Hormonal regulation of activities of 4-ene-5 beta- and 5 alpha-reductases and 17 beta-hydroxy-dehydrogenase in immature golden hamster testis

We have reported [1-3] in immature golden hamster testis that 5 beta-reductase is localized in the tubular nongerm cells, while 5 alpha-reductase is present in the interstitial tissue and that the 17 beta-hydroxy-dehydrogenase activity is found predominantly in the tubular nongerm cells. Hormonal regulation of these enzyme activities was examined in the present study. Male golden hamsters were hypophysectomized on day 22 after birth. The hypophysectomized hamsters in groups of 3-8 were injected daily with 10 micrograms NIH-LH-S19, 50 micrograms NIAMD-Rat-FSH-B-1, 8 or 16 micrograms NIAMD-oFSH-13, 8 micrograms NIAMD-oFSH-13 plus 5 or 10 micrograms NIH-LH-S19, 1 mg testosterone propionate or saline for 5 days starting from day 23. Testicular homogenates of the treated hamsters and intact hamsters on day 28 were incubated with [14C]4-androstene-3,17-dione and NADPH, and enzyme activity (nmol/testes/h) was estimated. The activities of 5 beta- and 5 alpha-reductases and 17 beta-hydroxy-dehydrogenase decreased significantly 6 days after hypophysectomy. In the hypophysectomized hamster testis, a distinct response to FSH but not to LH in the activities of 5 beta-reductase and 17 beta-hydroxy-dehydrogenase was found. The injection of LH in addition to FSH showed no significant additive effects on these enzyme activities. The 5 alpha-reductase activity was stimulated significantly by LH plus FSH but not by LH alone, FSH alone or androgen. These results show that 5 beta-reduction of 4-ene-3-ketosteroids takes place in the Sertoli cells under the influence of FSH while 5 alpha-reduction occurs in the interstitial cells under the influence of LH and FSH in immature hamster testis.     

Intratubular localization of 4-ene-5 beta-reductase and 17 beta-hydroxy-dehydrogenase in immature golden hamster testis

We have reported [1,2] in immature golden hamster testis that 5 beta-reductase is localized in the seminiferous tubules, while 5a-reductase is present in the interstitial tissue and that the 17 beta-ol-dehydrogenase activity is found predominantly in the seminiferous tubules. In the present study, we show the intratubular localization of these enzymes. The left testis of golden hamster was irradiated with 2000R or 8000R of X-rays at 22 days of age. The hamsters were killed at 28 days of age. Homogenates of the left irradiated and right intact testes were incubated with [14C]-4-androstone-3,17-dione and NADPH, and enzyme activity was estimated. Both testes were also examined histologically. The X-irradiation of the testis resulted in an almost complete disappearance of germ cells with a significant decrease in testis weight, but the interstitial tissue and tubular nongerm cells including Sertoli cells remained almost unchanged. However, the activities of 5 beta-reductase and 17 beta-ol-dehydrogenase expressed as nmol formed/testis/h did not decrease at all. These results show that 5 beta-reductase is localized in the tubular nongerm cells including the Sertoli cells and 17 beta-ol-dehydrogenase is present in the tubular nongerm cells and interstitial tissue in immature golden hamster testis.     

Effects of hyperprolactinemia on FSH- or LH-induced activities of 17 beta-oxidoreductase, 5 alpha-reductase, and 17-hydroxylase in hypophysectomized rat testes

Two pituitaries from 7-week-old female rats (Sprague-Dawley strain) were grafted under the capsule of the left kidney of a 49-day old male rat. The pituitary grafted and sham-operated rats were hypophysectomized at 56 days of age. The hypophysectomized rats were given daily injections of NIAMDD-oFSH-13 (20 micrograms/0.5 ml saline), NIAMDD-oLH-23 (9 micrograms/0.5 ml saline) or saline for 4 days starting from day 58. The treated rats and normal male rats were killed at 61 days of age. Testicular homogenates were incubated with [14C]4-androstene-3, 17-dione or [3H] progesterone, and enzyme activities per testes were estimated. Hypophysectomy caused significant decreases in activities of testicular 17 beta-oxidoreductase and 17-hydroxylase. The decreased activity of 17 beta-oxidoreductase was significantly stimulated by FSH or LH treatment, whereas the decreased 17-hydroxylase activity was stimulated only by LH treatment. Although pituitary grafts alone showed little or no effect on these enzyme activities in the hypophysectomized rats, the grafts significantly inhibited FSH-stimulated 17 beta-oxidoreductase activity and the LH-stimulated 17 beta-oxidoreductase and 17-hydroxylase activities but enhanced LH-induced 5 alpha-reductase activity. The present results confirm previous findings that an excess of prolactin directly inhibits LH-stimulated 17-hydroxylase activity but enhances LH-induced 5 alpha-reductase activity in the rat testis. The present results also demonstrate that the same grafts directly inhibit FSH-stimulated 17 beta-oxidoreductase activity but have no effect on FSH-induced 5 alpha-reductase activity.     

Effects of chronic administration of LH releasing hormone on age-dependent activities of testicular 4-ene-5 alpha-reductase and 17 beta-ol-dehydrogenase in mice

Male (WB X C57BL/6)F1 hybrid mice of 16, 26 and 66 days of age, 4 in each group, were injected daily with 0.2 micrograms/10 g body weight of LH releasing hormone (LHRH) or saline for 14 days. Testicular homogenates were incubated with [14C]4-androstene-3,17-dione and enzyme activities were examined. In mice treated with saline, testicular 17 beta-ol-dehydrogenase activity increased with age but 4-ene-5 alpha-reductase (5 alpha-reductase) activity decreased with age. LHRH treatment for 14 days starting from day 26 resulted in a delay in sexual maturation, as evidence by significant decreases (P less than 0.05) in seminal vesicle weight and testicular 17 beta-ol-dehydrogenase activity and by a significant increase (P less than 0.05) in 5 alpha-reductase activity. However, LHRH treatment starting from day 66 had no significant effect on these testicular enzyme activities.     

Hyperprolactinemia enhances LH-stimulated 4-ene-5 alpha-reductase activity but inhibits LH-induced 17-hydroxylase activity in testes of hypophysectomized immature rats

Two pituitaries from 7-week old female rats (Sprague-Dawley strain) were grafted under the capsule of the left kidney of 21-day old male rat. The pituitary grafted and sham-operated rats were hypophysectomized at 27 days of age. The hypophysectomized rats, in groups of 4, were given daily injections of 9 micrograms NIAMDD-oLH-23 (minimum effective dose) or saline for 3 days starting from day 29. Testicular homogenates were incubated with [3H]progesterone or [14C]4-androstene-3,17-dione, and enzyme activities per testes were estimated. Testicular HCG-binding sites were also measured. Hypophysectomy caused significant decreases in activities of testicular 5 alpha-reductase, 17-hydroxylase, and 17 beta-hydroxysteroid oxidoreductase. These decreased enzyme activities were significantly stimulated by LH treatment. Although pituitary grafts alone showed no effects on these enzyme activities in the testes of the hypophysectomized rats, the grafts significantly enhanced LH-stimulated 5 alpha-reductase activities but inhibited LH-stimulated 17-hydroxylase activity. Testicular LH/HCG receptors were significantly increased by the grafts, especially in the presence of LH, without affecting affinity for HCG. The present results demonstrate for the first time that hyperprolactinemia directly stimulates LH-stimulated 5 alpha-reductase activity in rat testes. The results also show that the same grafts directly inhibit LH-stimulated 17-hydroxylase activity, probably via postreceptor mechanisms.     

Differential effects of follicle-stimulating hormone and luteinizing hormone on Leydig cell function and restoration of spermatogenesis in hypophysectomized and photoinhibited Djungarian hamsters (Phodopus sungorus)

Effects of pure human follicle-stimulating hormone (hFSH) and ovine luteinizing hormone (oLH) on testicular function were investigated in long-term hypophysectomized or photoinhibited Djungarian hamsters. hFSH (5 IU) or oLH (5 micrograms) or a combination of FSH and LH (5 IU and 5 micrograms, respectively) were injected s.c. twice daily for 7 days to hypophysectomized and photoinhibited hamsters. Other photoinhibited hamsters were treated for 14 and 21 days with FSH and LH (3 IU and 3 micrograms, respectively) in a similar way. LH alone had little, if any, effect on testicular weights; FSH, when injected alone or in combination with LH (FSH/LH), caused a significant increase in testes weights at each time point. On the other hand, LH or FSH/LH, but not FSH alone, caused a significant increase in the accessory organ weights. FSH had no effect on intratesticular testosterone (T) or on 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) activity but enhanced the in vitro response of interstitial cells to hCG. LH and FSH/LH had pronounced effects on intratesticular T, 3 beta-HSD activity, and in vitro response of interstitial cells to human chorionic gonadotropin. Treatment with FSH or FSH/LH caused regrowth of the testis and restoration of tubular lumen and tubular diameter and restored complete spermatogenesis. However, LH had little effect on spermatogenesis in spite of increased intratesticular and peripheral T levels. These results indicate that although LH can cause a full redifferentiation of Leydig cells in photoinhibited hamsters, it has only minor effects on tubular function. On the other hand, FSH alone induces full restoration of tubular function in these animals and has no direct effect on Leydig cell steroidogenesis, but may enhance the Leydig cell responsiveness to LH.     

Cell-type-specific localization of transforming growth factor-beta 2 and transforming growth factor-beta 1 in the hamster ovary: differential regulation by follicle-stimulating hormone and luteinizing hormone.

Cell-type-specific localization and gonadotropin regulation of transforming growth factor-beta 1 (TGF-beta 1) and transforming growth factor-beta 2 (TGF-beta 2) in the hamster ovary were evaluated immunohistochemically under three conditions: (1) during the estrous cycle (Day 1 = estrus; Day 4 = proestrus); (2) after the blockade of periovulatory gonadotropin surges by phenobarbital, and (3) after FSH and/or LH treatment of long-term hypophysectomized hamsters. Ovarian TGF-beta 1 activity was primarily localized in theca and interstitial cells. The activity increased moderately but significantly after the preovulatory LH surge and reached a peak at 0900 h, Day 2 h; oocytes showed considerable activity. TGF-beta 1 immunoreactivity subsequently fell to low levels in theca-interstitial cells through 0900 h, Day 4. Significant TGF-beta 2 immunoreactivity appeared after the surge, mainly in the granulosa cells of both preantral and antral follicles; a few interstitial cells surrounding preantral follicles showed discrete staining. TGF-beta 2 immunoreactivity in granulosa cells and in interstitial cells next to preantral follicles reached a peak at 0900 h, Day 1, and persisted up to 0900 h, Day 2; oocytes showed no staining. Phenobarbital treatment blocked the appearance of TGF-beta 1 and TGF-beta 2 immunoreactivities at 1600 h, Day 4; however, a rebound in immunoreactivities was observed with the onset of the surge after a 1-day delay. Replacement of LH to long-term hypophysectomized hamsters resulted in a marked increase in TGF-beta 1 immunoreactivity in the interstitial cells, but FSH, although it induced follicular development, did not influence ovarian TGF-beta 1 activity. Treatment with FSH, however, induced a massive increase in TGF-beta 2 immunoreactivity in the granulosa cells of newly developed antral and preantral follicles but not in the interstitial cells; LH, on the other hand, had no significant effect on TGF-beta 2 activity. Treatment with FSH and LH combined resulted in a dramatic increase in TGF-beta 2 immunoreactivity in granulosa and interstitial cells and in TGF-beta 1 in theca and interstitial cells comparable to their peak activity in intact animals. Western analyses substantiated the presence of TGF-beta 1 and TGF-beta 2 in the hamster ovary and the specificity of immunolocalization. These studies, therefore, provide critical evidence that TGF-beta 1 and TGF-beta 2 in the hamster ovary are expressed in specific cell types and that their expression is differentially regulated by LH and FSH, respectively.     

Effects of hypothalamic neurohormones on prolactin release from pituitary allografts in the hamster

Recent reports indicate that luteinizing hormone-releasing hormone (LHRH) releases prolactin (PRL) under some circumstances. We examined the chronic effects of LHRH, growth hormone-releasing hormone (GHRH), and corticotrophin-releasing hormone (CRH) on the release of PRL, luteinizing hormone (LH), and follicle-stimulating hormone (FSH) by pituitary allografts in hypophysectomized, orchidectomized hamsters. Entire pituitary glands removed from 7-week-old-male Golden Syrian hamsters were placed under the renal capsule of hypophysectomized, orchidectomized 12-week-old hamsters. Beginning 6 days postgrafting, hamsters were injected subcutaneously twice daily with 1 microgram LHRH, 4 micrograms GHRH, or 4 micrograms CRH in 100 microliter of vehicle for 16 days. Six hosts from each of the four groups were decapitated on Day 17, 16 hr after the last injection. Prolactin, LH, and FSH were measured in serum collected from the trunk blood. Treatment with LHRH significantly elevated serum PRL levels above those measured in the other three groups, which were all similar to one another. Serum LH levels in hosts treated with vehicle were elevated above those measured in the other three groups. Serum FSH levels in hosts treated with LHRH were greater than FSH levels in any of the other three groups. These results indicate that chronic treatment with LHRH can stimulate PRL and FSH release by ectopic pituitary cells in the hamster.     

Effects of progesterone or luteinizing hormone on folliculogenesis in intact or hypophysectomized immature hamsters

Since exogenous progesterone (P4) causes superovulation in hypophysectomized (hypoxed) cyclic hamsters treated with gonadotropins, the current study was performed to evaluate the roles of P4 and luteinizing hormone (LH) as a folliculotropic complex in the immature hamster. Intact or hypoxed immature hamsters were injected daily, beginning on Day 23, with 1 mg P4 and/or 20 micrograms LH for 4 days. Treatment with P4 alone or combined with LH in intact immature hamsters increased the number of antral follicles (6.7 and 4.3, respectively, vs. 1.5 per ovary in controls), but neither treatment maintained large follicles in hypoxed animals. In contrast, in hypoxed hamsters, the number of small preantral follicles was enhanced by P4 or LH (406 and 409, respectively, compared to 302 per ovary in untreated controls), but with no additive effects by combined treatment. The stimulatory effect of P4 in intact hamsters was unrelated to serum levels of follicle-stimulating hormone, estradiol, or LH. Moreover, in the hypoxed hamster, P4 or LH acts directly to increase the numbers of small preantral follicles with 2 to 5 layers of granulosa cells, whereas equally large doses of stilbestrol or estradiol cyclopentylpropionate are ineffective. In the hypoxed or intact hamster, the effects of P4 or LH may involve either recruitment of smaller follicles into larger stages or prevention of atresia. The present experimental design can not distinguish between these possibilities.     

Effects of FSH and LH on incorporation of [3H]thymidine into follicular DNA

To assess the roles of FSH and LH on follicular growth, after various experimental manipulations, hamster follicles were sorted into 10 stages and incubated for 4 h with [3H]thymidine. Stages 1-4 correspond to follicles with 1-4 layers of granulosa cells, respectively; Stage 5 = 5 or 6 layers of granulosa cells plus theca; Stage 6 = 7-8 layers of granulosa cells plus theca; Stage 7 = early formation of the antrum; Stages 8-10 = small, intermediate and large antral follicles, respectively. Phenobarbitone sodium injected at 13:00 h on pro-oestrus blocked the normal rise of blood FSH and LH concentrations at 15:00 h and prevented the increase of [3H]thymidine incorporation into follicles of Stages 1-9. The optimal treatment to reverse the effects of phenobarbitone was 1 microgram FSH and 2 micrograms LH injected i.p. at 13:00 h which restored DNA replication to follicles of Stages 2-10: FSH acted primarily on Stages 2-5 and LH on Stages 5-10. Injection of phenobarbitone at 13:00 h on prooestrus followed by 2.5 micrograms FSH at 22:00 h restored DNA synthesis by the next morning to follicles at Stages 1-8. In hamsters hypophysectomized at 09:00 h on the day of oestrus (Day 1), injection on Day 4 of 2.5 micrograms FSH restored DNA synthesis 6 h later to Stage 2-6 follicles. Unilateral ovariectomy on Day 3 resulted 6 h later in an acute rise in FSH and LH and change of follicles from Stage 4 to Stage 5 but, paradoxically, there was decreased synthesis of DNA in follicles of Stages 5-10.(ABSTRACT TRUNCATED AT 250 WORDS)     

Regulation of the pituitary 5 alpha-reductase activity by gonadotropin releasing hormone and testosterone in the adult male rat

Intact or castrated adult male rats were treated for nine days with GnRH (10 micrograms/day), the synthetic GnRH goserelin (100 micrograms/day) or the GnRH-antagonist Org 30276 (250 or 500 micrograms/day). In some series, 1 mg testosterone propionate was administered alone, or in combination with goserelin or Org 30276. The in vitro metabolism of [1 alpha,2 alpha-3H]testosterone by pituitary and hypothalamic homogenates was investigated in combination with the estimation of plasma concentrations of testosterone and gonadotropins. No qualitative or quantitative differences were observed in hypothalamic testosterone metabolism or in the pituitary 17 beta-hydroxysteroid dehydrogenase activity. Testosterone administration to intact male rats decreased the pituitary 5 alpha-reductase activity and LH, while administered to castrated rats, it was able to suppress totally the castration-induced increase of the 5 alpha-reductase activity and of the gonadotropin secretion. The drastic decrease of the plasma levels of testosterone, observed after a prolonged treatment with GnRH, goserelin or Org 30276 was not accompanied by an increased pituitary 5 alpha-reductase activity. Injected to castrated rats, it was observed that the castration-induced increase of the pituitary 5 alpha-reductase was further stimulated by GnRH, totally suppressed by goserelin and partially suppressed by Org 30276. Concomitant administration of goserelin or Org 30276 and testosterone propionate to castrated rats resulted in a further decrease of the pituitary 5 alpha-reductase activity, compared to the castrated, GnRH-analogue treated rats. These data indicate that the pituitary 5 alpha-reductase enzyme system is controlled by both direct steroidal and indirect GnRH-mediated mechanisms.     

Down regulation of prolactin receptors in the hamster ovary by the preovulatory gonadotropin surge

Binding of 125I-prolactin (Prl) to hamster ovarian homogenates was found to decrease markedly at the time of the preovulatory gonadotropin surge (PGS). Saturation analysis revealed that the decrease was due to a reduction in the number of available Prl receptors and not due to a change in binding affinity. Loss of Prl receptors following the PGS was not affected by treatment with ergocryptine to block the release of pituitary Prl, indicating that the reduction in the number of available Prl receptors was not due to increased occupancy by endogenous Prl. Loss of Prl receptors was prevented by treatment with phenobarbital (Phen) to block the normal luteinizing hormone (LH)/follicle-stimulating hormone (FSH) surge; whereas, an injection of 50 micrograms of LH or 50 micrograms FSH (but not 100 micrograms Prl) induced a marked decrease in Prl receptors in Phen-treated hamsters. To determine whether Prl receptor loss induced by 50 micrograms FSH might be due to LH contamination, Phen-treated hamsters were injected with minimal ovulatory doses of LH and FSH. Injection of 5 micrograms or 2.5 micrograms LH induced a loss of Prl receptors in 90% and 70% of Phen-treated hamsters, respectively. In contrast, injection of 5 micrograms or 2.5 micrograms FSH induced a loss of Prl receptors in 0% and 20% of Phen-treated hamsters, respectively. These results indicate that the PGS causes an acute heterologous down regulation of ovarian Prl receptors and suggest that this down regulation may be due principally to the action of LH.     

Estrogen regulation of progestin biosynthetic enzymes in cultured rat granulosa cells

The mechanism by which estrogens enhance gonadotropin-stimulated ovarian progestin production was investigated by studying the modulation of pregnenolone biosynthesis as well as the activities of 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) and 20 alpha-hydroxysteroid dehydrogenase (20 alpha-HSD) in cultured rat granulosa cells. Cells from immature hypophysectomized, estrogen-treated rats were cultured for 3 days with follicle-stimulating hormone (FSH) and/or estrogens. Pregnenolone production was measured in the presence of cyanoketone which inhibits 3 beta-HSD activity. Activities of 3 beta-HSD and 20 alpha-HSD were determined in cell homogenates by direct enzyme assays. Some cells were also primed with FSH to induce luteinizing hormone (LH) receptors for studies on the effects of estrogens on LH-modulated parameters. Pregnenolone production by cultured granulosa cells was stimulated by FSH, while treatment with diethylstilbestrol (DES) or estradiol further enhanced the gonadotropin action. Treatment with FSH increased 3 beta-HSD activity. Similarly, concomitant treatment with DES further enhanced 3 beta-HSD activity in a dose-dependent manner with an apparent ED50 of 10(-8) M. Also, treatment with estrogens alone increased 3 beta-HSD activity. The increases in enzyme activity induced by estrogen alone or in combination with FSH were not associated with changes in the apparent Km values. FSH also stimulated 20 alpha-HSD activity by 2-fold in these cells, while concomitant treatment with DES did not affect the FSH action. In FSH-primed cells, LH stimulated pregnenolone production while the LH action was enhanced by concomitant treatment with the estrogens. Likewise, LH stimulated the activity of 3 beta-HSD, while concomitant DES treatment further augmented LH action. LH did not stimulate 20 alpha-HSD activity when added alone or in combination with DES. Thus, the estrogen enhancement of the gonadotropin-stimulated progesterone production in cultured rat granulosa cells is associated with increases in pregnenolone biosynthesis and the activity of the 3 beta-HSD enzyme, without affecting the 20 alpha-HSD activity.     

Follicle-stimulating hormone inhibits granulosa cell 5 alpha-reductase activity. Possible role of 5 alpha-reductase as a steroidogenic pubertal switch.

In order to elucidate the role of 5 alpha-reductase in the ovarian pubertal transition from 5 alpha-reduced to non-5 alpha-reduced steroids, we examined the characteristics and regulation of granulosa cell (GC) 5 alpha-reductase activity. Maximum activity was observed at 37 degrees C and at a pH of 6.5-8.0. Synthetic 4-aza-3-oxosteroids proved to be potent inhibitors (76% inhibition at 0.1 microM) of ovarian 5 alpha-reductase activity, and 20 alpha-DHP was a better substrate than either progesterone or testosterone (4- or 7-fold higher affinity constants, respectively). The Km (20 alpha-DHP) of the enzyme was 0.50 +/- 0.03 microM and 0.75 +/- 0.20 microM in homogenates of whole ovaries and GC, respectively. 17 beta-Estradiol was a non-competitive inhibitor (KI = 6.97 microM). 5 alpha-Reductase activity was 22-fold (immature) to 68-fold (mature) higher in liver than ovary and 4-fold higher in theca-interstitial shells than in isolated GC. Ovarian 5 alpha-reductase activity decreased markedly with age (greater than 60% inhibition in mature, randomly cycling rats as compared to immature rats). In vivo administration of follicle-stimulating hormone (FSH) to immature rats produced a dose-dependent decrease in GC 5 alpha-reductase activity (36 +/- 1.1% and 46 +/- 5.9% inhibition following 12 micrograms and 24 micrograms FSH, respectively). Similarly, the in vitro provision of FSH (100 ng/ml) to cultured GC from immature rats resulted in (36-59%) inhibition in 5 alpha-reduced steroids. Inasmuch as FSH promotes GC development and the advancement of puberty, its ability to "switch-off" ovarian 5 alpha-reductase activity may enhance the formation of biologically potent (i.e. non-5 alpha-reduced) progestins as well as the availability of aromatizable androgens, in the best interests of pubertal steroidogenesis.     

Stimulatory and inhibitory effects of progesterone on follicular development in the hypophysectomized follicle-stimulating hormone/luteinizing hormone-treated hamster

Cyclic hamsters hypophysectomized at estrus (Day 1 of the cycle) and injected with 5 micrograms follicle-stimulating hormone (FSH) on Day 1 and 20 micrograms luteinizing hormone (LH) in polyvinylpyrrolidone (PVP) from Days 1-4 ovulated 15.3 ova, in response to 30 IU human chorionic gonadotropin (hCG) administered at 1500 h on Day 4 (Kim and Greenwald, 1984). When 1 mg progesterone (P4) was administered daily from Days 1-4 concurrent with the above regimen, ovulation increased to 38 ova, a clearcut superovulatory response. However, daily injection of 1, 10, or 100 micrograms P4 plus FSH and LH reduced the number of antral follicles present on the afternoon of Day 4 to 3-4 per ovary, compared to 9 per ovary after FSH-LH alone, and the ovulation rate was drastically reduced with most animals being anovulatory. Substituting 1 mg 17 alpha-hydroxyprogesterone or estradiol cyclopentylpropionate for P4 on Days 1-4 did not alter the number of antral follicles on Day 4 from FSH-LH alone, whereas 1 mg androstenedione or 1 mg testosterone cyclopentylpropionate reduced the number of antral follicles to 3 or less. Hence, the stimulatory effects of 1 mg P4 are not attributable to its conversion to other P4 derivatives. After the concurrent injection of 1 mg P4 and FSH-LH, on the afternoon of Day 3, an average of only 1.8 large preantral follicles was present per ovary. By the morning of Day 4, however, the ovary contained 14 large preantral and early antral follicles in addition to 8 large antral follicles. Injection of hCG at this time resulted in the ovulation of 14.5 ova.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hormonal changes associated with estrogen induced luteolysis in the pregnant hamster: a reevaluation of the luteotropic complex

Hamsters were injected sc on Day 1 of pregnancy (sperm positive) with 50 micrograms estradiol cyclopentylpropionate (ECP) or peanut oil. On Day 5, serum progesterone (P4) was 10.6 ng/ml in controls vs 3.1 ng/ml after ECP. In the ECP group, serum prolactin (PRL) and follicle stimulating hormone (FSH) did not differ from controls but serum luteinizing hormone (LH) was significantly lower than that of the controls, and usually below the sensitivity of the radioimmunoassay (RIA). After ECP, structural signs of luteolysis (weight and histology) and absence of antral follicles characterized the ovary. Injection of an anti-LH serum on Day 4 halved serum P4 levels on Day 5 in control animals but caused no further lowering of P4 in ECP-treated hamsters. Treatment on Days 1-5 with 1.0 IU hCG or 10 micrograms LH plus ECP on Day 1 restored, by the afternoon of Day 5, serum P4 to the control range (9-10 ng/ml) and antral follicles were now present. The results indicate that a large dose of ECP causes luteolysis by reducing LH levels and reinforce the concept of a luteotropic complex in the hamster with PRL and FSH constituting the minimal components and LH serving as a synergist.     

Prolactin involvement in regulation of testicular 5 alpha-reductase activity in the immature rat

Treatment of intact immature (25-day-old) rats with bromoergocryptine (BR), which suppressed prolactin (Prl) secretion, decreased testicular 5 alpha-reductase activity, whereas treatment with Prl increased the enzyme activity in BR-treated animals. Serum luteinizing hormone (LH) concentrations were not reduced by BR treatment or elevated by Prl, suggesting that the BR and Prl effects on enzyme activity were not due to alterations in LH secretion. Hypophysectomy (at 21 days of age) caused a dramatic decrease in testicular 5 alpha-reductase activity, and treatment with LH partially reversed this effect. Treatment of hypophysectomized animals with Prl alone had no effect on the enzyme activity but enhanced the effect of LH. Testosterone propionate, given to hypophysectomized animals in a regimen that increased testicular testosterone to concentrations at least as high as those in intact (sham-hypophysectomized) controls, had no effect on enzyme activity, whether given alone or in combination with LH. These results indicate that Prl is involved, along with LH, in maintaining the high 5 alpha-reductase activity of the prepubertal rat testis; the action of Prl, apparently requiring the presence of LH, may be to decrease the rate of degradation of the enzyme. The data also suggest that the action of LH on testicular 5 alpha-reductase activity is not mediated by its stimulation of testosterone production.     

Role of 5 alpha-reduction in progesterone's ability to release FSH in estrogen-primed ovariectomized rats.

In ovariectomized estrogen-primed rats, progesterone as well as 5 alpha-dihydroprogesterone (5 alpha-DHP) are capable of inducing the release of gonadotropins. This study examined the need of 5 alpha-reduction as a prerequisite for the action of progesterone. The 5 alpha-reductase inhibitor, N,N-diethyl-4-methyl-3-oxo-4-aza-5 alpha-androstane-17 beta-carboxamide was injected at a 1 or 2 mg dose/rat 2 h prior to an injection of 0.4 or 0.8 mg progesterone/kg body weight at 0900 h to immature ovariectomized, estrogen-primed rats and serum was analyzed for LH and FSH at 1500 h. Pituitary and hypothalamic 5 alpha-reductase activity was measured at the time of progesterone administration and at the time of the surge by incubating tissue homogenates with [3H]progesterone. Substrate, ([3H]progesterone) and product ([3H]5 alpha-DHP), were separated by reverse phase HPLC. The pituitary 5 alpha-reductase activity was not blocked at 1500 h. However, both pituitary and hypothalamic 5 alpha-reductase was blocked at the time of progesterone administration. No effect was seen by acute administration of the 5 alpha-reductase inhibitor upon either the 0.4 or 0.8 mg progesterone/kg-induced release of LH and FSH. There was, however, a specific, significant inhibition of progesterone-induced FSH but not LH release when the 5 alpha-reductase inhibition was sustained throughout the afternoon of the gonadotropin surge. These results indicate a biologically significant role for the irreversible 5 alpha-reduction of progesterone in the modulation of the release of FSH.     

Effects of estrogen, pituitary gonadotropins and prolactin on immunohistochemical localization of inhibin subunits in the ovary of hypophysectomized female rats.

Effects of estrogen, pituitary gonadotropins and prolactin on immunohistochemical localization of alpha- and beta A-subunits in the ovaries of hypophysectomized female rats were investigated. Hypophysectomy resulted in disappearance of immunoreactive inhibin subunits in the ovary. Administration of DES, FSH and LH to hypophysectomized rats provoked growth of follicles, and resulted in positive immunostaining for inhibin subunits in the granulosa cells. In contrast to follicle-stimulating hormone (FSH) and luteinizing hormone (LH), prolactin administration failed to demonstrate positive staining for inhibin subunits in the ovary. The present in vivo results suggest that several hormones which are known to stimulate granulosa cell growth and maturation, such as estrogen, FSH and LH, enhance inhibin subunit production by the ovary. The morphologic aspect of inhibin subunit production by the ovary in response to several hormones has been demonstrated in the present study.     

LH contamination may explain FSH effects on rat Leydig cells

Treatment of immature, hypophysectomized male rats with 50 micrograms ovine FSH (NIH-FSH-S12) twice a day for 5 days stimulated the maximum quantity of 17 beta-hydroxyandrogen produced by isolated Leydig cells in response to hCG. Pretreatment of the FSH preparation with an LH antiserum in one study markedly reduced and in another study completely abolished this stimulatory effect of FSH, but only slightly impaired the capacity of the hormone to stimulate the Sertoli cell in vivo (epididymal androgen-binding protein). Administration of another highly potent FSH preparation (LER-1881) had no discernible effects on the dose-response characteristics of the Leydig cells but was superior to the NIH-FSH-S12 in its capacity for stimulating the Sertoli cell. When all hormone preparations were tested for their ability to stimulate steroid secretion from normal Leydig cells in vitro, a close correlation was obtained between their Leydig cell-stimulating activity (a measure of LH contamination) and their capacity to alter Leydig cell responsiveness after in-vivo treatment. FSH treatment had no effects on specific LH binding per 10(6) Leydig cells. It is concluded that the stimulatory influence of FSH on rat Leydig cells may to some extent be a result of the LH contaminating the hormone preparation.