The role of calcitriol in stimulating intestinal active Ca²⁺ absorption during postnatal life was studied in newborn, suckling, and weaned control (Con) piglets and piglets suffering from inherited calcitriol deficiency (Def piglets). In addition, a group of Def piglets was treated with vitamin D₃ (Def-D₃ piglets), which normalized plasma calcitriol levels. Regardless of age, duodenal calbindin-D_9k concentrations ranged between 1,839 and 2,846 µg/g mucosa in Con piglets, between 821 and 1,219 µg/g mucosa in Def piglets, and between 2,960 and 3,692 µg/g mucosa in Def-D₃ animals. In weaned animals, active Ca²⁺ absorption as calculated from in vitro ⁴⁵Ca²⁺ flux rate measurements in Ussing chambers could be related to calbindin-D_9k levels. Thus active Ca²⁺ absorption was completely absent in Def animals but was reconstituted in Def-D₃ animals. In contrast, in newborn Def piglets active Ca²⁺ absorption functioned normally despite the low plasma calcitriol and mucosal calbindin-D_9k levels and could not be affected by treatment with vitamin D₃. Similar results were obtained from suckling Def piglets. The microtubule-disrupting agent colchicine caused significant inhibition of transepithelial net Ca²⁺ absorption in duodenal epithelia from newborn piglets without exerting an effect in suckling and weaned animals. Colchicine had no effect on Ca²⁺ uptake across the brush border membrane of mucosal enterocytes or on glucose-dependent electrogenic net ion flux rates in duodenal preparations from newborn Con piglets. In conclusion, our findings reveal intestinal active Ca²⁺ absorption during early postnatal life of pigs that involves calcitriol-independent mechanisms and that may include intact microtubule actions.