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Treatment of intact adult male rats with 10 mg. perphenazine (Trilafon)/kg. resulted in a decrease in the prolactin content of the pituitary within 1 hr. This decrease was probably due to suppression of the hypothalamic prolactin-inhibiting factor (PIF). Subsequent intracarotid infusion of neutralized acid extracts of rat hypothalamus restored the pituitary prolactin content. This effect was dose-dependent within a range of -2 hypothalami. Infusion of extracts of cerebral cortex failed to increase pituitary prolactin. The response to the hypothalamic extracts is considered to be specific for PIF and is proposed as an assay method for PIF.