This study investigated the role of endogenous inhibin in regulating FSHβ mRNA levels subsequent to the gonadotropin surge in the immature, estradiol (E2)-treated female rat. Rats which undergo FSH surges on day 29 have low to undetectable levels of FSH3 mRNA at 0900 h on day 30, whereas those treated simultaneously with E2 and progesterone (P) implants to block these surges have considerably higher levels of FSHg mRNA. In view of the profound inhibitory effect of inhibin on FSHβ mRNA, we examined the possibility that increased inhibin secretion is responsible for the decline in FSHβ mRNA levels on the morning after the FSH surge by immunoneutralization of endogenous inhibin. Twenty-eight day-old rats which received E2 and blank (Bl) or P implants were injected iv with 0.4 ml of a potent anti-rat inhibin serum (anti lα, prepared in sheep against rat inhibin α(l–26)-Tyr27 coupled to human a-globulins) or normal sheep serum at 1700 to 1830 h on dav 29 and were killed at 0900 h on day 30. Animals which received the inhibin antiserum showed significantly (P< 0.001) elevated serum FSH levels (22.9 ± 1.9 ng/ml [E2 + Bl] and 17.1 + 0.6 ng/ml [E2 + P]) compared to those which received normal serum (4.4 + 0.1 [E2 + Blj and 4.2 ± 0.1 [E2 + PJ). Serum LH was undetectable (P2 + Bl-treated rats but was significantly suppressed by P after injection of either normal serum or anti la. Total pituitary RNA was extracted and hybridized to cDNA probes for rat FSHβ, LHβ, and the common a-subunit by Northern blot analysis; RNA levels were normalized with β-actin or cyclophilin probes. As expected, in rats which received normal serum, FSHβ mRNA levels were about 4-fold higher after treatment with E2 + P implants than after treatment with E2 + Bl implants. However, injection with anti-inhibin serum resulted in a striking elevation of FSHβ mRNA levels: 13-fold in animals treated with E2 + Bl implants and 5-fold in animals treated with E2 + P implants. There were no significant differences in levels of LHβ or α-subunit mRNAs between rats which received anti-inhibin or normal serum although there was a 30-40% decrease in α mRNA after P treatment. These results, taken together with our previous results showing higher serum inhibin concentrations in rats undergoing FSH surges than in those in which the surge is blocked by P, indicate that endogenous inhibin regulates FSHβ mRNA levels in the rat. In addition, progesterone suppresses both secretion of free α-subunit and α mRNA levels. At present these conclusions are limited to the immature female rat; further studies will be necessary to determine if they generalize to the adult female and/or male rat.