Abstract A single injection of estradiol benzoate (10 mug), while highly effective in ovariectomized female guinea-pigs, does not prime castrated males to display progesterone-facilitated lordosis. In contrast, adult males and females exhibit the same, high degree of progesterone-facilitated lordosis when primed with two, 2.0 mug injections of free estradiol-17ss (pulse regimen). We compared neural progestin receptor induction after these different estradiol treatments by in vitro radioligand binding assays and immunocyto-chemistry. Binding assays confirmed previous observations of lower concentrations of cytosol progestin receptors in the mediobasal hypothalamus-preoptic area in estradiol benzoate-treated males than in females. No such sex difference was observed in animals that had been exposed to the behaviorally effective estradiol pulse regimen; rather, hypothalamic-preoptic area progestin receptor concentrations in these animals did not differ from the low levels observed in males treated with the behaviorally ineffective estradiol benzoate regimen. Immunocytochemical analysis revealed progestin receptor-immunoreactivity in fewer cells in the medial preoptic nucleus-anterior hypothalamic nucleus, periventricular preoptic area and arcuate nucleus of estradiol pulse- as compared to estradiol benzoate-treated males and females. Estradiol benzoate treatment induced progestin receptor-immunoreactivity in more cells in the medial preoptic area and ventrolateral hypothalamus than estradiol pulses in males, but not in females. Surprisingly, in these regions estradiol benzoate-treated males had significantly more progestin receptor-immunoreactive cells than females. These experiments yield two major findings: First, as has been shown in rats, the display of progesterone-facilitated lordosis is not inflexibly differentiated according to sex in guinea-pigs. Furthermore, reduced concentrations of estradiol-induced progestin receptors in the hypothalamus and preoptic area cannot account for the lack of progesterone-facilitated lordosis that is observed following priming with estradiol benzoate in males. Secondly, in the medial preoptic area and ventrolateral hypothalamus of female guinea-pigs, estradiol pulses are as effective as estradiol benzoate in inducing progestin receptors. These observations, taken together with the finding of equal behavioral efficacy of the two estradiol treatments, are consistent with the hypothesis that estradiol-induced progestin receptors in these regions of the brain are involved in progesterone-facilitated lordosis in female guinea-pigs.