Identification of Inhibin-Like Activity in Ovarian Venous Plasma of Rats during the Estrous Cycle

Abstract

These studies determined whether ovarian venous plasma (OVP) contains nonsteroidal material which has FSH-inhibiting activity (FSH-IA) and, if so, whether such activity varies inversely with plasma FSH concentrations during the 4-day rat estrous cycle. Ovarian venous blood was collected for a 10-min period from cyclic rats at 0900 and 1600 h on diestrous days 1 and 2 (Dl and D2); on proestrus (Pr) at 0900, 1200, 1400, 1600, and 1800 h; and on estrus (E) at 0900, 1200, and 1600 h. A peripheral plasma sample from the abdominal aorta was taken after the ovarian vein collections. Peripheral and ovarian venous plasma were radioimmunoassayed for concentrations of 17β- estradiol (E₂), progesterone (P), LH, and FSH. FSH-IA was assessed in terms of its ability to suppress the 24-h secretion of FSH in a dispersed pituitary cell culture system. The addition of untreated OVP obtained from rats at 0900 and 1600 h on Dl and D2 and at 0900, 1200, and 1400 h Pr produced a 43–56% suppression of 24-h FSH secretion by cultured pituitary cells. During this interval, peripheral plasma LH, FSH and P concentrations were basal but plasma E₂ increased during D2 to reach maximal concentrations by 0900 h on Pr. Concomitant with the preovulatory surge of LH and FSH (1600- 1800 h on Pr), FSH-IA declined in OVP and produced only 29% and 21% inhibition, respectively, in FSH secretion in vitro. Also during this interval, peripheral plasma E2 declined and P increased in concentration. At 0900 h on E, plasma FSH was elevated above Dl levels, but plasma LH and P were basal and FSH-IA had further declined to produce only a 9.5% inhibition of FSH secretion by cultured pituitary cells. By 1600 h on E, when FSH approached basal Dl peripheral concentrations, FSH-IA increased in OVP and produced a 39% inhibition of FSH secretion in vitro. In attempts to remove steroids, OVP was diluted with water and treated with charcoal. In one study, the water was removed by lyophilization and the OVP sample was reconstituted with medium to its original concentration. Such treatment removed all detectable FSH-IA from the OVP samples. Similar dilution of porcine follicular fluid before charcoal treatment also markedly reduced FSH-IA when this material was tested for its ability to suppress FSH secretion in vitro. The responsiveness of the cell culture system to E₂, P, and testosterone in the presence of plasma obtained from 2-h ovariectomized rats (OVX) was also evaluated. The addition of OVX rat plasma and E₂ (1 × 10^-8 M) to the culture medium did not significantly affect the 24-h secretion of LH or FSH. When E₂ (1 × 10^-8 M) was added with P (1 × 10^-7 M) and OVX plasma to the culture medium, an increase in FSH but not LH secretion occurred (28%). Similarly, the addition of a mixture of OVX plasma, E₂, P, and testosterone (1 × 10^-7 M) produced a further stimulation of FSH (41.8%) but not of LH secretion. OVP was extracted with diethyl ether (which removes 93% of the [³H]P added to OVX plasma) and the N2-dryed residue was redissolved in culture medium. Aliquots of this material, when added to the culture system, had no suppressive effects on FSH secretion. The different concentrations of E₂, P, LH, and FSH in OVP (due to peripheral plasma hormonal variations) which were added to the culture system do not account for OVP-induced changes in the secretion of FSH in vitro. Rather, these studies provide evidence that OVP contains FSH-IA which varies inversely with peripheral plasma FSH concentrations.