Increased esophageal blood flow may protect against damaging refluxed gastric juices. We have shown that mast cells, histamine, and nitric oxide increase esophageal blood flow in the opossum during acid perfusion. This study examined the roles of substance P and calcitonin gene-related peptide on acid-induced hyperemia and whether the effects of substance P are mediated by mast cells. The opossum distal esophagus was perfused with saline, acid, or capsaicin while blood flow and histamine release were determined. Neuropeptides and neurokinin antagonists were administered parenterally. Only acid or calcitonin gene-related peptide (not substance P or capsaicin) significantly increased blood flow, which was prevented by neurokinin or calcitonin-gene-related peptide antagonists. Acid, substance P, and capsaicin all increased histamine release. Pretreatment with neurokinin antagonists did not affect acid-induced histamine release. We conclude that calcitonin-gene-related peptide is an important mediator of acid-induced esophageal hyperemia, while substance P plays an indirect role.