In the pregnant mare, luteal estrogen production increases at the onset of equine chorionic gonadotropin (eCG) secretion by endometrial cups. In previous studies, we have demonstrated that eCG stimulates luteal androgen and estrogen production in pregnant mares. To further elucidate the regulation of steroidogenesis within the equine corpus luteum (CL) of pregnancy, we examined the expression of 3 beta -hydroxysteroid dehydrogenase (3 beta -HSD), cytochrome P450 17 alpha -hydroxylase/17,20 lyase (P450(17 alpha)) and cytochrome P450 aromatase (P450(arom)) in luteal tissue samples collected during diestrus (Days 7 to 10) and pregnancy before (Days 29 to 35) and after (Days 42 to 45) the onset of eCG secretion. Immunoblot analyses revealed a single protein per enzyme with molecular weights of 48 kDa (3 beta -HSD), 58 kDa (P450(17 alpha)) and 56 kDa (P450(arom)). Steady-state levels of 3 beta -HSD were lower in luteal tissue of diestrus than pregnancy, but expression did not change during pregnancy. Steady-state expression of P450(17 alpha) in CL of diestrus was not significantly different from that of pregnancy. During pregnancy, P450(17 alpha) expression was significantly higher after the onset of eCG secretion. Steady-state expression of P450(arom) in CL of diestrus was not significantly different from that of pregnancy. During pregnancy, luteal expression of P450(arom) was significantly lower after the onset of eCG secretion. These data support the hypotheses that eCG has a differential effect on the expression of luteal steroidogenic enzymes, that the eCG-induced increase in luteal estrogen production is the result of an increase in available aromatizable androgen due to an increase in P450(17 alpha) expression and activity, and that increased luteal estrogen production is not due to an increase in aromatase expression. (C) 2001 by Elsevier Science Inc.