Oxytocin (OT) binding sites are modulated by estrogens in several brain regions including the ventromedial hypothalamic nucleus (VMN) in both male and female rats. To further study steroid regulation of OT receptor binding, we examined the effect of androgen replacement in castrated male rats on OT binding with quantitative autoradiographic methods. Castrated adult male rats were treated with either 250 micrograms testosterone propionate (TP) or oil for 2 days and killed 48 h after the last injection. Brain sections through the preoptic area and VMN were labeled with 5.0 nM[3H]-OT +/- 5.0 microM unlabeled OT or 1.0 microM[Thr4,Gly7]OT and apposed to tritium-sensitive film for 7 weeks. Results of this study show that TP increased [3H]-OT binding up to 5-fold in the ventrolateral VMN and 4-fold in the bed nucleus of the stria terminalis. In addition [Thr4,Gly7]OT completely displaced [3H]-OT binding in the VMN indicating that binding in this brain region was specific to OT receptors. Because estrogens also increase OT receptor binding in male rats, it is possible that TP affects OT binding after being converted by aromatase to estradiol.