Inhibition of FSH augmented adenylate cyclase activity in porcine granulosa cells by ovarian protein.

We sought to evaluate the effects of a fraction of porcine follicular fluid, termed follicle regulatory protein (FRP), on FSH-induced adenylate cyclase activity in porcine granulosa cell membranes using Gpp(NH)p and forskolin as pharmacological probes of adenylate cyclase activity. Without FSH treatment, the addition of 100 micrograms/ml of the FRP fraction induced a significant decrease in Gpp(NH)p stimulated cyclase activity while maximal inhibition of cAMP formation was achieved with 1 mg/ml of FRP. Granulosa cells cultured with FSH reached a maximum in adenylate cyclase activity at 20 min which returned to baseline by 45 minutes. FRP induced a reduction in adenylate cyclase activity during this same interval of time. Adenylate cyclase activity of cells treated with FRP was unchanged in the presence of methyl-isobutal-xanthine. Further, when FRP was heated (56 degrees C, 45 min) or precipitated with 10% TCA, it was unable to inhibit adenylate cyclase. The 50% inhibitory dose (ID50) for FRP inhibition of adenylate cyclase activity in cells stimulated with Gpp(NH)p was 80 micrograms/ml and 500 micrograms/ml when granulosa cells were preincubated with FSH prior to Gpp(NH)p stimulation. The ID50 for the FRP inhibition of forskolin stimulated adenylate cyclase activity was 500 micrograms/ml. Gpp(NH)p stimulated adenylate cyclase activity was more sensitive than forskolin stimulated activity to inhibition by FRP. In conclusion, the data presented here demonstrate that a partially purified fraction of porcine follicular fluid inhibited FSH responsive adenylate cyclase activity in porcine granulosa cells.