Effects of estrogen on 3H-leucine uptake by the hypothalamus and pituitary: correlation with hypothalamic and serum LHRH and LH.

An injection of estradiol benzoate (EB, 5 microgram/rat) to ovariectomized rats suppressed serum LH levels 2, 4, 8 and 24 h later; serum LHRH levels were unaltered but the LHRH content in the medial basal hypothalamus (MBH) was significantly elevated at 24 h. EB treatment also modified the sensitivity of the pituitary to LHRH administration: a rapid decrease at 4 h followed by an augmentation in LH release at 8 and 24 h. 3H-Leucine incorporation into proteins of the pituitary and neural tissues was unaffected for 4 h following EB treatment at the time when serum LH and the responsiveness of the pituitary to LHRH were clearly depressed. However, a marked increase in 3H-leucine incorporation into the proteins of the pituitary alone occurred at 8, 16 and 24 h after EB treatment, coincident with the enhanced pituitary sensitivity to LHRH. These studies showed that estrogen promoted storage of LHRH in the MBH and alteration in protein metabolism may be an important initial step in the estrogen-induced augmentation of the pituitary responsiveness to LHRH.     

Short-term pituitary desensitization to LH-RH after pulsatile LH-RH in the ovariectomized rat: an in vivo experiment

The development of acute insensitivity of pituitary LH secretion to LH-RH after a short exposure to LH-RH is described. In the first experiment, ovariectomized (OVX), phenobarbital-pretreated rats were given pulses of LH-RH (1.25 or 6.25 ng/100 g body weight (b.w.), intravenously). In rats given 1.25 ng at time 0, 6.25 ng at 60 min, 1.25 ng at 80 min and 1.25 ng at 120 min, there was a substantial increase in plasma LH after the first two injections, no increase after the third injection and a relatively small increase after the fourth one. In other rats treated identically but not given a 1.25-ng dose at 80 min, the plasma LH rise in response to the 1.25-ng dose at 120 min was comparable to that seen after the 1.25-ng dose given at time 0. If the 1.25-ng LH-RH pulses given at times 0 and 80 min were replaced by a rat pituitary extract, the plasma LH rise in response to the 1.25-ng dose at 120 min was comparable to that seen after administration of pituitary extract. In the second experiment, OVX phenobarbital-pretreated rats were given 1.25 ng LH-RH/100 g b.w. at t = 0. They were then divided into three groups, each receiving 1.25, 3.75 or 6.25 ng LH-RH/100 g b.w. at t = 60 min. Each of these three groups was again divided into three groups which received 1.25 ng LH-RH/100 g b.w. at 80, 100 or 120 min.(ABSTRACT TRUNCATED AT 250 WORDS)     

Synthesis of LH-RH by rat hypothalamic tissue in vitro. II. Effect of short-term orchiectomy and estradiol benzoate therapy.

Normal, castrate and estradiol benzoate (EB; 1 microgram/100 g b.w. x 6d, s.c.)-treated castrate male rats were utilized to study the short term effects of these treatments upon the incorporation of 3H-glycine into LH-RH in vitro. The rise in serum LH and FSH observed in the untreated castrates was prevented by EB therapy. The treatment had no significant effect upon serum LH-RH levels or the incorporation of 3H-gly into LH-RH. EB treatment markedly (p less than 0.01) increased the hypothalamic LH-RH concentration and significantly lowered the specific activity (nCi/ng) of the 3H-gly-LH-RH (p less than 0.05). These results suggest that EB acts both on the pituitary and hypothalamus to suppress gonadotropin (Gn) release, but that, at the time period studied, neither castration or EB-treatment had any apparent effect upon hypothalamic LH-RH synthesis.     

Luteinizing hormone-releasing hormone (LH-RH) antagonist Cetrorelix down-regulates the mRNA expression of pituitary receptors for LH-RH by counteracting the stimulatory effect of endogenous LH-RH

The mechanisms through which LH-RH antagonists suppress gonadotroph functions and LH-RH receptor (LH-RH-R) production are incompletely understood. To elucidate these mechanisms, we investigated the effects of Cetrorelix on the mRNA expression of pituitary LH-RH-R and luteinizing hormone (LH) secretion in three experimental systems with different pituitary LH-RH environments. Ovariectomy induced 3.61-fold and 6.34-fold increases in the mRNA expression of pituitary LH-RH-R in rats after 11 and 21 days, respectively. After (5 h) a single injection of 100 microg Cetrorelix, no significant decrease occurred in the mRNA levels of pituitary LH-RH-R in ovariectomized (OVX) rats with high pituitary exposure to LH-RH, but there was a significant 23.2% reduction in cycling rats with normal hypophysial LH-RH environment. Prolonged treatment for 10 days with a Cetrorelix depot formulation releasing 100 microg/day decreased the concentration of mRNA for pituitary LH-RH-R by 72.6% in OVX rats, but only by 32.9% in normal rats. The decline in serum LH was 98.7% in OVX rats and 63.2% in normal rats, resulting in a minimal 0.1--0.2 ng/ml LH concentration in both groups. A continuous exposure of pituitary cells to 100 nM Cetrorelix in the superfusion system, which is devoid of LH-RH, did not cause any significant changes in LH-RH-R mRNA level. These studies demonstrate that prolonged exposure to Cetrorelix in vivo, but not in vitro, down-regulates the mRNA expression of the pituitary receptors for LH-RH. Our findings indicate that LH-RH antagonists exert their inhibitory effects on the gene expression of pituitary LH-RH-R by counteracting the stimulatory effect of endogenous LH-RH.     

Steroid priming of the luteinizing hormone response to luteinizing hormone releasing hormone.

Perifusion experiments were performed to study the stimulatory effects of luteinizing hormone releasing hormone (LH-RH) on the release of LH from anterior pituitary tissue. Exposure of pituitary tissue from normal male rats to LH-RH (5 ng/ml for 5 min) induced a small release of LH; in tissue from ovariectomized rats receiving no pretreatment, the release was more than three times greater and in tissue from gonadectomized male or female rats pretreated with oestradiol benzoate and progesterone, the release was six times greater than that observed in normal rats. Further exposure of pituitary tissue from gonadectomized steroid-pretreated male and female rats to LH-RH (5 ng/ml) induced an increase in the level of LH even greater than that seen after the initial exposure (priming action of LH-RH); in tissue from ovariectomized rats receiving no pretreatment, less LH was released than after the first exposure to LH-RH and in tissue from normal male rats the response was unchanged.     

Sexual differentiation of positive feedback: effect of hour of castration at birth on estradiol-induced luteinizing hormone secretion in immature male rats

In the male rat, a dramatic increase in hypothalamic testosterone and estradiol concentrations occurs during the first few hours of postnatal life. These experiments sought to determine whether such increases participate in the defeminization of positive estrogen feedback effects on LH secretion. Newborn male rats were castrated either in utero (0 h males), or 10 or 24 h after birth. Some males were castrated at 0 h in utero and injected at the time of surgery with 1,2.5, or 5 micrograms testosterone propionate. A group of females was ovariectomized at 0 h in utero (0 h females). The control group consisted of male and female rats sham gonadectomized at 0 h in utero which were either gonadectomized at 21 days of age or left intact. The experimental groups were challenged before puberty to determine if estrogen induced a release of LH using two different types of estrogen treatment. The first treatment consisted of an injection of 0.2 microgram estradiol benzoate (EB) on day 28 followed by a second 10 micrograms injection of EB on day 29. This treatment resulted on the afternoon of day 30 in a surge of LH in intact females. Normal males, 0 h males, or females castrated at 21 days did not have a significant LH surge. The second test consisted of the daily injection of 0.05 microgram EB on days 23-27; on day 28 the rats were injected with 2.5 micrograms EB. Zero hour male and female rats showed a large LH surge on the afternoon of day 29; sham castrated males never responded to this treatment. No sex difference was observed in the mean size of the LH surge providing the males were castrated at 0 h in utero. The effect of the hour of castration on the day of birth also was studied. Males castrated at 10 or 24 h after birth showed either no LH surge or the magnitude of the surge was greatly reduced compared to that obtained in the 0 h males (P less than 0.001). The fact that 0 h males injected with 1 microgram testosterone propionate never showed an LH surge after prepuberal treatment with estrogen suggests that 0 h is a time during which the newborn is sensitive to the defeminizing effect of androgens. These results are consistent with the idea that the testicular hyperactivity which occurs at the time of birth could influence the defeminization of the LH surge mechanisms.     

Sexual differences in the serotonergic control of prolactin and luteinizing hormone secretion in the rat

The effects of serotonin on PRL and LH release were investigated in female and male rats under different experimental conditions. At a dose of 5 mg/kg ip, serotonin increased serum PRL titers in intact males and in females during diestrus and estrus; the levels attained in the male rats were much higher than in the females. At a lower dose (2.5 mg/kg) the PRL-releasing effect of serotonin was only evident in male rats. Thus, we chose this dose for the following experiments to investigate the apparent sexual difference. To evaluate the importance of the hormonal status characteristic of male and female in conditioning the serotonin effect, an experiment was performed in gonadectomized rats, untreated or treated with estradiol benzoate (EB), or testosterone propionate (TP). In the three hormonal conditions the sexual difference was maintained: serotonin released PRL in males and failed to do so in females. However, if males were castrated within 24 h of birth, and females androgenized by a single perinatal injection of TP, the sexual difference in adulthood were reversed; thus, androgenized females responded to serotonin and males castrated at birth failed to do so. These results suggest that a male differentiated brain is more sensitive to the PRL-releasing effect of serotonin, irrespective of the hormonal environment of the rat. On the other hand, serotonin increased serum LH in female rats in estrus and in adult ovariectomized rats treated with EB; but not in females in diestrus or in ovariectomized rats, treated with TP or untreated. Neither did it modify serum LH titers in male rats whether intact, orchidectomized, or orchidectomized plus steroids. However, if male rats were castrated a few hours after birth and then treated in adulthood with EB, serotonin effectively released LH. Thus, two components, estradiol and a feminine differentiated brain, may be necessary for the facilitatory action of serotonin on LH release. Since no sex differences were observed in the increase of serum serotonin after the injection of 2.5 mg/kg of the drug, it can be discounted that the differences described for the endocrine effect of the drug could be due to different levels of circulating indolamine achieved in male and female rats. Taken together, our results indicate that serotonergic control of anterior pituitary secretion is sexually differentiated and that it presents individual characteristics for PRL and LH release.     

The effect of systemic administration of dopamine and apomorphine on Plasma LH and prolactin concentrations in conscious rats.

The effect of systemic administration of various doses of dopamine (DA) and apomorphine (APM) on plasma gonadotropin and prolactin (Prl) concentrations in ovariectomized (OVX) as well as in ovariectomized, estrogen-progesterone (OEP)-primed rats bearing indwelling jugular venous catheters was evaluated. Intravenous (i.v.) infusion or pulse injection of 0.9% NaCl had no significant effect on plasma titers of LH or Prl. I.v. infusion of DA at 4 micrograms/kg-b.w./min induced a progressive increase in circulating LH concentration in OEP rats while infusion at a similar dose in OVX animals had no effect on plasma LH. I.v. injection of 100 micrograms DA or APM significantly increased LH at 15 min in OVX rats. Similarly, in OEP rats 100 micrograms of DA elevated plasma LH at 30 and 90 min while APM induced a significant elevation of plasma LH at 15 min after injection. In OVX rats injection of DA i.p. at a dose of 5 mg/kg-b.w. did not alter plasma LH levels, but a dose of 50 mg/kg-b.w. produced a significant reduction in plasma LH concentration. APM injected i.p. at either 5 or 50 mg/kg-b.w. doses was nearly equally effective in lowering plasma LH and the suppressive effect was significantly greater than with similar doses of DA. A single injection of LH-RH (100 ng in 0.2 ml of 0.9% NaCl) in animals pretreated 15 min earlier with an effective dose of APM (5 mg/kg-b.w.) produced a peak increase in LH titers 15 min after injection. The increment in plasma LH following LH-RH in APM-treated rats was comparable to that in rats which had received saline instead of APM. Prl levels were significantly lowered by each dose of DA and APM in OVX as well as in OEP rats. There was no significant change in plasma FSH titers induced by either drug in any of the experiments. It is concluded that DA may have different actions depending upon the dose and the endocrine state of the animal. Thus, i.v. infusion of low doses of DA in OEP animals or by pulse injection in both OVX and OEP rats can elevate plasma LH by activating the release of LH-RH from the hypothalamus, while large doses of DA in OVX animals may suppress the release of LH-RH.     

Developmental changes in pituitary responsiveness to luteinizing hormone-releasing hormone (LHRH) in the female rat: ovarian-adrenal influence during the infantile period.

Pituitary LH and FSH repsonses to synthetic LHRH as estimated by increases in plasma FSH and LH 15 and 45 min following its iv injection were enhanced during the first 2 weeks of life, reaching a maximum around day 10-15 and declining thereafter. No AM.-PM. variations in pituitary responsiveness were observed at any age studied. The increased pituitary response found in infantile rats did not appear to be caused by a slower rate of disappearance of LHRH in blood of the younger animals. Ovariectomy-adrenalectomy. (Ovx-Adrx) or Ovx at day 10, but not Adrx alone, resulted in elevated LH and FSH levels 5 days later and almost complete obliteration of the FSH response to LHRH. The LH response was not altered. Treatment with 5alha-dihydrotestosterone (DHT) but not with estradiol benzoate (EB) or testosterone propionate (TP) suppressed the post-Ovx-Adrx rise in plasma LH and FSH. Progesterone (P) potentiated the effect of DHT. Restoration of basal plasma LH and FSH levels (by DHT and/or P) restored FSH responsiveness to exogenous LHRH. EB and TP were ineffective. The LH response was slightly depressed by EB + DHT. It is concluded that the elevated plasma FSH levels in the infantile female rat may be due at least in part to a high degree of pituitary responsiveness to LHRH and/or FSH-RF brought about by steroidal signal of ovarian origin. DHT and P appear to be the steroids responsible for such a stimulatory action.     

Relation of gonadal hormones to differential LH response to naloxone in prepubertal male and female rats

Naloxone (NAL) has been shown to induce LH release in female but not in male rats 10-25 days of age. The purpose of this study was to examine the role of neonatal gonadal hormones on NAL-induced LH release in male and female rats 15, 25, and 35 days of age. On each of these days rats received a s.c. injection of either NAL (5 mg/kg) or physiological saline, and blood was collected 30 min later by decapitation. At 15 days of age, NAL induced LH release in intact and ovariectomized (OVX) female rats, and in male rats castrated (CAST) on the 1st day of life (neonate CAST males). Injection of 10 micrograms estradiol benzoate (EB) 24 h prior to NAL administration blocked NAL-induced LH release in these rats. NAL had no effect on LH release in 15- or 25-day-old intact and CAST male rats or in female rats given 2 mg testosterone propionate at 3 days of age (androgenized female rats). At 35 days of age, NAL induced LH release in intact, OVX, and OVX-EB treated female rats, and in neonate CAST and neonate CAST-EB treated male rats. NAL had no effect on serum LH levels in androgenized female rats. NAL induced LH release in intact and CAST 35-day-old male rats, but pretreatment with estrogen prevented NAL from eliciting LH release. These results indicate that neonatal exposure to androgen is responsible for the sex difference in the LH response to NAL observed in prepubertal male and female rats before 30 days of age.(ABSTRACT TRUNCATED AT 250 WORDS)     

Estrogen inhibits luteinizing hormone (LH), but not follicle-stimulating hormone secretion in hypophysectomized pituitary-grafted rats receiving pulsatile LH-releasing hormone infusions

The differential feedback actions of estrogen (E2) on gonadotropin secretion were studied by means of an in vivo isolated pituitary paradigm. Adult female rats were hypophysectomized (hypox) and the next day received single anterior pituitary transplants (graft) under the kidney capsule. At the same time rats underwent bilateral ovariectomy. On the third day each animal was fitted with a catheter system which allowed for intermittent infusions of LHRH (250 ng/5 min.h) and chronic blood sampling. Rats received LHRH infusions for 7 days. On the sixth day of LHRH infusions blood samples were collected for 4 h 5, 15, 25, 35, 45 min after each hourly LHRH pulse. After 1 h of sampling, animals received sc injections of 2 micrograms estradiol benzoate (EB; n = 5) or oil vehicle (n = 5). Plasma LH, FSH, E2, and PRL levels in samples from all groups were determined by RIA. In hypox/graft rats LH release, but not FSH release, was pulsatile in response to the hourly LHRH infusions. Injection of EB in the hypox/graft rats significantly (P less than 0.05) suppressed LH release within 3 h by 57%, while FSH was unaffected. PRL levels were elevated by approximately 10-fold in the hypox/graft animals compared to those in pituitary-intact rats. These levels, however, were not changed as a function of steroid treatment and, therefore, could not account for the effects of EB on LH secretion. On the basis of these observations we conclude that 1) a major inhibitory effect of an acute injection of EB on LH secretion is exerted by a direct action on pituitary gonadotropes, and 2) E2 can differentially affect the release of LH and FSH by an intrapituitary mechanism. It is hoped that development of this model will allow for further investigation of the cellular mechanisms that mediate feedback actions of E2 on pituitary gonadotropes exposed to intermittent LHRH stimulation.     

The effect of short-term starvation on pituitary and plasma LH,plasma estradiol and progesterone,and on pituitary response to LH-RH in the laying hen (Gallus domesticus)

The effect of starvation on pituitary and plasma LH, plasma estradiol and progesterone levels, and pituitary response to LH-RH in the laying hen were studied. Deprivation of food for 7 days prevented egg laying and significantly decreased plasma LH, estradiol, and progesterone concentrations, but did not significantly change pituitary LH concentration. Resumption of feeding raised plasma LH and progesterone to their initial levels, but increased plasma estradiol to a higher level than its initial level within 3–4 days of refeeding. A single injection of 20 μg LH-RH to intact laying hens before starvation significantly increased plasma LH. LH-RH at 20-μg dose failed to increase plasma LH on the third day of food withdrawal, but increased it to some extent on the seventh day. The hen recovered initial sensitivity to LH-RH in increasing plasma LH after 7 days of refeeding. These results indicate that starvation caused follicular atresia in the hen probably due to the decrease of gonadotrophin secretion from the pituitary, and that starvation may reduce pituitary gonadotrope sensitivity to LH-RH in the hen at least in the early stage of fasting.     

Developmental changes in the positive feedback effect of estrogen on luteinizing hormone release in ovariectomized female rhesus monkeys

To examine developmental changes in the LH response to estrogen, eight neonatally ovariectomized monkeys received repeated injections (sc) of 50 micrograms/kg estradiol benzoate (EB) at approximately 4-month intervals starting at age 8-12 months and ending at 49-52 months. Serum samples were obtained 24 before and 0, 6, 12, 24, 36, 48, 60, 72, 94, 108, and 120 h after each EB injection. Serum LH and estradiol levels were measured by RIA. The baseline LH level before EB injection during the prepubertal period (greater than 20 months of age) was 14.4 +/- 2.2 ng/ml, and it increased progressively to 115.3 +/- 13.5 ng/ml at 41-44 months, the age shortly before the first ovulation in our intact colony animals, then declined slightly. EB first induced a typical LH response, which consisted of a negative phase (suppression) followed by a positive phase (surge), at the average age of 29.3 +/- 1.9 months (n = 8). This is similar to the age of menarche in our colony animals. The baseline LH level before EB injection at the time of the first typical response (with negative and positive phases) was 36.7 +/- 6.7 ng/ml, a level 2.5 times higher than that of the prepubertal age. The magnitude of LH suppression by EB was significantly correlated with the baseline level of LH; the higher the baseline LH before EB injection, the greater the degree of LH decrease (r = 0.968; P less than 0.001). Similarly, the amplitude of the LH peak from the trough of the negative phase was significantly correlated with the baseline LH; the higher the LH level before EB injection, the higher the LH increase (r = 0.863; P less than 0.001). The latency to the LH peak was shortest when baseline LH was highest; the peak latency (34.4 +/- 1.6 h) of the LH surge at 41-44 months of age was significantly shorter than the latency (46.5 +/- 2.7 h) of the first LH response occurring at 29.3 +/- 1.9 months of age (P less than 0.001). Finally, the pattern of circulating levels of estradiol after EB injection did not differ across the developmental stages examined. These results are interpreted to mean that an increase in LH release, presumably LHRH release, starts at the onset of puberty and continues until the age of first ovulation, and that the levels of LHRH release during the pubertal period may determine the effectiveness of estrogen on the LH surge.(ABSTRACT TRUNCATED AT 400 WORDS)     

In vivo LH-RH output of ovariectomized rats following estrogen treatment

In the present experiment we examined the effect of estrogen upon in vivo luteinizing hormone releasing hormone (LH-RH) release from the medial basal hypothalamus of freely moving ovariectomized female rats during the period of the LH surge. Ovariectomized females (10-20 day) received two subcutaneous injections of oil or estradiol benzoate (EB; 25 micrograms/rat) at 48 and 24 h prior to push-pull perfusion and were perfused between 10:00 and 18:00 h on the 3rd, day. Unexpectedly, the activity of the LH-RH pulse generator in ovariectomized rats was characterized by a very low mean LH-RH output. Administration of EB increased the activity of the LH-RH pulse generator as indicated by a significant increase in overall mean LH-RH release of EB- versus oil-treated females. In addition, EB-treated females demonstrated a diurnal variation in LH-RH release with marginal, but significant, increases in LH-RH release during the period of the LH surge (14:10-18:00 h) as compared with levels obtained between 10:00 and 14:00 h. This increase in the overall mean release during the period between 14:10 and 18:00 h appears to be attributable to an EB-induced increase in the frequency of LH-RH release, since the amplitude did not change between the two 4-hour sampling periods.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of estradiol and tamoxifen on GnRH self-priming in perifused rat adenohypophysial cells

Columns of enzymatically dispersed rat pituitary cells have been used to examine the effects of estradiol and the estrogen receptor antagonist, tamoxifen, on the responsiveness of the gonadotropes to repeated challenges with GnRH. Synthetic GnRH stimulated readily the release of LH from the pituitary cells. When challenged repeatedly with the same submaximal dose (8 nmol/l) of the releasing hormone, cells from animals in diestrus 2 or pro-estrus, like those from juvenile rats, showed a marked progressive increase in their sensitivity, but those from rats in estrus or diestrus 1 did not. Adenohypophysial cells removed from adult rats ovariectomized 14 days previously also failed to exhibit sensitization even when estradiol-17 beta (1 and 10 nmol/l) was included in the perifusion fluid. However, those from ovariectomized rats treated with appropriate doses of the steroid (1.5 microgram/day sc for 14 days or 5 micrograms sc 22 h before decapitation) did. The capacity of pituitary cells from intact rats to exhibit priming on the anticipated day of pro-estrus was unaffected by the administration of the estrogen receptor antagonist, tamoxifen (700 micrograms/100 g sc on the morning of diestrus 1 and 24 h later). Similarly, addition of tamoxifen (1 mumol/l) to the incubation medium failed to influence the capacity of pituitary cells from juvenile rats to exhibit priming in vitro. The results support the concept that the ability of GnRH to 'prime' the gonadotropes is dependent on estradiol and indicate that the exposure of the hypothalamo-pituitary complex to critical levels of estradiol prior to autopsy is necessary for the gonadotropes to retain their capacity to exhibit priming in vitro.     

Effects of p-nonylphenol and 4-tert-octylphenol on the anterior pituitary functions in adult ovariectomized rats

p-Nonylphenol (NP) and 4-tert-octylphenol (OP) are known to mimic the action of estrogens as endocrine disruptors. However, their acute effects on the pituitary and the hypothalamus functions in vivo have been uncertain. We therefore determined their effects on the anterior pituitary, in particular, gonadotropin secretion. Two weeks after ovariectomy, the rats were given a subcutaneous injection of 10 mg NP, 10 mg OP, 10 mg bisphenol A, 1 microg 17beta-estradiol, or sesame oil alone as control. Twenty-four hours after the treatment, the expression of progesterone receptor mRNA in the anterior pituitary and the level of luteinizing hormone (LH), follicle-stimulating hormone, and prolactin were determined. The expression of progesterone receptor mRNA in the anterior pituitary was significantly increased by either NP, OP, bisphenol A, or estradiol, but bisphenol A was less effective. The level of LH was significantly decreased by either NP or OP, but not by bisphenol A and estradiol. Only estradiol significantly increased the level of prolactin. The level of follicle-stimulating hormone was unchanged by any of the treatments. To check the effects of NP and OP on pulsatile LH secretion, blood samplings were done at 6-min intervals for 3 h. Twenty-four hours after treatment in ovariectomized adult rats, we found that the injection of NP significantly decreased the amplitude of LH pulses and the mean LH concentrations, but not the frequency of LH pulses. The injection of OP significantly decreased the mean LH concentrations without affecting the frequency and amplitude of the LH pulses. Finally, the rats given an injection of NP or sesame oil were intravenously injected with 50 ng of gonadotropin-releasing hormone (GnRH) to check whether NP affected the LH secretory responsiveness of the anterior pituitary to GnRH. We found that the responsiveness to GnRH in NP-injected rats was significantly attenuated compared to the sesame oil-injected rats. The present study suggests that NP, even with a single injection, suppresses the pulsatile LH secretion in adult ovariectomized rats, probably by affecting the anterior pituitary level.     

Effects of active and passive immunization with LH-RH on gonadotrophin secretion and reproductive function in female rats.

In order to get information about the physiological role played by LH-RH in the regulation of tonic and phasic secretion of gonadotrophins, and about the existence of FSH-RH distinct from LH-RH, we attempted to neutralize endogenous LH-RH by passive and active immunization with LH-RH in female rats. Anti-LH-RH serum prevented pre-ovulatory gonadotrophin surges at proestrus and ovulation, and it suppressed gonadotrophin increase induced by oestradiol benzoate or progesterone in ovariectomized rats. Elevated serum gonadotrophin concentrations in long-term ovariectomized rats were lowered by anti-LH-RH serum injection. The decrease in FSH levels was less than that in LH levels. Serum FSH rose without significant changes in serum LH level for 6 h after ovariectomy on pro-oestrus or dioestrus. The post-ovariectomy rise of FSH was not suppressed by the anti-LH-RH serum which was enought to inhibit serum LH to undetectable levels. Active immunization with LH-RH resulted in decreasing LH levels but failed to alter FSH levels in both serum and pituitary. Seven out of 10 rats immunized with LH-RH became constantly di-oestrous. The weights of anterior pituitary, ovary and uterus of the LH-RH immunized rats were significantly smaller than those of BSA immunized controls. Ovaries of LH-RH immunized rats contained few fresh corpora lutea. These results indicate that LH-RH plays a significant role in the control of both plastic and tonic secretion of LH and FSH. The existence of FSH-RH distinct from LH-RH or mechanism which specially controls the basal FSH secretion is indicated.     

Estradiol potentiation of hypothalamic uptake of LH-RH from the CSF.

Ovariectomized (Ovx) rats were treated (s.c.) with estradiol benzoate (E2B) for 7 days, and then 10, 25 or 100 ng LH-RH were microinjected into the 3rd ventricle. Intraventricular LH-RH elevated plasma LH at 10 and 30 min in a dose-response manner. In experiment 2, Ovx rats were treated with E2B or oil for 7 days and then intraventricularly injected with 25 ng LH-RH. Two days later, 100 ng LH-RH were systemically administered. E2B treatment resulted in a greater release of LH in response to the intraventricular administration of LH-RH but not to the systemic injection. A direct measure of median eminence (ME) uptake of LH-RH was used in experiment 3. Ovx-E2B or oil-treated rats were decapitated 10 min after intraventricular injection of 125I-LH-RH, 125I or 3H-glycine. The ME region of the hypothalamus, cortex, anterior pituitary (AP) and plasma were solubilized and their radioactivity determined. E2B increased the radioactivity in the ME following injection of 125I-LH-RH but not in other tissues. Tissue uptake of 125I and 3H-glycine were similar in E2B or oil-treated rats. These data indicate E2B facilitates the incorporation of LH-RH into the ME from the cerebrospinal fluid (CSF).     

Effects of anterior hypothalamic deafferentation on plasma LH response to LH-RH in the baboon (author's transl)]

We reported previously that a biphasic LH release (within 30 min and 90 approximately 150 min after LH-RH injection) was observed in normal menstruating baboons (non-human primates). Plasma immunoassayable LH-RH reached a maximum within 4 min after injection and was undetectable within 60 min. Plasma estrogen and progestin were elevated within 45 min. No plasma LH release with increment of plasma estrogen and progestin was observed in saline injected baboons. This study was performed to investigate the mechanism of biphasic LH release by LH-RH injection. Synthetic LH-RH (100 microgram) was injected sc into four female retrochiasmatic deafferented baboons. Blood samples collected 30 and 2 min before injection and 5, 10, 15, 30, 45, 60, 90, 120, 150 and 180 min after injection were assayed for LH, estrogen and progestin. In four baboons plasma LH peak was observed within 30 min after injection and plasma estrogen and progestin were elevated within 45 min. However, plasma LH peak within 90 approximately 150 min was not observed. These results infer that exogenous LH-RH exerts an effect on the pituitary to release LH within 30 min after injection. Increased estrogen and progestin exert the effect on the higher brain area (extrahypothalamic area) and the anterior hypothalamus to facilitate release of endogenous LH-RH, which subsequently release LH within 90 approximately 150 min in cooperation with exogenous LH-RH which may be considered to participate LH synthesis in the pituitary. Thus, it seems likely that the extrahypothalamic area (for example; limbic system) and the anterior hypothalamus have an important role in regulating LH-RH secretion and subsequent LH release in the baboon.     

Effect of actinomycin D on the pituitary response to LH-RH.

The effect of Actinomycin D (Act D) on the release of LH and FSH induced by LH-RH was investigated in rats. Immature male rats received an iv infusion over a period of 3-4 h or a quick iv injection of synthetic LH-RH. Infusion of LH-RH significantly increased serum LH and FSH levels at 1, 2, 3 and 4 h after the initiation of infusion. Pre-treatment with 100 mug/100 g.b.w. Act D failed to affect the rise of serum LH and FSH levels 1 h after the infusion but significantly suppressed the response at 2, 3 and 4 h. The increase in serum LH and FSH levels after a quick injection of LH-RH was unaffected by pre-treatment with Act D whether the antibiotic was injected 1 or 2 h before LH-RH. The results suggest that the initial phase of the pituitary response to LH-RH does not require DNA-dependent RNA synthesis, whereas that in the later period does. RNA synthesis may be necessary only to maintain the increased secretion of both LH and FSH during a continuous stimulation with LH-RH.