Previous studies have provided evidence for a discrete localization of two types of vasopressin (AVP)-labeled binding sites in the rat brain, i.e., regions labeled preferentially with AVP (putative AVP receptors) and regions labeled with AVP as well as oxytocin (OT). The latter binding sites are considered here as putative OT receptors. In the present study the effect of estradiol on the number of these putative receptor sites for OT and AVP was investigated in rat brain after daily subcutaneous administration of the steroid (10 µg/100 g body weight) to ovariectomized rats. Specific binding of [3H]-OT and [3H]-AVP was determined after in vitro incubation of frozen brain sections, autoradiography and quantitation of the images with computer-assisted densitometry. Estradiol increased the number of OT receptors at least 4-fold in the ventromedial nucleus of the hypothalamus, regions of the olfactory tubercle, the nucleus accumbens and occasionally in the organum vasculosum laminae terminalis. A smaller increase (two-fold) was noted in the central amygdala, while a tendency to a decrease in OT receptor number was noted in the olfactory nucleus and the ventral subiculum. Estradiol treatment permitted an estimation of binding constants of [3H]-OT-binding to a membrane fraction of microdissected ventromedial hypothalamic region {Kd: 1.3 nM, Bmax: 19.9 fmol/mg protein). The number of putative AVP receptors in the lateral septum and in the nucleus tractus solitarii was not affected by estradiol. In conclusion, the OT receptor system is subject to modulation by estradiol in some discrete brain regions, but not in others. One of the principal sites of this interaction is located in the ventromedial nucleus, which is a hypothalamic region known to contain a high density of estradiol receptors and to respond to estrogens by changes in ultrastructure, protein synthesis, electrophysiology and sexual behavior. The results show that estradiol may sensitize these processes to OT and AVP by increasing the number of putative OT receptors.