We studied the effects of vitamin D metabolites on parathyroid hormone (PTH) secretion. Test materials were injected into the cranial thyroid artery of the dog, and immunoreactive PTH was measured frequently in serum samples from the inferior thyroid vein and the femoral vein. This model for the study of secretion had previously been validated with the use of known modulators on PTH secretion. In control experiments, injection of 100% ethanol, the vehicle in which cholecalciferol (D3) metabolites were suspended, resulted in no change in PTH secretion. Likewise, native vitamin D3, in doses ranging from 250 to 1,250 ng had no effect on PTH secretion. 25-Hydroxycholecalciferol, 25-(OH)D3, in doses of 125-240 ng, caused complete suppression of PTH secretion. When 24,25-dihydroxycholecalciferol, 24,25-(OH)2D3, was injected in doses of 50-250 ng, suppression of PTH secretion was again complete; in doses of 5 ng, injection of this metabolite resulted in significant but incomplete suppression of secretion. In doses of 50-250 ng, 1,25-(OH)2D3 strongly stimulated PTH secretion, but in a dose of 5 ng this metabolite had no effects. Injection of equal doses of 1,25-(OH)2D3 and 24,25-(OH)2D3 resulted in significant suppression of PTH secretion. Hypocalcemia-induced stimulation of PTH secretion was suppressed by 24,25-(OH)2D3 while hypercalcemia-induced suppression of PTH secretion was stimulated by 1,25-(OH)2D3. In all experiments showing suppression of PTH secretion, peripheral PTH decreased. Arguments are presented for considering the suppressive effects of D3 metabolites as physiologic modulators. However, this stimulating effect of 1,25-(OH)2D3 occurred only in pharmacologic doses and hence probably has no physiologic relevance.