Recent investigations have indicated that luteal cells of pregnant rats become capable of synthesizing androgen and estradiol when stimulated by sustained low levels of LH/hCG. In this investigation we sought 1) to determine whether hCG causes the induction/activation of the luteal enzymes responsible for the conversion of progesterone to estradiol, 2) to investigate the time course of hCG action, and 3) to compare the effect of hCG on luteal and follicular P45017 alpha activities. To determine first the minimum stimulatory dose of hCG, pregnant rats were treated with 0, 1.5, 3, 9, or 30 IU hCG twice on days 12 and 13 and once on day 14. Two hours after the last injection, rats were bled from the ovarian vein, and corpora lutea were isolated and incubated for the determination of in vitro steroid production. Exposure of rats to twice daily injections of 1.5 IU hCG caused a dramatic increase in the ovarian secretion and luteal production of both testosterone and estradiol. However, an inverse dose-related response was observed with higher doses of hCG. To determine the time course of hCG action, pregnant rats were injected with a single dose of 3 IU hCG, and steroid production was determined at different times thereafter. There was no increase in either in vivo or in vitro testosterone and estradiol production until 12 h after hCG administration, at which time a dramatic increase in the secretion of both steroids was observed. hCG administration did not affect the capacity of luteal cells to secrete progesterone, nor did it enhance aromatase activity. It did, however, increase P45017 alpha activities; lyase and hydroxylase activities were stimulated 5- and 1.7-fold, respectively. To compare the effects of hCG on luteal and follicular P45017 alpha, both corpora lutea and follicles were isolated from ovaries of pregnant rats treated with either 1.5 IU hCG or vehicle between days 12-14. In vivo hCG administration caused at least a 1000-fold increase in the specific activity of follicular 17 alpha-hydroxylase and 17,20-lyase. The hCG-induced increase in the specific activities of both hydroxylase and lyase in the follicle far exceeded that in the corpora lutea. However, total lyase and hydroxylase activities in each follicle were only 3- and 5.5-fold higher, respectively, than those in the corpus luteum. In summary, the results of the present investigation have revealed that hCG stimulation of luteal estradiol production is due to a selective effect of this gonadotropin on P45017 alpha.(ABSTRACT TRUNCATED AT 400 WORDS)