Since male hamsters castrated as adults respond to exogenous estradiol-17β (E2) treatment by displaying female-like behavior, we wondered whether males, like females, would release gonadotropin following E2 treatment. To determine this, we performed 2 experiments. In experiment 1, blood samples from adult gonadectomized hamsters of both sexes were collected at 3 h intervals for 48 h starting the day after a midafternoon injection of estradiol benzoate (EB). Serum levels of E2, LH and FSH were quantified by radioimmunoassay. Males injected with either 50 or 200 µg of EB showed no LH or FSH surge at any time during the ensuing 2 days. Females, on the other hand, showed midafternoon peaks of LH on both days following the EB injection. In experiment 2, gonadectomized hamsters of both sexes were pretreated with E2-containing implants that elevated serum E2 levels to 170 ± 20 pg/ml (mean ± SEM). The injection of 50 µg EB 1 week later induced a midafternoon surge of both gonadotropins in females but not in males. These experiments show that in hamsters the response of CNS centers that control gonadotropin release to E2 are sexually dimorphic. Furthermore, they suggest that the CNS structures that mediate gonadotropin release in males are more sensitive to masculinizing effects of prenatal androgens than those that mediate sexual behavior.