The effects of a conceptus-derived substance on the activity of 3β-hydroxysteroid dehydrogenase (3β-HSD) and 20α-HSD in the ovary were studied in the rat. On day 7 of pregnancy (day 1 = insemination), rats were laparotomized and the desired number of conceptuses was aspirated from the uterus; thus, rats carrying one, two, three, four, five to seven or eight to ten conceptuses were prepared. They were autopsied on day 15 and 3β-HSD and 20α-HSD activity in the corpus luteum (CL) or non-luteal ovarian tissue (NLO) was determined. Conceptus number was directly related to 3β-HSD and inversely related to 20α-HSD activity in the CL. The serum progesterone level and CL weight were also directly related to conceptus number. Neither 3β-HSD nor 20α-HSD activity in the NLO was affected by conceptus number. These results indicated that 3β-HSD and 20α-HSD in the CL are probably regulated by placental hormone secreted in proportion to the number of conceptuses; in the NLO these enzymes may be controlled by a different mechanism.