Pseudo-Pregnancy
Anoestrus
Diagram to Illustrate Moncestrous Cycle.
One oestrous period per breeding season. A complete cycle occu-
pies the whole breeding season, which is separated from the next by
anoestrus (as found in dog).
Y
CEstrus
Pregnancy
Metoestrus
Dioestrus
TD
seudo-Pregnancy
Prooestrus
Single Dioistrous Cycle, which is repeated a number
OF TIMES during THE BREEDING SEASON.
In the rat and the mouse pseudo-pregnancy is dependent upon
sterile copulation, while in the guinea-pig dioestrus is really pseudo-
pregnancy (see p. 35).
The general account given above has assumed ovulation to be
spontaneous, i.e. to occur automatically at oestrus irrespective
of copulation. In two common mammals, however, the rabbit
and ferret, ovulation occurs only after copulation. In the
absence of the male, oestrus, including the presence of mature
follicles in the ovar}^ persists right through the breeding season,
at the end of w^hich follicular atresia takes place. In these
26 INTERNAL SECRETIONS OF THE OVARY
two animals, therefore, no cycle exists in the unmated animal,
except the alternation of anoestrus and breeding season.
Anoestrus
1 i
(Estrus (with coitus)
1
Qistrus (without coitus)
Pregnancy Pseudo-Pregnancy
1 ! ■ ; 1
Anoestrus
Metoestrus
1
i
Diagram of Cycle where Ovulation is dependent on
Copulation, as in the Rabbit and Ferret.
As has been pointed out above, cyclic changes occur in the
secondary organs in correlation with those in the ovary. The
extent to which the various accessory organs participate in the
oestrous cycle shows much specific variation. All species which
have been studied show considerable uterine changes, but
vaginal and mammary cycles have been less frequently described.
In the ferret, guinea-pig, rat, and mouse, however, vaginal
changes at least as marked as the uterine ones are well known,
while cyclic mammary changes in the unmated female have been
described in the rat (472, 605), guinea-pig (402), cow (265), and
human (159); in other species, apparently, they may be absent.
The extent to which the vagina participates in the cycle entirely
determines the application to any particular species of the
vaginal smear technique (see p. 95).
The exact nature of the changes taking place in the accessory
organs varies very much from animal to animal, but they are
roughly comparable; a comparison of the ovarian and uterine
cycles is given on p. 27.
For descriptive purposes it is convenient to use Loeb's (400)
division of the cycle into two main phases: (a) the follicular
phase, during which the Graafian folhcles mature and ovulate
(this includes prooestrus and oestrus) and {b) the luteal phase
(including dioestrus, pseudo-pregnancy and pregnancy) during
which the corpus luteum dominates ovarian activity.
SEXUAL PERIODICITY IN THE FEMALE MAMMAL 27
Uterine cycle (and
Phase
Ovarian cycle
vaginal cycle
where occurring)
Anoestrus
Rest
Rest
Prooestrus
Maturation of fol-
licles
Growth
CEstrus
Ovulation
Degeneration
Copulation
Pseudo-
Development of
Extensive pseudo-
Pregnancy
corpus luteum
pregnant development
Anoestrus
Rest
Rest
Metoestrus
Formation of cor-
pus luteum
Recuperation
Dioestrus
Transitory deve-
Transitory or no deve-
lopment of cor-
lopment
pus luteum
Prooestrus
Maturation of fol-
licles
Growth
The diversity in the details of the cyclic changes in the female
genitalia makes any sort of classification of the types of cycle
extremely difficult. Scarcely two mammals, except the rat and
mouse, are entirely comparable, and species which are closely
related zoologically may show entirely different features in the
sexual cycle. The rat and the rabbit, for instance, are extra-
ordinarily different in their reproductive phases. The division
of animals into monoestrous and polyoestrous is unsatisfactory
from a descriptive point of view because {a) the condition varies
under domestication, and (b) the only common true monoestrous
Eutherian is the dog.
Certain animals, however, have various salient points in
common. The rabbit and ferret, for instance, both of which
remain on oestrus throughout the breeding season in the absence
of copulation, and do not ovulate spontaneously, are comparable
to some extent. In the description of the oestrous cycles of the
chief mammals, comparable species are as far as possible
discussed together.
CHAPTER IV
TYPES OF (ESTROUS CYCLE
{a) MARSUPIALS
Dasyurus. In Dasyurus vivierriniis, studied by Hill and
O'Donoghue (300), O'Donoghue (476), and Sandes (544), the
anoestrous period lasts more than half the year. This is termi-
nated by the onset of the active phase of the reproductive cycle,
which is divided by Hill and O'Donoghue into four stages — pro-
oestrus, oestrus, postoestrus, and pregnancy or pseudo-pregnancy.
During prooestrus, which lasts from four to twelve days, char-
acteristic changes occur both in the uterus and in the external
genitalia. The marsupial pouch, for instance, enlarges, but
according to O'Donoghue no mammary changes occur during
oestrus. The uterine mucosa increases in thickness and becomes
very vascular, the glands lengthen and become convoluted,
while the epithelium tends to thicken. These processes are
continued during oestrus, which lasts only for one or two days,
and at which time copulation takes place. Ovulation is delayed
until the next stage, to which Hill and O'Donoghue gave the
name of 'postoestrus'. The length of this phase appears to be very
variable, but the authors state that ovulation does not occur
until five or six days after the end of oestrus. Ovulation is
spontaneous and is remarkable because of the large number of
ova liberated. Pregnancy lasts not less than eight and not more
than fourteen days. In the absence of conception, changes occur
which are essentially the same, and of the same duration, as
those during pregnancy. Corpora lutea, indistinguishable from
those of pregnancy, are formed in the ovary (544). Typical
changes in the mammary glands, the uterus, and the marsupial
pouch also occur. As a result of a detailed study of the
growth changes in the mammary apparatus, O'Donoghue
came to the conclusion that no difference is observable
28
TYPES OF (ESTROUS CYCLE
29
between the development of pregnancy and of pseudo-
pregnancy.
The Opossum. The opossum was originally thought to be
monoestrous, like Z)(^s\7/r//s, but Hartman's (271, 278] detailed
study of the cycle shows it to be polyoestrous. The di-
oestrous cycle in the opossum is, however, complicated by a
CEstrus Ovulation
— I — I — r~
Graafian
follicle
Parturition
CEstrus
+
0
L— J \ \ 1 1 1 1 1 I I L_L_1 I I I I \ I I I I I I I
10
15
20
25 days
Fig. 14. — Diagram of CEstrous Cycle in the Opossum.
(From Hartman).
definite period of pseudo-pregnancy which, as a rule, is not
well marked in polyoestrous animals. The recent work on the
opossum has been greatly facilitated by the fact that this Mar-
supial, like the rat, mouse, and guinea-pig, shows characteristic
oestrous changes in the vagina. With the aid of a vaginal smear
technique, Hartman has been able to study in detail the various
phases of the cycle, the total length of which is about twenty-
eight days. He points out, however, that its duration may be
30 INTERNAL SECRETIONS OF THE OVARY
disturbed by follicular atresia. The opossum is remarkable
among polyoestrous animals for the fact that pregnancy, which
lasts only about thirteen days, occurs without disturbing the
normal periodicity of oestrus.
The ovarian cycle consists of the rapid growth of Graafian
follicles just before oestrus, and ovulation about one day later.
The uterine cycle is characterized by growth at oestrus, which
is continued during the pseudo-pregnant period or during
pregnancy. During anoestrus the uterus shows a reduction of
both the mucosaand themuscular layer. The glands are straight
or only very slightly coiled, while the lumen is small and the
epithelium has one layer. At prooestrus, when the Graafian
follicles have attained about a quarter of their maximum size,
the uterus undergoes distinct changes. The gross size becomes
greaterowing to increase in the various elements, to increased vas-
cularity, and, above all, to infiltration of lymph into the mucosa.
These changes begin in early prooestrus and culminate some days
after oestrus. By the time that oestrus sets in, the uterus is
considerably swollen and the glands greatly coiled. The
lumen has become much enlarged. After ovulation the growth
of the uterus continues, and is characterized by further hyper-
trophy of the mucosa and glands. At about the eleventh day
after ovulation, in the absence of pregnancy, the uterus under-
goes atrophy. The mucosa collapses owing to the withdrawal
of lymph, and the epithelial glands undergo degeneration. The
uterus then returns to the dioestrous condition in which the
epithelial glands consist of low columnar or cuboidal cells, and
degenerating material is found in the lumen.
The vaginal cycle shows equally obvious changes. During
anoestrus, the epithelial lining is thin and there is no cellular
debris in the lumen. At prooestrus the vagina grows in diameter
and the epithelium becomes thickened. At the end of this stage
the mucosa is at least twelve to fifteen cells thick. No
leucocytes are to be seen, and the epithelium and the vaginal
smear consist of large, flat, nucleated cells. At oestrus these
nucleated cells are replaced in the vaginal smear by true
cornified cells which arise from the cornified mucosa of the
vagina. After ovulation, leucocytes begin to appear in
the vaginal smear, and within three or four days the smear
TYPES OF (ESTROUS CYCLE 31
consists of leucocytes with only a few cornified cells. Imme-
diately after this, nucleated epithelial cells appear again.
During dicestrus the vaginal smear consists of nucleated
epithelial cells with a number of leucocytes. The opossum is
remarkable in having lateral vaginal canals which undergo a
similar cycle.
The mammary glands of the opossum have a very clear cycle.
Growth begins at prooestrus, and continues uniformly until the
end of pseudo-pregnancy or the end of true pregnancy. It is
necessary to suppose that the beginning of this growth is under
the control of the oestrus-promoting stimulus and that later the
corpora lutea are responsible. The growth occurring during
prooestrus is, however, comparatively slight compared with that
found during pseudo-pregnancy. At the end of pseudo-preg-
nancy or true pregnancy, atrophy sets in, and the lowest point
of development is reached about twenty-three days after the
previous ovulation or some live days before the next ovulation
is due.
{b) DOG
Some observations on the ovary of the dog were made by
Bischoff (66) as early as 1845. Bouin and Ancel (75) and Van
der Stricht (602) later studied the ovarian cycle. Fried-
laender (233) appears to have been the first to deal with
the uterine mucosa, while Retterer (533) and Keller (319)
published more detailed work. The whole subj ect of the oestrous
cycle in the dog has been investigated in detail by Marshall and
Jolly (450), Marshall and Hainan (449), and more recently by
Gerlinger (240, 241) and Evans and Cole (185).
With the exception of the vagina, the cyclic changes in the
reproductive organs are very definite. During anoestrus, the
ovary is small and contains neither large follicles nor functional
corpora lutea. Primordial follicles are, however, developing.
The uterus is thin and anaemic. The mucosa is shallow, and the
glands and vessels are few. Prooestrus is characterized by
follicular growth in the ovary, while thickening of the mucosa,
accompanied by congestion and multiplication of the stromal
capillaries, takes place in the uterus. The glands of the mucosa
begin to secrete. According to Marshall and Hainan, the
32 INTERNAL SECRETIONS OF THE OVARY
mammary tissue in the virgin bitch shows no growth character-
istic of prooestrus or oestrus.
During oestrus, ovulation takes place and the growth phase in
the uterus is superseded by a regressive one, which is char-
acterized by the breakdown of the capillary walls and the
extravasation of blood into the stroma. Aided by a certain
amount of destruction of the superficial epithelium, the blood
poly III./ \®^^ ^ ^ _^\ ^^
Fig. 15. — Procestrous Uterine Mucosa of Dog.
(From Marshall and Jolly).
ex.bl. extravasated blood corpuscles; polym. polymorph; sec. cells
probably indicating secretory activity.
corpuscles find their way into the lumen of the uterus and thence
to the exterior, giving rise to the external bleeding characteristic
of early oestrus in the dog. During metoestrus the corpora lutea
are beginning to form in the ovary, and the uterine mucosa
regenerates. If pregnancy does not occur, pseudo-pregnant
changes take place, correlated with the development and
persistence of the corpus luteum in the ovary. Pseudo-
pregnancy is characterized by considerable enlargement of the
uterus and by growth of the mammary glands. Gerlinger
distinguishes two layers of the uterine stroma during this period ;
TYPES OF (ESTROUS CYCLE
33
a deep spongy foundation, and a compact superficial layer
crowded with shallow crypts. At about eight to nine weeks
after oestrus, correlated with the atrophy of the corpus luteum,
regressive changes take place in the accessory organs. These
^.r\i>; . f\:.y
Fig. i6. — Uterine Mucosa of Dog at the end of
Pseudo-Pregnancy.
Extra vasated blood is seen in the stroma.
(From Marshall and Hainan).
changes lead to the secretion of milk in the mammary glands, and
to the breakdown of capillaries and to the extravasation of blood
into the stroma of the uterus. In spite of a similarity to the
prooestrous phenomenon, this pseudo-pregnant degeneration
in the uterus is readily distinguished by a difference in the
p.s.o. c
34 INTERNAL SECRETIONS OF THE OVARY
epithelium and glands. At the end of pseudo-pregnant
degeneration, anoestrous quiescence supervenes.
In the early stages of pregnancy, changes similar to those of
pseudo-pregnancy are found. Subsequently, of course, the
development of both uterus and mammary glands is in excess
of that found in the non-pregnant animal. Lactation occurs
in what would be the next anoestrous period, so the compli-
cation of coincident oestrus and lactation does not appear.
For the same reason the bitch, unlike rodents, cannot suckle
and gestate at the same time.
{c) GUINEA-PIG AND COW
The guinea-pig and cow both have a dioestrous cycle contain-
ing a luteal phase, which has been shown experimentally to be
under the control of the corpus luteum. They are also similar in
having a relatively long period of gestation, so that the young
of both are born in an advanced stage of development.
Guinea-pig. Early workers on the oestrous cycle in the guinea-
pig include Bischoff (67), Reichert (528), Hensen (289), Rein
(532), and Rubaschkin (539). These workers all recognised that
'heal' occurred very soon after parturition and that ovulation
was spontaneous. They failed, however, as did early workers on
the rat and mouse, to trace the cycle without taking parturition
as the starting-point. During the last twenty years very
extensive investigations on the uterine, ovarian, and mammary
cycles in the guinea-pig have been carried out by Loeb (379,
389, 391, 393), while more recently the whole subject has been
reinvestigated by Stockard and Papanicolaou (599), who
elucidated the vaginal cycle and introduced the vaginal smear
technique. According to Stockard and Papanicolaou the length
of the dioestrous cycle is about sixteen days (or rather longer in
winter); Loeb's observations agree roughly with this figure,
except that he found greater variability and a tendency towards
long cycles after copulation. Voss (625) gives the length of the
whole cycle as varying between thirteen and twenty days, of
which dioestrus occupies nine to sixteen days.
Sterile copulation exerts no such influence on the time of
appearance of the next oestrous period as it does in the rat
TYPES OF (ESTROUS CYCLE 35
and mouse. This is probably correlated with the fact shown by
Loeb that the corpora lutea function in postponing the next
oestrus, even in the unmated cycle of the guinea-pig.
The guinea-pig is similar to the rat and mouse in having an
immediate post-partum cestrous period, but it differs from these
two animals, according to Loeb and Kuramitsu (406), in that
inhibition of oestrus is not found during the rest of lactation.
The second post-partum oestrus in the guinea-pig occurs in
about sixteen days, i.e. at the normal interval. Lactation, in
other words, fails to cause unusual persistence of the corpora
lutea from the immediate post-partum ovulation. Since the
mouse requires to be suckling more than two young for this
result to be produced, the difference in the guinea-pig is probably
accounted for by the relatively small demand made on the
mother by the smaller-sized, more m.ature litter.
The ovarian cycle in the guinea-pig is rather remarkable,
according to Loeb (398), who has described waves of follicular
growth during both dioestrus and pregnancy. Atresia, however,
not ovulation, results from this growth. Following ovulation,
small follicles become medium-sized in some six days and large
in another two, after which atresia sets in. The next wave of
follicular growth culminates at the following oestrus and results
in ovulation of some of the mature follicles and atresia of the
remainder. During pregnancy there are two or more waves of
follicular growth ending in atresia. The corpus luteum be-
comes fully formed about five days after ovulation. Regressive
changes are first visible on the tenth day and accelerate rapidly
until the next oestrus. By ten days after the new ovulation,
the corpora lutea of the previous oestrus are reduced to small
vacuolar bodies surrounded by a connective tissue capsule.
During pregnancy the developmental stage of the corpora lutea
lasts longer and results in the formation of larger structures.
Degenerative changes, similar to those found in the dioestrous
corpora lutea, are observed only after the fortieth day.
The post-ovulation condition of the uterus has been described
at length by Loeb. According to his account, there is no marked
hypertrophy such as takes place during the luteal phase in the
dog, but definite changes occur in the epithelium and stromal
glands of the uterus, and in its physiological condition.
36
INTERNAL SECRETIONS OF THE OVARY
The vaginal cycle of the guinea-pig is remarkable for the fact
that during the dioestrous interval the vaginal orifice is normally
closed by an epithelial membrane. This is ruptured at the
approach of procestrus by the turgidity of the vulva. Stockard
and Papanicolaou have divided procestrus and oestrus (which
together last about twenty-four hours) into four stages according
to the nature of the vaginal contents (Table 2).
Table 2. — Vaginal changes during (Estrus
IN THE GuINEA-PiG
(After Stockard and '.
Papanicolaou).
Staee
Nature of contents
Duration
Condition
Condition of
of Vagina
(hrs.)
of Ovary
Vagina
I
jMucous secretion
6-12 hrs.
Mature
Vaginal epithel-
Epithelial cells
follicles
ium being shed.
(cornifiecl)
Infiltration of leu-
cocytes beginning
II
Cheesy. Great num-
bers of epithelial
cells (nucleated)
2-4 hrs.
Ovulation
Leucocytes in-
creasing. Desqua-
mation of epithe-
lium continuing.
III
Fluid thinner. Leu-
cocytes beginning
to appear
4-6 hrs.
Congestion. Leu-
cocytes passing to
lumen.
IV
Large numbers of
1-2 hrs.
Corpus
Rupture of few
leucocytes. Some
luteum
capillaries, fol-
haemorrhage
forming
lowed by regenera-
tion of epithelium.
Copulation, accompanied by the formation of the vaginal plug,
takes place during the first stage, and the corpora lutea are fully
formed four to five days later. Stockard and Papanicolaou con-
sider the breakdown of the vaginal epithelium to be due to the
removal of the protective influence of the corpora lutea of the
previous ovulation, and to be homologous with human mens-
truation. The time relation of ovulation is, however, radically
different from that found in the human.
Loeb and Hesselberg (402) have described a growth cycle in
the mammary glands of the non-pregnant guinea-pig corres-
ponding with the dioestrous cycle, a burst of growth accom-
panying each oestrus. Following oestrus, the glands regress and
there is no reactivation during the luteal phase of the cycle.
TYPES OF (ESTROUS CYCLE 37
Thus, during the time of maximum development of the corpora
lutea in the non-pregnant animal, the mammary gland is at its
minimum, so that the luteal phase of mammary growth is
presumably missing in the normal non-pregnant guinea-pig.
Prolongation of the life of the corpora lutea by hysterectomy or
by deciduomata formation results in considerable mammary
proliferation.
Cow. The importance of the cow from an agricultural point
of view has attracted a large number of workers, among whom
may be mentioned Schmaltz (547), Kiipfer (336), Zeitzschmann
(645), Murphey and co-workers (468-9), Frei and Metzger (232)
and more especially Hammond (265), from whose work the
following account is largely compiled.
The complete dicestrcus cycle in the cow lasts about three
weeks, of which oestrus (as determined by discharge of mucus
and willingness to copulate) occupies twelve to twenty-four hours.
Much variation is, however, found according to breed, age,
condition of animal, and time of year. Ovulation is spontaneous,
and occurs 0-24 hours after the beginning of heat. Only one
ovum is normally produced at each ovulation. The ovaries
generally ovulate alternately, though this is not invariable. The
rupture of the follicle is accompanied by a small hcnemorrhage.
Three days after the beginning of heat the ovaries contain the
young corpus luteum, an old corpus luteum from the last cycle
(usually in the opposite ovary), and a number of small follicles.
Eight days after 'heat,' the old corpus luteum has become
insignificant, the new one is fully formed, and a new large
follicle has appeared (usually in the same ovary as the old corpus
luteum). Ten days later, i.e. three days before the beginning of
the next heat, the ovaries are in much the same condition,
except that the old corpus luteum has practically disappeared,
and slight growth has occurred in the new large follicle. In
another two days (one day before heat) degenerative changes,
accompanied by decrease in size, have appeared in the recent
corpus luteum, and rapid follicular growth is taking place.
The ovarian cycle in the cow is shown diagrammatically in
Fig. 17, taken from Hammond. This diagram suggests that the
follicle which will ovulate at the next oestrous period can already
be distinguished from the 'reserve' of small follicles at the time
38
INTERNAL SECRETIONS OF THE OVARY
o
U
H
<
Z
O
W
Ph
o
w
u
o
« S
< o
O t
o
TYPES OF GESTROUS CYCLE 39
of the previous ovulation. Thus the cow differs from the mouse
(see p. 137) where the fohicles which will ovulate at the next
oestrous period become distinguishable only during the first half
of the preceding dioestrus.
The cyclic changes in the accessory organs of the cow do not
appear to have been entirely worked out. Hammond describes
the uterus as secreting a large amount of fluid at oestrus (as
occurs in other mammals) and showing congestion during the
eight days following heat. According to Murphey the vaginal
epithelium undergoes development up to eighteen days after the
beginning of the previous heat, and this is followed by general
desquamation and reformation of the epithelium during the next
week. Haemorrhage from the vaginal stroma, accompanied by
leucocytic infiltration, is usual after heat. A more detailed
description is given by Frei and Metzger, whose illustrations of
the vaginal contents suggest that some prooestrous cornification
may occur, as in the rat and mouse. The degree of cornification
must be inconsiderable, however, compared with that found in
the rodents.
Hammond has described cyclic changes in the mammary
gland of the virgin animal correlated with the oestrous cycle.
Before puberty the glands consist merely of ducts, but after the
first ovulation there occurs development of lobules of alveoli
associated with, but lagging behind, the development of the
corpus luteum. The gland never returns to the pre-pubertal
state, but regression of the alveoli takes place during the second
half of each dioestrus. Hammond does not describe any growth
characteristic of the actual period of heat.
Lactation in the cow does not appear to have any effect upon
the recurrence of the cycle in the ovary. Cows are normally put
to the bull soon after parturition, and milking is continued
until six to eight weeks before the next calf is due.
{d) HORSE, SHEEP AND PIG
These animals under conditions of domestication have a
dioestrous cycle which probably contains a luteal phase con-
trolled by the corpus luteum, but precise experimental infor-
mation is lacking.
40 INTERNAL SECRETIONS OF THE OVARY
Horse. Until recently very little accurate information was
available as to the oestrous cycle in the mare. Heape (287) made
the general observation that the length of the dioestrous cycle
is three to four weeks, while Marshall (444) states that the dura-
tion of oestrus is about a week, becoming shorter, according to
Ewart (i8g), as the season advances.
Lately, Seaborn and Champy (553), Seaborn (552), and Aitken
(4) have brought forward much more data.
Seaborn gives the length of the dicestrous cycle as twenty-four
days, while Aitken found the average to be twenty-two or
twenty-three days, with a normal variation from twenty to
twenty-five days. The duration of prooestrus and oestrus is three
days each, according to the former authors. Copulation takes
place only during cestrus. Aitken gives the duration of oestrus
as seven days, with normal variation from four to eleven days.
Both agree that ovulation occurs towards the end of oestrus.
The ovary of the horse is rather remarkable for its size and
fibrous nature. Ovulation, which is spontaneous at oestrus,
takes place from an ovulation fossa. According to Aitken,
germinal epithelium covers this part of the ovary, while the rest
is covered by peritoneum. Aitken gives the diameter of the
mature folhcle as about 6 cms. and of the fully developed corpus
luteum as 4 cms. An ovary containing a ripe follicle weighs
about 300 gms. and one during dioestrus about 60 gms.
Ovulation is not necessarily alternate, and Aitken found a high
proportion of double ovulations, often from one ovary. The
maturation of the folhcle, and its differentiation from the group
of small follicles, seems to occur very rapidly before ovulation.
Regressive changes take place in the corpus luteum during the
second half of dioestrus.
The uterine cycle has been roughly described by Seaborn (552) ,
the central point being the usual proliferation of the mucosa
at prooestrus. No vaginal cycle has so far been described
and Aitken found that vaginal smears showed no definite cycle.
As Marshall states that the mammary gland becomes congested
and enlarged during cjestrus, it would appear that a cycle
exists in this organ in the non-pregnant animal.
Sheep. The sheep is a good example of the transition from
the monoestrous to the polyoestrous condition. Most wild
TYPES OF (ESTROUS CYCLE 41
species, according to Marshall (444), are probably monoestrus,
having only one oestrous period in the short breeding season. In
captivity, however, and particularly under conditions of domes-
tication, two or more dioestrous cycles appear during the breed-
ing season, while the Australian Merino is said to experience
an unbroken series of dioestrous cycles throughout the year in
the absence of pregnancy. In Great Britain the breeding season
lasts from two to three months. According to Marshall the length
of the cycle is about fifteen or sixteen days, though this doubt-
less varies according to breed and nutrition . Other authors have
given from two to four weeks as the length of the cycle.
Neither vaginal nor mammary cycles have been described in the
sheep, but Marshall (440) has given a full description of the
changes taking place in the non-pregnant uterus. These changes
consist essentially of the phases of rest, growth, destruction and
regeneration, such as have been described in the dog.
During the growth phase the mucosa increases in thickness,
the stroma undergoes cell division, and uterine congestion
begins. The period of destruction is characterized by the
breaking dow^n of some of the capillaries and extravasation of
blood into the stroma. Bleeding into the uterine cavity does not
usually occur. The extra vasated blood usually remains under
the endometrium, giving rise to pigmentation. This period of
destruction, corresponding to the end of prooestrus, is followed
by a period of recuperation. Subsequently the uterus returns to
the condition of rest.
Pig. The duration of the dicestrous cycle in the sow varies
from two to four weeks, but is usually three weeks. Struve (604),
in a statistical investigation, found the average to be 20-66 ± -205
days. The external signs of heat are excitement of the animals,
and swelling of the vulva, from which there may be a slight
(possibly sanguinary) discharge. According to Corner (122),
however, the blood is of vulval rather than internal origin.
CEstrus lasts about three days.
The ovarian cycle in the pig has been described in detail by
Corner (122). About three days before the onset of oestrus, the
follicles due to ovulate undergo rapid growth and finally attain
a diameter of 8-10 mms. Ovulation occurs towards the end of
oestrus according to Lewis (362), but Corner and Amsbaugh
42 INTERNAL SECRETIONS OF THE OVARY
(128) think that it is probably on the second day. The ova
reach the uterus in about four days, and, if fertihzation has
taken place, become implanted eight to ten days after ovula-
tion. The corpus luteum attains its maximum size in the non-
pregnant animal in about ten days, at which time it is 8-9 mms.
in diameter. The corpus luteum of pregnancy may enlarge to
a diameter of lo-ii mms.
CEstrus
CEstrus
Days 0 2 4 6 8 10 i: 14 16 lb 20 22 24 26 28 30 32 34 36 38 40 42
Fig. 1 8. — Diagram of Ovarian Cycle in Non-Pregnant Sov\\
(From Corner).
At about the tenth day after ovulation in the non-pregnant
animal a comparatively sudden regression takes place in the
corpus luteum, which in two or three days shrinks to less than
half its previous volume, largely owing to the disappearance of
the granulosa lutein cells. This degeneration is followed by the
growth of the next group of follicles. The ovarian cycle in the
unmated animal is shown by Corner as in Fig. i8.
The changes in the uterine mucosa corresponding to the
ovarian cycle have been studied in detail by Corner. During
oestrus the cells are of low columnar type and are closely packed
together. Mitosis is frequent, and the mucosa has the appear-
ance of active proliferation. The glands, however, do not show
this. The sub-epithelial connective tissue is crowded with
polymorph leucocytes. The uterine stroma is oedematous.
During the first week after cestrous growth, a further pro-
liferation takes place in the mucosa cells, which, by the end of the
first week, have become columnar. There is also a burst of
mitosis in the gland cells, beginning three to four days after
TYPES OF (ESTROUS CYCLE 43
ovulation. All mitosis, however, ceases about the end of the
first week. At about the tenth day the mucosa cells are again
reduced to the low columnar type, with the extrusion of
cytoplasmic processes, and the gland epithelium returns to
normal. During the last few days of dicestrus the mucosa cells
become more elongated and large numbers of leucocytes appear
in the stroma.
This building up of the uterine mucosa during the first half of
dioestrus, though comparatively slight, is clearly correlated with
the development of the corpus luteum, and is unaffected by the
fate of the ova. It may be considered, in the non-pregnant
animal, as an abbreviated pseudo-pregnancy.
Vaginal and mammary changes during the dioestrous cycle in
the pig do not seem to have been described.
As in the cow, mare, and guinea-pig, lactation has no effect on
the recurrence of the cycle, except that, according to Marshall
(444), there is an interval of five weeks after parturition before
the next oestrus. Struve (604), however, states that oestrus
recurs four to nine days after parturition.
(e) MOUSE AND RAT
In spite of the ease with which the animals can be kept for
observation, accurate data on the oestrous cycle of the rat and
mouse have only recently become available. Earlier workers
were much handicapped by the difficulty of external diagnosis
of oestrus, and were compelled to use the immediate post-partum
oestrus as their starting-point. Sobotta (583), working from this
period, put the length of the cycle in the mouse at about twenty
days, while Lataste (358), from the intervals between sterile
copulations, placed the length of the cycle at about twelve days.
Long and Smith (427), and Smith (572) estimated the length to be
sixteen to nineteen days. In the white rat Morau (463), Lataste
(358), Long and Quisno (426). and Heape (287), all put the length
of cycle at ten days. The discovery, however, that a very defi-
nite vaginal cycle exists in both these animals has made it pos-
sible to analyse completely the length of the oestrous cycle. In
the ordinary unmated mouse and rat oestrus occurs about every
five days. Long and Evans (425) for the rat give an average length
44 INTERNAL SECRETIONS OF THE OVARY
of cycle of 5-4 days for 1,999 cycles. Of these 82 per cent, had
an average of 4-6 days. In the mouse the length of the cycle is
given by Allen (6) as between four and five days, while the
present author (498) found a mean length of 6-2 days for 1,000
cycles. The first oestrus occurs in the rat when about ten to eleven
weeks old, while in the mouse the cycle begins at the age of
about seven weeks. The earlier cycles appear to be slightly
longer than those occurring in the fully mature animal. In the
mated mouse, in the absence of pregnancy, the interval between
sterile copulations is about twelve days, and this increase is due
to the occurrence of a pseudo-pregnant period. Long and Evans
found the interval in the rat to be about thirteen days and also
showed that the mechanical stimulation of the cervix by the
vaginal plug results in the ovarian changes associated with the
pseudo-pregnant period (see also p. 171). Since copulation was
originally used as the criterion of oestrus, this lengthening of the
cycle after sterile copulation explains the error in the original
estimates.
(Estrus occurs within twenty-four hours after parturi-
tion and is then in abeyance during the whole of lactation,
provided that a normal-sized litter is being suckled. In the
mouse this dioestrous period during lactation lasts for about
three weeks (491). In the rat, it lasts rather longer, twenty-five
to forty days according to Long and Evans, varying with the
number of young suckled. This disappearance of oestrus during
lactation is correlated with ovarian changes which result from
lactation. The abnormal prolongation of lactation does not
result in further prolongation of the dioestrous period. By
foster-mothering young litters, it is possible to prolong lactation
in the mouse for forty to fifty days without materially extending
the length of the lactation dioestrus (495). If one or two young
only are being suckled, inhibition is not set up by lactation and
the cycle after parturition is normal. If pregnancy supervenes
from the immediate post-partum oestrus the female mouse and
rat may be both pregnant and lactating at the same time.
Under these conditions the length of pregnancy in the mouse
is increased from the normal nineteen or twenty days up to as
much as twenty-eight days. According to Kirkham (323) this
is due to delay in implantation of the embryos.
TYPES OF (ESTROUS CYCLE
45
The period of oestrus is marked by very definite clianges in
tlie vagina and uterus, and by follicular maturation in the
ovaries. Changes in the mammary gland may also occur during
the ordinary unmated cycle.
Ovarian cycle. The morphological picture of the ovarian cycle
is complicated by the fact that the corpora lutea survive
histologically long after their function is in abeyance, and in the
non-mated adult animal four or five sets of corpora lutea.
n.c.l.
Fig. 19. — Ovary of Mouse (Pasini Stain), showing two
SETS OF Corpora Lutea.
n.c.l. new corpora lutea; o.c.l. old corpora lutea.
representing previous ovulations, may be found at one time.
A large number of medium-sized follicles are present in the
ovary of the mouse at the beginning of dioestrus, and in this
large group it is difficult or impossible to pick out the set destined
to ovulate at the next oestrous period. At about the middle of
dioestrus, however, certain of the follicles undergo a rapid phase
of growth. This growth phase becomes more rapid during
prooestrus, and culminates in ovulation during the period of
oestrus — late in oestrus according to Allen (6), but earlier
according to other workers (82). Long and Evans state that in
the rat ovulation may occur any time after eighteen hours from
46
INTERNAL SECRETIONS OF THE OVARY
the beginning of vaginal cornification. The passage of the ova
down the tube is facihtated in both animals by the accumulation
of fluid in the periovarian cavity at oestrus. During the next
two days the corpora lutea develop, and in the unmated normal
rat they attain their maximum growth about three days after
oestrus. This set of corpora lutea may be distinguished histo-
logically from the previous sets without the aid of vital staining,
but the previous sets can only be separated reliably by means of
this technique, or by special histological examination of the
Fig. 20. — Uterus of Mouse : (a) during Dicestrus, (b) during
(EsTRUs, showing typical distension of Uterus and Ovarian
Capsule during Qistrus.
lutein granules. According to Long and Evans, degeneration can
be detected by the latter technique in the new corpora lutea at
the beginning of the next oestrus. During pseudo-pregnancy
growth of the follicles is postponed until about the usual time
before the next oestrus. This postponement of growth is un-
doubtedly brought about by the activity of the corpora lutea,
which undergo greater development during pseudo-pregnancy
than in unmated animals. In the same way ovulation is entirely
inhibited during true pregnancy, and follicular maturation,
beginning when the corpora lutea atrophy just before parturi-
tion, is completed only in time for the post-partum ovulation.
No ovulation takes place between the oestrous period occur-
ring immediately after parturition and that marking the end
of the lactation dioestrus.
Uterine cycle. The uterine cycle in both the mouse and rat is
closely synchronized with the ovarian cycle. During dioestrus
TYPES OF (ESTROUS CYCLE
47
the uterus is constricted and anaemic. At prooestrus dilation
of the lumen has begun and at the time of ovulation the uterus
has increased to about twice the normal diameter; the stroma
and muscular material are then very much attenuated. The
increase in size is caused solely by the distension of the lumen.
i?^'*^
Fig. 21. — Uterus of Mouse during Dicestrus.
During metoestrus the uterus gradually returns to its dioestrous
size, and in the unmated animal no further change takes place
until the next prooestrus. During pseudo-pregnancy, however,
growth of the stroma takes place, and at the end of pseudo-
pregnancy, correlated with the atrophy of the corpora lutea, a
certain amount of haemorrhage into the lumen may occur.
Vaginal cycle. The vaginal changes during the oestrous cycle
are also very pronounced and are now used for the detection of
the oestrous cycle in the intact animal. During prooestrus the
48
INTERNAL SECRETIONS OF THE OVARY
vaginal mucosa undergoes growth and becomes many layers
thick. The subsequent degeneration of this growth takes the
form of keratinization of the cells, which are subsequently
sloughed off into the lumen and can be collected for examination.
This stage is followed by the infiltration of vast numbers of
.-r^-^'^""^?:-:. '...d^i^*-'-^^— «^'^'^*<^^,^ ^ •*> "^^.^
kiwi's; ..
Fig. 22. — Uterus of Mouse during CEstrus, showing
distension of lumen.
leucocytes from the vaginal stroma. The vaginal contents,
therefore, show typical cyclic changes, which for the mouse
may be summarized as follows :
Dicestrus. Epithelial cells, largely nucleated, and leucocytes
constitute most of the vaginal smear during this stage. The
smear tends to be fairly fluid in the unmated animal.
Prooestrus. During this stage the leucocytes disappear
altogether from the vaginal contents and the smear is made up
entirely of lightly staining nucleated epithelial cells. The
TYPES OF (ESTROUS CYCLE
49
vaginal contents are serous during this stage, which lasts some
twelve hours in the normal mouse.
Qistrus. Towards the end of procestrus the nucleated
epithelial cells begin to be mixed with the cornified mucosa cells
and the latter gradually supersede the former in the vaginal
contents. The cornified cells are typicall}^ non-nucleated and
eosinophil at this stage, w^hile the vaginal contents as a whole
have a granular appearance. Copulation occurs at this stage,
Fig. 23. — Vagina of Mouse during Dicestrus.
/.;;. leucocytes and nucleated epithelial cells in the lumen.
usually within the first day. As in the rat, the formation of the
vaginal plug from the vesicular secretion of the male provides a
reliable means of detecting copulation. The plug, surrounded
by an envelope of cornified vaginal epithelium, is shed in about
eighteen to twenty-four hours. In the unmated animal this stage
lasts about two days and is terminated by the appearance cf
leucocytes in the smear.
Metoestrus. During metoestrus the vagina returns to the
dioestrous condition. The smear consists of cornified cells with
a gradually increasing number of leucocytes. Finally, nucleated
epithelium appears, and the sm.ear becomes of the normal dioes-
trous type.
P.S.O. D
50 INTERNAL SECRETIONS OF THE OVARY
Pregnancy. During the whole of pregnancy the smear
consists of a few nucleated epithelial cells and leucocytes. Much
mucus is present all the time, and blood appears in both the rat
and mouse at about the twelfth day owing to uterine haemorr-
hage (the ' placental sign ' of Long and Evans).
c.e.
Fig. 24. — Vagina of Mouse during (Estrus, showing
cornification of the epithelium.
C.e. cornified epithelium sloughed off into the lumen.
Pseudo-pregnancy . Following sterile copulation, i.e. during
pseudo-pregnancy, the vaginal smear is of the dicestrous type,
except that much mucus is present. About three days before
the next oestrus is due a smear containing blood is usually
obtained.
In the rat the vaginal cycle is similar to that of the mouse.
Long and Evans (425) summarize the changes during the cycle
in the rat as in Table 3.
The nature of the leucocytes in the vaginal smear has been
studied by Post (518) and Guttmacher (254).
TYPES OF (ESTROUS CYCLE
51
fyiu
n.c. —
-^jr^:
M ..)
f5
»^^«-
iM;^,
(^)
.»
" <%
f^:
i-k
■» *1%
%>■' *
'1' fi :*>L
-c.e.
ii.e.
Tf ' •
&Jiftx-.. ''-<;xVtv
Fig. 25. — Vaginal Smears of the Mouse during the CEstrous Cycle.
(a) dioestrus, (6) late prooestrus, {c) oestrus, {d) late metoestrus, {e) pregnancy
c.e. cornified epithelial cells; /. leucocytes; n.e. nucleated epithelial cells.
52
INTERNAL SECRETIONS OF THE OVARY
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TYPES OF (ESTROUS CYCLE
53
Mammary Cycle. The mammary cycle in the unmated rat
and also the development of the gland during pregnancy have
been dealt with by Myers (472; and Sutter (605). The former
states: ' If pregnancy does not occur with the iirst ovulation the
mammary glands undergo slight retrogressive changes; how^-
ever, as the next ovulation approaches the mammary gland
takes on a new development. Similar changes can be observed
with each succeeding ovulation until pregnancy occurs.'
(/) RABBIT AND FERRET
The rabbit and ferret correspond in two peculiarities; oestrus
persists throughout the breeding season in the absence of
Fig. 26. — Uterus of Rabbit in CEstrus.
The stroma is glandular, but compact.
54
INTERNAL SECRETIONS OF THE OVARY
copulation, and, as originally shown by Heape (288) in the
rabbit, and Marshall (441) in the ferret, ovulation is dependent
upon copulation. Both animals have a well-developed pseudo-
pregnant period after sterile copulation, during which the ovary,
uterus, and mammary glands undergo growth similar to that
found during at least the first half of pregnancy.
.^
■'-.^■i.'i^^fV'r
Fig. 27. — Uterus of Pseudo-Pregnant Rabbit.
The stromal glands are greatly developed.
Rabbit. The histological and physiological changes occurring
during the fertile cycle of the rabbit have been extensively
studied by Ancel and Bouin (27, 29, 30-4, 75-7) and by Ham-
mond (264). The changes which take place are very marked in
the uterus, ovaries, and mammary glands. The vaginal cycle is
less clear, though changes have been described by Tsu (617) and
Courrier (139). Since oestrus is persistent, the only cycle is that
of anoestrus, oestrus, pregnancy, and lactation. If the animal
becomes pregnant at the beginning of the breeding season a
condition of cestrus will reappear immediately after parturition
and will continue during lactation. Pregnancy, however, does
not usually occur during lactation, as the blastocysts fail to
TYPES OF (ESTROUS CYCLE
55
become embedded and are reabsorbed. This sterility set up by
lactation in the rabbit is clearly a continuation of the conditions
found in the rat and mouse, where lactation causes a delay in the
implantation of the blastocysts, but not a permanent inhibition
Sterile copulation in the rabbit is followed by a condition of
pseudo-pregnancy, during which the changes in the ovary,
Fig. 28. — Photograph of Mammary Gland of
Pre-Pubertal Rabbit.
The gland is limited to a few ducts around the nipple.
(After Hammond).
the uterus, and the mammary glands are similar to those
occurring during the first half of pregnancy. Pseudo-preg-
nancy in the rabbit, though not so prolonged sls in Dasyurus or
the ferret, is nevertheless a far more distinct phase than the
pseudo-pregnant period in the mouse and the rat. The female
rabbit is atypical among lower mammals in allowing copula-
tion at other times than oestrus (264).
The ovarian cycle consists in the maturation of Graafian
follicles at the beginning of the breeding season and persistence
of mature follicles until copulation takes place, or until the
breeding season ends. In the absence of mating, atresia sets in
at the end of the breeding season. Ovulation takes place about
56
INTERNAL SECRETIONS OF THE OVARY
ten hours after copulation. The subsequent development of the
corpora lutea is continued for some fourteen days in the non-
pi egnant animal.
Fig. 29. — Photograph of .Mammary Gland of Rabbit in
FIRST QiSTRUS.
The ducts have developed out radially from the nipple.
The uterine cycle consists of growth and vascularization at the
beginning of the breeding season, and, following ovulation, in
certain pregnancy or psuedo-pregnancy changes. During
oestrus the uterine stroma, though hypersemic, is not highly
glandular and the epithelium is straight and continuous. After
ovulation, under the influence of the corpus luteum, immense
growth takes place in the glands, which supply a secretion
responsible for the initial nutrition of the embryos. During
TYPES OF (ESTROUS CYCLE
57
lactation the uterus becomes much atrophied, presumably owing
to the drain on the metabohsm. This atrophy during lactation
is in marked contrast to the conditions found in the rat and
mouse, in which the uterus is active during lactation owing to
the persistence of the corpora lutea.
Fig. 30. — Photograph of Mammary Gland of Rabbit
TWELVE DAYS PsEUDO-PrEGNANT.
Clumps of alveoli have developed along the ducts.
(After Hammond.)
The cyclic changes in the mammary glands of the rabbit are
also very obvious. Prior to the first oestrous period the gland
consists merely of a few ducts in the neighbourhood of the
nipple. At the time of the first oestrous period these ducts grow-
out radially for a distance of an inch or an inch and a half round
58
INTERNAL SECRETIONS OF THE OVARY
the nipple. At this stage, however, the development of the
gland stops until ovulation has taken place and luteal activity
begins. Under the influence of the corpus luteum the third
phase of mammary development, the appearance of bud-like
Fig. 31. — Photograph of Mammary Gland of Rabbit
AFTER Pseudo-Pregnancy.
The lobules of alveoli are undergoing atrophy, but complete return
to the virgin condition (Fig. 29) is not found.
Structures at the end of the ducts and the growth of nests of
alveoli, is started. In the absence of pregnancy no further
development than this occurs, and the degenerative phase,
which coincides with the atrophy of the corpora lutea at the end
of pseudo-pregnancy, results in the appearance of a small
TYPES OF (ESTROUS CYCLE
59
amount of milk in the ducts. During pregnancy the growth of
the mammary glands for the first fourteen days is similar to that
found during pseudo-pregnancy and is lateral and radial only.
Fig. 32. — Photograph of Mammary Gland of Rabbit
TWENTY-THREE DAYS PrEGNANT.
Great hypertrophy of the gland, characterized by secondary
thickening, has begun.
In the second half of pregnancy an entirely new phase of
mammary growth is initiated, i.e. a thickening of the gland,
during which the different nipple areas become confluent. The
tissue reaches a thickness of some •3--4 cms., and the glands
weigh about 100 gms. at the end of pregnancy.
6o
INTERNAL SECRETIONS OF THE OVARY
Ferret. Reproduction in the ferret has been studied by
Marshall (441) and Hammond and Marshall (267) . The breeding
season in the ferret lasts from about March to August, and
ovulation can take place at any time during this period, provided
that copulation occurs. Ripe follicles are always present in the
unmated female during the breeding season.
r.a
,_ c.t.
m.
Section of Lobule of Mammary Gland of
Rabbit during Pseudo-Pregnancy.
Fig. 33
c.t. connective tissue; m. muscle; r.a. rudimentary alveoli
The pseudo-pregnant period which follows sterile copulation
is remarkable in that it lasts six weeks, as long as true pregnancy.
In this characteristic the ferret is without parallel among known
Eutheria, except possibly the dog. CEstrus returns about eight
weeks after sterile copulation.
The ferret is also peculiar in showing a prodigious growth of
the vulva at oestrus. This swelling, which begins at the onset of
cestrus, reaches its maximum size in two to three weeks, and is
maintained during the whole of the breeding season in the
absence of copulation. The hypertrophy, which enlarges the
oestrous vulva to about fifty times the anoestrous size, dis-
appears at the beginning of pregnancy, pseudo-pregnancy, or
anoestrus.
The uterus also shows well marked development during
oestrus, but this is continued and increased after ovulation.
During anoestrus the uterus is very small and its glands com-
paratively undeveloped. At the beginning of cestrus the uterus
enlarges, and the mucosa and glands are rather better developed.
The uterus remains static in this condition during the whole
TYPES OF (ESTROUS CYCLE
6i
cEstrous period. In pseudo-pregnancy, development continues
up to the fifth or sixth week, after which degeneration takes
place.
In virgin ferrets, the mammary glands are entirely unde-
veloped both in ancestrus and oestrus. During pseudo-pregnancy
Fig. 34. — Section of Mammary Gland of twenty-nine
DAYS PREGNANT RaBBIT.
c.t. connective tissue; I. a. lobules of alveoli.
the ducts grow out round the nipple, and develop bud-like
terminations consisting of secretory alveoli.
{g) PRIMATES
The most obvious stage of the oestrous cycle in Primates is, of
course, the menstrual period, and most of the early work on the
Primate cycle was directed to analysing the significance of this
phenomenon. The correlation of the ovarian and uterine cycles
has been attempted systematically only during recent Vears.
Little success has been attained, and Heape's original observa-
62
INTERNAL SECRETIONS OF THE OVARY
tion on Semnopithecus entellus, that a regular menstrual cycle
may be maintained in the apparent absence of an ovarian cycle,
has been fully substantiated.
The uterine cycle. The uterine cycle in at least two species of
monkey and in the human has been fully worked out. The
writings of Heape on Seninopithecus (284) and Macacus rhesus
in.
Fig. 35. — Endometrium of human Uterus on the first day of
Menstruation, showing destruction of Epithelium and
superficial Stroma.
g. gland; /. lumen; ni. muscle; s. stroma.
(After Shaw).
(285), of Corner (123) and Allen (12, 14, 15, i6j on Macacus, of
Van Herwerden (297) on Cercocehus} and of Hitschmann and
Adler (307), Webster (632), Corner (125, 126), and Shaw (259)
on the human should be consulted for details and references.
The uterine cycles in monkeys and women appear to be
identical in all essential features, and Milnes Marshall's (456)
original division of the phases in the human, identical with
^ More probably Macacus cynomologos.
TYPES OF (ESTROUS CYCLE 63
Heape's division in Scninopithccits and Macaciis, distinguished
four main stages :
(a) period of rest.
{b) period of growth.
(c) period of destruction.
{d) period of regeneration.
Of these stages in woman the first is said by WilHams (638)
to last twelve days, the second (the stage of premenstrual
congestion) five days, the third (the actual period of menstrua-
tion) four days and the last seven days. According to other
workers, however, the initial premenstrual changes begin much
earlier than this, the period of quiescence being correspondingly
shorter.
Shaw states that the earliest signs of activity in the inter-
menstrual endometrium become apparent about fourteen days
after the beginning of the previous menstrual period. The
surface epithelium becomes more columnar and then hyper-
trophied and dilated. Typical translucent areas appear behind
the nuclei of the gland cells, which begin to secrete b}' the
twentieth day. By this time also, the general hypertrophy
has become much greater. After the twentieth day other
changes become evident. Three la^^ers can now be distinguished
in the stroma :
{a) A dense layer immediately below the surface epithelium;
the cells are tightly packed together round the gland
ducts in this area.
(b) An oedematous layer which surrounds the glands and in
which the cells are separated, but the capillaries dilated.
(c) A basal layer in which the characteristic oedema and
hypertrophy are absent.
These changes are accentuated until the 28th day when, if
pregnancy fails to supervene, degeneration sets in. This
consists essentially in the disorganization and disintegra-
tion of the glands of the superficial and middle zones of the
endometrium. Coincidently, blood is extravasated from the
capillaries and, having formied small hasmatomata, finally
reaches the lumen through the disintegrated epithelium. The
extent to which the endometrium is destroyed and shed varies
64 INTERNAL SECRETIONS OF THE OVARY
very greatly in different individuals. Subsequently, the
endometrium re-forms and returns to the resting stage.
The uterine changes in monkeys, as described by Heape,
Corner, and other writers, are essentially similar. Heape
divided the whole cycle into eight stages: i. resting stage, 2.
growth of stroma, 3. increase of vessels, 4. breakdown of vessels,
5. formation of lacunae, 6. rupture of lacunae, 7. formation of
menstrual clot, 8. recuperation stage. Corner gives the following
description of the menstruating uterus of Macacus: ' In general
the menstrual process consists of an extravasation of blood in
the more superficial part of the endometrium. This leads to the
necrosis and collapse of the stroma and glands in the affected
region, and here and there to the lifting away of small sheets of
epithelium by formation of haematomata under them. It
appears that most of the surface is denuded of epithelium; that
portions of the glands may be lost, and that the stroma is
necrosed and cast off to a depth of roughly one-fourth of the
entire thickness of the endometrium. ... As far as can be made
out in these specimens regeneration of the epithelium is by
growth from the glands and perhaps from the undisturbed islets
of epithelium. The result is a smooth healing of the wound-
surface with a single layer of very low epithelium.'
Information as to the time relations of the various phases of
the uterine cycle in monkeys does not seem to be available,
though the actual duration of menstruation (four to six days) is
similar to that of the human, as is the total length of cycle.
The ovarian cycle. The difficulties in the way of obtaining
suitable human material of known history, together with the
untrustworthiness of the methods employed, led many of the
earlier workers, such as Steinhaus (597), Jackson (314), Leopold
(360) and Aveling (52), to doubt the existence of a regular
ovarian cycle in man. This conclusion was supported by the
erratic behaviour of the ovaries of Heape's monkeys. More
recent work, however, and our general knowledge of mammalian
reproduction, makes it necessary to suppose that a regular
ovarian cycle exists in the normal human, as in lower mammals.
In monkeys, on the other hand, under conditions of captivity in
temperate climates, the ovarian cycle very often appears to be
in abeyance. In both cases information as to the details of the
TYPES OF (ESTROUS CYCLE 65
cycle is very meagre, the exact time of ovulation not being
known, even in the human.
A tubal ovum has not yet been recovered in man and this
method of determining the time of ovulation is not therefore
available. Certain data are provided, however, by the history
of the corpus luteum. The presence of newly-formed corpora
lutea in the human ovary at fifteen to seventeen days after the
beginning of the previous menstrual cycle led Meyer (459),
Schroder (548) and Novak and TeLinde (475), among others, to
conclude that ovulation takes place between these times, i.e.
in the middle of the intermenstrual interval. Shaw's material
enabled him to narrow the time down to between the thirteenth
and seventeenth days. By the nineteenth day of the cycle the
corpus luteum is mature, and it remains unaltered until the
twenty-seventh day of the cycle. By the twenty-eighth day
signs of degeneration, synchronizing with the onset of menstrua-
tion, have appeared, and eventually, some eight months later
(560), a corpus albicans is produced.
In Macaciis, ovulation (deduced from the recovery of tubal
ovaj has been found by Corner (123) to occur fourteen to fifteen
days, and by Allen (9) ten to fourteen days after the onset of
the previous menstruation. Apart from some notes by Allen on
the corpora lutea, little further appears to be known of the
ovarian cycle in monkeys.
Vaginal and mamma rv cycles. Both Allen and Corner have
dealt with the vaginal cycle in Macacus. Their work makes it
evident that, while slight changes occur in the vaginal epithelium
and in the proportions of leucocytes in the smear, no striking
cyclic changes occur such as are found in the mouse and rat.
In other monkeys, notably Papio sps., extraordinary swelling of
the vulva occurs during the follicular phase.
In the human, Papanicolaou (486) has claimed to be able to
detect all kinds of pathological conditions, as well as the normal
cyclic stages of the uterus and ovaries, by changes in the vaginal
smear. Dierks (160) has also described slight vaginal changes
during the menstrual cycle. King (322), however, obtained
negative results.
In monkeys, mammary changes do not appear to have been
described during the ordinary menstrual cycle, though Heape
P.S.O. E
66 INTERNAL SECRETIONS OF THE OVARY
refers to enlargement of the nipples concurrent with menstrua-
tion. In the human, mammary hypertrophy during the im-
mediate premenstrual phase is well known and has been
considered in detail by Dieckmann (159).
The interpretation of the Primate cycle. Much controversy still
exists as to the precise significance of the menstrual cycle in
Primates. There can be little doubt that it is strictly compar-
able with the cycle in lower mammals, though the changes are
greatly exaggerated at the uterine haemorrhage stage. Apart
from hypotheses which maintain that no correspondence exists
between the menstrual cycle and the cycle in lower mammals,
three different theories have been maintained at one time or
other:
(a) Most clinicians and embryologists have supposed that the
premenstrual growth of the uterine endometrium is
designed to facilitate the implantation of the embryo,
and is thus a postoestrous phase, corresponding to the
luteal or pseudo-pregnant phase of lower mammals.
Menstruation on this view is supposed to represent
solely the destruction of the prepared endometrium
when the fertilized ovum has failed to materialize : in
other words, to represent the abortion of an unwanted
decidua. There is much evidence in favour of this view.
Premenstrual congestion begins after ovulation, and
the breakdown of the endometrium coincides with
retrogressive changes in the corpora lutea. Degene-
ration of pseudo-pregnant congestion, sometimes ac-
companied by haemorrhage, has been described in the
rabbit by Hammond (264) and Ancel and Bouin (31),
in the dog by Gerlinger (241), in the sow by Corner
(122), and also occurs in the mouse.
Further evidence in support of this view is that
X-ray sterilization stops the menstrual cycle in the
human, whereas in lower mammals such treatment
causes the disappearance of the luteal phase, but not of
the follicular phase (see p. 177). Seitz and Wintz (557)
even state that X-irradiation during the first half of the
inter-menstrual period (i.e. early enough to stop the
formation of the corpus luteum) leads to immediate
TYPES OF CESTROUS CYCLE 67
amenorrhoea, while irradiation during the second half
{i.e. after the formation of the corpus luteum) does not
interfere with the next menstruation.
The essential feature of the interpretation of the
menstrual period as purely pseudo-pregnant degenera-
tion is that preliminary activity of the corpus luteum is
presupposed. This conflicts with the well-established
fact that some humans (123, 314, 360), and many
monkeys, have been known to menstruate without
previous ovulation. Unless in such conditions men-
struation is purely pathological (which its regularity
makes very doubtful), its occurrence in non-ovulating
animals is strong support of the second theory of
menstruation.
{b) From his studies on monkeys, Heape was led to suppose
that menstruation was analogous with the prooestrous
haemorrhage which takes place in such animals as the
dog, as described by Marshall and Jolly, and to a lesser
extent in the cow and sheep. A similar view was
initially held by Marshall. In general, the period of
cEstrus in mammals is characterized by destruction of
the epithelium of the genital tract, such as the cornifi-
cation of the vaginal epithelium in the rat, mouse, and
guinea-pig. This merely represents the disintegration
of a short prooestrous growth, and its identification with
menstruation (as attempted by Stockard and Papani-
colaou for the guinea-pig) is diflicult. Such a point of
view is strengthened, however, by the occurrence of
menstruation without ovulation, and by Allen's (12)
induction of menstruation in Macacus (a) by destruc-
tion of ripe follicles and (b) by injection into the
ovariectomized animal of the cestrus-producing
hormone.
The view that menstruation is prooestrous degenera-
tion is difficult to reconcile with the time relations of
the uterine and ovarian cvcles. It is generally agreed
that ovulation takes place about fourteen days after
the beginning of the previous menstruation. In the
lower mammals ovulation and oestrus are strictly
68
INTERNAL SECRETIONS OF THE OVARY
o
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TYPES OF (ESTROUS CYCLE 69
synchronized, and some extraordinary distortion must
have taken place in the human cycle if fourteen days
(i.e. half the total length of cycle) elapse between
prooestrus and ovulation.
(c) More recently, Marshall has suggested that menstrua-
tion represents both pseudo-pregnant and prooestrous
degeneration telescoped into one (448). In animals
such as the guinea-pig with a dioestrous cycle con-
taining a luteal phase, the end of the latter (pseudo-
pregnancy) is rapidly followed by prooestrus, and a
further contraction of the cycle at this point might
result in these stages becoming merged. This view of
Primate menstruation is satisfactory in explaining
many difficulties. It admits the essential pseudo-
pregnant nature of the premenstrual growth of the
uterus, and explains menstruation in non-ovulating
animals by the supposition that the part of menstrua-
tion occurring under such conditions is the prooestrous
stage only. Corner's (126) observation that menstrua-
tion without ovulation is not preceded by the typical
premenstrual growth of the endometrium is therefore
of great interest. Allen's (12) results can also be
explained on the grounds that the rupture of large
follicles and the cessation of injection of the oestrus-
producing hormone precipitate the destruction of
prooestrous growth.
Even this view, however, has the difficulty that
the destruction of prooestrous growth will have ended
some nine days before ovulation, and it does not
explain such abnormal phenomena as menorrhagia
associated with cystic follicles.
CHAPTER V
THE OVARY AS AN ORGAN OF INTERNAL
SECRETION
The synchronization found between development in the ovary
and activity in the accessory reproductive organs immedi-
ately suggests that one controls the other, and it has become
abundantly clear that the ovary is responsible for the growth
and cyclic changes of the uterus, vagina, and mammary
glands. Furthermore, the evidence shows that the nervous
system plays a very minor part in this correlation, which
is maintained by means of internal secretions. This view is
based primarily upon ovariectomy and transplantation experi-
ments, and in a lesser degree upon cases of abnormal sexuality.
Such evidence in itself is not quite complete, and the ultimate
proof (extraction of the active substances themselves from the
ovary) is only just being obtained after a long period of very
indifferent work.
The effects of ovariectomy and ovarian transplantation, and
the earlier experiments on the use of ovarian preparations,
are discussed below.
{a) EFFECTS OF OVARIECTOMY
Gonadectomy in both mammals and birds leads to the
development of a type which is intermediate between the two
sexes, and which may conveniently be regarded as approxi-
mating to the neutral embryonic condition. In birds this
neutral type tends to resemble superficially the male (the
homozygous sex).
In birds, ovariectomy still leaves intact the rudimentary right
gonad, which may subsequently develop into testis tissue. This
results in sex-reversal, which should not be confused with
70
OVARY AS ORGAN OF INTERNAL SECRETION 71
the effects of complete gonadectomy. The same probably
applies to the assumption of male characters which may
happen in mammals when the ovary is interfered with by
':i^>P>^\^^'\:^.
^.
o
V'^ri'h''''y^&
■:f.
W
M0y^'
Fig. 36. — Normal Uterus of Rat.
(From ^larshall and Jolly.)
malignant growth, or bacterial action, or at the menopause.
Complete gonadectomy of the female mammal does not lead to
the assumption of male secondary characters.
Effects on secondary characters. As indicated above, owing to
the negative nature of the female secondary sexual characters,
the superficial effects of ovariectomy are not as obvious as those
72 INTERNAL SECRETIONS OF THE OVARY
of castration. Certain effects, however, unconnected with the
accessory organs, do follow ovariectomy, particularly of the
immature female. These include:
(a) The subcutaneous fat which is laid down at puberty in
the normal female fails to appear, and, subsequently,
adipose tissue is laid down indiscriminately.
(b) The long bones undergo increased growth owing to the
non-ossification of the epiphyses.
(r) In correlation with the increased growth of the skeleton,
a greater wxight is attained. Experimental work on
this has been performed by Wang, Richter, and Gutt-
macher (630).
^d) The sex instinct is suppressed, together w4th such sex
reflexes as the ' tail erection ' of the female rodent
before copulation. With the suppression of the sex
instinct comes a general state of lethargy. Advantage
is taken of this fact in the spaying of farm stock before
fattening for market.
{e) According to Loewy and Richter (424) the basal meta-
bolism falls after ovariectomy. Blair Bell (56) reports
decrease in calcium excretion, increase in phosphorus
excretion, and no change in nitrogen metabolism after
ovariectomy.
(/) Much confusion exists as to the effect of ovariectomy on
the other ductless glands. Biach and Hulles (64) state
that ovariectomy of the young animal causes atrophy
of the pineal. This has not been confirmed. Hammond
(262) and Blair Bell (56) found hypertrophy and
hyperactivity of the pituitary after ovariectomy.
Considerable hypertrophy of the thyroid is known to
occur during various phases of the sexual cycle, and
Blair Bell reports hyperactivity after ovariectomy.
According to the same author, an increase in the zona
reticularis of the adrenal is found. According to
Masui and Tamura (457), Miller (460), and Deanesly
(154), sex dimorphism is found in the adrenal cortex
of the mouse. Castration causes changes in the male
cortex which obliterate the sex difference, but ovariec-
tomy produces no effect on the female gland.
OVARY AS ORGAN OF INTERNAL SECRETION 73
It will be noted that man\' of these effects of ovariectomy
are general results of de-sexing the animal and occur after
castration of the male. Similar effects are also found in
chronic insufficiency brought on by delayed development or
hypo-function of the gonad.
Effects on the accessory organs. Ovariectomy causes profound
changes in the accessory organs of reproduction, the effect being
more striking when the ovaries are removed before puberty.
The operation at this time results in complete failure of the
organs to undergo their normal development, and in non-
FiG. 37. — Uterus of Rat after Ovariectomy, showing
DEGENERATIVE CHANGES.
(From Marshall and Jolly.)
appearance of the cyclic changes of cestrus. Post-pubertal
ovariectomy leads immediately (but see p. 137) to the stoppage
of all cyclic activity in the uterus, vagina, and mammary
glands, but the actual degeneration of the organs is slow, and
the degree of atrophy found at autopsy is usually proportional
to the time after operation. The general effects on the acces-
sory organs are suggestive of the changes occurring at the
menopause, and ovariectomy is often spoken of clinically as
producing an ' artificial menopause.'
Carmichael and Marshall (114) have reported extensive
experiments on the adult rabbit. Six months after ovariectomy
the uterine stroma had entirel}^ degenerated, the glands,
mucosa, and muscle layers having undergone atrophy. De-
generation had also extended to the Fallopian tubes. Similar
74 INTERNAL SECRETIONS OF THE OVARY
results were obtained by Bucura (109) on the rabbit, by
Marshall and Jolly (452, 453) on rats, by Tandler and Keller
(609) on cows, and by Halban (260) on guinea-pigs. This last
author also states that the mammary gland fails to develop
after pre-pubertal ovariectomy.
Ovariectomy in the human leads to the degeneration of
uterus, vagina, and mammary glands, or, if the operation is
pre-pubertal, to the non-development of these organs. In the
™ Ob **.*•
Fig. 38. — Uterus of Mouse after Pre-Pubertal Ovariectomy,
SHOWING rudimentary CONDITION.
case of the mammary glands, however, it is necessary to make a
qualification; there is some evidence that if the gland is secreting
at the time of the operation, the secretion may be prolonged.
In the mouse (508) the effects of pre-pubertal ovariectomy
(at three weeks old) are remarkable. Two months after the
operation the uterus is not only undeveloped, but has actu-
ally degenerated from the pre-pubertal state, being only about
•I mm. in diameter. Microscopically it consists of a ' stroma,'
three to four cells thick, surrounded by a few layers of muscle
fibres. The changes in the vagina are equally definite. Though
about the same size as the pre-pubertal organ, the vagina
after ovariectomy is collapsed, the component layers are
atrophic, and the lumen is blocked by debris. No effects on
OVARY AS ORGAN OF INTERNAL SECRETION 75
the clitoris are produced by ovariectomy of the immature
mouse, a fact which emphasizes the vestigial nature of the
clitoris in this animal.
These effects of ovariectomy are now almost universally
ascribed to the removal of ovarian influence. Early workers,
such as Hofmeier (308), were inclined to attribute the effect to
■^ > > > > y
Liquor foUiciiU
Maximum purity
0-13 (r.u.)
Doisy, Ralls, Allen, and
Johnston (167)
Minimum purity
19-5 (r.u.)
> > > > > >
Unstated
•00002 (m.u.)
Lipschiitz et al {^yz)
Cow
I2-I-2-6I (m.u.)
Parkes and Beller% (503)
Pig
2I-5-I4-2 (m.u.)
) > y > ) >
Horse
9-4 (m-u.)
I » ) ) ) )
Pig
•0I--02 (r.u.)
Ralls, Jordan, and Doisy
(524)
Residual ovariaji
tissue
Cow
I6-9-9-0 (m.u.)
Parkes and Bellerby (503)
Pig
35-0-IO-5 (m.u.)
J > >> > >
Horse
19-6 (m.u.)
> > >> > 1
Placenta
Human
25-0 (r.u.)
Doisy, Ralls, Allen, and
Johnston (167)
> f
277-3-83 (m.u.)
Parkes and Bellerby (506)
Cow
I9-2-I-9 (m.u.)
y y J ) ) >
Sheep
2I7-6-25 (m.u.)
> > ) ) > >
Unstated
•02 (m.u.)
Steinach and co-workers
(593)
Human
•I--OI (m.u.)
Laqueur and co-workers
)t
•006 (m.u.)
Glimm and Wadehn (246)
})
•0008 (m.u.)
Laqueur, Hart, and de
Jongh (355)
>>
•003 (r.u.)
> > > > ) >
>)
•010-26 (r.u.)
Allan, Dickens, Dodds,
and Howitt (5)
io6 INTERNAL SECRETIONS OF THE OVARY
Table 6. — Weight of the Unit of CEstrin.
(continued).
Weight of mouse
Source of Extract
unit or rat unit
Authority
(Mgms.)
Female urine
Pregnancy
•001 (m.u.)
Zondek (648)
y >
•22--o6 (r.u.)
V'eler and Doisy (621)
> >
•0016 (m.u.)
Marrian and Parkes (438)
Plant sources
Yeast
•I (m.u.)
Glimm and Wadehn (247)
Unstated
lo-o (r.u. or m.u.)
Coward and Burn (146)
2-25
J
Frank and Gustavson
(229)
found in the amounts required. Pratt and Allen (519) indeed
claim to have produced clinical effects with very small doses.
The purity of an extract may best be stated in terms of the
weight of the unit. The weights given by various authors are
collected in Table 6. Such figures, of course, can be only ap-
proximate. The variety of techniques used for assay, and
unavoidable inaccuracies, make strict comparison impossible.
(/) DISTRIBUTION
Extracts were originally made only from animal tissues,
but recently the presence of the oestrus-producing hormone has
been detected in body fluids. In addition, oestrin or oestrin-hke
substances have been obtained from various plant sources.
Distribution m the ovary. The ovarian follicle was originally
used for extraction of the hormone and has long been considered
its essential site of origin. Allen and his co-workers (24) by
the use of centrifuged and filtered material showed that oestrin
is actually present in the liquor folliculi and not merely in the
cellular contents which are aspirated at the same time. Zondek
and Aschheim (650) claim to have shown by their implantation
method (see p. 91) that the hormone is present in the theca
THE GESTRUS-PRODUCING HORMONE 107
interna cells, especially in atretic follicles, and absent in the
follicular granulosa, the ovarian stroma, and the germinal
epithelium. The hormone has, however, been shown to occur
in residual ovarian tissue, after removal of large follicles and
corpora lutea, in much larger amounts than could be accounted
for by the remaining follicles (158, 503). Various workers have
noted oestrin in ovarian cysts (24, 488). Much controversy has
raged around the alleged presence of oestrin in the corpus
luteum. Early workers were almost unanimous in supposing
that oestrin could be obtained from the corpus luteum.
Thus, Iscovesco (313), Seitz, Wintz and Fingerhut (558),
Fellner (192), and Herrmann (292) all obtained active extracts
from this source. Some of these workers used growth of the
rabbit uterus as a test-object, so that the possibility of other
than oestrus-producing substances giving the reaction is not
excluded. Their illustrations, however, give no suggestion of
any reaction except the oestrous condition. Okinschitz (481)
failed to obtain positive results from corpus luteum. More
recently, positive results have been obtained by Frank and
Gustavson (229), Fraenkel and Fonda (218), by Glimm and
Wadehn (245), and Zondek and Aschheim (654). Allen and
Doisy failed to obtain positive results with corpora lutea from
the cow and pig, except in one case of tertiary liquor folliculi,
while Johnston and Gould (316) failed with pig corpora lutea.
Brouha and Simonnet (93) also obtained negative results from
cow corpora lutea. Allen and Doisy did, however, find a slight
activity in the hollow human corpora lutea during the early
stages of development. The discrepancy in these observa-
tions is probably due to the fact that, in the cow particularly,
many corpora lutea contain a fluid centre derived from the
remains of the liquor folliculi. This fluid might reasonably be
supposed to contain oestrin. It is unlikely that the corpus
luteum itself, an organ whose development is always asso-
ciated with the absence of oestrus, would produce the
oestrous hormone. Parkes and Bellerby (507) found that in
the cow's corpus luteum the fluid enclosed in the centre of
hollow specimens contained oestrin, but that the tissue of
solid corpora lutea possessed no activity whatever. If, as this
work suggests, the oestrin content of the corpus luteum is
io8 INTERNAL SECRETIONS OF THE OVARY
restricted to the fluid centre, it seems probable that the
corpus luteum tissue does not elaborate the hormone. As
regards the ovary, therefore, it may be said that the liquor
folliculi of all mammals which have been examined contains this
hormone, as does the stromal tissue, but that the corpus luteum
contains it incidentally or doubtfully.
Distribution in other animal tissues. It has been known for
a long time that the placenta contains large amounts of
oestrin. Eels (202) failed to find it before the second month
of pregnancy, but Allen, Pratt, and Doisy (25) found it in
placentae of the third month, and Eellner (191, 192), Herrmann
(292), Eraenkel and Fonda (218), Zondek and Brahn (661),
Frank and co-workers (229, 231), Allen, Doisy and co-
workers (23, 25, 167), Hartmann and Isler (281), Suzuki
(606), Zondek and Aschheim (651), Parkes and Bellerby (506),
Ceresoh (117), and Dickens, Dodds, and Wright (158) have
all found it at later stages. (Estrin has also been found in
the foetal membranes (25), amniotic fluid (506, 132, 93, 203),
and umbihcal cord (25), but extracts of the foetus itself have
given uniformly negative results (25, 167, 506).
Various other body tissues have been examined either with a
view to ascertaining the distribution of oestrin in the body or
with a view to providing control experiments on the extrac-
tion of ovaries. Negative results have been obtained from
liver (230), brain (230), thymus (230), pancreas (167),
pituitary (538), adrenal (654), thyroid (651), spleen (654), and
muscle (503).
Distribution in body fluids. Since it is obvious that the
hormone must pass from its site of origin to its site of activity
by way of the circulating blood, one would expect to be able to
detect its presence in the blood provided that sufftciently small
amounts could be recognized. Also, since its action is cyclic, it
might be expected that periodic variation in amount in the blood
could be demonstrated. Its presence in the blood of the non-
pregnant female has been shown by Loewe (408), Frank and
co-workers (225, 227), and Smith (574). Frank and co-workers
(223) found oestrin in the blood of the oestrous sow, and in the
human in amounts varying with the stage of the cycle.
According to these workers, it is present in greatest amount
THE (ESTRUS-PRODUCIXG HORMONE 109
about the first day of menstruation, after wliich it rapidly
decreases. It is present in menstrual blood in larger amounts
than in the circulating blood. Frank also uses the blood oestrin
test to determine intersexual conditions. Eels (202) and
Trivino (615) found oestrin in the circulation only during
pregnancy. Aschheim (36) reports negative results from pre-
menstrual blood, inter-menstrual blood, and blood during
labour.
During pregnancy, however, very considerable amounts of
oestrin are present in human blood, as shown by Zondek and
Aschheim (657), Eels (202), Trivino (615) and Smith (574). Eels
describes it as increasing rapidly in the circulating blood after
the sixth month of pregnancy, while Aschheim (36) found it
only after the fourth month. It is then so plentiful that 2 c.c.
of blood serum will give a positive reaction.
The detection of oestrin in blood soon led to an examina-
tion of the possibility of its excretion. The presence of
oestrin in small amounts has been described in the urine
of the non-pregnant female (413), varying according to stages
of the menstrual cycle, but during pregnancy relatively
enormous amounts have been found by various workers,
including Aschheim and Zondek (38), Slotta (570), Zondek (648),
and Veler and Doisy (621) . Aschheim and Zondek (40) state that
it is doubtfully present during the first two months of pregnancy,
and as small amounts are found in the urine of the non-pregnant
female, this is not essentially a characteristic of pregnancy.
They described (38) the concentration in the urine of late
pregnancy as 1,000 m.u. per litre. Subsequently, the same
authors (40) gave the yield as 4,000-10,000 m.u. per litre
(see p. 168). The human female thus seems to excrete about
a million mouse units during pregnancy. The comparatively
rapid disappearance of oestrin from the urine after parturition
has been noted by Aschheim and Zondek (see p. 167) and
Veler and Doisy (621). The latter give their results as in
Table 7 (p. no).
Occurrence in the male. The presence of oestrin in the testis
and also in the male body fluids has been reported by various
authors. Eellner (195), Laqueur and de Jongh (357). Brouha
and Simonnet (106) and Robinson and Zondek (538) have all
no INTERNAL SECRETIONS OF THE OVARY
claimed to have secured active extracts from the testis, but
Allen and his co-workers (21) failed to obtain this result.
Hirsch (301) and Frank and Goldberger (228) obtained a
number of positive results from male blood.
Table 7. — CEstrin in Post-Partum Urine.
Time Post-Partum
R.u. per
(hrs.)
Litre
8-5
640
37
500
48
80
72
22
96
20
M4
<5
192
<5
264
<4
Several authors (162, 348, 422, 437) have reported the
preparation of oestrin from male urine. Loewe (422) even
states that oestrin can be fractionated from the male sex
hormone in the urine. It is certain that small amounts (1-3
m.u. per litre) of a substance which will cause cornification of the
vagina of the rat and mouse can be obtained from male urine,
but in view of recent doubts of the absolute specificity of this
test, it is necessary that every possible criterion of oestrus should
be used before the substance is definitely identified as oestrin
(437)-
Distribution in animals other than mammals. Little is known
about the non-mammalian distribution of this hormone. Fellner
(198) claims that it may be detected in the eggs of hens and fish,
but Doisy and co-workers (167) were unable to confirm this.
Occurrence in plants. Various workers have claimed that an
oestrous reaction may be produced by the use of extracts of
various plants. Thus, Loewe, Lange, and Spohr (418) say that
an extract of willow catkins gives a positive reaction, and that
as much as 200 mouse units may be obtained from a kilogram of
fresh material. The stigmata of willow blooms contain a
small amount «I4 m.u. per kg.), while the stalks and flowers of
THE (ESTRUS-PRODUCING HORMONE
III
Table 8. — Yields of (Estrin from various Sources.
Yield per kg.
Source
or litre (mouse
units or rat units)
Authority
Whole Ovaries
Cow
293 m.u.
Parkes and Bellerby (503)
,, (immature)
73-350 m.u.
> ) > > > >
Pig
219 m.u.
> > y ) J y
,, (immature)
166-273 m.u.
> J > ) y >
Pig
120 (r.u.)
Doisy, Ralls, Allen, and
Johnston (167)
Sheep (anoestrous)
203 (m.u.)
Parkes and Bellerby (503)
Liquor Follicidi
Human
433-7000 (r-u.)
Allen, Pratt, and Doisy
1 7Z\
Cow
37-788 (m.u.)
Parkes and Bellerby (503)
Horse
113 (m.u.)
> > >> > J
Pig
23-75 (m.u.)
> > > y ) >
, , (average)
878 (r.u.)
Ralls, Jordan, and Doisy
(524)
>>
600-1600 (m.u.)
Laqueur, Hart, de Jongh,
and Wijsenbeek (356)
>i
167 (r.u.)
Dickens, Dodds, and
Wright (158)
Residual ovarian
tissue
Pig
225 (r.u.)
Dickens, Dodds, and
Wright (158)
Cow
150-326 (m.u.)
Parkes and Bellerb}' (503)
Pig
227-865 (m.u.)
> > y y > >
Horse
27 (m.u.)
->> >» »>
Corpora lutea
Human
3700 (r.u.)
Allen, Pratt, and Doisy
(25)
Pig
>8 <25 (r.u.)
Allen and Doisy (21)
Cow (unsorted)
16 (m.u.)
Parkes and Bellerby (507)
,, (solid)
no yield
y y >> > J
(tissue of
hollow)
II (m.u.)
y y y y >>
,, (fluid of
hollow)
184 (m.u.)
y > > > > >
112 INTERNAL SECRETIONS OF THE OVARY
Table 8. — Yields of Q^strin from various Sources.
(continued) .
Yield per kg.
Source
or litre (mouse
units or rat units)
Authority
Placenta
Human
192-2123 (m.u.)
Parkes and Bellerby (506)
> >
400-700 (r.u.)
Doisy, Ralls, Allen, and
Johnston (167)
Cow
Parkes and Bellerby (506)
(maternal)
203-3200 (m.u.)
y > } > > >
,, (foetal)
143-782 (m.u.)
y > > y ) )
Sheep
183-308 (m.u.)
> > ) > >>
Human
1500 (m.u.)
Aschheim (36)
} }
52-270 (r.u.)
Allan, Dickens, Dodds,
and Howitt (5)
Female blood
Late pregnancy
1000 (m.u.)
Aschheim (36)
Female Urine
Pregnancy
1000 (m.u.)
Aschheim and Zondek
(38)
> }
4-io,ooo(m.u.)
,,(40)
> )
470-1240 (r.u.)
Veler and Doisy (621)
Plant Sources
Willow catkins
200 (m.u.)
Loewe (418)
Press Yeast
30 (m.u.)
Glimm and Wadehn (247)
Brewers' Yeast
50 (m.u.)
) ) ) y } >
the water lily, though giving a definitely positive reaction,
have only about i m.u. per kg. According to Glimm and
Wadehn (247), and to Dohrn, Poll, and Blotevogel (163) oestrin
is present in appreciable quantities in yeast and potatoes. It
should be emphasized again, however, that the production of
oestrin from such anomalous sources must be confirmed in
every possible manner before being fully accepted, especially
as the prooestrous smear is considered to indicate a positive
result by various workers, including Loewe.
THE (ESTRUS-PRODUCING HORMONE 113
(g) SITE OF ORIGIN
The coincidence of follicular maturation with the occurrence
of oestrus, together with the fact that the initial extractions
were made from follicular fluid, led to the supposition that
the oestrus-producing hormone was essentially elaborated by the
mature follicle. More recently, however, the oestrous hormone
has been found in situations where it cannot possibly have been
elaborated, so that its occurrence in any particular site is not
evidence of its origin there. At the time of the first oestrous period
no corpora lutea are present in the ovary, and these structures
cannot, therefore, be considered as the essential site of origin.
In the same way the placenta, also an abundant source of the
hormone, is clearly not the essential site of origin. Further, it
has been shown that elimination of the Graafian follicles by
exposure to X-rays does not prohibit the formation of the
oestrus-producing hormone, and it would seem, therefore, that
under certain conditions, if not normally, the stromal tissue of
the ovary can elaborate the hormone.
Any hypothesis as to the site of origin of the hormone is
necessarily influenced by the view^ taken as to the exact nature of
its function (see Chapter VH).
{h) PHARMACOLOGICAL PROPERTIES
The pharmacological properties, like the chemical pro-
perties, can only be arrived at from negative results ; any
positive effect may be caused by impurities.
Effect on circulation. According to Allen and co-workers (24),
Fraenkel (212), Laqueur (346), Doisy, Ralls, and Jordan (166)
and Brouha and Simonnet (93), the effect of oestrin on blood
pressure is negligible, though crude extracts have a depressor
effect. The heart rate is unaltered. Laqueur (346) found no
action on the isolated frog heart.
Respivation. Laqueur (346) found no effect on the respiration
of narcotized cats, and later (357) no effect after injection
of 2,000 m.u. into the intact dog.
Metabolism. Laqueur, Hart, and de Jongh (346) found no
effect on the blood sugar, but a slight increase in the basal
P.S.O. H
114 INTERNAL SECRETIONS OF THE OVARY
metabolism of ovariectomized females following injection of
oestrin. No effect was observed in castrated males. Fraenkel
(212) failed to raise the total metabolism of women by similar
treatment. Dickens, Dodds, and Wright (158) have reported an
antagonistic action of fat-soluble oestrin to the hypoglycaemic
effect of insulin.
Effect on groivth and activity. Bugbee and Simond (112) state
that the injection of oestrin causes a dechne in weight of normal
and gonadectomized animals of both sexes. Brouha and
Simonnet (90) and Allen and Doisy (24), however, found no
effect on the growth of the immature animal.
Slonaker (566) and Wang, Richter and Guttemacher (630)
have shown that the spontaneous activity of rats undergoes
cyclic variation in correlation with the cestrous cycle, the
maximum activity occurring at oestrus. This increased activity
stops after ovariectomy, but can be brought about in the
ovariectomized animal by the injection of oestrin (m). Con-
tinuous injection leads to continuous increased activity.
CHAPTER VII
THE FUNCTION OF (ESTRIN
{a) ACTION OX TEST ANIMALS
Ik test animals, the effect of injection of oestrin in adequate
amounts is to produce all the extra-ovarian histological and
ph3^siological symptoms of the normal oestrous period, except
possibly the mating instinct. The uterus of the ovariectomized
rabbit undergoes hypertrophy after injection, until a condition
similar to that of the normal oestrous uterus is attained. In the
immature uterus this involves a very considerable increase in
size. Injection of the ovariectomized mouse, rat, and guinea-pig
with oestrin leads to growth of the vaginal mucosa, which ends in
the keratinization and sloughing off of the surface layers. In
the uterus, typical cestrous changes are also produced, including,
in the mouse and rat, the characteristic distension of the lumen
and attenuation of the uterine wall. The reactions of these
animals have been observed by a large number of workers and
are beyond dispute. The administration of excessive doses leads
to an exaggeration of the oestrous symptoms analogous with
that found in nymphomania. Other animals in which the
action of oestrin has been tested after ovariectomy include the
opossum and Macacus rhesus.
In view of these reactions of test animals there can be no
doubt whatever that oestrin is responsible for the extra-ovarian
changes typical of prooestrus and oestrus. The actual
mechanism of its growth-promoting action in the organs is not
known.
(b) ACTION ON NORMAL ANIMALS
The injection of oestrin into the normal animal just before
oestrus is due, leads to an intensification of the normal changes
in the accessory organs without alteration of the ovarian cycle
115
ii6 INTERNAL SECRETIONS OF THE OVARY
(but see 436). During the time when oestrus is normally in
abeyance (except in the very young animal), the injection of the
intact animal with adequate amounts of oestrin results in the
appearance of extra-ovarian oestrus At certain times, how-
ever, notably when functional corpora lutea are present in
the ovary, considerable dosage is required to bring on this effect.
No ovarian changes characteristic of oestrus occur during such
an induced period (see p. 119 for non-effect of oestrin on follicular
maturation).
. Prenatal period. Since the administration of large amounts of
oestrin during pregnancy leads to reabsorption or abortion, it-
is not possible to subject the foetuses to an excessive additional
oestrous stimulus, even if it is assumed that oestrin will cross the
placenta. Working with amounts insufficient to produce
abortion, Courrier (133) claimed to find growth effects on the
foetal uteri after injection of the pregnant guinea-pig. Allen,
Francis, and Craig (23), and Parkes and Bellerby (504) were
unable to confirm these results on the rat and mouse, while
negative results have also been obtained by Loeb (404) on the
guinea-pig. Eels (202) claims to have found oestrin in foetal
blood. In view of the large amounts of oestrin in the circulating
blood of the pregnant female (see p. 109), the passage of oestrin
across the placenta would mean that all foetuses, including the
males, would develop under the influence of this hormone.
Courrier 's result is therefore improbable. Parkes and Bellerby
(504), in fact, found that the uterus of the new-born mouse did
not react to oestrin.
The pre-pubertal animal. The effect of oestrin injection on the
normal pre-pubertal animal has already been indicated in
discussing the use of such animals for testing (see p. 93 and
also p. 97). In the rat and mouse rapid proliferation of the
vaginal epithelium is produced and the vaginal orifice appears.
The uterus shows the typical distension, with change of the
undeveloped epithelium to a low columnar type. According to
Brouha and Simonnet (90) the mammary gland shows no
change. Since the same oestrous reaction is found in the ovariec-
tomized immature animal, it is clearly independent of any
primary effect upon the existing ovary.
Loeb (405) has produced similar oestrous changes by injection
THE FUNCTION OF (ESTRIN
117
of oestrin into the immature guinea-pig, while Laqueur and de
Jongh (357) have caused growth in the uterus of the immature
dog..
The ancestrons animal. Asdell and Marshall (45) report the
induction of early procestrous changes in the anoestrous dog by
oestrin injection. Courrier (134) induced oestrous conditions in
the hibernating h'cdgehog by the same means.
Dioestrus
Normal oestrus
Induced oestrus following'
injection of oestrin
Death
lU Jays horizontal
scale
Fig. 44. — Diagram showing the Growth Curves and CEstrous
Cycle Histories of Rats injected with CEstrin during
Vitamin B deficiency Ancestrus.
Where ancestrus has been induced in the normal animal by
dietary deficiency, it is also possible to cause oestrous changes in
the accessory organs by injection of oestrin. Thus Parkes (497)
was able to bring about such changes during the anoestrous
periods caused by vitamin B deficiency. No ovarian stimulation
occurred.
The dicestrotis or pseudo-pregnant animal. The dioestrous
interval of the rat and mouse is too short for accurate experi-
mental work to be done during this phase, but the dioestrous
phase in the accessory organs can be almost eliminated by
ii8 INTERNAL SECRETIONS OF THE OVARY
continued cestrin injection (see p. 126). Both Mahnert and
Siegmund (436) and Brouha and Simonnet (91) found that
continued injection of the normal animal leads to long periods of
cornification separated by short dioestrous intervals. Since the
ovarian cycle is disturbed only slightly, if at all, and since
persistent cornification is easily produced in the normal im-
mature animal, this result may be due to the cyclic development
of the corpus luteum. There can be little doubt that the
continuous injection of large doses would lead to persistent
cornification in the normal animal.
The pregnant animal. Since oestrus is invariably absent
during pregnancy, the artificial induction of oestrus during
gestation would seem to promise results of interest. Striking
effects have actually been obtained. Allen, Francis, and Craig
(23) obtained a positive oestrus smear during early pregnancy
as the result of injection of oestrin, but Brouha and Simonnet
(91) failed to do so in the later stages. Smith (573) showed that
pregnancy in the rat could be interrupted in its early stages by
the injection of the oestrus-producing hormone, while Parkes
and Bellerby (504) found that pregnancy in the mouse could be
terminated at all stages by the administration of an adequate
dose. The amount required during the later stages was twice as
great as in the early stages. Loeb and Kountz (405) failed to
interrupt pregnancy in the guinea-pig by similar means, but this
was probably due to inadequate dosage. Fraenkel (212) also
failed to produce the effect in the rabbit. Zondek and Aschheim
(659) produced abortion in the mouse even in the last half of
pregnancy by using large doses (10 m.u.), and Eels (205) reports
the same result. The facts seem to be well authenticated as re-
gards rats and mice. The effect may be brought about by either
or both of two actions. In the earlystages of pregnancy the effort
of the uterus to assume an oestrous condition may result in the
failure of the embryos to become implanted, or, on the other
hand, the injection of cestrin, by overriding the action of the
persistent corpora lutea, may bring about a state analogous with
that produced by removal of the corpora lutea during pregnancy.
Abortion has also been reported by Engle and Mermod (179)
as a result of the artificial production of oestrus during pregnancy
by the injection of the oestrus-stimulating preparations of the
THE FUNCTION OF (ESTRIN 119
anterior pituitary body. Zondek and Aschheim (659), however,
produced ovulation but not abortion by the same means. The
bulk of the evidence suggests most strongly that oestrus and
pregnancy are incompatible, and that the artificial production of
an oestrous phase during pregnancy will result in its termination.
The lactating animal. During lactation, when oestrus is in
abeyance in the rat, mouse, and guinea-pig (except for the
immediate post-partum period), injection of oestrin will result
in the appearance of oestrous symptoms in the accessory organs
(23, 505, 565). Parkes and Bellerby (505) came to the following
conclusions:
[a) The amount of cestrin required to produce oestrous
changes is roughly proportional to the number of
young suckling. It is well known that if only one or
two young are suckling, spontaneous oestrus will appear
during lactation. With three or four suckling several
m.u. of oestrin are required, while with seven suckling
not less than 10-12 units will produce oestrus. This is
probably a corpus luteum effect (see p. 183).
(h) The induction of oestrus does not materially affect the
efficiency of lactation, but a slight break in the growth
curve of the young was found following injection of the
mother.
The senile animal. Slonaker (568), Steinach, Heinlein, and
Wiesner (594) and Laqueur, Hart, and de Jongh (345) report the
production of oestrous changes in the senile anoestrous mouse by
the injection of oestrin. The last authors obtained the result
with 2 m.u. Later, Steinach, Kun, and Hohlweg (596) report the
complete rejuvenation of the senile mouse by oestrin, including
the recommencement of follicular maturation and ovulation.
Effect of oestrin on follicular maturation. Injection of the
immature animal has been described by Loeb (405) and Truffi
(616) as stimulating the development of the follicles, without,
however, causing ovulation. Frank, Kingery, and Gustavson
(224) originally claimed to have caused ovulation in the im-
mature rat, but their results were probably due to the use of
animals approaching the first oestrous period. All other
workers appear to agree that oestrin injection does not cause
120 INTERNAL SECRETIONS OF THE OVARY
ovulation when it would not otherwise occur, either in the
immature or the adult animal. Allen and Doisy (19) , Brouha and
Simonnet (90), and Zondek and Aschheim (652) have all
arrived at this conclusion. Parkes and Bellerby (504-5) found
that copulation at an oestrous period induced either during preg-
nancy or lactation never resulted in conception, and thus con-
cluded that ovulation did not occur.
Similar results have been obtained by Slonaker (568), and
Steinach, Heinlein,and Wiesner (594) on the senile mouse. After
the menopause, when the cycle has stopped, injection of oestrin
will bring about cornification of the vagina without the cor-
responding ovarian changes. Exhaustion of oocytes may have
been responsible for the absence of ovulation under these con-
ditions.
According to Mahnert and Siegmund (436) injection of cestrin
into the normal adult tends to retard ovulation.
Clinical results. Owing to the difficulty of administering
adequate doses of the oily extracts to women, clinical
research with oestrin has not progressed far. With the new
water-soluble preparations it should be possible to make a great
advance in this direction. Even so, however, it is far from
obvious what part oestrin may be able to play in the correction
of reproductive disorders in the human. The premenstrual
growth of the uterus is an effect of the corpus luteum, and at
least a part of the ensuing menstruation is pseudo-pregnant
degeneration (see p. 66). Fraenkel (213-4) in particular has
emphasized the improbability that oestrin plays a dominant
part in the human cycle. Nevertheless, a variety of positive
results have been reported. Wintz (640) and Seitz, Wintz, and
Fingerhut (558) record a number of clinical tests of corpus
luteum preparations — ' lipamin' and ' luteolipoid,' some of
which appeared to give positive results, but it is very difficult to
assess this early work. The same applies to the clinical work
of Herrmann (292).
Using definitely oestrus-producing preparations, Pratt and
Allen (519) claim to have produced enlargement of the human
uterus in cases of both primary and operative amenorrhoea.
Menstruation was not produced. Zondek (646) obtained more
satisfactory results, while Brouha and Simonnet (104) claim to
THE FUNCTION OF (ESTRIN 121
have successfully treated ovariectomy atrophy, amenorrhoea,
sterility, dysmenorrhoea, infantile conditions of the genitalia,
and menopause symptoms. Laqueur (357) reports some
success in treating amenorrhoea.
It is clear that in considering all such clinical work, due
attention must be paid to the difficulty of adequately controlhng
work'on the human, and also to the psychological factor involved.
Little can be deduced from the results at present, but it seems
probable that oestrin will have some clinical use, especially in
expediting labour (see p. 205).
Murphey and his co-workers (471) have reported some trials of
the value of oestrin in veterinary practice.
Action on the male. Various authors have investigated the
effects of oestrin on the male, particularly upon the testis.
Adverse effects were reported by Herrmann and Stein (295) who
found that the attainment of maturity was delayed in young
male rats and rabbits by injections of ovarian lipoids. Heavier
doses produced degenerative changes in the testis similar to
those caused by X-rays. Fellner (195) confirmed these
results, but found that the administration of testis lipoids had
the same effect, which was therefore probably not specific.
Gould and Doisy (see Allen and Doisy, 21) arrived at a similar
conclusion. Bugbee and Simond (112) failed to find any adverse
effects on the male. Fels (204) has described an inhibiting action
of blood serum of pregnancy on male genitalia, probably due to its
oestrin content. Laqueur and co-workers (345) have recently
produced degenerative testis changes with a comparatively pure
preparation of oestrin, and some 'anti-masculine' action may
therefore occur. It is difficult to reconcile this conclusion with
the presence of oestrin-like substances in the male urine (see
p. no) and possibly in the testis.
Action on non-mammals. Riddle and Tange (535-6), and
Loewe, Voss and Paas (419) have investigated the action of
oestrin in birds, but no definite results have been obtained.
[c] LIMITS OF ACTION
Many workers have suggested that oestrin is the one and only
ovarian hormone, and that it controls all the changes of the
122 INTERNAL SECRETIONS OF THE OVARY
complete reproductive cycle. In addition, the pre-pubertal
development of the accessory organs is usually ascribed to the
action of the same hormone. This view supposes that the corpus
luteum elaborates the cestrus-producing hormone, at least in the
early stages of its life, and that the placenta is also an actual site
of origin. The germ 'gestational gland' has been coined by
Frank and Gustavson (229) to cover the action of Graafian
follicle, corpus luteum, and placenta in successively elaborating
the 'female sex hormone.' Allen also supposes that oestrin
performs all the ovarian functions. ' The continuous availability
of this material from the placenta, present in amounts increasing
with placental growth with the advance of gestation, is the most
logical explanation of the growth of the uterus and mammary
glands, and the absence of menstruation during pregnancy '
(12). Allen and his co-workers (24), however, failed to obtain
oestrin from the corpora lutea, except in the human, and his
view therefore implies that the corpus luteum performs no
endocrine action whatever (except a temporary one in the
human) , and is, in fact , merely a histological ornament . Wadehn
(627) also supposes that the changes of pregnancy are due to
quantitative variation in oestrin, while Zondek and Aschheim
come to much the same conclusion (654).
The whole problem really depends on whether the oestrus-
producing hormone is responsible for the changes in the post-
oestrous phase of the cycle, or whether these changes are under
the control of the corpus luteum. The view that oestrin controls
the post-ovulation changes is based largely on two sets of facts:
(a) that the hormone causes hypertrophy of the accessory organs,
including the mammary glands, and (b) that the hormone is
present in very large amounts during pregnancy. It has yet to
be definitely shown, however, that the hypertrophy which can be
produced by oestrin is greater than that occurring during oestrus,
and so far we have no evidence as to the significance of the
abundance of the hormone during pregnancy.
Animals such as the rabbit and the ferret, which stay on
oestrus indefinitely in the absence of copulation, provide strong
evidence that the action of the oestrus-producing hormone
relates to oestrus only. During the persistent oestrus in these
animals the accessory organs will remain in a static condition for
THE FUNXTIOX OF (ESTRIX 123
months. Immediately ovulation takes place and the corpora
lutea are formed an entirely new phase of growth is initiated.
A large mass of work begun by Fraenkel (208-9) and extended by
Ancel and Bouin (30-2), Loeb (400 ), Marshall (444), and Ham-
mond (264), shows fairly definitely that the corpus luteum
itself controls the changes of the post-oestrous phase b}' some
specific internal secretion distinct from that associated with the
production of oestrus. This work is dealt with in Chapter X, but
one further point may be mentioned here. Functional correla-
tion suggests that the corpus luteum and the oestrus-producing
substance are to some extent antagonistic. The injection of the
oestrus-producing hormone during pseudo-pregnancy will over-
ride this stage of the cycle and result in the reappearance of
oestrus. In the same way its injection during pregnancy has
been shown to lead to abortion or reabsorption of the foetuses.
If the apparent antagonism between the oestrus-producing sub-
stance and the corpus luteum is a genuine effect, it is difficult
to see how the one hormone can control the whole of the
fertile cycle.
The controversy about the limitations of the action of oestrin
therefore centres round three problems:
ia) Is oestrin responsible for the pre-pubertal development of
the accessory organs before the first oestrus?
(h) Is oestrin responsible for the post-ovulation changes in the
vagina, uterus, and mammary glands?
(c) What is the significance of the abundance of oestrin in the
placenta and body fluids during pregnancy?
These problems are discussed below.
[d) ATTAINMENT OF PUBERTY
It has been pointed out above (p. 20) that the attainment of
puberty consists of two phases; the gradual pre-pubertal growth
of the accessory organs and the sudden appearance of the first
oestrous period. There is clearly no reason to distinguish
between the causative mechanism of this first oestrous period
and that of any subsequent one. The underlying mechanism
must be the same, although it is operating for the first time at
124 INTERNAL SECRETIONS OF THE OVARY
puberty. It has been very clearly demonstrated that injections
of oestrin will cause the immature animal to show abruptly all the
extra-ovarian symptoms of oestrus (19, 652). To some extent
one is justified in calling this a condition of 'precocious puberty.'
On account of this reaction of the immature animal, it has been
supposed by many authors that cestrin is responsible for the
initial pre-pubertal development of the accessory organs. The
induction of precocious cestrus is, however, no real simulation of
the slow and steady growth which takes place in the pre-pubertal
organs, and these experiments only show that the first oestrus is
produced, like the subsequent ones, by the action of cestrin.
It is possible, nevertheless, that oestrin is the factor concerned
in the development of the accessory organs, and that it is present
before puberty in amounts sufficient to produce uterine and
vaginal growth, but insufficient to cause oestrous symptoms.
This view is supported by Laqueur and co-workers (350) who
found that uterine growth may be caused by much smaller
amounts than are required to produce oestrus. Thus, -000007
mgm. per day for five days, or about -i m.u. in all, caused
growth of the immature rat uterus. Zondek and Aschheim (652)
found that treatment with their water-soluble oestrin for 14 days
increased the weight of the immature genitalia of mice from
15-27 mgms. to 75-92 mgms.; hence they conclude that oestrin
is the initial growth stimulus. Other evidence is lacking, and
Eels (202) was unable to find oestrin in the ovary ofthe new-born
child, in which the uterus is already undergoing development.
There is, moreover, some reason to suppose that a separate
ovarian factor (presumably hormonic) is required for the initial
development and subsequent maintenance of the accessory
organs. Marshall (444) states: '. . . The conclusion to be drawn
... is that the ovarian hormone which produces oestrus or heat
is different from that which is responsible for maintaining the
normal uterine nutrition.' Schroder (549) reached a similar
conclusion.
There is no doubt that in many animals all the extra-ovarian
histological and physiological symptoms of oestrus can be
produced in the ovariectomized animal b}^ injection of oestrin,
but Asdell and Marshall (45) found that in anoestrous dogs
injection brought about the changes characteristic of prooestrus,
THE FUNCTION OF (ESTRIN . 125
without those of complete oestrus. Even in rodents, oestrus
induced in the ovariectomized animal is incomplete, in the
experience of most workers, in that copulation does not take
place. Allen and his collaborators (24) report that seven out
of eleven rats copulated at induced oestrus, but Parkes, Fielding
and Brambell (510J found that of 92 induced oestrous periods in
mated ovariectomized mice only seven w^ere accompanied
by copulation. Of these seven, three occurred in mice
which were found to have regenerated ovarian tissue. Copula-
tion takes place freely during induced oestrus in ovariectomized
mice with regenerated ovarian tissue, and during oestrus
induced in the normal animal. In the writer's experience, also,
ovariectomized rabbits, receiving heavy injections of oestrin, do
not show the tail erection reflex, and will not copulate. Lacas-
sagne and Gricouroff (338), however, state that oestrin will
produce copulation in the ovariectomized rabbit. So far as
these results go, copulation is clearly not usual at induced
oestrus in the ovariectomized mouse, and the missing factor may
well be that responsible for the initial development of the
accessory organs.
On this view, the ovary probably controls the initial develop-
ment of the accessory organs by means of some basic internal
secretion analogous to that produced by the interstitial tissue
of the testis. To this constant endocrine activity, the cyclically
active hormones of the oestrous cycle are added at puberty.
The irregular occurrence of ovarian interstitial tissue and its late
appearance even where present, make it highly improbable
that it can elaborate this basic ovarian secretion in a manner
analogous with the endocrine activity of the interstitial tissue of
the testis. Some such assumption is, however, implied by the
use of the term puberty gland ' by Steinach (590) and others to
denote both ovarian and testicular interstitial tissue.
{e) (ESTRIN AND THE CHANGES OF THE
POST-OVULATION PHASE
If the changes of the whole cycle are under the control of
oestrin, obviously the continuous injection of oestrin ought not
to result in the prolongation of oestrous symptoms, but in the
126 INTERNAL SECRETIONS OF THE OVARY
appearance of changes characteristic of the post-ovulative phase.
Animals such as the rabbit and the ferret which remain on
oestrus indefinitely (in the absence of copulation) without show-
ing any other changes, lead one to suppose that prolonged
exertion of the oestrous stimulus does not cause changes
characteristic of the luteal phase. This supposition can be
confirmed experimentally — continued injection of oestrin results
in continued oestrus, probably with exaggerated symptoms, but
in nothing more. Analysis of the results produced in the
various accessory organs by prolonged administration of oestrin
fails to show the production of any changes characteristic of the
luteal phase.
Changes tn the vagina. Cornification of the immature or
ovariectomized rodent vagina has been prolonged for nine days
by Frank, Kingery and Gustavson (224), thirteen days by
Parkes and Bellerby (505), two to three weeks by Zondek and
Aschheim (652), fifteen days by Mahnert and Siegmund (436),
and thirteen days by Tuisk (619), who concludes 'that we shall
always find a prolonged oestrus if in any way the threshold
concentration of the follicular hormone is maintained in the
blood.' Failure to secure persistent cornification in the normal
adult with the doses employed may be due to the cyclic activity
of the corpus luteum. The evidence is thus against the view
that continued injection of oestrin in the ovariectomized animal
will produce anything but prolonged cornification in the vagina,
which contrasts definitely with the disappearance of cornified
cells and the great infiltration of leucocytes which follow
ovulation in the normal animal. Continued action of oestrin
is therefore antagonistic to the vaginal changes characteristic of
the luteal phase.
(Estrin and the post-ccstrous uterus. The growth of the uterus
in the non-pregnant animal may be divided into two phases,
namely that occurring at oestrus and that occurring after
ovulation, presumably as a preparation for the reception of a
fertilized ovum. In most animals these two phases are quite
distinct and the changes characteristic of oestrus pass off before
the onset of those correlated with the luteal phase of the cycle.
It is known quite definitely that the oestrous changes in the
uterus are brought about by the oestrus-producing hormone, and
THE FUNCTION OF (ESTRIN 127
accordingly it seems highly improbable that a further set of
different changes can be produced by the same agency. In the
normal mouse and rat the injection of oestrin after copulation
leads to the return of oestrous symptoms, and to the failure of
the embryos to become implanted. There is sufficient evidence
to state, therefore, that in the rat and mouse the oestrus-
producing hormone is not concerned with the production of
post-oestrous changes; the experimental prolongation of its
activity is directly antagonistic to the changes of the luteal
phase. In the guinea-pig and the cow, as shown respectively by
Loeb and by Hammond, the antagonism between the folli-
cular phase and the post-oestrous phase has been well demon-
strated. These authors showed that removal of the corpora
lutea after ovulation led to a much earlier return of oestrus, and
in these animals, therefore, the action of the corpus luteum is
clearly directly opposed to that of the oestrus-producing
substance. Loeb (405J, too, has shown that the pre-decidual
changes in the guinea-pig uterus are inhibited by the injection
of oestrin, and further, that such treatment inhibits the uterine
sensitivity to mechanical irritation necessary for the formation
of deciduomata. In the rabbit uterus, the prolonged period of
oestrus leads to none of the changes w^hich are characteristic of
pseudo-pregnancy. Many workers have figured the rabbit
uterus after heavy or prolonged dosage w^th oestrin, and in no
case has any appearance of pseudo-pregnancy been produced.
Thus Fellner's early figures (192) and a very recent illustration
by Laqueur (357) both show the typical oestrous condition,
which, as pointed out before (p. 54), is so obviously distinguish-
able from that of pseudo-pregnancy. The same condition is
shown in Courrier's (145) illustrations. Long and Evans (425)
have found that the onset of the next oestrus is fatal to the exist-
ence of deciduomata, which are known to be directly under the
influence of the corpus luteum. Ovulation, therefore, does not
merely transfer the elaboration of the oestrus-producing hormone
from the follicle to the corpus luteum.
The most obvious post-ovulation changes in the uterus are of
course found in Primates, and evidence is now beginning to
accumulate as to the effect of various experimental procedures
upon this pseudo-pregnant development. Allen (12) has
128 INTERNAL SECRETIONS OF THE OVARY
described remarkable experiments on Macaciis rhesus.. His
experimental results may be summarized as follows:
(i) Ovariectomy before the next menstruation is due leads
to its premature appearance. The same effect is
produced by the experimental rupture of the large
follicles in the ovary.
(2) The injection of the follicular hormone into the ovariec-
tomized female results in the rapid appearance of the
local colouration found at certain stages of the cycle
in the female animal.
(3) When the ovariectomized female has been regularly
injected for some considerable period, the cessation of
injection is followed by the appearance of menstruation.
The protocols, however, suggest that this last reaction is less re-
gular and less intense than is found in the normal female.
Allen explains these results by the assumption that the follicular
hormone is responsible for the pre-menstrual congestion of the
uterus, and that its secretion is carried on after ovulation by the
corpus luteum, and, in the event of pregnancy, by the placenta.
These assumptions would explain why menstruation in the
normal animal does not appear until some time after ovulation^
and why it disappears entirely during pregnancy. If the pro-
duction of the follicular hormone is stopped prematurely, owing
to {a) atrophy of the corpus luteum and the failure of placental
tissue to appear, (b) experimental damage to large follicles^ or
double ovariectomy, (c) the cessation of injection in the ovariec-
tomized animal, uterine retrogression resulting in menstruation
sets in. Allen's explanation of his experimental results assumes
a continued activity of oestrin from the beginning of the follicular
phase to the end of pregnancy.
The whole problem, however, is connected with the question
of the interpretation of the menstrual cycle of the Primates.
This has been considered in Chapter V, where it was concluded
that the phenomenon of menstruation is partly pseudo-pregnant
degeneration, and partly a prooestrous occurrence. It is pro-
bable, therefore, that the menstruation produced by Allen in
the injected monkeys was the prooestrous phenomenon, and it
seems possible to interpret all his experimental results on this
THE FUNCTION OF (ESTRIN 129
basis. Allen was inducing that type of menstruation which is
found in non-ovulating monkeys and humans, and which has
been show^n by Corner (126) to be entirely independent of the
true pre-menstrual congestion of the uterus.
(Estyi)i and the mammary gland. Some evidence has been
brought forward in support of the view that prolonged injection
of oestrin will cause complete development of the mammary
glands. The development of the mammary gland in castrated
male guinea-pigs feminized by an ovarian graft (see p. 78) is
sometimes interpreted as being due to the action of cestrin
derived from the graft. The condition produced in the cast-
rated male has been likened by Lipschiitz (371) to prolonged
oestrus, but, even if no organized corpora lutea occur in the
graft, lutein cells are undoubtedly formed by the atretic
follicles of the graft (see p. 76), and it is impossible, therefore,
to exclude the possibility of luteal activity.
Mammary growth has occasionally been reported as accom-
panying abnormal persistence of oestrus in polyoestrous animals.
Thus Courrier (136) has described a guinea-pig in w^hich nympho-
mania, accompanied by cystic ovaries and prolonged cornifica-
tion of the vagina, was associated with development of the
mammary gland. There is no evidence, however, that the
development was equal to that of pregnancy. Probably an
exaggeration of the oestrous development of the glands, which
is considerable in the guinea-pig, had occurred.
Very early in the course of work on ovarian extracts it was
reported that their injection would produce hypertrophy of
the mammary tissue in the ovariectomized or immature
rabbit. Since there can be little doubt that all the early
extracts of ovary contained only the oestrus-producing
substance as the active principle, this finding seems to suggest
some connection between the oestrus-producing hormone and
the mammary gland. Herrmann (291) and Fellner (192)
originally reported the effect in the ovariectomized female
and castrated male rabbit, and quite recently Vintemberger
(624) has confirmed this result. Aschner (41), Frank and
Rosenbloom (231), and Loewe (407) have all reported similar
effects in various animals. According to Hartman, Dupre, and
Allen (279) the reaction of the mammary tissue to oestrin in the
P.S.O. I
130 INTERNAL SECRETIONS OF THE OVARY
opossum is very obvious and can actually be detected b}^
palpation of the intact animal. Allen (12) also has described
hypertrophy of the mammae in Macacus rhesus after the
injection of cestrin, while Loeb (405) has dealt with the same
effect in the guinea-pig. Laqueur and co-workers (351) have
recently, by injecting oestrin, produced hypertrophy of the
mammae in normal and ovariectomized female, and normal
and castrated male guinea-pigs, in young male dogs, and in
castrated monkeys. They consider oestrin to be the normal
stimulus for the entire development of the mammary gland.
Haterius (283) obtained hypertrophy of the mammary tissue of
male guinea-pigs by the same treatment. Similar results on the
ovariectomized guinea-pig have been reported by Steinach and
co-workers (593).
Superficially, all this would appear to be evidence against the
great mass of observational and experimental data which seems
to show that the corpus luteum is responsible for the develop-
ment of the mammary gland. Actually, however, it is highly
probable that no such interpretation can be placed upon these
results. In the first place, few workers claim to have produced
sufficient development in the mammary gland to lead to the
secretion of milk. Fellner (192) specifically states that the
growth was not enough to allow of milk secretion. It seems
most probable that the degree of development of the mammary
gland which can be produced by the injection of oestrin is only
comparable to that which has been shown to occur normally
at oestrus in many species and to be quite independent of the
main development during pregnancy, which is under the control
of the corpus luteum. In Chapter III it was pointed out
that a cycle in the mammary gland in the unmated animal may
be just as typical of the oestrous cycle as is the cycle in the
uterus and the vagina. Thus, in the guinea-pig Loeb and
Hesselberg (402) have shown that proliferation in the mammary
gland occurs at oestrus, while Myers (472) has described cyclic
mammary changes in the non-pregnant rat. Marshall (444)
describes hyperplasia of the mammary gland in the mare at
oestrus. Hartman (271) in discussing the oestrus cycle in the
opossum also emphasizes the fact that the amount of growth in
the mammary gland at the time of oestrus, although consider-
THE FUNCTION OF (ESTRIN 131
able, is negligible compared with that found during pregnancy or
pseudo-pregnancy. In this animal, however, the growth from
prooestrus to the end of pseudo-pregnancy does appear to be
continuous. Hartman's results show that injection of folli-
cular extract into the ovariectomized opossum produces growth
only equal to that of oestrus. In the rabbit the distinction be-
tween the oestrous proliferation of the mammary gland and
that occurring under the influence of the corpus luteum is most
obvious. Ancel and Bouin (34) and Mntemberger (624)
distinguish very definitely between the slight mammary pro-
liferation characteristic of cestrus, which can be produced by the
injection of oestrin, and the extensive hypertrophy which is
characteristic of pregnancy and pseudo-pregnancy. The latter
phase of growth does not occur during even the most prolonged
oestrus and cannot be caused by the injection of the oestrus-
producing hormone. Fellner's (192) extensive illustrations of
the effects of his ovarian extracts on the mammary gland
make it perfectly obvious that in no case was greater develop-
ment induced than is normally found at oestrus.
The comparative influence of oestrin on the mammae of the
rat and guinea-pig is also instructive. In the former animal the
growth produced by oestrin is negligible (565) while in the latter
it is considerable (230). This comparison corresponds exactly
with the conditions in the normal animals at oestrus.
The evidence that oestrin can cause complete development of
the mammary gland is therefore quite inconclusive. Probably,
at most, an exaggeration of the proliferation characteristic of
oestrus can be produced.
(/) SIGNIFICANCE OF DISTRIBUTION
The occurrence of oestrin-like substances in the male cannot
be discussed until their identity with oestrin is demonstrated.
As regards the non-pregnant female, the distribution of oestrin
presents few problems. Its presence in the follicles and stromal
tissue of the ovary, its very doubtful presence in the corpus
luteum, and the small and varying quantities in the blood and
urine, are quite in accordance with expectation.
Significance of occurrence in body fluids during pregnancy.
132 INTERNAL SECRETIONS OF THE OVARY
The most difficult problem in connection with the distribution of
oestrin is to explain its abundance in the body fluids during
pregnancy, i.e. during the luteal phase, and to explain why this
abundance does not result in the appearance of cestrus and
abortion. A priori the excess of oestrin during pregnancy
suggests strongly that it is responsible for the changes of the
luteal phase. Two explanations more in keeping with the mass
of evidence discussed above may be put forward:
(a) In spite of the large amounts of oestrin in the body fluids,
the corpus luteum may still be dominant.
{h) There is some evidence (see p. i88) that the preliminary
action of oestrin is necessary for the effective action of
the luteal hormone. In this case the presence of oestrin
during pregnancy may be a necessary complement to
the action of the corpus luteum.
In either case the idea of a balance between oestrin and the
corpus luteum (see p. 182) is a necessary assumption. The
refinement of methods for obtaining the luteal hormone from
body fluids should enable this point to be decided. The source
of the large excess of oestrin during pregnancy has usually been
put down to its elaboration by the placenta, but Fellner
(194) considers it to be due at least in part to the activity of the
ovarian interstitial tissue.
Significance of occurrence in placenta. The abundance of
oestrin in the placenta has led many workers, notably Allen (12)
and Aschheim (36) to conclude that elaboration of the hormone
is carried on in this organ. It is, however, almost as difficult
to assume that oestrin is elaborated by the placenta as to assume
the opposite. The only evidence in favour of its elaboration by
the placenta is its abundance in the organ, and this is far from
conclusive. On the other hand, various workers have shown that
the injection of oestrin during pregnancy leads to reabsorption
or abortion, and its secretion by the placenta would thus be a
definite anomaly. Weight for weight, the placenta contains
as much oestrin as the ovaries, and since the weight of the
placenta may be anything up to 500 times as much as the ovaries,
it contains about 500 times as much oestrin. If the placenta
elaborates oestrin, therefore, and at the same rate as the ovaries.
THE FUNCTION OF (ESTRIN 133
it would produce 500 times as much as the ovaries; yet
no symptoms of oestrus occur during pregnancy, and the in-
jection of oestrin during pregnancy leads to abortion. If,
on the other hand, the presence of oestrin in the placenta in
such large amounts is not due to its elaboration there, it is
necessary to have some working hypothesis as to the reason for
its occurrence. It has been suggested (506) that the placenta
absorbs oestrin from the maternal circulation in order to protect
the male foetuses from its action. As Lillie (363) has pointed
out, the male foetus must be protected in some manner from the
sex hormones of the mother, and in view of the reported 'anti-
masculine' action of oestrin it is not improbable that some such
mechanism does exist.
CHAPTER VIII
THE PERIODICITY OF (ESTRUS
{a) ROLE OF THE CYCLIC STRUCTURES OF THE OVARY
The intimate correlation between the cyclic changes in the
ovary and those in the accessory organs naturally led to the
conclusion that the periodic development of the cyclic structures
of the ovary was responsible for the cyclic changes in the
accessory organs. At first considerable attention was given to
the corpora lutea as the main regulators of oestrous periodicity;
it was held that their development after each ovulation inhibited
a further appearance of oestrus during their functional lifetime.
This view was supported by the various experiments in which
the removal of the corpora lutea was found to expedite the
appearance of the following oestrous period. There are many
reasons, however, for concluding that oestrus is not necessarily
preceded by the atrophy of corpora lutea. (a) The appearance
of oestrus at puberty or after anoestrus occurs when no corpora
lutea at all are present in the ovary, (b) In many animals the
atrophy of the corpus luteum at the end of pregnancy is not
followed immediately by oestrus (rodents, of course, are an
exception to this). It may be concluded, therefore, that while
the corpora lutea, when caused to persist, undoubtedly have
the effect of delaying the next oestrous period, they do not
regulate the essential periodicity of oestrus. It follows that
if any cyclic structure of the ovary has this function, it
must be the mature Graafian follicle, and many observations
tend to support this view.
{b) RELATION BETWEEN THE GRAAFIAN FOLLICLE AND
THE PRODUCTION OF CESTRUS
Functional correlation. In the normal animal a regular
connection is found between the maturation of Graafian
134
THE PERIODICITY OF (ESTRUS 135
follicles and the appearance of cestrous symptoms in the acces-
sory organs. This correlation is as follows:
{a) (Estrus and ovulation are synchronized throughout the
entire reproductive life of the animal. The first oestrus
appears with the first ovulation at puberty and the last
oestrus is synchronized with the last ovulation at the
menopause. The bat, as pointed out by Courrier (131),
is an exception to this rule. In this animal oestrus
occurs in the autumn and ovulation in the spring.
{h} Where the breeding season is limited, the beginning and
end of the ovarian cycle are correlated wath the begin-
ning and end of uterine activity.
(c) In the ferret and rabbit, where ovulation only occurs after
copulation, mature follicles and the oestrous condition
persist together indefinitely in the absence of coitus.
(d) During the luteal phase of the cycle, particularly during
pseudo-pregnancy and pregnancy, when no follicles
normally mature, no symptoms of oestrus occur.
(e) A condition of persistent oestrus in animals which nor-
mally ovulate spontaneously is sometimes found in
conjunction with persistent cystic follicles in the
ovary.
Since no uterine effect on the maturation of the follicles has
been demonstrated, this functional correlation throughout the
whole lifetime of the animal could most easily be explained on
the grounds that the oestrous cycle is regulated by the periodic
maturation of Graafian follicles.
The tendency to emphasize the importance of the Graafian
follicle in the production of oestrus w^as accentuated when the
liquor folliculi w^as found to contain large amounts of the
oestrus-producing substance. Allen (11) claims to have shown
that the amount of the oestrus-producing substance which can
be obtained from liquor folliculi varies according to the stage of
maturation of the follicle. In this connection he remarks (8)
' Its presence and absence, due to the periodic development of
successive sets of follicles, is sufficient to explain the mechanism
of oestrous phenomena. ' This author and his co-workers actually
maintain that the Graafian follicle elaborates the oestrus-produc-
136 INTERNAL SECRETIONS OF THE OVARY
ing hormone under the influence of the ovum itself. They
state (24) ' From a functional analysis of the follicle through
its various stages of growth, it seems probable that the produc-
tion of this hormone is referable ultimately to the metabolism
of the ovum itself as the dynamic centre of the follicle/ while
Hartman (271) in discussing the production of oestrus in the
opossum says, 'What element of the ovary constitutes the source
of stimuli that lead to procestrus and to cestrus? The
opossum affords an unequivocal answer which is in full accord
with the clear and succinct statements by both Allen and
Robinson, in which they make out a case for the Graafian
follicles. Their reasons I consider conclusive.' Robinson's (537)
work on the ferret led him to conclude that 'the phenomena of
procestrus and oestrus only appear when a group of follicles
has attained a stage of development which may be called
pre-inseminal maturity, and the phenomena are due to some
secretion produced by the follicles in that phase of their develop-
ment.' In a recent paper Zondek and Aschheim (650) conclude
that the oestrus-producing hormone is elaborated by the theca
interna of the follicle. These workers implanted into ovariecto-
mized mice various portions of the human ovary; only the theca
interna implants caused a positive reaction. Even very recently
Hammond and Marshall (267) have stated in connection with
the vaginal oestrous changes in the ferret, ' This outward sign
of the production of the oestrous hormone we consider is due to
the presence of ripe follicles in the ovaries since it is absent
during anoestrus, when only small follicles are present.' All
these statements, however, are made on the evidence of
functional correlation, and the whole hypothesis that the mature
follicle is responsible for the production of oestrus has resulted
from the elimination of other probabilities rather than from
experimental work.
Time relation of follicular maturation and operation of the
oestrus-producing stimulus. Evidence that the maturing Graa-
fian follicle is not the causative factor in the production of
oestrus is forthcoming from the fact that the real maturation
of the follicle only begins after the oestrus-producing stimulus
has become active. It has been known for some time (425) that
double ovariectomy may be followed shortly afterwards by the
THE PERIODICITY OF OESTRUS 137
appearance of oestrus, although no subsequent recurrence is
found unless ovarian regeneration takes place. A similar
observation was made by Coward and Burn (146). The signi-
ficance of this discovery was not, however, appreciated im-
mediately. Brambell and Parkes (82) found that oestrus in the
mouse may occur up to 36-48 hours after double ovariectomy;
the only possible explanation of this phenomenon is that the
oestrus-producing stimulus becomes operative at about 48 hours
before its effect can be discerned by examination of the vaginal
smear. Histological examination of the ovaries removed from
mice coming into oestrus within the two days following the opera-
tion revealed the fact that the real maturation growth of the
follicle had not taken place. The average volume of Graafian
follicles not due to ovulate at the next oestrous period is about
three million /u^. This same size is maintained until about
halfway through the dioestrus preceding ovulation. When the
oestrus-producing stimulus becomes operative the follicles about
to ovulate have increased in size to an average of 3I million ju^.
During the two days preceding ovulation, enormous follicular
growth takes place, so that a follicle at the time of ovulation has
a volume of between eight and nine million /ul^. These facts
show adequately that in the mouse, at any rate, the maturation
of the follicle does not occur until after operation of the oestrus-
producing stimulus, and cannot, therefore, be responsible for
this stimulus.
(c) OCCURRENCE OF CESTRUS AFTER FOLLICULAR
ABLATION
The real test of w^hether or not the maturation of the Graafian
follicle is the causative factor of oestrus is the effect of total
ablation of the follicles. The difficulty of completely destroying
the follicular system of the ovary has delayed such experimental
work. Marshall and Runciman (454) failed to inhibit the onset
of oestrus by rupturing the maturing follicles of the dog, and
hence considered that the presence of mature follicles was not
essential for the production of oestrus. Later, however,
Marshall and Wood (455) were unable to confirm these results.
Various doubtful points may be raised with regard to these
1 38 INTERNAL SECRETIONS OF THE OVARY
experiments. If the initial growth of the uterus of the dog had
begun when the folhcles were ruptured, the removal of the
follicles might have led to the immediate onset of prooestrous
degeneration. The inhibitory effects produced in the later
experiments might be due to the severity of the operation or to
the formation of luteal tissue by the ruptured follicles (see p,
184). There is thus no direct experimental evidence that the
mature Graafian follicle is the essential factor in the production
of oestrus.
Evidence is now available which shows quite definitely that
the maturation of Graafian follicles is not necessary for the
occurrence of oestrous symptoms in the accessory organs. Blair
Bell (56) long ago reported experiments where the grafting of
rabbit ovaries from which the cortical areas had been removed
resulted in the appearance of oestrus in the host. Since the
cortical areas would contain the majority, if not all, of the
Graafian follicles, these experiments provided a hint that
the follicles were not essential for the appearance of oestrus.
The really critical experiment, namely, the investigation of the
results of entire obliteration of the follicular system of the ovary,
has only recently been attempted. Possible means whereby
the ovarian follicles can be eliminated are few, and exposure to
X-rays is by far the most certain technique. The action of
X-rays on the ovary has been studied to some extent physiolo-
gically, and in detail histologically, for many years, and it was
soon found that the Graafian follicles and, indirectly, the
corpora lutea could be eliminated. Their elimination has been
shown to have no effect in inhibiting the development of the
accessory organs in the immature animal, or in causing atrophy
of the accessory organs in the adult. The early w^orkers on the
effects of X-ray sterilization appear to have made no observa-
tions on its effects on the occurrence of the oestrous cycle.
Recently, however, this problem has been investigated in detail
(492-5), and it is possible to state quite definitely that the en-
tire elimination of the whole follicular system neither inhibits
oestrus, nor interferes with its normal periodicity.
Histological effects of exposure to X-rays. Early work on the
histological effects of exposure of the ovary to X-rays was
carried out by Halberstadter (261) and by Fellner (190) on the
THE PERIODICITY OF (ESTRUS
139
rabbit. Bergonie and his co-workers (61-3) showed that while
the Graafian fohicles were caused to undergo complete atrophy,
existing corpora lutea were not affected. Bouin, Ancel, and
Villemin (78) found that the interstitial tissue remained intact,
and according to Steinach and Holznecht (595) the interstitial
'^^iib
Fig. 45. — Ovary of Mouse sterilized by Exposure to
X-RAYS when three WEEKS OLD.
The ovary is largely composed of new tissue derived from the
germinal epithelium.
f.y. follicular remains; ;/./. new tissue.
tissue is actually augmented from the degenerated follicles.
Increase in the interstitial tissue w^as likewise found by Hiissy
and Wallart (309) in the human ovary after irradiation. The
effects of X-rays on the ovary have also been described in detail
by Reifferscheid (529-531).
In the mouse the changes following X-rays have been des-
cribed in detail by Brambell, Fielding, and Parkes (83-6). In
the young animal irradiated before puberty the Graafian
follicles undergo complete atrophy and are entirely reabsorbed.
This atrophy, which includes both the ovum and the membrana
granulosa, eventually involves the theca interna in follicles
where it is differentiated. Finally, the degenerated follicles are
140 INTERNAL SECRETIONS OF THE OVARY
represented merely by small cavities and remnants of zona
pellucida. In a few cases, larger follicles may become filled with
blood and form cysts, or the cells of the theca interna and the
membrana granulosa may grow and invade the antrum to form
a corpus luteum atreticum. These corpora lutea atretica appear
Fig. 46. — Ovary of Mouse sterilized when three weeks
OLD, SHOWING CaVITIES WITH REMAINS OF OVA.
b.v. blood vessel; f.c. follicular cavity.
to persist indefinitely, although no physiological function can
be attributed to them. Concurrently with these changes in the
follicles, the old interfollicular tissue atrophies and proliferation
takes place from the germinal epithelium. This proliferation,
forming a parenchymatous tissue, constitutes almost the whole
THE PERIODICITY OF OESTRUS
141
of the usual type of ovary resulting from sterilization before
puberty. Subsequently, a second proliferation from the germinal
epithelium may take place in the form of cords resembling
small anovular follicles or embryonic testis tubules. The weight
of evidence favours the view that they are of the former nature.
In certain abnormal mice the first proliferation from the
germinal epithelium becomes extremely luteal-like, and, since
this tissue forms almost the whole of the sterilized ovarv, the
J" '
>- ^^
- - t
"V;--
Fig. 47. — Ovary of Mouse sterilized by Exposure to
x-rays when adult.
The ovary is largely composed of tissue derived from the old follicles.
appearance of a single large corpus luteum is produced. This
abnormal type of sterilized ovary results more commonly from
sterilization immediately after birth than at three weeks old.
iVnimals with the abnormal type of sterilized ovary show
physiological abnormalities, including inhibition of oestrus.
Sterilization of the adult mouse leads roughly to the same
kind of histological change, but the details are somewhat
different. In the irradiated adult, the elements already forming
the ovarian cortex at the time of irradiation persist in a changed
142 INTERNAL SECRETIONS OF THE OVARY
form and constitute the bulk of the sterihzed ovary. Im-
mediately after irradiation of the adult, the small follicles be-
come atretic without, however, undergoing complete degenera-
tion; the larger follicles (with the dosage used] survive and may
eventually ovulate. Thus mice may become pregnant up to
so
?
'''$ 9
\»^
*% i> ^ I
$
^1
Fig. 48. — Ovary of Mouse sterilized at Birth.
This shows an abnormal condition of luteinization found in a
small percentage of sterilized ovaries.
some ten days after the application of a dose of X-rays suffi-
cient to result in complete sterility later. The remaining tissues
of the smaller follicles become entirely disorganized, and the
ovary ultimately consists of a more or less uniform tissue
derived from granulosa cells. Small anovular follicles are also
present in the ovaries of the irradiated adult. They are formed,
however, not by proliferation from the germinal epithelium, but
by the degeneration of the ovum and the growth of the mem-
THE PERIODICITY OF (ESTRUS 143
brana granulosa cells of primordial follicles. The adult ovary
does not show proliferations from the germinal epithelium after
sterilization. Such an ovary differs, therefore, from that of the
sterilized immature animal in consisting of tissue of follicular
derivation. No signs of periodic change have been observed in
this tissue once a stable condition has been reached; the histo-
logical periodicity appears to be destroyed as completely in the
adult as in the immature animal. The abnormal luteal type of
Fig. 49. — Uterus of Mouse sterilized at weaning time by
Exposure to X-rays, showing typical cestrous Condition
{cf. fig. 22).
ovary is found onl}^ rarely in the sterilized adult. The effects
of exposure to X-rays in adequate dosage may therefore be
summarized as follows:
(a) Destruction, and in the immature animal complete
reabsorption, of the Graafian follicle, and indirect
elimination of the corpora lutea.
(h) Complete elimination of cyclic histological changes.
Effect of X-ray sterilization on the cestrous cycle. Mice in which
the entire follicular system has been destroyed by exposure to
X-rays, show the typical cestrous changes in the accessory organs
at the same periodicity as the normal female. Sterilization (492-5)
has been described at three ages — {a) just before or at the time
of birth, (h) at weaning, [c] at maturity. In mice irradiated at
weaning time, puberty occurs at the normal date, namely, 6-7
144 INTERNAL SECRETIONS OF THE OVARY
weeks of age, and the subsequent periodicity of oestrus, though
shghtly erratic in certain instances, is reasonably normal. In
mice sterilized at an earlier age, the oestrous cycle tends to be
more abnormal. This is due to the fact that abnormal histo-
logical effects, such as the luteinization of the cells derived from
the germinal epithelium, are more often found. In such animals
the cycle is either absent or ceases after a transitory appearance.
In most sterilized adults the cycle in the accessory organs
persists quite unchanged. Table 9 shows the length of oestrus
and dioestrus in twenty mice before and after irradiation.
5 7 9 11 13 IS 17
Leng-th in days of cycle
19
21
23 25
Fig. 50. — Frequency Polygons for length of Oistrous Cycle
before and after x-ray sterilization.
No significant change is observable.
In all these animals histological examination of the ovaries
showed that no Graafian follicles whatever were present. The
component parts of the cycle, namely dioestrus, prooestrus,
metoestrus, and oestrus, were all found to be normal. The only
unusual feature of the post-irradiation cycle is its slightly
greater variability in length; it is quite evident that this in no
way detracts from the general conclusion that the complete
destruction of the follicular system of the ovaries does not
inhibit the occurrence of oestrus. Zondek (647) has reported
similar results.
Analysis of the experimental results from X-rayed animals
shows that the oestrus-producing hormone is probably elaborated
by the first post -irradiation proliferation from the germinal
epithelium in animals sterilized while immature, and by the
amorphous tissue of follicular derivation in those sterilized when
adult. In the former group the similarity of the proliferation
THE PERIODICITY OF (ESTRUS
145
c
u +->
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el
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>
Difference
after
irrad.
i66|m(v}m6666666 6 66 6
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6
+
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rOO 0
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Difference
after
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+
-f-MHCOCOMOWMCOMMO'^OOOlOOM
+ + + + +1 1 1 1 I+ + + + +I++I +
After
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Before
irrad.
coo 0 M t^OO CTn 0 ^ CO M 00 7^o *7< 9 v^ 9 ^ 9
f-N lo ino 00 iJoKc^K»Jn-t-uo'i-uoJr^ iJno iJn 10
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Animal
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H H H OJ M COCO^^-^^^^VOUO
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P.S.O.
K
146 INTERNAL SECRETIONS OF THE OVARY
to the so-called interstitial tissue of the ovary suggests a com-
parison with the supposed endocrine activity of this latter tissue.
On the other hand, the irregularity of the occurrence of ovarian
interstitial tissue, together with our complete ignorance as to
its endocrine significance, makes it difficult to arrive at any
conclusions.
Significance of occurrence of cestrus after follicular ablation.
Allen (12) has doubted whether any real information regarding
the normal animal can be obtained from these experiments on
sterilized mice. This objection might be of importance if the
sterilized mouse were separate and distinct from the norrral
animal. In animals irradiated when adult, however, the
change to the sterilized condition is very gradual. Even after
a dose of X-rays which will lead to complete sterility, the
degeneration of the follicles is comparatively slow, and during
the transition period when the Graafian follicles are gradually
disappearing, the oestrous cycle shows no abnormality. Since
it is unlikely that the regulation of oestrous periodicity could
be taken over imperceptibly by a different mechanism while the
sterilization changes are proceeding, it is highly probable that
the same periodic mechanism is at work in the sterilized animal
as in the normal. Three general conclusions may be drawn from
these experiments on the occurrence of the oestrous cycle after
X-ray sterilization:
(a) The Graafian follicle is probably not the essential source of
the oestrus-producing hormone;
[h) The periodicity of oestrus is not governed by the periodic
maturation of follicles;
(c) Since the elimination of the corpora lutea of ovulation has
also no effect on the periodicity of oestrus, they can
perform no such inhibitory function in the unmated
mouse as has been demonstrated by Hammond (265) in
the cow and by Loeb (390, 400) in the guinea-pig.
In those animals in which the corpora lutea of ovulation have
been shown to inhibit the next oestrous period, it is probable
that the elimination of the follicular system would, by indirectly
eliminating the corpora lutea, shorten the dioestrous interval.
The same result would be expected in the mouse under condi-
THE PERIODICITY OF (ESTRUS 147
tions where pseudo-pregnancy should normally be produced,
namely, after sterile copulation. Experiments showed, however
that this did not occur in all cases (500). It seems probable
that the mechanism concerned in the stimulation of luteal
tissue after sterile copulation also affects the tissues of the
sterilized ovary. Some X-rayed mice, when mated, will
actually copulate every four or five days, namely, at the
periodicity which is characteristic of oestrus in the unmated
mouse. In the majority, however, this is not found, and
after one or two oestrous periods attended by copulation, the
oestrous cycle gradually fades out.
That the Graafian follicle is not necessarily the essential
source of oestrin is emphasized by quantitative examination of
the liquor folliculi and of the residual tissue of the ovary after
removal of all large follicles (see p. 107). This examination has
shown that oestrin is fairly equally divided between the fol-
licles and the stroma (but see also 613), and it is obvious
that the stroma has at least as good a claim to be considered
the site of origin of oestrin as has the follicular tissue. The
X-ray work reveals the ovary in quite a new light, and two main
problems are raised:
(a) If follicular maturation does not initiate oestrus, by what
means can the synchronization be arranged ?
(b) How is the periodicity of oestrus regulated ?
Periodic action of oestrin. It is possible that oestrin is produced
at intervals and so achieves its periodic action. It must be re-
membered, however, that the oestrus-producing hormone can be
extracted from the ovary even when oestrus is completely in
abeyance, as during anoestrus and pregnancy. If, as seems
probable, the production of oestrin by the ovary is continuous,
some periodic mechanism, such as the cyclic attainment of a
threshold value, must exist. It has also been suggested that the
accessory organs have a periodic increase and decrease in
sensitivity to the oestrus-producing hormone. This supposition
was based on the idea that continuous injection of oestrin would
not prolong the oestrous changes indefinitely. More recent work,
however, has shown that cornification can be prolonged at will
and therefore that no periodic uterine sensitivity occurs (see
148 INTERNAL SECRETIONS OF THE OVARY
p. 126). The difficulties in the way of explaining the periodicity
of oestrus on the ground that the ovary regulates its own
periodicity, complicated the explanation of the results of X-ray
sterilization. Recently, however, the discovery of the pituitary-
ovary mechanism has made possible a simpler explanation.
Synchronization of oestrus and follicular maturation. It was
originally thought from the X-ray experiments that the
oestrus-producing hormone caused the maturation of the follicle
as the oestrous change in the ovary, in the same way as it caused
oestrous changes in the accessory organs. This possibility was
supported by the discovery that in the mouse the maturation
growth of the follicle does not start until the cestrus-producing
stimulus has become active (see p. 136). On this view the
oestrous changes in the accessory organs and the maturation of
the follicle would be synchronized by their both being
due to a common stimulus, namely the oestrus-producing hor-
mone. It has not been possible, however, to substantiate this
hypothesis, owing to the fact that follicular maturation cannot
be induced by the injection of the oestrus-producing hormone at
times when it would not otherwise occur, such as during the
lactation dioestrus, during pseudo-pregnancy, or during preg-
nancy (see p. 119).
{d) OCCURRENCE OF FOLLICULAR MATURATION
WITHOUT (ESTRUS
The opposite result to the occurrence of oestrus after follicular
ablation, i.e. the occurrence of follicular maturation without
oestrus, has been very ingeniously produced by Zondek (647),
who, by feeding thallium to mice, was able to suppress all cyclic
changes in the accessory organs. This suppression probably
means that the production of cestrin was inhibited; but never-
theless the histological ovarian cycle was unaffected, ovulation
and the formation of corpora lutea taking place. The matura-
tion of the Graafian follicle in these animals, therefore, cannot
have been dependent upon the production of oestrin. It is thus
possible to reach the conclusion that the production of oestrin
is not dependent upon the maturation of the Graafian follicles,
and, conversely, that the maturation of the Graafian follicles is
THE PERIODICITY OE CESTRUS 149
not dependent upon the production of oestrin. Zondek (647)
draws attention to this absence of correlation as follows: ' Wir
haben also festgestellt: das Ei beherrscht nicht das Hormon, das
Ovarialhormon beherrscht aber auch nicht das Ei. Ei und
Ovarialhormon stehen nebeneinander, sind koordiniert, sind
gleichberechtigt. Sie stehen aber unter der Herrschaft einer
zentralen Regulation. . . .'
x-\ll this initial work paved the way for the discovery of
the influence exerted on the ovary by the anterior pituitary
body, and it is possible to assert definitely at this stage that the
regulation of ovarian periodicity is controlled by the anterior
pituitary, and that ovarian regulation is therefore external to
the ovary itself. The same conclusion is indicated by numerous
experiments on ovarian grafts.
(e) REASONS FOR SUPPOSING 0\^\RIAN REGULATION
TO BE EXTERNAL
The hypothesis that the periodicity of ovarian endocrine
activity depended upon the periodicity of its cyclic structures
seemed so well-founded that small attention was originally
paid to the facts which implied that the regulation of the ovary
might be to some extent external. Recent work on the mechanism
of the oestrous cycle, however, has brought these experiments
into prominence.
Ovarian grafts. As long ago as 1900 it was shown by Foa (207)
that extraordinary effects were obtained by grafting the ovaries
from one animal into an animal of a different age. He found
that the ovary of the immature animal grafted into an ovariec-
tomized adult underwent rapid development and attained a
state of maturity long before it would have done so in its
original environment. Many experiments have been made in
confirmation of Foa's original observations (365, 367) ,and further
details have been added. Converse experiments have show^n that
an adult ovary grafted into an ovariectomized immature female
loses both its histological and endocrine cyclic activity. Control
operations have shown quite adequately that neither effect is
due to manipulation, and it must be concluded, therefore, that
the ovarian age is regulated by that of the soma. Parallel
150 INTERNAL SECRETIONS OF THE OVARY
results are obtained when an ovary is grafted into a cas-
trated male, though where a mature ovary is grafted into a
mature male, the periodicity of the ovary is lost and a state
which has been compared to persistent oestrus appears. Ham-
mond (264) sums up these facts by stating that 'the age of
puberty is determined by the nutritive state of the soma of the
animal and not by age changes in the ovary itself.'
Compensatory hypertrophy of the ovary. It has been known
for many years that the removal of one ovary will result in the
immediate hypertrophy of the remaining ovary. This
hypertrophy usually proceeds to a degree which makes possible
the maturation of the usual number of follicles (35, 42, 115, 148).
It is possible to obtain the same result after removing the whole
of one ovary and the greater part of the other. Thus, a small
ovarian fragment will undergo extensive hypertrophy and
in time produce as many follicles as would both original
ovaries. This process of hypertrophy appears to be limited
only by the supply of oocytes. Lipschiitz (375) has shown
quite definitely that after a time hypertrophied ovarian tissue
may become almost denuded of oocytes owing to the com-
paratively small number left in the original fragment. This
observation is very strong evidence against the view that germ-
cells may be re-formed in the adult female. The hypertrophy of
ovarian fragments is paralleled by the hypotrophy of super-
fluous ovarian tissue. An extra ovary grafted into an animal
does not function — three ovaries produce only the same
number of mature follicles as do two ovaries.
The fact that an animal will ripen the normal number of
follicles from minute amounts of ovarian tissue shows definitely
that some limiting factor is at work to prevent the wholesale
maturation of follicles from the normal ovaries. This has been
expressed by Lipschiitz (370) as the ' law of follicular constancy.'
In the same way the endocrine activity of one ovary becomes as
efficient as that of the initial two ovaries. Thus, the length of
cycle after unilateral ovariectomy has been shown (425, 491,
599) to be indistinguishable from the normal.
The 'generative ferment.' Heape (288) put forward the
hypothesis that some substance required for both growth and
reproduction is present in the body in such small amounts
THE PERIODICITY OF (ESTRUS 151
that only one process can proceed at a time. Hammond (264)
has tentatively accepted this hypothesis to explain the
maturation of immature ovaries grafted into the mature animal,
and the mechanism of compensatory hypertrophy of the ovary.
On this view, the hypothetical substance in question, to which
the name 'generative ferment' was given, is used in the young
animal for growth, none being available for the reproductive
processes, which are thus in abeyance. When body growth
stops, however, the substance can be utilized by the repro-
ductive organs, and their activity begins. Similarly, the sub-
stance may be regarded as essential for follicular maturation,
and the amount present at any one time is only sufficient to
ripen a certain number of follicles. When one ovary is removed
the entire supply is available for the use of the other, which is
therefore able to mature twice the ordinary number of follicles.
A similar view has been adopted by Lipschlitz (369) and others,
who, how^ever, have used the more reasonable term ' X-sub-
stances' to denote the hypothetical factor. An extension of this
hypothesis has been put forward to explain the absence of
ovarian activity during pregnancy, the use of the substance for
uterine and foetal growth preventing its use by the ovary.
Though purely speculative, this theory of the somatic control of
the ovary appears to have received some vindication from recent
work on the effect of anterior pituitary substances on the ovary.
CHAPTER IX
THE RELATION BETWEEN THE OVARY AND THE
ANTERIOR PITUITARY BODY
[a) INTRODUCTION
The observations and experiments recorded in Chapter VIII
make it evident that the ovary does not regulate its own
periodicity, and it is necessary, therefore, to look to some
somatic tissue for the source of this regulation. The evidence
suggests that a Somatic endocrine organ is responsible.
For many years some connection has been assumed between
the gonads and the other endocrine organs: thus in experimental
and clinical studies, the thyroid, thymus, adrenals, and pitui-
tary body have all shown some correlation with the ovary
and testis. The evidence is not sufficiently definite to indi-
cate that the thyroid (656), thymus (444), or adrenals (154)
could be the seat of the regulation of ovarian periodicity. On
the other hand, very striking experiments have recently de-
monstrated a close relationship between the anterior pituitary
body and the ovary.
Various authors, including Frohlich (234) and Cushing (150),
have pointed out that disorders of the anterior pituitary result
in marked aberration of sexual function, as well as in bodily
abnormalities such as gigantism. Both hypo- and hyper-
pituitarism have been described in association with amen-
orrhoea and infantile sex organs in the human female. The
experimental attack on the problem is comparatively recent,
and may be said to have begun with Evans' (182) observations
on the effect of pituitary extracts on the ovary, and with
Smith's (575) work on the effects of hypophysectomy on the
oestrous cycle.
152
OVARY AND THE ANTERIOR PITUITARY BODY 153
{b) LUTEINIZATIOX OF THE CxRAAFIAX FOLLICLE
Evans (182) found that large amounts of a saline extract of
ox anterior pituitary, when injected daily into the normal rat,
resulted in the disappearance of the oestrous cycle during the
whole time that injections were carried out. This suppression
of oestrus was found to be associated with remarkable changes
in the ovary. All Graafian follicles which had reached the size
at which the antrum appears had undergone luteinization,
namely, had formed corpora lutea atretica without the inter-
mediate act of ovulation. As the result of this, the ovaries of the
injected animals came to consist mainly of a large mass of corpora
latea in which were embedded the remains of the ova. At the
cessation of injection the oestrous cycle returned after varying
periods of time. It has since been shown (502, 610) that the
luteal tissue produced as a result of this treatment is remarkably
healthy and will perform all the functions normally associated
with the corpus luteum.
Preparation of extract. Evans' early extracts were merely
made with saline, but his technique was subsequently im-
proved and elaborated as follows (610): Anterior lobes of ox
pituitaries were carefully dissected out, washed, and partially
sterilized in 70 '\j alcohol. The tissue was then ground in a
mortar and extracted over-night on ice with O'lA^ sodium
hydroxide. After extraction, the macerated preparation was
neutralized to phenol red with 0-2 A^ acetic acid. After centri-
fuging, the supernatant fluid was used for injection in amounts
of about I c.c. per day, equivalent to about i gm. of original
tissue. The extracts were injected intraperitoneally by Evans
and his collaborators, and their results have been criticized on the
grounds that the effects ma}^ have been due to the introduction
of irritating material into the peritoneal cavity. It has been
shown, however, that the same result can be produced by
subcutaneous injection (500), and adequate controls with other
tissues have shown quite definitely that the luteinizing effect is
due to some principle of the anterior lobe of the pituitary.
Histological effects. The first histological effect on the ovary
of the injection of such extracts is swelling of the follicular
epithelium, with corresponding crushing in of the antrum
154 INTERNAL SECRETIONS OF THE OVARY
(500). The ovum itself then shows atrophic changes, such as
fragmentation of the nucleus and general shrinkage. The con-
tinuing hypertrophy of the follicular epithelium gradually
closes the cavity of the follicle, until the latter becomes solid.
Mitoses are not observed during this process and the increase in
Fig. 51. — Ovary of adult Mouse injected with Sodium
hydroxide extract of anterior pituitary body.
Large numbers of atretic corpora lutea are present, and few-
follicles.
the amount of tissue can be accounted for solely by the growth,
mainly cytoplasmic, of the individual cells. Allowing for the
restricted space and the presence of liquor folliculi and the ovum,
the process appears to be analogous with that normally occurring
after ovulation. The corpora lutea atretica thus formed are
crowded closely together, but do not become confluent; they
remain as discrete bodies having definite lines of demarcation.
Since the action of this type of extract depends in the normal
OVARY AND THE ANTERIOR PITUITARY BODY 155
m.g.l.
Fig. 52. — Early Stage in Luteinization of the Graafian
Follicle of Mouse by Sodium hydroxide extract of anterior
Pituitary Body.
l.f. liquor folliculi; m.g.l. membrana granulosa undergoing luteiniza-
tion; 0. remains of ovum.
Fig. 53. — Later Stage of Luteinization.
{cf. fig. 52).
156 INTERNAL SECRETIONS OF THE OVARY
animal upon the presence of follicles, it was originally thought
that injection of an animal sterilized by exposure to X-rays
Fig. 54. — Atretic Corpus Luteum in untreated Mouse.
{cf. fig. 52 ).
Fig. 55. — Ovary of Mouse injected with Sodium Hydroxide
extract of anterior Pituitary at three weeks old.
C.I. a. corpus luteum atreticum; 0. remains of ovum.
would not cause the typical inhibition of oestrus, in other words,
would not produce luteinization of the ovary. This expectation,
0\^\RY AND THE ANTERIOR PITUITARY BODY 157
however, was not confirmed (500). Injection of the sterilized
mouse possessing no Graafian folhcles results in the luteiniza-
tion of the whole of the first post-irradiation proliferation (see
p. 140J, and in the consequent inhibition of oestrus in the
accessory organs. This histological alteration of the ovary was
Fig. 30. — Ovary of Rabbit injected with Sodium Hydroxide
EXTRACT of ANTERIOR PiTUITARY, SHOWING LARGE NUMBERS OF
ATRETIC Corpora Lutea.
not permanent, however, and the cessation of injection w^as
followed by the reversion of the luteinized X-rayed ovary to the
normal X-rayed type and by the recurrence of oestrus.
The action of these pituitary extracts has also been investi-
gated in the rabbit. In this animal, also, atretic corpora lutea
are produced on a large scale, but in many cases blood follicles
are formed (502). Many of the atretic corpora lutea are incom-
pletely solidified and contain a fluid centre consisting of blood
and liquor folliculi. In spite of these slight differences in histo-
158 INTERNAL SECRETIONS OF THE OVARY
logical effect, the usual characteristic luteal activity is found in
the ovaries of injected rabbits.
In the immature animal sim.ilar luteinization of the follicles
occurs, neither ovulation nor oestrous changes in the accessory
organs taking place. Brouha (88) found that the action of
oestrin in the immature anima] is not materiall}^ inhibited by
such luteinization.
T , .-*vM:t^v.-?^"\. ^ t .•■'.'''_, ^. ,- . /^ ■
1~ '■^K.
¥\G. 57. — Luteal Tissue of Ovary of Fig. 56, showing
HEALTHY CONDITION.
(c) THE PRODUCTION OF OVULATION
The amount of evidence suggesting that the ovary does not
regulate its own periodicity led Zondek and Aschheim in 1927
(656) to carry out most thorough researches on the effects of
other endocrine organs and body tissues upon the female
reproductive organs. As a result of these researches they came
to the following conclusions:
(a) Implantation of male or female anterior pituitary into the
immature mouse brings about precocious oestrus,
including the ovarian changes.
OVARY AND THE ANTERIOR PITUITARY BODY 159
(b) The effect is exerted purely through the ovary, which is
caused to undergo precocious maturation and to
elaborate the oestrus-producing hormone.
At about the same tim.e, Smith (S7S), having found that hypo-
physectomy in the rat stopped the oestrous cycle, had begun
to investigate the effects of 'implantations' of the pituitary body
^ *f^^-i^<
Fig. 58. — Luteal Tissue produced in sterilized Ovary of
Mouse by anterior Pituitary Preparations (cf. fig. 48).
(576-9). As in Zondek and Aschheim's experiments, the im-
plants were not grafts, but rather intramuscular injections of
macerated tissue. Subcutaneous injection of saline suspensions
of anterior pituitary tissue have the same effect. Later Smith, in
collaboration with Engle, arrived at results substantially the
same as those of the German workers, i.e. that implantation of
anterior pituitary tissue into the immature animal causes the
rapid appearance of the uterine, vaginal, and ovarian changes
characteristic of oestrus. The duration of treatment necessary
is inversely proportional to the age of the animal. The im-
i6o INTERNAL SECRETIONS OF THE OVARY
plants, like the NaOH extracts, have no effect whatever on the
accessory organs in the ovariectomized animal. Smith and
Engle (581) found that the immature ovary after implant-
^■■■
c"*-^.
""V
/
^Wtii'H
f.-
(«)
(6)
Fig. 59. — Effect of anterior pituitary Implants on Ovary
OF immature Rat.
(a) control litter mate; (6) follicular maturation induced on
twenty-ninth day by eight daily implants of two mouse hypophyses
(From Smith and Engle).
ation of anterior pituitary might be ten times as large as the
control in the rat or nineteen times in the mouse. This was
not due to any increase in size of mature follicles as compared
with the normal adult, but to a great increase in their number
and in that of the corpora lutea produced by ovulation.
OVARY AND THE ANTERIOR PITUITARY BODY i6i
The result of injection of suspensions of the anterior pituitary
tissue is therefore essentially the rapid maturation of Graafian
follicles in the ovar}^ with the coincident appearance of oestrous
^-tSA
Fig. 6o. — Effect of anterior Pituitary Implants on Ovary
OF Immature Rat.
(a) Control litter mate; (b) Mature follicles and corpora lutea found
after four daily implantations of anterior lobe of rabbit. (From Smith
and Engle.)
changes in the accessory organs. The production of oestrin by the
stimulated immature ovary in such experiments has been attri-
buted to the hypertrophy caused in the Graafian follicles, but the
P.S.O. L
l62 INTERNAL SECRETIONS OF THE OVARY
same effect has been observed (500) when the injected immature
animals have been sterihzed by exposure to X-rays and there-
fore possess no Graafian folhcles. In such cases the production
of cestrus, and therefore of oestrin, must have been dependent on
the stimulation of the tissue of the irradiated ovary. In the
ovary of the sterilized adult, no histological changes were
r
>4l^.
Fig. 61. — Group of Tubal Ova following Super-ovulation
DURING precocious CEsTRUS IN THE MoUSE.
Forty-eight ova were present in this tube. (From Smith and Engle.
observed following the injection of anterior pituitary suspen-
sions, but the duration of cornified vaginal smears was
prolonged.
The only important difference between the work of Zondek
and Aschheim and that of Smith and Engle seems to be that
whereas the latter authors always found that normal follicular
maturation occurred, the former found that atretic corpora lutea
and blood follicles were also produced. Smith and Engle (581)
actually state: ' In contrast to the large number of follicles
which are found undergoing atresia in the normal animal, we
OVARY AND THE ANTERIOR PITUITARY BODY 163
rarely find any atretic follicles in the precociously matured
animals.' According to Eels (206) the formation of atretic cor-
pora lutea is more frequent than ovulation. In view of the earlier
work of Evans on the luteinizing effects of anterior pituitary
this distinction is of considerable interest. The problem of
whether one or two anterior pituitary substances are involved
is discussed below.
The pioneer work of Zondek and Aschheim, and of Smith and
Engle has now been confirmed- by many authors, including Eels
(206), Brouha and Simonnet (100), Loewe and co-workers (423)
and Siegmund (564). Brouha and Simonnet, however, consider
that another pituitary substance, fat-soluble and producing
oestrus in castrated animals, can be demonstrated.
{d) ARE TWO ANTERIOR PITUITARY SUBSTANCES
CONCERNED IN THE REGULATION OF THE OVARY?
The difference in the effects following injection of Evans'
sodium hydroxide extract and injection of the macerated fresh
tissue, led to the tentative supposition that two different
anterior pituitary principles were involved, one causing the
conversion of the follicular granulosa to luteal cells and another
causing the burst of growth preceding follicular maturation.
Doubt has recently fallen on this supposition. In the first
place, the method of obtaining the two preparations is funda-
mentally different, {a) The macerated suspensions are fresh and
correspond only to minute amounts, 5 to 20 mgms., of fresh
tissue daily; the sodium hydroxide extracts, on the other hand,
made from ox pituitaries, may not be really fresh and the daily
amount injected corresponds to about i gm. of original tissue.
(b) The later preparations used by Evans (182) were subjected to
a fairly drastic chemical treatment, namely, extraction with
sodium hydroxide. His earlier extracts, however, were only
made with saline; and similar suppression of ovulation has
been obtained by Walker (629) who administered the fresh
substance to fowls.
More serious criticism of the view that two anterior pituitary
substances are involved is forthcoming from the work of Zondek
and Aschheim (656). These authors, in investigating the effects
i64 INTERNAL SECRETIONS OF THE OVARY
of implants or extracts on the immature animal, found that
three ovarian reactions were characteristic of this treatment.
(a) In the majority of cases foUicles matured and ovulated, as
found by Smith and Engle.
(b) Other follicles, however, in the same ovary, became atretic
and underwent luteinization without ovulation in a
manner corresponding essentially with that described
by Evans.
(c) A small number of follicles in the same ovary would
become cystic and appear as ' Blutpunkte' on the
surface of the ovary.
Zondek considers these results to show that only one anterior
pituitary hormone is involved, and that the complete luteiniza-
tion caused by the sodium hydroxide extracts is due to the
injection of relatively large amounts, while the characteristic
ovulation effect of the suspensions results from the substance
being present in comparatively small amounts. Zondek's
aqueous extract of anterior pituitary can apparently produce
the three reactions he describes in any proportion, according to
the amounts administered.
Evans and Long (i86) failed to produce oestrus in the im-
mature rat by the injection of the alkali preparation. This
would be expected, since Zondek and Aschheim (658) found
that the ovulation-stimulating principle is destroyed by alkali.
On the other hand, if there are two anterior pituitary sub-
stances affecting the ovary, the mixed ovulation and luteini-
zation obtained by Zondek and Aschheim might be due to the
presence of both substances in the implants. This would, how-
ever, apply equally to Smith and Engle's preparations.
With the evidence available at present, it is difficult to decide
whether two different substances are concerned in these effects,
but basing a conclusion on Evans' results with alkali extracts,
and Zondek and Aschheim's observation that the ovulation -
producing substance is destroyed by alkali, it seems not im-
probable that two are concerned, as supposed by Bellerby (58).
There is little doubt that the problem will soon be cleared up by
administration of (a) the NaOH extracts in smaller amounts,
(b) the two types of preparation simultaneously.
OVARY AND THE ANTERIOR PITUITARY BODY 165
Since recent detailed work on the relation between the
anterior pituitary body and the ovary has been largely con-
cerned with the production of ovulation, the following
account will be restricted to this aspect of the problem.
{e) ASSAY OF ANTERIOR PITUITARY EXTRACTS
It is essential that anterior pituitary extracts should be tested
on the intact animal, as contrasted with oestrin which must be
assayed on the ovariectomized animal. The effects of oestrin
a
c d
Fig. 62. — Effect of CEstrin and anterior Pituitary
Extracts on Uterus and Ovary of young Mouse.
{a) Normal; (6) oestrin — action on uterus only; (c) implants of
anterior pituitary — action on ovary and thence on the uterus; {d)
NaOH extract of anterior pituitary — action on ovary only.
and anterior pituitary on the intact immature animal are
superficially similar, i.e. the production of precocious oestrus.
Actually, of course, the result is quite different. CEstrin, acting
directly on the accessory organs, causes the reaction in the
ovariectomized animal; anterior pituitary preparations,
primarily affecting the ovary, will act only on the intact animal.
When testing for the anterior pituitary substance in fluids
which might contain oestrin {i.e. the urine of pregnancy),
Aschheim and Zondek (39) give a control dose to ovariecto-
mized animals, and, if this is negative, test again on intact
i66 INTERNAL SECRETIONS OF THE OVARY
animals. They use the 6-8 gms. immature mouse, and give six
doses over forty-eight hours. The result appears in about lOO
hours. They define the mouse unit of anterior pituitary hormone
as the amount which given in six doses will produce precocious
oestrus, including the ovarian changes, in the 6-8 gms. mouse
within 100 hours.
(/) PREPARATION AND PROPERTIES
Zondek and Aschheim have obtained aqueous extracts of the
anterior pituitary body which have the same effect as the
implants. Further, they have described (658) the preparation
from urine as follows: The urine is acidified with acetic acid,
filtered, and evaporated down to half the volume. It is then
extracted with ether to remove oestrin. The watery residue is
dialysed and evaporated to dryness. By further purification
(details not given) a whitish amorphous water-soluble powder is
obtained. These authors give a table of the comparative
chemical properties of oestrin and the anterior pituitary sub-
stance (Table 10).
Table 10. — Comparison of Properties of (Estrin
AND THE Anterior Pituitary Substance.
Property.
(Estrin.
Anterior pituitary-
substance.
Thermostability
Resistance to acid and alkali
Solubility in water
Solubility in lipoid solvents
Very stable
Very resistant
Soluble
Very soluble
Destroyed at 6o°C.
Easily destroyed
Soluble
Insoluble
Smith (577) has reported that anterior pituitary substance is
inactive when administered by mouth; probably the active prin-
ciple is destroyed by the digestive enzymes.
{g) DISTRIBUTION OF THE ANTERIOR PITUITARY
HORMONE
After the discovery of the characteristic effects of the injection
of anterior pituitary preparations, attempts were soon made to
OVARY AND THE ANTERIOR PITUITARY BODY 167
discover whether other tissues and body fluids possessed the same
active principle. Zondek and Aschheim (658) found that the
implantation of o-i gm. placenta produced ovulation, while blood
serum and urine of pregnancy were active in amounts of 0-5 c.c.
and 1-2 c.c. respectively. The blood of a pregnant cow was
found to contain even larger amounts (659). Zondek and Asch-
heim thus suppose that elaboration of the hormone is more rapid
during pregnancy, and since it is absent in the urine of the
End of
iincnstruation
Monti) of preg-nancy
Days
postpartum
Anterior pituitary = Ovarian hormone
hormone (oeatrin)
Fig. 63. — Diagram of amount of CEstrin and anterior Pituitary
Substance in the Urine of Pregnancy.
(From Aschheim and Zondek.
non-pregnant female, its occurrence in the urine of early preg-
nancy is used as a test of this condition in the early stages (39).
By means of their technique (see p. 165) for separating and
testing the anterior pituitary hormone and oestrin in the urine,
they are able (40) to give the relative concentrations in the urine
during the course of pregnancy as shown in Table 11 and fig. 63.
Eels (204) also reports the hormone in the blood during
pregnancy. The significance of the occurrence of this large
amount of anterior pituitary substance in the urine of pregnancy
is not as yet understood.
i68 INTERNAL SECRETIONS OF THE OVARY
Aschheim and Zondek (38) have also detected the anterior
pituitary hormone in the decidua, in the corpus luteum of
pregnancy, in navel blood, and in tubal mucous membrane.
Table ii. — (Estrin and Anterior Pituitary Hormone
IN Urine of Pregnancy.
(After Aschheim and Zondek.)
Stage
Qistrin
m.u. per litre
Anterior pituitary
hormone
m.u. per litre
1-8 weeks
3-7 months
7-10 months
300-600
5000-7000
6000-10000
3000-5000
3000-6000
2OCO-3COO
Negative results were given by a large number of other tissues
and fluids, including male urine.
(h) ACTION ON THE NORMAL ANIMAL
Prenatal period. The anterior pituitary hormone would
appear to be unable to traverse the placenta — otherwise, in
view of its abundance in the blood during pregnancy, the
foetuses would presumably be born in a state of sexual maturity.
Prepubertal period. Smith and Engle (581), using mice rather
younger than those used by Zondek and Aschheim, have
secured the ovulation reaction at fifteen days of age, five
days after the beginning of injections. As a result of the
treatment, follicles mature rapidly in the ovary and the antrum
appears, together with the cumulus oophorus. Ovulation then
takes place and corpora lutea are formed. Smith and Engle
imply that the follicles and corpora lutea are all normal, but
according to Zondek and Aschheim atretic and blood follicles
are also formed. These ovarian changes result in the elaboration
of oestrin and in the production of oestrous changes in the
accessory organs. Owing, presumably, to the formation of
corpora lutea, continued administration does not lead to
persistent oestrus, but great hypertrophy of the accessory organs
is produced. The continued injection of both anterior pituitary
OVARY AND THE ANTERIOR PITUITARY BODY 169
substance and oestrin does, however, lead to prolonged oestrus
(658 j. Zondek and Aschheim found that mice in premature
puberty will not mate owing to disparity of size. Smith
(580), however, reports that they will copulate at nineteen days
old if precocious puberty has been induced. Anterior pituitary
implants have a stimulating action on the testis of the young
male, with consequent acceleration of growth in the accessory
organs (626).
Adult animal. Smith and Engle (581) have described the
effect of the anterior pituitary substance on the adult animal.
' Super-ovulation ' of large numbers of follicles takes place in
the ovary, and oestrous changes occur in the accessory organs,
followed by an interval in which an irregular type of vaginal
smear is found. The ovaries of injected animals are found to
contain large numbers of corpora lutea, but these differ essen-
tially from the corpora lutea produced by sodium hydroxide
extracts in that they are normal corpora lutea resulting from
normal maturation and ovulation of Graafian follicles. Pro-
longed dosage will lead to the disappearance by ovulation of all
large-sized follicles from the ovary.
Smith and Engle and also Zondek and Aschheim have found
very numerous true corpora lutea in ovaries of animals thus
treated, and the former workers have found up to forty-eight
ova in one Fallopian tube. Engle (177), however, found, in the
early stages of pregnancy, twenty-nine embryos at most after
such super-ovulation. Much thinning out must take place both
before and after implantation.
Marrian and Parkes (439) have found that the anoestrous
period brought about in the rat by inanition or vitamin B
deficiency may be terminated by an oestrous period following the
administration of anterior pituitary substance. The typical
ovarian and extra-ovarian changes were produced.
The pregnant animal. Zondek and Aschheim (659) and Engle
and Mermod (179) have described the effects of administering
anterior pituitary substance during pregnancy. The latter
found that pregnancy could be readily interrupted in the middle
third, though less easily later on. Ovulation occurred soon after
abortion, but not if pregnancy continued. Zondek and Asch-
heim found that although abortion could be caused, with
170 INTERNAL SECRETIONS OF THE OVARY
suitable dosage ovulation might be induced without terminating
gestation. It is possible that ovulation may be caused by
amounts of anterior pituitary insufficient to produce enough
oestrin to cause abortion. The abortion induced by anterior
pituitary implants is clearly comparable with that produced
by oestrin injection (see p. ii8).
The senile animal. Zondek and Aschheim (658) have reported
that oestrus can be induced in the senile mouse after the
cessation of the cycle by anterior pituitary treatment. In this
case the result differs essentially from that of oestrin injection
(see p. 119) in that the ovary is stimulated to ovulation.
Similar results on mice showing spontaneous ovarian deficiency
have been reported by Loewe, Voss and Pass (423).
(0 THE MECHANISM OF OVARIAN REGULATION
Influence of the anterior pituitary on the ovary. The work
described above makes it evident that the anterior pituitary
body produces a substance or substances which have a most
potent action upon the ovary, and it is thus reasonable to
suppose that the anterior pituitary plays some part in regulating
the normal ovarian cycle. The fact that precocious oestrus can
be induced in the ovaries and accessory organs of the normal
immature animal by administration of anterior pituitary
substance, suggests that the first oestrus of puberty is brought
about by some action of the pituitary. This, in itself, would not
explain how the first oestrus is precipitated: the problem is
merely transferred from the ovary to the anterior pituitary.
Since anterior pituitary bodies from male or female, young or
mature animals are all efficacious, it is difficult to explain how
the first stimulus to the ovary is liberated from the anterior
pituitary. Smith and Engle (581) ' believe that the hypothesis
of the periodic liberation of gonad-stimulating hormone of the
pituitary may explain the periodic ripening of groups of follicles
more satisfactorily than any previously advanced. ' Whether one
or two substances are secreted by the anterior pituitary for
the regulation of the ovary is still uncertain, but if two occur it
would seem that the first is connected with the maturation of
the follicle and the second with the transformation of the
OVARY AND THE ANTERIOR PITUITARY BODY 171
ruptured follicle into luteal tissue. If one substance only is
involved, quantitative variation probably produces the different
effects.
It has been shown by Engle (178) that the hypertrophy of the
remaining ovary after unilateral ovariectomy is greatly
expedited by the injection of anterior pituitary substance. The
author considers this result as evidence that the factor limiting
the number of follicles ovulated at any one time, the factor
governing the law of follicular constancy, variously called
'generative ferment' or X-substance, is merely the follicle-
stimulating principle of the anterior pituitary.
Influence of changes in the accessory organs on the anterior
pituitary. It is necessary to mention here some theoretical
considerations upon which no work has been carried out. If
the anterior pituitary is directly responsible for the changes in
the ovary, then some means must exist whereby events in the
accessory organs can influence the anterior pituitary. For
instance, since ovulation does not take place during pregnancy,
some mechanism must cause the anterior pituitary at this time
to stimulate the corpus luteum and not the follicle. Similarly,
it must be concluded that the absence of oestrus during pseudo-
pregnancy in the mouse indicates that the anterior pituitary
body reacts to sterile copulation and exerts a stimulating
effect upon the corpora lutea. Since the effect of sterile
copulation can be produced by mechanical irritation of the
uterine cervix, it would seem that such stimulation can react
upon the anterior pituitary.
A similar conclusion is reached by another argument. The
fact that cervical stimulation will activate the corpora lutea in
a grafted ovary in the rat (425) makes it fairly certain that the
effect is not direct. The intermediate point, where the stimulus
changes from nervous to endocrine, may reasonably be supposed
to be the anterior pituitary. In the rabbit copulation probably
causes ovulation (see p. 54) by stimulation of the anterior
pituitary. Since the action of copulation can only be nervous
in nature, ovulation in the rabbit would appear to occur as the
result of a vulva-pituitary-ovary chain of stimulation, in which
the first link is nervous and the second endocrine.
Similarly, the fact that lactation in the rat and mouse
172 INTERNAL SECRETIONS OF THE OVARY
causes the corpora lutea of the immediate post-partum ovulation
to persist means presumably that lactation influences the
anterior pituitary body, causing it to exert the luteinizing
stimulus. Then again, premature weaning apparently causes
the anterior pituitary body to exert its stimulus to follicular
maturation.
Influence of events in the ovary on the anterior pitnitary. It is
well known that the ovarian cycle can be expedited by various
means such as puncturing the maturing follicles (478) or
removing young corpora lutea (265). This might theoretically
be explained in either of two ways:
[a) That the alteration in the ovary has expedited a cycle in
the anterior pituitary.
[h) That anterior pituitary substance is made available for
corpora lutea by the destruction of follicles, and vice
versa.
The latter view would presuppose the presence of only one
anterior pituitary substance, and also the absence of endocrine
cycle in the anterior pituitary.
The occurrence of the cycle in X-ray sterilized animals
shows that it is not brought about by follicles and corpora lutea
alternately utilizing a single anterior pituitary substance.
Hence, hypothesis [h) is invalidated and hypothesis (a) must be
accepted. Thus, cyclic endocrine activity exists in the anterior
pituitary, even if only one substance is produced. It is
evident, therefore, that events in the ovary may, under certain
conditions, influence the cycle in the anterior pituitary.
CHAPTER X
THE INTERNAL SECRETION OF THE
CORPUS LUTEUM
{a) INTRODUCTION
The facts discussed in previous chapters make it clear that the
ovary possesses some periodicity which is quite independent of
the periodic production of Graafian folHcles and of corpora
lutea, and which is probably under the control of the anterior
pituitary body. This basic periodicity appears in the unmated
cycle of the normal mouse, because in this animal the complete
ablation of the periodic ovarian structures does not alter the
periodicity of oestrus. In most species, however, even in the
unmated animal, this basic cycle is disturbed by the transient
development of corpora lutea after each ovulation, namely, by
the interpolation of a luteal phase. When pregnancy leads to
the full development of the corpora lutea the derangement of the
cycle is much greater. The conditions in animals, such as the
rabbit and ferret, in which oestrus persists in the absence of
mating, are somewhat difficult to explain by this conception
of the ovarian function; it is necessary to suppose that the
factor which causes periodic oestrus in animals such as the rat
and mouse is persistently operative in the unmated rabbit and
ferret.
The functions of the corpus Jut cum. The analysis of the
mechanism controlling the luteal phase of the sexual cycle has
not yet proceeded as far as the analysis of the mechanism of the
follicular phase. Nevertheless, a great deal of work has been
carried out on the functions of the corpus luteum, and, in spite
of the present tendency to minimize its importance in the
oestrous cycle, certain definite functions can be ascribed to it.
The subsequent history of the corpus luteum formed after
173
174 INTERNAL SECRETIONS OF THE OVARY
ovulation depends both on the species of animal in question, and
also upon the occurrences which take place in the accessory
organs. In the short five-day cycle of the unmated rat and
mouse it may be asserted that the corpus luteum performs no
function. After sterile copulation, however, in these animals,
the corpora lutea, now the corpora lutea of pseudo-pregnancy,
undergo a greater development and the postponement of the
next oestrous period for about twelve days is correlated with the
development of other luteal functions during this pseudo-
pregnant period. Both the rat and the ntouse, for instance,
develop sensitivity of the uterus. This is characteristic of the
activity of the corpus luteum and is always found in the
normal cycle in the guinea-pig, and also during lactation in the
rat and the mouse. In the guinea-pig a prominent luteal phase
corresponding to the pseudo-pregnant period in the rat and
the mouse occurs in the ordinary unmated cycle. In the
Eutheria, the corpora lutea attain their maximum growth and
their full functional activity during pregnancy.
In the rat and mouse, therefore, the history of the corpus
luteum of ovulation depends upon (a) whether copulation has
taken place, and (b) whether copulation is fertile and
followed by the implantation of embryos. In the rabbit and
ferret, no corpora lutea are found until copulation has taken
place; the corpora lutea then undergo great development,
resulting in a period of pseudo-pregnancy which is much more
obvious than that in the guinea-pig, rat, or mouse. In the dog,
where ovulation is spontaneous at oestrus, a well-marked
development of the corpus luteum also takes place quite
irrespective of pregnancy, and a definite pseudo-pregnant
period is found. There exists, therefore, a reciprocal co-ordina-
tion between the accessory organs and the corpora lutea. In
the rat and mouse the corpus luteum of ovulation needs the
stimulus provided by the act of copulation before it can develop
to a functional stage. Further, as in all higher mammals, it
requires the stimulus provided by implantation of embryos
before developing to the fully mature state. Following parturi-
tion in the rat and mouse, lactation causes the corpora lutea of
the post-partum ovulation to become persistent for a period of
about three weeks. The nature of the stimulus exerted by
INTERNAL SECRETION OF CORPUS LUTEUM 175
these occurrences in the accessory organs is not precisely known,
but the anterior pituitary body is presumably concerned (see
p. 171) . Having been stimulated, the corpus luteum elaborates
the internal secretion which performs its various functions.
Many experiments have been carried out on the experimental
ablation and stimulation of the corpora lutea, and as a result of
these it is possible to state that four functions are performed by
the corpora lutea of pregnancy, pseudo-pregnancy, or lactation.
These functions may be summarized as follows:
(a) The inhibition of ovulation and of oestrous changes in the
accessory organs.
{b) The sensitization of the uterus for the implantation of
fertilized ova.
(c) The development of the mammary glands from the
condition in which they are found at oestrus to that
characteristic of the end of the luteal phase.
(d) The maintenance of pregnancy.
Methods of removing the corpora lutea. Experimental work on
the functions of the corpora lutea has dealt mainly with the
effects of their removal. The exact methods by which this has
been achieved have varied considerably and some importance
attaches to this point. Most workers have performed double
ovariectomy in order to remove the corpora lutea, but this
method has the great disadvantage that all ovarian activity is
eliminated; it is therefore quite useless in work dealing with
the return of oestrus. In large monotocous animals, such as the
cow, the corpora lutea can readily be squeezed out from the rest
of the ovary, but in smaller animals, such as the rabbit,
surgical dissection or cauterization has to be employed if the
corpora lutea only are to be eliminated. Such an operation is
very severe and may easily lead to post-operative effects which
may be confused with those due to ablation of the corpora lutea.
As regards the operation during pregnancy, for instance, several
workers have found that their control experiments (cutting the
ovary, etc.) gave almost the same results. In an animal as small
as the mouse even these methods are impracticable. The ideal
subject for experiments of this nature would be a small mono-
tocous animal in which the ovary containing the corpus luteum
176 INTERNAL SECRETIONS OF THE OVARY
could be readily removed, leaving intact the second ovary
containing no corpus luteum, to carry on the other ovarian
functions. Recently a technique has been elaborated whereby
the mouse can be converted to this type (499). This technique
consists in unilaterally sterilizing the young animal by X-rays.
When adult, corpora lutea are present only in one ovary, which
can be removed surgically without any adverse after-effects.
The other ovary, without corpora lutea, is capable of carrying
on the ovarian functions other than those associated with the
corpora lutea (see p. 143).
(b) INHIBITION OF OVULATION AND CESTRUS
The idea that the corpus luteum performs the function of
suppressing ovulation during pregnancy appears to have been
put forward originally by Beard (54) and by Prenant (520).
These authors based their conclusion on the general functional
correlation which is known to exist between the development of
the corpus luteum and the absence of oestrus. This correlation,
of course, is not found during the period of anoestrus, through-
out which ovarian activity, both follicular and luteal, is in
abeyance. This, however, is a special condition, and so far as the
ordinary ovarian cycle of the regular polyoestrous animal is
concerned, the persistence of the corpus luteum is invariably
associated with the absence of oestrus.
A few authentic cases of superfoetation have been recorded
(see Smith, 571). These seem to show that ovulation may
occur during pregnancy, but the condition is very rare, and in
the normal animal the presence of a functioning corpus luteum
prevents ovulation. The exact extent to which follicular
maturation is inhibited by the functional corpus luteum seems
to show specific variation. In the guinea-pig, for instance, Loeb
(389) describes waves of follicular growth even during pregnancy.
This growth ends, however, in atresia, inhibition of ovulation
occurring in all species. During recent years the hypothesis put
forward by Beard and Prenant has been extended by a variety
of experimental studies, the more important of which are
summarized below.
Functional correlation. In certain animals, such as the cow
INTERNAL SECRETION OF CORPUS LUTEUM 177
and the guinea-pig, the removal of the corpora lutea of ovulation
expedites the appearance of the next oestrous period. Thus,
Loeb (390) found that their removal in the guinea-pig led to the
appearance of the next oestrus at about the fourteenth day
instead of at the twentieth day. Loeb (400) also found that
the removal of the corpora lutea during pregnancy did not
always result in the immediate termination of gestation.
Ovulation, however, very soon followed the operation whether
or not the foetuses remained temporarily.
Hammond (265), by squeezing out the corpus luteum from the
ovary of the cow, was able to cause the next oestrous period to
occur at the ninth day instead of the twentieth day. In the
mouse (494), however, indirect elimination of the corpora lutea
by exposure to X-rays does not bring about an earher appear-
ance of the next oestrus. This is due to the fact that in this
animal the dioestrous interval is very short, and practically no
development of the corpora lutea to a functional stage takes
place in the ordinary unmated cycle. In other words, the
unmated mouse possesses no luteal phase in the cycle, and
therefore the elimination of the corpora lutea cannot suppress
such a phase. After sterile copulation, however, when the
cycle in the mouse does possess a luteal phase, the ehmination of
the corpora lutea hastens the reappearance of oestrus. Thus,
in certain mice, ovulation may temporarily be in abeyance at
oestrus so that no corpora lutea are formed. In such animals
pseudo-pregnancy is not found (500) . In the cow and the guinea-
pig the luteal phase is prominent and its elimination, therefore,
leads to the earlier appearance of the next oestrus.
Similar conclusions may be drawn from converse experiments,
namely, from the experimental prolongation of the functional
life of the corpus luteum. Correlated with this there is found
a prolonged disappearance of oestrous changes both in the ovary
and in the accessory reproductive organs. Thus, Loeb (^01)
found that hysterectomy in the guinea-pig causes the corpora
lutea to remain intact and functional for a long period. This
condition is accompanied by a cessation of the oestrous cycle. In
the cow various workers, including Hess (298), Wilhams (638),
and Tandler (608), have found that the persistence of corpora
lutea results in sterihty, owing to the suppression of ovulation.
P.S.O. M
178 INTERNAL SECRETIONS OF THE OVARY
The expulsion of such abnormally long-lived corpora lutea usually
brings about the return of cestrus. In the same way the human
cycle may cease when the corpora lutea persist abnormally and
the removal of such abnormal corpora lutea is followed, accord-
ing to Ochsnier (482), by the return of the cycle. Quite recently
it has been found possible to prolong the functional life of
ovarian luteal tissue almost indefinitely by the injection of
sodium hydroxide extracts of the anterior pituitary body. Thus,
in the mouse and rat the immense production of luteal tissue
which follows such treatment is associated with the complete
absence of oestrus and ovulation, while in the X-rayed animal
the luteinization of the tissue of the sterilized ovary by similar
treatment also brings about the suppression of oestrus.
Mechanism of cestrus inhibition. Definite information is
lacking as to how the corpus luteum brings about the suppression
of oestrus. It is clear that it cannot be merely a local mechanical
effect in the ovary itself ; the presence of a corpus luteum
in one ovary is sufficient to inhibit the oestrus-producing
activity of both ovaries. In the cow and other usually mono-
tocous animals only one corpus luteum at a time is normally
present. In polytocous animals, the same condition can be
produced experimentally by eliminating the Graafian follicles,
and hence the corpora lutea, of one ovary by exposure to X-rays.
Preparation of a-strus-inhibiting extracts. By analogy with
other ovarian functions, it is probable that the oestrus-inhibiting
action of the persistent corpus luteum is brought about by some
endocrine activity. Only very recently, however, have extracts
been prepared from the corpus luteum which have any genuine
oestrus-inhibiting activity. Corner and Hurni (129) reported
negative results from the injection of rats with corpus luteum
preparations, while Loeb (400) working on the guinea-pig, was
unable to produce regularly positive eftects. Pearl and Surface
(512) claim to have succeeded in stopping ovulation in laying
hens by injection of extracts of a commercial preparation of
corpora lutea, while Kennedy (320) reported positive results on
the rabbit by the injection of sahne extracts of similar material.
Pearl and Surface, however, state that toxic effects were
produced by their extracts and the same appears probable in
Kennedy's experiments from the fact that ovulation was
INTERNAL SECRETION OF CORPUS LUTEUM 179
suppressed in some animals for months after the end of treat-
ment. Clearly no physiological action comparable with the
normal activity of the corpus luteum can have caused such a
prolonged inhibitory effect. Haberlandt (259), however, has
more recently described oestrus-inhibiting effects from the
inj-ection of extracts of both ovary and placenta. In the rabbit,
the inhibition and recovery is described by him as consisting of
three stages; (a) complete inhibition of both ovulation and
mating instincts, (b) inhibition of ovulation, though copulation
will take place, (r) restoration of full ovulation and mating
instinct. Since a rabbit in good condition will copulate during
pseudo-pregnancy and pregnancy, one would imagine that
toxic effects also influenced these experiments. The neces-
sity for considering the possible toxicity of tissue extracts
has been emphasized by Kohler (334), and by Herrmann and
Stein (295), who obtained inhibition of oestrus by the injection
of irritant organic substances.
Loewe (417) has reported the inhibition of cestrus in the
mouse by injection of a commercial extract of corpus luteum.
Quite recently various workers have concentrated on the pre-
paration of corpus luteum extracts capable of causing the
suppression of oestrus in the normal animal. Papanicolaou
(487 j injected lipoid extracts into the guinea-pig and brought
about the suppression of oestrus for a considerable period.
No method of preparation, however, was given by this author.
Johnston and Gould (316) were unable to inhibit the action of
oestrus-producing extracts by the simultaneous injection of
extracts of corpus luteum. On the other hand, Parkes and Bel-
lerby (507) inhibited oestrus in the unmated mouse by the injec-
tion of extracts of corpus luteum made with fat solvents.
Corpora lutea of the cow were dissected and all hollow specimens
rejected. The tissue of the solid corpora lutea was then minced
and ground up with anhydrous sodium sulphate. The mixture
was extracted with ether tw^o or three times in the cold, the
ether extracts evaporated down to small bulk, and acetone added
to precipitate the phosphatides. The acetone extract, when
evaporated down, gave a brownish oil which, emulsified with
J ^0 'Jodium bicarbonate and injected subcutaneously, was found
to be active in inhibiting oestrus. Large amounts of this oil had
i8o INTERNAL SECRETIONS OF THE OVARY
to be injected to produce positive results, but control injections
showed that the administration of even larger amounts of inert
fat emulsions had no effect on the cycle.
Beginning work from another viewpoint Hisaw (303) was able
to prepare extracts of corpus luteum which had the remarkable
property of dissolving the pubic ligaments of the guinea-pig and
the pocket gopher. This extract was afterwards found to have
other properties characteristic of luteal activity. Thus, its
injection into the normal animal inhibited oestrus and produced
the sensitivity of the uterus to mechanical stimulation. The
improvement of the extraction of the active substance from the
corpus luteum is still being carried on by Hisaw and his
co-workers, but the most recent information shows that their
extract is prepared essentially as follows. The corpora lutea of
the sow are used, preferably those of a pregnant animal. The
solid tissue only is employed and after grinding in a mortar,
twice the volume of acidified ethyl alcohol is added (98 c.c. of 95 %
alcohol — 2 c.c. of HCl). The mixture is shaken thoroughly and
allowed to stand for twelve hours. The alcohol is then decanted
and a second extract made. This second extract is removed by
means of a press and combined with the first extract. The
alcoholic extract may be used as a stock solution and keeps
well. In the further stages of purification, the alcohol is filtered
and evaporated in a vacuum at a low temperature. The aqueous
residue after removing the alcohol is neutralized to a pH. of 5-4
with a 15% solution of NaOH. A heavy precipitate is formed
which should be filtered off and re-extracted. The salts pre-
sent may be removed by dialysis.
Gley (243) has described the preparation of an oestrus-
inhibiting extract by the following method. Corpora lutea of
the cow are extracted with tartaric acid and the extract treated
with lead acetate. This brings down various toxic substances,
but not the hormone, which is water-soluble. Further purifica-
tion is effected with Cu(0H)2, which is subsequently removed
by HgS. After neutralization the solution is protein free and may
be injected. It causes congestion of the uterus and suppression
of oestrus. Payne, Peenan, and Cartland (511) have described
the preparation of an oestrus-inhibiting substance from corpora
lutea by saponification.
INTERNAL SECRETION OF CORPUS LUTEUM i8i
Distribution of the ccstnts-i)ihibiting Jwnnonc. According to
Hisaw (304), the corpus luteum is the primary source of the
oestrus-inhibiting hormone, but it may be detected in the blood
of the pregnant rabbit, guinea-pig, sow, cat, dog and mare,
and in the maternal and foetal sides of the rabbit placenta.
Properties of the cestr its-inhibiting hormone. Although little
precise experimental work has yet been performed, it is possible
to deduce from the methods which give active extracts certain
of the fundamental properties of the oestrus-inhibiting hormone.
There can be little doubt that the active principle is soluble in
organic solvents such as alcohol, and there is good reason for
supposing that its thermo-stability is low. Hisaw (304) states
that the resistance to acid and alkali is poor, while Parkes and
Bellerby's original extracts seem to show that oxidization is very
rapid. It is fairly evident, therefore, that this oestrus-inhibiting
hormone is not easy to handle, and its delicate nature probably
accounts for the failures of early investigators to obtain active
extracts.
Action of the cestr ns-i}ihihiting hormone. The ovaries of
animals in which oestrus has been inhibited are described by
Papanicolaou (487) as showing complete absence of corpora
lutea, and the presence of a large number of medium-sized
follicles, in which the theca interna was enlarging preparatory
to atresia. The action of the hormone on the ovary and uterus
has been studied by Hisaw and co-workers (305) and Gley (244).
The former state that a condition analogous to that found
during pseudo-pregnancy is set up in the uterus. Gley, on the
other hand, describes effects similar to those following ovariec-
tomy. This result, however, is highly improbable if a true
corpus luteum effect is being reproduced.
Assay of the cestr us-inhihiting hormone. Papanicolaou (487)
suggested that the oestrus-inhibiting hormone should be assayed
by its power to inhibit oestrus in the guinea-pig for a period of
five days, one unit being the amount required to do this. Such a
method, however, would clearly be both inaccurate and cumber-
some. Since it is not possible to say precisely when the next
oestrous period of an animal is due, the degree of inhibition of this
period cannot be gauged precisely. Hisaw (304) and his co-
workers assay their extract by its ability to relax the pubic
i82 INTERNAL SECRETIONS OF THE OVARY
ligaments of the virgin guinea-pig in full cestrus. The smallest
amount that will do this is taken as a unit. Fifteen to twenty
of these units are required daily to inhibit oestrus in a rat. It
seems probable, however, that the most satisfactory way of
assaying the oestrus-inhibiting hormone will be to test it against
a known quantity of the oestrus-producing hormone, but this,
of course, would involve the accurate assay of the oestrus-
producing substance and it cannot be said at the moment that
such accuracy has been achieved.
Scope of function. It has been found by Weichert (633) that
the oestrus-inhibiting hormone also performs the function of
sensitizing the uterus to mechanical stimulation (see p. 184),
and it is thus possible that the same hormone is responsible for
all functions of the corpus luteum.
TJic ijiteyaction of ccstrus-producer and ccstnis-inhihitoy. The
work described above shows that results are being obtained which
indicate that the effects of luteal activity may be reproduced
by extracts. The oestrus-inhibiting action of the corpus luteum
would thus appear to be definitely endocrine in nature. In this
case the interaction of the oestrus-producing hormone and the
oestrus-inhibiting hormone affords a rich field for experimental
work. Preliminary research in this direction has been carried out
by a number of workers, who have injected the oestrus-producing
hormone during the time when the corpus luteum dominates
ovarian activity, as for instance, during pregnancy or pseudo-
pregnancy. In such experiments it has been shown that the
injection of the oestrus-producer in sufficient amounts will
override the inhibiting action of the persistent corpus luteum.
Smith (573), and Parkes and Bellerby (504) were able to override
the activity of the persistent corpora lutea of pregnancy in the
mouse by the injection of the oestrus-producing hormone, while
Engle and Mermod (179) produced the same result by injection
of the oestrus-stimulating extracts of anterior pituitary. Some
doubt exists as to whether the effects produced were due to
overriding the oestrus-inhibiting power of the corpus luteum of
pregnancy, or to local action on the uterus. It is hardly possible,
therefore, to claim these experiments as demonstrating the
interaction of the corpus luteum and the oestrus-producing
hormone. The ordinary dioestrous period of the rat and mouse
INTERNAL SECRETION OF CORPUS LUTEUM 183
is so short that no adequate time is available for experiment and
the same applies in a lesser degree to the pseudo-pregnant
period. During the prolonged inhibition of oestrus which is
found during lactation there is, however, an adequate oppor-
tunity for experimental work. Parkes and Bellerby (505),
studying the effects of injection of the oestrus-producing
hormone into the lactating mouse (see p. 117), showed that a
very considerable oestrus-inhibiting action is set up by lactation.
By means of two other experiments, inhibition was found to be
directly due to the persistent corpora lutea of lactation. In the
first, lactating mice were ovariectomized and the amount of
oestrin required to produce oestrus determined ; the ovariec-
tomized lactating mouse required but very little more oestrin to
produce oestrus than the ordinary ovariectomized mouse. In
other words, the oestrus-inhibition set up by lactation had
disappeared following ovariectomy (505), showing that the
inhibition is set up through the ovary and is not merely due
to the heavy drain upon the metabolism which must result from
lactation. Since the suckling mouse produces between one-fifth
and one-quarter of its own weight per day of milk (505), this
drain upon the suckling mouse is enormous. The second set of
experiments was performed on unilaterally sterilized mice (500).
Such mice were allowed to become pregnant and to suckle their
litter in the ordinary way. During lactation, the ovary contain-
ing the corpora lutea was removed and the sterilized ovary
without corpora lutea left. This sterilized ovary (see p. 143
and p. 175), is capable of carrying on all ovarian endocrine
functions other than those performed by the corpus luteum. In
the lactating mouse containing the sterilized ovary only, the
oestrus inhibition was negligible. These experiments showed
clearly that the oestrus-inhibition set up by lactation is per-
formed through the corpora lutea, which are caused to become
persistent by the act of lactation.
(c) SENSITIZATION OF THE UTERUS
Post-cestroits development of the uterus. The classic work of
Fraenkel (208-11) on the rabbit made it known that the presence
of the corpus luteum is necessary for the attachment of the
i84 INTERNAL SECRETIONS OF THE OVARY
fertilized ovum to the uterine mucosa, and also for the subse-
quent maintenance of foetal nutrition. Fraenkel's work
was confirmed and extended by the experiments of Ancel and
Bouin (29-32) on the same animal. This dependence of the ovum
on the influence of the corpus luteum for attachment is clearly
correlated with the post-ovulative changes which occur in the
uterine endometrium. These changes may be either very
obvious histologically, as in the dog, ferret, and rabbit, or they
may be less perceptible and indicated mainly by physiological
sensitivity, as in the guinea-pig. After sterile copulation in the
rabbit and the consequent formation of the corpora lutea, the
uterus shows growth, vascularization, and particularly glandular
increase in a manner comparable to the growth changes during
pregnancy. During this pseudo-pregnant period the six folds of
the rabbit endometrium proliferate and become so infiltrated with
convoluted glands that in cross section they present a fern-
like appearance. This typical change during pseudo-pregnancy
has been definitely shown to be under the control of the corpus
luteum. Thus, Ancel and Bouin (31), by ablation of the
corpora lutea after ovulation, prevented the typical pseudo-
pregnant changes, while O'Donoghue (478), having produced
luteal tissue experimentally by puncturing the Graafian follicles,
was able to bring about the pseudo-pregnant changes without
the preliminary act of true ovulation. In the dog similar post-
oestrous activity takes place in the uterus, and there can be
little doubt that this is due to the activity of the corpus luteum,
though experimental evidence is lacking. In the same way
post-oestrous changes occur in Dasyurus, and Corner (122) has
described in the uterus of the sow a certain amount of post-
ovulation activity.
Production of deciduomata. In other mammals the actual
histological changes are less obvious, but, nevertheless, in many
cases it has been shown that a peculiar sensitivity to mechanical
irritation is present. This reaction of the post-oestrous uterine
mucosa was originally shown by Loeb (380) in the guinea-pig.
This author, by cutting the endometrium, was able to cause the
production of large blocks of decidual cells, to which the name
placentomata or deciduomata has been given. Loeb (382-3)
found that this sensitivity of the post-oestrous uterus v/as
INTERNAL SECRETION OF CORPUS LUTEUM 185
entirely dependent upon the presence of the corpora lutea and
could be prevented by their removal after ovulation.
The removal of both ovaries has a similar effect in pre-
venting this response of the uterus to mechanical irritation.
Loeb also found that the sensitivity appeared at a certain
definite time after ovulation. These deciduomata were entirely
reabsorbed before the occurrence of the next oestrous period and
■r::33?^^I}^^»
Fig. 64. — Uterus of Mouse (after sterile copulation)
WITH Deciduoma.
/. ]umen; d. deciduoma.
were shown by Loeb to have no effect upon the length of
functional life of the corpus luteum. Pregnancy, however, by
prolonging the life of the corpora lutea, prolongs the life of
deciduomata. Corner and Warren (130) and Frank (219) were
able to produce the same reaction in the rat during lactation.
Long and Evans (425), also working on the rat, found that the
uterus in this anim.al was quite unable to respond to mechanical
stimulation at any stage of the normal unmated cycle. These
authors, however, confirmed the report that the sensitivity did
appear in the rat during pseudo-pregnancy and lactation,
when the corpora lutea undergo a degree of development not
i86 INTERNAL SECRETIONS OF THE OVARY
found in the ordinary unmated cycle. Long and Evans also
introduced the technique of inserting a small loop of surgical
silk transversely through the uterus as a means of providing the
mechanical stimulation. If the stitch passes through the mucosa,
the stimulus which it set up is found to be a§ effective as that
of an actual incision through the muscle and mucosa. The
uterus was found to be most sensitive in the rat about four
days after ovulation. These results have been extended and
confirmed by various workers. Loeb (381), Gasbarrini (236),
Hammond (263), and Nielsen (474) have induced deciduomata
formation during pseudo-pregnancy in the rabbit, while Krainz
(335) has obtained a like effect in the bitch. Results agreeing
exactly with those of Long and Evans (425) on the rat have been
obtained (501) on the mouse, although in this animal the
maximum sensitivity occurs somewhat sooner after copulation.
Evidence of luteal control. The evidence that the corpora
lutea are responsible for this sensitization of the uterus is so
strong that its absence during the ordinary unmated cycle in the
rat is additional evidence that the corpus luteum of ovulation
in this animal does not function. This fact is comparable with
the lack of change in the periodicity of oestrus following oblitera-
tion of the corpora lutea in the unmated mouse. In the rat and
mouse, therefore, the corpus luteum of the unmated animal
possesses neither a sensitizing nor an oestrus-inhibiting function.
Additional evidence of the luteal control of the sensitization of
the uterus has recently been obtained from the fact that
sensitivity does not appear even after copulation in sterilized
mice possessing no corpora lutea (501 ) . Interesting experiments
have been reported by Teel (610) showing that the corpus
luteum in the rat is responsible for the sensitization of the
uterus. It has been described above how the injection of
anterior pituitary extracts will bring about the luteinization of
the ovary. Teel was able to show that with this treatment
the consequent luteinization of the ovary of the rat made
possible the production of deciduomata even in the unmated
animal. The deciduomata were produced most readily when
the operation was made on the fifth day of injection. The
reaction, however, did not occur in the ovariectomized animal
and cannot, therefore, have been directly due to the anterior
INTERNAL SECRETION OF CORPUS LUTEUM 187
pituitary extract. There can be little doubt that the sensitivity
was induced, when normally it would have been absent, by the
Fig. 65. — Uterus of Mouse with Deciduoma.
In this case anterior pituitary extract was used t^ produce the
necessary luteal stimulation.
/. lumen; d. deciduoma.
hypertrophied luteal tissue of the ovary stimulated by the
anterior pituitary preparation. Similar results have since been
i88 INTERNAL SECRETIONS OF THE OVARY
described for both the rat (87) and the mouse (501). In the
mouse the injection of the sodium hydroxide extract of anterior
pituitary made possible the production of deciduomata in the
unmated animal, but not in the unmated sterilized mouse in
spite of the conversion of the irradiated ovary into luteal tissue.
The nature of the activity by which the corpus luteum sensi-
tizes the uterine mucosa is just beginning to be understood.
Loeb (383) originally reported two interesting facts, {a) that the
sensitization is specific to the uterus, other tissues not being
affected, (b) that the sensitization is equally well induced in
grafted uterine tissue. These observations made it clear that
the sensitization is chemical in nature, but initial attempts at
preparing from the corpus luteum an extract capable of causing
this sensitivity, when it would otherwise be absent, were not
successful. Loeb (383) obtained only negative results from the
injection into ovariectomized animals of corpora lutea extracts
and also by the injection of blood from animals in the stage of
uterine sensitivity. These results, however, were merely incon-
clusive. Recently Weichert (633) has been successful in sensi-
tizing the uterus of the rat during the unmated dioestrous cycle
by the injection of the oestrus-inhibiting extract of corpus
luteum as prepared by Hisaw and his co-workers. A similar
result followed injection of corpus luteum extract in the ovariec-
tomized animal, provided that an artificial oestrous period had
been first induced by the injection of the oestrus-producing
hormone. This work of Weichert is of extraordinary interest in
showing that the preliminary activity of the oestrus-producing
hormone may be necessary for the later action of the corpus
luteum secretion.
(d) DEVELOPMENT OF THE MAMMARY GLANDS
The facts which have been recorded in previous chapters make
it clear that two stages of development take place in the
mammary glands even before the appearance of the first corpus
luteum. These two stages are (a) a slight pre-pubertal develop-
ment from the time when the female mammary gland differen-
tiates from its male analogue, and (b) a burst of growth at the
first and subsequent oestrous periods (see p. 130). This growth
INTERNAL SECRETION OF CORPUS LUTEUM 189
still leaves the gland in a rudimentary condition and only during
pregnancy (and to a lesser degree during pseudo-pregnancy)
does the real mammary development occur.
The initial development of the female mammary glands is
clearly endocrine in nature (see p. yS), and the control during
pregnancy must also be of the same nature since transplantation
of mammary tissue to abnormal sites does not affect its normal
development (534). The foetus and placenta, as well as the
ovary, have been suggested as possible sources of the stimulus
required during pregnancy, but a wealth of evidence has now
accumulated to show that the corpus luteum is the responsible
factor.
The mammary gland in pseudo-pregnancy. After ovulation,
correlated with the development of the corpus luteum, an
entirely new phase of mammary growth sets in. The extent to
which this growth takes place in the non-pregnant animal varies
with the species, and with the intensity of the luteal phase in the
non-pregnant animal. In the unmated rat and mouse, where the
luteal phase of the cycle is missing, growth of the mammary
gland appears to take place only at oestrus (see p. 53).
Where the luteal phase becomes pronounced, as during
pseudo-pregnancy in Dasyurus, and in the rabbit and the ferret,
very considerable development of the mammary gl-and takes place
even in the absence of foetuses. The withdrawal of the stimulus
at the end of pseudo-pregnancy in these animals results in at
least a temporary secretion of milk. In the rabbit the develop-
ment of the mammary glands during pseudo-pregnancy and
during pregnancy has been studied in very considerable detail
by Anceland Bouin (30, 32) and Hammond (264). Even when
oestrus has lasted for some months the continued activity of
oestrus-producing hormone causes no development of the
mammary glands other than the slight growth normally
associated with oestrus. Immediately ovulation takes place,
however, and the corpora lutea are formed, development of the
mammary gland begins and even in the absence of pregnancy
continues for some fourteen days, that is, as long as the corpus
luteum of pseudo-pregnancy is functional. This proliferation
consists in the lateral extension and swelling of the ducts.
Clumps^ of alveoli also develop at the ends of the milk ducts.
igo INTERNAL SECRETIONS OF THE OVARY
Ancel and Bouin (30), by removal of the corpora lutea after
sterile copulation, were able to show that this growth of the
mammary gland during pseudo-pregnancy is entirely dependent
upon these structures.
Marshall and Hainan (449) have described the development
of the mammary glands of the dog during pseudo-pregnancy;
in this animal the constructive phase proceeds so far that the
breakdown process at the end of pseudo-pregnancy actually
leads to lactation. In Dasyurus the development of the
mammary glands during pseudo-pregnancy is indistinguishable
from that which occurs during true pregnancy; as in other
species the growth of the gland is correlated with that of the
corpus luteum. In the guinea-pig, according to Loeb and Hes-
selberg (402), the mammary tissue undergoes very little develop-
ment during the luteal phase of the ordinary dioestrous cycle in
the unmated animal, but there is appreciable growth when the
corpora lutea are caused to become abnormally persistent by
hysterectomy. During the dioestrous cycle, even when there
is a definite luteal phase, as in the guinea-pig, the develop-
ment of the gland is not normally carried far enough to result in
the actual secretion of milk. Woodman and Hammond (644),
however, report that virgin heifers after a series of dioestrous
cycles may occasionally secrete a small quantity of milk.
Dieckmann (159) has described the growth which takes place
in the mammary gland during the luteal phase of the human
menstrual cycle. None of the changes characteristic of the
luteal phase are found after ovariectomy or removal of the
corpora lutea. The control of this mammary development
during pseudo-pregnancy is known definitely to be endocrine in
nature.
Our knowledge of the mammary gland during pseudo-
pregnancy makes it evident {a) that the presence of foetuses is
not essential for at least the initial phases of mammary develop-
ment, and (h) that since the only ovarian change in pseudo-
pregnancy is the development of corpora lutea, it is reasonable
to suppose that these bodies are the actual site of origin of the
stimulus required. The almost synchronized appearance of the
katabolic changes in the corpus luteum and in the mammary
tissue further supports this view. As a result of his study of the
INTERNAL SECRETION OF CORPUS LUTEUM 191
mammary gland of Dasyurus, in which pseudo-pregnancy and
pregnancy are of the same length, O'Donoghue (476) came to the
conclusion 'that the corpus luteum is a ductless gland producing
a secretion which is the inciting cause of the growth of the
mammary gland.' Up to the present, however, administration
of corpus luteum preparations to the ovariectomized animal
has not induced a degree of mammary development com-
parable to that found in the normal animal during the luteal
phase of the cycle. This, however, is doubtless due to the failure
of the extracts employed up to the present to contain the active
principle. Preliminary experiments with the oestrus-inhibiting
extracts of the corpus luteum have given negative results.
Loeb and Hesselberg (403) failed to cause mammary develop-
ment by the injection of aqueous extracts of corpus luteum.
Champy and co-workers (119), however, have recently reported
the induction of mammary growth by the injection of luteal
extracts, while Bencan, Champy, and Keller (59) claim that the
corpus luteum substance can be obtained from placentae as
well as from the corpus luteum. The dependence of the
mammary gland on the corpus luteum is further shown by
experiments on artificial pseudo-pregnancy in the rabbit
(Parkes, 502). In these experiments rabbits were injected with
the sodium hydroxide extract of anterior pituitary and corpora
lutea were caused to appear in the ovary in the absence of
copulation. Under these conditions the growth in the mammary
gland typical of pseudo-pregnancy occurred.
The development of the mammary gland during pregnancy. In
the rabbit, the growth of the mammary gland in the first four-
teen days of pregnancy is exactly the same as occurs during
pseudo-pregnancy. After this period, however, a completely
new phase of growth begins, consisting essentially in thicken-
ing of the gland as opposed to lateral extension. That a very
real difference exists between these two stages of pregnancy is
shown by the fact that pseudo-pregnancy in a parous rabbit, in
which the ducts are already well developed, produces no greater
growth of the gland than pseudo-pregnancy following the first
ovulation.
Woodman and Hammond (644), and Asdell (43) have shown
that two phases of m.ammary development occur during preg-
192 INTERNAL SECRETIONS OF THE OVARY
nancy in the cow and the goat. In nulHparae a great develop-
ment of the alveoH begins about midway through pregnancy and
this is correlated with a change in the type of secretion found in
the gland. Before this stage the secretion is of a serous nature
and has the characteristics of diluted milk. At the midway
stage the secretion changes abruptly to a thick pigmented fluid
containing up to 40% of solids, mostly globulin. Drummond-
Robinson and Asdell (171) found that the removal of corpora
lutea in the goat did not result in milk secretion, unless the
operation was performed after the globulin stage had been
reached.
In the guinea-pig, also, two phases of mammary growth
during pregnancy have been described by Loeb and Hesselberg
(403) . For the first twenty-four days of pregnancy (out of sixty)
the mammary gland is more or less quiescent (as in the luteal
phase of the dioestrous cycle) ; from this stage onwards
continuous growth takes place.
The cause of the final development during pregnancy. The
initial growth of the mammary gland during pregnancy,
corresponding to the growth found during pseudo-pregnancy, is
clearly under the influence of the corpus luteum, but for various
reasons it has often been thought that the growth taking place
during the second half of pregnancy requires some additional
stimulus. It has been supposed, for instance, that the corpus
luteum does not function during the later stages of pregnancy
and cannot, therefore, exert an influence on the mammary gland.
It has also been pointed out that the growth of the mammary
gland during pseudo-pregnancy, when no foetuses are present,
does not equal that occurring during true pregnancy. For these
reasons the theory has been held that some foetal or other extra-
ovarian stimulus is required for the complete growth of the
mammary glands. Lane-Claypon and Starling (341) reported
experiments on the injection of foetal and placental extracts into
virgin rabbits. These experiments seem to show that some
growth could be produced by such means, but their illustrations
make it quite clear that the amount of development produced
was far less than that normally found during pseudo-pregnancy
and consequently less than takes place under the influence of the
corpus luteum alone. Ancel and Bouin (33) ascribe the later
INTERNAL SECRETION OF CORPUS LUTEUM 193
development in pregnancy to a uterine myometrial gland.
Hammond (264), however, was unable to find such a gland with
any regularity. The evidence that no foetal factor is required
for the complete development of the glands has always been
strong and has recently become conclusive. In the egg-laying
mammals where no intra-uterine development of the embryo
takes place and where, therefore, no foetal hormone can function,
the mammary glands proceed to their full development and
function normally. Loeb and Hesselberg (403) found no mam-
mary development in the rare cases where pregnancy persisted
for some time after removal of the corpora lutea, but, as these
authors point out, abortion finally took place before extensive
proliferation would have occurred even in normal pregnancy.
Hammond (263) has shown that the presence of decidual tissue
during pseudo-pregnancy in the rabbit does not increase the
mammary development. The real answer to the question,
however, is to be found by prolonging pseudo-pregnancy to
the length of normal pregnancy. In the rabbit the slighter
mammary development during pseudo-pregnancy, as com-
pared with that during true pregnancy, might be due to the
much shorter duration of the former period. In the ferret, where
pseudo-pregnancy has the same duration as true pregnancy, the
development of the mammary glands during both periods is
identical (267). Loeb (401) was able to prolong the life of the
dioestrous corpora lutea in the guinea-pig to the duration of
pregnancy by the operation of hysterectomy. In such animals,
subjected to prolonged luteal action, the mammary glands
underwent development comparable,. if not quite equal, to that
found in pregnancy.
Recently it has been possible to prolong pseudo-pregnancy in
the rabbit to the same length as true pregnancy (502). It was
thought that this could be done by the continuous injection of
the virgin animal with the sodium hydroxide luteinizing pre-
parations of the anterior pituitary body. In practice, however,
prolonged administration of such crude preparations affected
the animal adversely, and it was found preferable to start the
injections at the end of pseudo-pregnancy and to prolong the
life of the corpora lutea so as to stretch out pseudo-pregnancy to
the length of true pregnancy. Under these conditions, in spite
P.S.O. N
l94 INTERNAL SECRETIONS OF THE OVARY
of the absence of foetuses, the mammary glands undergo the
same development as during normal pregnancy. These ex-
FiG. 66. — Photograph of Mammary Gland of Rabbit during
Pseudo-Pregnancy prolonged by anterior Pituitary
Extracts.
Complete growth as found at the end of pregnancy takes place
during prolonged pseudo-pregnancy.
periments show quite conclusively that the complete develop-
ment of the mammary gland can be brought about in the absence
of foetuses and is not, therefore, dependent upon any foetal
INTERNAL SECRETION OF CORPUS LUTEUM 195
stimulus. Similar results appear to have been obtained b}^
Grueter (251), though details are not given by this author.
The failure of Loeb and Hesselberg (403) to induce mammary
growth by injections of corpus luteum preparations was doubt-
less due to the mode of preparation (saline suspensions of
commercial dried tissue). Where autopsy is performed some
days after the cessation of the anterior pituitary injections a
copious secretion of milk is found. This shows definitely that
Fig. 67. — Section of Gland shown in Fig. 66.
secretion is precipitated by removal of the luteal influence and
not by the removal of a hormone present during pregnancy, as
postulated by Gaines and Davidson (235).
Abnormal mammary secretion. Instances in which the
mammary glands function in quite abnormal situations and
at abnormal times are not uncommon. The new-born infant of
either sex quite often shows some abnormal mammary develop-
ment leading to the appearance of a certain amount of milk in
the ducts. The meaning of this is not entirely known, but it
possibly represents some activity on the part of the maternal
stimulus operating through the placenta upon the foetus.
Secretion by virgin animals is also not unknown (622). More
diificult to explain are the cases of mammary secretion in
the adult male (152). Many of these cases are doubtless of
196 INTERNAL SECRETIONS OF THE OVARY
problematical authenticity, but at the same time it is possible
that some at least are genuine.
{e) MAINTENANCE OF PREGNANCY
As pointed out in a previous section there can be little doubt
that the corpus luteum is necessary for the sensitization of
the uterus, and thus probably for the implantation of ova.
Authors are not agreed, however, as to how long the corpus
luteum continues to be essential for the maintenance of preg-
nancy. Fraenkel (211) came to the conclusion that it was only
required during the early stages and that subsequently the
corpus luteum could sometimes be removed with impunity.
Even so, Fraenkel's work showed that the corpus luteum was
necessary for some time after implantation had taken place,
and, therefore, it was not merely concerned with the sensitiza-
tion of the uterus. Marshall and Jolly (451) for the dog and the
rat, and Kleinhaus and Schenk (324) for the rabbit, came to the
same conclusion. Blair Bell (56) and Essen-Moller (181) report
clinical cases which suggest that in the human the removal of
the corpus luteum of pregnancy during the later stages may
produce no adverse effect. Ask-Upmark (46) has collected
similar instances. Herrick (290) found that in some cases
pregnancy would continue in the guinea-pig after double
ovariectomy. In spite of this large amount of evidence showing
that the corpus luteum is not essential during the later stages
of gestation, many authors have come to exactly the opposite
conclusion and find that the removal of the corpora lutea or
ovaries at any stage of pregnancy results in the termination of
gestation. Blair Bell and Hick (57), Hammond (264), Wey-
meersch (635), Mcllroy (430), and Dick and Curtis (155) have
reported that in the rabbit the removal of the ovaries during
pregnancy is inevitably followed by the abortion or reabsorp-
tion of the foetuses. Mulon (466) and Daels (151) state that
ovariectomy at any stage terminates pregnancy in the guinea-pig.
In the cow the removal of the corpora lutea was found by Hess
(298) , by Wester (634) , and by Schmaltz (547) to be incompatible
with the continuance of pregnancy. Similar results have been
described for the goat by Drummond- Robinson and Asdell (171) ,
INTERNAL SECRETION OF CORPUS LUTEUM 197
for the opossum by Hartman (272), for the spermophile by
Drips (170), and for the mouse and the rabbit by Harris (270)
and by Corner (127).
Since the opossum is aplacental, Hartman's (272) results are
of particular interest in showing that the effect of the corpus
luteum is not solely concerned with facilitating implantation.
In addition to the above, cases have been recorded by Hammond
(264) and Hartman (277) in which abnormal degeneration of
the corpora lutea during pregnancy has led to foetal death.
By means of the unilateral sterilization technique (described
on p. 176), it has been possible to get very definite results on
mice (499). Unilaterally sterilized mice become pregnant quite
readily from the untreated ovary, which undergoes considerable
compensatory hypertrophy. In such pregnant mice the removal
of the untreated ovary containing the corpora lutea invariably
results in the termination of pregnancy. The abortion following
the operation takes place 24-48 hours later, so that when the
operation is performed at the seventeenth day of pregnancy,
i.e. two days before parturition would normally take place, the
operative abortion and normal birth coincide. Removal of the
sterilized ovary containing no corpora lutea has no effect on the
gestation. From these experiments it may be concluded that in
the mouse the corpora lutea are necessary throughout pregnancy
until undergoing regression at about the seventeenth day,
forty-eight hours before parturition. There is thus considerable
discrepancy between the results of different workers in this
field, which may to some extent be due to the variation in the
methods used to. remove the corpora lutea and also to the
variety of animals employed. As regards the latter point, it is
improbable that closely related species, such as the rat and
mouse, would show any great difference in the necessity for the
presence of the corpus luteum, and moreover contradictory
results have been obtained by different workers on the same
species of animal. The w^eight of the evidence, however, favours
the view that the corpora lutea are essential during the whole
of pregnancy.
198 INTERNAL SECRETIONS OF THE OVARY
(/) THE FUNCTIONAL RELATION BETWEEN THE CORPUS
LUTEUM AND THE INTERSTITIAL TISSUE
Many workers have supposed that the interstitial tissue and
the corpora lutea are to some extent functionaUy interchange-
able, and, if the former is derived exclusively from degenerate
membrana granulosa, the hypothesis might be supported on the
ground that both are of common origin. It has been shown
(see p. 157) that functional luteal tissue can be produced from
follicles without ovulation, and even from tissue which has
never formed part of an organized follicle; hence the normal
origin from an ovulated follicle is not essential. Further, luteal
and interstitial cells resemble each other to some extent, and
there are thus some histological grounds for supposing that the
interstitial tissue might function as a corpus luteum.
It is necessary to consider, therefore, if there is any evidence
that the four functions of the corpus luteum can be carried on by
the interstitial tissue. As regards the inhibition of oestrus,
Haberlandt (256) found that the graft of a third ovary into a
normal rabbit caused prolonged inhibition of ovulation. Sub-
sequent examination of the graft showed that no corpora lutea
were present, but that the interstitial tissue, as is usual in such
grafts, was highly developed. The inhibition of ovulation was
ascribed to the abundance of interstitial cells.
Pseudo-pregnant development of the uterus has also been
described as being initiated by interstitial tissue. Steinach and
Holzknecht (595) have described pregnancy changes in the uteri
and mammae of virgin guinea-pigs following destruction of the
ovarian follicles by X-rays, and they ascribe the result to the
action of the large amount of interstitial tissue resulting from
follicular degeneration. This explanation was, however, based
on the idea that the interstitial tissue (forming the 'puberty
gland') is the chief endocrine tissue of the ovary, that it
produces the sole ovarian hormone, and that the changes of
pregnancy result from increased production of the one ovarian
hormone. Their illustrations show a perfectly developed
pseudo-pregnant uterus in an irradiated animal possessing
no corpora lutea.
Contrary results are found in the mouse, where X-irradiation
INTERNAL SECRETION OF CORPUS LUTEUM 199
does not interfere with the occurrence of cestrus, and pregnancy
changes are not usually produced. In a few exceptional animals
(508), however, hypertrophy of the mouse uterus is produced by
X-irradiation, and is correlated with the occurrence in the
sterilized ovary of what Lipschiitz and others would call
interstitial tissue, but which w^as considered (84; to have a
greater resemblance to luteal tissue.
As regards the stimulation of the mammary glands by
interstitial tissue, it is possible to quote the well-known feminiza-
tion of the male guinea-pig by an ovarian graft. No corpora
lutea or normal follicles are found in the graft, so that the femini-
zation is usually attributed to the large amount of interstitial
tissue produced by the graft (see p. 78). Since complete
development of the mammary glands is found in such feminized
males, the interstitial cells would seem to be capable of perform-
ing this function of the corpus luteum. This explanation of
hyperfeminization of the male is more probable than the
hypothesis that it depends on the production of cestrin by the
graft.
x\ccording to Biedl (65) the human corpus luteum undergoes
degeneration at the end of the first half of pregnancy, and its
functions are then taken over by the interstitial tissue which is
increased during pregnancy by follicular atresia.
On the other hand, interstitial tissue present before puberty
clearly performs none of the functions of the corpus luteum. In
many animals the interstitial tissue is said to be most abundant
before puberty — i.e. when none of the characteristic luteal
effects have yet appeared. In the rabbit, the changes in the
uterus and mammary glands characteristic of luteal activity do
not appear before the first ovulation, although the pre-pubertal
ovary is full of interstitial tissue.
The whole problem is complicated by the lack of any agreed
definition of interstitial tissue and by the uncertain behaviour
of atretic folHcles. It seems clear that the granulosa cells of
atretic follicles in grafted ovaries produce all gradations of luteal
and interstitial cells (Steinach, 591), and until it is ascertained
whether an absolute difference exists between luteal and in-
terstitial cells further discussion can be of little value.
CHAPTER XI
PARTURITION
Numerous theories of the cause of parturition have been put
forward, and while many subsidiary factors (separation of the
placenta, etc.) no doubt play a part, it is fairly clear that the
physiological changes which initiate the act are due to
hormonic stimuli.
(a) CORRELATION WITH OVARIAN CYCLE
Conditions in pseudo-pregnancy . All the evidence tends to
show that parturition takes place when some particular stage
of the ovarian cycle is reached. Thus, the immediate forerunner
of parturition is the retrogression of the corpora lutea of preg-
nancy. The view that this retrogression of the corpora lutea is
the actual cause of parturition is supported by the fact that at
the end of pseudo-pregnancy some animals show certain
symptoms normally associated with parturition. Thus, the
rabbit plucks its fur and makes a nest, while Dasyurus also
makes a nest and cleans its marsupial pouch. It may be
assumed, therefore, that a certain phase of the Qvarian cycle is
correlated with the occurrence of parturition. No ovarian
extracts, however, have been shown to have any really signifi-
cant direct action upon uterine contraction, so it is probable that
some indirect mechanism is at work.
The relation between oestrus and parturition. Since in many
animals a period of oestrus follows very soon after parturition,
it is necessary to discuss the relation between these two events.
In the mouse, rat, rabbit, and guinea-pig, oestrus follows within
a few hours of parturition. In other mammals, however, a
greater delay is found. ExperimentaHy, it has been shown by
injection of oestrin, or by the implantation of anterior pituitary
200
PARTURITION 201
(see p. 169), that the induction of oestrus during pregnancy leads
to abortion or reabsorption. It is not improbable, therefore,
that the recurrence of the oestrus-producing stimulus, associated
with the atrophy of the corpora lutea at the end of gestation, is
connected with the mechanism of parturition.
Prolongation of gestation by luteal stimulation. Since the
atrophy of the corpora lutea is a necessary prelude to parturition,
it would seem that the injection of active luteal extracts when
this atrophy is beginning should inhibit the onset of parturition.
Data on this point appear to be lacking, but a similar experiment,
i.e. the prolongation of luteal activity, has recently been carried
out by Teel on the rat (611). This author stimulated the luteal
tissue during pregnancy by the injection of NaOH extracts of
the anterior pituitary body (see p. 153 and p. 186), and ap-
parently, as a consequence of this, the period of gestation was
lengthened by two to six days. Teel's conclusions are as
follows: (a) the increase in the gestational period is due to a
delay in implantation of the embryos, (6) the foetuses eventually
die in ittero owing to the failure of the parturition mechanism,
and are expelled still-born, (c) this intra-uterine death is due to
severance of the placental attachment. The delay in the
implantation of the fertilized ovum is contrary to what would
be expected, but even if luteinization of the ovary during
pregnancy does not make possible a longer development of
the fcetuses, it clearly interrupts the normal mechanism of
parturition.
[h) DIRECT ACTION OF THE OVARY ON SPONTANEOUS
UTERINE CONTRACTION
In view of the clear connection between the ovarian cycle and
parturition, it might be supposed that some ovarian secretion
produced at a certain time in the cycle acted directly on the
uterus to cause the activity resulting in parturition. This
possibility is substantiated by the fact that the isolated uterus
shows in its spontaneous contractions a cyclic variation which
is correlated with the ovarian cycle. It has not been possible,
however, to show that ovarian extracts act on the uterus to a
degree necessary to produce parturition. C\C A /
202 INTERNAL SECRETIONS OF THE OVARY
Cyclic variation in the spontaneous activity of the uterus. It is
well known that the uterus of the guinea-pig during cestrus
shows remarkable spontaneous contractions, which make the
organ at that time useless for the standardization of drugs. The
cyclic variation in spontaneous contraction of the uterus has
been studied in the rat by Blair (68), Frank and co-workers
(222), and Clark, KnausandParkes (120); andin thesow by Keye
(321), Corner (124), Seckinger (554), and Wislocki and Gutt-
macher (642). The general conclusion reached is that during
dioestrus the uterus shows rapid feeble contractions, while
cestrus is characterized by fewer but much more powerful con-
tractions. Seckinger, however, obtained rather contrary
results from the Fallopian tube, and Clark, Knaus, and Parkes
found the variation in the rate of conduction of a contraction to
be the most significant factor. Frank and his co-workers (222)
state that after ovariectomy the uterus shows contractions
similar to those occurring during dioestrus. Knaus (327-9) has
extended this type of work to the rabbit uterus during preg-
nancy. This author has carried out very careful experiments,
in which any possible effect of enlargement of the muscle fibres
on the uterine properties was eliminated by the use of a sterilized
cornu containing no foetuses. His results show that during the
first half of pregnancy the uterus is practically inactive owing
to loss of contractihty, while during the second half there occurs
a continuous rise in spontaneous activity, which reaches a
climax at parturition. Knaus ascribes this comparative
quiescence of the uterus during most of gestation to the action
of the corpus luteum.
Direct effect of ovarian extracts on uterine contraction. Acting
on the idea that the greater amplitude of the spontaneous
contractions of the uterus during oestrus is due to the influence
of oestrin, various workers have endeavoured to reproduce the
effect artificially. This has been attempted in two ways, {a)
injection of the ovariectomized or dioestrous female before
preparing the isolated uterus, (h) subjection of the isolated
uterus to oestrin. Frank and co-workers (222) and Brouha and
Simonnet (101-2) state that the contractions of the uterus of the
ovariectomized or dioestrous animal can be altered to those of
oestrus by preliminary injection of the animal. Fellner (192),
PARTURITION
203
in one of his early papers, stated that contraction of the isolated
uterus was caused by the oestrus-producing extract. Laqueur
and his co-workers (356) showed that oestrin stimulated the
isolated uterus, and Brouha and Simonnet (103) later stated
that the contractions of the uterus after ovariectomy can be
caused to change to those typical of oestrus by the addition of
liquor folliculi to the bath in vitro. They ascribe this result to
the presence of an oxytoxic substance in the follicle rather than
to the action of oestrin.
Fraenkel (212), however,
found no effect on the iso-
lated uterus, and Bourne
and Burn (79) found a
direct action of cestrin on
uterine contraction in vitro
only when the uterus was
particularly sensitive. It
may also be pointed out
that Frank's view of the
continued action of oestrin
all through gestation (see
p. 122) is difficult to re-
concile with his results on
uterine contraction. In
any case it seems certain
that the direct effect of
oestrin on uterine contraction is not sufficient to be of
any importance in parturition. The action of active corpus
luteum extracts on the uterus does not seem to have been
investigated.
Fig. 68. — Effect of (Estrin on the
contraction of an isolated guinea-
PiG Uterus.
(From Bourne and Burn).
(c) ROLE OF OXYTOCIN
Since direct action of the ovary on uterine contraction is
insufficient to account for parturition, it is natural to con-
sider if any part may be played by the oxytoxic principle
of the posterior pituitary body. Combining the evidence of
the correlation of the ovarian cycle with parturition, and the
evidence of the action of oxytocin on the uterus, two alternative
204 INTERNAL SECRETIONS OF THE OVARY
theories may be evolved: (a) that the ovary in its immediate
pre-partum state exerts a stimulating action on the posterior
pituitary which then secretes more oxytocin, or (b) that the
immediate pre-partum ovary increases the sensitivity of the
uterus to the posterior pituitary.
Influence of the ovary on the posterior pituitary body. With
the former of these ideas in view Dixon and Marshall (i6i) in-
vestigated the effects on the posterior pituitary body secretion of
extracts of ovaries at various stages of the cycle. These experi-
ments were performed upon the dog, and consisted in the
collection of the cerebro-spinal fluid before and after the
injection of ovarian extracts, and in the assay of the cerebro-
spinal fluid samples for their oxy toxic action. As a result of
these experiments, Dixon and Marshall concluded that the ovary
at a certain stage of its cycle, when the corpora lutea were in
regression, elaborated some secretion which stimulated the
posterior pituitary body. They considered a similar action at
the end of pregnancy to be a vital factor in the mechanism
of parturition. Many criticisms have, however, been levelled
against these experiments, for instance by Swale Vincent
(623). In the first place, doubt exists as to whether the
posterior pituitary body actually secretes into the cerebro-spinal
fluid, and in any case Dyke and Kraft (174) found no changes in
the oxytoxic properties of the cerebro-spinal fluid during labour.
Secondly, various tissue extracts may affect the secretion of the
posterior pituitary, and thirdly, Blau and Hancher (69)
entirely failed to confirm Dixon and Marshall's results. Addis
(i), however, appears to have obtained some clinical results
in keeping with the hypothesis.
Effect of the corpus luteuni on sensitivity of the uterus to oxytocin.
The evidence that the pre-partum ovary stimulates the secretion
of the posterior pituitary is not, therefore, conclusive. On the
other hand, the evidence that the uterus has a cyclic suscepti-
bility to oxytocin does seem to be conclusive. Knaus (326)
found that abortion can only be produced in the rabbit by the
injection of posterior pituitary substance during the last few
days of pregnancy; earlier than this no result follows the
administration of even large amounts. In confirmation of this
result, Knaus (328), as an extension of the work referred to
PARTURITION
205
earlier (see p. 202), has shown that the uterus of the rabbit is
practically insensitive to oxytocin during the first eighteen days
of pregnancy. During the second half of pregnancy the sensi-
tivity increases slowly, until just before parturition, when the
increase is very rapid.
Fitiiitary
Eyctract
oou.
O'OSu
Otsttih.
OO/u
Fig. 69. — Action of CEstrin in sensitizing an isolated
Uterus to Oxytocin.
(From Bourne and Burn).
On Knaus' view^, therefore, parturition is due, at least in part,
to the uterus developing a far greater sensitivity to oxytocin
towards the end than is found during the rest of pregnancy.
This insensitivity during the major part of pregnancy Knaus
ascribes to the action of the persistent corpora lutea of gestation.
Effect of cestrin on sensitivitv of the uterus to oxytocin. In
comparison with Knaus's work on the effect of the corpus
luteum, interesting experiments have recently been performed
on the effect of oestrin on uterine sensitivity. Brouha and
Simonnet (102) and Miura (461) have show^n that the preliminary
injection of the animal wdth oestrin greatly increases the
sensitivity of the uterus to oxytocin. Bourne and Burn (79)
extended this result by showing that the initial treatment of the
uterus in vitro with oestrin greatly increases the response to
oxytocin. Actually a synergism exists between oestrin and
oxytocin.
2o6 INTERNAL SECRETIONS OF THE OVARY
It is clear, therefore, that the dedine of the corpus luteum and
the reassertion of the oestrus-producing stimulus both tend to
increase the sensitivity of the uterus to oxytocin. It is difficult
to estimate at the moment just how important this effect may
be in the causation of parturition, but it would seem that it must
be a highly important, if not crucial, factor.
(d) RELATION OF PARTURITION TO EFFECTS OF
OVARIECTOMY DURING PREGNANCY
It is interesting to consider the effects of double ovariectomy
and removal of corpora lutea during pregnancy in connection
with the theories of the mechanism of parturition. The fact
that double ovariectomy leads to abortion means that either this
effect is not comparable to true parturition, or else that parturi-
tion is not a positive ovarian action. In other words, if the
experimental abortion is comparable to parturition it is difficult
to explain how the latter, as on Dixon and Marshall's results,
can be due to some internal secretion of the ovary. If, on the
other hand, Knaus's work is accepted as the basis of a theory
of the mechanism of parturition, it is clearly possible to compare
the abortion which follows ovariectomy or removal of corpora
lutea during pregnancy with true parturition. Assuming that
the corpora lutea have a de-sensitizing effect on the uterus, their
removal by ovariectomy would permit the reappearance of
uterine sensitivity to oxytocin; if the corpora lutea only were
removed the sensitivity would be increased by the action of the
oestrus-producing hormone of the ovarian stroma. On the whole,
therefore, it seems probable that the termination of pregnancy
which follows removal of the corpora lutea or double ovariec-
tomy can be compared to some extent with true parturition.
This, of course, only applies to the later stages of pregnancy.
The interruption of pregnancy in the early stages, either by
ovariectomy, removal of corpora lutea, or injection of
oestrin, is probably due to adverse uterine changes.
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2o8 INTERNAL SECRETIONS OF THE OVARY
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584. SoBOTTA. 'Ueber die Bildung des Corpus luteum bei der Maus.' Arch, f
mikro. Anat., 47, 1896.
585. SoBOTTA. ' tjber die Entstehung des Corpus luteum der Saugetiere.' Ergebn
d. Anat. u. Entw. Gesch., 8, 1898.
586. SoBOTTA. ' tJber die Entstehung des Corpus luteum der Saugetiere.' Ergebn
d. Anat. u. Entw. Gesch., 11, 1901.
587. SoBOTTA. ' tJber die Bildung des Corpus luteum beim Meerschweinchen.
Anat. Hefte, 32, 1906.
588. SoKOLOFF. ' Ueber den Einfiuss der Ovarien-Exstirpation auf Structurver-
anderungen der Uterus.' Arch. f. Gyn., 51, 1896.
589. SoNNENBERG. 'Die Brunst und ihre Ursache.' Berlin-Tierdrzt. Wochen.,
No. 39, 1907.
590. Stein ACH. ' Umstimmung des Geschlechtscharakters bei Saugetieren durch
Austausch der Pubertatsdriisen.' Zentralbl. f. Phys., 25, 1911.
591. Steinach. ' Willkiirliche Umwandlung von Saugetier-Mannchen.' Pflilgers
Arch., 144, 1912.
592. Steinach. ' Feminierung von Mannchen und Maskulierung von Weibchen.'
Zentralbl. f. Phys., 27, 1913-
593. Steinach, Dohrn, Schoeller, Hohlweg, and Faure. ' tJber die biologischen
Wirkungen des weiblichen Sexualhormons.' Pfliigers Arch., 219, 1928.
594. Steinach, Heinlein, and Wiesner. ' Auslosung des Sexualzyklus, Entwick-
lung der Geschlechtsmerkmale, reaktivierende Wirkung auf den senilen
weiblichen Organismus durch Ovar- und Placentaextrakt.' Pfliigers Arch.,
210, 1925.
595. Steinach and Holzknecht. ' Erhohte Wirkungen der inneren Sekretion bei
Hypertrophie der Pubertatsdriisen.' Arch. f. Entwick.-mech., 42, 1917.
396. Steinach, Kun, and Hohlweg. ' Reaktivierung des senilen Ovars und des
Gesamtorganismus auf hormonalen Wege.' Pfliigers Arch., 219, 1928.
597. Steinhaus. Menstruation und Ovulation. Leipzig, 1890.
598. Stockard. ' The general morphological and physiological importance of the
oestrous problem.' Amer. Jour. Anat., 32, 1923.
599. Stockard and Papanicolaou. ' The existence of a typical oestrous cycle in
the guinea-pig — with a study of its histological and physiological changes.'
Amer. Jour. Anat., 22, 1917.
600. Stockard and Papanicolaou. ' The vaginal closure membrane, copulation,
and the vaginal plug in the guinea-pig, with further considerations of the
oestrous rhythm.' Biol. Bull., 37, 1919.
601. VAN DER Stricht. ' La rupture du follicule ovarique et I'histogenese du corps
jaune.' C.R. Assoc. Anat., 3, 1901.
602. VAN DER Stricht. ' La structure de I'oeuf de la chienne et la genese du corps
jaune.' C.R. Assoc. Anat., 10, 1908.
BIBLIOGRAPHY 231
603. VAN DER Stricht. ' Sur le processus de I'excretion des glandes endocrines:
Le corps jaune et la glande interstitielle de I'ovaire.' Arch. Biol., 27, 1912.
604. Struve. ' Die Perioden der Brunst bei Rindern, Schweinen, und Pferden.'
Filhling's lavdwirtschaft. Zeitiiyig, 60, 191 1.
605. Sutter. ' Cyclic changes in the mammary gland of the rat associated with the
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606. Suzuki. ' On the influence of the placental extracts upon the female genital
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607. Tamura. ' The effects of implantation upon ovarian grafts in the male mouse.'
Proc. Roy. Soc. Edin., 47, 1927.
608. Tandler. ' Ueber den Einfluss der innersekretorischen Anteile der Geschlechts-
driisen auf die aussere Erscheinung des Menschen.' Wien. klin. Woch.,
23, 1910.
609. Tandler and Keller. ' Die Korperform der weichlichen Friihkastraten des
Rindes.' Arch. f. Entwick.-mech., 31, 1910.
610. Teel. ' The effects of injecting anterior hypophysial fluid on the production
of placentomata in rats.' Anier. Jour. Phys., 79, 1926.
611. Teel. ' The effects of injecting anterior hypophysial fluid on the course of
gestation in the rat.' Amer. Jour. Phys., 79, 1926.
612. Thayer, Jordan, and Doisy. ' Improved procedure for the extraction of the
ovarian hormone. II. Some corrections and additions.' Jour. Biol.
Chem., 79, 1928.
613. Thayer and Doisy. ' The distribution of the ovarian hormone between liquor
folliculi and the residual tissue.' Eridocrin., 12, 1928.
614. Trevan. ' The error of determination of toxicity.' Proc. Roy. Soc, B loi,
1927.
615. Trivino. ' tJber Wachstumsteigerung des Uterus durch gravidenserum.'
Klin. Woch., 5, 1926.
616. Truffi. ' L'azione del liquido follicolare e la secrezione interna dell' ovario.'
Boll. Soc. med.-chir. di Pavia, 6, 1926.
617. Tsu. ' Le rythme vaginal chez la lapine.' C.R. Soc. Biol., 89, 1923.
618. TuFFiER. ' Etude chirurgicale sur 230 greffes ovariennes.' Bull. Acad. Med.,
86, 1921.
619. TuiSK. ' Protracted oestrus induced by ovarian extracts.' Jour. Phys., 63,
1927.
620. * Uhlmann. ' Etude critique des methodes detitrage de I'hormone ovarienne.'
La Gynecologic, 26, 1927.
621. Veler and Doisy. ' Extraction of ovarian hormone from urine.' Proc. Soc.
Exp. Biol, and Med., 25, 1928.
622. Velich. ' La secretion du lait sans fecondation.' Le Lait, 6, 1926.
623. * Vincent. ' Current views on " internal secretion." ' Phys. Rev., 7, 1927.
624. Vintemberger. 'Action des injections de liquide folliculaire sur la glande
mammaire.' Arch. Biol., 35, 1925.
625. Voss. ' tJber weibliche Sexualhormone (Thelytropine). XIV. Beitrage zur
Physiologic der vaginalen Brunstvorgange des Meerschweinchens.'
Pfliigers Arch., 216, 1927.
626. Voss and Loewe. ' Geschlechtspragende Wirkungen des Hypophysenvorder-
lappens am Mannchen.' Pfliigers Arch., 218, 1928.
627. Wadehn. ' iJber Sexualhormone, insbesondere das Feminin.' Zeits. f. ang.
Chemie, 41, 1928.
232 INTERNAL SECRETIONS OF THE OVARY
628. Waldeyer. Eierstock tuid Ei. Leipzig, 1870.
629. Walker. 'An inhibition in ovulation in the fowl by the intraperitoneal
administration of fresh anterior hypophyseal substance.' Am^r. Jour.
Phys., 74, 1925.
630. Wang, Richter, and Guttmacher. 'Activity studies on male castrated rats
with ovarian transplants, and correlation of the activity with the histology
of the grafts.' Amer. Jour. Phys., 73, 1925.
631. Watrin. ' Recherches nouvelles sur les injections de liquide folliculaire.'
C.R. Soc. Biol., 93, 1925.
632. Webster. Hitman Placentation. Chicago, 1901.
633. Weichert. ' Production of placentomata in normal and ovariectomized
guinea-pigs and albino rats.' Proc. Soc. Exp. Biol, and Med., 25, 1928.
634. Wester. Eierstock und Ei. Berlin, 192 1.
635. Weymeersch. ' Etude sur le mecanisme de I'avortement apres ovariotomie
double, et sur la restauration uterine consecutive.' Jour. A nat. et Phys., 47,
191 1.
636. WiESNER. 'Die Phasennatur des Sexualzyklus.' Arch. f. Fraitenkunde, 13,
1927.
637. Williams, W. L. Veterinary Obstetrics. Ithaca, 1917.
638. Williams, J. W. Obstetrics. New" York, 4th ed., 1922.
639. Winiwarter. ' Recherches sur I'ovogenese et I'organogenese de I'ovaire des
mammiferes.' Arch. Biol., 17, 1900.
640. Wintz. ' Die physiologisch-chemische Wirkung des Follikelsaftes.' Arch. f.
Gyn., 113, 1920.
641. Wintz. ' Experimentelle Untersuchungen zur inneren Sekretion von Corpus
luteum und Plazenta.' Deut. med. Woch., 50, 1924.
642. WiSLOCKi and Guttmacher. ' Spontaneous peristalsis of the excised whole
uterus and Fallopian tubes of the sow wath reference to the ovulation
cycle.' Bull. Johns Hopkins Hospital, 35, 1924.
643. WoLBACH and Howe. 'Vitamin A deficiency in the guinea-pig. Arch.
Path, and. Lab. Med., 5, 1928.
644. Woodman and Hammond. ' Note on the composition of a fluid obtained from
the udders of virgin heifers.' Jour. Agri. Sci., 12, 1922.
645. Zeitschmann. ' tJber Funktionen des weiblichen Genitale bei Saugetier und
Mensch . ' A rch . f. Gyn ., 115, 1 92 1 .
646. Zondek. ' Das Ovarialhormon und seine klinische Anwendung.' Klin.
Wochen., 5, 1926.
647. Zondek. ' Ei und Hormon.' Arch. f. Gyn., 132, 1927.
648. Zondek. ' Darstellung des weiblichen Sexualhormons aus dem Harn, insbeson-
dere dem Harn von Schwangeren.' Klin. Wochen., 7, 1928.
649. Zondek. ' Weibliche Sexualhormone.' Text. Beil. zur Klin. Wochen., 1928.
650. Zondek and Aschheim. ' Experimentelle Untersuchungen iiber die Funktion
und das Hormon des Ovariums.' Arch. f. Gyn., iiy, 1925.
651. Zondek and Aschheim. ' Experimentelle Untersuchungen liber die Funktion
und das Hormon des Ovariums.' Klin. Wochen., 4, 1925.
652. Zondek and Aschheim. ' Ovarialhormon, Wachstum der Genitalien, sexuelle
Friihreife.' Klin. Wochen., 5, 1926.
653. Zondek and Aschheim. ' Der Scheidenzyklus der weissen Maus als Test-
objekt zum Nachweis des Ovarialhormons.' Klin. Wochen., 5, 1926.
654. Zondek and Aschheim. ' Zur Funktion des Ovariums.' Klin. Wochen., 5,
1926.
655. Zondek and Aschheim. ' Ei und Hormon.' Klin. Wochen., 6, 1927.
656. Zondek and Aschheim. ' Hypophysenvorderlappen und Ovarium.' Arch.
f. Gyn., 130, 1927.
BIBLIOGRAPHY 233
657. Zondek and AscHHEiM. ' Das Hormon des Hypophysenvorderlappens. Test-
objekt zum Xachweis des HorniDns.' Klin. Wochen., 6, 1927.
658. Zondek and Aschheim. ' Das Hormon des Hypophysenvorderlappens.
Darstellung, chemische Eigenschaften, biologische Wirkungen.' Klin.
Wochen., 7, 1928.
659. Zondek and Aschheim. ' Ovulation in der Graviditat— ausgelost durch
Hypophysenvorderlappenhormon.' Endokyinologie, 1, 1928.
660. Zondek and Bernhardt. ' Biologische Priifung von Ovarialpraparaten.'
Klin. Wochen., 4, 1925.
661. Zondek and Brahn. ' Uber Darstellung des Ovarialhormons in wassriger
Losung.' Klin. Wochen., 4, 1925.
INDEX OF AUTHORS
Adamberg, 95, 105.
Addis, 204.
Adler, 62, 84.
Aime, 1 1 .
Aitken, 40, 121.
Allan, 88, 102, 105, 112.
Allen, 4, 9, 44, 45, 62, 65, 67, 69, 90-92,
95, 103-114. 118-130, 132, 135, 136,
146.
Amsbaugh, 41.
Ancel, II, 31, 66, 123, 131, 139, 184,
189, 190, 192.
Aral, 150.
Aschheim, 86, 91, 106, 107-109, 112,
118, 119, 120, 122, 124, 126, 132, 136,
152, 158, 163, 164-170.
Aschner, 85, 129.
Asdell, 117, 124, 150, 191, 192, 196.
Ask-Upmark, 196.
Athias, 12, 76.
Aveling, 64.
Von Baer, 8.
Beard, 176.
Beigel, 12.
Bell, 72, 78, 138, 196.
Bellerby, 86, 88, 103, 105, 107, 108,
III, 112, 116, 118, 119, 120, 126, 133,
164, 179, 182, 183.
Bencan, 191.
Bergonie, 139.
Beuttner, 81, 83.
Biach, 72.
Biedl, 199.
Bischoff, 31, 34.
Blair, 202.
Blau, 107, 204.
Blotevogel, 97, 112.
Bonham, 85, 97, 202.
Borchardt, 86, 124, 130.
Bouin, II, 31, 66, 123, 131, 139, 184,
189, 190, 192.
Bourne, 203, 205.
Brahn, 87, 108.
Brambell, 2, 4, 18, 23, 45, 74, 80, 95,
100, 125, 137, 139, 199.
Brinkworth, 88.
Brouha, 86, 90, 95-97, 107-109, 113,
114, 116, 118, 120, 158, 163, 188, 202,
203, 205.
Brown-Sequard, 80.
Bucura, 74, 81, 83.
Bugbee, 86, 103, 104, 114, 121.
Burn, 92, loi, 104, 106, 137, 203, 205.
Burr, 92.
Camboulas, 80.
Carmichael, 73, 75, 150.
Cartland, 91, 180.
Castle, 98.
Ceresoli, 108.
Champy, 40, 85, 191.
Clark, 202.
Cole, 31.
Colgate, 91, 92, 95, 103, 106, 107, 113,
114, 122, 125, 136.
Corner, 5, 9, 41, 62, 65-67, 69, 81, 129,
178, 184, 185, 197, 202.
Courrier, 81, 86, 95, 108, 116, 117, 127,
129, 135-
Coward, 92, loi, 104, 106, 137.
Craig, 108, 116, 118, 119.
Crew, 2, 80, 150.
Curtis, 196.
Cushing, 152.
Daels, 196.
Darwin, 195.
Davenport, 99.
Davidson, 195.
Deanesly, 72, 152.
Dick, 196.
Dickens, 86, 88, 90, 102, 105, 107, 108,
III, 112, 114.
Dieckmann, 26, 66, 190.
Dierks, 65.
Dingemanse, 86, no, 130.
Dixon, 204.
Dodds, 86, 88, 90, 102, 105, 107, 108,
III, 112, 114.
Dohrn, 88, 105, no, 112, 130.
Doisy, 89-91, 95, 103-114, 120-122, 124,
125. 136.
Doncaster, 2.
235
236
INTERNAL SECRETIONS OF THE OVARY
Drips, 197.
Drummond-Robinson, 192, 196.
Duncan, 24.
Dupre, 129.
Van Dyke, 204.
Engle, 118, 160, 162, 168-171, 182.
Essen-Moller, 196.
Evans, 20, 31, 43, 50, 81, 92, 97, 127,
136, 150, 15-^, 153. i(>3, 164, 171, 185,
186.
Ewart, 40.
Faure, 86, 88, 91, 105, 130.
Fee, no.
Fellner, 84, 85, 107-109, no, 121, 127,
129-132, 138, 202.
Eels, 107-109, 116, 118, 121, 124, 163,
166.
Fielding, 100, 125, 139, 199.
Fingerhut, 84, 107, 120.
Foa, 149.
Fonda, 89, 107, 108.
Fraenkel, L., 11, 113, 114, 118, 120,
123, 183, 203, 211.
Fraenkel, S., 84, 89, 107, 108.
Francis, 91, 92, 95, 103, 106, 107, 108,
113, 114, 116, 118, 119, 122, 125,
136.
Frank, M. L., 85, 108.
Frank, R. T., 86, 90, 97, 99, 106-108,
no, 119, 122, 126, 129, 131, 185,
202.
Frei, 37.
Friedlaender, 31.
Frohlich, 152.
Gaines, 195.
Gasbarrini, 186.
Gatenby, 5, 9.
Gerlinger, 31, 66.
Gibson, 91, 92, 95, 103, 106, 107, 113,
114, 122, 125.
Gley, 180, 181.
Glimm, 87, 89, 105-107, 112.
Goldberger, 85, 86, 99, 108, 110.
Goldschmidt, 2, 80.
Gould, 107, 121, 179.
Gricouroff, 125.
Grigorieff, 76.
Grueter, 195.
Gsell-Busse, 97.
Gustavson, 85, 90, 97, 106-108, 119,
122, 126, 131, 202.
Guttmacher, 50, 72, 114, 202.
Haberlandt, 179, 198.
Halban, 74, 78.
Halberstadter, 138.
Hainan, 31, 190.
Hammond, 4, 9, 26, 37, 60, 66, 72, 123,
136, 146, 150, 151, 172, 177, 186, 189,
190, 191, 193, 196, 197.
Hancher, 204.
Hannan, 92.
Hanson, 10 1.
Harris, 197.
Hart, 86, 87, 90-92, 95, 102, 105, no,
III, 1 13, 1 19, 121, 203.
Hartman, 4, 7, 12, 29, 129, 130, 136,
197.
Hartmann, 89, 108.
Haterius, 100, 130.
Heape, 22, 40, 43, 62, 150.
Heinlein, 119, 120.
Hensen, 34.
Herrick, 196.
Herrmann, 84, 107, 108, 120, 121, 129,
179.
Van Herwerden, 62.
Hess, 177, 196.
Hesselberg, 26, 36, 130, 190, 191-193
195-
Heys, loi.
Hick, 196.
Hill, 5, 9, 28.
Hirsch, no.
Hisaw, 180, 181.
Hitschmann, 62.
H-ofmeier, 75.
Hohlweg, 88, 105, 119, 130.
Holloway, 85, 90, 108, 131.
Holzknecht, 139, 198.
Howe, 97.
Howitt, 88, 105, 1 12.
Hulles, 72.
Hurni, 178.
Hiissy, 139.
Hyndman, 85, 90, 108, 131.
Iscovesco, 84, 107.
Isler, 89, 108.
Jackson, 64, 67.
Jentzer, 81, 83.
Johnston, 91, 92, 95, 103, 105-108,
110-114, 122, 125, 136, 179.
Jolly, 31, 74, 76, 78, 83, 196.
dejongh, 86, 87, 90-92, 95, 102, 105,
109-111, 113, 117, 119, 121, 124, 127,
130, 203.
Jordan, 89, 90, 91, in, 113.
Kahnt, 103.
Keller, 31, 74, 191.
Kennedy, 178.
Keye, 202.
King. 65.
Kingery, 85, 119, 126.
Kirkham, 44.
INDEX OF AUTHORS
237
Kleinhaus, 196.
Knauer, 78.
Knaus, 202, 204.
Kohler, 179.
Kountz, 86, 91, 92, 95, 103, 106, 107,
113, 114, 118, 122, 125, 136.
Kraft, 204.
Krainz, 186.
Kreuger, 85, 90, 108, 131.
Kun, 119.
Kiipfer, 37.
Kuramitsu, 35.
Lacassagne, 125.
Lane-CIaypon, 12, 192.
Lange, 86, 91, 95, 97, 109, no.
Laqueur, 86, 87, 90-92, 95, 102, 109-
111, 113, 117, 119, 121, 124, 127, 130,
203.
Lataste, 43.
Lee, 23.
Leopold, 64, 67.
Levin, 96.
Lewis, 41.
Lillie, 133.
Limon, 78.
Lipschiitz, 12, 76, 86, 95, 105, 129, 149,
150, 151-
Loeb, 7, 26, 34, 35, 36, 79, 81, 86, 116,
118, iig, 123, 127, 130, 146, 176, 177,
178, 184, 186, 188, 190-195.
Loewe, 86, 87, 91, 95, 97, 108, 109, no,
112, 121, 129, 163, 170, 179.
Loewy, 72, 80.
Long, 20, 43, 50, 127, 136, 150, 164,
171, 185, 186.
Luciani, 4.
McIlroy, 196.
McXutt, 121.
Magian, 21.
Mahnert, 116, 118, 120, 126.
Marrian, 102-104, 106, no, 169.
Marshall, A. Milnes, 62.
Marshall, F. H. A., 8, 23, 31, 40, 41, 43,
60, 69, 73-76, 78, 83, 117, 123, 124,
130, 136, 137, 150, 152, 190, 193, 196,
204.
Masui, 72.
Mermod, 118, 169, 182.
Metzger, 37.
Meyer, 65, iSi.
Miller, 72.
Miura, 205.
Moore, 76.
Morau, 43.
Morris, 76.
Mulon, 196.
Murphey, 37, 121.
Myers, 26, 53, 130.
Neumann, 80.
Nielsen, 186.
Novak, 65.
OCHSNIER, 178.
O'Donoghue, 5, 11, 12, 28, 172, 184,
191.
Okinschitz, 84, 107.
Paas, 86, 97, 121, 163, 170.
Paladino, 12.
Papanicolaou, 4, 34, 65, 86, 107, 130,
179, 181.
Parkes, 20, 2^, 44, 45, 74, 86, 88, 95,
100-120, 125, 126, 133, 137-139, 143.
147. 150, 153. 154. 157. 162, 169, 176,
^77. 179, 182, 183, 186, 188, 191, 193,
197, 199, 202.
Payne, 91, 180.
Pearl, 178.
Peenan, 91, 180.
Pettinari, 76, 78.
Pfliiger, 5.
Phillips, 98.
Policard, 12.
Poll. 112.
Post, 50.
Pratt, 106, 108, III, 120.
Prenant, 176.
QUISNO, 43.
Rabl, 12.
Ralls, 89, 90, 91, 105, loS, 1 10, III,
112, 113.
Recamier, 139.
Regaud, 12.
Reichert, 34.
Reifferscheid, 139.
Rein, 34.
Retterer, 31.
Ribbert, 76, 78, 79, 189.
Richter, 72, 80, 114.
Riddle, 121.
Robertson, 91, 92, 95, 103, 106, 107
113, 114, 122, 125, 136.
Robinson, A., 136.
Robinson, M. R., 96, 108, 109.
Rosenbloom, 85, 108, 129.
Rubaschkin, 34.
Runciman, 137.
Sainmont, 12.
Salazar, 6.
Sand, 76.
Sandes, 28.
Schaeffer, 11.
Schenk, 196.
238
INTERNAL SECRETIONS OF THE OVARY
Schmaltz, 37, 196.
Schoeller, 88, 105, 130.
Schroder, 65, 124.
Schron, 12.
Schultz, 76.
Seaborn, 40, 85.
Seckinger, 97, 202.
Seitz, 66, 84, 107, 120.
Shaw, 9, 62, 65.
Siegmund, 116, 118, 120, 126, 163.
Simond, 86, 103, 104, 114, 121.
Simonnet, 86, 90, 95-97, 107-109, 113,
114, 116, 118-120, 131, 163, 202, 203,
205.
Simpson, 153.
Slonaker, 114, 119, 120.
Slotta, 89, 109.
Smith, B., 176.
Smith, H. P., 43.
Smith, M. G., 108, 109, 118, 182.
Smith, P. E., 152, 159, 160, 162, 166,
168-170.
Sobotta, 8, 43.
Sokoloff, 75.
Sonnenberg, 84.
Spohr, 86, no.
Starling, 192.
Stein, 121, 179.
Steinach, 12, 76, 88, 105, 119, 120, 125,
130, 139. 199-
Steinhaus, 64.
Stockard, 34, 150.
Van der Stricht, 8, 12, 31.
Struve, 41, 43.
Surface, 178.
Sutter, 26, 53.
Suzuki, 108.
Tamura, 72, 76.
Tandler, 74, 177.
Tange, 121.
Tause, 86.
Teel, 133, 186, 201.
Te Linde, 65.
Thayer, 150.
Trevan. 102.
Tribondeau, 139.
Trivino, 109.
Trufh, 119.
Tsu, 54.
Tuisk, 95, 105, 126.
Uhlmann, 93.
Vas, 80.
Veler, 106, 109, 112.
\"elich, 195.
Vesnjakov, 95, 105.
Villemin, 139.
Vincent, 204.
\'intemberger, 129, 131.
Voss, 34, 86, 97, 1 10, 121, 163, 170.
Wadehn, 87, 89, 90, 105, 106, 107, 112,
122.
Wahner, no.
Waldeyer, 5, 12.
Walker, 163.
Wallart, 139.
Wang, 72, 1 14.
Warren, 185.
Webster, 62.
Weichert, 181, 182, 188.
Wester, 196.
Weyerts, 85, 108.
Weymeersch, 196.
White, 85, 90, 108, 131.
Wiesner, 119, 120.
Wijsenbeek, 86, 87, 90, 105, iii, 203.
W^illiams, J. W., 177.
Williams, W. L., 12, 63.
De Winiwarter, 5.
Wintz, 66, 84, 85, 107, 120.
Wislocki, 202.
Wolbach, 97.
Wood, 137.
Woodman, 190, 191.
Wright, 86, 90, 102, 107, 108, in, 114.
Zeitschmann, 37.
Zondek, 86, 87, 91, 96, 106-109, 112,
118-120, 122, 124, 126, 136, 144, 148,
149, 152, 158, 163-170.
Zupp, 121.
INDEX OF SUBJECTS
Abortion, produced by oestrin, ii8;
produced by anterior pituitary im-
plants, i6g.
Accessory reproductive organs, 12 ;
atrophy at menopause, 20 ; effect of
ovariectomy on, 73 ; grafts of, 79 ;
relation with anterior pituitary body,
171.
Activity, variation in during oestrous
cycle, 114.
Adrenal gland, 72, 152.
Amenorrhoea, effect of oestrin on, 121 ;
in pituitary disorders, 152
Anoestrus, zz ; effect of oestrin during,
117 ; effect of anterior pituitary ex-
tracts during, i6g. {See also Oestrous
cycle.)
Anterior pituitary body, relation to
ovary, Chap. IX. ; luteinizing ex-
tract of, 153 ; effect of extract on
X-rayed ovary, 157 ; implantation
of, 158 ; production of ovulation,
159 ; comparison of extracts of, 163 ;
assay of extracts of, 165 ; properties
of extracts of, 166 ; distribution of
hormone of, 166 ; action of implants
on normal animal, 168.
Bat, corpus luteum of, 8 ; time of
ovulation in, 135.
Birds, effects of ovariectomy on, 70 ;
presence of oestrin in, 1 10 ; action of
oestrin on, 121.
Blood, oestrin in, 108 ; anterior
pituitary hormone in, 167.
Breeding season, definition, 22 ; in
various animals, 2^.
Carotene, 7.
Cercocebus, menstrual C3''cle of, 62.
Circulation, effect of oestrin on, 113.
Clitoris, comparative structure, 15.
Corpus albicans, 11.
Corpus luteum, formation of, 7 ; struc-
ture of in various mammals, 8-10 ;
of human, 65 ; extracts of, 84 ; pre-
sence of oestrin in, 107 ; relation to
occurrence of oestrus, 134 ; internal
secretion of, Chap. X. ; functions of,
173-175 ; specific variation in devel-
opment, 174 ; methods of removing,
175 ; inhibition of ovulation and
oestrus by, 176 ; effects of removal,
177, 196 ; effect on deciduoma pro-
duction, 186 ; control of mammary
gland, 191 ; functional relation to
interstitial tissue, 198 ; effect on
sensitivity of uterus to oxytocin, 204.
{See also Oestrus-inhibiting hormone.)
Corpus luteum atreticum, 10 ; result-
ing from X-rays, 140 ; caused by
anterior pituitary extracts, 154.
Corpus luteum spurium, 10.
Cow, corpus luteum of, 9 ; Fallopian
tube of, 13 ; vagina of, 14 ; mam-
mary gland of, 16 ; breeding season in,
23 ; oestrous cycle in, 37 ; period of
gestation, 68 ; age of puberty in, 68 ;
mammary gland during pregnancy
in, 192.
Dasvitrus, theca interna in, 5 ; corpus
luteum, 10 ; oestrous cycle in, 28,
68 ; period of gestation, 68 ; post-
oestrous changes in uterus, 184.
Deciduoma, 184 ; in various mammals,
185 ; method of producing, 186 ;
sensitization of uterus to, 186.
Didelphis. See Opossum.
Dioestrus, 24 ; effect of oestrin during,
117. {See also Oestrous cycle.)
Discus proligerus, 4.
Dog, Fallopian tube of, 13 ; breeding
season in, 23 ; oestrous cycle in, 31 ;
period of gestation, 68 ; causation of
oestrus in, 137 ; post-oestrous changes
in uterus, 184 ; deciduoma in, 186;
mammary gland during pseudo-
pregnancy in, 190.
Dysmenorrhoea, effect of oestrin on,
121.
239
240
INTERNAL SECRETIONS OF THE OVARY
Elephant, mammary gland of, 16.
Endocrine organs, effect of ovariectomy
on, 72.
Fallopian Tube, comparative struc-
ture, 13.
' Feminin,' see Oestrin.
Ferret, corpus luteum of, 10, 24 ;
vagina of, 14 ; oestrous cycle in, 26 ;
period of gestation, 68 ; history of
corpus luteum in, 174 ; post-oestrous
changes in uterus, 184.
Fertility, of various mammals, 68.
Fish, presence of oestrin in, no.
Follicular phase,' 26.
Folliculin,' see Oestrin.
' Generative Ferment,' 150.
Germinal epithelium, 4, proliferation
after X-irradiation, 140, 142.
Gestation, see Pregnancy.
Goat, breeding season in, 23 ; oestrous
cycle in, 68 ; mammary gland during
pregnancy in, 192.
Graafian follicle, structure of, 4 ; atre-
sia of, 6 ; maturation changes in, 6 ;
polyovular, 7 ; anovular, 7, 141 ;
effect of oestrin on, 119 ; relation to
oestrus, 134-148 ; growth of, in
mouse, 137 ; effect of X-rays on,
138-140 ; effects of anterior pituitary
extracts on, 154-163 ; growth during
pregnancy, 176.
Growth, effect of oestrin on, 114.
Guinea-pig, 8 ; vagina of, 15 ; mam-
mary gland of, 16 ; oestrous cycle
in, 34 ; period of gestation, 68 ;
function of corpus luteum in, 174;
post-oestrous changes in uterus, 184 ;
deciduoma in, 184.
Horse, oestrous cycle in, 40, 68 ;
period of gestation in, 68.
Hyperfeminization, 78 ; mammary
gland during, 199.
Hypophysectomy, effect on oestrous
cycle, 159.
Hypopituitarism, 152.
Intersexuality, 80.
Interstitial tissue, 11, 144 ; source of , 12;
effect of X-rays on, 139 ; functional
relation to corpus luteum, 198.
Lactation, effect on oestrous cycle, 39,
43, 44, 54, 68 ; effect of oestrin dur-
ing, 119, 183 ; inception of, 195.
Liquor folliculi, 4, weight of unit of
oestrin from, 105 ; oestrin content of ,
III, 135-
Luteal phase,' 26.
Macaciis rhesus, menstrual cycle of, 62 ;
time of ovulation in, 65 ; vaginal
cycle in, 65 ; period of gestation, 68 ;
pseudo-pregnancy in, 69.
Mammary gland, comparative struc-
ture of, 15 ; types of, 16 ; cyclic
changes in, 26, 28, 29, 32, 36, 39, 40,
5?)< 57, 61, 130, 189, 192 ; effect of
oestrin on, 129 ; development of by
corpus luteum, 175 ; development
of, 188 ; during pseudo-pregnancy,
189 ; and corpus luteum, 191 ; dur-
ing pregnancy, 191 ; cause of iinal
development, 192 ; in hyperfemini-
zation, 199.
Man, corpus luteum of, 9, 65 ; Fallo-
pian tube of, 13 ; uterus of, 14 ;
vagina of, 14 ; clitoris of, 15 ;
mammary gland of, 16 ; breeding
season in, 23 ; uterine cycle in, 62 ;
time of ovulation in, 65 ; mammary
cycle in, 66 ; period of gestation, 68 ;
pseudo-pregnancy in, 69.
Marsupials, oestrous cycle in, 28.
Membrana granulosa, 4, 5 ; effect of
X-rays, 139, 142.
' Menformon,' see Oestrin.
Menopause, 20 ; in human, 21 ; in
mouse, 22 ; effect of oestrin during,
121.
Menstrual cycle, of Primates, 61 ; in-
terpretation of, 66 ; after X-ray
sterilisation, 66.
Menstruation, 63, 66 ; relation of
oestrin to, 128.
Metabolism, effect of oestrin on, 113.
Metoestrus, 24. {See also Oestrous
cycle.)
Mouse, corpus luteum of, 8, 24 ; Fallo-
pian tube of, 13 ; uterus of, 14 ;
vagina of, 14 ; clitoris of, 15 ; mam-
mary gland of, 16 ; oestrous cycle in,
43 ; age of puberty in, 68 ; size of
litter in, 68 ; period of gestation, 68 ;
growth of Graafian follicle in, 137 ;
effects of anterior pituitary prepara-
tions, 154-163 ; function of corpus
luteum in, 174.
Oestrin, history of preparation of, 83 ;
preparation of, 84-88 ; chemical
properties of, 88-91 ; administration
of, 91-93 ; methods of testing, 93-98 ;
effect on rabbit uterus, 93, 126 ;
vaginal smear testing, 95 ; specificity
INDEX OF SUBJECTS
241
of vaginal test, 96, 97 ; effect on im-
mature animal, 97 ; effect on uterine
contractions, 97 ; standardization of,
101-106 ; unit of, 102 ; weight of
unit, 104, 105 ; distribution of, 106-
112 ; specificity of, 108 ; occurrence
of, in male, 109 ; yields of, 11 1 ; site
of origin, 113 ; pharmacological pro-
perties, 113, 114; effect on growth
and activity, 114; effect on test
animals, 115 ; effect during im-
maturity, 116, 124 ; effect during
anoestrus, 117 ; effect during dioes-
trus, 117 ; effect during pregnancy,
118; effect during lactation, 119;
effect on follicular maturation, 119 ;
clinical results with, 120 ; action on
male, 121 ; action on birds, 121 ;
limits of action, 1 21-123 ; ^^d copu-
lation, 125 ; relation to post-ovula-
tion changes, 1 25-131 ; effect on
vagina, 126 ; effect on uterus, 126 ;
relation to menstruation, 128 ; effect
on mammary gland, 129 ; signifi-
cance of distribution, 131-133 ; pres-
ence in liquor folliculi, 135 ; action
of during lactation, 183 ; effect on
sensitivity of uterus to oxytocin, 205.
Oestrous cycle, essential features of, 23;
types of , 25, Chap. I\'. ; comparative
changes during, 27 ; in Dasyuriis, 28;
in opossum, 29 ; in dog, 31 ; in
guinea-pig, 34 ; during lactation, 39,
43, 44, 54 ; in horse, 40 ; in sheep,
40 ; in pig, 41 ; in mouse, 43 ; in
rat, 43 ; in rabbit, 54 ; in ferret, 60 ;
nature of, in various mammals, 68 ;
mammary changes during, 130 ;
length after X-irradiation, 144 ; re-
gulation of, 146-148 ; effect of
thallium feeding, 148 ; length after
unilateral ovariectomy, 150.
Oestrus, 22 ; criteria of, 94 ; relation
of Graafian follicle to, 134 ; relation
to corpus luteum, 134; precocious,
158 ; periodicity of. Chap. VIII. ;
inhibition of, 176 ; relation to par-
turition, 200. {See also Oestrin.)
Oestrus-inhibiting hormone, prepara-
tion of, 179, 180 ; distribution of,
181 ; properties of, 181 ; assay of,
181. {See also Corpus luteum.)
Oestrus-producing hormone, see Oestrin.
Opossum, theca interna in, 5 ; oestrous
cycle in, 29 ; period of gestation, 68 ;
mammary gland, 130 ; source of
oestrin in, 136.
Ornithorhynchus, theca interna in, 5 ;
corpus luteum in, 9 ; development of
mammary glands of, 193.
Ovarian capsule, 13.
Ovarian cycle, in Dasyurus, 28 ; in
opossum, 30 ; in dog, 31 ; in guinea-
pig. 35 ; in cow, 37 ; in horse, 40 ;
in pig, 41 ; in mouse and rat, 45 ;
in rabbit, =f^ ; in man, 63 ; corre-
lation with parturition, 200 ; effect
on oxytocin production, 204.
Ovarian extracts, early work on, 80, 83.
Ovarian graft, histology of, 76 ; effects
of, 78; toanimalsof different age, 149.
Ovariectomy, effects of, 70 ; in birds,
70 ; effects on secondary sexual
characters, 72 ; effects on ductless
glands, 72 ; effects on accessory
organs, 73, 74 ; pregnancy after, 98 ;
effect during pregnancy, 196, 206.
Ovary, structure of, 4 ; accessory, 12 ;
regeneration of, 98 ; weight of unit
of oestrin from, 105 ; yield of oestrin
from, III ; grafts of, 149 ; hyper-
trophy of, 150 ; hypotrophy of, 130 ;
relation with other endocrine organs,
152 ; effects of anterior pituitary ex-
tracts on, 153-164 ; influence on
anterior pituitary body, 172. {See
also Ovarian cycle.)
Ovulation, changes preceding, in fol-
licle, 6 ; time of, in Primates, 65 ;
time of, in bat, 135 ; production of
by anterior pituitary extracts, 158^
inhibition of by corpus luteum, 175.
{See also Ovarian cycle.)
Ovum, structure of, 6 ; multinucleate,
7 ; effect of X-rays on, 139.
Oxytocin, role in parturition, 203 ;
effect of ovary on production of, 204 ;
changes in uterine sensitivity to, 205.
Papio, \-ulval cycle in, 65.
Para-lutein cells of corpus luteum, 9.
Parturition, Chap. XI. correlation with
ovarian cycle, 200 ; relation to oes-
trus, 200 ; inhibition of, 201 ; role
of oxytocin, 203 ; relation to effects
of ovariectomy, 206.
Pig, theca interna in, 5 ; corpus luteum
of, 9 ; breeding season in, 23 ; oes-
trous cycle in, 41 ; age of puberty in,
68 ; size of litter in, 68 ; period of
gestation, 68.
Pineal gland, 72.
Pituitary body, after ovariectomy, 72.
{See anterior pituitary body, and
oxytocin.)
Placenta, extracts of, 84 ; weight of
unit of oestrin from, 105 ; yields of
oestrin from, 112 ; presence of oes-
trin in, 132 ; anterior pituitary
hormone in, 167.
242
INTERNAL SECRETIONS OF THE OVARY
Placentoma, see Deciduoma.
Plants, presence of oestrin in, no.
Platypus, see Ornithovhyuchus.
Polar body, 6.
Precocious puberty,' induction by
oestrin, 124 ; induction of, by anterior
pituitary extracts, 158-163, 168.
IVegnancy, length of, in various mam-
mals, 68 ; after ovariectomy, 98 ;
effect of oestrin during, 118 ; mam-
mary gland during, 191 ; mainten-
ance of, 196.
Primates, mammary gland of, 16 ;
menstrual cycle of, 61 ; period of
gestation in, 68.
Prooestrus, 24. {See also Ocstrous
cycle.)
Pseudo-pregnancy, 25 ; in opossum,
30 ; in dog, 32 ; length of, in various
mammals, 68 ; in man, 69 ; in
Macacus rhesus, 66 ; mammary
gland during, 189. {See also Oes-
trous cycle.)
Puberty, 18 ; age of, in various mam-
mals, 68 ; precocious, 158.
' Puberty gland,' 125.
Rabbit, interstitial tissue of, 11 ;
uterus of, 14 ; mammary gland of,
16 ; oestrous cycle in, 26 ; age of
puberty in, 68 ; size of litter in, 68 ;
period of gestation, 68 ; effect of
oestrin on uterus, 126 ; effects of
anterior pituitary extracts on, 157 ;
history of corpus luteum in, 174 ;
post-oestrous changes in uterus, 184 ;
deciduoma in, 186 ; mammary gland
during pseudo-pregnancy in, 189.
Rat, Fallopian tube of, 13 ; uterus of,
14 ; vagina of, 14 ; mammary gland
of, 16 ; age of puberty, 20, 68 ;
ocstrous cycle in, 43 ; size of litter
in, 68 ; period of gestation, 68 ;
effects of anterior pituitary prepara-
tions, 154-163 ; function of corpus
luteum in, 174 ; deciduoma in, 185.
Respiration, effect of oestrin on, 113.
Rodents, breeding season in, 23.
Secondary Sexual Characters, 2 ;
effect of ovariectomy on, 72.
Semnopithecus eatelhts, menstrual cycle
of, 62.
Sexual differentiation. Chap. I.
Sexual periodicity. Chap. III.
' Sexual season,' 22.
Sheep, oestrous cycle in, 4 ; corpus
luteum of, 8 ; Fallopian tube of, 13 ;
breeding season in, 23 ; age of pu-
berty, 68 ; period of gestation, 68.
Sterility, effect of oestrin on, 121.
Superfoetation, 176.
* Super-ovulation,' production of, by
anterior pituitary implants, 169.
Testis, occurrence of oestrin in, 109 ;
effect of oestrin on, 121 ; effect of
testis lipoids on, 121.
Theca externa, 4.
Theca interna, 4, 5 ; oestrin in, 106 ;
effect of X-rays on, 139.
Theca-lutein cells of corpus luteum, 9.
Thymus, relation with ovary, 152.
Thyroid, 72 ; relation with ovary, 152.
Urine, weight of unit of oestrin from,
106 ; presence of oestrin in, 109 ;
yields of oestrin from, 109, 112 ;
anterior pituitary hormone in, 167.
Uterine cycle, in Dasyurus, 28 ; in
opossum, 29 ; in dog, 31 ; in guinea-
pig. 35 ; in cow, 39 ; in sheep, 41 ;
in pig, 42 ; in mouse and rat, 46 ;
in rabbit, 56 ; in ferret, 60 ; in
Primates, 62 ; in man, 63.
Uterus, comparative structure, 14 ;
effect of oestrin on, 126 ; sensiti-
zation of, 175 ; post-oestrous changes
in, 184 ; sensitization to deciduoma
formation, 186 ; cyclic variation in
contractility, 202 ; changes in sen-
sitivity to oxytocin, 204. {See also
Uterine cycle.)
Vagina, comparative structure, 14 ;
effect of oestrin on, 126. {See also
Vaginal cycle, and Vaginal smear.)
Vaginal cycle, 26 ; in opossum, 29 ;
in guinea-pig, 36 ; in cow, 39 ; in
inouse and rat, 47 ; in rabbit, 54 ;
in ferret, 60 ; in Macacus rhesus, 65.
Vaginal plug, in guinea-pig, 36 ; in rat
and mouse, 49.
Vaginal smear, limitations of, 26 ; in
opossum, 29 ; in guinea-pig, 36 ; in
mouse, 48 ; in Primates, 65 ; tech-
nique, 95 ; effect of vitamin A
deficiency on, 97.
Vitamin A, effect of deficiency on
vaginal cornification, 97.
Vulva, structure, 14 ; cycle of, in
Papio, 65.
X-RAYS, effect on menstrual cycle, 66 ,
histological effects on ovary, 138-143;
effects on oestrous cycle, 143-148 ;
unilateral sterilisation, 176.
Zona Pellucida, 6.
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BY ROBERT MACLEHOSE AND CO. LTD.
THE UNIVERSITY PRESS. GLASGOW.