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Facilitation of ovulation by steroids in immature rats primed with pregnant mare serum gonadotropin
237
FACILITATION
OF OVULATION IN IMMATURE
PRIMED
WITH
Shao-Yao
PREGNANT
Ying2
BY STEROIDS
RATS
MARE
SERUM
and Roland
GONADOTROPIN'
K. Meyer
Department of Zoology University of Wisconsin Madison, Wisconsin 53706 Received: 3/21/72 ABSTRACT
Two mg of 2Oa- or 208-hydroxy-4-pregnen -3-one, deoxycorticosterone, corticosterone, 118-hydroxyprogesterone and 12a-hydroxyprogesterone3 administered at 10:00 AM at 24 days of age markedly increased the incidence of ovulation in immature rats given a single injection of 12 IU of pregnant mare serum gonadotropin (PMS) two days earlier. However, the same amount of Sa-pregnane-3, 11, 20-trione and l-2 mg of cortisone given at the same time did not increase the incidence of ovulation over that of the PMS treatment alone.
INTRODUCTION
Y'ing
and Meyer
of ovulation
(4) have
in 25-day-old of a single
immature
various
doses
maximum
at 3 and 30 IU while
did not induce of ovulation
ovulation.
in animals
was due to an increase
reported
injection
that the incidence
rats treated
of PMS reached
doses between
They postulated treated
with
in the amount
with a
10 and 20 IU that the failure
12 IU or 15 IU of of estrogen
PMS
secretion
STEROIDS
238
and thus a decrease This deficiency
Injection
of neuropharmacologic
agents
3 IU PMS
by these (7).
were undertaken by steroids with
of various
and chlorpromazine,
lation produced with
prior
deoxycorticosterone
gesterone,
other
such
such as pro-
the administration
as phenobarbital, the block
in immature a series
of ovu-
rats primed of experiments
the ovulation-inducing
than progesterone
by
to the critical
prevents
In this paper,
hormone.
can be corrected
steroids
after
agents
to determine
of ovulating
hormone
of progesterone
period (5,6).
atropine,
in the release
of ovulating
administration
20:3
in immature
ability
rats primed
12 IU PMS.
MATERIALS
AND METHODS
Twenty-one-day-old female rats weighing 45-50 g were obtained from the Badger Research Corp., Madison, Wisconsin and were caged in a light controlled (14 hr light and 10 hr room. Rockland rat chow darkness), air-conditioned(75-8OoF) and water were provided -ad lib. All animals were injected subcutaneously with 12 IU of Chicago) in 0.25 ml of PMS (Equinex, Ayerst Laboratories, physiological saline on Day 22 between 7:30 and 8:00 AM. Steroids except progesterone were suspended in 0.25 ml of methylcellulose (MC) and were injected subcutaneously at Progesterone was dissolved in 0.1 ml 10:00 AM on Day 24. Controls receiving vehicle only were included corn oil. The amounts of steroids are given in in each experiment. Tables l-3. Autopsies were performed between 8:00 AM and 12:OO noon on Day 25. The oviducts were removed and compressed The ova were counted under a between two glass slides. low power light microscope.
STEROIDS
Sept. 1972
239
RESULTS
PMS injected of progesterone ovulating (Table
on Day 22 followed
resulted
an average
PMS followed
in 75 to 100% of the animals
of 19 and 30 ova,
Ovulation
1).
by 2.0 or 0.5 mg
did not occur
by corn oil or MC.
respectively
in rats receiving
A mixture
or 2.0 mg 20a- and 208-hydroxy-4-pregnen and 2013-OH) was as effective 75 to 100% ovulation can be seen that whereas
When
Table
tive.
1.0 mg of 20a-OH
was observed
2 shows
desivatives, ducers;
average
that
and caused
was administered
in all animals.
llB-hydroxyprogesterone
appears
7 of 8 rats to shed
ovulation
unlike when
and its
also act as into be more
Two mg of 118-hydroxyprogesterone
Sa-pregnane-3,11,20-trione,
It
in 80% of the
or 20B-OH
llf3-hydroxyprogesterone
able to produce
(20c-OH
was not effective
and 12a-hydroxyprogesterone
on Day 24 caused
1.0
from 23 to 38 ova.
ovulation
2.0 mg of 20a-OH
ovulation
-3-one
as progesterone,
1.0 mg 20I3-OH induced
animals. alone,
with
of 0.5,
given
effec-
at 1O:OO AM
an average
of 11 ova.
progesterone,
was un-
2.0 mg was given
at 1O:OO AM
on Day 24. The data on facilitation are presented
in Table
3.
of ovulation
Deoxycorticosterone
1.0 or 2.0 mg,
and 2.0 mg corticosterone
in all animals
with
respectively.
Neither
ovulation,
0.5 mg aldosterone
shed
but
21 ova.
by corticoids
an average
at doses
induced
of
ovulation
of 50, 18, and 11 ova,
1.0 nor 2.0 mg of cortisone caused
induced
1 of 5 rats
to
STEROIDS
240
20:3
DISCUSSION McCormack
and Meyer
of progesterone when
injected
the compounds
we wanted
to compare
dissolved
in a solution
data obtained,
potent
steroids
of MC.
corticosterone; (d) two methyl
activity
derivatives
of quanti-
of the most
can be arranged
as follows:
corticosterone;
tested,
to correlate
(c) 20a-OH;
C-4,5-double
terone
are important with
bond
of the small
group
ring D eliminates
their
ovulation-
number
of com-
group
C-3-keto
group
of proges-
of ovulation.
attached
at ring D,
did not induce
to 12 IU PMS-treated
that a hydroxy
struc-
can be only tentative.
for facilitation
17- or 16a-hydroxyprogesterone, administered
with
and C-20 keto
an hydroxy
17-acetate).
the molecular
that the characteristic
group,
Corticoids
steroids
the interpretations
It may be concluded
deoxy-
(2a-methyl-llB-hydroxypro-
Because
capacity.
in
(a) progesterone,
and ba-methyl-17-hydroxyprogesterone
inducing
were
of 118-hydroxyprogesterone
ture of the progestational
dicates
some of
ll$-hydroxyprogesterone,
(b) 208-OH,
It was of interest
when
potency
ovulation
and 12a-hydroxyprogesterone
pounds
effects
with progesterone
On the basis
the relative
of 20a-OH:208-OH,
gesterone
similar
in corn oil, all compounds
in causing
of decreasing
mixture
that suspensions
at 1O:OO AM on Day 24, and since
tative
terms
shown
in MC or corn oil have
w.ere not easily suspended
(6) have
attached
the ovulation-inducing
rats.
ovulation This in-
at C-16 or C-17 of activity.
STEROIDS
Sept. 1972
McCormack
and Meyer
progesterone
(6) showed
acetate
acts as an inducer
241
that
6a-methyl-17-hydroxy-
(MAP or medroxyprogesterone)
of ovulation,
approximately
that of progesterone.
Another
17-acetoxyprogesterone
also facilitates
efficiently that
as progesterone
the acetylation
ring D probably ovulation.
would
compound
(8).
These
at the hydroxy restore
also equal
to
6-chloro-6-dehydroovulation results
group
the ability
as indicate
attached
to
to facilitate
242
20:3
STEROIDS
Table
1
Facilitation of ovulation by progesterone, 2Ou-OH or 20B-OH in immature rats pretreated with 12 IIJ PMS
Xv.
No. Ova/ Ovul. Rat + S.E.
Treatment
No. Rats Ovul. (%>
P?/ISControl
o/9
(
Progesterone* 0.5 mg
4/4
(100)
64.3
rl 3.7
30.2 t 2.5
2.0 mg
7/s
( 88)
51.6 + 4.5
19.0 + 5.0
ZOa-hydroxy4-pregnen3-one 1.0 mg
l/5
( 20)
50.9 f 5.3
2.0 mg
4/4
(100)
49.3 f 1.1
17.5 f 4.5
20B-hydroxy4-pregnen 3-one 1.0 mg
4/5
( 80)
64.8 t 6.1
34.0 k10.9
2.0
mg
4/4
(100)
63.0 + 2.4
32.9 + 3.8
20 (cX+S)hydroxy-4pregnen-3one 0.5 mg
4/4
(100)
54.2 + 4.5
22.8 + 4.1
1.0 mg
3/4
( 75)
45.8 k 4.1
34.3 + 4.9
2.0 mg
4/4
( 75)
60.0 + 3.8
38.0 + 1.8
_.___. *
___
_ ~.
_.
0)
Av. Ov. Wt. (mg) ? S.E.
52.8 i 6.1
_
The other steroids Progesterone was dissolved in corn oil. were suspended in methylcellulose. Steroids were given subcutaneously at 10:00 A?[, Day 24.
STEROIDS
Sept. 1972
Table
243
2
Facilitation of ovulation steroids in 12 IU PYS-treated
by various immature rats
Av. Ov. Wt. (mg) k S.E.
Av. No. Ova/ Ovul. Rat t S.E.
Treatment
No . Rats Ovul. (%)
PMS Control
O/l0
c
0)
52.3 I!Z5.9
__
2-methyl-llf3hydroxyprogesterone 2.0 mg
7110
(703
52.3 ir 4.1
31.1 + 6.1
1.0 mg
5/10
(50)
46.3 k 2.1
28.0 k 6.1
6e-methyl-118hydroxyprogesterone 2.0 mg
7110
(70)
50.4
L!z4.1
29.6 t 4.2
llB-hydroxy-4pregnene-3,20dione 2.0 mg
7/8
(88)
43.9
+ 2.6
11.3 -+ 4.6
12a-hydroxyprogesterone 2.0 mg
5/7
(71)
65.3 rt 2.7
32.8 * 7.1
5a-pregnane-3,l.l, 20-trione 2.0 mg
O/l0
( 0)
48.0 F 4.8
16a-h,ydroxy-4pregnene-3,20dione 2.0 mg
O/8
17-hydroxy-4pregnene-3,20 'dione 2.0 mg Steroids
were
injected
(
01
t 3.1
-_
44.6 It 3.9
--
46.3
subcutaneously
at lo:00 AV, Day 24.
STEROIDS
244
Table Facilitation
203
3
of ovulation
by corticoids
Treatment
No. Rats Ovul. (%)
Av. Ov. Wt. (mg) F S.E.
Av. No. Ova/ Ovul. Rat t S.E.
PMS Control
O/15
(
47.8 t 3.6
__
Deoxycorticosterone 2.0 mg
8/8
(100)
54.4
1.0 mg
8/8
(100)
56.3 _+ 5.5
49.8
Corticosterone 2.0 mg
7/7
(100)
50.1 + 2.1
10.7 + 2.9
Cortisone 2.0 mg
O/5
(
0)
66.4 i 4.9
__
1.0 mg
O/5
(
0)
54.4 ? 6.4
-_
Aldosterone 0.5 mg
l/5
(
25)
46.5 5 2.2
0)
i: 2.9
17.9 + 3.8
21.0
+ 3.1
STEROIDS
Sept. 1972
245
REFERENCES
1.
This study was supported by The Ford Foundation Grant No . 63-505 and by PHS Training Grant No. 1 TO1 HD 00104-04 of the National Institute of Child Health and Human Development.
2.
Present Address: Laboratory of Human Reproduction and Reproductive Biology and Department of Anatomy, Harvard 02115. Medical School, Boston, Massachusetts
3.
IIJPAC names
of compounds
referred
to:
-3-one 20e-OH: 20a-hydroxy-4-pregnen -3-one 208-OH: 20B-hydroxy-4-pregnen 118-hydroxyprogesterone: llB-hydroxy-4-pregnene3,20-dione 2a-methyl-ll8-hydroxyprogesterone:2a-methyl-ll8-hydroxy4-pregnene-3,20-dione 6amethyl-llR-hydroxyprogestesone:6a-methyl-ll8-hydroxy4-pregnene-3,20-dione 12e-hydroxyprogesterone: 12a-hydroxy-4-pregnene-3,20-dione 17-hydroxyprogesterone: 17-hydroxy-4-pregnene-3,20-dione 16a-hydroxyprogesterone: 16a-hydroxy-4-pregnene-3,20-dione
4.
Ying, S.Y. and Meyer, MED. 130, 40 (1969).
5.
6.
R.K.,
PROC.
SOC. EXPTL.
BIOL.
McCormack, C.E., 3,300 (1963).
and Meyer,
R.K.,
GEN. COMP.
ENDOCR.
McCormack, C-E., (1965).
and Meyer,
R.K.,
FERTIL.
STERIL.
-16,384
7.
Ying,
8.
J. Harting,
S.Y.
and Meyer, personal
R.K.,
ENDOCRINOLOGY
communication.
-84:1466
(1969)
FACILITATION
OF OVULATION IN IMMATURE
PRIMED
WITH
Shao-Yao
PREGNANT
Ying2
BY STEROIDS
RATS
MARE
SERUM
and Roland
GONADOTROPIN'
K. Meyer
Department of Zoology University of Wisconsin Madison, Wisconsin 53706 Received: 3/21/72 ABSTRACT
Two mg of 2Oa- or 208-hydroxy-4-pregnen -3-one, deoxycorticosterone, corticosterone, 118-hydroxyprogesterone and 12a-hydroxyprogesterone3 administered at 10:00 AM at 24 days of age markedly increased the incidence of ovulation in immature rats given a single injection of 12 IU of pregnant mare serum gonadotropin (PMS) two days earlier. However, the same amount of Sa-pregnane-3, 11, 20-trione and l-2 mg of cortisone given at the same time did not increase the incidence of ovulation over that of the PMS treatment alone.
INTRODUCTION
Y'ing
and Meyer
of ovulation
(4) have
in 25-day-old of a single
immature
various
doses
maximum
at 3 and 30 IU while
did not induce of ovulation
ovulation.
in animals
was due to an increase
reported
injection
that the incidence
rats treated
of PMS reached
doses between
They postulated treated
with
in the amount
with a
10 and 20 IU that the failure
12 IU or 15 IU of of estrogen
PMS
secretion
STEROIDS
238
and thus a decrease This deficiency
Injection
of neuropharmacologic
agents
3 IU PMS
by these (7).
were undertaken by steroids with
of various
and chlorpromazine,
lation produced with
prior
deoxycorticosterone
gesterone,
other
such
such as pro-
the administration
as phenobarbital, the block
in immature a series
of ovu-
rats primed of experiments
the ovulation-inducing
than progesterone
by
to the critical
prevents
In this paper,
hormone.
can be corrected
steroids
after
agents
to determine
of ovulating
hormone
of progesterone
period (5,6).
atropine,
in the release
of ovulating
administration
20:3
in immature
ability
rats primed
12 IU PMS.
MATERIALS
AND METHODS
Twenty-one-day-old female rats weighing 45-50 g were obtained from the Badger Research Corp., Madison, Wisconsin and were caged in a light controlled (14 hr light and 10 hr room. Rockland rat chow darkness), air-conditioned(75-8OoF) and water were provided -ad lib. All animals were injected subcutaneously with 12 IU of Chicago) in 0.25 ml of PMS (Equinex, Ayerst Laboratories, physiological saline on Day 22 between 7:30 and 8:00 AM. Steroids except progesterone were suspended in 0.25 ml of methylcellulose (MC) and were injected subcutaneously at Progesterone was dissolved in 0.1 ml 10:00 AM on Day 24. Controls receiving vehicle only were included corn oil. The amounts of steroids are given in in each experiment. Tables l-3. Autopsies were performed between 8:00 AM and 12:OO noon on Day 25. The oviducts were removed and compressed The ova were counted under a between two glass slides. low power light microscope.
STEROIDS
Sept. 1972
239
RESULTS
PMS injected of progesterone ovulating (Table
on Day 22 followed
resulted
an average
PMS followed
in 75 to 100% of the animals
of 19 and 30 ova,
Ovulation
1).
by 2.0 or 0.5 mg
did not occur
by corn oil or MC.
respectively
in rats receiving
A mixture
or 2.0 mg 20a- and 208-hydroxy-4-pregnen and 2013-OH) was as effective 75 to 100% ovulation can be seen that whereas
When
Table
tive.
1.0 mg of 20a-OH
was observed
2 shows
desivatives, ducers;
average
that
and caused
was administered
in all animals.
llB-hydroxyprogesterone
appears
7 of 8 rats to shed
ovulation
unlike when
and its
also act as into be more
Two mg of 118-hydroxyprogesterone
Sa-pregnane-3,11,20-trione,
It
in 80% of the
or 20B-OH
llf3-hydroxyprogesterone
able to produce
(20c-OH
was not effective
and 12a-hydroxyprogesterone
on Day 24 caused
1.0
from 23 to 38 ova.
ovulation
2.0 mg of 20a-OH
ovulation
-3-one
as progesterone,
1.0 mg 20I3-OH induced
animals. alone,
with
of 0.5,
given
effec-
at 1O:OO AM
an average
of 11 ova.
progesterone,
was un-
2.0 mg was given
at 1O:OO AM
on Day 24. The data on facilitation are presented
in Table
3.
of ovulation
Deoxycorticosterone
1.0 or 2.0 mg,
and 2.0 mg corticosterone
in all animals
with
respectively.
Neither
ovulation,
0.5 mg aldosterone
shed
but
21 ova.
by corticoids
an average
at doses
induced
of
ovulation
of 50, 18, and 11 ova,
1.0 nor 2.0 mg of cortisone caused
induced
1 of 5 rats
to
STEROIDS
240
20:3
DISCUSSION McCormack
and Meyer
of progesterone when
injected
the compounds
we wanted
to compare
dissolved
in a solution
data obtained,
potent
steroids
of MC.
corticosterone; (d) two methyl
activity
derivatives
of quanti-
of the most
can be arranged
as follows:
corticosterone;
tested,
to correlate
(c) 20a-OH;
C-4,5-double
terone
are important with
bond
of the small
group
ring D eliminates
their
ovulation-
number
of com-
group
C-3-keto
group
of proges-
of ovulation.
attached
at ring D,
did not induce
to 12 IU PMS-treated
that a hydroxy
struc-
can be only tentative.
for facilitation
17- or 16a-hydroxyprogesterone, administered
with
and C-20 keto
an hydroxy
17-acetate).
the molecular
that the characteristic
group,
Corticoids
steroids
the interpretations
It may be concluded
deoxy-
(2a-methyl-llB-hydroxypro-
Because
capacity.
in
(a) progesterone,
and ba-methyl-17-hydroxyprogesterone
inducing
were
of 118-hydroxyprogesterone
ture of the progestational
dicates
some of
ll$-hydroxyprogesterone,
(b) 208-OH,
It was of interest
when
potency
ovulation
and 12a-hydroxyprogesterone
pounds
effects
with progesterone
On the basis
the relative
of 20a-OH:208-OH,
gesterone
similar
in corn oil, all compounds
in causing
of decreasing
mixture
that suspensions
at 1O:OO AM on Day 24, and since
tative
terms
shown
in MC or corn oil have
w.ere not easily suspended
(6) have
attached
the ovulation-inducing
rats.
ovulation This in-
at C-16 or C-17 of activity.
STEROIDS
Sept. 1972
McCormack
and Meyer
progesterone
(6) showed
acetate
acts as an inducer
241
that
6a-methyl-17-hydroxy-
(MAP or medroxyprogesterone)
of ovulation,
approximately
that of progesterone.
Another
17-acetoxyprogesterone
also facilitates
efficiently that
as progesterone
the acetylation
ring D probably ovulation.
would
compound
(8).
These
at the hydroxy restore
also equal
to
6-chloro-6-dehydroovulation results
group
the ability
as indicate
attached
to
to facilitate
242
20:3
STEROIDS
Table
1
Facilitation of ovulation by progesterone, 2Ou-OH or 20B-OH in immature rats pretreated with 12 IIJ PMS
Xv.
No. Ova/ Ovul. Rat + S.E.
Treatment
No. Rats Ovul. (%>
P?/ISControl
o/9
(
Progesterone* 0.5 mg
4/4
(100)
64.3
rl 3.7
30.2 t 2.5
2.0 mg
7/s
( 88)
51.6 + 4.5
19.0 + 5.0
ZOa-hydroxy4-pregnen3-one 1.0 mg
l/5
( 20)
50.9 f 5.3
2.0 mg
4/4
(100)
49.3 f 1.1
17.5 f 4.5
20B-hydroxy4-pregnen 3-one 1.0 mg
4/5
( 80)
64.8 t 6.1
34.0 k10.9
2.0
mg
4/4
(100)
63.0 + 2.4
32.9 + 3.8
20 (cX+S)hydroxy-4pregnen-3one 0.5 mg
4/4
(100)
54.2 + 4.5
22.8 + 4.1
1.0 mg
3/4
( 75)
45.8 k 4.1
34.3 + 4.9
2.0 mg
4/4
( 75)
60.0 + 3.8
38.0 + 1.8
_.___. *
___
_ ~.
_.
0)
Av. Ov. Wt. (mg) ? S.E.
52.8 i 6.1
_
The other steroids Progesterone was dissolved in corn oil. were suspended in methylcellulose. Steroids were given subcutaneously at 10:00 A?[, Day 24.
STEROIDS
Sept. 1972
Table
243
2
Facilitation of ovulation steroids in 12 IU PYS-treated
by various immature rats
Av. Ov. Wt. (mg) k S.E.
Av. No. Ova/ Ovul. Rat t S.E.
Treatment
No . Rats Ovul. (%)
PMS Control
O/l0
c
0)
52.3 I!Z5.9
__
2-methyl-llf3hydroxyprogesterone 2.0 mg
7110
(703
52.3 ir 4.1
31.1 + 6.1
1.0 mg
5/10
(50)
46.3 k 2.1
28.0 k 6.1
6e-methyl-118hydroxyprogesterone 2.0 mg
7110
(70)
50.4
L!z4.1
29.6 t 4.2
llB-hydroxy-4pregnene-3,20dione 2.0 mg
7/8
(88)
43.9
+ 2.6
11.3 -+ 4.6
12a-hydroxyprogesterone 2.0 mg
5/7
(71)
65.3 rt 2.7
32.8 * 7.1
5a-pregnane-3,l.l, 20-trione 2.0 mg
O/l0
( 0)
48.0 F 4.8
16a-h,ydroxy-4pregnene-3,20dione 2.0 mg
O/8
17-hydroxy-4pregnene-3,20 'dione 2.0 mg Steroids
were
injected
(
01
t 3.1
-_
44.6 It 3.9
--
46.3
subcutaneously
at lo:00 AV, Day 24.
STEROIDS
244
Table Facilitation
203
3
of ovulation
by corticoids
Treatment
No. Rats Ovul. (%)
Av. Ov. Wt. (mg) F S.E.
Av. No. Ova/ Ovul. Rat t S.E.
PMS Control
O/15
(
47.8 t 3.6
__
Deoxycorticosterone 2.0 mg
8/8
(100)
54.4
1.0 mg
8/8
(100)
56.3 _+ 5.5
49.8
Corticosterone 2.0 mg
7/7
(100)
50.1 + 2.1
10.7 + 2.9
Cortisone 2.0 mg
O/5
(
0)
66.4 i 4.9
__
1.0 mg
O/5
(
0)
54.4 ? 6.4
-_
Aldosterone 0.5 mg
l/5
(
25)
46.5 5 2.2
0)
i: 2.9
17.9 + 3.8
21.0
+ 3.1
STEROIDS
Sept. 1972
245
REFERENCES
1.
This study was supported by The Ford Foundation Grant No . 63-505 and by PHS Training Grant No. 1 TO1 HD 00104-04 of the National Institute of Child Health and Human Development.
2.
Present Address: Laboratory of Human Reproduction and Reproductive Biology and Department of Anatomy, Harvard 02115. Medical School, Boston, Massachusetts
3.
IIJPAC names
of compounds
referred
to:
-3-one 20e-OH: 20a-hydroxy-4-pregnen -3-one 208-OH: 20B-hydroxy-4-pregnen 118-hydroxyprogesterone: llB-hydroxy-4-pregnene3,20-dione 2a-methyl-ll8-hydroxyprogesterone:2a-methyl-ll8-hydroxy4-pregnene-3,20-dione 6amethyl-llR-hydroxyprogestesone:6a-methyl-ll8-hydroxy4-pregnene-3,20-dione 12e-hydroxyprogesterone: 12a-hydroxy-4-pregnene-3,20-dione 17-hydroxyprogesterone: 17-hydroxy-4-pregnene-3,20-dione 16a-hydroxyprogesterone: 16a-hydroxy-4-pregnene-3,20-dione
4.
Ying, S.Y. and Meyer, MED. 130, 40 (1969).
5.
6.
R.K.,
PROC.
SOC. EXPTL.
BIOL.
McCormack, C.E., 3,300 (1963).
and Meyer,
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