Decrease in serum calcium level leading to hypocalcemia during the first week of life is a frequent finding in premature neonates. Eight premature neonates presenting with such an episode of hypocalcemia in the course of their first 4 days of life were studied. They were fed with a phosphate-enriched human milk and given vitamin D3 (2,100 IU/day per os). We have evaluated the effect of a 24-hour pharmacologic calcium infusion on the circulating levels of calcium, inorganic phosphate, magnesium, 25-hydroxycalciferol (25-OHD), 1 alpha,25-dihydroxycalciferol [1,25(OH)2D] and immunoreactive parathyroid hormone (iPTH). After the infusion, circulating iPTH and Pi levels dropped significantly (p less than 0.025 and p less than 0.005 respectively) whereas serum Ca and 25-OHD (p less than 0.005) increased. Mg and 1,25(OH)2D serum levels remained unchanged. Our data show that an increased calcium supply sustained for 24 h induces an appropriate response in iPTH secretion. Effects on circulating levels of 1,25(OH)2D were variable and probably reflected individual differences in half life of 1,25(OH)2D or in set points in the feedback mechanisms involved in the control of 1,25(OH)2D synthesis.