The effects of synthetic thyrotropinreleasing hormone (TRH) were measured on release of prolactin by normal rat pituitary and pituitary tumor tissue in vitro, and in vivo on pituitary and serum prolactin concentrations in intact and thyro—parathyroidectomized male rats. TRH in doses of 3 and 30 ng was incorporated into medium 199 containing a half anterior pituitary from normal male rats. No significant effect was observed on prolactin release by the end of 4, 8 or 12 hr of incubation when compared with control pituitary halves. Similar doses of TRH were incubated with rat pituitary tumor (MtT.WlS) tissue for the same periods, and again no effect was observed on prolactin release. A single iv injection of 5 or 7.5 μg of TRH was given to male rats and blood samples were collected IS and 30 min later. No change was observed in serum prolactin levels. When 50 μg TRH was injected iv daily for 6 days into intact male rats, there was a significant increase in pituitary prolactin concentration and a small but insignificant elevation in serum prolactin. Neither TRH nor TSH had any effect on these parameters when injected into thyro—parathyroidectomized rats. Thyro—parathyroidectomy significantly reduced pituitary prolactin concentration and resulted in a small but insignificant decrease in serum prolactin. These results indicate that synthetic TRH does not stimulate prolactin release by the normal rat pituitary or by a rat pituitary tumor in vitro, and that multi—injections of TRH increase pituitary prolactin concentration in intact rats via the TSHthyroid system.(Endocrinology91: 1540, 1972)