Pituitary receptors for gonadotropin-releasing hormone in relation to changes in pituitary and plasma luteinizing hormone in ovariectomized-hypothalamo pituitary disconnected ewes. I. Effect of changing frequency of gonadotropin-releasing hormone pulses

Studies were undertaken to determine if changes in the amplitude of luteinizing hormone (LH) pulses that occur in response to changes in the frequency of gonadotropin-releasing hormone (GnRH) pulses are due to an alteration in the number of GnRH receptors. Ewes were ovariectomized (OVX) and the hypothalamus was disconnected from the pituitary (HPD). Ewes were then given pulses of GnRH at a frequency of 1/h or 1/3 h. Two control groups were included: OVX ewes not subjected to HPD, and HPD ewes that were not OVX. At the end of one week of treatment, blood samples were collected to determine the amplitude of LH pulses. The treated ewes were killed just before the next scheduled pulse of GnRH, and the content of LH and number of GnRH receptors were measured in each pituitary. The amplitude of LH pulses was highly correlated with the amount of LH in the pituitary gland (r = 0.71, p less than 0.01), and both LH content and pulse amplitude (mean + SEM) were higher in ewes receiving GnRH once per 3 h (189.7 +/- 39.3 microgram/pituitary, 10.3 +/- 1.1 ng/ml, respectively) than in ewes receiving GnRH once per h (77.8 +/- 11.4 microgram/pituitary, 5.2 +/- 1.3 ng/ml). The pituitary content of LH was highest in the OVX ewes (260.2 +/- 57.4 micrograms/pituitary) and lowest in the nonpulsed HPD ewes (61.7 +/- 51.2 micrograms/pituitary). The number of GnRH receptors was similar in all groups, and was not correlated with any other variable.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of estradiol in inducing an ovulatory-like surge of luteinizing hormone in sheep

Two experiments were performed to examine the effect of estradiol on secretion of luteinizing hormone (LH) and on the number of receptors for gonadotropin-releasing hormone (GnRH) after down regulation of GnRH receptors in ovariectomized ewes. In the first experiment, ovariectomized ewes were administered one of four treatments: Group 1) infusion of GnRH i.v. for 40 h; Group 2) injection of 100 micrograms estradiol i.m.; Group 3) infusion of GnRH i.v. for 16 h followed immediately by an injection of 100 micrograms estradiol i.m.; and Group 4) infusion of GnRH i.v. for 40 h plus injection of 100 micrograms estradiol i.m. after the 16th h of infusion. Ewes in Groups 1, 3 and 4 responded to the infusion of GnRH with an immediate increase in serum concentrations of LH, with maximum values occurring between 2 and 4 h after the start of infusion; serum concentrations of LH then began to decline and were approaching the pretreatment baseline within 16 h. Administration of estradiol resulted in a surge of LH regardless of whether the pituitary had been desensitized by infusion of GnRH or not. In all cases the magnitude of the surge was similar to that induced by the initial infusion of GnRH. In Groups 2 and 3 the surge of LH began at 12.3 +/- 0.1 and 11.9 +/- 0.1 h after administration of estradiol. In contrast, the ewes in Group 4 had a surge of LH beginning 3.7 +/- 0.1 h after administration of estradiol.(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.     

Direct effects of estradiol-17 beta on the number of gonadotropin-releasing hormone receptors in the ovine pituitary

Concentrations of pituitary receptors for gonadotropin-releasing hormone (GnRH) are affected by GnRH and gonadal steroids. To test the hypothesis that estradiol-17 beta (E2) directly affects the number of GnRH receptors in the pituitary, independent of GnRH secretion, ovariectomized ewes with hypothalamic-pituitary disconnections (HPD) were given 25 micrograms (i.m.) of E2 (HPD + E2, n = 5) or oil (HPD + OIL, n = 5). Ovariectomized control ewes, with intact hypothalamic-pituitary axes (INT), also received either E2 or oil (INT + E2, n = 6; INT + OIL, n = 6). Blood samples were taken hourly for analysis of serum concentrations of luteinizing hormone (LH) from 4 h prior to until 16 h after treatment. Pituitaries were collected 16 h after treatment for analysis of GnRH receptors. Treatment with E2 increased concentrations of LH in serum beginning 12.7 +/- 0.6 h after injection in INT ewes but not in HPD ewes. Compared to INT + OIL ewes, E2 treatment increased (p less than 0.001) the number of GnRH receptors by 2.5-fold in INT ewes and by 2.0-fold in HPD ewes. These results suggest that although GnRH is necessary for secretion of gonadotropins, E2 alone can directly increase the number of GnRH receptors in the pituitary.     

Evidence for a direct negative effect of estradiol at the level of the pituitary gland in sheep

The purpose of this experiment was to determine if pituitary stores of LH could be replenished by administration of GnRH when circulating concentrations of both progesterone and estradiol-17 beta (estradiol) were present at levels observed during late gestation. Ten ovariectomized (OVX) ewes were administered estradiol and progesterone via Silastic implants for 69 days. One group of 5 steroid-treated OVX ewes was given GnRH for an additional 42 days (250 ng once every 4 h). Steroid treatment alone reduced (p less than 0.01) the amount of LH in the anterior pituitary gland by 77%. Pulsatile administration of GnRH to steroid-treated ewes resulted in a further decrease (p less than 0.01) in pituitary content of LH. Compared to the OVX ewes, concentrations of mRNAs for alpha- and LH beta-subunits were depressed (p less than 0.01) in all steroid-treated ewes, whether or not they received GnRH. The ability of the dosage of GnRH used to induce release of LH was examined by collecting blood samples for analysis of LH at 15 days and 42 days after GnRH treatment was initiated. Two of 5 and 3 of 5 steroid-treated ewes that received pulses of GnRH responded with increased serum concentrations of LH after GnRH administration during the first and second bleedings, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Estrogen and progesterone receptor content in the pituitary gland and uterus of progesterone-primed and gonadotropin releasing hormone-treated anestrous ewes

The objective of this work was to investigate the effect of progesterone (P) and gonadotropin-releasing hormone (GnRH) treatment on estrogen receptor (ER) and P receptor (PR) concentrations in the pituitary gland and uterus of anestrous ewes. Ewes were either not treated (group C, n = 4); were treated with 0.33 g P-controlled internal drug release (P-CIDR) for 10 days (group P, n = 4), with GnRH, 6.7 ng i.v. injections every 2 h for 18 h followed by a 4 microg bolus administration of Receptal at 20 h (group GnRH, n = 4), or with a combination of the P and GnRH treatment (group P + GnRH, n = 3). Ewes were humanely killed either at the beginning of the experiment (group C), when the CIDR was removed (group P), or 24 h after the GnRH bolus treatment (groups GnRH and P + GnRH). Progesterone treatment increased serum P concentrations, indicating that the treatment was effective. All GnRH treated ewes had similar luteinizing hormone (LH) surges, which lasted 8 h. At slaughter, estradiol (E2) concentrations in the GnRH group were higher than in groups C, P, and P + GnRH. Treatment with GnRH increased more than 10-fold the content of ER and PR in the pituitary gland without altering steroid receptor concentrations in the uterus. When GnRH was combined with P the uterine receptor contents were higher than with P treatment alone. The treatment with P decreased ER and PR content in the uterus, but had no effect on the pituitary gland. The results show that regulation by P and GnRH of ER and PR content in anestrous ewes is tissue-specific.     

Progesterone does not affect the amount of mRNA for gonadotropins in the anterior pituitary gland of ovariectomized ewes

To evaluate the effect of progesterone on the synthesis and secretion of gonadotropins, ovariectomized ewes either were treated with progesterone (n = 5) for 3 wk or served as controls (n = 5) during the anestrous season. After treatment for 3 wk, blood samples were collected from progesterone-treated and ovariectomized ewes. After collection of blood samples, hypothalamic and hypophyseal tissues were collected from all ewes. Half of each pituitary was used to determine the content of luteinizing hormone (LH) and follicle-stimulating hormone (FSH), and the number of receptors for gonadotropin-releasing hormone (GnRH). The amounts of mRNA for LH beta subunit, FSH beta subunit, alpha subunit, growth hormone, and prolactin were measured in the other half of each pituitary. Treatment with progesterone reduced mean serum concentrations of LH (p less than 0.001) but ot FSH (p greater than 0.05). Further, progesterone decreased (p less than 0.05) the total number of pulses of LH. We were unable to detect pulsatile release of FSH. Hypothalamic content of GnRH, number of receptors for GnRH, pituitary content of gonadotropins and mRNA for LH beta subunit, FSH beta subunit, alpha subunit, growth hormone, and prolactin were not affected (p greater than 0.05) by treatment with progesterone. Thus, after treatment with progesterone, serum concentrations of LH (but not FSH) are decreased. This effect, however, is not due to a decrease in the steady-state amount of mRNA for LH beta or alpha subunits.     

Effect of number of receptors for gonadotropin-releasing hormone on the release of luteinizing hormone

The relationship between number of receptors for gonadotropin-releasing hormone (GnRH) and the ability of the anterior pituitary gland to release luteinizing hormone (LH) was examined in ovariectomized ewes. A GnRH antagonist was used to regulate the number of available receptors. The dose of GnRH antagonist required to saturate approximately 50 and 90% of GnRH receptors in ovariectomized ewes was determined. Thirty min after intracarotid infusion of GnRH antagonist, ewes were killed and the number of unsaturated (i.e., those available for binding) pituitary GnRH receptors was quantified. Infusion of 10 and 150 micrograms GnRH antagonist over a 5-min period reduced binding of the labeled ligand to approximately 50 and 12% of controls, respectively. The effect of reducing the number of GnRH receptors on release of LH after varying doses of the GnRH agonist, D-Ala6-GnRH-Pro9-ethylamide (D-Ala6-GnRH) was then evaluated. One of four doses of D-Ala6-GnRH (0.125, 2.5, 50 and 400 micrograms) was given i.v. to 48 ovariectomized ewes whose GnRH receptors had not been changed or were reduced to approximately 50 or 12% of control ewes. In ewes with a 50% reduction in GnRH receptors, total release of LH (area under response curve) was lower than that obtained for controls (P less than 0.01) at the 0.125-micrograms dose of D-Ala (6.1 +/- 0.7 cm2 vs. 13.5 +/- 0.7 cm2) but was not different at the 2.5-, 50- or 400-micrograms doses of D-Ala6-GnRH.(ABSTRACT TRUNCATED AT 250 WORDS)     

Steroid feedback inhibition of pulsatile secretion of gonadotropin-releasing hormone in the ewe

The long-term negative feedback effects of sustained elevations in circulating estradiol and progesterone on the pulsatile secretion of gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) were evaluated in the ewe following ovariectomy during the mid-late anestrous and early breeding seasons. GnRH secretion was monitored in serial samples of hypophyseal portal blood. Steroids were administered from the time of ovariectomy by s.c. Silastic implants, which maintained plasma concentrations of estradiol and progesterone at levels resembling those that circulate during the mid-luteal phase of the estrous cycle; control ewes did not receive steroidal replacement. Analysis of hormonal pulse patterns in serial samples during 6-h periods on Days 8-10 after ovariectomy disclosed discrete, concurrent pulses of GnRH in hypothalamo-hypophyseal portal blood and LH in peripheral blood of untreated ovariectomized ewes. These pulses occurred every 97 min on the average. Treatment with either estradiol or progesterone greatly diminished or abolished detectable pulsatile secretion of GnRH and LH, infrequent pulses being evident in only 3 of 19 steroid-treated ewes. No major seasonal difference was observed in GnRH or LH pulse patterns in any group of ewes. Our findings in the ovariectomized ewe provide direct support for the conclusion that the negative-feedback effects of estradiol and progesterone on gonadotropin secretion in the ewe include an action on the brain and a consequent inhibition of pulsatile GnRH secretion.     

Differential modulation of gonadotropin secretion by selective estrogen receptor 1 and estrogen receptor 2 agonists in ovariectomized ewes

The objectives of this study were to determine whether activation of estrogen receptor 1 (ESR1; also known as ERalpha), or estrogen receptor 2 (ESR2; also known as ERbeta), or both are required to: 1) acutely inhibit secretion of LH, 2) induce the preovulatory-like surge of LH, and 3) inhibit secretion of FSH in ovariectomized (OVX) ewes. OVX ewes (n = 6) were administered intramuscularly 25 micrograms estradiol (E2), 12 mg propylpyrazoletriol (PPT; a subtype-selective ESR1 agonist), 21 mg diaprylpropionitrile (DPN; a subtype-selective ESR2 agonist), or PPT + DPN. Like E2, administration of PPT, DPN, or combination of the two rapidly decreased (P < 0.05) secretion of LH. Each agonist induced a gradual, prolonged rise in secretion of LH after the initial inhibition, but neither agonist alone nor the combined agonists was able to induce a "normal" preovulatory-like surge of LH similar to that induced by E2. Compared with E2-treated ewes, the beginning of the increase in secretion of LH occurred earlier (P < 0.01) in DPN-treated ewes, later (P < 0.05) in PPT-treated ewes, and at a similar interval in ewes receiving the combined agonist treatment. Like E2, PPT decreased (P < 0.05) secretion of FSH, but the duration of suppression was much longer in PPT-treated ewes. DPN did not alter secretion of FSH in this study. Modulation of the number of GnRH receptors by PPT and DPN was examined in primary cultures of ovine pituitary cells. In our hands, both PPT and DPN increased the number of GnRH receptors, but the dose of DPN required to stimulate synthesis of GnRH receptors was 10 times higher than that of PPT. We conclude that in OVX ewes: 1) ESR1 and ESR2 mediate the negative feedback of E2 on secretion of LH at the level of the pituitary gland, 2) ESR1 and ESR2 do not synergize or antagonize the effects of each other; however, they do interact to synchronize the beginning of the stimulatory effect of E2 on secretion of LH, 3) ESR1 and ESR2 may mediate at least partially the positive feedback of E2 on LH secretion by increasing the number of GnRH receptors, and 4) only ESR1 appears to be involved in the negative feedback of E2 on secretion of FSH.     

Pulsatile GnRH administration stimulates the number of pituitary GnRH receptors in seasonally anoestrous ewes

Twenty seasonally anoestrous ewes were pretreated with progesterone for 4 days and divided into four equal groups. Ewes in Group 1 received no GnRH treatment and were slaughtered immediately after progesterone removal. Ewes in Groups 2, 3 and 4 received i.v. injections of 250 ng GnRH every 2 h for 36 h starting at the time of progesterone removal. Ewes in Group 2 were slaughtered immediately after the 36 h GnRH pulsing, while ewes in Groups 3 and 4 were given a bolus injection of 125 micrograms GnRH at this time and were slaughtered 2 and 10 h after the bolus injection, respectively. Blood samples were collected every 30 min from ewes in Group 4 only, from 4 h before the start of GnRH treatment until 10 h after the bolus injection. Pulsing with GnRH resulted in episodic release of LH, and the bolus injection of GnRH was immediately followed by a preovulatory type LH surge in those ewes in which an endogenous surge had no already begun. The pituitary GnRH receptor numbers were significantly higher for the ewes in Group 2 than for any of the other treatment groups, while there was no significant difference in the receptor numbers between Groups 1, 3 and 4. The results suggest an up-regulation of GnRH receptors resulting from pulsatile GnRH therapy.     

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)     

Negative feedback as an obligatory antecedent to the estradiol-induced luteinizing hormone surge in ovariectomized pigs

In ovariectomized pigs, estradiol treatment induces a preovulatory-like luteinizing hormone (LH) surge, but only after serum LH concentrations are suppressed for 48 h. This inhibition of LH release is attributable in large part to inhibition of gonadotropin-releasing hormone (GnRH) release. The present report examines the dependency of the estradiol-induced LH surge on this preceding phase of negative feedback. Ten ovariectomized gilts were given an i.m. injection of estradiol benzoate (10 micrograms/kg BW). Beginning at the time of estradiol treatment, 5 of these gilts received 1-microgram GnRH pulses i.v. every 45 min for 48 h, i.e. during the period of negative feedback. The remaining 5 control gilts received comparable infusions of vehicle. Estradiol induced the characteristic biphasic LH response in control gilts. On the other hand, the inhibitory LH response to estradiol was prevented and the ensuing LH surge was blocked in 4 of the 5 gilts given GnRH pulses during the negative feedback phase. These results indicate that suppressing release of GnRH and/or LH is an important antecedent to full expression of the LH surge in ovariectomized pigs. Assimilation of this observation with the existing literature provides novel insights into the neuroendocrine control of LH secretion in castrated and ovary-intact gilts.     

No evidence for pituitary priming to gonadotropin-releasing hormone in relation to luteinizing hormone (LH) secretion prior to the preovulatory LH surge in ewes

The purpose of this study was to determine the occurrence of and the regulatory mechanisms involved in priming of the pituitary to GnRH before the preovulatory LH surge in sheep. Experiment 1: Forty-two ewes had progestagen devices removed after 14 days and were assigned to luteal (Lut) or follicular (Foll) groups. Fifteen days later, blood sampling was initiated either immediately or 36 h after induced luteolysis in groups Lut and Foll, respectively. After 4 h, ewes were administered either saline (n = 5) or 250 ng (n = 8) or 10 microg (n = 8) of GnRH. Five ewes per treatment group were killed 1 h later, while remaining animals were blood sampled for a further 7 h. Experiment 2: Eighteen ewes were allocated to Lut and Foll groups (described above). Blood samples were collected from 2 h before GnRH (10 microg) treatment until 7 h after. Despite up-regulated GnRH-R mRNA levels in Foll ewes, pituitary content and plasma levels of LH and LHbeta mRNA levels were similar between groups. Mean FSHbeta mRNA and plasma FSH levels were elevated in Lut ewes but declined after GnRH treatment. Inversely, plasma estradiol and inhibin-A concentrations were higher in Foll ewes and declined after GnRH treatment. Fewer LH(+ve)/secretogranin II(-ve) (SgII(-ve)) granules were present in gonadotropes of Foll ewes, coincident with increased basal LH levels. Fewer smaller sized granules were present after GnRH treatment. In conclusion, there was no evidence of self-priming before onset of the preovulatory LH surge. Constitutive release of LH(+ve)/SgII(-ve) granules may maintain basal LH levels while smaller sized, presumably mature granules may be preferentially released after GnRH stimulation.     

Roles of estradiol and gonadotropin-releasing hormone in controlling negative and positive feedback associated with the luteinizing hormone surge in ovariectomized pigs.

Ovariectomized gilts (n = 63) were given estradiol benzoate (estradiol), antiserum to neutralize endogenous GnRH, and pulses of a GnRH agonist (GnRH-A) to stimulate release of LH. GnRH-A was given as 200-ng pulses hourly from 0 to 54 h and as 100- or 200-ng pulses every 30 or 60 min from 54 to 96 h after estradiol. Estradiol alone suppressed LH from 6 to 54 h and elicited an LH surge that peaked at 72 h. When GnRH-A was given every 30-60 min from 0 to 96 h, estradiol suppressed LH for 6-12 h, but then LH returned to pre-estradiol concentrations. When pulses of GnRH-A were given only between 54 and 96 h after estradiol, the surge of LH was related positively to dose and frequency of GnRH-A. We conclude that 1) estrogen acts at the hypothalamus to inhibit release of GnRH for 54 h and then causes a synchronous release of GnRH; 2) estrogen acts at the pituitary to block its response to GnRH for 6-12 h and enhances the accumulation of releasable LH; and 3) magnitude of the LH surge is dependent on the amount of GnRH stimulation.     

Dynamic changes in the gonadotrope cell subpopulations during an estradiol-induced surge in the ewe

Whether estradiol targets a subpopulation of gonadotrope cells was investigated in this study. Ovariectomized ewes (OVX) or OVX ewes immunized against GnRH and treated with hourly pulses of GnRH analogue (OVX-IMG) were killed at 6, 12, 16, and 24 h after administration of 50 microg of 17beta-estradiol (E(2)). Control ewes received no E(2) treatment. In OVX or OVX-IMG ewes killed 6 h after E(2) injection, a decrease in gonadotropin plasma levels was observed compared with non-E(2)-treated ewes. In contrast, a surge in gonadotropin plasma concentrations occurred in ewes killed 16 h after injection. The percentage of total immunoreactive gonadotrope cells among the pituitary cells was lower in E(2)-treated ewes compared with nontreated animals. The proportion of monohormonal LH cells was constant throughout the experiment, except at the surge peak, where it was enhanced. In the OVX ewes, the proportion of bihormonal LH/FSH cells was lower in the E(2)-treated ewes compared to the nontreated ewes (P: < 0.001), with a more pronounced decrease 16 h after E(2) injection. A slight increase occurred 12 h after E(2) injection compared with 6 h after injection (P: < 0.05). A similar pattern was observed in the OVX-IMG ewes, except at 12 h after E(2) injection, when no increase occurred. In both OVX and OVX-IMG ewes, injection of E(2) decreased FSHbeta mRNA expression but did not alter the relative levels of LHbeta mRNA. These data suggest that the negative feedback of E(2) on LH and FSH secretion mainly targets the bihormonal cells and occurs, at least in part, directly at the pituitary level. During the gonadotropin surge, the sustained FSH release from the bihormonal cells would induce a switch from bihormonal cells to monohormonal LH cells by depleting these cells of FSH.     

Effects of chronic treatment with a GnRH agonist on oestrous behaviour and on the secretion of LH and progesterone in the ewe

A study was carried out to investigate a novel approach to oestrus synchronization in the ewe by treatment with a gonadotrophin releasing hormone (GnRH) agonist. Groups of ewes were initially treated on Day 2, 10 or 14 of the oestrous cycle with 10 mug GnRH analogue (D-Ser(Bu(t)) 6 des Gly GnRH ethylamide) per ewe per day for 14 days. Behavioural oestrus was inhibited during GnRH agonist treatment and recurred from 8 to 38 days after the treatment in an unsynchronized manner. Luteal activity during treatment was not impaired but reduced progesterone concentrations occurred in cycles after the treatment. The rhythm of ovarian function, generally characterized by prolonged follicular development, was impaired. During the treatment and subsequent recovery period, integrity of pituitary function was examined by measuring luteinizing hormone (LH) after GnRH agonist was injected, and after stimulation test doses of 150 ng natural GnRH were administered. During treatment there was, with time, a decline in pituitary response to the agonist which suggested that pituitary release of LH was exhausted. After the 14-day treatment the stimulation test with GnRH revealed a gradual return to normal responsiveness although this was not complete three weeks after the treatment when compared to control ewes. This lowered pituitary activity could cause the impaired ovarian function.     

Does the seasonal increase in estradiol negative feedback prevent luteinizing hormone surges in anestrous ewes by suppressing hypothalamic gonadotropin-releasing hormone pulse frequency?

To test the hypothesis that the anestrous increase in estradiol negative feedback prevents estrous cycles by suppressing hypothalamic gonadotropin-releasing hormone (GnRH) pulse frequency, a variety of regimens of increasing GnRH pulse frequency were administered to anestrous ewes for 3 days. A luteinizing hormone (LH) surge was induced in 45 of 46 ewes regardless of amplitude or frequency of GnRH pulses, but only 19 had luteal phases. Estradiol administration induced LH surges in 6 of 6 ewes, only 3 having luteal phases. Anestrous luteal phase progesterone profiles were similar in incidence, time course, and amplitude to those of the first luteal phases of the breeding season, which in turn had lower progesterone maxima than late breeding season luteal phases. In the remaining ewes, progesterone increased briefly or not at all, the increases being similar to the transient rises in progesterone occurring in most ewes at the onset of the breeding season. These results demonstrate that increasing GnRH pulse frequency induces LH surges in anestrus and that the subsequent events are similar to those at the beginning of the breeding season. Finally, they support the hypothesis that the negative feedback action of estradiol prevents cycles in anestrus by suppressing the frequency of the hypothalamic pulse generator.     

Prepubertal changes in the hypothalamic-pituitary axis of Holstein bulls

We investigated the nature and sites of changes in the hypothalamic-pituitary axis associated with the onset of high-frequency, high-amplitude discharges of luteinizing hormone (LH) in young bulls during the transition from the infantile to the prepubertal phase of development. Blood serum and neuroendocrine tissues from bulls killed at 1, 6, 10, 14, or 18 wk of age were evaluated. Concentrations of LH in serum from bulls 1 or 6 wk old averaged less than 0.25 ng/ml and only one episodic discharge of LH was detected for 10 bulls. At 10, 14, or 18 wk, 14 of 15 bulls had episodic discharges of LH. Concentrations of testosterone in serum were progressively higher at 10, 14, and 18 wk, but the concentration of estradiol was maximal at 6 wk. The concentrations of gonadotropin-releasing hormone (GnRH) in the anterior hypothalamus, posterior hypothalamus, or median eminence were not influenced by age. However, concentration of GnRH receptors in the anterior pituitary gland increased 314% between 6 and 10 wk and the concentration of LH increased 67%. Between 6 and 10 wk, concentrations of estradiol receptors in the anterior and posterior hypothalamus declined by 68% and 46%, but the concentration of estradiol receptors in the anterior pituitary gland increased by 103%. For most characteristics, there was no major change between 10 and 18 wk. We postulate that between 6 and 10 wk of age, there is 1) removal of an estradiol-mediated block of GnRH secretion and 2) an estradiol-mediated, and possibly GnRH-mediated, increase in pituitary GnRH receptors. Together, these changes result in greatly increased stimulation of the anterior pituitary gland by GnRH between 6 and 10 wk of age and stimulation of the discharges of LH characteristic of bulls in the early prepubertal phase of development.