The ontogenesis of the 1,25-dihydroxyvitamin D3 specific binding activity in intestine was examined in vitamin D-deficient and replete rats. The absence of binding activity in intestines during the first two postnatal weeks was not influenced by vitamin D supplementation. The concentration of binding sites peaked on day 18 in vitamin D-replete rats and preceded that in the deficient group by approximately 1 wk. The influence of glucocorticoids on 1,25-dihydroxyvitamin D3-binding protein levels was examined by sequential hydrocortisone administration and adrenalectomy. Subcutaneous hydrocortisone administration before day 14 postpartum did not induce binding activity. The concentration of binding sites was significantly increased to 369 +/- 60 fmol/mg of protein by hydrocortisone injections from days 15 to 17 postpartum when compared with an average of 182 +/- 16 fmol/mg of protein in littermate controls. Hydrocortisone administration did not further increase receptor levels in rats injected from days 19 to 21. Bilateral adrenalectomy on day 17 postpartum significantly decreased the concentration of binding sites. It is concluded that adrenal glucocorticoids play an important role in the developmental appearance of 1,25-dihydroxyvitamin D3 specific binding activity in the postnatal rat intestine.