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.     

Human chorionic gonadotropin treatment prevents depressed 17 alpha-hydroxylase/C17-20 lyase activities and serum testosterone concentrations in 2,3,7,8-tetrachlorodibenzo-p-dioxin-treated rats

It is known that administration of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) causes decreased serum testosterone concentrations in the rat. Previous studies in this laboratory have shown that in rats TCDD exposure results in decreased 17 alpha-hydroxylase and C17-20 lyase activities. The decreases in these activities paralleled decreases in testicular microsomal heme and cytochrome P450 contents. As reported herein, neither testicular mitochondrial cytochrome P450 content nor the activity of cholesterol side-chain cleavage was altered in rats exposed to TCDD. Since the production of testosterone in the testis is dependent on LH, it is important to determine the early effects of TCDD on serum LH concentrations in the rat. Male Sprague-Dawley rats were given a single, oral dose of TCDD (50 micrograms/kg). Serum LH concentrations were determined by RIA on Days 1, 2, 3, 5, and 7 following TCDD treatment. Rat serum LH concentrations were decreased to 60% of controls as early as Day 1 and continued to be depressed on Days 2 and 3 at 53% and 59% of control values, respectively. Rat serum LH returned to control values by Day 5 in spite of continued depression of serum testosterone concentrations. The early depression in serum LH levels caused by TCDD may be related to the subsequent androgenic deficiency in the rat. Treatment of rats with hCG was found to be able to prevent the depression of the activities of testicular microsomal 17 alpha-hydroxylase and C17-20 lyase and serum testosterone concentrations caused by TCDD. These data indicate that TCDD decreases serum testosterone by decreasing P450(17 alpha) and C17-20 but not P450sec activities and that hCG treatment prevents the TCDD-induced decrease.     

Unexpected effects of nalmefene, a new opiate antagonist, on the hypothalamic-pituitary-gonadal axis in the male rat

In order to gain additional information on the role of brain opioid peptides in the regulation of the hypothalamic-pituitary-gonadal axis, we studied the effects of nalmefene, a new opiate antagonist, on gonadotropin and testosterone secretion in male rats. The results were compared with those obtained with naloxone, a well-studied antagonist. Acute injections of either nalmefene or naloxone (2 mg/kg) produced 4-fold increases in LH and testosterone secretion. In castrated male rats treated with testosterone propionate (TP), nalmefene (10 mg/kg) reversed the androgen negative feedback on LH secretion; surprisingly, when higher doses (25 and 50 mg/kg) were injected, the compound lost its ability to antagonize the testosterone-induced inhibition of LH levels. In contrast, naloxone was able to increase LH levels in TP-treated castrated rats even at the highest dose tested (50 mg/kg). Chronic administration of these antagonists resulted in suppression of the acute release of LH and T secretion in nalmefene-treated but not in naloxone-injected animals. These data are consistent with previous observations suggesting that opioid peptides a) exert a tonic inhibitory effect on LH and testosterone production and b) participate in the negative androgen-induced feedback control of LH secretion. Our results also show that the antagonistic action of nalmefene, but not naloxone, is reversed when higher doses are used or following chronic administration.     

Pituitary desensitization and the regulation of pituitary gonadotropin-releasing hormone (GnRH) receptors following chronic administration of a superactive GnRH analog and testosterone

We recently demonstrated that chronic daily administration of a superactive GnRH analog to intact rats resulted in an initial stimulation of serum LH levels with a subsequent return of LH levels to baseline at a time when testosterone levels were marked decreased. These data demonstrated pituatary desensitization following chronic GnRH analog treatment. Administration of GnRH analog with a dose of testosterone which did not markedly lower serum LH levels when administered alone prevented the stimulation of LH secretion by analog. The present studies were undertaken to determine the effects of GnRH analog and testosterone administration on the regulation of pituitary GnRH receptors. Pituitary GnRH receptor binding was increased by analog treatment alone at 20 days and returned to control levels at 40 and 60 days of treatment in parallel to the observed changes in serum LH, demonstrating that one mechanism by which chronic GnRH analog treatment leads to pituitary desensitization is down-regulation of pituitary GnRH receptors. Testosterone administration alone decreased pituitary GnRH receptor binding. Combined GnRH analog and testosterone administration prevented the increase in pituitary GnRH receptors observed with analog administration alone. These studies demonstrate that changes in pituitary GnRH receptor binding correlate with changes in serum LH and that the stimulatory effects of analog administration on LH are sensitive to inhibition by small doses of testosterone.     

Influence of a prolonged tamoxifen administration on steroidogenesis by incubated rat testes

Tamoxifen was administered i.m. for 9 days to adult male rats in a daily dose of 100 micrograms or 1 mg. The treatment resulted in a significant reduction of the plasma levels of testosterone and LH, without modification of the plasma levels of FSH and of the testes weight. Upon incubation, the testes from the tamoxifen-treated rats produced less testosterone and 7 alpha-hydroxytestosterone, but metabolized [4-14C]testosterone in the same way as the control animals. Small doses of hCG (0.5 i.u. for 9 days) were unable to modify the tamoxifen effect on the testicular function, while tamoxifen significantly inhibited the increase of the plasma levels of testosterone induced by the administration of moderate doses of hCG (1.5 i.u. or 2.5 i.u. for 9 days) to hypophysectomized rats. Tamoxifen treatment, however, did not modify significantly the reactivity of the testes towards high doses of hCG (10 i.u.), administered either 2 h before sacrifice or for 9 days. It is concluded that a prolonged administration of tamoxifen in the rat has, besides an indirect effect resulting from a decrease of the LH levels, a direct inhibitory influence on the testicular testosterone formation, which can be reversed by high doses of hCG.     

Effects of age and luteinizing hormone antiserum on human chorionic gonadotropin-stimulated testosterone secretion in young male rats

The steroidogenic capacity of young male rats of different ages was studied. Two days prior to sacrifice at 5, 10, 15, 20, 25 and 30 days of age, the rats in treatment groups were given intramuscularly either human chorionic gonadotropin (HCG) at 20 I.U. twice daily/rat or luteinizing hormone (LH) antiserum (AS) at 0.25 ml twice daily/rat. Either saline or normal sheep serum (NSS) was given to control rats. The serum and testicular testosterone concentrations in the control rats averaged 0.85 +/- 0.03 ng/ml and 1.35 +/- 0.06 ng/mg testicular protein, respectively. At day-15 the serum and testicular testosterone concentrations in the HCG-treated rats had significantly increased to 9.30 +/- 0.85 ng/ml and 11.92 ng/mg of testicular protein, respectively. At the same age, the HCG-induced higher levels of serum and testicular testosterone concentrations were significantly reduced to 2.80 +/- 0.70 ng/ml and 6.02 +/- 1.00 ng/mg protein by concomitant administration of LH/AS and HCG. Our results suggest that the testosterone production in response to HCG stimulation is age-related. It was also determined that neutralization of circulating gonadotropin in LH/AS-treated rats decreased the sensitivity of Leydig cells to gonadotropin stimulation. This in vivo model should provide an excellent opportunity for the investigation of the testicular function in developing young males.     

Chronic corticosterone treatment impairs Leydig cell 11beta-hydroxysteroid dehydrogenase activity and LH-stimulated testosterone production.

The effects of excess corticosterone on luteinizing hormone (LH)-stimulated Leydig cell testosterone production and activity of 11beta-HSD was studied. Adult male rats (200-250 g body weight) were treated with corticosterone-21-acetate (2 mg/100 g body weight, i.m., twice daily) for 15 days. Another set of rats was treated with corticosterone (dose as above) plus LH (ovine LH 100 microg/kg body weight, s.c., daily) for 15 days. Corticosterone administration significantly increased serum and testicular interstitial fluid (TIF) corticosterone but decreased testosterone levels. Administration of LH with corticosterone partially prevented the decrease in serum and TIF testosterone. The oxidative activity of 11beta-hydroxysteroid dehydrogenase (11beta-HSD) was significantly decreased in Leydig cells of rats treated with corticosterone alone and in combination with LH. The direct effect of corticosterone on Leydig cell steroidogenic potency was also studied in vitro. Addition of corticosterone to Leydig cell culture showed a dose dependent effect on LH-stimulated testosterone production. Corticosterone at 50 and 100 ng/ml did not alter LH-stimulated testosterone production, but at high doses (200-400 ng/ml), decreased basal and LH-stimulated testosterone production. Basal and LH-stimulated cAMP production was not altered by corticosterone in vitro. It is concluded from the present study that elevated levels of corticosterone decreased the oxidative activity of 11beta-HSD and thus resulting in impaired Leydig cell steroidogenesis and the inhibitory effects of corticosterone on testosterone production appear to be mediated through inhibition of LH signal transduction at post-cAMP level.     

Effect of neonatal testosterone and oestradiol treatment on the development of the hypothalamo-hypophysial axis in the female rat.

The hypothalamic LH-RH content and the concentrations of pituitary and plasma LH were measured at various ages in female rats treated daily with 10 micrograms testosterone propionate or 10 micrograms oestradiol-17beta from birth to Day 15. Persistent vaginal oestrus was induced in all the treated rats. Both hormones significantly reduced the hypothalamic LH-RH content and pituitary and plasma LH concentrations. Hypothalamic LH-RH increased after cessation of treatment but pituitary LH did not return to normal levels. Plasma LH levels were significantly lower than those in control rats. It is concluded that testosterone propionate and oestradiol-17beta (1) have a direct negative feed-back influence on the hypothalamus in the neonatal female rat; (2) alter the normal pattern of plasma and pituitary LH in developing female rats; (3) prevent the cyclic secretion of plasma LH after maturity; and (4) probably cause a chronic impairment in the release of LH-RH.     

Inhibition of estrogen biosynthesis and its consequences on gonadotrophin secretion in the male

Of the gonadal steroids in the male, testosterone is the most important regulator of gonadotrophin secretion. However, whether testosterone affects gonadotrophin secretion directly or whether it must first be aromatized to estrogens is controversial. We have reported extensively on the endocrine and anti-tumor effects of the non-steroidal aromatase inhibitors CGS 16949A and CGS 20267 in adult female rats. In these animals, both inhibitors potently and selectively inhibit estrogen biosynthesis. Thus these agents can be effectively used in studying estrogen-dependent processes. CGS 16949A was administered for 14 days to adult male rats, over a dose range which in females suppresses estradiol and elevates LH. In male rats a suppression of estradiol was seen, however, there was no significant effect on either serum LH or on the weights of androgen-dependent organs. CGS 16949A, when administered to healthy men at a dose of 1 mg b.i.d. for 10 days, causes a significant fall in plasma estradiol and significant elevations of plasma FSH and testosterone. Dose-dependent suppression of serum estradiol and an increase in serum testosterone and LH are seen after administration of single oral doses of CGS 20267. These results indicate that in the male rat, inhibition of aromatization of testosterone to estrogens does not influence gonadotrophin secretion whereas in men the negative feedback exerted by testosterone on gonadotrophin secretion is dependent on the aromatization of testosterone to estrogens.     

The effects of ethylene dimethane sulphonate (EDS) on rat Leydig cells: evidence to support a connective tissue origin of Leydig cells

Ethylene dimethane sulphonate (DS) administered to adult male rats in a single dose of 75 mg/kg body weight results in a rapid destruction of Leydig cells which, in turn, is associated with a marked decline in levels of serum testosterone. For 24-72 h after treatment with EDS (post-EDS) the Leydig cells undergo degenerative changes consisting of chromatin condensation and cytoplasmic vacuolation, and testicular macrophages progressively remove Leydig cells from the intertubular tissue by phagocytosis. This results in the total absence of Leydig cells on Days 7-14 and the absence of any detectable specific 125I-hCG binding to testis homogenates. Associated with the low levels of serum testosterone, levels of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in serum rise, LH to levels found in castrate rats. Morphometric and 125I-hCG binding studies indicate that a new generation of Leydig cells develop from Day 21 and reach control levels by Day 49. Morphologic observations suggest that the Leydig cells arise by differentiation from a pool of connective tissue cells that includes fibroblasts, lymphatic endothelial cells and pericytes. The new Leydig cells, which appear around Day 21 post-EDS, have the features of fetal Leydig cells. The latter appear to transform into Leydig cells typical of normal adult rats between 35-49 days post-EDS. The differentiation of new Leydig cells is associated with a reestablishment of normal levels of testosterone 21 days post-EDS. Serum LH and FSH return to normal at 28 days and 49 days respectively.     

Differential response of testis and serum gonadotropins to testosterone in rats treated with a gonadotropin releasing hormone antagonist or estradiol 17 beta.

Adult rats treated with a GnRH antagonist (Ac D2Nal1, D4Cl Phe2, DTrp3, DArg6, DAla10 GnRH; code: 103-289-10, National Institutes of Health, USA) for 5 weeks (250 micrograms/kg b.w) showed multiple degrees of impairment and atrophy of the genital organs concomitant with decreased serum levels of testosterone, LH and FSH. Inhibition of spermatogenesis was characterized by germ cell degeneration and overall decline in different cell numbers and in particular, spermatids of any kind were completely absent. Testosterone supplementation (60 micrograms/rat/day, sc) to GnRH antagonist treated rats, for the same period, significantly elevated the weights of the sex organs, and the serum levels of hormones. Spermatogenesis was improved both qualitatively and quantitatively; albeit failed to be restored back to control levels. Treatment with estradiol 17 beta (1 microgram/rat/day) for 5 weeks had insignificant effect on spermatogenesis but the weights of the genital organs (seminal vesicles by 19% and ventral prostate by 40%) and the levels of serum hormones (LH by 24%, FSH 22% and testosterone by 25%) were otherwise reduced. Administration of testosterone either alone or in combination with estradiol 17 beta had only a marginal effect on spermatogenesis or on other reproductive parameters. The results indicate a positive shift in the response of the testis and serum levels of gonadotropins to testosterone supplementation in rats treated with either GnRH antagonist or estradiol 17 beta.     

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.     

Bisphenol A inhibits testicular functions and increases luteinizing hormone secretion in adult male rats

Effects of a xenobiotic estrogen, bisphenol A (BPA), on reproductive functions were investigated using adult male rats. BPA was dissolved into sesame oil and injected s.c. every day (1 mg/rat) for 14 days. Animals were killed by decapitation after the final administration of BPA, and the trunk blood, pituitary, and testes were collected. Plasma concentrations of prolactin were dramatically increased and pituitary contents of prolactin were slightly increased in the BPA group compared to the control group. Plasma concentrations of testosterone were decreased and plasma concentrations of LH were increased in BPA-treated rats compared to control rats. Testicular contents of inhibin were decreased in BPA-treated rats compared to control rats, although plasma concentrations of inhibin were not changed after administration of BPA. The testicular response to hCG for progesterone and testosterone release was decreased in BPA-treated rats. Administration of BPA did not change the pituitary response to luteinizing hormone-releasing hormone (LH-RH) in castrated male rats treated with testosterone. Male sexual behavior also was not changed as a result of BPA treatment. These results suggest that BPA directly inhibits testicular functions and the increased level of plasma LH is probably due to a reduction in the negative feedback regulation by testosterone. The testis is probably a more sensitive site for BPA action than the hypothalamus-pituitary axis.     

Effects of cocaine hydrochloride on the male reproductive system

The reproductive system effects of cocaine were studied in male rats. The analysis included measurements of circulating levels of luteinizing hormone (LH) and testosterone (T) by radioimmunoassay (RIA). The weights of the testes and sex accessory organs were also assessed and compared with control animals. Dosage level, duration of treatment, and interval between injection and sacrifice were the parameters examined. Following a single intraperitoneal (IP) injection, LH levels decreased over a 3-hour period. At a high dosage (40 mg/kg), cocaine caused a significant elevation in serum T followed by a significant depression of T for at least 2 hours. When administered chronically for 15 days, the low dose group (10 mg/kg) did not vary significantly from the vehicle controls. However, the high dose group had lower LH and T levels, as well as correspondingly lighter weight seminal vesicles and epididymis. No changes were noted in the weights of the ventral prostate or testes. This research suggests that cocaine acts primarily at the hypothalamic-hypophyseal axis with a possible secondary action at the gonadal level.     

Prostacyclin (PGI2) effects on anterior pituitary hormones in the rat in vivo.

Intravenous injection of 600 microgram PGE2 or PGI2 significantly increased serum LH and prolactin levels in estradiol treated ovariectomized rats. There was no effect on serum FSH concentration. PGE2 and PGI2 stimulated LH release in a non-dose dependent manner, while prolactin levels were positively correlated with the dose administered following PGI2 treatment. 6-keto-PGF1 alpha at a comparable dose had no effect on pituitary hormone levels. Subcutaneous administration of 1 mg/kg or 60 mg/kg PGI2 for seven days significantly depressed serum LH level both in male and female rats. These doses had no effect on serum FSH or prolactin levels.     

Inhibition of pituitary-gonadal function in male rats by a potent GnRH antagonist

The inhibitory effects of the potent GnRH antagonist, [Ac-D-pCl-Phe1,2,D-Trp3,D-Arg6,DAla10]GnRH (GnRHant) upon pituitary-gonadal function were investigated in normal and castrated male rats. The antagonist was given a single subcutaneous (s.c.) injections of 1-500 micrograms to 40-60 day old rats which were killed from 1 to 7 days later for assay of pituitary GnRH receptors, gonadal receptors for LH, FSH, and PRL, and plasma gonadotropins, PRL, and testosterone (T). In intact rats treated with low doses of the antagonist (1, 5 or 10 micrograms), available pituitary GnRH receptors were reduced to 40, 30 and 15% of the control values, respectively, with no change in serum gonadotropin, PRL, and T levels. Higher antagonist doses (50, 100 or 500 micrograms) caused more marked decreases in free GnRH receptors, to 8, 4 and 1% of the control values, which were accompanied by dose-related reductions in serum LH and T concentrations. After the highest dose of GnRHant (500 micrograms), serum LH and T levels were completely suppressed at 24 h, and serum levels of the GnRH antagonist were detectable for up to 3 days by radioimmunoassay. The 500 micrograms dose of GnRHant also reduced testicular LH and PRL receptors by 30 and 50% respectively, at 24 h; by 72 h, PRL receptors and LH receptors were still slightly below control values. In castrate rats, treatment with GnRHant reduced pituitary GnRH receptors by 90% and suppressed serum LH and FSH to hypophysectomized levels. Such responses in castrate animals were observed following injection of relatively low doses of GnRHant (100 micrograms), after which the antagonist was detectable in serum for up to 24 h. These data suggest that extensive or complete occupancy of the pituitary receptor population by a GnRH antagonist is necessary to reduce plasma gonadotropin and testosterone levels in intact rats. In castrate animals, partial occupancy of the available GnRH receptor sites appears to be sufficient to inhibit the elevated rate of gonadotropin secretion.     

Effect of bromocriptine on the hypothalamo-pituitary function in adult male rats.

Daily oral administration of bromocriptine (50 μg/kg) to adult male rats, suppressed serum prolactin levels. The pituitary prolactin levels remained unaltered. Serum FSH levels as well as pituitary FSH levels showed no significant change as compared to the controls. Serum LH levels were significantly decreased in spite of the high pituitary LH levels, in bromocriptine treated rats. In the drug treated rats, $\text{in vitro}$ sensitivity of the pituitary to the exogenous LH-RH was not altered; whereas hypothalamic LH-RH content was considerably lowered. These observations suggest the possible effect of bromocriptine on the synthesis of LH-RH in the hypothalamus which leads to the accumulation of LH in the pituitary and decline of serum LH.     

Reduced ability of naloxone to stimulate LH and testosterone release in aging male rats; possible relation to increase in hypothalamic met5-enkephalin

Blood luteinizing hormone (LH) and testosterone levels are lower in old than in young male rats. The specific opiate antagonist, naloxone, previously shown to increase serum LH in mature male rats, exhibited relatively little ability to raise serum LH and testosterone levels in old (18–20 mo) as compared to young (4–5 mo) male rats. The brain opiate, met⁵-enkephalin, which depresses LH, was found to be significantly higher in the hypothalamus of old than of young male rats. These observations suggest that hypothalamic opiates may be partially responsible for the lower serum LH and testosterone levels in old male rats, and for reduced release of these hormones in response to naloxone administration.     

Ethanol inhibits the naloxone-induced release of luteinizing hormone-releasing hormone from the hypothalamus of the male rat

It has been inferred that ethanol suppresses the secretion of luteinizing hormone (LH) in the male by depressing the release of LH-releasing hormone (LH-RH) from the hypothalamus. Direct support for this inference has been difficult to obtain, however, because of significant technical difficulties in measuring LH-RH release under in vivo conditions. To circumvent these problems, we made use of the opiate antagonist naloxone, as a neuroendocrine probe, to elicit the release of LH-RH under in vivo conditions. We found that ethanol was a potent suppressor of the increase in serum LH levels evoked by naloxone at extremely low blood ethanol concentrations ( less than 60 mg/dl). Furthermore, we observed that the antagonism between ethanol and naloxone appeared to be competitive in nature since a fixed dose of ethanol (1 g/kg, blood ethanol concentration 60 mg/dl) shifted the naloxone dose-response curve significantly to the right and high doses of the antagonist overcame ethanol's effects. Finally, we found that the interaction between ethanol and naloxone took place at the level of the hypothalamus. Our results, therefore, seem to provide the first in vivo evidence supporting the widely-held hypothesis that ethanol reduces serum LH levels by depressing the hypothalamically-medicated release of LH-RH. The mechanisms underlying ethanol's depression of naloxone-induced increases in the release of LH-RH are not fully understood at this time, but one prominent possibility is that ethanol enhances the synthesis or release of endogenous opioids which in turn override naloxone's effects.     

Physiological role of putative testicular gonadotrophin releasing hormone (GnRH)

Evidence suggests that exogenous GnRH and agonist analogues have short-term stimulatory effects on rat Leydig cell function - when administered intratesticularly. Since rat Leydig cells possess GnRH receptors and their endogenous ligand has not yet been identified the physiological importance of the observations for testis function is unknown. To address this issue we have determined the consequences of blockade of testis GnRH receptors on Leydig cell function under both normogonadotrophic and hypogonadotrophic stimulation of the testis in vivo. A GnRH antagonist (ANT) was used to achieve receptor blockade but during continuous systemic infusion ANT occupied pituitary GnRH receptors and markedly reduced serum LH, FSH, testosterone, and intratesticular testosterone in adult and 30 d old immature male rats. These results were similar to those obtained by administration of a GnRH antiserum which did not bind to testis GnRH receptors. Thus, blockade of testis GnRH receptors during hypogonadotrophism did not produce additional inhibition of steroidogenesis by Leydig cells. However, direct continuous infusion of ANT into one testis produced greater than 90% occupancy of GnRH receptors while reducing GnRH receptors by only 50% in the contralateral testis. Unilateral intratesticular infusion did not reduce serum LH, FSH, Prolactin or testosterone levels despite 75% occupancy of pituitary GnRH receptors. Thus, both ANT infused and saline infused testes were exposed to the same gonadotrophic stimulants but in the former GnRH-R were essentially non-existent. Compared to the control testis, the ANT infused testis showed a 20-30% reduction in LH, FSH, lactogen receptors and 30-40% fall in testosterone content. Identical results were obtained in adult and 30 d-old male rats.(ABSTRACT TRUNCATED AT 250 WORDS)