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Several studies suggest that nitric oxide (NO) and cyclic guanosine monophosphate (cGMP) modulate testicular function. In this study, we examined the expression of cGMP-dependent protein kinase G-I (PKG-I), and cyclic nucleotide-gated channel 1 (CNG-1), 2 known mediators of cGMP action, and the expression of soluble guanylyl cyclase (sGC) subunits in the rat testis. Immunohistochemical analysis revealed that the alpha subunit of sGC was expressed in the blood vessels and Leydig cells of adult rat testes. In addition, the sGC alpha subunit was observed in the acrosomal structures of spermatids undergoing the middle and later stages of spermiogenesis, but not in mature spermatozoa. Similar localization and expression patterns were seen for the sGC beta subunit, indicating coexpression of the sGC subunits. PKG-I was expressed in blood vessels and in the acrosomal region of spermatids during the early and middle stages of spermiogenesis but was not observed in Leydig cells or in mature spermatozoa. In contrast to sGC and PKG-I, CNG-1 was expressed only in cytoplasm and the residual bodies of late-stage (17-19) spermatids, with no staining observed in blood vessels and Leydig cells. These results demonstrate that sGC, PKG-I, and CNG-1 are expressed in a stage- and cell-specific manner in the rat testis. The distinct temporal patterns of expression of these components of cGMP signaling pathways suggest different physiological roles for sGC, PKG-I, and CNG-1 in spermiogenesis and steroidogenesis.

Objective: To demonstrate the expression of soluble guanylyl cyclase (sGC) alpha and beta subunits in rat granulosa cells and determine the effects sGC activation on levels of cyclic GMP (cGMP), E2, and cAMP., Design: Basic research study., Setting: University research laboratory., Animal(s): Estrogen-treated immature Sprague-Dawley female rats from which primary cell culture of granulosa cells was obtained., Intervention(s): Functionally immature rat granulosa cells were incubated for 48 hours with media alone, FSH, or FSH plus YC-1, a specific activator of sGC., Main Outcome Measure(s): Expression of sGC alpha and beta subunits was determined by immunoblot analysis. Media concentrations of E2, cAMP, and cGMP were measured by radioimmunoassays., Result(s): Immunoblot analysis of granulosa cells revealed the expression of sGC alpha and beta subunits. While cGMP accumulation was low in cells incubated with media alone or with FSH, cotreatment with FSH plus YC-1 increased cGMP levels approximately five-fold. Incubation of cells with FSH stimulated E2 production in a dose-dependent manner. However, cotreatment of cells with FSH plus YC-1 significantly decreased E2 concentrations at all doses of FSH tested. Similarly, while FSH increased cAMP accumulation from granulosa cells, cotreatment with YC-1 markedly inhibited FSH-stimulated cAMP levels., Conclusion(s): These findings demonstrate the expression of sGC subunits in rat granulosa cells and indicate that activation of sGC increases cGMP levels, which are associated with inhibition of FSH-stimulated E2 production and cAMP accumulation.

The present study examined the gonadotropin surge-inducing actions of estradiol (E(2)), both alone and with progesterone (P(4)), in middle-aged, early persistent-estrous (PE) female rats that had become PE within 35 days. In addition, we also assessed the effect of P(4) on the mating-induced gonadotropin surges in these acyclic animals. Early PE rats were ovariectomized and received E(2) implants (Day 0). On Day 4, an s.c. injection of P(4) (0.5 mg/ 100 g body weight) at 1200 h markedly increased plasma P(4) and elicited both LH and FSH surges, whereas vehicle-treated controls displayed no rise in P(4) or gonadotropins. This observation confirms that at middle age, female rats no longer respond to the positive-feedback stimulation of E(2) on gonadotropin surges whenever the estrous cyclicity ceases. As PE continued, such a surge-inducing action of E(2) plus P(4) became diminished after 75 days of PE and disappeared thereafter. When caged with males, vehicle-treated early PE rats display a mating-induced increase in P(4) from the adrenal along with small gonadotropin surges. The amplitude of these mating-induced gonadotropin surges was enhanced by supplementation with exogenous P(4) in early PE rats. Our findings indicate that during the early phase of PE, the surge-inducing action of E(2) and P(4) remains intact but deteriorates as PE continues. Thus, a deficiency in P(4) secretion during aging may contribute to the diminished gonadotropin surge response in the hypothalamic-pituitary axis and the subsequent cessation of estrous cyclicity.

Soluble guanylyl cyclase (sGC) is activated by nitric oxide (NO) and carbon monoxide, resulting in cGMP production. Recent studies indicate that NO and cGMP influence ovarian functions. However, little information is available regarding the ovarian expression of sGC. The present study examined sGC alpha(1) and beta(1) subunit protein levels in the ovary during postnatal development, gonadotropin-induced follicle growth, ovulation, and luteinization as well as in cultured rat granulosa cells. In postnatal rats, sGC alpha(1) subunit immunoreactivity was high in granulosa cells of primordial and primary follicles on Day 5 but low in granulosa cells of larger follicles on Days 10 and 19. Theca cells of developing follicles, but not stromal cells, also demonstrated moderate sGC alpha(1) immunoreactivity. In gonadotropin- treated immature rats, intense sGC alpha(1) subunit staining was similarly observed in granulosa cells of primordial and primary follicles, but such staining was low in granulosa cells of small antral follicles and undetectable in granulosa cells of large antral and preovulatory follicles. Following ovulation, corpora lutea expressed moderate sGC alpha(1) immunoreactivity. Similar ovarian localization and expression patterns were seen for sGC beta(1), indicating regulated coexpression of sGC subunits. Immunoblot analysis revealed no change in total ovarian sGC alpha(1) and beta(1) subunit protein levels during gonadotropin treatment. Similarly, no effect of FSH on sGC subunit protein levels was apparent in cultured granulosa cells. These findings indicate regulated, cell- specific patterns of sGC expression in the ovary and are consistent with roles for cGMP in modulating ovarian functions.

Objective: To determine the effects of nitric oxide (NO) and cGMP on inhibin A and inhibin subunit mRNA levels from cultured rat granulosa cells., Design: Basic research study., Setting: University research laboratory., Animal(s): Primary cell culture of granulosa cells obtained from estrogen-treated, immature Sprague-Dawley female rats., Intervention(s): Functionally immature rat granulosa cells were incubated for 48 hours with media alone; FSH; forskolin; the NO generator DETA/NO; an inhibitor of soluble guanylyl cyclase (ODQ); and/or a cell-permeable cGMP analog., Main Outcome Measure(s): Media concentrations of inhibin A were measured by solid-phase immunosorbent assay. Inhibin alpha and betaA subunit mRNA levels were determined by Northern and slot blot analyses., Result(s): Whereas FSH caused a 20-fold increase in inhibin A levels compared with untreated granulosa cells, the NO generator DETA/NO significantly inhibited FSH-stimulated inhibin A concentrations. Similarly, cotreatment with FSH plus dibutyryl cGMP significantly attenuated inhibin A concentrations, compared with those in cells treated with FSH alone. Incubation with forskolin (FSK) stimulated inhibin A levels sevenfold, whereas cotreatment with FSK plus DETA/NO or FSK plus dibutyryl cGMP effectively decreased inhibin A concentrations. The effects of NO on inhibin A levels were not prevented by cotreatment with an inhibitor of soluble guanylyl cyclase. In addition, there was no influence of DETA/NO or dibutyryl cGMP on inhibin subunit mRNA levels., Conclusion(s): These findings indicate that NO and cGMP can attenuate inhibin A concentrations through actions at one or more post-FSH receptor sites. These influences may reflect inhibition of inhibin A secretion, rather than gene expression and protein synthesis. In addition, NO decreases inhibin A concentrations through both cGMP-dependent and -independent pathways. These results suggest local roles for NO and cGMP in the regulation of granulosa cell function.

Maintenance of female reproductive competence depends on the actions of several hormones and signaling factors. Recent reports suggest roles for bone morphogenetic proteins (BMPs) in early stages of folliculogenesis. A role for the type I BMP receptor BmprIB as a regulator of ovulation rates in sheep has been described recently, but little is known about the roles of BMP signaling pathways in other aspects of reproductive function. We report here that BMPRIB is essential for multiple aspects of female fertility. Mice deficient in BmprIB exhibit irregular estrous cycles and an impaired pseudopregnancy response. BmprIB mutants produce oocytes that can be fertilized in vitro, but defects in cumulus expansion prevent fertilization in vivo. This defect is associated with decreased levels of aromatase production in granulosa cells. Unexpectedly, levels of mRNA for cyclooxygenase 2, an enzyme required for cumulus expansion, are increased. BmprIB mutants also exhibit a failure in endometrial gland formation. The expression of BmprIB in uterine linings suggests that these defects are a direct consequence of loss of BMP signaling in this tissue. In summary, these studies demonstrate the importance of BMP signaling pathways for estrus cyclicity, estradiol biosynthesis, and cumulus cell expansion in vivo and reveal sites of action for BMP signaling pathways in reproductive tissues.

This study examined the influences of aging and reduced ovarian follicular reserve on estrous cyclicity, estradiol (E(2)) production, and gonadotropin secretion. Young virgin and middle-aged (MA) retired breeder female rats were unilaterally ovariectomized (ULO) or sham operated (control). Unilateral ovariectomy of young rats reduced the ovarian follicular reserve by one-half, to a level similar to that found in MA controls. Unilateral ovariectomy of MA females reduced the follicular pool further, to one half of MA controls. The incidence of regular cyclicity was significantly lower in MA ULO females than in young controls, with intermediate cycle frequency in young ULO and MA controls. Among cyclic rats, the magnitude of the proestrous LH surge was highest in young controls, intermediate in young ULO rats and MA controls, and lowest in MA ULO females. Similarly, ovulation rates were highest in young controls, intermediate in young ULO rats and MA controls, and lowest in MA ULO females. While young ULO rats exhibited augmented secondary FSH surges on estrous morning, middle-aged ULO females displayed secondary FSH levels comparable to young controls. The effects of age and reduced follicle number on estrous cyclicity and gonadotropin secretion were not due to altered E(2) secretion, as preovulatory E(2) levels were similar among all groups. Thus, experimental reduction in the follicular reserve exerts acute effects on the preovulatory LH surge, ovulation rate, and estrous cyclicity in both young and MA rats. However, decreased follicle number increases FSH levels only in young rats, indicating aging-related alterations in the feedback regulation of FSH.

WT1 is a zinc finger protein with transcriptional repressor activity on several growth factor and growth factor receptor genes. In the ovary, a potential role for WT1 in the suppression of the development of immature follicles has been demonstrated. Here, gel retardation assays further showed that recombinant WT1 protein interacted with consensus DNA sequences in the inhibin-alpha gene promoter. We investigated the pattern of WT1 expression in a wide variety of species and also over the reproductive life span in rats. In chicken ovaries, Northern blot analysis revealed the presence of WT1 transcript in small healthy white follicles (1-5 mm in diameter) and its absence in small yellow (6-12 mm in diameter) or larger follicles (F1-F5). In pig and monkey ovaries, WT1 expression was limited to granulosa cells of preantral follicles, as shown by in situ hybridization analysis. In rats, Northern blot analyses demonstrated the presence of WT1 transcript in the ovaries of young (3-mo-old) and middle-aged (9-mo-old) rats on the proestrous day, with a decrease in old (12-mo-old) rats in persistent estrus. In situ hybridization analysis further suggested that the decrease in WT1 expression in aging ovaries was associated with fewer immature follicles. Thus, WT1 expression is restricted to immature follicles in diverse avian and mammalian species and over the reproductive life span in rats. These data demonstrated that WT1 is a marker for immature follicles and suggested a potential role of this transcriptional repressor in the slow growth of early follicles.

The present study examined the effects of progesterone (P4) treatments on estrous cyclicity and the loss of ovarian follicles during aging. Young rats received repeated treatments with P4 or empty implants between 3.5 and 8 mo of age. At 8 mo, ovaries were obtained from some animals to determine the numbers of resting follicles, and estrous cycle patterns and hormone levels were determined from other groups of treated females. In contrast to the cyclic increases in P4, estradiol (E2), LH, and FSH in control animals, P4-implanted rats exhibited elevated serum P4 but low E2, LH, and FSH levels. After implant treatments, the follicular reserve was significantly (p < 0.05) larger in P4-treated females (2012 +/- 297 resting follicles per ovary, n = 5 rats per group) than in regularly cyclic control rats (713 +/- 226 follicles per ovary, n = 7). The effects of P4 implants on the follicular reserve were associated with a subsequently higher incidence of regular estrous cycles after P4 treatment. These results demonstrate that P4 prevents cyclic increases in E2 secretion and is associated with a conservation of the ovarian follicular reserve and the maintenance of regular estrous cycle patterns, indicating a protective effect of P4 on the age-related loss of ovarian follicles.

The stimulatory effects of gonadotropins on antral and preovulatory follicles are well known, but conflicting results have been reported regarding the gonadotropin responsiveness and dependency of preantral follicles. Taking advantage of the relatively uniform development of the first wave of follicles in the postnatal rat ovary, we evaluated the role of endogenous and exogenous gonadotropins on preantral follicle development. Reduction of the high levels of gonadotropins present in juvenile rats by either hypophysectomy (at Day 15) or GnRH antagonist treatment (starting from Day 11 of age) resulted in decreased ovarian weight at Day 19 of age that was associated with a reduced number of developing follicles and increased atresia of remaining follicles. In contrast, treatment with FSHctp (a long-acting FSH agonist) in intact (Days 5-19 of age), hypophysectomized (Days 15-19), or GnRH antagonist-treated (Days 11-19) animals resulted in increased ovarian weight and follicle development as determined histologically and by inhibin-alpha expression. A dose-dependent stimulatory effect of hCG on ovarian weight was seen when animals were cotreated with FSHctp and the GnRH antagonist. At low doses of hCG, augmentation of antral follicle formation occurred, whereas higher doses of hCG led to morphological signs of luteinization. These findings demonstrate the important role of endogenous gonadotropins in preantral follicle development and indicate that preantral follicles are highly responsive to exogenous gonadotropins.

Objective: To evaluate follicular FSH and LH requirements during suppression of endogenous gonadotropins with the GnRH antagonist Nal-Glu and whether LH-like activity could be supplied by administering subcutaneous hCG., Design: Randomized clinical trial., Participants: Thirty-two normally cycling females in the late follicular phase (dominant follicle mean diameter > or = 14 mm)., Intervention: Twelve normal women were randomized to receive 150 IU IM FSH with or without 75 IU SC hCG; 11 subjects were randomized to receive 225 IU FSH with or without 50 IU SC hCG; 9 women received 150 or 225 IU IM hMG. Subjects returned the next day for repeat blood sample and ultrasound., Results: Continued follicular maturation, as evidenced by rising E2 levels, correlated with serum immunoactive and bioactive FSH levels and was unrelated to bioactive LH-hCG. Two hundred twenty-five international units of exogenous FSH consistently supported follicular maturation. There was a similar increase in mean follicular diameter in women with an E2 rise versus those with a plateau or fall. In subjects receiving SC mini-dose hCG, serum bioactive LH-hCG levels were increased significantly and were similar to levels before Nal-Glu., Conclusions: During administration of a GnRH-a, the maturing follicle appears to require only FSH support. In markedly hypogonadotropic women, mini-dose hCG may be a more practical alternative to recombinant LH to promote normal follicle maturation.

On the basis of earlier observations of changing testicular LH receptor levels during postnatal development of the rat, we analyzed the levels of LH receptor mRNA transcripts in testes of rats at ages 5-70 days. Extracted testis RNA, prepared for Northern blotting, was hybridized with a specific LH receptor cRNA probe derived from subcloned cDNA corresponding to the extracellular domain of the receptor. Six LH receptor mRNA transcripts with molecular sizes of 7.8, 7.0, 4.2, 2.5, 1.8, and 1.2 kb were identified. Of these, the 1.2- and the 1.8-kb mRNA transcripts presumably code for truncated forms of LH receptor. At 5 days, only the 1.8- and the 4.2-kb mRNA transcripts were observed. Additional 7.0- and 1.2-kb transcripts appeared at 10 and 15 days, respectively. From the age of 25 days through adulthood, all six mRNA transcripts were observed. Densitometric analyses revealed that the amounts of the 7.0- and 1.8-kb mRNA transcripts correlated well with LH receptor levels, while the 4.2-kb transcript showed high levels earlier in life with poor correlation to LH receptor number. Because the 1.8 kb receptor transcript lacked transmembrane domains, the present results suggest the 7.0-kb LH receptor transcript as the likely candidate to encode the functional receptor. These data provide the basis for future analyses of the molecular regulation of LH receptor expression.

Follitropin (FSH) is a pituitary glycoprotein hormone that is essential for the development of ovarian follicles and testicular seminiferous tubules. FSH is used clinically to stimulate follicular maturation for in vitro fertilization and treatment of anovulatory women. One issue regarding the clinical use of FSH is its short half-life in the circulation. To address this point, we constructed chimeric genes containing the sequence encoding the C-terminal peptide of the chorionic gonadotropin beta subunit (CG beta) fused to the translated sequence of the human FSH beta subunit (FSH beta). This region of CG beta is important for maintaining the prolonged plasma half-life of human CG dimer. The presence of the C-terminal peptide sequence did not significantly affect assembly of FSH beta with the alpha subunit or secretion of the dimer. In vitro receptor binding and steroidogenic activity of dimer bearing the FSH beta-C-terminal peptide chimera were the same as wild-type FSH. However, both the in vivo potency and half-life in circulation of the dimer bearing either one or two C-terminal peptide units were enhanced. Dimers containing FSH beta-CG beta chimeras could serve as potent FSH agonists for clinical use, and the present strategy may have wide applications for enhancing the in vivo half-life of diverse proteins.

Objective: To examine the possible impact of abnormal adrenal steroidogenesis on the ovarian dysfunction seen in polycystic ovarian disease (PCOD)., Design: Prospective analysis of blood sampling monthly for 6 months, then three times weekly for 90 days., Setting: Tertiary institutional outpatient care., Participants: Six anovulatory women with a diagnosis of PCOD., Intervention: Six-month suppression with gonadotropin-releasing hormone agonist (GnRH-a) followed by suppression with dexamethasone (DEX) for 90 days., Main Outcome Measures: Serum levels of testosterone (T), androstenedione (A), dehydroepiandrosterone (DHEA), dehydroepiandrosterone sulfate (DHEAS), cortisol, estradiol (E2), progesterone (P), follicle-stimulating hormone (FSH), luteinizing hormone (LH) and bioactive LH., Results: Gonadotropin-releasing hormone agonist administration suppressed greater than 60% of the circulating levels of T and A, suggesting an ovarian origin. Minimal changes of DHEA, DHEAS, and cortisol were seen. With the addition of DEX, there was greater than 90% suppression of the total circulating A, T, DHEA, DHEAS, and cortisol, supporting the adrenal origin of the non-GnRH-a suppressible androgens. Excessive ovarian T and A secretion returned during the 90-day recovery study period in spite of rises of FSH concentrations that changed the ratio of FSH to LH in all subjects. Four of the six women failed to ovulate. In comparison of the women who did and did not ovulate during recovery, no differences in absolute levels or changes in concentrations of steroids or gonadotropins could be detected., Conclusions: Using sequential and simultaneous administration of GnRH-a and DEX, we were able to delineate the contributions of the ovaries and adrenals to the abnormal steroidogenesis seen in PCOD. Despite prolonged suppression of ovarian and then adrenal steroidogenesis, ovarian dysfunction, evidenced by abnormal androgen production, returned with cessation of agonist administration.

Regularly cyclic, middle-aged female rats exhibit a decreased incidence of fertility, and those females that are fertile produce smaller litters. This decreased litter size is directly related to a reduced number of normal blastocysts available for implantation. Recent evidence indicates that embryonic abnormalities in middle-aged rats become apparent as early as Day 2 of pregnancy. Inasmuch as the semicircadian secretion of prolactin (PRL) is essential for the rescue of corpora lutea during early gestation and luteal production of progesterone (P) and estradiol (E2) in sufficient quantities is obligatory for embryonic development and implantation, the present study examined the profiles of plasma PRL, P, and E2 during the first 3 days of pregnancy in both young and middle-aged rats and assessed the embryonic development in these same animals. Regularly cyclic, middle-aged (9-11 mo) and young (4-5 mo) rats were cannulated via the right jugular vein on Diestrus Day 2 and mated with fertile males on proestrus. The next morning, sperm in the vaginal lavage confirmed mating, and that day was designated Day 1 of pregnancy. Beginning at 1400 h on Day 1 and continuing to 2400 h on Day 2, serial blood samples were taken at 2-h intervals for PRL assay. In the first experiment, samples were also collected at 8-h intervals during Days 1-3 for measurement of plasma P.(ABSTRACT TRUNCATED AT 250 WORDS)

Down-regulation of plasma membrane receptors by homologous hormones has been found in diverse cell types. In testicular Leydig and ovarian luteal cells, treatment with LH/hCG decreases LH receptor content. Although suppression of LH-binding sites may result from ligand-induced receptor internalization, sequestration, and/or phosphorylation, the gonadotropins may also regulate receptor mRNA levels. We examined the regulation of testis LH receptor mRNAs in adult rats that received 10 or 200 IU hCG, using cRNA probes derived from the 5' extracellular domain (EC) or the 3' transmembrane domain (TM) of the rat receptor cDNA. Probe EC hybridized to predominant signals of 7 and 1.8 kilobases (kb) and weaker signals of 4.2 and 2.5 kb. However, probe TM hybridized to the three larger forms of the LH receptor mRNA, but not to the 1.8-kb species, suggesting that the latter form lacks the transmembrane domain. After 6 and 12 h of treatment with 200 or 10 IU hCG, respectively, hybridization to the larger mRNA species decreased by more than 60%, preceding decreases in testicular [125I]hCG binding. These transcripts were further inhibited (greater than 93%) between 24-72 h after hCG treatment and returned to 40% and 100% of control levels by days 6 and 9, respectively. In contrast, the truncated 1.8-kb LH receptor transcript was not affected by hCG treatment, indicating a differential suppressive effect of the ligand on its receptor mRNA levels. In the ovary, hybridization to probe EC revealed four transcripts with similar sizes as those found in the testes, with a predominant 7-kb species.(ABSTRACT TRUNCATED AT 250 WORDS)

Regularly cyclic, middle-aged female rats exhibit a decreased incidence of fertility, and those females that are fertile produce small litters. These decreases in fertility and litter size are associated with reduced numbers of normal blastocysts formed and implanted, suggesting that pre- and/or peri-implantation failures may be the causes for these aging-related reproductive declines. The present study examined the relationships and influence of circulating estradiol (E2) and progesterone (P) levels on early embryonic development and implantation in middle-aged rats. Serial blood samples obtained from cannulated, middle-aged pregnant rats revealed minor decreases in plasma P and increases in E2 levels during Days 2-4 of pregnancy, compared to young pregnant rats, resulting in significantly (p less than 0.001) decreased plasma P/E2 ratios. These alterations in endogenous hormone secretion in middle-aged pregnant rats were associated with fewer normal blastocysts on Day 5 of pregnancy and reduced numbers of normally implanting embryos. Correlation analysis further revealed a significant (p less than 0.05) inverse relationship between mean circulating E2 levels and numbers of normal conceptuses on Day 12 of gestation. Moreover, s.c. administration of P implants (in Silastic) to middle-aged pregnant rats increased serum P levels by about 34-40 ng/ml, and significantly (p less than 0.05) reduced the incidence of abnormal embryos before implantation. In contrast, treatment with E2 minipumps produced a sustained rise in serum E2 (by about 7-15 pg/ml) and resulted in the complete absence of embryos in the reproductive tracts by Day 5 of pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)

Recent reports suggest that activin (the dimer of inhibin beta subunits with FSH-releasing activity) has specific receptors on ovarian granulosa cells. The present study examined the effects of purified porcine activin on inhibin secretion and mRNA levels in granulosa cells obtained from immature, estrogen-treated rats. Cells were cultured for 48 h in culture media, or media containing FSH (10 ng/ml) and/or activin (30 ng/ml). Western blot analyses performed with affinity-purified antisera to inhibin alpha- and beta A-subunits revealed that treatment with either FSH or activin increased the secretion of inhibin alpha beta dimer (Mr 30,000), with a further increase after cotreatment. These results were confirmed by an inhibin alpha-subunit RIA, which revealed 7-, 14-, and 71-fold increases in the secretion of immunoreactive inhibin-alpha by activin, FSH, and activin plus FSH, respectively. TGF beta, a structural homolog of activin, also stimulated inhibin release, whereas follistatin was ineffective. Total RNA from cultured cells was hybridized with 32P-labeled inhibin alpha-subunit cRNA or beta-actin cDNA probes, and inhibin-alpha message levels were normalized with beta-actin mRNA levels. Northern blot analysis revealed that treatment with FSH and activin increased hybridization of a 1.5 kilobase (kb) message, corresponding to the inhibin alpha-subunit mRNA. Slot blot analyses indicated a 6- and 8-fold stimulation of inhibin alpha-subunit mRNA levels by FSH and activin, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

We have recently reported that successive treatments of young virgin rats with progesterone (P) implants produce elevated circulating P and consistently low estradiol (E2) concentrations, and subsequently delay the aging-associated reproductive decline. Inasmuch as E2 has been implicated in causing the loss of regular estrous cyclicity in aging rats, the present study examined if the concomitant presence of moderately increased circulating E2 levels could counteract the effects of P implants on reproductive aging. Starting at 3 1/2 mo and continuing to 8 mo of age, regularly cyclic, virgin rats received either s.c. Silastic implants of P (P-implanted), blank Silastic implants (virgin controls), or P + E2 implants (P + E2-implanted) for 3 wk, followed by implant removal for 1 wk. Each of these implant treatments was repeated in the same female rats 5 times. Blood samples were obtained on different days of the estrous cycle from the control group and on Day 11 of successive treatments with P or P + E2 implants for measurements of serum P and E2 values. At 8 1/2 and 10 mo of age, estrous cyclicity of these same virgin rats was again monitored, and 10-mo-old regularly cyclic females from each treatment group were mated with young fertile males to complete term pregnancies. While virgin controls showed cyclic increases in E2 and P secretion during the estrous cycle, P-implanted virgins exhibited consistently low serum E2 and moderately increased P levels during 5 successive treatments. The latter indicates a potent inhibition of ovarian E2 secretion by P implants.(ABSTRACT TRUNCATED AT 250 WORDS)

Inoculation of cyclic female rats with the prolactin (Prl)/growth hormone-secreting pituitary tumor, MtT.W15, resulted in a cessation of estrous cyclicity within 5--10 days. Associated with this acyclicity was a persistently low serum concentration of estradiol and marked increases in both circulating Prl and progesterone. At Day 26 of acyclicity, basal serum luteinizing hormone (LH) values measured in samples taken every 20 min from 0900--1100 h were significantly reduced when compared to cyclic, nontumor animals on diestrus Day 2. There was no difference in basal follicle-stimulating hormone (FSH) concentrations. In a separate group of acyclic, tumor-bearing females 42--56 days after transplantation, a single s.c. injection of 20 micrograms estradiol benzoate (EB) at 1030 h elicited significant increases in both serum LH and FSH values between 1700 and 1830 h on the next day. The magnitude of the LH surge was reduced and that of FSH was increased in tumor-bearing animals when compared to cyclic, nontumor females given a similar EB injection on diestrus Day 1. These results demonstrate that chronic hyperprolactinemia is associated with inhibition of basal LH secretion and ovarian estrogen production and an increase in circulating progesterone concentrations. Nevertheless, the stimulatory feedback effects of estrogen on LH and FSH release are still present and functioning in acyclic female rats under chronically hyperprolactinemic conditions. These data suggest that the cessation of regular ovulatory cycles associated with hyperprolactinemia may be due to a deficiency of LH and/or estrogen secretion, but not to a lack of central nervous system response to the stimulatory feedback action of estrogen.

In multiparous rats, the incidence of regular estrous cyclicity and fertility decreases markedly at middle age. However, recent studies have shown that repeated pregnancies or progesterone (P) implants can subsequently cause retired breeder females to maintain regular cyclicity for an extended period of time; these results suggest a P-mediated deceleration of reproductive aging. In the present study, we examined the relation of ovarian steroid levels in young virgin females to their subsequent estrous cyclicity and reproductive function during aging as compared to multiparous females. Beginning at 4 mo of age and continuing to 6 mo of age, regularly cyclic virgin rats received either consecutive P implants (n = 41) or no implants (controls, n = 45) for 3 wk, followed by implant removal for 1 wk. Additional females (n = 72) were mated and allowed to undergo repeated pregnancies at 4, 6 1/2, and 8 mo of age. Blood samples were obtained throughout the estrous cycle (virgin females), during pregnancy (multiparous rats), and on Day 11 of successive treatments with P implants (virgins with P implants) for P, estradiol (E2), and testosterone (T) measurements. Subsequently, regularly cyclic females from all three groups were mated with fertile males to undergo term pregnancies at 10 and 12 mo of age. While the virgin controls showed cyclic increases in P, T, and E2 secretion during their estrous cycles, the P-implanted females had persistently low E2 and high P and T levels during treatment, which indicates an inhibition of ovarian E2 synthesis by P.(ABSTRACT TRUNCATED AT 250 WORDS)