ABSTRACT

Plasma and ovarian concentrations of inhibin were determined at 3-h intervals throughout the 4-day oestrous cycle of rats by a radioimmunoassay (RIA) based on a bovine RIA. Plasma concentrations of LH, FSH, progesterone, testosterone, oestradiol-17β, and pituitary contents of FSH and LH were also determined during the cycle. Plasma levels of inhibin showed a marked increase on the morning of oestrus and the afternoon of metoestrus, and a further increase was noted on the morning of pro-oestrus. These increases in plasma levels of inhibin were probably due to the following three events in the ovary, follicular recruitment on the morning of oestrus, selection of follicles on the day of metoestrus, and final maturation of follicles for ovulation on the morning of pro-oestrus with an increase in oestradiol-17β secretion. A striking decrease in inhibin secretion occurred during the process of ovulation after the preovulatory gonadotrophin surge on the afternoon of pro-oestrus. Basal levels of plasma FSH gradually decreased from metoestrus to pro-oestrus as plasma levels of inhibin increased. There was a significant inverse relationship between plasma levels of FSH and inhibin throughout the oestrous cycle (r = −0·51). The present findings suggest that changes in the plasma levels of inhibin during the oestrous cycle provide a precise indicator for follicular recruitment, selection and ovulation, and that changes in concentrations of oestradiol-17β in the plasma are associated with follicular maturation.

Journal of Endocrinology (1990) 126, 151–157

ABSTRACT

The present study was undertaken to determine whether hypothalamic differentiation is involved in the selective release of FSH during the periovulatory period using adult male rats castrated and implanted with an ovary.

Adult male rats (70–90 days old) were castrated and an ovary obtained from a prepubertal female rat (26 days old) was immediately grafted subcutaneously. Four weeks later, human chorionic gonadotrophin (hCG, 10 i.u.) was injected i.v. into the experimentally manipulated rats to induce ovulatory changes in the grafted ovaries. Another group of similarly prepared rats was injected with 0·9% (w/v) NaCl solution as controls. After injection of hCG, plasma concentrations of FSH increased significantly by 6 h, reached peak values at 12 h and declined to control levels at 36 h. On the other hand, plasma concentrations of LH were reduced by 6 h and decreased further during the next 36 h. An abrupt fall in plasma concentrations of oestradiol-17β occurred within 3 h of the administration of hCG. Histological examination revealed that ovulatory changes and luteinization of follicles were induced in grafted ovaries by 18 h after the injection of hCG. Thirty-six hours after treatment with hCG, a set of newly formed corpora lutea was observed in grafted ovaries and plasma concentrations of progesterone were raised. Treatment with oestradiol-17β did not inhibit the selective release of FSH after the administration of hCG, suggesting that the abrupt decrease in secretion of oestradiol-17β from the grafted ovary is not involved in the occurrence of the FSH surge.

These results indicate that a selective release of FSH can be induced in castrated male rats bearing an ovarian transplant probably due to decreased secretion of inhibin by the luteinized follicles in the grafted ovaries. Sex differentiation of the hypothalamus is not, therefore, involved in the selective surge of FSH.

J. Endocr. (1985) 106, 31–36

The relationship between hypothyroidism and disturbance of the hypothalamo-hypophysial-adrenal axis was investigated using adult male rats. Hypothyroidism was produced by administration of 4-methyl-2-thiouracil (thiouracil) in the drinking water for 2 weeks. Hypothyroidism decreased adrenal weights to 57% of controls and plasma concentrations of corticosterone to 48% of controls. The changes in the weight of adrenals recovered to control levels by administration of thyroxine. The pituitary responsiveness to corticotrophin-releasing hormone (CRH) and arginine vasopressin (AVP) for ACTH release markedly increased in hypothyroid rats as compared with euthyroid rats. In vivo release of CRH and AVP in median eminence significantly increased in hypothyroid rats as compared with euthyroid rats. There were no significant differences in hypothalamic concentrations of CRH and AVP. These results indicate that hypothyroidism causes adrenal dysfunction directly and results in hypersecretion of ACTH mediated by increases in synthesis of CRH and AVP in the hypothalamus.

Abstract

Plasma and ovarian levels of inhibin were determined by a radioimmunoassay (RIA) at 3-h intervals throughout the 4-day oestrous cycle of hamsters. Plasma concentrations of FSH, LH, progesterone, testosterone and oestradiol-17β were also determined by RIAs. In addition, hamsters were injected at various times with human chorionic gonadotrophin (hCG) to determine the follicular development. The changes in plasma concentrations of FSH after injection of antisera to oestradiol-17β (oestradiol-AS) and inhibin (inhibin-AS) on the morning of day 2 (day 1=day of ovulation) were also determined.

Plasma concentrations of inhibin showed a marked increase on the afternoon of day 1, remained at plateau levels until the morning of day 4, then increased abruptly on the afternoon of day 4 when preovulatory LH and FSH surges were initiated. A marked decrease in plasma concentrations of inhibin occurred during the process of ovulation after the preovulatory gonadotrophin surges. An inverse relationship between plasma levels of FSH and inhibin was observed when the secondary surge of FSH was in progress during the periovulatory period. Plasma concentrations of oestradiol-17β showed three increase phases and these changes differed from those of inhibin. Changes in plasma concentrations of oestradiol-17β correlated well with the maturation and regression of large antral follicles. Follicles capable of ovulating following hCG administration were first noted at 2300 h on day 1. The number of follicles capable of ovulating reached a maximum on the morning of day 3 (24·8± 0·6), and decreased by 0500 h on day 4 (15·0 ± 1·1), corresponding to the number of normal spontaneous ovulations. Plasma concentrations of FSH were dramatically increased within 6 h after inhibin-AS, though no increase in FSH levels was observed after oestradiol-AS. These findings suggest that changes in the plasma levels of inhibin during the oestrous cycle provide a precise indicator of follicular recruitment, and that the changes in plasma concentrations of oestradiol-17β are associated with follicular maturation. These findings also suggest that inhibin may play a major role in the inhibition of FSH secretion during the oestrous cycle of the hamster.

Journal of Endocrinology (1995) 146, 169–176

ABSTRACT

The role of basal FSH secretion during the rat oestrous cycle in regulating ovulation was examined by suppressing FSH secretion using charcoal-treated porcine follicular fluid (pFF). Although 0·5 ml pFF given at 05.00 and 11.00 h on the day of pro-oestrus had no effect on ovulation, 0·5 ml pFF given five times at 6-h intervals from 11.00 h on dioestrus to 11.00 h on pro-oestrus completely eliminated ovulation on the morning of the next oestrus. When 0·25 ml pFF was given on the same schedule, all animals ovulated a significantly decreased number of oocytes (9·0 ± 0·8) at the next oestrus. During the period of pFF treatment, the number of follicles capable of ovulating in response to human chorionic gonadotrophin (hCG) decreased (7·6 ± 0·7 at 05.00 h on pro-oestrus) and plasma levels of oestradiol showed a peak level 6 h later than in controls treated with 0·5 ml steroid-free porcine serum. Treatment with pFF suppressed plasma FSH concentrations in a dose-dependent manner, although plasma LH was inhibited irregularly. Supplementary administration of LH throughout the treatment period with 0·5 ml pFF resulted in ovulation of one to three oocytes in response to hCG in only three out of ten animals. These results suggest that basal secretion of FSH during the rat oestrous cycle plays an essential role in follicular development and maturation towards ovulation, and that the levels of FSH secretion may contribute to the maturation of normal numbers of follicles for ovulation.

Journal of Endocrinology (1993) 139, 287–293

ABSTRACT

The relationship between a surge of FSH and the initiation of follicular maturation was examined using rats with a 4-day oestrous cycle. When antiserum against LH-releasing hormone (LHRH-AS) was given at 13.00 h on the day of pro-oestrus (day 0), surges of FSH and LH were blocked. Plasma FSH and LH were maintained at low basal levels without a surge release until the next spontaneous surge occurred on the afternoon of day 4, the predicted day of pro-oestrus. Follicular responsiveness to an injection of human chorionic gonadotrophin (hCG) indicated that preovulatory follicles, present at the time of treatment with LHRH-AS, were capable of ovulating on day 1 but had regressed by day 2. Subsequently, as shown by the ovulatory response to hCG, a new set of follicles had begun to mature by the morning of day 3 without a preceding surge of FSH. Changes in oestradiol-17β levels in the plasma throughout the oestrous cycle were the same in rats injected with LHRH-AS or non-immune control serum. The mature follicles in the rats treated with LHRH-AS thus retained the capacity to ovulate after losing their ability to secrete oestrogen.

These results suggest that an FSH surge is not essential for initiation of follicular maturation and that basal levels of FSH may be enough to initiate follicular maturation in the absence of newly formed corpora lutea.

J. Endocr. (1986) 110, 279–285

ABSTRACT

Concentrations of inhibin in peripheral blood were determined by radioimmunoassay in pseudopregnant, pregnant and lactating rats and correlated with the status of follicular development, luteal function in the ovaries and placental function. In pseudopregnant rats, plasma concentrations of inhibin remained relatively high from days 2 to 12 of pseudopregnancy and no significant changes occurred throughout pseudopregnancy. In pregnant rats, a marked decline in plasma concentrations of inhibin occurred between days 6 and 14 followed by an abrupt increase on days 20 and 21. In lactating rats, plasma concentrations of inhibin were consistently lower in dams nursing eight pups than in dams nursing two pups. Plasma concentrations of inhibin remained low during the early stages of lactation followed by a significant increase during the late stages in dams nursing eight pups. These changes in inhibin during the period of lactation correlated with those of plasma concentrations of LH, but not with those of oestradiol-17β, testosterone and progesterone. Throughout pseudopregnancy, pregnancy and lactation, on the other hand, plasma concentrations of FSH were within the range of dioestrous levels.

These findings indicate that concentrations of inhibin in peripheral plasma vary with the number of healthy antral follicles and no positive correlation was observed with luteal function during the luteal phase in rats. The results also suggest that inhibin is a more direct index of follicular development than oestradiol-17β in rats.

Journal of Endocrinology (1989) 121, 545–552

The high- and low-avoidance animal (HAA and LAA respectively) strains of Hatano rats were originally selected and bred from Sprague-Dawley rats for their performance in the shuttle-box task. The present study focused on the activity of the hypothalamo-pituitary-adrenocortical (HPA) axis of HAA and LAA rats in response to restraint stress. The restraint stress induced an elevation in plasma concentrations of ACTH, prolactin, corticosterone and progesterone. Peak levels of plasma ACTH during stress conditions were significantly higher in HAA rats than in LAA rats, while peak levels of prolactin were significantly lower in HAA rats than in LAA rats. Under stress conditions, ACTH and prolactin synthesis in the anterior pituitary glands was significantly higher in HAA rats compared with LAA rats. The peak plasma concentrations of corticosterone, during restraint stress, were significantly higher in LAA rats compared with HAA rats. These results indicate that the response of the hypothalamo-pituitary axis to acute restraint stress is greater in HAA rats than in LAA rats, whereas the ACTH-induced adrenal response of corticosterone release is higher in LAA rats than in HAA rats. On the other hand, prolactin secretory activity is higher in LAA rats compared with HAA rats. These differences in endocrine responses to stress may be involved in the regulation of the avoidance responses in the shuttle-box task.

The ontogeny of inhibin secretion in the testis of rats was investigated. Testicular localization, content of immunoactive and bioactive inhibin and its molecular size in fetal and neonatal rats (from 16 days of gestation to 5 days of age) were determined. Strong immunostaining with an antiserum against a polypeptide of porcine inhibin alpha-subunit was noted in testicular interstitial cells from 16 days of gestation. Co-localization of inhibin alpha-subunit and 3 beta-hydroxysteroid dehydrogenase (3 beta HSD) was observed in the interstitial cells until 2 days of age. Immunoreactive inhibin alpha-subunit in the interstitial tissue had disappeared by 5 days of age, although 3 beta HSD-positive cells were still detected. Weak immunostaining for the inhibin alpha-subunit was detected in the seminiferous tubules, probably in the cytoplasm of Sertoli cells, from 20 days of gestation onward. No inhibin alpha-subunit immunostaining was observed in germ cells throughout the experimental period. Testicular inhibin was detected at 16 days of gestation (49.5 +/- 6.7 pg per testis) by RIA. Testicular immunoreactive inhibin showed a tendency to increase during fetal life and levels were maintained at a similar value after birth (697.0 +/- 46.9 pg per testis at 5 days of age). Inhibin bioactivity and its molecular size in testicular homogenate was examined at 17 days of gestation and 0 and 5 days of age. Although no bioactivity was detected at 17 days of gestation, bioactivity was noted at 0 and 5 days of age (177.7 and 1303.9 pg per testis respectively). Immunoblot analysis with antiserum against inhibin alpha-subunit revealed only approximately 40 kDa molecular masses in the testis at 17 days of gestation, probably inhibin-related proteins, but not inhibin. At 0 and 5 days of age, a protein of 30 kDa molecular mass, possibly inhibin, was detected as well as material of approximately 40 kDa molecular mass. FSH in the plasma was first detected at 19 days of gestation (1197.0 ng/l), increased towards birth, and thereafter decreased (4588.5 +/- 572.3 ng/l at 21 days of gestation and 2400.0 +/- 179.6 ng/l at 5 days of age). These results indicate that Leydig cells in fetal and neonatal rats produce inhibin-related substances with no inhibin bioactivity, whereas Sertoli cells begin to produce inhibin during the perinatal period as a possible regulator of FSH secretion.

ABSTRACT

We have characterized α₁-adrenergic receptor subtypes in functional rat thyroid cells, FRTL, with relation to iodide efflux, and have also examined the effect of TSH on α₁ receptor subtypes. FRTL cells grown in a medium containing 5 mU TSH/ml (6H cells) had five times the number of α₁ receptors of those maintained in TSH-free medium (5H cells) (11·2 fmol/10⁶ cells compared with 2·0 fmol/10⁶ cells). Pretreatment with chlorethylclonidine (CEC; 10 μmol/l), which inactivates only α_1b receptors, caused 98·8% and 97·0% decreases in the density of specific [³H]prazosin-binding sites in 5H and 6H cells respectively. LIGAND computer program analysis of the displacement curves for 2-(2,6-dimethoxyphenoxyethyl)-aminomethyl-1,4 benzodioxane (WB4101) showed that FRTL cells contained mostly low-affinity WB4101 sites. Using the phenoxybenzamine inactivation method, we found a linear relationship between α₁ receptor density and the cytosolic free Ca²⁺ concentration response in FRTL cells. Pre-exposure of intact FRTL cells to CEC caused a 98·7% decrease in noradrenaline-stimulated maximal increase in cytosolic free Ca²⁺. Also, CEC and 3,4,5-trimethoxybenzoic acid 8-(diethylamino) octyl ester (TMB-8), but not nicardipine, inhibited noradrenaline-stimulated iodine efflux. The results suggest that FRTL cells contain mostly the α_1b-adrenergic receptor subtype; that the α_1b receptors mediate cytosolic free Ca²⁺ and iodide efflux responses, and that TSH enhances these responses by increasing the α_1b receptor density without affecting the post-receptor mechanism.

Journal of Endocrinology (1990) 124, 433–441