A bolus iv injection of 150 IU purified human urinary FSH (hFSH) or saline was given to 21 normal women during 3 different luteal phases (early luteal phase, n = 7; midluteal, n = 8; late luteal, n = 6), and the responses of ovarian steroids and gonadotropins were analyzed sequentially for 24 h after hFSH injection. A rapid and dramatic increase in serum FSH concentrations occurred after each hFSH injection, followed by a gradual decrease. Serum LH concentrations, including hourly fluctuations, were significantly higher after FSH administration than after NaCl injection at the same time during the preceding cycle. Neither serum progesterone nor testosterone levels were affected by the hFSH injection in any of the 3 luteal phases, but in the early and midluteal phases serum estradiol levels were higher than the preinjection values and those during the control cycles. These results indicate that FSH stimulates aromatization of luteal tissue in vivo as it does in vitro. Thus, FSH in addition to LH as the indispensable luteotropin may have a permissive role in the regulation of corpus luteum function.