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Summary. Inhibin activity was measured by bioassay in follicular fluid of 99 individual ovine follicles ranging from 1\m=.\4to 6\m=.\8mm diameter (used to calculate volume) and in various stages of atresia. Treatment of samples before assay with charcoal concentrations of > 1 mg/ml resulted in significant loss of inhibin activity.The inhibin content of follicular fluid from individual follicles varied with follicular fluid volume but not with the degree of atresia, as assessed by morphological criteria.Inhibin concentration was not related to atresia, but was correlated with follicular fluid volume. However, aromatase activity in granulosa cells and oestradiol-17\g=b\ concentration of follicular fluid, considered to be good indices of atresia, were highly correlated with both inhibin content and concentration in follicles \ m=ge\ 3\m=.\5mm diameter.Inhibin in ovine follicular fluid shows marked variation between follicles and it is suggested that this reflects a combination of the number and activity of granulosa cells within the follicle and the exit rate of inhibin from the follicle.

The effects of PMSG treatment on ovarian and circulating inhibin concentrations in immature female rats has been examined. Sixty-four hours after injection of 10, 20 or 40 IU of PMSG the animals were anesthetized with ether; ovaries, uteri and blood from the abdominal aorta were collected. Steroid-free extracts of ovary and serum samples were prepared and assayed quantitatively for inhibin activity in an in vitro bioassay system. PMSG treatment elevated (p less than 0.001) both uterine and ovarian wt, and ovarian and peripheral concentrations of inhibin. A dose-related increase occurred ovarian wt, and in peripheral and ovarian content of inhibin. Ovarian inhibin concentration increased with dose of PMSG until the highest dose, where a significant decline and luteinization were seen. Peripheral FSH levels were significantly lowered at all doses of PMSG treatment; in contrasts, LH was significantly elevated, due to cross-reaction of PMSG in the LH assay. These results show that both ovarian and circulating levels of inhibin are related to the degree of gonadotropic stimulation, supporting the view that inhibin is involved in folliculogenesis and in the feedback regulation of FSH.

Two forms of inhibin with molecular weights of 65,000 and 30,000 (65 and 30 kD) were isolated from ovine follicular fluid using a combination of gel permeation chromatography, reversed-phase high-performance liquid chromatography and preparative polyacrylamide gel electrophoresis. The 65 kD form was partially purified approximately 315-fold whilst the 30 kD form was isolated as two isoforms (29 and 30 kD) of similar biological activity and in greater than 95% purity (1210-fold purification and 4.2% recoveries). On reduction the 30 kD form resolved into four components of 36, 31, 20-21 and 16 kD of which the 20-21 and 16 kD components were similar to the corresponding inhibin subunits isolated from porcine and bovine follicular fluid. The 36 kD component was established as a non-reducible inhibin-like material, based on its binding to antiserum raised against bovine 58 kD inhibin. The nature of the remaining non-reducible 31 kD component is unknown. Two NH2-terminal amino acid sequences (first 13 amino acids) identified in purified 30 kD inhibin were identical to the corresponding subunit amino acid sequences of bovine 31 kD inhibin.

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