Effects of oxytocin on steroidogenesis by bovine theca and granulosa cells

Oxytocin (OT) is secreted during the final stages of bovine follicular development. To test OT's potential role as a regulator of follicular steroidogenesis, theca and granulosa cells were isolated from bovine preovulatory follicles 48 h after initiation of luteolysis with prostaglandin F2 alpha, and cultured with graded doses of OT (0, 0.5, 5, 50, and 500 mIU/ml). Granulosa cells were cultured with testosterone (0.5 microM) in either defined medium or medium containing 10% fetal bovine serum in the presence or absence of FSH (300 ng/ml); medium was collected and replaced daily for 5 days. In defined medium, oxytocin alone significantly increased progesterone production by granulosa cells (P less than 0.001) in a dose-dependent manner; over 5 days, doses of 0.5, 5, 50, and 500 mIU/ml OT caused 1.7-, 2.0-, 2.2-, and 2.6-fold increases. FSH enhanced progesterone 5-fold, but no dose of OT increased progesterone in the presence of FSH. OT also elevated progesterone in serum-containing medium (P less than 0.005), but the magnitude of its effects was lower (1.07-, 1.1-, 1.2-, and 1.4-fold increases with 0.5, 5, 50, and 500 mIU/ml OT). OT had little effect on estradiol secretion by granulosa cells cultured with or without FSH. To test the specificity of OT's effects on progesterone production by granulosa cells, granulosa cells were treated with graded doses of an OT antagonist (0, 1, 10, 100, and 1000 ng/ml) in the presence or absence of OT (5 and 50 mIU/ml). Progesterone production by granulosa cells in the presence of the antagonist alone was similar to production in control cultures. The stimulatory effects of 5 and 50 mIU OT were completely abolished in the presence of 100 or 1000 ng antagonist, respectively (P less than 0.01). Preparations of theca interna were cultured in defined medium with graded doses of OT (0, 0.5, 5, 50, and 500 mIU/ml) in the presence or absence of LH (300 ng/ml), with collection and replacement of medium at 3, 6, 12, 24, 48, and 72 h. LH alone increased both progesterone (12-fold) and androstenedione (4-fold) production over controls. However, no dose of OT significantly affected either progesterone or androstenedione production. These results show that OT stimulates progesterone production by granulosa cells, and thus, suggest that OT regulates steroidogenesis in bovine granulosa cells in vivo.