The in vitro effect of 1 alpha,25-dihydroxyvitamin D3 on the function of beta cells of the endocrine pancreas was investigated. Neonatal islets maintained in serum-free medium, or medium supplemented with 0.5% fetal bovine serum achieved a 2.5-fold increase in medium insulin levels in response to 10(8) M 1 alpha,25-dihydroxyvitamin D3 (P less than 0.001). The effect of 1,25-dihydroxyvitamin D3 required at least 96 h treatment to become evident and was similar at medium glucose concentrations of 10 and 20 mM. Cell-associated insulin was increased in 1 alpha,25-dihydroxyvitamin D3-treated cultures maintained in 0.5% serum. These data suggest that 1 alpha,25-dihydroxyvitamin D3 may have a direct effect in the beta cell.