Effect of stress-like concentrations of cortisol on the feedback potency of oestradiol in orchidectomized sheep

The effect of stress-like concentrations of cortisol on oestradiol-induced change in LH secretion and GnRH receptor expression was evaluated in orchidectomized sheep (wethers). Twenty-four wethers were assigned at random to one of the four treatment groups in a 2x2 factorial design (n=6 wethers/group). Wethers received cortisol (90 microg/kg/h; groups 2 and 4) or a comparable volume of cortisol delivery vehicle (groups 1 and 3) by continuous infusion for 48 h. During the final 24 h of infusion, wethers received oestradiol (6 ng/kg/h; groups 3 and 4) or oestradiol delivery vehicle (groups 1 and 2). The pattern of LH secretion was assessed during a 3-h period of intensive blood collection beginning 21 h after initiation of oestradiol infusion. Although neither cortisol nor oestradiol alone affected (P>0.05) mean serum concentration of LH or LH pulse frequency, serum LH and the frequency of secretory episodes of LH were significantly reduced (P<0.05) in wethers receiving cortisol and oestradiol in combination. Anterior pituitary tissue was collected at the end of the infusion period. Oestradiol increased (P<0.05) tissue concentrations of GnRH receptor and GnRH receptor mRNA. Although cortisol alone did not affect (P>0.05) basal concentrations of receptor or receptor mRNA, the magnitude of oestradiol-induced increase in GnRH receptor and GnRH receptor mRNA was significantly reduced in wethers receiving cortisol and oestradiol concurrently. Conversely, steady-state concentrations of mRNA encoding the LHbeta and FSHbeta subunits were increased (P<0.05) in wethers receiving cortisol. These observations demonstrate that stress-like concentrations of cortisol act in concert with oestradiol to suppress LH secretion. In addition, cortisol blocks oestradiol-dependent increase in pituitary tissue concentrations of GnRH receptor and GnRH receptor mRNA.     

Gonadotropin-releasing hormone (GnRH) receptor dynamics and gonadotrope responsiveness during and after continuous GnRH stimulation

Gonadotrope responsiveness, serum and tissue levels of luteinizing hormone (LH), and tissue concentration of gonadotropin-releasing hormone (GnRH) receptors in ovariectomized rats were determined during and after continuous GnRH stimulation. Intraperitoneal placement of GnRH-containing osmotic minipumps for 96 h established a rate of GnRH delivery (1 microgram/h) that resulted in stable serum levels of GnRH (500-700 pg/ml). Secretion of LH increased 8-fold within 6 h; however, serum LH returned to pretreatment levels by 24 h, even with continued GnRH stimulation. Tissue concentration of LH was depressed within 48 h of initiation of treatment but levels were restored by 96 h. Tissue levels of GnRH receptor remained elevated during the first 6 h of treatment but were reduced by 60% within 24 h and remained depressed for the duration of treatment. Gonadotrope responsiveness 48 h and 96 h after initiation of treatment was reduced by 50% and 90%, respectively. Removal of the GnRH delivery vehicle resulted in rapid disappearance of GnRH from serum. Dramatic reduction (75%) in circulating levels of LH, and a 2-fold increase in tissue levels of LH and in GnRH receptor concentration were noted within 6 h of minipump removal. Although tissue concentration of GnRH receptor returned to pretreatment levels within 48 h of minipump removal, both basal LH secretion and gonadotrope responsiveness remained depressed even 96 h after cessation of continuous GnRH stimulation. These data indicate that GnRH can "down regulate" its receptor, gonadotrope responsiveness is not obligatorily linked to receptor concentration, and desensitization that follows hyperstimulation represents effects directed at post-receptor loci.     

Gonadotrope function in ovariectomized ewes actively immunized against gonadotropin-releasing hormone (GnRH)

The gonadotrope cells of the ovine anterior pituitary were insulated from hypothalamic inputs by imposing an immunologic barrier generated by active immunization of ovariectomized ewes against gonadotropin-releasing hormone (GnRH) conjugated to keyhole limpet hemocyanin (KLH) through a p-aminophenylacetic acid bridge. All GnRH-KLH animals immunized developed titers of anti-GnRH that exceeded 1:5000. The antisera were specific for GnRH and cross-reacted with GnRH agonists modified in position 10 to an extent that was less than 0.01%. Ewes actively immunized against GnRH-KLH displayed levels of basal and GnRH agonist-induced gonadotropin secretion that were markedly lower (p less than 0.05) than comparable parameters in ewes actively immunized against KLH. In contrast, basal and thyrotropin-releasing hormone (TRH)-induced prolactin (PRL) secretion were not compromised by active immunization. Immunization against the GnRH-KLH conjugate, but not KLH alone, prevented expression of the positive feedback response to exogenous estradiol (E2). Pituitary stores of immunoactive luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were significantly (p less than 0.001) reduced in ewes immunized against GnRH-KLH but stores of PRL were not affected by such immunization. Further, the biopotency of the residual LH stores in tissue of animals from the anti-GnRH group was significantly (p less than 0.05) lower than LH biopotency in anti-KLH animals. Serum levels of LH in anti-GnRH ewes were restored by circhoral administration of a GnRH agonist that did not cross-react with the antisera generated. Pulsatile delivery of GnRH agonist in anti-GnRH ewes significantly (p less than 0.05) elevated serum LH within 48 h and reestablished LH levels comparable to anti-KLH ewes within 6 days of treatment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Progesterone directly inhibits pituitary luteinizing hormone secretion in an estradiol-dependent manner.

We recently demonstrated that progesterone and estradiol inhibit pituitary LH secretion in a synergistic fashion. This study examines the direct feedback of progesterone on the estradiol-primed pituitary. Nine ovariectomized (OVX) ewes underwent hypothalamic-pituitary disconnection (HPD) and were infused with 400 ng GnRH every 2 h throughout the experiment. After 7 days of infusion, estradiol was implanted s.c. Four days later, estradiol implants were exchanged for blank implants in 4 ewes and for progesterone implants in 5 ewes. These implants remained in place for another 4 days. Blood samples were collected around exogenous GnRH pulses before and 0.5 to 96 h after implant insertion and exchange. Serum LH and progesterone concentrations were determined through RIA. One month later, 4 of the HPD-OVX ewes previously implanted with steroids were reinfused with GnRH and the implantation protocol was repeated using blank implants only. In estradiol-primed ewes, progesterone significantly lowered LH secretion after 12 h of implantation and LH secretion remained inhibited while progesterone implants were in place (p less than 0.05). Removing estradiol transiently lowered LH secretion, and this effect was significant only 24 h after estradiol withdrawal (p less than 0.05). These data suggest that progesterone has a direct, estradiol-dependent inhibitory effect on pituitary LH release and that estradiol may sustain pituitary gonadotrope response to GnRH.     

Thermoregulatory and endocrine effects of a low dose of danazol in postmenopausal women: interaction with the effect of naloxone

Before and on the 30th day of danazol administration (200 mg/day), in six postmenopausal women the activity of endogenous opioid peptides has been indirectly evaluated by the effect on LH secretion and body temperature (measured as rectal temperature) exerted by the infusion of the opioid antagonist naloxone (1.6 mg/h x 4 h preceded by 1.6 mg iv bolus). Before and during danazol administration a GnRH test (100 mcg iv bolus) was also performed to evaluate possible variations in pituitary responsiveness to GnRH. Danazol significantly reduced mean plasma levels of LH and FSH (p less than 0.01), and their response to GnRH stimulus (p less than 0.05). Either before or during danazol administration mean plasma LH and FSH levels did not vary during the infusion of naloxone, while body temperature significantly decreased (p less than 0.01). The decrease in body temperature was significantly greater (p less than 0.05) during danazol than before treatment. The present data suggest that in postmenopausal women a low dose of danazol exerts an antigonadotropic effect mainly reducing the pituitary responsiveness to GnRH. The enhanced hypothermic response to naloxone observed during danazol administration also seems to suggest that in postmenopausal women a low dose of danazol enhances the thermoregulatory role of endogenous opioid peptides.     

Estrogenic effects of phenol red on rat pituitary cell responsiveness to gonadotropin-releasing hormone

This study investigated whether phenolsulfonphthalein (PR), a common pH indicator in tissue culture media, affects luteinizing hormone (LH) secretion from rat pituitary cells or 17 beta-estradiol (E2) augmentation of pituitary responsiveness to gonadotropin-releasing hormone (GnRH). PR enhanced GnRH-stimulated LH secretion and shifted the GnRH dose-response curve leftward with a relative potency ratio of 0.24 +/- 0.09 (+/- SE; p less than 0.01). The effect of E2 on LH release was significantly diminished by PR, which elevated GnRH-stimulated LH secretion in the absence of E2. This phenomenon was elicited by PR from different sources and was inhibited by the antiestrogen Cl628. Thus, PR exerted estrogen-like effects on rat pituitary cells and caused an underestimation of the degree to which E2 enhanced GnRH-stimulated LH secretion.     

Effect of stress-like concentrations of cortisol on follicular development and the preovulatory surge of LH in sheep

Stress-like concentrations of cortisol increase the negative feedback potency of oestradiol in castrated male sheep. A similar cortisol-dependent response in female sheep might be expected to suppress gonadotrophin secretion and impair follicular development and ovulation. The oestrous activity of 21 female sheep was synchronized using progestogen-treated vaginal pessaries to test this hypothesis. Stress-like concentrations of cortisol (60-70 ng ml-1) were established by continuous infusion of cortisol (80 micrograms kg-1 h-1; n = 13) beginning 5 days before, and continuing for 5 days after, pessary removal. Control animals (n = 8) received a comparable volume of vehicle (50% ethanol-saline) over the 10 day infusion period. Serum concentrations of oestradiol increased progressively in control sheep during the 48 h immediately after pessary removal. This increase in serum oestradiol was blocked or significantly attenuated in sheep receiving stress-like concentrations of cortisol. Preovulatory surge-like secretion of LH was apparent in control animals 58.5 +/- 2.1 h after pessary removal. In contrast, surge-like secretion of LH was not observed during the 5 days after pessary removal in 54% (7 of 13) of sheep receiving cortisol. Moreover, the onset of the surge was significantly delayed in the cortisol-treated ewes that showed surge-like secretion of LH during the infusion period. The ability of episodic pulses of exogenous GnRH to override the anti-gonadal effect of cortisol was examined in a second study. Oestrous activity of 12 ewes was synchronized using progestogen-containing pessaries as described above. Ewes were randomly assigned to one of three treatment groups (n = 4 ewes per group). Animals received cortisol (100 micrograms kg-1 h-1; groups 1 and 2) or a comparable volume of vehicle (group 3) beginning 5 days before, and continuing for 2 days after, pessary removal. Pulses of GnRH (4 ng kg-1 h-1, i.v.; group 1) or saline (groups 2 and 3) at 1 h intervals were initiated at pessary removal and continued for 48 h. Serum concentrations of oestradiol were not significantly increased after pessary removal in sheep receiving cortisol alone. Conversely, serum concentrations of oestradiol increased progressively during the 48 h after pessary removal in control ewes and in ewes receiving cortisol and GnRH. At the end of infusion, serum concentrations of oestradiol did not differ (P > 0.05) between control (7.7 +/- 0.8 pg ml-1) ewes and ewes receiving cortisol and episodic GnRH (6.4 +/- 1.3 pg ml-1). Moreover, these values were significantly greater (P < 0.05) than the serum concentrations of oestradiol in animals receiving cortisol (1.0 +/- 0.4 pg ml-1) alone. Collectively, these data indicate stress-like concentrations of cortisol block or delay follicular development and the preovulatory surge of LH in sheep. In addition, episodic GnRH overrides cortisol-induced delay in follicular maturation.     

Testosterone inhibits gonadotropin-releasing hormone pulse frequency in the male sheep

The objective was to determine the effect of chronic testosterone (T) treatment on GnRH and LH secretion in wethers. Rams were either castrated only or castrated and immediately treated with Silastic implants containing T. Several weeks later, a device for collecting hypophyseal-portal blood was surgically implanted. Six to seven days later, blood samples were collected simultaneously and continuously from the portal vessels and jugular vein of pairs of conscious animals. Samples were divided at 10-min intervals for 6-12 h. One hour before the end of collection, all animals received i.v. injections of 250 ng of GnRH. In samples collected simultaneously from 6 pairs of animals, T reduced the frequency of both GnRH pulses (1.8 +/- 0.2 vs. 0.9 +/- 0.3/h, p less than 0.03) and LH pulses (1.6 +/- 0.1 vs. 0.8 +/- 0.3/h, p less than 0.03). T did not alter amplitude of either GnRH or LH pulses. Testosterone reduced mean GnRH (9.7 +/- 0.6 vs. 7.9 +/- 0.5 pg/ml, p less than 0.05), whereas mean LH was not significantly reduced (9.6 +/- 1.4 vs. 6.1 +/- 1.8 ng/ml, p = 0.16). These results support the hypothesis that T reduces GnRH pulse frequency.     

Continuous infusion of GnRH reduces the LH response to an intravenous GnRH injection but does not inhibit endogenous LH secretion in cows

Plasma LH concentrations were monitored in 6 Hereford X Friesian suckled cows at about 80 days post partum, before and during a 14-day period of continuous s.c. infusion of GnRH (20 micrograms/h). Blood samples were collected at 10-min intervals on Days -2, -1, 1, 2, 3, 4, 7, 10, 13 and 14 (Day 1 = start of infusion). Plasma LH concentrations rose from mean pretreatment levels of 1.3 +/- 0.20 ng/ml to a maximum of 17.1 +/- 3.09 ng/ml within the first 8 h of GnRH infusion, but returned to pretreatment levels by Day 2 or 3. In 4/6 animals, the initial increase was of a magnitude characteristic of the preovulatory LH surge. In all animals, an i.v. injection of 10 micrograms GnRH, given before the start and again on the 14th day of continuous infusion, induced an increase in LH concentrations but the increase to the second injection was significantly (P less than 0.01) less (mean max. conc. 6.4 +/- 0.76 and 2.3 +/- 0.19 ng/ml). Mean LH concentrations (1.0 +/- 0.08, 1.1 +/- 0.08 and 0.9 +/- 0.06 ng/ml) and LH episode frequencies (3.3,4.3 and 3.2 episodes/6 h) did not differ significantly on Days -2,7 and 13. However, the mean amplitude of LH episodes was significantly lower (P less than 0.05) on Day 13 (1.3 +/- 0.10 ng/ml) than on Day -2 (1.8 +/- 0.16 ng/ml). Therefore, although the elevation in plasma LH concentrations that occurs in response to continuous administration of GnRH is short-lived and LH levels return to pre-infusion values within 48 h of the start of infusion, these results show that the pituitary is still capable of responding to exogenous GnRH, although the LH response to an i.v. bolus injection of GnRH is reduced. In addition, this change in pituitary sensitivity is not fully reflected in endogenous patterns of episodic LH secretion.     

Naloxone evokes large-amplitude GnRH pulses in luteal-phase ewes

Ewes were sampled during the mid-late luteal phase of the oestrous cycle. Hypophysial portal and jugular venous blood samples were collected at 5-10 min intervals for a minimum of 3 h, before i.v. infusions of saline (12 ml/h; N = 6) or naloxone (40 mg/h; N = 6) for 2 h. During the 2-h saline infusion 2/6 sheep exhibited a GnRH/LH pulse; 3/6 saline infused ewes did not show a pulse during the 6-8-h portal blood sampling period. In contrast, large amplitude GnRH/LH pulses were observed during naloxone treatment in 5/6 ewes. The mean (+/- s.e.m.) amplitude of the LH secretory episodes during the naloxone infusion (1.07 +/- 0.11 ng/ml) was significantly (P less than 0.05) greater than that before the infusion in the same sheep (0.54 +/- 0.15 ng/ml). Naloxone significantly (P less than 0.005) increased the mean GnRH pulse amplitude in the 5/6 responding ewes from a pre-infusion value of 0.99 +/- 0.22 pg/min to 4.39 +/- 1.10 pg/min during infusion. This episodic GnRH secretory rate during naloxone treatment was also significantly (P less than 0.05) greater than in the saline-infused sheep (1.53 +/- 0.28 pg/min). Plasma FSH and prolactin concentrations did not change in response to the opiate antagonist. Perturbation of the endogenous opioid peptide system in the ewe by naloxone therefore increases the secretion of hypothalamic GnRH into the hypophysial portal vasculature. The response is characterized by a large-amplitude GnRH pulse which, in turn, causes a large-amplitude pulse of LH to be released by the pituitary gland.     

Regulation of gonadotropin secretion and number of gonadotropin-releasing hormone receptors by inhibin, activin-A, and estradiol.

Primary cultures of ovine pituitary cells were used to characterize the effects of inhibin and activin on the secretion of gonadotropins and on the regulation of number of GnRH receptors in the presence or absence of estradiol. Number of GnRH receptors was determined by the specific binding of a saturating dose of [125I]des-Gly10-D-Trp6-GnRH-ethylamide (GnRH-A). Recombinant human inhibin-A (rh-inhibin-A) or inhibin in porcine and bovine follicular fluid (pFF and bFF, respectively) decreased secretion of FSH in a dose-dependent manner, with maximum inhibition at an inhibin concentration of approximately 0.1 nM. Neither pFF or bFF affected secretion of LH, although rh-inhibin-A caused a modest decrease (p less than 0.05) in secretion of LH. Treatment of cells with rh-inhibin-A, bFF, or pFF approximately doubled the number of GnRH receptors. Scatchard analysis indicated that increases in GnRH-A binding were due to an increase in receptor number rather than a change in affinity. Additionally, rh-inhibin-A, at a dose that doubled numbers of GnRH receptors, increased GnRH-induced LH release above that caused by GnRH alone, indicating that the increase in receptor number leads to increased responsiveness to GnRH. Recombinant human activin-A (rh-activin-A) increased secretion of FSH but did not affect secretion of LH. Number of GnRH receptors was not affected by lower concentrations of rh-activin-A but was decreased (p less than 0.05) by 3.0 nM activin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of progesterone on gonadotropin-releasing hormone receptor concentration in cultured estrogen-primed female rat pituitary cells

Acute (0.5–4 h) treatment of estradiol (E)-primed female rat pituitary cells with progesterone (P) augments gonadotropin-releasing hormone (GnRH)-induced LH release, whereas chronic (48 h) P-treatment reduces pituitary responsiveness to the hypothalamic decapeptide. Dispersed E-primed (48 h, 1 nM) rat pituitary cells were cultured for 4 or 48 h in the presence of 100 nM P to assess the effects of the progestagen on GnRH receptors and on gonadotrope responsiveness to the decapeptide. P-treatment (4 h) significantly augmented GnRH-receptor concentrations (4.44 ± 0.6 fmol/10⁶ cells) as compared to cells treated only with E (2.6 ± 0.5fmol/10⁶ cells). Parallel significant changes in GnRH-induced LH secretion were observed. The acute increase in GnRH-receptor number was nearly maximal (180% of receptor number in cells treated with E alone) within 30 min of P addition. Chronic P-treatment (48 h) significantly reduced pituitary responsiveness to GnRH as compared to E-treatment. The GnRH-receptor concentrations (3.9 ± 0.6 fmol/10⁶ cells), however, remained elevated above those in E-primed cells. GnRH-receptor affinity was not influenced by any of the different treatments. These results indicate that the acute facilitatory P-effect on GnRH-induced LH release is at least chronologically closely related to an increase in GnRH-receptor concentration. The chronic negative P-effect on pituitary responsiveness to GnRH, however, shows no relation to changes in available GnRH receptors.     

Effect of clomiphene citrate on pituitary responsiveness to gonadotropin releasing hormone in rams and wethers

The objective of this study was to determine the effect of clomiphene citrate (clomid) on pituitary responsiveness to gonadotropin releasing hormone (GnRH) in rams and wethers. Doses of 200 mg clomid per ram and 1 mug GnRH per 50 kg body weight were used in studies on 12 rams and 4 wethers. The experimental design involved bleeding each animal at 15-minute intervals for 6.5 hours. At the end of the first hour, GnRH was injected IV. The second GnRH challenge was administered 0.5 hours after an injection of clomid or vehicle (4.5% sorbitol solution) which was given on the third hour. The relative response to clomid or vehicle was calculated as the mean increase in concentration of LH during the two-hour period after the second GnRH injection. Each treatment (clomid and vehicle) was given to all animals with a 14-day recovery period between treatment days. The relative response for the rams receiving vehicle (1.80 +/- 0.65) was greater (P < 0.05) than the response during clomid treatment (0.34 +/- 0.22). This suppression of LH response by clomid was observed in 10 of the 12 rams. In contrast to the rams, the concentrations of LH in wethers after the second GnRH injection were lower than those observed after the first GnRH injection. Similar to the rams, the relative response following clomid treatment of wethers (0.04 +/- 0.04) was less than the relative response (P > 0.05) following vehicle (0.40 +/- 0.16). The results suggest that clomid at this dosage inhibits GnRH-induced release of LH from the pituitary of rams but not of wethers.     

The effect of opioid peptides on ovine pituitary gonadotropin secretion in vitro

Although a hypothalamic site of action has been firmly established for opiate-mediated gonadotropin regulation, there have been several reports which indicate the possibility of a direct influence on the pituitary gland. The objective of this study was to further investigate this possibility in an in vitro pituitary perifusion system utilizing ovine tissue. Treatment with gamma-endorphin (GE) or human beta-endorphin (hBE) resulted in elevated basal LH release (p less than 0.05), followed by an inhibition in the response to a subsequent GnRH challenge (p less than 0.05). The stimulatory effect of hBE was found to be dose-responsive (p less than 0.01). PRL secretion was not similarly stimulated. Ovine beta-endorphin (oBE) had no effect on LH secretion, even though it differs from hBE by only 2 amino acids and contains the active GE sequence. Met-enkephalin also did not influence gonadotropin secretion. Naloxone pretreatment did not reverse the effects of hBE on gonadotropin release. It was found, however, that [D-pGlu1, D-Phe2, D-Trp3,6]-GnRH, a specific GnRH receptor antagonist, did reduce hBE-induced LH and FSH release (p less than 0.05). Naloxone pretreatment alone suppressed the response to GnRH (p less than 0.05). These data indicate that certain opioid peptides can influence ovine gonadotropin secretion in vitro by activating the GnRH receptor. Furthermore, a facilitory role is suggested for endogenous opiates in the local regulation of pituitary gonadotropin secretion.     

Membrane-initiated actions of estradiol (E2) in the regulation of LH secretion in ovariectomized (OVX) ewes

Background  

We demonstrated that E2 conjugated to BSA (E2BSA) induces a rapid membrane-initiated inhibition of LH secretion followed hours later by a slight increase in LH secretion. Whether these actions of E2BSA are restricted to the pituitary gland and whether the membrane-initiated pathway of E2BSA contributes to the up-regulation of the number of GnRH receptors during the positive feedback effect of E2 were evaluated here. We have shown that the suppression of LH secretion induced by E2 and E2BSA is the result of a decreased responsiveness of the pituitary gland to GnRH. In this study we further tested the ability of E2BSA to decrease the responsiveness of the pituitary gland to GnRH under the paradigm of the preovulatory surge of LH induced by E2.     

Suppression of pulsatile luteinizing hormone secretion by gonadotrophin-releasing hormone antagonist does not affect episodic progesterone secretion or corpus luteum function in ewes.

Progesterone secretion has been observed to be episodic in the late luteal phase of the oestrous cycle of ewes and is apparently independent of luteinizing hormone (LH). This study investigated the effects of suppressing the pulsatile release of LH in the early or late luteal phase on the episodic secretion of progesterone. Six Scottish Blackface ewes were treated i.m. with 1 mg kg-1 body weight of a potent gonadotrophin-releasing hormone (GnRH) antagonist on either day 4 or day 11 of the luteal phase. Six ewes received saline at each time and acted as controls. Serial blood samples were collected at 10 or 15 min intervals between 0 and 8 h, 24 and 32 h, and 48 and 56 h after GnRH antagonist treatment and daily from oestrus (day 0) of the treatment cycle for 22 days. Oestrous behaviour was determined using a vasectomized ram present throughout the experiment. Progesterone secretion was episodic in both the early and late luteal phase with a frequency of between 1.6 and 3.2 pulses in 8 h. The GnRH antagonist abolished the pulsatile secretion and suppressed the basal concentrations of LH for at least 3 days after treatment. This suppression of LH, in either the early or late luteal phase, did not affect the episodic release of progesterone. Daily concentrations of progesterone in plasma showed a minimal reduction on days 11 to 14 after GnRH antagonist treatment on day 4, although this was significant (P < 0.05) only on days 11 and 13. There was no effect of treatment on day 11 on daily progesterone concentration, and the timing of luteolysis and the duration of corpus luteum function was unaffected by GnRH antagonist treatment on either day 4 or day 11. These results indicate that the episodic secretion of progesterone during the luteal phase of the oestrous cycle in ewes is independent of LH pulses and normal progesterone secretion by the corpus luteum can be maintained with minimal basal concentrations of LH.     

Feedback effects of ovarian steroids on the hypothalamic-hypophyseal axis in the rabbit

The feedback effects of two ovarian steroids, estradiol-17 beta (E2) and 20 alpha-hydroxypregn-4-en-3-one (20 alpha OH), were examined in both intact (INT) and ovariectomized (OVEX) does. We measured steroid-induced alterations in endogenous gonadotropin-releasing hormone (GnRH) from sequential 10-min samples of hypothalamic perfusates, simultaneous changes in peripheral plasma luteinizing hormone (LH) and follicle-stimulating hormone (FSH), and the modification of pituitary responsiveness, i.e., increments in plasma LH (delta LH) and plasma FSH (delta FSH), after 50 ng, 250 ng, and 1 microgram of exogenous GnRH in individual does of 6 treatment groups. The groups were: INT does, OVEX does, OVEX does receiving either one (1 E2) or two (2 E2) E2-filled Silastic capsules, OVEX does receiving a 20 alpha OH-filled capsule (20 alpha OH), and OVEX does receiving both capsules of E2 and 20 alpha OH (1 E2 + 20 alpha OH). Ovariectomy enhanced the pulsatile release of hypothalamic GnRH and pituitary LH and FSH, and increased the LH response (delta LH) to exogenous GnRH (OVEX vs. INT, p less than 0.05). Replacement of E2 at the time of ovariectomy prevented the increased GnRH and gonadotropin secretion as well as the enhanced delta LH that were observed in untreated OVEX does. The release of hypothalamic GnRH in the 20 alpha OH group was lower (p less than 0.05) than that in the OVEX group and not different from that in the INT group. The release of pituitary LH and FSH and the delta LH in the 20 alpha OH group was not different from that in the OVEX group, but these parameters were greater (p less than 0.05) than those in the INT group. The hypothalamic GnRH pulse frequency in the 1 E2 + 20 alpha OH group was lower (p less than 0.05) than that in either the 1 E2 or the 20 alpha OH group, but the delta LH in the 1 E2 + 20 alpha OH group was not different from that in either the 1 E2 or the 20 alpha OH group. The highest dose (1 microgram) of exogenous GnRH stimulated a modest increase in FSH in the OVEX, 20 alpha OH, 1 E2 + 20 alpha OH, and 1 E2 groups; but a steroid effect on delta FSH among these 4 groups was not apparent.(ABSTRACT TRUNCATED AT 250 WORDS)     

Induced ovulation and changes in pituitary responsiveness to continuous infusion of gonadotropin-releasing hormone during the ovarian cycle in the bullfrog, Rana catesbeiana

Chronic (2-4 days) constant-rate infusions of mammalian gonadotropin releasing hormone (GnRH) were performed in female bullfrogs, Rana catesbeiana. The magnitude and temporal relationship of profiles of plasma follicle-stimulating hormone (FSH), luteinizing hormone (LH) and sex steroids [testosterone (T), estradiol-17 beta (E2) and progesterone (P)] during GnRH infusion were dependent on ovarian stage. However, in all females, the same biphasic increase in plasma gonadotropins was apparent and initial elevations in gonadotropins were accompanied by correlated increments in plasma T and E2. Complete pituitary "desensitization" to chronic GnRH infusion was not observed. Females in early follicular stages were relatively unresponsive to infusions of 1.0-10.0 micrograms/h GnRH; elevations in plasma LH were marginal and FSH was unchanged. Females with fully developed (preovulatory) ovaries were more responsive: infusion of 1.0 micrograms/h GnRH produced significant elevations in plasma LH by 2 h followed by even larger increases ("surges") after 12 h. This LH "surge" was preceded by a decline in plasma T and E2 and was accompanied by abrupt elevations in plasma P and by ovulation. Postovulatory females showed a more gradual and smaller increase in plasma LH. Infusion of GnRH in the female bullfrog establishes a clear relationship between pituitary responsiveness and the ovarian cycle not evident from acute GnRH injection; GnRH was most effective immediately before ovulation. These data are also the first to detail periovulatory changes in plasma gonadotropins and ovarian steroids in an amphibian.     

Hormone responses to exogenous GnRH pulses in post-partum dairy cows

Nine Friesian dairy cows were treated with 2.5 micrograms GnRH i.v. at 2-h intervals for 48 h commencing between Days 3 and 8 post partum. Hormone concentrations were measured in jugular venous plasma. An episodic pattern of LH release was induced in all animals and there was no significant change in amplitude during treatment. However, cows treated between Days 7 and 8 ('late') showed higher LH episode peaks than did those treated between Days 3 and 6 ('early'). Plasma FSH concentrations showed a less clear episodic pattern in response to GnRH injection. The mean height of FSH responses to GnRH tended to be higher in the 'early' group than in the 'late' group, as did mean FSH concentrations during the pretreatment sampling period. Although clear episodic changes were not observed, GnRH treatment induced a rapid sustained rise in plasma oestradiol-17 beta concentrations, indicating the responsiveness of ovarian follicles to gonadotrophin stimulation early in the post-partum period. There was no difference in oestradiol-17 beta concentrations between the 'early' and 'late' groups during the treatment period. Only one cow exhibited preovulatory-type LH, FSH and oestradiol-17 beta surges during the 96-h post-treatment sampling period. It is concluded that: (1) responsiveness to GnRH pulses increases significantly and FSH responsiveness tends to decrease with time post partum, (2) ovarian follicles are able to secrete oestradiol-17 beta in response to GnRH-induced LH and FSH release during the early post-partum period and there is no time-dependent change in responsiveness; and (3) the lack of preovulatory surges, except in one cow, may reflect a temporary defect in the positive-feedback mechanism by which high concentrations of oestradiol-17 beta induce preovulatory gonadotrophin release.     

Inhibition of luteinizing hormone secretion by localized administration of estrogen, but not dihydrotestosterone, is enhanced in the ventromedial hypothalamus during feed restriction in the young wether

The ability of steroids to inhibit LH secretion is enhanced during undernutrition. To identify potential hypothalamic sites at which this enhancement may occur, we examined LH secretion in feed-restricted or fed young wethers treated with locally administered metabolites of testosterone. In experiment 1, microimplants containing crystalline estradiol-17beta (E) or cholesterol were administered via chronic guide tubes directed to the preoptic area (POA) or ventromedial hypothalamus (VMH) in fed or feed-restricted wethers. E treatment in the VMH decreased LH pulse frequency, pulse amplitude, and mean LH concentration in feed-restricted, but not fed, wethers. E may act in the POA to suppress LH under feed restriction, but definite conclusions cannot be drawn because of steroid-independent effects of feed restriction on LH pulse frequency. In experiment 2, the effect of dihydrotestosterone (DHT) in the VMH was determined. DHT administration to the VMH did not alter LH secretion in either feed-restricted or fed wethers. Thus the VMH is one site wherein E negative feedback is enhanced during feed restriction in the wether. In contrast, we found no evidence for enhanced responsiveness to androgen negative feedback within the VMH of feed-restricted wethers. We suggest that increased sensitivity within the VMH to E, but not to DHT, is important for suppressing LH secretion in undernourished male sheep.