Antiprogestagen RU486 prevents the LH-dependent decrease in the serum concentrations of inhibin in the rat

Summary 1. In the rat, the LH-dependent ovarian progesterone rise mediates several actions of the primary surge of LH on the ovary. This experiment was aimed at elucidating the effects of the antiprogestagen RU486 on the LH-dependent decrease in both the serum concentrations and the ovarian content of inhibin.2. All rats in this experiment were treated with an antagonist of LHRH (1 mg/200 µl saline at 0800 h in proestrus) to supress the endogenous release of LH. One group of rats received 32 µg LH/250 µl saline at 1200 h in proestrus. Other group was given 4 mg RU486/200 µl oil at 0800 h in proestrus. The third group was injected with both RU486 and LH. Rats from the control group were injected with 250 µl saline and 200 µl oil. Animals were decapitated at 1700 h in proestrus and trunk blood and ovaries collected to determine the serum concentrations of LH, FSH, progesterone, 17ß-estradiol and inhibin as well as the ovarian content of inhibin.3. The ovulatory dose of LH in LHRHa-treated rats decreased both the serum concentrations and the ovarian content of inhibin and increased the serum concentrations of FSH. The administration of RU486 blocked the effect of LH on the serum concentrations of inhibin but not that on the ovarian content of inhibin.4. Since the antiprogestagen RU486 blocked the effect of LH on the serum concentrations of inhibin, we conclude that ovarian progesterone, besides mediating the effects of the primary LH surge on the ovulatory process and luteinization, participates in the LH-dependent drop in the serum concentrations of inhibin in proestrous afternoon.     

Interaction between ovarian and adrenal steroids in the regulation of gonadotropin secretion

Recent work from our laboratory suggests that a complex interaction exists between ovarian and adrenal steroids in the regulation of preovulatory gonadotropin secretion. Ovarian estradiol serves to set the neutral trigger for the preovulatory gonadotropin surge, while progesterone from both the adrenal and the ovary serves to (1) initiate, (2) synchronize, (3) potentiate and (4) limit the preovulatory LH surge to a single day. Administration of RU486 or the progesterone synthesis inhibitor, trilostane, on proestrous morning attenuated the preovulatory LH surge. Adrenal progesterone appears to play a role in potentiating the LH surge since RU486 still effectively decreased the LH surge even in animals ovariectomized at 0800 h on proestrus. The administration of ACTH to estrogen-primed ovariectomized (ovx) immature rats caused a LH and FSH surge 6 h later, demonstrating that upon proper stimulation, the adrenal can induce gonadotropin surges. The effect was specific for ACTH, required estrogen priming, and was blocked by adrenalectomy or RU486, but not by ovariectomy. Certain corticosteroids, most notably deoxycorticosterone and triamcinolone acetonide, were found to possess "progestin-like" activity in the induction of LH and FSH surges in estrogen-primed ovx rats. In contrast, corticosterone and dexamethasone caused a preferential release of FSH, but not LH. Progesterone-induced surges of LH and FSH appear to require an intact N-methyl-D-aspartate (NMDA) neurotransmission line, since administration of the NMDA receptor antagonist, MK801, blocked the ability of progesterone to induce LH and FSH surges. Similarly, NMDA neurotransmission appears to be a critical component in the expression of the preovulatory gonadotropin surge since administration of MK801 during the critical period significantly diminished the LH and PRL surge in the cycling adult rat. FSH levels were lowered by MK801 treatment, but the effect was not statistically significant. The progesterone-induced gonadotropin surge appears to also involve mediation through NPY and catecholamine systems. Immediately preceding the onset of the LH and FSH surge in progesterone-treated estrogen-primed ovx. rats, there was a significant elevation of MBH and POA GnRH and NPY levels, which was followed by a significant fall at the onset of the LH surge. The effect of progesterone on inducing LH and FSH surges also appears to involve alpha 1 and alpha 2 adrenergic neuron activation since prazosin and yohimbine (alpha 1 and 2 blockers, respectively) but not propranolol (a beta-blocker) abolished the ability of progesterone to induce LH and FSH surges. Progesterone also caused a dose-dependent decrease in occupied nuclear estradiol receptors in the pituitary.(ABSTRACT TRUNCATED AT 400 WORDS)     

Hypersecretion of follicle-stimulating hormone (FSH) on estrous afternoon in rats treated with RU486 in proestrus

Summary 1. Intact or ovariectomized (OVX) cyclic rats injected or not with RU486 (4 mg/0.2 ml oil) from proestrus onwards were bled at 0800 and 1800h on proestrus, estrus and metestrus. Additional RU486-treated rats were injected with: LHRH antagonist (LHRHa), estradiol benzoate (EB) or bovine follicular fluid (bFF) and sacrified at 1800 h in estrous afternoon. LH and FSH serum levels were determined by RIA.2. RU486-treated intact or OVX rats had decreased preovulatory surges of LH and FSH, abolished secondary secretion of FSH and hypersecretion of FSH in estrous afternoon. The latter was decreased by LHRHa and abolished by EB or bFF. In contrast, EB induced an hypersecretion of LH in RU486-treated rats at 1800h in estrus.3. It can be concluded that in the absence of the proestrous progesterone actions, the absence of the inhibitory effect of the ovary in estrus evoked a LHRH independent secretion of FSH.     

A possible dual mechanism of the anovulatory action of antiprogesterone RU486 in the rat

The purpose of these experiments was to investigate the mechanism of the anovulatory action of antiprogesterone RU486 (RU486) in rats by studying its effects on follicular growth, secretion of gonadotropins and ovarian steroids, and ovulation. Rats with 4-day estrous cycles received injections (s.c.) of either 0.2 ml oil or 0.1, 1, or 5 mg of RU486 at 0800 and 1600 h on metestrus, diestrus, and proestrus. At the same times, they were bled by jugular venipuncture to determine serum concentrations of luteinizing hormone (LH), follicle-stimulating hormone (FSH), 17 beta-estradiol (E), and progesterone (P). On the morning of the day after proestrus, ovulation and histological features of the ovary were recorded. Rats from each group were killed on each day of ovarian cycle to assess follicular development. Rats treated similarly were decapitated at the time of the ovulatory LH surge and blood was collected to measure LH. The serum levels of LH increased and those of FSH decreased during diestrus in rats treated with RU486. Neither E nor P levels differed among the groups. Treatment with RU486 caused both a blockade of the ovulation and an increase in ovarian weight in a dose-dependent manner. At the time of the autopsy (the expected day of ovulation), rats treated with 1 mg RU486 had ovaries presenting both normal and post-ovulatory follicles and unruptured luteinized follicles. Rats treated with 5 mg RU486 presented post-ovulatory follicles without signs of luteinization. The number of follicles undergoing atresia increased in rats treated with RU486. Rats treated with 5 mg RU486 exhibited a significant decrease in ovulatory LH release. The mechanism by which RU486 produces the ovulatory impairment in rats seems to be dual: first, by inducing inadequate follicular development at the time of the LH surge and second, by reducing the amount of ovulatory LH released. The physiological events-decreased basal FSH secretion and follicular atresia-that result from use of RU486 cannot be elucidated from these experiments and should be investigated further.     

Hormone secretion in the asian elephant (Elephas maximus): characterization of ovulatory and anovulatory luteinizing hormone surges.

In the elephant, two distinct LH surges occur 3 wk apart during the nonluteal phase of the estrous cycle, but only the second surge (ovLH) induces ovulation. The function of the first, anovulatory surge (anLH) is unknown, nor is it clear what regulates the timing of these two surges. To further study this observation in the Asian elephant, serum concentrations of LH, FSH, progesterone, inhibin, estradiol, and prolactin were quantified throughout the estrous cycle to establish temporal hormonal relationships. To examine long-term dynamics of hormone secretion, analyses were conducted in weekly blood samples collected from 3 Asian elephants for up to 3 yr. To determine whether differences existed in secretory patterns between the anLH and ovLH surges, daily blood samples were analyzed from 21 nonluteal-phase periods from 7 Asian elephants. During the nonluteal phase, serum LH was elevated for 1-2 days during anLH and ovLH surges with no differences in peak concentration between the two surges. The anLH surge occurred 19.9+/-1.2 days after the end of the luteal phase and was followed by the ovLH surge 20.8+/-0.5 days later. Serum FSH concentrations were highest at the beginning of the nonluteal phase and gradually declined to nadir concentrations within 4 days of the ovLH surge. FSH remained low until after the ovLH surge and then increased during the luteal phase. Serum inhibin concentrations were negatively correlated with FSH during the nonluteal phase (r = -0.53). Concentrations of estradiol and prolactin fluctuated throughout the estrous cycle with no discernible patterns evident. In sum, there were no clear differences in associated hormone secretory patterns between the anLH and ovLH surge. However, elevated FSH at the beginning of the nonluteal phase may be important for follicle recruitment, with the first anLH surge acting to complete the follicle selection process before ovulation.     

Role of progesterone in the modulation of the preovulatory surge of gonadotropins and ovulation in the pregnant mare's serum gonadotropin-primed immature rat and the adult rat

The role of progesterone in the regulation of the preovulatory surge in gonadotropins and ovulation was examined in this study by use of a potent antagonist of progesterone, RU 486 (17 beta-hydroxy-11 beta-[4-dimethyl-aminophenyl]-17 alpha- [prop-1-ynyl]estra-4,9-diene-3-one). The immature rat primed with pregnant mare's serum gonadotropin (PMSG) and the cycling adult animal were the models used to verify the role of progesterone. When RU 486 (200 micrograms/rat) was given as a single dose on the morning of proestrus, there was a significant reduction in the preovulatory surge levels of gonadotropins and ovulation in both animal models. Serum progesterone levels in both models at the time of death on the evening of proestrus were unaltered upon treatment with RU 486. RU 486 did not have any effect on gonadotropin levels in immature rats 7 days after castration. These results show that the actin of RU 486 on the preovulatory gonadotropin surge is due to an antagonism of the action of progesterone on the hypothalamic-pituitary axis. Thus, a role for progesterone in modulating the preovulatory surge of gonadotropins and, consequently, ovulation is strongly suggested.     

Gonadotropin surge induces two separate increases in messenger RNA for progesterone receptor in bovine preovulatory follicles

In mice deficient in progesterone receptor (PR), follicles of ovulatory size develop but fail to ovulate, providing evidence for an essential role for progesterone and PR in ovulation in mice. However, little is known about the expression and regulation of PR mRNA in preovulatory follicles of ruminant species. One objective of this study was to determine whether and when PR mRNA is expressed in bovine follicular cells during the periovulatory period. Luteolysis and the LH/FSH surge were induced with prostaglandin F(2alpha) and a GnRH analogue, respectively, and the preovulatory follicle was obtained at 0, 3.5, 6, 12, 18, or 24 h after GnRH treatment. RNase protection assays revealed a transient increase in levels of PR mRNA, which peaked at 6 h after GnRH and declined to the time 0 value by 12 h and a second increase at 24 h. The second objective was to investigate the mechanisms that regulate PR mRNA expression through in vitro studies on follicular cells of preovulatory follicles obtained before the LH/FSH surge. Theca and granulosa cells were isolated and cultured with or without a luteinizing dose of LH or FSH, progesterone, LH + progesterone, or LH + antiprogestin (RU486). Levels of PR mRNA increased in a time-dependent manner in granulosa cells cultured with LH or FSH and in theca cells cultured with LH, peaking at 10 h of culture. In contrast, progesterone (200 ng/ml) did not upregulate mRNA for its own receptor, and neither progesterone nor RU486 affected LH-stimulated PR mRNA accumulation. Furthermore, RU486 completely blocked LH-stimulated expression of oxytocin mRNA, indicating that PR induced by LH in vitro is functional. These results show that the gonadotropin surge induces a rapid and transient increase in expression of PR mRNA in both theca and granulosa cells of bovine periovulatory follicles followed by a second rise close to the time of ovulation and that the first increase in PR mRNA can be mimicked in vitro by gonadotropins but not by progesterone. These results suggest multiple and time-dependent roles for progesterone and PR in the regulation of periovulatory events in cattle.     

Inhibin activity and secretion of gonadotropin during the period of follicular maturation

To evaluate the relative contributions of the ovarian inhibin and estradiol-17 beta (E) on the regulation of FSH secretion, inhibin and E in ovarian venous plasma (OVP) and FSH and LH in peripheral plasma were simultaneously measured using superovulating rats with special reference to follicular maturation. By the transplantation of a pituitary gland from adult male rats under the kidney capsule between 1100 and 1200 hr on diestrus-1 in cyclic rats, superovulation was successfully induced on the morning of the next estrus without any additional treatment with human chorionic gonadotropin (hCG). The number of maturing follicles capable of ovulating in response to hCG significantly increased at 12 hours after the grafting as compared with sham-operated controls and further increases occurred until the afternoon of proestrus. In the superovulating rat, first and second surges of FSH were completely blocked and an LH surge was also partially suppressed during the periovulatory period when surges of FSH and LH were normally observed in controls. Contents of FSH as well as LH in the animal's own pituitary gland were suppressed significantly after the grafting as compared with controls. A marked increase in inhibin activity in OVP of rats with a pituitary transplant occurred concomitantly with an increase in the number of follicles capable of ovulating whereas E levels in OVP did not so. Inhibin activity in OVP at each point was much higher in the pituitary grafted rats than in controls but this was not true for E levels. These results suggest that ovarian inhibin derived from the maturing follicles rather than E may be a primary factor for regulation of FSH secretion, and high levels of endogenous inhibin can suppress synthesis of LH as well as FSH in the pituitary gland of the female rat.     

Mechanism of the stimulatory action of antisteroid RU486 on follicle-stimulating hormone secretion in the cyclic Rat

These experiments explored the mechanism underlying FSH hypersecretion on estrous afternoon in rats injected with RU486 (RU) on proestrus. Four-day cyclic rats were injected with RU at 12:00 h on proestrus (1 or 4 mg/0.2 ml oil; s.c.), and its effects on LH and FSH secretion at 18:30 h on estrus were compared with those of antiprogestagens ZK299 (ZK) (1 or 4 mg/0.2 ml oil; s.c.) and Org31806 (OR) (2 or 8 mg/0.2 ml oil; s.c.). Additionally, rats treated with RU or nembutal (PB) (60 mg/kg; i.p. at 13:00 h on proestrus) were injected with an LHRH antagonist (LHRHa) at 10:00 h on estrus (1 mg/0.2 ml saline; s.c.) or progesterone (P) (7.7, 15.5 or 30.9 mg/0.2 ml oil; s.c.) on proestrus at 10:00 h in RU-injected rats and at 14:00 h in PB-injected rats. Animals were killed by decapitation at 18:30 h on estrus and serum LH and FSH concentrations were determined. Rats treated with 1 or 4 mg of RU or Org or 4 mg of ZK recorded increased serum FSH on estrous afternoon, while 1 mg ZK had no effect. PB increased mainly serum LH levels and, to a lesser extent, FSH levels. P decreased serum FSH concentrations in both RU- and PB-injected rats. LHRHa reversed the effects of PB on FSH secretions, but reduced FSH hypersecretion induced by RU only. These results are interpreted to mean that, in the absence of proestrous afternoon P-inhibitory action of the neural stimulus controlling LHRH release, FSH secretion on estrous afternoon involves two components: one is LHRH dependent while, in contrast to LH secretion, the other is LHRH independent, and only expressed in a low estrogen background.     

Decreased luteinizing hormone-stimulated progesterone secretion by preovulatory follicles isolated from cyclic rats treated with the progesterone antagonist RU486.

Since administration of the antiprogesterone RU486 to cyclic female rats at metestrus and diestrus results in increased serum levels of LH, estradiol, and testosterone at proestrus, we investigated whether RU486 affects follicular steroidogenesis. Female rats with a 4-day estrous cycle, induced experimentally by a single injection of bromocriptine on the morning of estrus, were given RU486 (2 mg) twice daily (0900 and 1700 h) on metestrus and diestrus. At proestrus the preovulatory follicles were isolated and incubated for 4 h in the absence and presence of LH. In the absence of LH, accumulation of estradiol, testosterone, and progesterone in the medium was not different for RU486-treated rats and oil-treated controls. In contrast, LH-stimulated estradiol, testosterone, and progesterone secretions were significantly lower in RU486-treated rats compared with controls. Addition of pregnenolone to the incubation medium resulted in a significantly lower increase of progesterone in follicles from RU486-treated rats compared with those from oil-treated controls. This suggests that 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) activity is decreased by administration of RU486 in vivo. Aromatase and 17 alpha-hydroxylase/C17-20 lyase activities were not affected: addition of substrate (androstenedione and progesterone respectively) did not affect differently the amount of product formed (estradiol and testosterone) in RU486- and oil-treated rats. However, LH-stimulated pregnenolone secretion was lower in follicles from RU486-treated rats compared with follicles from oil-treated controls, suggesting that either cholesterol side-chain cleavage activity or LH responsiveness is decreased. At proestrus the preovulatory follicles from RU486- and oil-treated rats were not morphologically different.(ABSTRACT TRUNCATED AT 250 WORDS)     

Rapid recovery of estrogen-induced pituitary gonadotropin surges after luteectomy in rhesus monkeys

In the presence of a functional corpus luteum, positive estrogen feedback on the surge modes of gonadotropin secretion is blocked in rhesus monkeys. We investigated the effects of luteectomy (Lx) on the time required for recovery of pituitary responsiveness (LH/FSH surges) to positive estrogen feedback. Estradiol-17 beta-3- benzoate (EB, 50 microgram/kg sc) was given: 1) 24th prior to, 2) the day of, or 3) 24 h after luteal ablation. Daily measurements of serum follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol-17 beta (e2) and progesterone (P) were made on each monkey for 5 days. Serum P fell to undetectable levels within 24 h after Lx, whereas E2 levels in circulation peaked within 24h after injection of EB. Among early follicular phase monkeys, this EB treatment results in typical midcycle type LH/FSH surges within 48h. Lx alone was not soon followed by significant changes in pituitary gonadotropin secretion. When circulating P levels were undetectable the pituitary responded fully to EB; that is, typical midcycle type FSH/LH surges occurred. When serum P was in the midst of declining after Lx, gonadotropin surges were present, but attenuated. However, when P levels remained elevated for more than 24 h after EB injection, the surge modes of FSH/LH secretion remained fully blocked. These results demonstrate that the suppressive influence of luteal secretions (principally progesterone) on positive estrogen feedback regulation of the surge modes of pituitary gonadotropin secretion is quite transient in these primates.     

Circulating radioimmunoassayable inhibin during periods of transient follicle-stimulating hormone rise: secondary surge and unilateral ovariectomy

We have measured changes in circulating immunoreactive (ir-) inhibin in male and female rats using an RIA with an antiserum raised against porcine inhibin alpha (1-26)-Gly-Tyr. The same synthetic peptide was used for standards and for the preparation of tracer. Serum ir-inhibin levels were significantly higher in intact female than in intact male rats (p less than 0.001). Immunoreactive inhibin was significantly reduced in both sexes 24 h after bilateral gonadectomy (p less than 0.0001). Unilateral ovariectomy (ULO) of female rats on metestrus caused a transient decrease in serum inhibin 8 h after surgery, but levels were not significantly different from those of sham-operated controls at later times after surgery. Increases in serum FSH and LH were observed for 8-18 h after ULO. Serum ir-inhibin levels were also measured on the early morning of estrus during the secondary FSH surge. At this time, ir-inhibin levels were low, while FSH levels were high and LH levels were low. These results show that serum ir-inhibin levels in rats are decreased at times when serum FSH levels are high.     

Detailed examination of the mechanism and site of action of progesterone and corticosteroids in the regulation of gonadotropin secretion: hypothalamic gonadotropin-releasing hormone and catecholamine involvement.

Progesterone and certain corticosteroids, such as deoxycorticosterone (DOC) and triamcinolone acetonide (TA), can stimulate gonadotropin surges in rats. The mechanism of these steroids could involve a pituitary or hypothalamic site of action, or both. Progesterone and TA did not alter the ability of GnRH to release LH or FSH either before, during, or after the gonadotropin surge induced by these steroids in estrogen-primed ovariectomized female rats. Furthermore, progesterone, TA and DOC were unable to induce a gonadotropin surge in short-term estrogen-primed castrated male rats. These results suggested a hypothalamic rather than a pituitary site of action of progesterone and corticosteroids in the release of gonadotropins. Since progestin and corticosteroid receptors are present in catecholamine neurons, a role for catecholamine neurotransmission in progesterone and corticosteroid-induced surges of LH and FSH in estrogen-primed ovariectomized rats was examined. Catecholamine synthesis inhibitors and specific alpha 1 (prazosin), alpha 2 (yohimbine), and beta (propranolol) receptor antagonists were used to determine the role of catecholamine neurotransmission in the steroid-induced surges of LH and FSH. Both of the catecholamine synthesis inhibitors, alpha-methyl-p-tyrosine HCl (alpha-MPT), a tyrosine hydroxylase inhibitor, and sodium diethyldithiocarbamate (DDC), an inhibitor of dopamine-beta-hydroxylase, attenuated the ability of progesterone, TA, and DOC to induce LH surges when administered 3 h and 1 h, respectively, before the steroid. DDC also suppressed the ability of progesterone, TA, and DOC to induce FSH surges. Rats treated with alpha-MPT had lower mean FSH values than did steroid controls, but the effect was not significant. Both the alpha 1 and alpha 2 adrenergic antagonists, prazosin and yohimbine, significantly suppressed the ability of progesterone, TA, and DOC to induce LH and FSH surges. In contrast, the beta adrenergic receptor blocker, propranolol, had no effect upon the ability of progesterone, TA, or DOC to facilitate LH and FSH secretion. Finally, the stimulatory effect of progesterone and TA upon LH and FSH release was found to be blocked by prior treatment with a GnRH antagonist, further suggesting hypothalamic involvement. In conclusion, this study provides evidence that the stimulation of gonadotropin release by progesterone and corticosteroids is mediated through a common mechanism, and that this mechanism involves the release of GnRH, most likely through catecholaminergic stimulation. Furthermore, catecholamine neurotransmission, through alpha 1 and alpha 2 but not beta receptor sites, is required for the expression of progesterone and corticosteroid-induced surges of LH and FSH in estrogen-primed ovariectomized rats.     

Integration of the effects of estradiol and progesterone in the modulation of gonadotropin secretion

Estradiol secreted by the maturing follicle is the primary trigger for the surge of gonadotropins leading to ovulation. Progesterone has stimulatory or inhibitory actions on this estrogen-induced gonadotropin surge depending upon the time and dose of administration. The administration of progesterone to immature ovariectomized rats primed with a low dose of estradiol induced a well-defined LH surge and prolonged FSH release, a pattern similar to the proestrus surge of gonadotropins. A physiological role of progesterone is indicated in the normal ovulatory process because a single injection of the progesterone antagonist RU 486 on the day of proestrus in the adult cycling rat and on the day of the gonadotropin surge in the pregnant mare's serum gonadotropin stimulated immature rat resulted in an attenuated gonadotropin surge and reduced the number of ova per ovulating rat. Progesterone administration brought about a rapid LHRH release and an decrease in nuclear accumulation of estrogen receptors in the anterior pituitary but not the hypothalamus. The progesterone effect was demonstrated in vitro in the uterus and anterior pituitary and appears to be confined to occupied estradiol nuclear receptors. In in vivo experiments the progesterone effect on estradiol nuclear receptors appeared to be of approximately 2-h duration, which coincided with the time period of progesterone nuclear receptor accumulation after a single injection of progesterone. During the period of progesterone effects on nuclear estrogen receptors, the ability of estrogens to induce progesterone receptors was impaired. Based on the above results, a model is proposed for the stimulatory and inhibitory effects of progesterone on gonadotropin secretion.     

Effects of supraphysiological concentrations of progesterone on the characteristics of the oestradiol-induced gonadotrophin surge in women

Intramuscular injections of oestradiol benzoate were given to 8 normally cyclic women in the early follicular phase of 3 different cycles. Progesterone was also injected in the second (low dose) and the third cycle (high dose). Oestradiol induced simultaneous surges of LH and FSH in all women and the onset of these surges was advanced by progesterone. Low-dose progesterone induced a significant increase in the amplitude and the duration of the LH and FSH surges, while high-dose progesterone decreased the duration significantly. In contrast to the oestrogen-only treatment cycles, when the women were treated with progesterone, basal LH and FSH concentrations were suppressed significantly not only before the onset but also after the end of the surge. The results suggest that progesterone affects the dimension of the oestradiol-induced gonadotrophin surge by exerting both a stimulatory and an inhibitory effect on pituitary gonadotrophin secretion. Supraphysiological concentrations of progesterone decreased the duration of the oestradiol-induced gonadotrophin surge significantly and this is possibly part of the mechanism which attenuates the endogenous LH surge in women superovulated for in-vitro fertilization.     

Effects of bovine follicular fluid inhibin on serum gonadotrophin concentrations in ewes during oestrus

Experiments were conducted with ewes to investigate the effects of an enriched bovine follicular fluid inhibin preparation (INH) on gonadotrophin secretion after the onset of oestrus. Administration of INH (10 mg) 1 h after the onset of oestrus did not significantly alter the preovulatory FSH and LH surges or the second FSH peak. To determine the effects of INH on the second FSH surge, ewes were treated with saline (N = 7) or INH (N = 10) at 4 h (10 mg) and 24 h (5 mg) after the peak of the preovulatory LH surge. The second FSH surge was delayed about 24 h (P less than 0.05) in ewes treated with INH; however, the delay did not alter the interval to the next oestrus. In a third experiment, 16 ewes were assigned to 4 groups in a 2 x 2 factorial with the main effects being ovariectomy at 4 h and INH treatment (10 mg) at 4, 20 and 36 h after the peak of the LH surge. Controls received sham ovariectomy and saline injection as appropriate. Ovariectomy resulted in a rapid increase in serum FSH but not LH and this was delayed (P less than 0.05) by INH treatment. These results indicate that inhibin has a selective inhibitory action on FSH secretion in ewes and suggests that the second FSH surge results from increased basal FSH secretion due to decreased endogenous inhibin levels.     

Luteal expression of cytochrome P450 side-chain cleavage, steroidogenic acute regulatory protein, 3beta-hydroxysteroid dehydrogenase, and 20alpha-hydroxysteroid dehydrogenase genes in late pregnant rats: effect of luteinizing hormone and RU486

A decrease in serum progesterone at the end of pregnancy is essential for the induction of parturition in rats. We have previously demonstrated that LH participates in this process through: 1) inhibiting 3beta-hydroxysteroid dehydrogenase (3beta-HSD) activity and 2) stimulating progesterone catabolism by inducing 20alpha-hydroxysteroid dehydrogenase (20alpha-HSD) activity. The objective of this investigation was to determine the effect of LH and progesterone on the luteal expression of the steroidogenic acute regulatory protein (StAR), cytochrome P450 side-chain cleavage (P450(scc)), 3beta-HSD, and 20alpha-HSD genes. Gene expression was analyzed by Northern blot analysis 24 and 48 h after administration of LH or vehicle on Day 19 of pregnancy. StAR and 3beta-HSD mRNA levels were lower in LH-treated rats than in rats administered with vehicle at both time points studied. P450(scc) mRNA levels were unaffected by LH. The 20alpha-HSD mRNA levels were not different between LH and control rats 24 h after treatment; however, greater expression of 20alpha-HSD, with respect to controls, was observed in LH-treated rats 48 h after treatment. Luteal progesterone content dropped in LH-treated rats at both time points studied, whereas serum progesterone decreased after 48 h only. In a second set of experiments, the anti-progesterone RU486 was injected intrabursally on Day 20 of pregnancy. RU486 had no effect on 3beta-HSD or P450(scc) expression but increased 20alpha-HSD mRNA levels after 8 h treatment. In conclusion, the luteolytic effect of LH is mediated by a drop in StAR and 3beta-HSD expression without effect on P450(scc) expression. We also provide the first in vivo evidence indicating that a decrease in luteal progesterone content may be an essential step toward the induction of 20alpha-HSD expression at the end of pregnancy in rats.     

Changes in the ovarian dynamics and endocrine profiles in goats treated with a progesterone antagonist during the early luteal phase of the estrous cycle

The aim of the present study was to determine the physiological role of endogenous progesterone in the regulation of ovarian dynamics, gonadotropin and progesterone secretion during the early luteal phase in the goat. Cycling Shiba goats received subcutaneously a vehicle (control group, n=5) or 50 mg of RU486 (RU486 group, n=4) daily from 1 to 7 days after ovulation (day 0) determined by transrectal ultrasonography. Ovarian dynamics were monitored by the ultrasonography and blood samples were collected daily until the subsequent ovulation for analysis of progesterone, luteinizing hormone (LH) and follicle stimulating hormone (FSH) secretion. Blood samples were also collected at 10 min intervals for 6 h on day 3 and day 7 for the analysis of pulsatile patterns of LH and FSH. The LH pulse frequency was significantly (P<0.05) higher in the RU486 group than in the control group on day 7 (4.8+/-1.1 pulses/6 h versus 1.2+/-0.4 pulses/6 h). The shape of the FSH pulses was unclear on day 3 and day 7 in both groups and the overall means of FSH concentration for 6 h on day 3 and day 7 were not significantly different between the RU486 and the control groups. The pattern of daily FSH concentrations showed a wave-like fluctuation in both groups. There was no significant difference in the inter-peak intervals of the wave-like pattern of daily FSH secretion between the RU486 and the control groups (4.1+/-0.6 days versus 4.5+/-0.6 days). The maximum diameter of the largest follicle that grew from day 1 to day 7 in the RU486 group tended to be greater than that in control goats (6.4+/-0.8 mm versus 5.0+/-0.8 mm, P=0.050), whereas no significant difference was detected in the size of the corpus luteum and progesterone concentrations between the control and RU486 groups on almost all days during the treatment period. These results indicate that the rise of the progesterone concentration suppresses the pulsatile LH secretion and follicular growth, whereas progesterone has no physiological role in the regulation of FSH secretion and luteal function during the early luteal phase of the estrous cycle in goats.     

Unilateral ovariectomy, flutamide treatment and HCG reverse the anovulatory action of antiprogesterone RU486 in rat.

Administration of antiprogesterone RU486 (4 mg/day) from estrus through proestrus to cyclic rats blocked ovulation. Moreover, RU486 increased basal serum concentrations of LH, PRL, testosterone and estradiol, while it decreased basal serum concentration of FSH. Both unilateral ovariectomy and antiandrogen flutamide treatment, as well as an ovulatory injection of HCG in the proestrus afternoon partially reversed, the ovulatory blockade of RU486. These results indicate that both the decreased FSH concentration and the increased testosterone concentration, as well as the reduced ovulatory LH release are responsible for the anovulatory effects of RU486.