Expression and regulation of mRNA for inhibin/activin alpha- and beta(A)-subunits in the granulosa layer of the two largest preovulatory follicles during the hen ovulatory cycle

In order to understand the role of inhibin and activin in regulating follicular development in the hen, the steady-state mRNA levels of inhibin/activin alpha- and beta(A)-subunits in the granulosa layer of the largest (F-1) and second largest (F-2) follicles of the hen were investigated at 4-hr intervals throughout the ovulatory cycle. In addition, because it was hypothesized that luteinizing hormone (LH) regulated beta(A)-subunit expression, the effect of in vivo administration of ovine LH (oLH) on the expression of these subunits during the early-and mid-ovulatory cycle was examined. Northern blot analysis, using P-32-labeled cDNA probes of chicken inhibin/activin alpha- and beta(A)-subunits and glyceraldehyde-3-phosphate dehydrogenase (GAPDH, internal control), revealed that in the F-1 follicle, the relative level of beta(A)-mRNA (n = 3) was low at 23.5 hr and increased (P < 0.05) at 19.5, 15.5, and 11.5 hr before the next predicted ovulation. It then decreased (P < 0.05) at 7.5 hr and was further reduced at 3.5 and 0.5 hr prior to ovulation. In the F-2 follicle, beta(A)-mRNA was maintained at a basal level throughout the sampling period except for a brief increase (P < 0.05) at 0.5 hr before ovulation. In contrast to the beta(A)-subunit, inhibin alpha-mRNA was abundantly expressed with no significant variations throughout the ovulatory cycle in either the F-1 or the F-2 follicle. When oLH was injected at 18 hr before ovulation, 200 but not 100 or 50 mu g/kg (n = 3 hens per dose) significantly (P < 0.05) reduced the beta(A)-mRNA level in the F-1 follicle by 2 hr after injection compared to the control (saline). The experiment was repeated at 12 hr before ovulation and both 100 and 200 but not 50 mu g/kg oLH significantly (P < 0.05) reduced the expression of beta(A)-subunit mRNA with no significant difference between 100 and 200 mu g/kg oLH. In contrast to the beta(A)-subunit, inhibin alpha-subunit mRNA was abundantly expressed and not affected by oLH treatment. Our data indicate that the expression of inhibin/activin beta(A)-but not alpha-subunit mRNA is developmentally regulated in the granulosa layer of the two largest follicles during the hen ovulatory cycle. In addition, LH may participate, directly or indirectly in negative regulation of the beta A-subunit. (C) 1997 Academic Press.