To assess the role of estradiol (E2) upon progesterone (P4) synthesis, a well defined human midluteal cell system was used. A dose-dependent inhibition of P4 synthesis with and without hCG was induced by E2. In addition, E2 had a dose related cumulative effect on pregnenolone as compared with control experiments (2-fold, P < 0.05) as well as in hCG-stimulated conditions (3-fold, P < 0.005). On the other hand, the concentrations of 20 alpha-hydroxyprogesterone obtained in all experimental conditions were similar to control values, indicating that the catabolism of P4 was not modified. 3 beta-Hydroxysteroid dehydrogenase activity was significantly diminished (P < 0.05) in the presence of E2. Finally, the kinetic studies on P4 synthesis from pregnenolone showed a competitive type of inhibition with a K1 of 2.22 x 10(-6) mol/L. These data indicate an inhibition of 3 beta-hydroxysteroid dehydrogenase on human corpus luteum by E2.