We previously identified a receptor protein for 1,25-dihydroxyvitamin D3 in rat liver nuclei. The present studies were undertaken to investigate the ontogenesis of the hepatic nuclear vitamin D receptor (nVDR) and the estrogen regulation of this receptor in the liver, small intestine, and kidneys. The hepatic nVDR was significantly elevated in adult female rats compared to prepubertal female rats, while in male rats, this increase was not observed. Oophorectomized rats contained significantly less hepatic nVDR than did intact female rats. Administration of estradiol to castrated male or oophorectomized rats increased the hepatic nVDR. Further studies demonstrated that the increase in the hepatic nVDR was observed only after 2 weeks of estradiol treatment and was positively correlated with circulating estradiol concentrations. Castration of male rats did not alter the hepatic nVDR compared to intact male rats nor did testosterone administration to castrated male rats for 4 weeks change the hepatic nVDR concentration. Unlike the liver, intact female rats contained significantly less renal nVDR than did kidneys from intact male or castrated male rats. Estradiol administration to oophorectomized rats significantly decreased the renal nVDR. Renal nVDR concentrations correlated inversely with the serum concentration of estradiol. Castration of male rats had no effect on the renal nVDR. Intestinal nVDR concentrations were unaffected by castration of male rats or by treatment of castrated male rats with estrogen for up to 4 weeks. These results indicate that estradiol increases the nVDR in liver, decreases the nVDR in kidney and does not change the nVDR in the intestine. Physiological concentrations of testosterone do not regulate the nVDR in these tissues. Estradiol regulation of this receptor is organ specific and, therefore, conclusions about the regulation of the nVDR in one tissue cannot be extrapolated to other tissues.