Numerous endogenous vasoactive agents have been shown to cause lymphatic smooth muscle contraction. In this study, we assessed the ability of serotonin (5-HT) to alter lymphatic smooth muscle activity and elucidated the receptor mechanisms of 5-HT's actions. Both intralymphatic and intra-arterial administration of 5-HT significantly increased lymphatic smooth muscle activity in lymphatics perfused at constant flow, as indicated by an increase in lymphatic perfusion pressure. The 5-HT-induced increase in lymphatic perfusion pressure is attenuated but not blocked by the intra-arterial infusion of phentolamine, suggesting the involvement of -adrenoreceptors and 5-HT receptors. Intralymphatic infusion of the 5-HT₂-receptor-agonist -methylserotonin significantly increased lymphatic perfusion pressure, either alone or when administered into an -receptor blocked preparation, whereas the 5-HT₁-receptor-agonist carboxyamidotryptamine maleate did not effect the prenodal lymphatics. These data indicate that the lymphatic smooth muscle contraction produced by 5-HT is mediated both by lymphatic -adrenoreceptors and 5-HT₂ receptors.