Pituitary and plasma levels of luteinizing hormone and follicle-stimulating hormone in male and female rabbit fetuses

To study the ontogenesis of fetal pituitary gonadotrophin synthesis and release, LH and FSH were measured by radioimmunoassay in fetal rabbit pituitary glands and blood of both sexes from day 18 of gestation until birth. Results on levels of testicular and plasma testosterone were also included. Immunoreactive LH was first detected on day 19 in the pituitary gland and on day 20 in the plasma of fetuses of both sexes. Pituitary FSH was first measurable in both male and female fetuses at 24 days of gestation. Levels of FSH could not be detected in the blood of male fetuses at any time during gestation. In females, FSH could be measured in the circulation from day 27 of gestation until birth. These results show that (1) the ontogeny of pituitary LH and the onset of testosterone secretion are closely correlated and take place between 18 and 20 days of gestation just before the beginning of differentiation of the male genital tract, and (2) the highest concentrations of pituitary LH and FSH are observed, in both sexes, in late gestation after days 24-25.     

Gonadal and sex steroid feedback regulation of gonadotrophin mRNA levels and secretion in neonatal male and female rats

Serum and pituitary LH and FSH, and their pituitary mRNA levels, were measured in neonatal male and female rats after gonadectomy and after gonadectomy with sex steroid replacement. The animals were gonadectomized on day 3 of life, and those given sex steroid replacement were implanted with silicone elastomer capsules containing testosterone for males and diethylstilboestrol for females. Sham-operated rats served as controls. The animals were killed 4 or 8 days latter and the sera and pituitaries collected. Pituitary contents of mRNAs for the alpha subunit, FSH-beta and LH-beta were determined by blot hybridization using corresponding cDNAs. Distinct sex differences were found in the mRNA responses to gonadectomy and steroid replacement. In the males, gonadectomy increased all mRNA levels at 7 days of age. In the females, a rise on day 7 was detected only for FSH-beta; the other mRNAs were increased on day 11 of age. The steroid replacements reversed all the post-gonadectomy increases of mRNAs in both sexes. Moreover, the common alpha and LH-beta mRNAs of the male animals were consistently suppressed below control levels. The serum concentrations of gonadotrophins increased after gonadectomy on day 7 in the males but only on day 11 in the females. The steroid replacements also suppressed the post-gonadectomy increases in serum gonadotrophins, but only the serum concentration of FSH in the females was reduced below controls. Pituitary gonadotrophin concentrations were not affected by gonadectomy, but the steroids suppressed LH in the males and FSH in the females.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dopamine agonist- and antagonist-induced modulation of pituitary gonadotrophin releasing hormone receptors are independent of changes in serum prolactin

The effect of drug-induced hypo- and hyperprolactinaemia on pituitary gonadotrophin releasing hormone receptors (GnRH-R), serum and pituitary gonadotrophins (LH and FSH) and prolactin was investigated in intact adult male and female rats. Hypoprolactinaemia (serum prolactin less than 20% of control values) resulting from dopamine agonist (bromocriptine) infusion (4 mg/kg per day for 7 days) was accompanied by a 40-50% increase in GnRH-R in both male and female animals, though this was not accompanied by any major change in serum or pituitary LH and FSH. Hyperprolactinaemia (serum prolactin greater than ten times control values) induced by the dopamine receptor antagonist metoclopramide (65 mg/kg per day for 7 days) increased GnRH-R between 35 and 45% in both male and female rats without altering serum gonadotrophins. Domperidone (1 mg twice daily for 14 days) also increased GnRH-R by 50% but only in female rats. Both dopamine antagonists significantly increased pituitary prolactin content. Pituitary FSH increased in female rats treated with both metoclopramide and domperidone. The stimulatory effects of bromocriptine and metoclopramide on GnRH-R in male rats were prevented by concurrent treatment with a GnRH antiserum, suggesting that the drug effects were mediated through alteration in endogenous GnRH secretion. Induction of massive (serum prolactin greater than 2000 micrograms/l) hyperprolactinaemia in male and female rats with a transplantable prolactin-secreting pituitary tumour did not reduce GnRH-R concentration, although serum gonadotrophins were suppressed and pituitary gonadotrophin content was increased.(ABSTRACT TRUNCATED AT 250 WORDS)     

In vitro basal and GnRH-stimulated secretion of gonadotrophins reflects long-lasting modulatory effects, and peripheral levels are not predicted by pituitary responsiveness to GnRH

OBJECTIVE: Production of the appropriate pattern of gonadotrophin levels is crucial to proper functioning of the female reproductive system. We aimed to establish whether the pituitary has invariant secretory characteristics when isolated from in vivo controls. We aimed to obtain information during both the rising and declining phases of the gonadotrophin surge. DESIGN: This study investigated factors that are directed at the pituitary by isolating it from the acute influences of the in vivo environment and studying gonadotrophin secretion in vitro. METHODS: Pituitaries of adult female rats were collected at selected times during the day of pro-oestrus and incubated in vitro, and at the same time blood was collected. Peripheral levels of LH and FSH were measured over the whole day of pro-oestrus, basal in vitro secretions of LH and FSH from pituitaries were measured, GnRH-stimulated LH and FSH secretion were assessed, and the responsiveness of LH and FSH secretion to GnRH were calculated. RESULTS: Peripheral levels of LH peaked at 1800 h (P<0.02) followed by a subsequent decline. In contrast, although FSH had a peak at 1800 h (P<0.01), serum levels were also high at the end pro-oestrus. The profile of basal LH and FSH secretion from the pituitary in vitro, in the absence of added secretagogue, resembled that of the peripheral blood levels of each gonadotrophin. Pituitaries collected at 1800 h secreted most LH (P<0. 02). FSH secretion was low early on the day of pro-oestrus and then increased to and was maintained at high levels in the last quarter of the day (P<0.01).When the pituitaries were stimulated with GnRH the patterns of LH release and FSH release approximated those observed for basal release. Responsiveness of the pituitaries to GnRH was calculated by determining the ratio of GnRH-stimulated release to basal release. However, low levels of gonadotrophin were secreted even from pituitaries which were highly responsive as determined from consideration of percentage increase in secretion induced by GnRH. CONCLUSIONS: The secretory activity was dependent on the time of day the pituitaries were collected. Since the secretion occurred after the tissue had been removed from the direct influence of the in vivo environment, the variations in secretion must reflect long-lasting components of the mechanism that regulate gonadotrophin concentrations. There were changes in both LH and FSH responsiveness to GnRH stimulation over the day of pro-oestrus. Delineation of the time courses and changing predominance of multiple processes is needed to assist understanding the mechanisms underlying the female reproductive cycle.     

Sex and age differences in the spontaneous release of gonadotropins from immature rat pituitary glands in vitro

The present study was carried out to characterize the spontaneous release of LH and FSH from the pituitary gland of infant rats in an in vitro system. In addition, the responsiveness of their pituitary glands to synthetic LHRH in vitro was examined. Wistar-Imamichi male and female rats, aged between 1 and 21 days and androgenized female rats at 7 and 21 days of age were used. One-day-old female rats were androgenized by a subcutaneous injection of 1 mg of testosterone propionate. Animals were killed by decapitation, trunk blood was collected, and the pituitary gland was dissected free and weighed. Pituitaries were placed in 9 ml-test tube with 2 ml Krebs-Henseleit solution and incubation was carried out in a shaking incubator for 6 hours at 37 degrees C under the gassing of 5% CO2 and 95% O2. After preincubation for 15 min, medium was replaced with 2 ml fresh medium and LH and FSH concentrations released during the first 3 hr-incubation period were assessed as for the spontaneous release and the second 3 hr-incubation period assessed for the response to LHRH (10(-6)M) stimulation. In an experiment, time course changes of the spontaneous release of LH and FSH were studied using 7-day-old rat pituitaries. An aliquot of 0.5 ml of medium was taken at 30, 60, 120 and 180 min during the incubation. Gonadotropin contents in the pituitaries were determined by adding the residue in the pituitary gland and the amounts released into medium. Spontaneous release of LH and FSH increased with age in both male and female rats, and the released amounts of LH as well as FSH in female rats tended to be higher than those in males at 1, 3, 7 and 21 days of age examined. But significant sex differences in the spontaneous release of LH and FSH were only seen at 21-day-old; Spontaneous release of LH in female rats was 7 times higher than that in age-matched males. Serum LH and FSH concentrations in female rats were significantly higher than those in males at all ages examined, except the LH level at 1-day-old. In contrast to LH, age and sex differences in the magnitude of the spontaneous release of FSH from the pituitary paralleled with the age and sex differences in serum FSH concentrations.(ABSTRACT TRUNCATED AT 400 WORDS)     

Hypothalamic luteinizing hormone releasing factor and corticotrophin releasing activity in relation to pituitary and plasma hormone levels in male and female rats.

Hypothalamic corticotrophin releasing (CR) activity and LH-releasing factor (RF) content, and pituitary and plasma LH, FSH and ACTH were measured in adult male and female Wistar rats maintained under 14 h light per day. Hypothalamic LH-RF and pituitary and plasma hormones were estimated by radioimmunoassay while CR-activity was assessed by the amount of ACTH released from hemipituitaries in vitro. Two experiments were carried out on male animals. In the first, some of the animals were kept in a room, distant from the animal house, in which the lighting was reversed with respect to the external environment. In animals exposed to the reversed lighting regime, hypothalamic LH-RF content and pituitary gonadotrophin concentrations were significantly lower than the values in male rats kept in the animal house where they were in close proximity to female rats. In the second experiment, which was carried out on animals which had all been kept in the animal house, there was no significant differences between the LH-RF contents measured at 3-4 h intervals throughout the day. Pituitary LH and FSH contents, but not concentrations, were significantly increased at 12.00 h. There was little differences between the experiments in CR-activity, plasma ACTH concentrations and profiles of pituitary ACTH content and concentration. As expected there was a diurnal rhythm in plasma corticosterone concentrations (determined by competitive protein-binding assay) with the peak occurring between 15.00 and 18.00 h. The profiles of plasma and pituitary ACTH were similar to that of plasma corticosterone. Corticotrophin releasing activity dropped significantly between 12.00 and 16.00 h, but remained steady at the other times. In female rats there were no significant differences between hypothalamic LH-RF content throughout the 4-day cycle. During pro-oestrus the mean LH-RF content rose to teach a high level at 18.00 h at which time plasma LH concentration had risen sharply to a level consistent with the peak of the preovulatory surge. Plasma FSH concentration also rose significantly between 15.00 and 18.00 h of pro-oestrus. At metoestrus and dioestrus, plasma FSH levels were lower in the morning than in the evening. These results suggest that (1) there is no diurnal rhythm in hypothalamic LH-RF, (2) there may be a diurnal rhythm in pituitary gonadotrophin content in the male and in plasma FSH concentration on the days of metoestrus and dioestrus in the female, (3) if a surge of LH-RF does occur on the afternoon of pro-oestrus, the rate of LH-RF synthesis exceeds its release, and (4) the mechanism which regulates gonadotrophin secretion in the male may be affected by factors in the environment other than daylength. The results provide further evidence for the view that the diurnal rhythm of corticosterone secretion is under hypothalamo-hypophysial control.     

Hypothalamic-pituitary function in neonatally oestrogen-treated male rats.

Neonatal oestrogen administration to male rats permanently impaired the function of the pituitary-testicular axis possibly by inhibiting neonatal gonadotrophin secretion. To analyse the hypothalamus and/or pituitary involvement in this inhibition, pituitary responsiveness to acute stimulation with LH-releasing hormone (LHRH) was studied in vivo and in vitro in Wistar male rats injected on day 1 of age with oestradiol benzoate (OB) or olive oil. FSH and LH pituitary content and plasma concentrations were reduced in oestrogenized male rats at days 10 and 16 of age. Likewise, the in-vivo increase in gonadotrophin plasma concentrations after acute stimulation with LHRH was almost completely suppressed in 10- and 16-day-old oestrogenized males. In vitro, the increased secretion of FSH after LHRH stimulation was abolished and the LH response strongly reduced in pituitaries from oestrogenized males. Finally, the effects of neonatal oestrogenization were not abolished by treatment from day 1 to day 15 with an LHRH agonist (0.01 microgram/kg per 12 h). We conclude that in male rats the effects of oestrogenization are due to both a reduction in LHRH endogenous secretion and a decrease in the pituitary responsiveness to LHRH.     

Daily administration of gonadotrophin-releasing hormone increases pituitary gonadotroph number and pituitary gonadotrophin content, but not serum gonadotrophin levels, in female rats on day 1 of dioestrus.

Gonadotrophin-releasing hormone (GnRH) has been shown to regulate the synthesis and release of gonadotrophins acutely, yet few studies have investigated the chronic effects of this agent on pituitary gonadotrophins. In the present study we determined the effect of chronic administration of GnRH on the female rat pituitary gland. Rats of 8 weeks of age were injected s.c. with various doses of GnRH daily for 30 days. After completion of the GnRH treatment, treated rats and age-matched controls were killed by decapitation at 09.00 h on the first day of dioestrus, as determined from vaginal smears. Treatment with 10 ng-10 micrograms GnRH/day increased pituitary contents of FSH and LH in a dose-dependent manner. The change in FSH content was much greater than that of LH content. The pituitary FSH content of rats treated with 40 micrograms GnRH was significantly less than that of rats treated with 10 micrograms GnRH. There was a marked increase in the number of cells which stained positively for FSH (266%) and LH (28%) in the anterior pituitary of rats given 10 micrograms GnRH, but there was no demonstrable change in the areas of single cells stained positively for FSH and LH. Serum levels of LH, FSH and oestradiol were not affected by the GnRH treatment. These data indicate that chronic administration of GnRH is capable of increasing the pituitary gonadotrophin content and numbers of FSH and/or LH-stained cells and that FSH cells are affected more than LH cells by the GnRH treatment. The increase in pituitary gonadotrophin content, however, does not necessarily produce an increase in circulating levels of gonadotrophins.     

Ontogeny of serum and pituitary gonadotrophins in male rats treated with low doses of 5 alpha-dihydrotestosterone

The effect of s.c. daily injections of 10 or 1000 ng 5 alpha-dihydrotestosterone (DHT)100 g body weight from birth to day 21, or from days 26 to 117 of age, on the changes in concentration of serum and pituitary gonadotrophins was investigated in male rats. Treatment with 10 ng DHT from days 1 to 21 depressed serum FSH, but not LH, at day 7, while 1000 ng DHT depressed both serum LH and FSH. Treatment with both doses of DHT reduced pituitary levels of LH and FSH at day 7, with FSH being more depressed than LH. Treatment with 10 ng DHT from days 26 to 117 increased serum FSH from days 82 to 117, while 1000 ng DHT did not have this effect. Treatment with 1000 ng, but not 10 ng, DHT between days 26 and 117 reduced pituitary levels of LH and FSH at day 40. Rats treated with the two doses of DHT from days 26 to 117 showed a difference in the responsiveness of the pituitary to LH-releasing hormone (LHRH). Treatment with 10 ng DHT enhanced LHRH-induced release of LH without affecting FSH release, while 1000 ng DHT depressed LHRH-induced release of FSH but not of LH. These findings support the view that DHT may play a modulatory role in the ontogeny of serum gonadotrophins and the responsiveness of the pituitary to LHRH during the onset of puberty in the male rat.     

Pituitary gonadotropin-releasing hormone receptor regulation in mice. II: Females

The regulation of pituitary GnRH receptors (GnRH-R) by gonadal steroids was examined in female mice housed in a constant environment (six to 8 per cage in same room as males). A 60% decrease in GnRH-R occurred 7 days after ovariectomy (OVX) (9.2 +/- 0.9 fmol/pituitary OVX vs. 25 +/- 2 for intact random estrous cycle controls). The receptor affinity (Ka 1.86 X 10(9) M-1) remained constant in intact and OVX female mouse pituitary particles. The pattern of GnRH-R fall after OVX was similar to that found in male mice, except that the GnRH-R decrease began some 6 h later than in males and serum LH also rose more slowly. Serum FSH was significantly elevated 6 h post OVX. In contrast to males, pituitary LH, in spite of a rapid fall (60%) at 12 h, regained the random, estrous cycle control value by 4 days post OVX and then increased to above this level. Pituitary FSH, unlike in males, remained at the intact value (3.1 +/- 0.24 micrograms/pituitary) up to 24 h post OVX and then gradually rose to 7.9 +/- 0.37 micrograms/pituitary on day 4 and 15.5 +/- 0.32 micrograms/pituitary on day 7. Treatment of OVX female mice with estradiol-17 beta (300 ng/day) attenuated the postcastration GnRH-R fall, and was more effective when combined with progesterone (375 micrograms/day). Progesterone alone was ineffective. The GnRH-R fall post OVX persisted for up to 2 months, despite elevated serum and pituitary LH and FSH levels. GnRH-R fell by 40% in lactating mice (20.6 +/- 0.95-lactating vs. 32.4 +/- 1.25 fmol/pituitary-random, estrous cycling females). Serum LH was reduced by 70% in lactating mice. These findings are qualitatively and quantitatively similar to those in lactating rats suggesting that, in this physiological situation, a similar mechanism may account for the receptor fall in both species. In sex reversed (Sxr) mice (genotypic female-phenotypic male) GnRH-R values were about 50% higher than those of intact normal male and normal, random estrous cycling, female values. This was the only situation in mice in which pituitary GnRH-R increases were observed to date. Serum and pituitary LH and FSH values in Sxr mice were elevated, especially when compared with normal, random estrous cycling female controls. The results indicate that pituitary GnRH-R of female mice fall in response to removal of gonadal steroid feedback, in the same way as males.(ABSTRACT TRUNCATED AT 400 WORDS)     

The impact of ovarian laser surgery on the gonadotrophin secretion in women with polycystic ovarian disease

To evaluate the effects of ovarian surgery on the deranged episodic gonadotrophin release of women with the polycystic ovarian disease (PCOD), we studied 11 patients with the clinical and endocrinological features of PCOD before and after laparoscopic laser coagulations of ovarian surfaces and cysts. During both occasions, blood was collected at 15-min intervals for 8 h to determine LH and FSH secretory profiles and additionally for 3 h during GnRH injections (25 micrograms twice within 2 h) to assess pituitary responsiveness. Serum testosterone, androstendione and oestrogen (oestrone, oestradiol) levels were markedly reduced (P less than 0.05 or less) after surgery. Mean LH concentrations declined (P less than 0.001), while FSH levels increased (P less than 0.01) following laser treatments. The LH pulse frequencies (by Cluster analysis) did not change after ovarian surgery, but the LH pulse amplitudes were markedly reduced (P less than 0.01). Lower (P less than 0.05 or less) LH concentrations were attained in response to GnRH challenges, and the stimulated FSH release also tended to decrease after laser treatments. Thus, ovarian surgery in PCOD women resulted in reduced serum sex steroid concentrations and in divergent effects on serum LH and FSH levels. The attenuated pituitary LH responsiveness after ovarian surgery suggests action of sex steroids primarily at the pituitary site, while the increase in FSH concentrations may be attributed to other factors selectively modulating FSH release.     

Effects of bovine 35 kDa FSH-suppressing protein on FSH and LH in rat pituitary cells in vitro: comparison with bovine 31 kDa inhibin

The mode of action of a recently isolated gonadal protein, termed FSH-suppressing protein (FSP) or follistatin, on basal and gonadotrophin-releasing hormone (GnRH)-stimulated release of FSH and LH and on pituitary cell content of FSH and LH was examined in rat pituitary cell cultures and compared with the previously reported effects of inhibin. Pituitary cells were cultured for 3-9 days in the presence of graded doses of FSP and the basal release rates and changes in cell contents of FSH and LH determined during this period. FSP suppressed both the basal release rate and the cell content of FSH with median inhibitory concentrations (IC50) of 135 and 161 pmol/l respectively. The corresponding effects of FSP on LH basal release rate and LH cell content (IC50 = 200 pmol/l) were limited compared with the effects on FSH. The effect of FSP on GnRH-stimulated release of FSH and LH during 4 h was determined in cells which had been preincubated with FSP for 3 days, and the GnRH-stimulated release of FSH and LH analysed as a percentage of the respective gonadotrophin available for release. FSP antagonized GnRH action with dose-related increases in the GnRH median effective (stimulatory) concentrations for FSH and LH release (EC50 values = 56 and 400 pmol/l respectively) and a suppression in the maximum release of FSH and LH by excess GnRH (IC50 values = 142 and 150 pmol/l respectively). The effect of FSP on FSH cell content after 3 days in culture was insensitive to the neutralizing effects of an inhibin antiserum.(ABSTRACT TRUNCATED AT 250 WORDS)     

The ontogeny of hypothalamic pituitary function in the pig. I. Pituitary LH and FSH in the fetus and neonate

Pituitaries were collected from fetal and postnatal pigs from day 55 p.c. until 6 weeks after birth at closely spaced intervals. LH and FSH in individual pituitaries were quantified by both homologous RIA and homologous RRA. No significant difference was found between results obtained by RIA and RRA. Both LH and FSH are first detected by RIA and RRA in the porcine fetal pituitary at 75 days p.c. Thereafter both LH and FSH pituitary content rises until term. LH pituitary concentration in both male and female fetuses and FSH pituitary concentration in males exhibit a peak just before birth. FSH pituitary concentration in females rises until birth and thereafter remains elevated. A statistically significant sex difference was found postnatally with regard to FSH content and concentration but not for LH.     

Modulation of rat pituitary gonadotrophin secretion by porcine granulosa cell 'inhibin', LH releasing hormone and sex steroids in rat anterior pituitary cells in culture

The incubation of female rat adenohypophysial cells in primary culture with porcine granulosa cell culture medium (GCM) led to the complete inhibition of responses of LH and FSH to LH releasing hormone (LHRH) as well as to the inhibition of spontaneous release of FSH. These effects of GCM suggest the specificity of the 'inhibin'-like activity of this material. Granulosa cell culture medium completely reversed the stimulatory effect of oestradiol-17 beta on the responses of LH and FSH to LHRH, as well as reversing the stimulatory effect of progesterone, oestradiol or a combination of both steroids on the spontaneous release of FSH, while not affecting the spontaneous release of LH. The antioestrogenic effects of progesterone observed on the response of LH to 0.3 nM-LHRH were amplified in the presence of GCM while the stimulatory effects of progesterone, oestradiol or both on the response of FSH to 0.3 nM-LHRH were completely reversed by the medium. Moreover, the presence of GCM led to an additive inhibitory effect with dihydrotestosterone on the response of LH to LHRH while it completely reversed the stimulatory effect of the androgen on spontaneous and LHRH-induced FSH release. The present data show that the presence of porcine granulosa cell 'inhibin' activity can exert marked interactions with sex steroids in the control of gonadotrophin secretion. This 'inhibin' activity reversed all the stimulatory effects and potentiated all the inhibitory effects of sex steroids on gonadotrophin secretion. Although the physiological role of 'inhibin' remains to be defined well, the importance of this activity is clearly demonstrated in anterior pituitary cells in culture.     

Releasing factor and hormonal changes in the hypothalamic-pituitary-gonadotrophin and -adrenocorticotrophin systems before and after birth and puberty in male, female and androgenized female rats.

The hypothalamic content of LH releasing factor (RF), pituitary ACTH and pituitary and plasma LH and FSH were measured by radioimmunoassay from foetal Day 15 to postnatal Day 65. Bioassayable corticotrophin releasing activity was also measured during the postnatal period. Hypothalamic LH-RF was detectable as early as foetal Day 15, increasing gradually until postnatal Day 2 and then steeply between Days 5 and 16. The levels of LH-RF were similar in both male and normal female rats until Day 41, after which the increase which had been occurring from Day 16 continued in the male but not the female. In female rats treated with testosterone propionate neonatally ('androgenized females') the hypothalamic content of LH-RF at Day 9 was significantly less than that in the male or normal female, levels reaching those found in the latter two groups by Days 16-22. The lower level of LH-RF in the androgenized female was associated with pituitary gonadotrophin and plasma FSH levels which were lower than in the normal female until Day 30. In the normal female, vaginal opening was associated with a marked drop in hypothalamic LH-RF content and in pituitary LH and FSH, but in the androgenized female, vaginal opening occurred while hypothalamic LH-RF and pituitary LH levels were still rising. The peaks in pituitary FSH and LH and in plasma LH seen on Days 22, 30 and 41, respectively, in the normal female were each delayed by 8-9 days in the androgenized female. In all three types of animal there was a significant drop in plasma FSH between Days 50 and 65 which was associated with a significant increase in pituitary FSH in the male and a significant decrease in pituitary FSH in the androgenized female rats. The day 17 foetal pituitary gland also contained ACTH, and again levels of this hormone rose steeply between Days 5 and 9. In contrast to the gonadotrophins, there was a marked divergence between the pituitary content and concentration of ACTH: content rose while concentration remained relatively steady after Day 9. There was no major difference in pituitary ACTH levels between the three types of animal throughout the study; however, around Days 16 and 50, corticotrophin releasing activity was higher in males and androgenized females compared with that in normal females.     

Effect of pulsatile gonadotropin-releasing hormone on the release of luteinizing hormone and follicle-stimulating hormone in vitro by anterior pituitaries from lactating and cycling rats

The ability of pituitaries from lactating animals to secrete LH and FSH in response to gonadotropin-releasing hormone (GnRH) was studied in vitro using a pituitary incubation system. Hemipituitaries were exposed to GnRH for 6 min during each hour of incubation. LH release by anterior pituitaries (APs) from day 5 postpartum rats nursing eight pups, in response to pulsatile exposure to GnRH, was significantly less than that released by APs from diestrous cycling females. Even though the amount of LH released by APs increased as lactation progressed, LH release by APs from day 15 postpartum rats nursing eight pups was still less than LH release by APs from diestrous females. In contrast pituitaries from lactating females nursing two pups released amounts of LH similar to that released by pituitaries from diestrous females, whereas females deprived of their litters for 48 h showed a greater response than diestrous females. Generally, there was a good quantitative relationship between the amount of LH released in vitro and plasma LH concentrations for all the intact groups studied. The ability of lactation to suppress the postcastration rise in serum LH also was demonstrated in vitro as pituitaries from ovariectomized or intact females nursing eight pups released similar amounts of LH on days 5 and 10 postpartum. However, by day 15 postpartum, even though serum LH concentrations were still very low, pituitaries from ovariectomized lactating females released LH in vitro at a rate similar to pituitaries from nonlactating rats. Serum FSH concentrations were not suppressed but similar in intact and cycling females. Also, the total amount of FSH released in vitro in response to GnRH by pituitaries from lactating and cycling females did not differ significantly, even though LH release differed greatly among these groups of animals. However, the patterns of GnRH-stimulating FSH secretion differed among intact lactating, ovariectomized lactating, and nonlactating females. Pituitary LH concentrations were similar on day 5 postpartum and diestrus and on day 15 postpartum and proestrus. Pituitary FSH concentrations on day 5 postpartum were similar to those during diestrus and proestrus and had increased 2-3 times by day 15 postpartum. Generally, there was no correlation between the amount of LH or FSH released by pituitaries in response to GnRH and pituitary gonadotropin content. In summary, the inability of pituitaries from lactating rats to respond adequately to large doses of GnRH in vitro suggests that the suckling stimulus indirectly suppresses pituitary responsiveness to GnRH. This suppression differentially affects basal LH secretion, but not basal FSH secretion, and may be the direct result of inadequate GnRH stimulation in vivo.     

Gonadotrophin-releasing activity of neurohypophysial hormones: I. Potential for modulation of pituitary hormone secretion in rats

Neurohypophysial hormones have been implicated in the control of anterior pituitary function, and oxytocin has been shown to stimulate gonadotrophin excretion and ovarian follicular development in certain species. To determine the role of neurohypophysial peptides in the control of gonadotrophin release, their actions on LH and FSH secretion were analysed in rats in vivo and in vitro. In adult female rats, administration of oxytocin during early pro-oestrus advanced the spontaneous LH surge and markedly increased peripheral LH levels at 15.00 h compared with control animals. In cultured pituitary cells from adult female rats, oxytocin and vasopressin elicited dose-related increases in LH and FSH release. Such responses were not affected by a potent gonadotrophin-releasing hormone (GnRH) antagonist that abolished GnRH agonist-induced release of LH and FSH. Oxytocin did not enhance GnRH agonist-stimulated gonadotrophin release to the same extent as it increased basal secretion, but at low concentrations of GnRH agonist the effects were additive. The gonadotrophin responses to oxytocin and vasopressin were inhibited by the specific neurohypophysial hormone antagonists, [d(CH2)5D-Ile2,Ile4,Arg8]vasopressin and [d(CH2)5Tyr (Me),Arg8]vasopressin. These results provide direct evidence that neurohypophysial hormones can stimulate gonadotrophin secretion through a receptor system distinct from the GnRH receptor. Such a mechanism could represent a complementary hypothalamic control system for long-term modulation of LH and FSH secretion by exerting a basal or tonic influence on gonadotrophin production.     

Mechanism of the first spontaneous gonadotrophin surge and that induced by pregnant mare serum and effects of neonatal androgen in rats.

The mechanism of the first (pubertal) preovulatory gonadotrophin surge was investigated in Wistar rats by measuring (1) LH releasing factor (LH-RF) in pituitary stalk plasma and in extracts of hypothalamic and preoptic tissue, (2) LH and FSH in peripheral plasma and extracts of anterior pituitary tissue and (3) the LH and FSH response to LH-RF. The mechanism of the first surge appeared broadly to resemble that in the adult. That is, the gonadotrophin surge which occurs on the afternoon of the day before vaginal opening was found to coincide with a surge of immunoreactive LH-RF in pituitary stalk plasma and a significant increase in pituitary responsiveness to LH-RF. The magnitude of the change in pituitary responsiveness was, however, less at puberty than in the adult while the peak of the LH-RF surge was higher at puberty. The surges of LH-RF in stalk plasma and gonadotrophin in peripheral plasma corresponded relatively precisely with a fall in the preoptic and hypothalamic content of LH-RF and in the pituitary content of LH and FSH, suggesting that, in contrast with the adult, the synthesis of LH-RF and gonadotrophin at puberty cannot keep up with their release. A significant increase in stalk plasma LH-RF concentration occurring concomitantly with a surge of LH could be induced on the afternoon of Day 32 of life by administering pregnant mare serum (PMS) ON Day 30 (about 10 days before vaginal opening). This, together with other evidence, suggests that the timing of the first gonadotrophin surge depends upon the capacity of the ovary to secrete oestradiol in the form of a surge. The fact that no significant increase in the pituitary responsiveness to LH-RF occurred in PMS-treated rats could account for the fact that the height of the gonadotrophin surge in these animals is only a third that of the spontaneous surge. No significant change in peripheral plasma LH or stalk plasma LH-RF concentrations was found around the time of vaginal opening or cornification in female rats given 1.25 mg testosterone propionate on Day 4 (androgenized female rats). Studies on the effect of ovariectomy and/or adrenalectomy suggested that the ovary and adrenal are involved in the timing of vaginal opening in normal but not in androgenized female animals.     

Relationship between gonadotrophin subunit gene expression, gonadotrophin-releasing hormone receptor content and pituitary and plasma gonadotrophin concentrations during the rebound release of FSH after treatment of ewes with bovine follicular fluid during the luteal phase of the cycle.

The modulation of FSH secretion at the beginning and middle of the follicular phase of the cycle represents the key event in the growth and selection of the preovulatory follicle. However, the mechanisms that operate within the pituitary gland to control the increased release of FSH and its subsequent inhibition in vivo remain unclear. Treatment of ewes with bovine follicular fluid (bFF) during the luteal phase has been previously shown to suppress the plasma concentrations of FSH and, following cessation of treatment on day 11, a rebound release of FSH occurs on days 12 and 13. When luteal regression is induced on day 12, this hypersecretion of FSH results in an increase in follicle growth and ovulation rate. To investigate the mechanisms involved in the control of FSH secretion, ewes were treated with twice daily s.c. injections of 5 ml bFF on days 3-11 of the oestrous cycle and luteal regression was induced on day 12 with prostaglandin (PG). The treated ewes and their controls were then killed on day 11 (luteal), or 16 or 32 h after PG and their pituitaries removed and halved. One half was analysed for gonadotrophin and gonadotrophin-releasing hormone (GnRH) receptor content. Total pituitary RNA was extracted from the other half and subjected to Northern analysis using probes for FSH-beta, LH-beta and common alpha subunit. Frequent blood samples were taken and assayed for gonadotrophins. FSH secretion was significantly (P less than 0.01) reduced during bFF treatment throughout the luteal phase and then significantly (P less than 0.01) increased after cessation of treatment, with maximum secretion being reached 18-22h after PG, and then declining towards control values by 32h after PG. A similar pattern of LH secretion was seen after bFF treatment. Pituitary FSH content was significantly (P less than 0.05) reduced by bFF treatment at all stages of the cycle. No difference in the pituitary LH content was seen. The increase in GnRH receptor content after PG in the controls was delayed in the treated animals. Analysis of pituitary mRNA levels revealed that bFF treatment significantly (P less than 0.01) reduced FSH-beta mRNA levels in the luteal phase. Increased levels of FSH-beta, LH-beta and alpha subunit mRNA were seen 16h after PG in the bFF-treated animals, at the time when FSH and LH secretion from the pituitary was near maximum.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effect of porcine follicular fluid preparations on gonadotrophin secretion by the mouse pituitary gland in vitro

The acute effects of pooled porcine follicular fluid (PFF), before and after various methods of processing to eliminate steroids, were studied on the luteinizing hormone releasing hormone (LH-RH)-induced release of FSH and LH by whole pituitary glands, from 34-day-old mice, incubated in vitro for 3-4 h. Charcoal treatment of PFF eliminated the steroids and reduced the inhibitory potency on gonadotrophin secretion. On the other hand, dialysis or ultrafiltration (mol. wt greater than 10,000) did not reduce the inhibitory activity on gonadotrophin secretion. Of the three steroids tested, only oestradiol at a concentration of 10(-10) mol/l inhibited FSH and LH secretion in vitro. This inhibitory effect was counteracted by the inclusion of the oestrogen antagonist tamoxifen in the incubation medium. The presence of tamoxifen did not decrease the suppression of FSH and LH induced by PFF, suggesting that the inhibition observed under the conditions of incubation was not due to oestrogen. Preincubation of mouse pituitary tissue for 1 h with PFF reduced the subsequent release of bioactive FSH and LH induced by LH-RH. The inhibitory effect of PFF was rapid and sustained. The continuous presence of PFF throughout the incubation period was not necessary for manifestations of the inhibitory effects on gonadotrophin release. The suppression of gonadotrophin secretion was related to the dose of PFF with the curve showing a biphasic pattern. The degree of FSH suppression was uniformly greater than that of LH, showing the preferential nature of the inhibitory effect of PFF. At high doses of PFF, the degree of FSH suppression was decreased significantly. This effect on LH release was less pronounced. The inhibition caused by PFF in the in-vitro incubation procedure was not due to destruction of LH-RH or the released gonadotrophins.