Follicle-stimulating hormone-induced estradiol and progesterone production by bovine antral and mural granulosa cells cultured in vitro in a completely defined medium.

Abstract

Functional subpopulations of granulosa cells exist in bovine follicles. This study was designed to compare the in vitro steroid production by cultured bovine antral and mural granulosa cells in response to various amounts of FSH. Antral and mural granulosa cells (600,000 viable cells/well) harvested from ovaries of PMSG-treated prepuberal calves were cultured in serum-free conditions for 4 d in wells containing 1 mL of defined Ham's F-12 medium, supplemented with 0, 2, or 10 ng/mL of FSH. Culture medium was collected and replaced each day. The mean concentration of estradiol in culture media of bovine granulosa cells decreased from d 1 to d 4 (P < .001). Granulosa cell production of estradiol increased in antral cells following addition of 2 ng/mL FSH (P < .001) but decreased following addition of 10 ng/mL FSH (P < .001) as determined on d 4 by RIA and thin layer chromatography. In contrast, there was no response to FSH stimulation in mural granulosa cells. Progesterone production increased (P < .01) in a dose-dependent manner following stimulation with 2 or 10 ng/mL FSH and was consistently higher (P < .001) in antral than in mural granulosa cells. Addition of LH on d 4 stimulated estradiol and progesterone production in antral (P < .01) but not in mural cells (P > .10). This suggests that FSH- and LH-induced estradiol and progesterone productions are considerably lower in mural than in antral bovine granulosa cells. This suggests that functional differences between these two cell compartments need to be considered in studies involving in vitro cultures of bovine granulosa cells.