The capacity of the medium eminence to transport thyrotropin-releasing hormone (TRH) from cerebrospinal fluid (CSF) to hypophysial portal blood, and the ability of TRH when introduced into a lateral ventricle to stimulate TSH release from the pituitary gland were investigated. Male rats were injected either intraventricularly or intravenously with 0, 1, 10, or 100 ng of TRH, and plasma TSH concentrations were determined at various times thereafter. TRH administration via both routes resulted in substantial release of TSH. Following intraventricular injection of TRH, there was a delay in reached maximal TSH concentration when compared with the faster elevation and faster decline in TSH concentrations which followed intravenous injection of the same dose of TRH. In a second experiment, 7 muCi of [3H]TRH were introduced intraventricularly or intravenously, and hypophysial portal and arterial blood were simultaneously collected and examined for the presence of radioactivity. The intraventricular injection of [3H]TRH resulted in a peak of radioactivity in portal blood within minutes, which was maintained for 20--30 min and then declined. The concentration of radioactivity in arterial blood from the same animals was considerably lower than that in portal blood. The intravenous administration of [3H]TRH resulted in radioactive peaks in both portal and arterial blood with a higher concentration of radioactive substances in arterial blood. However, the level of radioactivity in portal blood following intravenous injection of [3H]TRH comprised no more than 5--10% of that found following intraventricular administration of the saem dose. The data support the view that TRH is able to cross the medium eminence from CSF into hypophysial portal blood and that it is capable of stimulating the pituitary gland to release TSH.