Ken-ichi Ohshima

Dynamics of Messenger RNAs Encoding Inhibin/Activin Subunits and Follistatin in the Ovary During the Rat Estrous Cycle

Abstract Quantitative changes in ovarian inhibin/activin subunit and follistatin mRNAs during the rat estrous cycle were examined by ribonuclease protection assay using digoxygenin-labeled RNA probes. Levels of ovarian inhibin α subunit mRNA remained low throughout estrus, metestrus, and diestrus; abruptly increased on the morning of proestrus; then rapidly decreased when the primary gonadotropin surge occurred. A similar changing pattern was observed in inhibin/activin βA subunit mRNA. On the other hand, inhibin/activin βB subunit mRNA showed a different changing pattern. Levels of βB subunit mRNA remained constant during metestrus and diestrus, abruptly decreased on the afternoon of proestrus, then quickly recovered from the nadir by 1100 h on estrus. Throughout the rat estrous cycle, especially during the periovulatory period, α subunit mRNA levels were considerably higher than βA and βB subunit mRNA levels. In addition, changes in plasma concentrations of inhibin A and inhibin B were very similar to that in ovarian βA and βB subunit mRNA levels, respectively, with several-hour delays. These results suggest that levels of β subunit mRNAs restrict secretion of dimeric inhibins. Levels of follistatin mRNA remained low from the midnight of metestrus to the midnight of diestrus, then increased until initiation of the primary gonadotropin surge. Thereafter, follistatin mRNA decreased, reached the nadir at 0200 h on estrus, then increased abruptly at 1100 h on estrus. Afterward, follistatin mRNA levels remained high until the morning of metestrus. The changing pattern of ovarian follistatin mRNA was similar to, and preceded, the changes in plasma concentrations of progesterone, suggesting that ovarian follistatin may modulate progesterone secretion during the rat estrous cycle.

Changes in Expression of Inhibin Subunits in the Cyclic Golden Hamster (Mesocricetus auratus) and the Regulation of Inhibin α Subunit Production by Luteinizing Hormone

In the present study, changes in localization of each inhibin subunit in the ovary were investigated during the estrous cycle of the golden hamster. The effect of LH surge on changes in localization in inhibin α subunit in the ovary was also investigated. Inhibin α subunit was localized in granulosa cells of various stages of follicles throughout the estrous cycle. Inhibin α subunit was also present in numerous interstitial cells on days 1 and 2 (day 1 = day of ovulation), but the number of positive interstitial cells was fewer on days 3 and almost disappeared on day 4 of the estrous cycle. Newly formed luteal cells were also positive for inhibin α subunit on days 1 and 2. On the other hand, positive reactions for inhibin βA and βB subunits were only present in the granulosa cells of healthy antral follicles. However, a positive reaction for inhibin βB subunit in peripheral mural granulosa cells disappeared on days 3 and 4 of the estrous cycle. Treatment with LHRH-AS at 1100 h on day 4 completely blocked the luteinizing hormone (LH) surge and ovulation, although relatively high concentrations of plasma follicle-stimulating hormone (FSH) were maintained throughout the experiment. There were few positive reactions for inhibin α subunit in theca and interstitial cells 24 hr after LHRH-AS injection. The effect of LHRH-AS treatment was blocked by a single injection of 10 IU human chorionic gonadotropin. These results suggest that the major source of dimeric inhibin in the cyclic hamster was granulosa cells of healthy antral follicles. Different distribution pattern of inhibin βA from βB subunits in large antral follicles on days 3 and 4 of the estrous cycle suggests different secretion patterns of inhibin A from B on these days. Furthermore, the LH surge may be an important factor to induce production of inhibin α subunit in interstitial cells of the cyclic hamster.

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on serum inhibin concentrations and inhibin immunostaining during follicular development in female Sprague-Dawley rats

Intact and hypophysectomized immature rats were pretreated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; 0 or 32 microg/kg p.o.) and sacrificed throughout synchronized follicular development (0, 12, 24, 48, and 72 h after equine chorionic gonadotropin, eCG). TCDD administration to intact rats resulted in a premature elevation of serum FSH and LH by 12 h post-eCG. In intact rats pretreated with TCDD, the intensity of ovarian immunoreactivity for inhibin and the number of ovarian follicles staining for inhibin in midsaggital ovarian sections were decreased at the time of eCG administration (24 h post-TCDD) in comparison to controls. However, this decreased ovarian staining for inhibin was not associated with alterations in serum inhibin concentrations. Serum inhibin was suppressed in TCDD-treated rats when compared to intact controls only at 24 h post-eCG. Hypophysectomized animals exhibited no effect of TCDD on serum inhibin at any timepoint but did have decreased estradiol concentrations during follicular development. In summary, TCDD reduced serum concentrations of inhibin after the premature increases in FSH and LH suggesting that inhibin is not important in the initial elevation of FSH following exposure to TCDD.

Regulations of gonadotropin secretion by circulating inhibin, estradiol, and progesterone in cyclic hamsters

The physiological importance of gonadal hormones in feedback control of gonadotropin secretion during the estrous cycle in golden hamsters was investigated with immunoneutralization methods. Anti-inhibin serum (inhibin-AS) treatment always induced a drastic increase in follicle-stimulating hormone (FSH) secretion and occasionally raised luteinizing hormone (LH) secretion. Anti-estradiol-17β serum (estradiol-AS) treatment increased LH secretion typically. Although estradiol-AS elevated FSH secretion occasionally, the elevation was much less than that by inhibin-AS. Plasma FSH reached ovariectomized levels by a synergistic effect of both antisera. Elevated plasma LH with both antisera was much less pronounced than in ovariectomized animals. Plasma LH increased dramatically to the levels in the ovariectomized group when antibody against progesterone (progesterone-AB) was given together with inhibin-AS and estradiol-AS, although progesterone-monoclonal antibody alone did not alter plasma gonadotropin levels. These results indicate that in hamsters FSH secretion is mainly regulated by inhibin and LH secretion is regulated by estradiol-17β and progesterone during the estrous cycle.The physiological importance of gonadal hormones in feedback control of gonadotropin secretion during the estrous cycle in golden hamsters was investigated with immunoneutralization methods. Anti-inhibin serum (inhibin-AS) treatment always induced a drastic increase in follicle-stimulating hormone (FSH) secretion and occasionally raised luteinizing hormone (LH) secretion. Anti-estradiol-17beta serum (estradiol-AS) treatment increased LH secretion typically. Although estradiol-AS elevated FSH secretion occasionally, the elevation was much less than that by inhibin-AS. Plasma FSH reached ovariectomized levels by a synergistic effect of both antisera. Elevated plasma LH with both antisera was much less pronounced than in ovariectomized animals. Plasma LH increased dramatically to the levels in the ovariectomized group when antibody against progesterone (progesterone-AB) was given together with inhibin-AS and estradiol-AS, although progesterone-monoclonal antibody alone did not alter plasma gonadotropin levels. These results indicate that in hamsters FSH secretion is mainly regulated by inhibin and LH secretion is regulated by estradiol-17beta and progesterone during the estrous cycle.