Selective failure of androgens to regulate follicle stimulating hormone beta messenger ribonucleic acid levels in the male rat

FSH beta, as well as LH beta, and alpha-subunit mRNA levels were examined in the pituitary glands of male rats after sex steroid replacement at various times (7, 28, or 90 days) after orchiectomy. Testosterone propionate, dihydrotestosterone propionate, or 17 beta-estradiol benzoate (E) were administered daily for 7 days before killing, to assess the role of different gonadal steroids on gonadotropin subunit mRNA levels. Subunit mRNAs were determined by blot hybridization using rat FSH beta genomic DNA, and alpha and LH beta cDNAs. At all time points, alpha and LH beta mRNAs increased after gonadectomy and fell toward normal levels with either androgen or estrogen replacement. FSH beta mRNA levels increased variably postcastration: 4-fold at 7 days, 2-fold at 28 days, and 4- to 5-fold at 90 days. Although E replacement uniformly suppressed FSH beta mRNAs, neither testosterone propionate nor dihydrotestosterone propionate administration suppressed FSH beta mRNA levels at any time point after orchiectomy. These data demonstrate that there is a relative lack of negative regulation of FSH beta mRNA levels by androgens in a paradigm in which E administration results in marked negative regulation of FSH beta mRNA levels. Thus, in the male rat, estrogens negatively regulate all three gonadotropin subunit mRNA levels while androgens negative regulate LH beta and alpha-subunit but fail to suppress FSH beta mRNAs.     

Differential regulation of pituitary gonadotropin subunit messenger ribonucleic acid levels in photostimulated Siberian hamsters

FSH levels begin to rise 3-5 days after male Siberian hamsters are transferred from inhibitory short photoperiods to stimulatory long photoperiods. In contrast, LH levels do not increase for several weeks. This differential pattern of FSH and LH secretion represents one of the most profound in vivo examples of differential regulation of the gonadotropins. The present study was undertaken to characterize the molecular mechanisms controlling differential FSH and LH synthesis and secretion in photostimulated Siberian hamsters. First, we cloned species-specific cDNAs for the three gonadotropin subunits: the common alpha subunit and the unique FSHbeta and LHbeta subunits. All three subunits share high nucleotide and predicted amino acid sequence identity with the orthologous cDNAs from rats. We then used these new molecular probes to examine the gonadotropin subunit mRNA levels from pituitaries of short-day male hamsters transferred to long days for 2, 5, 7, 10, 15, or 20 days. Short-day (SD) and long-day (LD) controls remained in short and long days, respectively, from the time of weaning. We measured serum FSH and LH levels by RIA. FSHbeta, LHbeta, and alpha subunit mRNA levels were measured from individual pituitaries using a microlysate ribonuclease protection assay. Serum FSH and pituitary FSHbeta mRNA levels changed similarly following long-day transfer. Both were significantly elevated after five long days (2.3- and 3.6-fold, respectively; P < 0.02) and declined thereafter, but they remained above SD control values through 20 long days. Alpha subunit mRNA levels also increased significantly relative to SD control values (maximum 2-fold increase after seven long days; P < 0.03), although to a lesser extent than FSHbeta. Neither serum LH nor pituitary LHbeta mRNA levels changed significantly following long-day transfer. The results indicate that long-day-associated increases in serum FSH levels in Siberian hamsters reflect an underlying increase in pituitary FSHbeta and alpha subunit mRNA accumulation.     

Androgens positively regulate follicle-stimulating hormone beta-subunit mRNA levels in rat pituitary cells

We have shown previously that androgens negatively regulate LH alpha and beta-subunit mRNA levels, but have little or no effect on FSH beta mRNA levels in rats in vivo. In contrast, estrogen negatively regulates all three gonadotropin subunit mRNA levels in vivo. We have examined the effects of these sex steroids on gonadotropin subunit synthesis directly at the level of the pituitary gland by using cultured rat pituitary cells. Adult female and male rat pituitaries were dissected, dispersed enzymatically, and maintained in culture for 2 days. At that time, cells were treated for varying lengths of time with either medium alone or sex-steroid hormone treatments (estradiol or testosterone). Dose-response and time-course experiments were performed. Cells were then harvested and total RNA was extracted. Gonadotropin subunit mRNA levels were assessed by blot hybridization techniques. Sex-steroid hormones were added to achieve final concentrations ranging from 10(-12) to 10(-6) M for dose response experiments and 10(-8) M for time-course experiments. Testosterone treatment (10(-8) M) increased FSH beta mRNA levels 3-fold in females (P less than 0.01) and males (P less than 0.05), but had no effect on alpha or LH beta mRNA levels in either sex. Dose-related increases in FSH beta mRNA levels with increasing concentrations of testosterone were observed in both female and male pituitary cell cultures. Time-course studies revealed that the testosterone-stimulated increases in FSH beta mRNA levels are statistically significant by 12 h and 6 h after hormone addition in female and male cultures, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of estradiol on gonadotropin subunit messenger ribonucleic acid amounts during an induced gonadotropin surge in anestrous ewes

The preovulatory gonadotropin surge in the sheep was recently characterized by a divergent pattern of LH beta and FSH beta mRNAs immediately preceding this event. It is not clear whether this pattern is due to estradiol (E2), inhibin or other effectors. In this study, to determine if E2 may be involved in the divergent beta mRNA patterns seen during the surge, gonadotropin surges were induced in anestrous ewes (An) by E2 (An + E2) and several parameters were then measured. These included the amounts of alpha, LH beta, and FSH beta mRNAs, as assessed by solution hybridization assays, plus pituitary and serum gonadotropin concentrations. The values were compared with those observed in control, An ewes, to assess the effect of E2. The E2 treatment resulted in LH and FSH surges that appeared to be similar to the normal surges seen during the breeding season. Concomitantly, the E2 treatment lowered pituitary concentrations of FSH (P less than 0.05), while LH amounts did not change. Although the effect of E2 on gonadotropin subunit mRNA amounts varied depending upon the individual subunit, the changes that were observed paralleled changes reported during the preovulatory surge of the cycle. Specifically, alpha mRNA amounts increased significantly (P less than 0.001) while FSH beta mRNA amounts fell dramatically (P less than 0.001). Moreover, LH beta mRNA amounts were slightly increased, although not significantly by E2. These results demonstrate that E2 effects changes in the amounts of the gonadotropin subunit mRNAs during an induced gonadotropin surge in An ewes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Stimulatory effects of insulin-like growth factor 1 on expression of gonadotropin subunit genes and release of follicle-stimulating hormone and luteinizing hormone in masu salmon pituitary cells early in gametogenesis

Insulin-like growth factor-I (IGF-I) has been shown to be involved in pubertal activation of gonadotropin (GTH) secretion. The aim of this study was to determine if IGF-I directly stimulates synthesis and release of GTH at an early stage of gametogenesis. The effects of IGF-I on expression of genes encoding glycoprotein alpha (GPalpha), follicle-stimulating hormone (FSH) beta, and luteinizing hormone (LH) beta subunits and release of FSH and LH were examined using primary pituitary cells of masu salmon at three reproductive stages: early gametogenesis, maturing stage, and spawning. IGF-I alone or IGF-I + salmon GnRH (sGnRH) were added to the primary pituitary cell cultures. Amounts of GPalpha, FSHbeta, and LHbeta mRNAs were determined by real-time PCR. Plasma and medium levels of FSH and LH were determined by RIA. In males, IGF-I increased the amounts of all three subunit mRNAs early in gametogenesis in a dose-dependent manner, but not in the later stages. In females, IGF-I stimulated release of FSH and LH early in gametogenesis, whereas no stimulatory effects on the subunit mRNA levels were observed at any stage. IGF-I + sGnRH stimulated release of FSH and LH at all stages in both sexes, but had different effects on the subunit mRNA levels depending on subunit and stage. The present results suggest that IGF-I itself directly stimulates synthesis and release of GTH early in gametogenesis in masu salmon, possibly acting as a metabolic signal that triggers the onset of puberty.     

Inhibin, activin, and follistatin: regulation of follicle-stimulating hormone messenger ribonucleic acid levels

Primary pituitary cell cultures derived from adult male rats were used to explore the direct effects of purified porcine inhibin and follistatin, and recombinant human activin A on FSH beta, as well as LH beta and alpha-subunit mRNA levels. Subunit mRNAs were determined by blot hybridization using alpha, LH beta, and FSH beta cDNA and genomic fragments. Treatment with inhibin for 72 h significantly suppressed alpha and FSH beta mRNA levels with parallel changes in FSH secretion. No change in LH beta mRNA levels was observed. A decrease in FSH beta mRNA to undetectable levels was seen 4 h after inhibin administration. Recombinant human Activin A caused dose-dependent and parallel increases in FSH beta mRNA levels and FSH secretion. This increase was evident at 4 h after activin administration and maintained at longer times. alpha and LH beta mRNA levels remained unchanged. Follistatin addition to cultures for 72 h significantly reduced FSH beta mRNA levels. In a time-course experiment, a reduction in FSH beta mRNA to undetectable levels was observed 24 h after follistatin administration. There were no changes in alpha or LH beta mRNA levels. These data demonstrate that the actions of these gonadal peptides on FSH secretion may be accounted for, at least in part at the level of biosynthesis, by reductions in FSH beta mRNA levels directly at the level of the anterior pituitary gland.     

Divergent changes in the concentrations of gonadotropin beta-subunit messenger ribonucleic acid during the estrous cycle of sheep

Cyclic changes in the production of the pituitary gonadotrophic hormones, LH and FSH are essential events in the maintenance of the reproductive system of female mammals. While studies have examined changes in the secretion of LH and FSH during the estrous cycle and demonstrated the importance of these hormones in regulation of ovarian development and gametogenesis, considerably less is known concerning the regulation of the biosynthesis of these hormones. Although initial studies have examined changes in LH subunit mRNA concentrations during the rat and ovine estrous cycles, no information concerning the physiological regulation of FSH beta mRNA concentrations has been available. In the present study we have examined the relationship between pituitary concentrations of LH and FSH subunit mRNAs and the serum concentrations of these gonadotropins. The results demonstrate a very different pattern of change for FSH beta subunit mRNA than that observed for alpha and LH beta subunit mRNAs. In fact, FSH beta mRNA concentration decline substantially during the preovulatory period, reaching minimal values at a time when alpha and LH beta mRNA levels are near maximal. Furthermore, this decline in FSH beta mRNA amounts occurs when serum FSH concentrations are maximal. Thus, FSH beta mRNA concentrations follow a very different pattern than that of serum FSH. In contrast, LH beta mRNA and serum LH concentrations tend to increase at the same time. These findings provide evidence that concentrations of LH beta and FSH beta mRNAs are likely regulated by different mechanisms.     

Sequence analysis and expressional regulation of messenger RNAs encoding beta subunits of follicle-stimulating hormone and luteinizing hormone in the red-bellied newt, Cynops pyrrhogaster

Two distinct cDNAs encoding beta subunits of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were cloned from the cDNA library constructed for the pituitary of the red-bellied newt, Cynops pyrrhogaster, and sequenced. The newt FSHbeta and LHbeta cDNAs encode polypeptides of 129 and 131 amino acids, including signal peptides of 20 and 19 amino acids, respectively. The number and position of cysteine and N-glycosylation in each of the beta subunits of FSH and LH, which are considered essential for assembly of the alpha subunit, are well conserved between the newt and other tetrapods. The high homology (41.6%) between the beta subunits of newt FSH and LH imply less specificity of FSH and LH in gonadal function. One cDNA encoding the common polypeptide chain alpha subunit of FSH and LH was also isolated from the newt pituitary gland. The mRNAs of FSHbeta, LHbeta, and the alpha subunit were expressed only in the pituitary gland among various newt tissues. Double-staining with in situ hybridization and immunohistochemistry revealed coexpression of FSHbeta and LHbeta in the same newt pituitary cells. Ovariectomy induced a significant increase in FSHbeta mRNA levels, but there was no significant change in LHbeta or alpha subunit mRNA levels compared with those in control animals. Taken together, these data suggest that two kinds of gonadotropins, namely FSH and LH, are expressed in the same gonadotropin-producing cells in the pars distalis of the newt as well as in other tetrapods and that the expression of FSHbeta is negatively regulated by the ovaries.     

Divergent responses of gonadotropin subunit messenger RNAs to continuous versus pulsatile gonadotropin-releasing hormone in vitro

Episodic GnRH input is necessary for the maintenance of LH and FSH secretion. In the current study we have assessed the requirement of a pulsatile GnRH signal for the regulation of gonadotropin alpha- and beta-subunit gene expression. Using a dispersed rat pituitary perifusion system, GnRH (10 nM) was administered as a continuous infusion vs. hourly pulses. Secretion of free alpha-subunit, LH, and FSH were monitored over 5-min intervals for the entire 12-h treatment period before the responses of alpha, LH beta, and FSH beta mRNAs were assessed. Basal release of all three glycoproteins declined slowly over 6-8 h before reaching a plateau. The cells were responsive to each pulse of GnRH, but continuous GnRH elicited only a brief episode of free alpha-subunit, LH, and FSH release, followed by a return to unstimulated levels. Despite the similar patterns of secretion, differences were observed in the responses of gonadotropin mRNAs to the two modes of GnRH. alpha mRNA increased in response to continuous (1.6-fold) or pulsatile (1.7-fold) GnRH. FSH beta mRNA was suppressed to 48% of the control value after continuous GnRH, but was stimulated over 4-fold by the pulses. LH beta mRNA was unresponsive to either treatment paradigm. We conclude that in vitro 1) alpha mRNA levels are increased in response to GnRH independent of the mode of stimulation; 2) under the conditions studied, LH beta mRNA levels are unresponsive to either mode of GnRH input; and 3) the response of FSH beta mRNA to GnRH is highly dependent on the mode of administration, with levels depressed in response to continuous GnRH, but stimulated by pulsatile GnRH.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of inhibin from primate Sertoli cells and GnRH on gonadotropin subunit mRNA in rat pituitary cell cultures

Partially purified inhibin from primate Sertoli cell culture medium (pSCl) suppresses both LH and FSH secretion from cultured rat pituitary cells stimulated with GnRH. To examine the mechanism of action of pSCl, we have measured steady state levels of mRNAs for the gonadotropin subunits in pituitary cell cultures exposed to 10 nM GnRH for 6 h in control or pSCl-containing medium (short term) and after 72-h pretreatment with pSCl or control medium (long term). Messenger RNA levels were determined by Northern analysis using specific cDNA probes for rat FSH beta, LH beta, and the common alpha-subunit. In the long term experiments, pSCl inhibited GnRH-stimulated release of FSH (47.4 +/- 3.3% of control), LH (69.2 +/- 2.3%), and free glycoprotein alpha-subunit (74.2 +/- 4.5%), and intracellular FSH declined to 88.4 +/- 3.5% of control. Concentrations of the subunit mRNAs were all decreased: FSH beta to 54.4 +/- 5.0%, LH beta to 79.6 +/- 9.4%, and alpha to 70.8 +/- 8.7% of control. In the short-term experiments, pSCl also suppressed FSH, LH, and alpha-subunit secretion to 75.9 +/- 3.6%, 79.5 +/- 2.1%, and 90.9 +/- 1.8% of control, respectively. Intracellular LH and alpha-subunit levels were significantly increased in cells treated for 6 h with GnRH and pSCl (155 +/- 18%, 145 +/- 14% of control), while FSH was comparable to control. After 6 h, pSCl selectively reduced the level of mRNA for FSH beta (56.5 +/- 5.8% of control).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of testosterone metabolites on serum gonadotropin concentrations in immature male rats.

The ability of testosterone, androsterone, 5alpha-androstane-3alpha,17beta-diol, and 5alpha-androstane-3beta,17beta-diol to prevent the castration-induced rise in serum gonadotropin levels was investigated in immature male rats. Rats castrated at 30 days of age were treated once per day by subcutaneous injection of 12.5-100 mug of the steroid per 100 g body weight per day for 3 days, beginning on the day of castration. The animals were sacrificed 24 h after the last injection. Testosterone propionate, androsterone propionate, and 5alpha-androstane-3alpha,17beta-diol dipropionate were also tested at the approximate molar equivalent of 100 mug of the free alcohol form per 100 g body weight per day. Testosterone propionate and 5alpha-androstane-3alpha,17beta-diol were the only compounds tested that prevented the castration induced rise in luteinizing hormone (LH) concentrations. Testosterone propionate also inhibited the rise in follicle stimulating hormone (FSH) concentrations whereas 5alpha-androstane-3alpha,17beta-diol inhibited the rise in FSH in one but not in another experiment. These were the only compounds tested that affected serum FSH concentrations. The lower doses of testosterone tested significantly increased serum LH, but not FSH concentrations compared to castrate control animals. The highest dose tested partially inhibited the rise in serum LH concentrations. Both androsterone and androsterone propionate maintained ventral prostate weights. Although neither compound prevented the castration induced rise in serum LH, two groups receiving androsterone had serum LH concentrations significantly lower than the castrate control group. 5alpha-Androstane-3beta,17beta-diol and 5alpha-androstane-3alpha,17beta-diol dipropionate failed to maintain ventral prostate weights or prevent the rise in serum gonadotropin levels. These results indicate that 5alpha-androstane-3alpha,17beta-diol is capable of preventing the castration induced rise in serum LH concentrations in the immature male rat and thus may participate in the regulation of LH secretion in these animals.     

Possible involvement of inhibin in altered follicle-stimulating hormone (FSH) secretion during dissociated luteinizing hormone (LH) and FSH release: unilateral castration and experimental cryptorchidism

Male rats were either unilaterally or bilaterally castrated, or were rendered cryptorchid when they were either 15 or 45 days old. Subsequently, blood was sampled over the next several weeks and plasma luteinizing hormone (LH), follicle-stimulating hormone (FSH), testosterone (T), and immunoreactive inhibin-alpha (irI alpha) levels were measured by specific radioimmunoassays (RIAs). At the end of the experiment, gonadal expression of inhibin-alpha, inhibin-beta A, and inhibin-beta B subunits was measured by S1 nuclease analysis and in situ hybridization. In both age groups, bilateral castration (BC) produced the expected marked (p less than or equal to 0.01) increases in plasma LH and FSH levels, and concomitant decreases in T and irI alpha secretion within 1 - 2 days after surgery. In 15-day-old animals, unilateral castration (UC) significantly increased FSH and decreased circulating levels of irI alpha, but did not measurably alter LH or androgen production. At 7 days after surgery, the level of inhibin mRNA in the remaining testis was unchanged. In 45-day-old animals, UC caused a measurable increase in FSH, with little or no changes in the circulating levels of irI alpha. Plasma T levels were lowered (p less than or equal to 0.05) by UC; however, there were no statistical changes in LH levels in these UC rats. Finally, T administration markedly reversed UC-induced increase in FSH secretion in both age groups. Androgen therapy also interfered with inhibin release in 45-day-old, but not in 15-day-old rats. In rats 15 days old at the time of surgery, cryptorchidism produced a small but measurable increase (p less than or equal to 0.05) in LH release at Week 6 only, which was accompanied by a significant (p less than or equal to 0.01) decline in T secretion. Plasma FSH levels were elevated at all times in cryptorchid rats, and at 2, 4, and 6 wk, these levels were not statistically distinguishable (p greater than 0.05) from those of castrated animals. In this group of rats, cryptorchidism caused a transient increase (p less than or equal to 0.05) in irI alpha values 1 wk after surgery, but no changes at later times. Finally, measurement of testicular inhibin-alpha subunit messenger RNA (mRNA) levels showed an approximately 2-fold increase compared to total RNA levels in the testis. However, because of the significant decrease in total RNA levels per testis caused by cryptorchidism, the absolute change in inhibin-alpha subunit mRNA levels per testis corresponded to an approximately 3-fold decrease.(ABSTRACT TRUNCATED AT 400 WORDS)     

Modulation by sex steroids and [D-Trp6, Des-Gly-NH2(10)]luteinizing hormone (LH)-releasing hormone ethylamide of alpha-subunit and LH beta messenger ribonucleic acid levels in the rat anterior pituitary gland

LHRH and sex steroids play a major and direct regulatory role in the secretion of LH by the anterior pituitary gland. The aim of the present study was to investigate the interactions between sex steroids, more especially the potentiating effect of progesterone (P) in the presence or absence of a low dose of 17 beta-estradiol (E2) and/or dihydrotestosterone (D) on mRNA levels encoding the alpha- and beta-subunits of LH in both female and male rats. We also studied the effect of 2-week treatment with the LHRH agonist [D-Trp6, des-Gly-NH2(10)]LHRH ethylamide on the same parameters. After treatment with the LHRH agonist (5 micrograms daily), the accumulation of mRNA encoding the alpha-subunit was stimulated by approximately 3-fold while the LH beta mRNA concentration remained unchanged. Ovariectomy performed 14 days earlier, increased pituitary alpha and LH beta mRNA levels by 3.7- and 8.8-fold, respectively, while orchiectomy performed 14 days earlier increased alpha and LH beta mRNA levels by 6- and 6.5-fold, respectively. The present data demonstrate that although P alone exerts no effect on alpha and LH beta mRNA levels in castrated animals, treatment with P markedly potentiates the inhibitory effect of E2 on both mRNA levels in female as well as male rats. In addition, P potentiates the inhibitory effect of D on LH beta mRNA levels in castrated female rats. Furthermore, the present study illustrates the importance of the cumulative inhibitory effects of relatively low doses of E2 and D on mRNAs encoding both LH subunits. Moreover, the present observation of a differential modulation of alpha-subunit and LH beta mRNA levels after chronic treatment with an LHRH agonist offers an explanation for the high plasma levels of free alpha-subunit found in patients treated with LHRH agonists.     

Effects of starvation and refeeding on gonadotropin and thyrotropin subunit mRNAs in male Japanese quail

The contents of mRNAs encoding LH beta-, FSH beta-, TSH beta- and common alpha-subunit precursor molecules were measured in food-deprived and subsequently re-fed male Japanese quail. Pituitary LH beta, FSH beta and common alpha mRNA levels were decreased by starvation, and increased to the control levels by re-feeding. The rates of decreases of LH beta and common alpha mRNA levels were greater the corresponding rate for FSH beta levels. Pituitary TSH beta mRNA levels were not decreased by starvation, but increased transitorily by re-feeding. Plasma LH and triiodothyronine levels were decreased by starvation, and then increased to control levels by re-feeding, while plasma FSH and thyroxine levels did not show significant changes. Plasma LH and FSH levels showed positive correlations with pituitary common alpha and FSH beta mRNA levels, respectively, while plasma thyroxine levels showed a negative correlation with TSH beta mRNA levels. Hepatic weight was decreased slightly but significantly by starvation, and then showed a remarkable rebound after re-feeding was started. These results suggest that LH synthesis and secretion are more sensitive to starvation than FSH synthesis and secretion in Japanese quail, and that LH production recovered to initial levels within several days when birds were fully fed. Also, there is a possibility that the synthesis of TSH is accelerated transitorily by re-feeding. Furthermore, these results showed that there are different relationships between the plasma levels of LH, FSH, and TSH and the various hormone subunit mRNA levels. The remarkable change in hepatic weight leads us to assume that hepatic thyroid hormone metabolism is affected by starvation and re-feeding.     

Chronic administration of estradiol produces a triphasic effect on serum concentrations of gonadotropins and messenger ribonucleic acid for gonadotropin subunits, but not on pituitary content of gonadotropins, in ovariectomized ewes

To determine the acute and chronic effects of estradiol on synthesis and secretion of LH and FSH, ovariectomized ewes were administered estradiol via silastic capsules for 0 h, 12 h, 1 day, 2 days, 4 days, 8 days, 16 days, or 32 days (n = 5/group). Concentrations of GnRH in the median eminence began to decrease within 12 h and were lower (p less than 0.05) than in control ewes from 1 to 4 days after estradiol administration was begun. Serum concentrations of LH were decreased relative to pretreatment control levels from 1 to 10 h, elevated during a preovulatory-like surge from 11 to 22 h, and then decreased and remained below 1 ng/ml for the duration of the experiment. Serum concentrations of FSH followed a pattern similar to those for LH except that the magnitude of change was smaller. Treatment with estradiol initially (12 h) reduced (p less than 0.05) quantities of mRNA for alpha-, LH beta-, and FSH beta-subunits, after which the quantities of mRNA for the subunits returned to near or above control levels by Day 2. After 8 days of treatment the amounts of mRNAs for gonadotropin subunits were again less (p less than 0.05) than those of controls, and they remained suppressed through Day 32. Pituitary concentrations of LH and FSH decreased (p less than 0.05) during the first day of treatment and remained suppressed for the duration of the experiment. Thus, estradiol had a triphasic effect on secretion of gonadotropins and steady-state levels of mRNA for the gonadotropin subunits, but not on pituitary content of gonadotropins.(ABSTRACT TRUNCATED AT 250 WORDS)     

Ontogeny of follicle-stimulating hormone and luteinizing hormone gene expression during pubertal development in the female striped bass, Morone saxatilis (Teleostei)

Pubertal development in teleost fish is characterized by gonadal growth that is directly stimulated by the pituitary gonadotropins, FSH and LH. We used a quantitative ribonuclease protection assay to provide, for the first time, the developmental profiles of the alpha-, betaFSH-, and betaLH-subunit gene expression in a seasonal breeding fish, the female striped bass (3-yr study, n = 207). Two-year-old females were sexually immature, although a transient rise in all gonadotropin subunit mRNAs was measured in the pituitary. Pubertal ovarian development occurred in 65% of 3-yr-old females, characterized by the appearance of lipid droplets within the oocytes. This reproductive phase, termed pubertal development, was associated with a 34-fold increase in the mRNA levels of betaFSH and a rise in the pituitary concentration of LH. The first sexual maturation took place in 4-yr-old females and coincided with a 218-fold increase in the mRNA levels of betaFSH. During this time period, the mRNA levels of the alpha and betaLH subunits increased by 11- and 8-fold, respectively. At the final stages of vitellogenic growth, mRNA levels of betaFSH declined to basal levels, whereas the mRNA levels of the alpha and betaLH subunits remained elevated. Throughout the study, pituitary LH concentration was positively correlated to the mRNA levels of betaLH, but plasma levels of LH remained low and unchanged (0.4-0.8 ng/ml) despite increasing levels of pituitary LH concentration, suggesting a regulated secretion pathway. Taken together, the data show that the profiles of betaFSH and betaLH mRNAs appear to follow an annual rhythm that is associated with developmental events in the growing oocytes. In particular, increasing levels of betaFSH mRNA appear to underlie the first sexual maturity in the female striped bass.     

Influence of gonadotropin-releasing hormone pulse amplitude, frequency, and treatment duration on the regulation of luteinizing hormone (LH) subunit messenger ribonucleic acids and LH secretion

The effects of GnRH pulse amplitude, frequency, and treatment duration on pituitary alpha and LH beta subunit mRNA concentrations were examined in castrate-testosterone replaced male rats. Experimental groups received iv GnRH pulses (5, 25, or 125 ng) at 7.5-, 30-, or 120-min intervals for 8, 24, or 48 h. Saline pulses were given to control rats. Acute LH secretion was measured in blood drawn before and 20 min after the last GnRH pulse. In saline controls, alpha and LH beta mRNAs (150 +/- 14, 23 +/- 2 pg cDNA bound/100 micrograms pituitary DNA) fell to 129 +/- 14 and 18 +/- 2, respectively, after 48 h. In animals receiving GnRH pulses (7.5-min intervals), the 125-ng dose stimulated a slight increase (P less than 0.01) in alpha mRNA levels after 8 and 24 h and both LH subunit mRNAs were increased by the 25- and 125-ng doses after 48 h. The 30-min pulse interval injections (25- and 125-ng doses) increased LH beta mRNA levels after 8 h, but alpha mRNAs were not elevated until after 24 h. Maximum (3-fold) increases in alpha and LH beta mRNAs were seen in rats receiving 25-ng pulses every 30 min for 48 h. Using 120-min pulses, LH subunit mRNAs were not increased by any GnRH dose through 48 h. Acute LH release was not seen in rats receiving 5 ng GnRH pulses at any pulse interval.(ABSTRACT TRUNCATED AT 250 WORDS)     

Selective release of follicle-stimulating hormone and luteinizing hormone by 5 alpha-dihydroprogesterone and 3 alpha, 5 alpha-tetrahydroprogesterone in pregnant mare's serum gonadotropin-primed immature rats exposed to constant light

The effects of 5 alpha-dihydroprogesterone (5 alpha-DHP) and 3 alpha, 5 alpha-tetrahydroprogesterone (3 alpha, 5 alpha-THP) on follicle-stimulating hormone (FSH) and luteinizing hormone (LH) release were examined in the pregnant mare's serum gonadotropin (PMSG)-primed immature female rat (8 IU PMSG at 28 days of age) maintained in constant light. Control rats kept in 14L:10D conditions exhibited proestrous-like surges of LH and FSH release with peak levels attained at 1800 h on the second day after PMSG treatment. In rats exposed to constant light, the PMSG-induced surges of LH and FSH were not only delayed until 1000 h on the third day after PMSG, resulting in a delay in ovulation, but were also significantly attenuated when compared to the gonadotropin surges that occurred on Day 2 in rats kept under normal light-dark conditions. The administration of 5 alpha-DHP significantly enhanced the release of FSH at 1000 h on Day 3 when compared to constant light-exposed controls, but had no effect on LH. Treatment with 3 alpha, 5 alpha-THP selectively potentiated the release of LH at 1000 h on Day 3 and had an attenuating effect on FSH release on Days 2 and 3. These observations confirm earlier findings in the immature ovariectomized estrogen-primed rat and suggest that 5 alpha-DHP and 3 alpha, 5 alpha-THP may have significant roles in the regulation of FSH and LH secretion.     

Cloning and sequence analysis of FSH and LH in the giant panda (Ailuropoda melanoleuca)

The giant panda (Ailuropoda melanoleuca) is an endangered species and indigenous to China. It has been proposed that it has a highly specialized reproductive pattern with low fecundity, but little is known about its basic reproductive biology at the molecular level. In this report the genes encoding gonadotropin subunits alpha, follicle-stimulating hormone (FSH) beta and luteinizing hormone (LH) beta of the giant panda were amplified for the first time by RT-PCR from pituitary total RNA, and were cloned, sequenced and analyzed. The results revealed that the open reading region (ORF) of gonadotropin subunits alpha, FSH beta and LH beta are 363, 390 and 426 bp long, respectively. They displayed a reasonably high degree (74-94, 85-93, 75-91%, for alpha, FSH beta and LH beta subunits, respectively) of identity when deduced amino acids were compared with homologous sequences from partial available mammals including human, cattle, sheep, pig, rat, mouse. Three distinct differences were found at the site of 59 aa of the alpha subunit and 55 aa, 68 aa of FSH beta subunit. Our results provide an insight into understanding the mechanism of reproduction regulation and genetic characteristics of giant panda which will make an actual contribution to its conservation. In addition they lay a foundation for a further study towards producing recombinant panda FSH and LH which can be used in artificial breeding aimed to increase its captive reproductive efficiency.