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The distribution and source of oestrin in the pregnant mare.

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T H E DISTRIBUTION AND SOURCE O F OESTRIN I N
THE PREGNANT MARE
H. R. C A T C H P O L E A N D H. H. COLE
College o f Agriculture, University of California, Davis, California
Oestrin is present in mare serum between the one hundred
and fiftieth day of pregnancy and term in low concentration
(Cole and Hart, '30 and '30 a ) but extremely large amounts
are excreted in the urine (Zondek, '30); frequently the
hormonal content of the urine is ten times greater than that
of the pregnant woman. Glud et al. ( ' 3 3 ) have shown that
considerable amounts are likewise excreted in the feces. A
systematic study has been made by Cole and Saunders (unpublished) of the hormonal outputs of a number of individual
mares which have been available throughout pregnancy. They
have found that oestrin appears in the urine between the
fiftieth and one hundredth day of pregnancy. The range in
time of appearance of the hormone is undoubtedly less than
was indicated by Cole et al. ( '33). I n arriving at this earlier
estimate of a range extending from the sixtieth to the one
hundred and seventy-fifth day of pregnancy one case was
included in which, as was later disclosed, there was fetal
resorption and oestrin never appeared in the urine.
We have been interested in this hormone in relation to
other phenomena occurring in the mare in pregnancy. These
include : 1) the output of gonad-stimulating hormone during
the first half of pregnancy; 2) the hypertrophy of the interstitial cells of the gonads of the fetus and the finding in them
of fairly considerable amounts of female sex hormone; 3) the
behavior of the maternal ovaries which after regression both
of the primary corpus luteum of pregnancy, and also of the
secondary crop of corpora induced presumably by the gonad
335
336
H. R. C A T C H P O L E A N D H. H. COLE
stimulating hormone of the serum, become apparently quiescent during the last third of pregnancy. A brief review of
these findings has been reported by Cole et al. ( '33). The
fetal gonads reach their maximum size a t a fetal C. R. length
of 45 to 65 em., a time when the maternal ovaries appear
quiescent, and when the oestrin output in urine is high. This
tempted a suggestion that the fetal gonads might be one source
of the large amounts of hormone that are appearing in the
urine of the mare throughout this period.
Although aware from a parallel study of gonad-stimulating hormone of the limitations of this particular method in
dealing with questions of source of hormone, we were nevertheless sufficiently interested in the phenomenon of hormonal
distribution in this species to make a number of comparative
assays of various tissues for their oestrin content. These
data are of greater significance when correlated with data
which we have accumulated on the disappearance of oestrin
from mare and foal urine following parturition.
MATERIALS
Our source of materials was the same as described in a
preceding paper (Cole et al., '33). I n view of the labor involved in preparing extracts for the assay of oestrin, a very
limited series of tissues in which the fetal length varied from
28 em. to term were obtained. I t was, unfortunately, not
possible in all the cases to obtain a sample of urine from the
slaughtered animals. From the mares we took the ovaries
and samples of liver, kidney, endometrium and blood. From
the fetal side we took gonads, liver, kidneys, placenta and
blood. I n addition to the slaughter-house material we were
interested in studying the hormonal content of mare urine
preceding and following parturition. We also determined
the oestrin content of newborn foal urine. The placentas of
these mares, dropped at term, were also investigated f o r
oestrin content.
OESTRIN I N T H E PREGNANT MARE
337
METHODS
a. Preparation. o f extracts. The preparation of oily extracts containing the female sex hormone, oestrin, is possible
by a variety of methods relying essentially on the same
principle, namely, destruction of the tissue by rather drastic
chemical treatment, followed by extraction of the product
successively with various lipoid solvents. The method used
by Fevold, Hisaw and Leonard ('32) for the preparation of
progestin embodies these same principles. Allen and Meyer
('33) remark that the preparation of crude oestrin and crude
progestin extracts is carried out in exactly the same way, and
we had previously (Cole et al., '33) employed the rapid progestin method of Fevold for the preparation of oestrin from
fetal gonads. It was found convenient t o use this method in
the present study. An important feature of the method is
the initial boiling of the tissue with a fairly strongly acid
solution of 95 per cent ethyl alcohol. We have applied this
same process in the preparation of oestrin from mare urine
after initial concentration of the urine by evaporation, with
no indication that much hormone was being lost in the process,
and consider it entirely suitable for the rather rough assays
in the present instance. The method employed consisted in
the working up of usually 100 gm. tissue by the 'rapid' method
(Fevold et al., '32) a s described for progestin carrying the
process to the methyl alcohol stage for removal of most of
the cholesterol. The products were in every case thick brown
oils. The urine samples cited in tables 2 and 3 were not
treated chemically.
b. Assay of hormowe. The products were injected in three
doses at 8-hour intervals into 4-month-old spayed virgin
female rats. Vaginal smears were taken at 48,72 and 96 hours
after the first injection. The animals were used over again
a t intervals of 2 or 3 weeks, with the usual precaution of
initial smearing before injection. The doses corresponded
to a certain original tissue weight. After a preliminary experiment to determine the approximate concentration, the
rat unit was determined by halving the dose until the smallest
338
H. R. CATCHPOLE AND H. H. COLE
amount was found which would give a smear of nucleated
epithelial cells and cornified cells, or the latter alone, in at
least three out of four rats within 48 hours. I n the testing of
some extracts only two rats were used at each level and in
these cases the rat unit was designated as the minimal dose
at which both rats reacted.l I n the results, the weight of
original tissue corresponding to a rat unit will be stated.
The earlier paper (Cole et al., '33) deals with the oestrin
content of fetal ovaries, testes, livers and blood. Referred t o
original tissue weights, it was found that with the fetal gonads,
from 8 to 32 gm. were necessary to give a positive vaginal
reaction by the method of subcutaneous injection, although
by smearing the oils into the vaginae of the test animals as
little as 0.15 gm. was effective. This was explained by the
poor absorption of the injected preparations. It was found
that extracts made by the above procedure, whether intended
for oestrin or progestin were invariably strongly acid in
reaction. This acidity seems to be responsible for the skin
lesions so often reported after the use of these preparations.
If the extracts be properly neutralized, such lesions are never
produced, and absorption is good. This is reflected in a
decrease in the amounts of tissue required to produce a reaction. I n the case of fetal gonad extract, the minimal dose
fell to 1.5 to 3.0 gm. A discrepancy between the methods of
injection and vaginal smearing still exists, although to a less
degree. It must be attributed t o a localization of action in
the latter method whereby the hormone passes directly to its
site of action, without entering the general circulation, whence
it is probably rapidly excreted. I n the series reported in this
paper the extracts were injected subcutaneously in all
instances.
Conce&ration of oestrin in horse tissues
The activity of several maternal and fetal tissues from four
cases is shown in table 1. It may be seen that in very late
'Judging from the values given by Zondek ('31) f o r oestrin in mare urine,
our rat unit is equivalent to ten t o twenty of his mouse units.
339
OESTRIN I N T H E PREGNANT MARE
stages of pregnancy (case H12 with a fetus having a crownrump length of 90 em.) there is less oestrin present in all
tissues than in the cases taken a t earlier stages of pregnancy.
Oestrin is evidently distributed in both maternal and fetal
tissues. I t is clear that mare kidney, endometrium and fetal
placenta contain larger amounts than other tissues listed. The
high activity of the mare kidney accords with the high oestrin
content of the urine and must be due partly to included urine
and partly t o the presence of oestrin in the tubular or
TABLE 1
A s s a y of oestrin an mare and f e t a l tissues a t different stages of pregnancy
-
~~
.
~
~~
~~
..
..
-
~
.
.~
~
~~~
~-
I
E2
70
C A S E DESIGNATION
P X T A L C. R. LENGTH IN CENTIMETERS
,
I
H12
90
IMILLIGRAMS OF TISSUE NECESSARY TO EVOKE AN
OESTROUS REACTION I N T H E SPAYED
VIRGIN F E M A L E R A T
~
_.
hiarc urine
Mare kidney
1M are liver
Mare serum
Mare ovaries
~-
~
400
I
>3200
Endoinetriuin
Fetal placenta
Fetal gonads
Fetal liver
Fetal kidney
Fetal blood
800
3000
I
~
3200
3200
150
800
2000
>3200
1600
>3200
I
4000
>3200
500
1400
3400
1100
1700
>3200
1600
>3200
3200
>3200
>3200
- . -.
-
-
glomerular epithelium. It appears certain that the kidney is
removing oestrin selectively from the blood stream, since no
tissue yet investigated contains a concentration of oestrin
comparable with urine. The activity of the endometrium is
comparable to that of the mare kidney while that of the fetal
placenta is somewhat less. The high concentration of oestrin
in the endometrium and fetal placenta may be accounted for
in a number of ways: It is possible that oestrin is being
formed in one or both of these tissues. Evidence to be submitted later in this paper will have a bearing upon this assumption. O r it may be that oestrin is formed in the fetus
340
H. R. C A T C H P O L E A N D H. H. C O L E
and merely becomes concentrated in these tissues because
they are concerned with its excretion or transfer from the fetal
t o the maternal blood stream. I n another paper (Cole et al.,
'33) we suggested that possibly the fetal gonads were concerned with the production of oestrin. Although there is an
interesting correlation between the development of the fetal
gonads and the concentration of oestrin in the urine (the
maximum concentration of oestrin in the urine of the mare
occurs at the time when the fetal gonads reach their greatest
development, and the subsequent fall in the oestrin content of
the urine a t the time when the fetal gonads are regressing),
the evidence in this paper does not support such a theory. All
fetal tissues studied, gonads, livers and kidneys, contain
oestrin in about the same concentration and thus the presence
of oestrin in the fetal gonads loses the special significance
which we had previously attached t o it. Further, data t o be
given later on the disappearance of oestrin from foal urine
after parturition makes the assumption of oestrin production by the fetal gonads still less tenable. Finally, in explaining the high concentration of oestrin in endometrium and
fetal placenta, it might be assumed that oestrin is produced
by the maternal gonads and selectively stored or used by the
placental membranes. This is improbable for it has been
shown (Cole, Howell and Hart, '31) that the maternal ovaries
are in an apparently quiescent state during that phase of pregnaney when the oestrin content of the urine is greatest.
Further, we have shown here that the maternal ovaries contain relatively small amounts of oestrin. An experiment now
in progress involving the removal of the maternal gonads
during pregnancy should completely settle the question as to
their involvement in the oestrin secretion of the mare during
this period.
I n addition to the tissues listed in table 1we have extracted
and assayed three term placentas. In one an equivalent of
1600 mg. of fresh tissue and in another 3200 mg. represented
a rat unit; in the third 3200 mg. was negative.
341
OESTRIN IN T H E PREGNANT MARE
Oestrin in the urine
We have studied t,he oestrin content of the urine of four
mares for 2 months antepartum and shortly after this event
in the hope that it might throw additional light on the source
of the oestrin of pregnancy (table 2). Further, we have examined the urine of newborn foals. It may be seen that during the last 2 months of pregnancy there is a steady fall in
TABLE 2
Oestrin in mare urine preceding and following parturition
RAT UNITS PER LITER O F URINE I N THE FOLLOWING MARES
1
DAYS PRECEDINQ PARTURITION
c7
c9
c2
I
~
40-55
25-39
15-24
5-14
1-2
16000
8000
4000
4000
2000
8000
4000
4000
4000
1000
<zoo
Day of foaling
-I_____
16000
8000
~
I
I.
~
c3
16000
8000
~
1
4000
i
l--l
1
<loo
4000
'_ _ ~ _ _
TABLE 3
Oestrin in t h e urine of newborn foals
1
Day of foaling
1day after foaling
2 days after foaling
RAT UNITS O F OESTRIN PER LITTER O F URINE I N THE
FOLLOWING FOALS
c2 foal
-1
c3 foal
___2000
1
<loo
<~OO
the oestrin content of mare urine. I n two instances in which
samples were obtained 1 to 2 days before foaling it took Q to
1 cc. to produce a response in spayed rats. I n each of the
four cases cited in the table doses of 5 cc. of urine collected
after parturition were negative. This rapid disappearance
of oestrin from the urine of the mare is duplicated in the
foal (table 3 ) . I n two foals (c7 and c9) we were fortunate
in obtaining the first urine voided after birth. These samples
342
H. R. CATCHPOLE A N D H. H. COLE
contained oestrin whereas urine on subsequent days was
negative in the amounts administered. I n foal c2 the sample
of urine collected on the first day after parturition contained
oestrin. This urine was thick, dark brown, contained considerable mucus and had every appearance of the first voided
urine. It is quite likely that some foals do not urinate for
24 hours after birth for we have waited 12 to 14 hours t o
obtain the first voided urine in other instances. I t is significant that oestrin disappears from maternal and foal urine in
the course of a few hours following parturition. If either
the maternal or fetal gonads are concerned with the production of part or all of the oestrin excreted in the urine, their
activity is precipitously terminated by the expulsion of the
fetus and placenta. No corresponding morphological change
is apparent in the gonads at this time to account f o r such a
sudden change in physiological activity.
DISCUSSlON O F RESULTS
a. The placenta as an endocrine organ. Halban ( '05) suggested that the placenta takes over hormonal functions during
pregnancy, and a considerable number of investigators since,
relying frequently on merely finding hormone in this location,
have supported his theory. The most recent work on this
point has dealt with hormonal output following extirpation of
the ovaries during pregnancy. It has been known for many
years that pregnancy in woman will take a normal course after
bilateral ovariectomy. Waldstein ( '29) performed this operation on a woman in a case of bilateral dermoid cyst a t the
estimated thirty-fourth day of pregnancy. The pregnancy
was maintained and the birth was normal. The oestrin content of the blood in the eighth month was normal ; that of the
urine was lower than the usual very large amounts recorded
for normal pregnancies. This implies a considerable ovarian
contribution in normal pregnancy. I n the case of woman, it
may be remarked, there occurs no fibrous regression of the
ovary as in the mare. The hormonal content of the placenta
in Waldstein's case was normal, but even so contained only
OESTRIN IN T H E PREGNANT MARE
343
a fraction of the total daily excretion of hormone. I n this
case it is impossible to regard the hormone as coming out of
a placental reservoir, since ovariectomy was performed at
such an early age. Following birth, atrophy of the uterus
and, up to 7 months following birth, absence of menstruation
bore witness to the completeness of the operation. Waldstein
concludes that the placenta is to be regarded as a principal
source of the oestrous hormone-a conclusion concurred with
by Novak, J. Halban and 0. 0. Fellner. That the ovary, if
present, may continue to secrete hormone, and that it is necessary for furthering implantation during the first month of
gestation, is not denied by these investigators.
Szarka ('30) reports a similar case with bilateral ovariectomy at the third month. He found just previously to birth
1000 mouse units of oestrin in the urine as compared with a
normal of 5000 to 8000 M.U. This hormone might possibly
be proceeding from the fetus, which the observation of de
Snoo ('28) does not entirely rule out. The latter worker
finds excessive amounts of oestrin being excreted in a case of
chorioepithelioma of the tube, in which trophoblastic elements
alone are present. But the ovaries were also present, and the
extensive ovarian changes shown by Novak and Koff ( '30) to
occur in these conditions render this finding unreliable, not
only f o r the exclusion of fetal participation, but even for the
assumption of a chorionic role. Zondek ( '31) regards Waldstein's experiment as conclusive, and in the German literature,
rather more than in the English, the vicarious production of
oestrin, at least as far a s the placenta is concerned, seems to
be fully accepted.
b. Studies im the puerperiurn. Philipps' studies ( ' 3 0 ) show
oestrin to be present in the newborn up to 72 hours after birth.
Thereafter it diminishes and none is present after 6 days.
He speaks in favor of a placental origin, and considers that
since the conditions are the same in girls and boys a fetal
origin is excluded. I n the urine of post-pregnant women hormone disappears in the course of 8 days (Zondek, '31). Briihl
finds female sex hormone in the urines of newborn children
without distinction of sex up to the fourth day.
T H E ANATOMICAL RECORD, VOL. 5 9 , NO.
3
344
H. R. CATCHPOLE A N D H. H. COLE
The theory that the placenta is contributing to the large
amounts of hormone that are being produced in the pregnant
woman receives strong support from some of the foregoing
evidence. The actual amounts that are coming from the active
organs must wait on future pregnancy castration studies in
this and other forms.
c. Deductions from the present studies. Oestrin disappears
from the body of the mare even faster than it does in women,
and hormone has yet to be demonstrated in the postpartum
blood and urine of the mare. The actual daily production of
oestrin in the mare at 1 or 2 days antepartum-assuming a
daily production of 10 liters of urine and a rat unit in 0.5 to
1.00 cc.-is from 10,000 to 20,000 R.U. The day following
birth this figure has fallen t o one-fifth or less of its value, and
this abrupt failure in hormone production must be explained.
I n both woman and the mare the uniform concentration of
hormone in the blood is a striking feature. I n woman this
figure is not appreciably altered by castration, although the
excretion is reduced. I n both these forms there is an overproduction of hormone reflected in the excretion of the excess.
This over-production is specially marked in the mare, in which
the blood concentration of hormone is only a little higher than
in woman, whereas the excretion is ten times a s great.
I n the foal, all the hormone that is present in the body at
birth appears to pass completely into the urine during the
first day, and is excreted. Whatever its function in the mare
it is improbable that its presence in the fetus is more than
incidental. The findings with foal urine would appear to
rule out the fetal gonads as significant sources of the hormone
at this time. The morphological evolution of the fetal gonads
is not interrupted by the incident of birth, and were these
responsible for only a fraction of the amounts of hormone
found immediately antepartum it should be possible to recognize them in a state of hormonic activity beyond the point at
which we actually find hormone in the foal-namely at the
first urination, and at this point only. The fetal gonads in
later stages are much degenerated, and the interstitial cells
are vastly different from the healthy interstitial cell masses
OESTRIN IN T H E PREGNANT MARE
345
which are present at the time of maximal development; at this
latter time the possibility of their hormonic activity cannot
be denied. But the present demonstration of the approximate
equivalence of fetal body tissues in respect of hormonal content leave histological considerations as the sole support for
the theory of fetal gonadal participation in the active production of oestrin in the mare. The question of male sex hormone in the fetal gonads of the male was touched upon in the
earlier paper, but has not been further investigated. The
ovaries of the mare have been described as undergoing regression in the last third of pregnancy. During this time they
are found to be reduced in size, being tough and fibrous, with
no follicles grossly recognizable. Highly pigmented (orange
or red) flecks are often present in the stroma, representing
carotinoid remains of corpora. We would suggest that their
inactivity is absolute, as compared with the relative activity
of the human ovaries. This would explain the total failure
of oestrin in the mare following parturition.
There remains to be considered the placenta. The results
of the German investigators have shown that it possesses a
definite capacity for production of oestrin in the pregnant
castrated female. The case for placental production in the
mare is rendered strong by the above considerations that appear to rule out the ovary of the mare as a significant secretory organ in the latter part of pregnancy, and the fetal
gonad as an unlikely one.
The position of the maternal endometrium assumes in this
study an importance equal to that in the case of the gonadstimulating hormone ( Catchpole and Lyons, '34). The endometrium is the main absorber of the products of excretion of
the fetus and the allanto-chorion, and in the event that the
'placenta be regarded as an important source of oestrin in
this species, the activity of the maternal endometrium in the
two cases in which it was studied and found to be markedly
potent is immediately clarified, without recourse having to be
made to the very unlikely hypothesis that it possesses an internal secretory function of its own in respect of oestrin
production.
346
H. R . CATCHPOLE A N D H. H. COLE
SUMMARY AND CONCLUSIONS
1. A quantitative study has been made of the distribution
of oestrin in the tissues of four mares and their respective
fetuses. Hormone has been found in mare kidney, endometrium and ovary; in fetal gonads, liver and kidneys; in the
allanto-chorion, both during pregnancy and at term.
2. The amounts of oestrin in the urines of mares and foals
at parturition have been investigated.
3. These findings are discussed in relation to the histological pictures in the gonads of the fetus and the ovaries of
the mare over the period studied, and also in relation to the
excretion of oestrin in mare urine during pregnancy. Their
bearing on the source of the oestrous hormone of pregnancy
is discussed.
4. It is considered that the theory of placental formation of
hormone must be adopted in this form. A basis is thereby
given for the quantitative results.
L I T E R A T U R E CITED
ALLEN,W. M., AND R. K. MEYER 1933 The quantitative separation of progestin
from oestrin in extracts of the corpus luteurn. Am. J. Physiol.,
vol. 106, p. 53.
BRUHL, R. 1929 Das Vorkommen von weiblichen Sexualhormon und Hypophysenvorderlappenhormon im Blute und Urin von Neugeborenen.
Klin. Woch., Jahrg. 8, 8. 1766.
CATCHPOLE,
H. R., AND W. R. LYONS 1934 The gonad stimulating hormone of
pregnant mares. Am. J. Anat. ( I n press.)
COLE,11. H., AND G. H. HART 1930 The potency of blood serums of mares i n
progressive stages of pregnancy i n effecting the sexual maturity of
the immature rat. Am. J. Physiol., vol. 93, p. 57.
1 9 3 0 a Sex hormones i n the blood serum of mares. 11. The sera
of mares from the 222nd day of pregnancy t o the first heat period
post partum. Am. J. Physiol., vol. 94, p. 597.
COLE,11. H., C. E. HOWELL,
AND G. H. HART 1931 The changes occurring i n
the ovary of the mare during pregnancy. Anat. Rec., vol. 49, p. 199.
COLE,H. H., G. H. HART,W. R. LYONS,AND H. R. CATCHPOLE 1933 The development and hormonal content of fetal horse gonads. Anat. Rec., vol.
56, p. 275.
FEVOLD,
H. H., F. H. HISAW,AND S. L. LEONARD1932 Hormones of the corpus
luteum. Separation and purification of three active substances. J.
Am. Chem. Soc., vol. 54, p. 234.
OESTRIN I N T H E PREGNANT MARE
347
GLUD, P., K. PEDERSEN-RJERGAARD,
AND E. PORTMAN
1933 Uber Graviditatsreaktion bei der Stute. Endokrinologie, Jahrg. 13, S. 21.
HALBAN,
J. 1905 Die innere Secretion von Ovarium und Placenta, und ihre
Bedeutung f u r die Funktion der Milchdruse. Arch. f. Gynak., Jahrg.
75, s. 353.
NOVAK,E., AND A. K. KOFF 1930 The ovarian and pituitary changes associated
with hydatidiform mole and chorionepithelioma. Am. J. Obstet. and
Gynec., vol. 20, p. 481.
PHILIPP,
E. 1930 Hypophysenvorderlappen und Placenta. Zent. f . Gynak.,
Jahrg. 54, S. 450.
DE SNOO,E. 1928 Chorionepitheliom der Tube. Hormonbildung vom isolierten
Trophoblasten (Menformon). Zent. f. Gynak., Jahrg. 52, S. 2703.
SZARKA,
S. 1930 Hormcnale Verhaltnisse wahrend dcr Schwangerschaft bei
Fehlen der Eierstocke. Zent. f . Gynak., Jahrg. 54, S. 2211.
WALDSTEIN, E. 1929 Friihkastration in der Schwangerschaft. Zur Genese des
Ovarialhormons. Zent. f. Gynak., Jahrg. 53, S. 1305.
ZONDEK,B. 1931 Die Hormone des Ovariums und des Hypophysenvorderlappens.
Berlin. Verlag von Julius Springer.
1930 Hormonal Schwangerschaftsreaktion aus dem Harn bei Mensch
und Tier. Klin. Wchnschr., Jahrg. 9, S. 2285.
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