Replacing body stores of vitamin D in pregnant rats with radiolabelled cholecalciferol, enabled the measurement of cholecalciferol and its hydroxylated metabolites in fetal tissue. Elevated levels of 24, 25-dihydroxycholecalciferol were found to be present in fetuses, with highest accumulation in the skeleton. A similar finding was observed when tritiated 24,25-dihydroxycholecalciferol was administered continuously to pregnant rats. When tritiated 1,25-dihydroxycholecalciferol was administered, very little was transported into the fetuses, and out of the transported fraction a major portion was found to be esterified. A selectivity pattern was established for the lacteal transport of cholecalciferol and its hydroxylated metabolites, in the order: cholecalciferol > 25-hydroxycholecalciferol > 24,25-dihydroxycholecalciferol > 1,25-dihydroxycholecalciferol. Vitamin D sulfoconjugates were not detected in suckling rat pups, and over 80% of the lacteal-transported 1,25-dihydroxycholecalciferol in suckling pups was found to be esterified. It is suggested that rat fetuses and newborn pups do not require 1,25-dihydroxycholecalciferol, that a protective mechanism against vitamin D intoxication operates in fetuses and pups in the form of esterifying enzymes, and that 24,25-dihydroxycholecalciferol might be associated with bone metabolism.