The effects of progesterone and RU486, a synthetic anti-progesterone, on ovarian 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) activity, a key enzyme of progesterone production, were studied during ovulation in immature 22-day-old rats primed with pregnant mares' serum gonadotrophin (PMSG) and human chorionic gonadotrophin (hCG). Ovarian 3 beta-HSD activities had increased significantly 4 h after hCG injection. These increases were inhibited at 4 and 6 h after hCG when 20 mg RU486 kg-1 was administered 2 h before hCG. However, RU486 had no influence on the activity of 3 beta-HSD when administered at the same time as hCG injection. A histochemical study revealed that 3 beta-HSD activities in the granulosa cell layer, but not in the theca cell layer, were inhibited when RU486 was given 2 h before hCG. Serum progesterone concentrations, but not oestradiol concentrations, were significantly suppressed by RU486 treatment 4 and 6 h after hCG. The effect of progesterone on ovarian 3 beta-HSD activity was tested by administering graded doses of progesterone exogenously to rats 2 h before hCG injection. Ovarian 3 beta-HSD activity was increased in a dose-dependent manner, and more than 20 mg progesterone kg-1 significantly stimulated the activity. Although 10 mg progesterone kg-1 did not stimulate ovarian 3 beta-HSD activities, the RU486-inhibited activities were recovered by the concomitant administration of 10 mg progesterone kg-1 with RU486. These results indicate that ovarian 3 beta-HSD activity depends on progesterone concentrations, and suggest an autocrine regulation of progesterone production during ovulation in immature rat ovaries stimulated with PMSG and hCG.