A number of previous investigations have indicated that the pituitary may directly stimulate secretion of parathyroid hormone. Others have disagreed. With the recent development of an in vitro bovine parathyroid perfusion system, the direct effect of suspected secretagogues can be assessed on a dynamic, ongoing basis. A partially purified pituitary extract (preparation A) was injected into calves. The plasma calcium increased an average of 1.1 mg/100 ml plasma. No increase of immunoreactive parathyroid hormone (iPTH) was detected, however, in the peripheral plasma prior to the increase in plasma calcium concentration. Since the peripheral plasma iPTH concentration has been shown to be relatively insensitive to changes in the secretion rate, the inability to detect a change in the iPTH concentration does not preclude a direct stimulating effect of the pituitary on the parathyroid. When preparation A was tested on in vitro perfused bovine parathyroid glands, a 30% average increase in secretion of c-iPTH (carboxy terminus) and a 56% average increase in secretion of n-iPTH (amino terminus) was observed under normocalcemic conditions. Under conditions of hypercalcemia, there was an average increase in the c-iPTH secretion rate of 60% and an average n-iPTH secretion rate increase of 88%. A failure of TSH, LH, GH, ADH, oxytocin, and prolactin to stimulate iPTH was observed. Previous reports have eliminated ACTH, MSH, and lipotropin as possible parathyroid secretagogues. The concept of a parathyroid stimulating hormone (PTSH) located in the pituitary that can directly stimulate the parathyroid gland to secrete parathyroid hormone is consistent with the results of this investigation.