Endocrine-regulated and protein kinase C-dependent generation of superoxide by rat preovulatory follicles

The ovulatory LH surge results in follicular inflammation with an increase in cytokines and PGs. Reactive oxygen species (ROS) are also produced during inflammatory processes. To study ROS generation during the ovulatory cascade, preovulatory follicles were dissected from immature female rats primed with PMSG. Follicles were isolated, and ROS generation was assessed by luminol-amplified chemiluminescence. Immature rat granulosa cells were also subjected to luminometry after isolation from immature rats treated with diethylstilbestrol. Phorbol ester-stimulated ROS generation by follicular cells was completely suppressed by superoxide dismutase and the NADPH/NADH oxidase inhibitor diphenylene iodonium bisulfate, whereas catalase was without effect. Fractionation of granulosa cells with an antibody against leukocyte common antigen-1 showed that leukocyte-enriched cells produced more than 95% of the superoxide measured. In vivo treatment with LH produced a 5-fold increase in phorbol-stimulated superoxide production by isolated follicles. This response was maximal within 4 h and was blocked by indomethacin. In vivo administration of PGE(2) and PGF(2alpha) did not reverse the blockade by indomethacin; however, isolated follicles incubated with PGE(2) produced a time-dependent increase in phorbol-stimulated superoxide generation. Thus, a superoxide generator is present in the preovulatory follicle that is leukocytic in origin, hormone regulated, and activated by a protein kinase C-dependent pathway. The regulated generation of superoxide by preovulatory follicles may indicate a role for ROS in the periovulatory period.     

Changes in ornithine decarboxylase activity in ovarian follicles of the laying hen (Gallus domesticus) during the ovulatory cycle

Fowl ovarian ornithine decarboxylase activity was measured at 20, 10 and 3 h before an expected ovulation in granulosa and thecal tissues from follicles at various stages of development. An increase in the enzyme activity between 10 and 3 h before an expected ovulation was assumed to be caused by preovulatory increase in plasma LH concentration. The activity in granulosa tissue increased with increasing size of the follicle. In the largest (F1) follicle there was an 11-fold increase in granulosa ornithine decarboxylase specific activity between 10 and 3 h before ovulation. In the third (F3) and fifth (F5) largest follicles there was a 1.9- and 2-fold increase respectively. The enzyme activity in thecal tissue from the follicular hierarchy decreased with increasing size of the follicle and the F3 thecal preparation was the only tissue to respond to the preovulatory LH surge. In contrast, ornithine decarboxylase activity in thecal tissue from small (less than 5 mm) non-atretic follicles increased by two- to threefold after the preovulatory LH surge. The activity in atretic follicles of the same size was low and remained unchanged throughout the ovulatory cycle.     

Progesterone secretion by highly differentiated human granulosa cells isolated from preovulatory Graafian follicles induced by exogenous gonadotropins and human chorionic gonadotropin

Human granulosa-luteal cells were harvested from preovulatory Graafian follicles at the time of oocyte retrieval for in vitro fertilization after induction of follicle maturation by sequential injections of menopausal gonadotropins and hCG. Such highly differentiated granulosa cells produced large quantities of progesterone basally (6.8 pg/cell X 2 days) in monolayer culture. Human LH significantly increased progesterone biosynthesis after 6, 12, 48, 96, or 144 h in culture, with a maximal increase of 8- to 20-fold occurring at 96 h. The stimulatory effect of LH could be observed under serum-free conditions and was maximal in the presence of 4% serum. Human granulosa-luteal cells also exhibited significant stimulatory responses to hCG, prostaglandin E2, or the cAMP effectors 8-bromo cAMP, choleratoxin, or forskolin in serum-free incubations. Concentrations of 17 beta-estradiol that are attained physiologically in ovarian follicles in vivo markedly suppressed basal and LH (or cAMP)-stimulated progesterone production in vitro (maximal suppression, greater than 90%). The nonaromatizable androgen 5 alpha-dihydrotestosterone also inhibited progesterone production, but by no more than 45-50% even at supraphysiological concentrations. Estradiol's blockade of progesterone synthesis was associated with a corresponding increase in pregnenolone accumulation. The present studies indicate that human granulosa-luteal cells isolated from preovulatory follicles induced with exogenous gonadotropins and hCG secrete large quantities of progesterone in vitro. Such cells retain stimulatory responses to human LH, hCG, prostaglandin E2, and classical cAMP effectors in serum-free incubations. Moreover, physiological concentrations of 17 beta-estradiol suppress progesterone production, probably by inhibiting cellular conversion of pregnenolone to progesterone. Thus, the present in vitro system permits an investigation of hormone action in well differentiated, human granulosa-luteal cells isolated from preovulatory Graafian follicles that have a defined endocrine history of prior gonadotropin exposure in vivo.     

Preovulatory secretion of steroids by cultured cumulus oophorus complexes of the rat: effects of FSH and LH.

The main objective of the present study was to establish whether a shift in steroid production, previously observed for preovulatory follicles, also takes place in preovulatory cumulus oophorus complexes (COCs). Female Wistar rats, displaying a regular 4-day oestrous cycle, were killed in succession every 2 or 3 h on the day of prooestrus and oestrus until ovulation (11.00-24.00 h). From excised ovaries preovulatory follicles were isolated. After puncturing cumuli oophori were aspirated and subsequently cultured for 24 h either in hormone-free or FSH- or LH- or FSH plus LH-supplemented medium. Cultured COCs released only a small amount of androgens, the main steroid produced being oestradiol. Its secretion decreased before ovulation (24.00 h). Relatively high progesterone release occured only in cultures set up at 22.00 and 24.00 h, thus during cumulus expansion. FSH and LH present in the medium effected above all oestradiol release, stimulating it before the presumptive endogenous gonadotrophin surge and inhibiting it thereafter. The activity of delta5-3beta-hydroxysteroid dehydrogenase (3betaHSD) investigated in cryostat sections appeared in COCs at 22.00h and was still present at 24.00 h. However, the activity was much weaker than in the granulosa cells lining the basal lamina. This in vivo study confirms in vitro results on more intense progesterone synthesis during cumulus expansion. The results indicate that in preovulatory COCs a shift in steroid production occurs after which the main steroid synthesized is progesterone, while oestradiol secretion decreases. However, this switch in COCs takes place later than the previously established shift in whole follicles.     

Norepinephrine, active norepinephrine transporter, and norepinephrine-metabolism are involved in the generation of reactive oxygen species in human ovarian granulosa cells

The neurotransmitter norepinephrine (NE) is derived from the sympathetic nervous system and may be involved in the regulation of ovarian functions. Ovarian innervation increases in patients with polycystic ovarian syndrome (PCOS), prompting us to readdress a role of NE in the human ovary. In vitro fertilization-derived granulosa cells (GC), follicular fluids (FF), and ovarian sections were studied. NE was found in FF and freshly isolated GC, yet significantly lower levels of NE were detected in samples from PCOS patients. Furthermore, the metabolite normetanephrine was detected in FF. Together this suggests cellular uptake and metabolism of NE in GC. In accordance, the NE transporter and NE-metabolizing enzymes [catechol-o-methyltransferase (COMT) and monoamine oxidase A] were found in GC, COMT in GC and thecal cells of large human antral follicles in vivo and in cultured GC. Cellular uptake and metabolism of NE also occurred in cultured GC, events that could be blocked pharmacologically. NE, in the range present in FF, is unlikely to affect GC via activation of typical α- or β-receptors. In line with this assumption, it did not alter phosphorylation of MAPK. However, NE robustly induced the generation of reactive oxygen species (ROS). This action occurred even when receptors were blocked but was prevented by blockers of NE transporter, COMT, and monoamine oxidase A. Thus, NE contributes to the microenvironment of preovulatory human follicles and is lower in PCOS. By inducing the production of ROS in GC, NE is linked to ROS-regulated events, which are emerging as crucial factors in ovarian physiology, including ovulation.     

Preovulatory changes in ovarian expression of collagenases and tissue metalloproteinase inhibitor messenger ribonucleic acid: role of eicosanoids.

The preovulatory surge of gonadotropins activates a cascade of proteolytic enzymes resulting in the rupture of the follicular wall and the release of a fertilizable ovum during ovulation. In the rat the process is initiated by a rise in follicular tissue-type plasminogen activator, produced predominantly in granulosa cells. Recent studies revealed a preovulatory increase in ovarian collagenolytic activity in vivo and an increase in activatable collagenase in vitro. In view of the complicated control of mammalian collagenase synthesis and activity by local inhibitors and activators, we examined the expression of ovarian interstitial and type IV collagenases and tissue inhibitor of metalloproteinase (TIMP) mRNA after an ovulatory stimulus. Ovarian mRNA was isolated from immature PMSG-treated rats 3, 6, and 9 h after hCG stimulation. Northern blot analyses revealed a mRNA of 1.7 kilobases (kb) hybridizing with the human interstitial collagenase cDNA probe. The levels of this mRNA showed a 25-fold increase between 3-6 h after hCG stimulation. The human cDNA probe of collagenase IV hybridized with a mRNA of 3.1 kb, which showed only a 4-fold increase 9 h after hCG treatment. The interstitial collagenase mRNA was expressed in both granulosa cells of preovulatory follicles and the residual ovarian tissue, whereas the expression of collagenase IV mRNA was limited to the residual tissue. Inhibitors of eicosanoid synthesis, previously shown to block ovulation and the LH/hCG-induced rise in ovarian collagenolysis, suppressed the gonadotropic stimulation of interstitial collagenase mRNA, but slightly stimulated that of collagenase IV. The mouse cDNA probe of TIMP hybridized with a 0.9-kb mRNA, which was stimulated by hCG to reach a maximum (7- to 8-fold increase) between 6-9 h after stimulation. TIMP was expressed and stimulated in both the granulosa cells and the residual tissue. Inhibitors of eicosanoid synthesis did not affect the gonadotropic stimulation of TIMP mRNA. These data support the suggested role of interstitial collagenase in follicle rupture and the essential role of eicosanoids in the mediation of gonadotropic stimulation of interstitial collagenase production and action. The observed stimulation of TIMP mRNA expression by the gonadotropin and the lack of any effect of eicosanoid synthesis inhibitors on this action of LH/hCG offer an additional mechanism by which these inhibitors may block ovulation. Thus, the suppression of ovulation by inhibitors of eicosanoid synthesis may result from selective inhibition of interstitial collagenase expression and undisturbed gonadotropin-stimulated TIMP expression.     

Stage-specific expression of pituitary adenylate cyclase-activating polypeptide type I receptor messenger ribonucleic acid during ovarian follicle development in the rat

Expression of pituitary adenylate cyclase-activating polypeptide (PACAP), a neuropeptide with considerable homology to vasoactive intestinal peptide, has been shown to be stimulated by gonadotropins in the ovary. The present studies further evaluated the cell-type specific expression and gonadotropin regulation of PACAP type I receptor (PACAPR) messenger RNA in immature rat ovaries and in cultured preovulatory follicles. Northern blot analysis of ovaries obtained from prepubertal rats revealed the increased expression of PACAPR during prepubertal development. The major cell types expressing PACAPR messenger RNA were granulosa cells of large preantral follicles. Treatment of immature rats with PMSG caused a decrease in ovarian PACAPR expression. In contrast, treatment with human (h) CG at 2 days after PMSG treatment stimulated ovarian PACAPR messenger RNA within 3-6 h in granulosa cells of preovulatory follicles. Treatment of cultured preovulatory follicles in vitro with LH further confirmed the time- and dose-dependent stimulation of PACAPR by gonadotropins in granulosa cells of preovulatory follicles. Moreover, RNase protection assay revealed that the short variant of ovarian PACAPR was the predominant form stimulated during prepubertal development and by gonadotropins. These results demonstrate the expression of PACAPR messenger RNA in granulosa cells of growing follicles and of preovulatory follicles stimulated by gonadotropins, and suggest that PACAP may play a role in the growth of developing follicles and in ovulation as an autocrine/paracrine factor.     

Duration of the reproductive period decreases the frequency of preovulatory luteinizing hormone surges in heavy weight-sire line turkey hens

The frequency of preovulatory luteinizing hormone (LH) surges is an important determinant of ovulation and oviposition rates in turkeys. Egg production rate is relatively poor in heavy weight-sire line type turkey hens and declines with advancing duration of the reproductive period. The purpose of this study was to measure frequency and characteristics of preovulatory LH surges in turkey hens of a heavy weight-sire line type early, at peak of egg production (Early), and late, after egg production rate had declined (Late), in a reproductive period. The Early hens were photostimulated with a continuous photoperiod [24 h light (L):0 h dark (D)] at 40 weeks of age and sampled during peak egg production at about 47 weeks of age. The Late hens were photostimulated at 40 weeks of age with a long day photoperiod (14L:10D). After a 27-week egg production period, the Late hens were switched to the 24L:0D photoperiod and sampled at 74 weeks of age. Continuous lighting was used during blood sampling to allow the rhythm of preovulatory LH surges to free run. All hens were cannulated 3-5 days before starting sampling and hourly blood samples were collected for 200 h. All hens were necropsied and ovarian and oviductal morphologies were measured after serial bleeding. The Late hens had a longer interval between intra-clutch preovulatory LH surges than the Early hens, and a higher incidence of atretic ovarian follicles. The Early hens had higher baseline and surge amplitude LH concentrations but lower progesterone (P4) surge amplitude concentrations than the Late hens. The duration of preovulatory LH surges, incidence of "blind" preovulatory LH surges, baseline P4 concentrations, and overall estradiol-17beta (E2) concentrations were not different between Early and Late hens. In conclusion, a longer interval between preovulatory LH surges, lower LH baseline and surge amplitude concentrations, a higher incidence of atretic follicles, and higher P4 surge amplitude concentration were associated with the decline in egg production late in the reproductive period in a heavy weight-sire line of turkey hens.     

Clinical review 126: Roles and novel regimens of luteinizing hormone and follicle-stimulating hormone in ovulation induction

Although FSH is universally recognized as the key driver of ovarian follicle growth and maturation, the role of LH in these processes is more controversial. LH acts on theca cells to induce androgen substrate for estrogen conversion by the aromatase system; furthermore, LH can affect granulosa cell function starting in the mid- follicular phase, when these cells express LH receptors. The capacity of LH to stimulate granulosa cells in larger follicles (>10 mm diameter) may be the critical mechanism involved in the selection of the dominant follicle in the normal menstrual cycle. Furthermore, the addition of LH activity can shorten and optimize FSH ovulation induction and reduce the development of smaller preovulatory ovarian follicles that are associated with the severe complications of this procedure. Novel mixed gonadotropin administration regimens that incorporate graded amounts of LH and FSH activity may improve efficacy, safety, and cost of ovulation induction, particularly in the area of assisted reproduction.     

Luteinizing hormone-induced caspase activation in rat preovulatory follicles is coupled to mitochondrial steroidogenesis

Atresia and luteolysis are well-documented processes in which most of the growing ovarian follicles and all corpora lutea, respectively, are eliminated by apoptosis. We have previously reported that LH and FSH enhance caspase-3 and -7 activity and apoptosis in the theca-interstitial cells of rat preovulatory follicles in culture. Here we have used cultured follicles to examine whether LH-induced caspase activation is related to the ability of LH to stimulate steroid production. In these studies, we used three inhibitors of enzymes involved in steroid production: aminoglutethimide and ketoconazole, acting on cytochrome P450 side-chain cleavage (P450scc) located at the mitochondria, and epostane, acting on 3beta-hydroxysteroid dehydrogenase located at the endoplasmic reticulum. We found that treatment with either aminoglutethimide or ketoconazole, but not with epostane, significantly reduced LH-induced caspase-3 and -7 activation and apoptosis, suggesting the mediation of LH-induced caspase activation by P450scc. Supplementing pregnenolone, the product of P450scc catalysis, to follicles treated with aminoglutethimide did not restore LH-induced caspase activation. On the other hand, treatment with antioxidants inhibited LH-induced caspase activation. Moreover, LH treatment was associated with an increase in reactive oxygen species which was inhibited by aminoglutethimide. Thus, P450scc catalysis results in an increase in reactive oxygen species, which in turn may trigger/facilitate caspase-3 activation. Finally, we found that in rat corpora lutea in vivo, an increase in steroidogenesis was accompanied by an increase in caspase activity. Thus, this study reveals a linkage between two seemingly distinct processes in which LH-induced caspase activation in cultured rat preovulatory follicles is coupled to mitochondrial steroidogenesis via P450scc.     

Progesterone, androstenedione and oestradiol content of theca and granulosa tissues of the four largest ovarian follicles during the ovulatory cycle of the hen (Gallus domesticus)

The progesterone, androstenedione and oestradiol contents of the theca and granulosa tissues of the four largest follicles in the ovarian hierarchy of the hen were determined. The granulosa tissue contained significantly (P less than 0.05) more progesterone and less androstenedione and oestradiol than the theca tissue. The content of progesterone was greatest in the granulosa tissue of the first three follicles in the hierarchy and in each of these follicles there was a peak in progesterone content of the granulosa 4 h before ovulation. The theca of the second, third and fourth follicles and the granulosa of the third and fourth follicles contained significantly (P less than 0.05) more androstenedione than either tissue in the largest follicle. The content of androstenedione was maximal approximately 8 h before ovulation in both tissues of the second and third follicles. The content of oestradiol in the granulosa did not vary as follicles changed position within the hierarchy or during the ovulatory cycle. The oestradiol content of the theca tissue remained constant during the third and fourth positions in the hierarchy and declined throughout the second and first positions until a nadir was observed approximately 20 h before ovulation. It was concluded that the synthesis of androstenedione and oestradiol ceases in both follicular tissues after the follicle is exposed to the penultimate preovulatory surge of LH and that progesterone production is stimulated in the granulosa of the three largest follicles at the time of the preovulatory release of LH.     

Changes in the plasma concentrations of luteinizing hormone, progesterone, oestradiol and testosterone and in the binding of follicle-stimulating hormone to the theca of follicles during the ovulation cycle of the hen (Gallus domesticus)

The changes in the binding of FSH during follicular maturation were examined in the hen using 125I-labelled bovine FSH (bFSH) and unlabelled bFSH. The binding of 125I-labelled bFSH was not inhibited by bovine LH or chicken LH but was inhibited by extracts of chicken pituitary glands. The ovarian stroma, which contained both interstitial tissue and small follicles, bound the greatest amount of FSH. As the follicles progressed through the yolk-filled hierarchy of maturation, they bound decreasing amounts of FSH. In the two largest follicles of the hierarchy, there was a significant increase in the binding of FSH 12-16h before ovulation. There were two peaks in the concentrations of LH; a preovulatory peak occurred 4-6h before ovulation and a second peak occurred 14-16h before ovulation. Plasma concentrations of testosterone, oestradiol and progesterone began to rise 9, 8 and 6h, respectively, before ovulation. These data are consistent with the hypothesis that changes in the gonadotrophin concentration and binding regulate the order of the follicular hierarchy and the onset of preovulatory steroidogenesis in the hen.     

Effect of luteinizing hormone on respiration of the preovulatory cumulus oophorus of the rat.

The effect of luteinizing hormone (LH) on the respiration of isolated cumulus cell complexes (the oocyte surrounded by cumulus granulosa cells) obtained from immature Sprague-Dawley rats injected with 10 IU pregnant mare's serum gonadotropin on day 30 was investigated. The cell complexes were isolated from preovulatory follicles of rats killed at specific time intervals on the day preceding ovulation, i.e., on day 32. The samples were incubated in Eagle's tissue culture medium. Oxygen uptake was recorded either with the Cartesian diver technique or with a recently described microspectrophotometric technique using hemoglobin as an indicator. Exposure to exogenous bovine LH in vitro or in vivo or to endogenous LH, i.e., the preovulatory LH-surge, resulted in a marked decrease in respiratory activity of the cumulus cell complex, as revealed by both techniques. The cumuli exposed to LH showed an oxygen uptake of approximately 40-65% of the control cumuli. The results suggest that LH has a direct effect on this cell complex resulting in a decreased oxidative metabolism.     

Effects of stage of the cycle and estradiol-17 beta on oxytocin synthesis by ovine granulosa and luteal cells.

Individual ovine follicles or corpora lutea (CL) were obtained at different stages of the estrous cycle to compare the pattern of oxytocin synthesis with time in vitro. Granulosa cells from follicles in the early follicular phase produced minimal amounts of oxytocin whereas output from preovulatory (post LH surge) follicles increased to a peak of 540 pg/10(4) cells.24 h on days 4-7 in vitro declining to 180 pg/10(4) cells.24 h by day 11. Production from day 1 CL was also high, peaking at 1639 pg/10(4) cells.24 h. In contrast the capacity for oxytocin synthesis by day 2 CL had already declined, with peak output reaching only 185 pg/10(4) cells.24 h on days 3-4. Day 9 CL produced small amounts of oxytocin (50 pg/10(4) cells in the first 24 h) followed by a low output thereafter. The effect of estradiol-17 beta (E2 beta) on oxytocin synthesis was examined. The results were dependent on the stage of the cycle at which the cells were obtained. Oxytocin production was significantly stimulated in three and inhibited in four out of nine preovulatory follicles by the addition of 50 or 500 ng/ml E2 beta, whereas in days 1 and 2 CL E2 beta consistently inhibited oxytocin synthesis and in day 9 CL no response was found. These data indicate that the ovarian capacity to synthesize oxytocin varies markedly at different stages of the cycle, and that cells obtained close to ovulation do not experience the rapid down-regulation in oxytocin synthesis which occurs in vivo in the early luteal phase. E2 beta may switch from having a stimulatory to an inhibitory action on oxytocin synthesis shortly before ovulation.     

Induction of prostaglandin H synthase in rat preovulatory follicles by gonadotropin-releasing hormone.

Two distinct isoforms of prostaglandin (PG) endoperoxide synthase (PGS) have been identified in rat ovarian tissues: rPGSi (mol wt, 70,000-72,000) is induced by FSH and LH in preovulatory follicles, whereas the other isoform (mol wt, 69,000) is not. Induction of rPGSi is associated with LH-stimulated increases in PG biosynthesis obligatory for ovulation. Because GnRH, like LH, can also stimulate the synthesis of PGs and ovulation in the rat, this study was undertaken to determine which isoform of PGS might be induced by GnRH, in what cell type, and by what intracellular pathways. Results show that GnRH at relatively low concentrations (10(-8)-10(-7) M) induced the same isoform of PGS (rPGSi) in the same cell type (preovulatory granulosa cells) and within the same 5- to 7-h time course as did LH. Unlike LH and FSH, GnRH did not cause a major increase in cAMP, nor did GnRH induce luteinization. The effects of GnRH on rPGSi in preovulatory follicles were not mimicked by known activators of protein kinase-C (phorbol myristate acetate, bryostatin, diacyglycerol, and (+/-)ionomycin). Epidermal growth factor (but not basic fibroblast growth factor or platelet-derived growth factor), which activates a receptor-associated tyrosine kinase, caused a small increase in rPGSi. Genistein, a selective inhibitor of tyrosine kinases, blocked GnRH and LH induction of rPGSi. Taken together these results suggest that the mechanisms by which GnRH and LH selectively induce rPGSi in granulosa cells of preovulatory follicles before ovulation may converge at some step within a cellular tyrosine kinase cascade. Furthermore, the mechanisms responsible for inducing rPGSi are distinct from those required for cellular luteinization.     

Plasma gonadotropin concentrations in the cyclic female brushtail possum (Trichosurus vulpecula).

Changes in plasma concentrations of follicle-stimulating hormone (FSH) and luteinizing hormone (LH), and their relationship to antral follicle development and ovulation, were determined in female brushtail possums (Trichosurus vulpecula) in experiments in which pouch young were removed (RPY) from lactating females to promote ovarian activity. In Experiment 1 (n = 8), the development of preovulatory follicles and ovulation was monitored by laparoscopy. In Experiment 2 (n = 15) estrus and mating were monitored by cytology of urine. Ovulation occurred in 4/8 (Experiment 1) and 9/16 (Experiment 2) possums, and in these animals, plasma FSH concentrations fell progressively over the period of preovulatory follicle development and returned to pretreatment levels after ovulation. With the exception of samples taken at the time of the preovulatory gonadotropin surge, mean plasma LH levels remained basal. In those possums that failed to ovulate, plasma FSH concentrations were elevated while plasma LH concentrations were low; these patterns remained unchanged throughout the sampling period. It was not possible to distinguish between animals that would ovulate and those that would not ovulate after RPY on the basis of gonadotropin profiles at the time of RPY. A further group of possums (Experiment 3, n = 10) were blood-sampled at hourly intervals for 48 h to characterize preovulatory gonadotropin surges, using laparoscopy to monitor preovulatory follicular development and predict ovulation. A preovulatory LH surge (max. conc. 10.2-43.5 ng/ml, duration 7-9 h) was recorded in 4 animals, with a coincident preovulatory FSH surge (max. conc. 1.4-21.4 ng/ml, duration 3-11 h) observed in 3 of these possums. The patterns of gonadotropin secretion in the cycling brushtail possum conform to those reported for eutherians that ovulate spontaneously and appear to be regulated by similar mechanisms.     

Hormone concentrations and ovulatory response in rats treated with antiprogestagens

Administration of antiprogestagens (2 mg/day) to female rats for 21 days induces high serum prolactin levels. These levels stimulate luteal progesterone production and an increase in ovarian weight. Compared with RU486 (mifepristone) the increase in prolactin is less after treatment with ZK299 (onapristone), an antiprogestagen with lower antiglucocorticoid activity. To study whether cyclic ovulations occur in rats treated with antiprogestagens, 5-day cyclic rats were given daily injections of RU486 or ZK299 (2 mg) from metoestrus (day 1) to pro-oestrus. This treatment advanced the forthcoming ovulation by 1 day; however, the ovulation rate was low. Injection of 10 IU human chorionic gonadotrophin on the afternoon of pro-oestrus (day 3) increased the ovulation rate, but not to the level found in oil-treated rats. Serum LH concentrations measured from metoestrus to oestrus at 10.00 and 17.00 h were higher in antiprogestagen- than in oil-treated rats from day 2 (17.00 h) onwards. A low preovulatory LH surge was found in antiprogestagen-treated rats on the afternoon of pro-oestrus (day 3). Ovarian histology at the day of oestrus (day 4) confirmed the presence of a low LH surge as, besides ruptured follicles, unruptured follicles with dispersion of cumulus cells were present. The pro-oestrus surge of prolactin was also advanced by 24 h. The magnitude, however, was not different from that in oil-treated rats at day 4.(ABSTRACT TRUNCATED AT 250 WORDS)