Intracerebroventricularly administered 5-hydroxytryptamine (5HT) altered the release of dopamine into pituitary stalk blood and of PRL into the systemic circulation of male rats. The concentration of dopamine in pituitary stalk plasma of rats given 0.5 or 5.0 microgram 5HT was 0.32 +/- 0.06 (mean +/- SE) or 0.18 +/- 0.04 ng/ml, respectively, and was significantly less than that in vehicle-treated animals (0.80 +/- 0.04 ng/ml). Relative to the mean level of PRL in animals injected with the solvent vehicle, the mean concentration of PRL in central venous plasma of rats given 0.5 or 5.0 microgram 5HT was increased 4-fold or 13-fold, respectively. Other rats were infused iv with dopamine for 45 min. After 15 min of infusion, the mean concentration of dopamine in arterial plasma was several times that of dopamine in pituitary stalk plasma of rats not infused with dopamine. Even so, this high concentration of dopamine did not prevent the 5HT-induced release of PRL. We conclude that 5HT modulates the secretion of PRL through a stimulatory mechanism that is effective in the presence of a high plasma concentration of dopamine, a PRL-release inhibiting factor, and suggest that 5HT causes the release of a hypothalamic substance(s) that stimulates release of PRL, i.e. a PRL-releasing factor.