Alterations in intrafollicular levels of different molecular mass forms of inhibin during development of follicular- and luteal-phase dominant follicles in heifers

The experiment reported here characterized changes in serum concentrations of FSH, follicular fluid (FF) concentrations of immunoreactive alpha inhibins and dimeric inhibins, and different molecular forms of inhibin during development of dominant follicles in both luteal and follicular phases of the estrous cycle in heifers. Follicular status was determined daily by ultrasound examination of ovaries, and blood samples were taken at 4-6-h intervals. Heifers (5-6 per treatment) were ovariectomized on Days 0 (follicular phase, pre-LH surge), 1 - Ov (post-LH surge, preovulation), 1 + Ov (postovulation), 3 (selection phase), 6 (dominance phase), and 12 (atretic phase) of the estrous cycle. Follicles were classified on the basis of FF estradiol (E) to progesterone (Pi ratio and by ultrasound morphology. Concentrations of E, P, immunoreactive alpha inhibin, LH, and FSH were determined by RIA, and concentrations of dimeric inhibins were determined by a two-site dimeric-specific immunoradiometric assay (IRMA); the different molecular mass forms of inhibin were determined by immunoblot analysis. Serum FSH was transiently elevated during Days 0.5-1.5 and 8-10.5, in association with emergence of each new wave of follicular growth. The follicular-phase dominant follicle of heifers before the preovulatory LH/FSH surge had less immunoreactive alpha inhibin and 29-kDa inhibin hut more 110-kDa inhibin compared with concentrations after the surge (p < 0.05). During growth of the dominant follicle between Days 3 and 6, there were no changes in either proportions or amounts of any inhibin forms (p > 0.05). However, while concentrations of dimeric inhibins decreased between Days 1 and 6, the concentration of immunoreactive alpha inhibin increased (p < 0.05). Atresia of the luteal-phase dominant follicle during Days 6-12 was associated with a decrease in amounts and proportions of the 110- and > 160-kDa forms, an increase in amounts and proportions of the 34-kDa form, and an increase in dimeric inhibin concentration (p < 0.05). Alterations in FF E concentrations were positively correlated with changes in amounts of the 77- and 110-kDa precursor forms of inhibin, but negatively correlated with 34-kDa inhibin. The results of this experiment demonstrate that 1) seven different molecular mass forms of inhibin were present in bovine FF during all stages of development of follicular- and luteal-phase dominant follicles examined, 2) amounts and proportions of most inhibin forms are altered differently in follicular- and luteal-phase dominant follicles, 3) FF inhibins and E are regulated by different factors, and 4) RIA, IRMA, and immunoblot measurements of inhibin in bovine FF produced divergent results.