Hypothalamic site of progesterone action on gonadotropin release

Tonic gonadotropin secretion was monitored at 20 min intervals for a total of 9 hours in 3 female volunteers during the mid-luteal phase of an ovulatory cycle. This control period was followed by repeated LH-RH stimulation (12 micrograms LH-RH as i.v. bolus once every hour for another 5 hours). During the control period spontaneous albeit low-frequent pulsatile secretion was observed for LH (a pulse occurring once every 3-8 hours) but not for FSH. While intermittent exogenous LH-RH stimulation was being performed at circhoral LH-RH pulse frequency pulsatile gonadotropin release was established at synchronous episodicity and systemic gonadotropin levels consecutively increased. These data provide indirect evidence that the pituitary gland is not rendered refractory to LH-RH by luteal progesterone secretion but readily responds to LH-RH stimuli even when these simulate a follicular phase LH-RH pulse frequency. Thus, it is concluded that spontaneous pulsatile LH release at low frequency during the luteal phase of the cycle reflects low frequent LH-RH discharges from the hypothalamus. Underlying mechanisms are discussed.     

Negative feedback effects of chlormadinone acetate and ethynylestradiol on gonadotropin secretion in patients with prostatic cancer and male rats

Serum LH and FSH levels were determined before and after LH-RH injection (100 micrograms, i.m.) in patients with prostatic cancer who were chronically treated with either chlormadinone acetate (CMA, 100 mg/day) or ethynylestradiol (EE, 1 mg/day). In patients treated with EE, the levels of serum LH and FSH before and after injection of LH-RH were significantly lower than those in controls. On the other hand in patients treated with CMA, the basal levels of serum gonadotropins did not differ from those in controls, and the increase in gonadotropin after LH-RH injection was comparable to that in controls. To examine the effects of these steroids on the hypothalamo-hypophysial axis in the regulation of gonadotropin secretion, CMA or EE was implanted in castrated male rats. CMA, EE or cholesterol (control) was implanted in the hypothalamic median eminence-arcuate nucleus region through a stainless doublecannula. EE implantation resulted in a 75% decrease in serum LH (p < 0.001) and a 38% decrease in serum FSH (p < 0.05) from the control levels on day 5 of implantation. On the other hand, CMA implantation induced a 33% decrease in serum LH (p < 0.05) from the control level on day 3 of implantation, but no significant change in serum FSH levels. The injection of 2 micrograms/kg of LH-RH on day 7 of implantation induced significant lowering of LH and FSH levels. There was no significant difference between serum levels of the hormones 20 min after LH-RH injection for these two groups and those for the control group. These studies suggest that EE has a potent negative feedback effect on both LH and FSH secretion, and that CMA has a mild negative feedback effect on LH secretion.     

Stimulation of the estrogen synthesizing system of the immature rat ovary by exogenous and endogenous gonadotropins

Immature rat ovaries increase their secretion of estradiol (E₂) when stimulated by gonadotropins but only after a lag period of several hours. Once established, estrogen secretion can be maintained, or increased, by the continued presence of gonadotropin. A combination of ovine FSH+LH given at 2 hr intervals stimulated the estrogen synthesizing system ($\text{ESS}$) of the ovary and serum E₂ showed a pronounced rise between 16 and 20 hrs after the initial injection. When given every 2 hrs for 5 doses (0–8 hrs) serum E₂ was undetectable. However, it was increased if 20 IU PMS was injected at the time of the last dose of FSH+ LH. Endogenous FSH&LH, increased by hourly injections of LH-releasing hormone for a period of 8 hrs, stimulated the $\text{ESS}$; serum E₂ increased at the expected time when this treatment was followed by an injection of PMS.Anti-PMS antiserum given 12 hrs after PMS, prevented the expected rise in serum E₂ at 24 hrs. However, FSH, LH or a combination of the two given every 2 hrs beginning at the time of the anti-PMS produced an increase in E₂ secretion; the combination was more effective than either hormone alone.These results are consistent with the interpretation that a combined FSH-LH action is responsible for induction of the $\text{ESS}$ in the immature rat ovary. The combination of hormones is also very effective in maintaining estrogen secretion but some function appears possible with FSH or LH alone.     

Self-priming effect of LH-RH on pituitary gonadotropins in hyperprolactinemic women

To investigate how various concentrations of serum prolactin (PRL) influence the priming effect of luteinizing hormone releasing hormone (LH-RH) on the pituitary gland, 24 women with various blood PRL concentrations received intravenous injections of 100 micrograms of synthetic LH-RH twice at an interval of 60 minutes and their serum LH and follicle-stimulating hormone (FSH) were measured and analysed. In the follicular phase with a normal PRL concentration (PRL less than 20 ng/ml, n = 6), marked first peaks of the two hormones following the first LH-RH stimulation and enhanced second peaks after the second LH-RH administration were observed, indicating a typical priming effect of LH-RH on gonadotropins, though the second response of FSH was more moderate than that of LH. In hyperprolactinemia, in which the serum PRL concentration was higher than 70 ng/ml (n = 13), the basal concentration of gonadotropins was not significantly changed but the priming effect of LH-RH on LH and FSH was significantly decreased (p less than 0.01). No marked second peaks of LH and FSH were observed, suggesting an inhibitory effect of hyperprolactinemia on the second release of LH and FSH. In contrast, this effect was restored in a group of women whose serum PRL concentration was between 30 and 50 ng/ml (n = 5). Furthermore, enhanced second peaks of both LH and FSH were noted after successful bromocriptine therapy reduced hyperprolactinemia (PRL greater than 70 ng/ml) to less than 25 ng/ml (n = 5).(ABSTRACT TRUNCATED AT 250 WORDS)     

Hormonal induction of ovulation stimulates atresia of antral follicles in a vespertilionid bat, Scotophilus heathi

Scotophilus heathi is a seasonally monoestrous subtropical vespertilionid bat found at Varanasi, India. Although the antral follicles remain present in the ovaries of S. heathi from November till March, ovulation is delayed in this species until early March. In order to understand the mechanism of ovulation suppression during this period of delayed ovulation, the effects of human chorionic gonadotropin (hCG), pregnant mare's serum gonadotropin (PMSG), follicle stimulating hormone (FSH) and gonadotropin releasing hormone agonist (GnRH agonist) on ovarian morphology and steroid concentration were investigated. Hormonal treatments were given as a single i.p. dose 24 h after capture. The bats were sacrificed 48 h after the injection. Treatment with hCG, PMSG, FSH and GnRH agonist failed to induce ovulation in S. heathi, although these hormones produced a high degree of ovarian stimulation. The administration of hCG and PMSG induced ovarian enlargement, intense hyperemia, marked changes in the interstitial cells (ICs), development of several antral follicles and a varying degree of abnormalities in the oocytes of most of the antral follicles. In the bats treated with hCG, PMSG and GnRH agonist, androstenedione concentration increased significantly to extraordinarily high levels, whereas estradiol concentration decreased. Administration of FSH caused regression of ICs and pyknosis of granulosa cells in the majority of antral follicles. FSH did not enhance androstenedione concentration. The results of the present study suggest that the failure of hormonal treatments to induce ovulation during the period of delayed ovulation might be due to a seasonal desensitization of ovarian follicles in S. heathi. The hormonal treatment instead stimulated the ICs to produce a high level of androstenedione resulting in atretic changes of the antral follicles.     

Inactivation of follicle-stimulating hormone by a factor in a bovine serum albumin preparation

Bovine serum albumin (BSA) preparations are commonly employed as "carrier" or "protective" proteins in the solutions used to dissolve gonadotropin preparations. The present report describes a BSA preparation that was found to contain a factor that inactivated follicle-stimulating hormone (FSH). Four different BSA preparations (designated BSA1, BSA2a, BSA2b, BSA3) were studied. The FSH preparation (NIH-FSH-S16) was dissolved in 0.15 M NaCl, containing the various BSA preparations. The FSH solutions were injected subcutaneously, twice daily, for 5 days into hypophysectomized immature female rats bearing estrogen capsules. Twenty-four hours after the last injection, the rats were decapitated, and the ovaries were removed, trimmed and weighed. The FSH preparation produced ovarian weight gain when BSA1, BSA2b, or BSA3 was used, but not when BSA2a was used in the vehicle. In animals injected with the FSH dissolved in BSA1 vehicle and injected at a separate site with BSA2a solution, the FSH preparation was fully active, which indicates that contact of the BSA2a preparation with FSH was required for the inactivating factor to be operant. Indeed, after incubation in BSA2a solution, the radiolabeled FSH preparation exhibited a slight decrease in apparent molecular size when chromatographed on a Sephadex G-100 column. This result suggests that the BSA2a preparation contained a factor that may have inhibited FSH by degrading it.     

Prolonged intravenous infusions of LH-releasing hormone into normal men.

Five normal men received constant intravenous infusions of luteinizing hormone-releasing hormone (LH-RH), 0.2 mug/min, for 14-19 hours. Serum levels ofluteizining hormone (LH) revealed a biphasic pattern of increase, reaching maximal values by 4 hours after the infusions began, then remained near that level until the infusions ceased. Serum follicle stimulating hormone (FSH) levels rose gradually to maximal values by 6-13 hours and maintained this level until the end of the infusions. Testosterone (T) levels revealed gradual increases throughout the infusions. These results confirm an increase in serum T levels with prolonged endogenous gonadotrophin stimulation. This is in contrast to the inability of several previous studies to demonstrate an increase in T levels following the relatively short gonadotrophin elevation produced by single-shot LH-RH administration. The T increases produced, however, were quantitatively much less than those reported during prolonged LH-RH infusions in rams, suggesting that the human testis is less responsive to endogenous gonadotrophin stimulation than is that of the ram. In addition, prolonged LH-RH stimulation did not cause pituitary refractoriness in men as has been described in animals.     

Neonatal release of gonadotropins is essential for development of ovarian follicle-stimulating hormone receptors

In the rat, ovarian follicle-stimulating hormone (FSH) receptors increase markedly during the first two postnatal weeks, when serum gonadotropin levels are most elevated. This study was conducted to evaluate the hypothesis that these high gonadotropin levels, and in particular FSH, are involved in the acquisition of FSH receptors by the developing ovary. Gonadotropin release was suppressed by administration of several non-aromatizable androgens, among which dihydrotestosterone propionate (DHTP) was the most effective. In one series of experiments the steroids were administered from Days 5 to 11, and serum FSH and luteinizing hormone (LH) were measured on Day 12. Surprisingly, FSH receptor content was greater in rats with suppressed serum gonadotropins than in controls. The greatest increase in available receptors was observed in DHTP-treated rats in which serum FSH was reduced to 20% of control values and LH suppressed to undetectable values. DHTP failed to directly increase available FSH receptors in hypophysectomized immature rats. Magnesium chloride (MgCl2) treatment of ovarian membranes removed bound 125I-hFSH by 87% without affecting receptor viability. Exposure of control 12-day-old ovaries to MgCl2 increased available FSH receptors to a level similar to that of ovaries from DHTP-treated rats not exposed to MgCl2, suggesting that more receptors were available in DHTP-treated rats because serum FSH was suppressed. Earlier initation of DHTP treatment (postnatal Day 1) suppressed serum FSH and LH to undetectable values by Day 5 and decreased FSH receptor content below control values by Day 12. MgCl2 treatment only slightly increased available receptors in these DHTP-treated animals.(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in concentration of serum LH and FSH associated with estrogen-advanced ovulation in 4-day cyclic rats

Changes in concentration of serum luteinizing hormone (LH) and follicle stimulating hormone (FSH) associated with estrogen-advanced ovulation in 4-day cyclic rats were determined. 50 mcg of estradiol benzoate was administered sc on the 1st day of diestrus and LH and FSH were determined by radioimmunoassay 24, 29, and 47 hours postinjection. 48 hours postinjection, ovulation was determined by the presence of ova in the oviduct. Estradiol benzoate caused a significant (p less than .01) increase in serum LH after 24 and 29 hours and a drastic decrease after 47 hours postinjection. FSH increased (p less than .05) after 29 hours postinjection. These results suggest that an ovulatory gonadotropin surge occurs 24 hours ahead of schedule in estradiol-benzoate-treated rats.     

Site of negative feedback action of estrogen on gonadotropin secretion in normal women

We have reported that iv administration of conjugated estrogens results in no significant change in the plasma LH-RH level during the negative feedback phase of LH, suggesting that estrogen does not suppress LH by decreasing hypothalamic LH-RH. To determine the site of estrogen action during the negative feedback phase, we studied the pituitary response to a small amount of LH-RH after estrogen administration in normal cyclic women in the mid-follicular phase. The pituitary responses to an iv bolus of 2.5 micrograms of synthetic LH-RH were evaluated by measuring serum LH and FSH 2 h before and 8 h after administration of 20 mg of conjugated estrogens (Premarin). The mean levels of serum LH and FSH were significantly (p less than 0.05) decreased 8 h after the injection. The peak responses of LH and FSH to LH-RH were also significantly (p less than 0.05) reduced after Premarin administration. These findings suggest that the negative feedback effect of estrogen on gonadotropin secretion is caused by its direct suppression on the pituitary response to LH-RH.     

Inhibitory control of the pituitary LH secretion by LH-RH in male rats.

Luteinizing hormone-releasing hormone (LH-RH) was administered to prepubertal male rats (intact, castrate or castrate-adrenalectomized, 60 g body weight) for 28 days (1 microgram LH-RH/day, s.c.), at a 10-fold physiological dose, as compared to the minimal FSH-releasing dose of 100 ng/rat s.c. In intact rats, serum LH and weight of androgen-dependent organs (vented prostate, seminal vesicles) were reduced after 14 days of treatment. In castrate rats, the postcastration rise in serum LH was abolished by treatment. Pituitary LH content, FSH secretion and prolactin secretion were not suppressed. Hypothalamic LH-RH was increased at 14 and 21 days. In castrate adrenalectomized male rats, LH secretion was also suppressed by 1 microgram LH-RH s.c. x 28 days. The hypothalamic LH-RH content did not increase. The pituitary LH-RH receptor level was not down-regulated after 14 days treatment either in intact or castrate male rats. Pituitary inhibition (LH release) in rats by a supraphysiological dose of LH-RH given for 28 days indicates that the optimal regime for chronic treatment has to be determined by monitoring LH release at regular intervals. Direct pituitary inhibition by LH-RH may explain some of the unexpected antifertility effects observed with high doses of LH-RH.     

Basal and LH-RH-stimulated gonadotropin release after transport stress in male rats

A total of 120 male rats of the Sprague-Dawley-strain (6 weeks old) were used in this experiment. 5 groups of 12 animals each were treated intraperitoneally with 200 ng gonadotropin releasing hormone (LH-RH) per animal. 30 minutes later blood was sampled by heart puncture. Group I were animals without transport, group II immediately after, group III one day, group IV one week and group V six weeks after a standardised transport. Another 5 groups were subjected to the same protocol but received saline i.p. instead of LH-RH. Serum levels of LH and FSH were estimated by radioimmunoassay. LH and FSH serum levels could be stimulated by LH-RH in all groups. A significant rise of basal and LH-RH stimulated LH levels was observed until the first day after transport. Thereafter a drop was registered. No consistent patterns of basal as well LH-RH stimulated FSH-levels were noted. These data combine to suggest an elevation of LH-RH secretion as response to the stress. This results in a sensibilisation of the pituitary to exogenous LH-RH.     

Comparison of mammalian luteinizing hormone releasing hormone (LH-RH), and of an analog (ICI 118630), on luteinizing hormone and ovarian steroid (progesterone, oestradiol) secretions in laying hens. (Gallus domesticus)

This experiment was conducted to compare the luteinizing hormone (LH), progesterone (P4) and oestradiol (E2) release in response to injections of various doses of synthetic mammalian luteinizing hormone-releasing hormone (LH-RH) and of an LH-RH agonist, ICI 118630, administered to laying hens 4 to 9 hours after a mid-sequence ovulation. Plasma LH increased significantly within 10 minutes of injection of either compound whereas any increases in plasma steroid concentrations were discerned later, at approximately minutes post-injection. No dose-response relationship was found for either compound with respect to LH release, but ICI 118630 appeared more potent than LH-RH. This analog also produced a greater mean incremental rise in plasma progesterone, but not oestradiol, than LH-RH, and this was found in animals injected at a time when the largest ovarian follicle was not mature. These result suggest that ICI 118630 is a more potent releasing hormone in the hen at the level of the pituitary, and that it may have a stimulating effect on ovarian progesterone secretion.     

LH- and FSH response to long-term application of LH-RH analogue in normal males.

The effect of luteinizing hormone-releasing hormone (LHRH) analogue after subcutaneous application over a 12-day period was evaluated in 15 normal males to determine whether gonadotropin secretion is influenced by longterm application. After 1.25 mcg/day, LH increases were measurable 3 hours after daily receipt of the analogue. After 2.5 mcg/day, the release of LH secretion was even greater, although this stimulation was markedly reduced in the 2nd week. After 5 mcg in 2-day intervals, a clearly induced LH increase was observed on the day of administration and was maintained for 10 hours. Similar results were noted for follicle-stimulating hormone (FSH). There was a response of FSH after 1.25 mcg and 2.5 mcg, but this response decreased in the 2nd week. After application of 5 mcg every other day, there was a consistent FSH increase that subsided on the treatment-free day and continued into the 2nd week of treatment. These findings suggest that LHRH analogues should be administered in low doses subcutaneously or intranasally at daily intervals to enable a longterm effect on gonadotropin release.     

Studies on follicular growth in the immature rat and hamster: Effect of a single injection of gonadotropin or estrogen on the rate of3H-thymidine incorporation into ovarian DNAin vitro

Initiation of follicular growth by specific hormonal stimuli in ovaries of immature rats and hamsters was studied by determining the rate of incorporation of³H-thymidine into ovarian DNAin vitro. Incorporation was considered as an index of DNA synthesis and cell multiplication. A single injection of pregnant mare serum gonadotropin could thus maximally stimulate by 18 hr³H-thymidine incorporation into DNA of the ovary of immature hamsters. Neutralization of pregnant mare serum gonadotropin by an antiserum to ovine follicle stimulating hormone only during the initial 8–10 hr and not later could inhibit the increase in³H-thymidine incorporationin vitro observed at 18 hr, suggesting that the continued presence of gonadotropin stimulus was not necessary for this response. The other indices of follicular growth monitored such as ovarian weight, serum estradiol and uterine weight showed discernible increase at periods only after the above initial event. A single injection of estrogen (diethyl stilbesterol or estradiol-l7β) could similarly cause 18 hr later, a stimulation in the rate of incorporation of³H-thymidine into DNAin vitro in ovaries of immature rats. The presence of endogenous gonadotropins, however, was obligatory for observing this response to estrogen. Evidence in support of the above was two-fold: (i) administration of antiserum to follicle stimulating hormone or luteinizing hormone along with estrogen completely inhibited the increase in³H-thymidine incorporation into ovarian DNAin vitro; (ii) a radioimmunological measurement revealed following estrogen treatment, the presence of a higher concentration of endogenous follicle stimulating hormone in the ovary. Finally, administration of varying doses of ovine follicle stimulating hormone along with a constant dose of estrogen to immature rats produced a dose-dependent increment in the incorporation of³H-thymidine into ovarian DNAin vitro. These observations suggested the potentiality of this system for developing a sensitive bioassay for follicle stimulating hormone.     

Differential effects of LH-RH immunoneutralization on LH and FSH secretion in the ewe

Neutralization of LH-RH by injection of an ovine antiserum to LH-RH in ewes during the late follicular phase of the oestrous cycle resulted in an immediate blockade of pulsatile secretion of LH. Plasma concentrations of FSH gradually rose in the antiserum-treated ewes during the 36-h study period but levels declined in control ewes. These results show that, in the ewe, pulsatile LH secretion is dependent on LH-RH from the hypothalamus, while FSH is largely unresponsive to short-term reduction of LH-RH stimulation. Since reduction in LH secretion is likely to reduce ovarian function, the changes in FSH secretion may be attributed to the removal of a negative feedback influence of an ovarian factor, perhaps oestradiol, on FSH secretion.     

Ultrastructural observations of granule extrusion from rat anterior pituitary cells after injection of LH-releasing hormone

Summary In order to observe the ultrastructural aspects of granule extrusion from gonadotrophs, the authors injected pure, natural porcine LH-releasing hormone (LH-RH) into persistent-estrous (PE) rats. Clear-cut extrusions of secretory granules appeared from LH-gonadotrophs 2.5, 5 and 15 minutes after the injection. Massive extrusion was observed at 15 minutes. Serum LH and FSH were also estimated by radioimmunoassay on blood samples taken at the same times after the injections. LH was increased in the blood sera after the injection of LH-RH, but serum FSH was not significantly different among the various treated rats. The rise in serum LH after LH-RH injection was well correlated with the ultrastructural phenomenon of granule extrusion from LH-gonadotrophs.The study was supported by USPHS Grants AM 12583, AM 09094 and AM 07467.The authors acknowledge the able assistance of Mrs. Martha Castilleja in San Antonio and of Meredith M. Nickel in New Orleans.     

Validation of the mechanisms proposed for the stimulatory and inhibitory effects of progesterone on gonadotropin secretion in the estrogen-primed rat: a possible role for adrenal steroids.

The stimulatory and inhibitory effects of progesterone on luteinizing hormone (LH) and follicle-stimulating hormone (FSH) secretion were found to be dependent on the length of estrogen exposure in ovariectomized estrogen-primed rats. Progesterone suppressed LH and FSH secretion when administered 16 hours after a single injection of estradiol to ovariectomized rats. If the estradiol treatment was extended over 40 hours by two injections of estradiol 24 hours apart, progesterone administration led to a highly significant elevation of both serum LH and FSH levels 6 hours later. In addition to the direct stimulatory effect on LH and FSH release, progesterone, when injected 1 hour before, was able to antagonize the suppressive effect of a third injection of estradiol on LH and FSH release. In the immature ovariectomized estrogen-primed rat, 10 IU of ACTH brought about a release of progesterone and corticosterone 15 minutes later and LH and FSH 6 hours later. Progesterone, but not corticosterone, appeared to be responsible for the effect of ACTH on gonadotropin release. The synthetic corticosteroid triamcinolone acetonide brought about LH and FSH release in the afternoon, while cortisol, similar to corticosterone, was unable to do so. Nevertheless, triamcinolone acetonide and cortisol brought about increased secretion of FSH the following morning.     

Effect of prostaglandin F2alpha on ovarian and pituitary function in the mid-luteal phase.

The effects of PGF2alpha infusion in a dose of 25 micrograms/min for 5 hours on serum levels of estradiol-17beta, progesterone, LH, FSH, TSH and prolactin, and on the pituitary hormone responsiveness to LRH and TRH were studied in 10 apparently healthy cycling women in the mid-luteal phase. No systematic alteration was seen in the pituitary and ovarian hormone levels during PGF2alpha infusion, and the pituitary hormone responses to releasing hormones were unaffected. Ovarian steroid production increased in response to increased gonadotropin levels after LRH injection during PGF2alpha administration. These results confirm that PGF2alpha is not luteolytic in humans and no apparent relationship between PGF2alpha and pituitary hormone secretion exists.     

Familial hypothalamic hypogonadotropic hypogonadism

Clinical findings and endocrine studies are reported concerning six subjects (from two pedigrees) suffering from isolated hypogonadotropic hypogonadism. Their complete lack of any gonadotropin response to clomiphene stimulation, together with positive responses of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) to LH-releasing-hormone stimulation (LH-RH) provide evidence for a hypothalamic basis for this disorder. Additional data are presented indicating that variables such as circulating estrogen levels may influence the gonadotropin response to LH-RH.