BIOLOGY
OF REPRODUCTION
Pulsatile
18, 359 -364
Secretion
(1978)
of LH and Ovarian
Follicular
Estradiol
Phase of the Sheep DAVID
MRC
Unit
Department University
of
Estrous
During
the
Cycle
T. BAIRD Reproductive
Obstetrics of Edinburgh,
Biology,
and Gynaecology, 23 Chalmers Street,
of
Edinburgh ABSTRACT The concentration of LH in jugular venous plasma and the ovarian secretion of estradiol was measured every 10 mm for periods of 60 or 120 mm on 28 occasions in 13 ewes with utero-ovarian or ovarian autotransplants. Blood samples were collected at various times during the follicular phase; 2 days prior to esifus (estruS -2), estrus -1 and on the day of estrus and the results compared to those obtained during the luteal phase. During the luteal phase, episodic pulses of LH occurred at intervals of approximately 3 h 20 mm and each pulse was followed by a significant rise in the secretion of estradiol. During the follicular phase, the pulses of LH occurred with increased frequency (1/h) but with decreased amplitude. Although each pulse of LH was smaller, the peak of secretion of estradiol following each pulse was significantly greater during the follicular than during the luteal phase. By estrus, the secretion of estradiol declined in spite of LH values in the range of the preovulatory LH surge. It is suggested that the increased secretion of estradiol from the
preovulatory Graaflan follicle with increasing frequency.
is due to repeated
stimulation
INTRODUCTION
We
between the pituitary
from lated
maturing amounts
Cox Baird
1971; 1976b;
factors responsible stage of follicular are not
known.
estrogen
Barcikowski Karsch et for growth
stimulating and steroid
During
section,
this
the
period
1974;
Karsch
et al.,
1977).
of
If the
FSH
Salamonwhile the (Akbar et rise in LH
by maintaining lutefollicular development
is
that
inhibited
suggesting related
Accepted Received
(Baird
and
the
2 events
Scaramuzzi,
the
developing episodic
reported cycle,
that estrogen
Graafian pulses of
occur
during
the
secretion
follicles is stimuLH which occur
from the ovary of the phase of the cycle. AND
sheep
during
the
METHODS
Animals
Eight ewes with ovarian (Coding et al., 1967) and 5 ewes with utero-ovarian autotransplants (Harrison et al., 1968) were allocated to this experiment during the breeding season (October-February). The ewes were housed under natural lighting conditions in a covered pen at Dryden Field Station, Roshin, Midlothian, Scotland. The left ovary alone, or together with the oviduct, horn and body of the uterus, cervix and anterior vagina, had been autotransplanted to the neck at least 12 months previously. The left ovarian and uterine arteries and uteroovarian vein were anastomosed to the carotid artery and jugular vein which were exteriorized in separate skin tubes (Baird et al., 1976c). The right ovary and uterine horn were removed. The ewes with utero-ovarian transplants had regular estrous cycles (mean 17.8 ± 0.2 days) in contrast to those with ovarian transplants which failed to show heat due to persistence of the corpus luteum because of the absence of an adjacent uterine horn
of increased
concentration is prevented al levels of progesterone, causally
Experimental
this final secretion
concentration
of
MATERIALS
et al., 1974; al., 1977). The
actually falls (L’Hermite et al., 1972; sen et al., 1973; Pant et al., 1977) basal concentration of LH rises 4-fold al.,
estradiol follicular
Graafian follicle secretes increasof estradiol (Moore et al., 1969;
et al., et al.,
previously
phase
the by
pulses of LH which
about every 3 h (Baird et al., 1976a). The present study was initiated to determine the relationship between the frequency and amplitude of pulses of LH and the secretion of
secretion of gonadoand steroids from
the ovary is not clear. As the secretion of progesterone from the regressing corpus luteum declines on Day 14 or 15 (estrus -3 and -2), the ing
have
luteal
Although the endocrine events occurring during the periovulatory period in the sheep have been described in a number of studies, the exact relationship trophins from
by episodic
are
1976a).
September 9, 1977. July 11, 1977.
359
BAIRD
360 et
(Baird
al.,
1976c).
The
ewes
were
either
pure
Tasmanian
Merino or crossed with Finnish Landrace (F X M) or Scottish Blackface (BF X M). Heat was detected by testing with a vasectomized ram at least
and, on some occasions, more frequently. of the absence of cyclical ovarian funcewes with ovarian autotransplants were studied following premature luteal regression induced by i.m. injection (100 g) of a potent analogue of prostaglandin-F, a, ICI 80996 (cloprostenol) on Day 10. Luteal regression and estrus occurred Within 72 h (Baird and Scaramuzzi, 1975).
one in ovarian venous plasma, and hematocrit. No allowance was made for the volume of plasma trapped in the packed red cells, nor for the small amount of steroids (about 1096) adsorbed on the red cell surface.
twice/day
RESULTS
Because tion, the
Estrus
occurred
7 days of occasions autotransplants
were
occurred cloprostenol.
within
estrus Experimental
Design
On the estrus, both
of an experiment, after testing for jugular veins were cannulated as described previously (Collett et al., 1973). On the side of the urero-ovarian anastomosis, a silastic cannula (i.d. 2.0 mm, o.d. 3.2 mm; Dow Corning Corp., Medical Products, Michigan) was advanced so that the tip lay opposite the entrance of the utero-ovarian vein. After cannulation was complete, 5000 lU of heparin were injected i.v. and the ewes were returned to metabolism morning
at the
endocrine
studied,
luteal
within
regression
following
spontaneous
together
involving transplants
(Table
1 and
(Baird 1976b),
both have Fig.
of the
and
induced luteal regression 1976a; Baird et al.,
results of experiments and utero-ovarian
analyzed
time
72 h of the injection of the similarity of
Because
events
prostaglandin and Scaramuzzi, the an
expected
the experiment in all ewes. On the 12 on which the ewes with ovarian
ovaribeen 1).
The
crates.
After a rest period of 2 h, timed samples of ovarian venous blood (25 ml) were collected every 10 mm for period of 60 or 120 mm as previously described (Baird et al, 1 976a). The blood was collected under sterile conditions, centrifuged immediately at 4#{176}C and a 10 ml aliquot of plasma was aspirated for steroid analysis. The rest of the plasma and red cells were returned to the ewe via the cannula in the contra-lateral jugular vein. Midway between the collection of each ovarian vein sample, 3 ml of blood was removed from the jugular for the measurement of LH and progesterone. The plasma samples were stored at -20#{176} C until assay. The ovarian transplants were sampled on 12 occasions for 60 min, 28-30 h after the injection of cloprostenol to induce luteal regression. The uteroovarian transplants were sampled for a period of 120 mm on 16 occasions on Day 12-18 of the cycle, i.e. within 7 days of the onset of estrus. By testing the ewes for estrus twice daily, at 0800-1000 h and 1600-1700 h, it was possible, in retrospect, to group all 28 experiments into 2 on the day of estrus, 9 on estrus -1, 7 on estrus -2 and 10 in late luteal:estrus -7 to -3. Analytical
Baird
3
E
2
a’ C
9 6 I -J C
E a’
I 4
13 and 9.3% for respectively. LH, which ng/ml of NIH-LH-S14, was
were measured
in dupli-
radioimmunoassay
progesterone and estradiol, was expressed in terms of was measured by a double (Martensz
et
aL,
1976).
The intra- and inter-assay precision Was 8.1 and 12.0%, respectively. All samples from the same experiment were assayed in the same run. Calculation
The
of
ESTRUS
-I
n-13
3
the details of which have published elsewhere (Scaramuzzi et al., 1975; and Scaramuzzi, 1976). The inter-assay precision
antibody
LUTEAL n-6
2
Metbods
Progesterone and estradiol cate by radioimmunoassay, been
LATE 4
Results
ovarian secretion rate was calculated from a knowledge of the time taken to collect 25 ml of blood (blood flow), concentration of estradiol and progester-
o
-30 TIME
+
30
(Mm)
FIG. 1. The concentration of LH in jugular venous plasma and ovarian estradiol secretion in 13 ewes with ovarian or utero-ovarian transplants. For each group the results have been grouped around the episodic pulses of LH as defined in the text. Each point represents the mean ± standard error of between 4 and 13 observations. Significant differences from the mean basal values prior to the LH peak are indicated by asterisks (Student’s ‘t’ teat). P
OF REPRODUCTION
Pulsatile
18, 359 -364
Secretion
(1978)
of LH and Ovarian
Follicular
Estradiol
Phase of the Sheep DAVID
MRC
Unit
Department University
of
Estrous
During
the
Cycle
T. BAIRD Reproductive
Obstetrics of Edinburgh,
Biology,
and Gynaecology, 23 Chalmers Street,
of
Edinburgh ABSTRACT The concentration of LH in jugular venous plasma and the ovarian secretion of estradiol was measured every 10 mm for periods of 60 or 120 mm on 28 occasions in 13 ewes with utero-ovarian or ovarian autotransplants. Blood samples were collected at various times during the follicular phase; 2 days prior to esifus (estruS -2), estrus -1 and on the day of estrus and the results compared to those obtained during the luteal phase. During the luteal phase, episodic pulses of LH occurred at intervals of approximately 3 h 20 mm and each pulse was followed by a significant rise in the secretion of estradiol. During the follicular phase, the pulses of LH occurred with increased frequency (1/h) but with decreased amplitude. Although each pulse of LH was smaller, the peak of secretion of estradiol following each pulse was significantly greater during the follicular than during the luteal phase. By estrus, the secretion of estradiol declined in spite of LH values in the range of the preovulatory LH surge. It is suggested that the increased secretion of estradiol from the
preovulatory Graaflan follicle with increasing frequency.
is due to repeated
stimulation
INTRODUCTION
We
between the pituitary
from lated
maturing amounts
Cox Baird
1971; 1976b;
factors responsible stage of follicular are not
known.
estrogen
Barcikowski Karsch et for growth
stimulating and steroid
During
section,
this
the
period
1974;
Karsch
et al.,
1977).
of
If the
FSH
Salamonwhile the (Akbar et rise in LH
by maintaining lutefollicular development
is
that
inhibited
suggesting related
Accepted Received
(Baird
and
the
2 events
Scaramuzzi,
the
developing episodic
reported cycle,
that estrogen
Graafian pulses of
occur
during
the
secretion
follicles is stimuLH which occur
from the ovary of the phase of the cycle. AND
sheep
during
the
METHODS
Animals
Eight ewes with ovarian (Coding et al., 1967) and 5 ewes with utero-ovarian autotransplants (Harrison et al., 1968) were allocated to this experiment during the breeding season (October-February). The ewes were housed under natural lighting conditions in a covered pen at Dryden Field Station, Roshin, Midlothian, Scotland. The left ovary alone, or together with the oviduct, horn and body of the uterus, cervix and anterior vagina, had been autotransplanted to the neck at least 12 months previously. The left ovarian and uterine arteries and uteroovarian vein were anastomosed to the carotid artery and jugular vein which were exteriorized in separate skin tubes (Baird et al., 1976c). The right ovary and uterine horn were removed. The ewes with utero-ovarian transplants had regular estrous cycles (mean 17.8 ± 0.2 days) in contrast to those with ovarian transplants which failed to show heat due to persistence of the corpus luteum because of the absence of an adjacent uterine horn
of increased
concentration is prevented al levels of progesterone, causally
Experimental
this final secretion
concentration
of
MATERIALS
et al., 1974; al., 1977). The
actually falls (L’Hermite et al., 1972; sen et al., 1973; Pant et al., 1977) basal concentration of LH rises 4-fold al.,
estradiol follicular
Graafian follicle secretes increasof estradiol (Moore et al., 1969;
et al., et al.,
previously
phase
the by
pulses of LH which
about every 3 h (Baird et al., 1976a). The present study was initiated to determine the relationship between the frequency and amplitude of pulses of LH and the secretion of
secretion of gonadoand steroids from
the ovary is not clear. As the secretion of progesterone from the regressing corpus luteum declines on Day 14 or 15 (estrus -3 and -2), the ing
have
luteal
Although the endocrine events occurring during the periovulatory period in the sheep have been described in a number of studies, the exact relationship trophins from
by episodic
are
1976a).
September 9, 1977. July 11, 1977.
359
BAIRD
360 et
(Baird
al.,
1976c).
The
ewes
were
either
pure
Tasmanian
Merino or crossed with Finnish Landrace (F X M) or Scottish Blackface (BF X M). Heat was detected by testing with a vasectomized ram at least
and, on some occasions, more frequently. of the absence of cyclical ovarian funcewes with ovarian autotransplants were studied following premature luteal regression induced by i.m. injection (100 g) of a potent analogue of prostaglandin-F, a, ICI 80996 (cloprostenol) on Day 10. Luteal regression and estrus occurred Within 72 h (Baird and Scaramuzzi, 1975).
one in ovarian venous plasma, and hematocrit. No allowance was made for the volume of plasma trapped in the packed red cells, nor for the small amount of steroids (about 1096) adsorbed on the red cell surface.
twice/day
RESULTS
Because tion, the
Estrus
occurred
7 days of occasions autotransplants
were
occurred cloprostenol.
within
estrus Experimental
Design
On the estrus, both
of an experiment, after testing for jugular veins were cannulated as described previously (Collett et al., 1973). On the side of the urero-ovarian anastomosis, a silastic cannula (i.d. 2.0 mm, o.d. 3.2 mm; Dow Corning Corp., Medical Products, Michigan) was advanced so that the tip lay opposite the entrance of the utero-ovarian vein. After cannulation was complete, 5000 lU of heparin were injected i.v. and the ewes were returned to metabolism morning
at the
endocrine
studied,
luteal
within
regression
following
spontaneous
together
involving transplants
(Table
1 and
(Baird 1976b),
both have Fig.
of the
and
induced luteal regression 1976a; Baird et al.,
results of experiments and utero-ovarian
analyzed
time
72 h of the injection of the similarity of
Because
events
prostaglandin and Scaramuzzi, the an
expected
the experiment in all ewes. On the 12 on which the ewes with ovarian
ovaribeen 1).
The
crates.
After a rest period of 2 h, timed samples of ovarian venous blood (25 ml) were collected every 10 mm for period of 60 or 120 mm as previously described (Baird et al, 1 976a). The blood was collected under sterile conditions, centrifuged immediately at 4#{176}C and a 10 ml aliquot of plasma was aspirated for steroid analysis. The rest of the plasma and red cells were returned to the ewe via the cannula in the contra-lateral jugular vein. Midway between the collection of each ovarian vein sample, 3 ml of blood was removed from the jugular for the measurement of LH and progesterone. The plasma samples were stored at -20#{176} C until assay. The ovarian transplants were sampled on 12 occasions for 60 min, 28-30 h after the injection of cloprostenol to induce luteal regression. The uteroovarian transplants were sampled for a period of 120 mm on 16 occasions on Day 12-18 of the cycle, i.e. within 7 days of the onset of estrus. By testing the ewes for estrus twice daily, at 0800-1000 h and 1600-1700 h, it was possible, in retrospect, to group all 28 experiments into 2 on the day of estrus, 9 on estrus -1, 7 on estrus -2 and 10 in late luteal:estrus -7 to -3. Analytical
Baird
3
E
2
a’ C
9 6 I -J C
E a’
I 4
13 and 9.3% for respectively. LH, which ng/ml of NIH-LH-S14, was
were measured
in dupli-
radioimmunoassay
progesterone and estradiol, was expressed in terms of was measured by a double (Martensz
et
aL,
1976).
The intra- and inter-assay precision Was 8.1 and 12.0%, respectively. All samples from the same experiment were assayed in the same run. Calculation
The
of
ESTRUS
-I
n-13
3
the details of which have published elsewhere (Scaramuzzi et al., 1975; and Scaramuzzi, 1976). The inter-assay precision
antibody
LUTEAL n-6
2
Metbods
Progesterone and estradiol cate by radioimmunoassay, been
LATE 4
Results
ovarian secretion rate was calculated from a knowledge of the time taken to collect 25 ml of blood (blood flow), concentration of estradiol and progester-
o
-30 TIME
+
30
(Mm)
FIG. 1. The concentration of LH in jugular venous plasma and ovarian estradiol secretion in 13 ewes with ovarian or utero-ovarian transplants. For each group the results have been grouped around the episodic pulses of LH as defined in the text. Each point represents the mean ± standard error of between 4 and 13 observations. Significant differences from the mean basal values prior to the LH peak are indicated by asterisks (Student’s ‘t’ teat). P
