Superoxide radical (O2-.) combines with nitric oxide (NO) to form peroxynitrite, thereby nullifying the biological activity of NO. Superoxide dismutase (SOD) prevents this reaction by converting O2-. to H2O2. We tested the hypotheses that the antioxidant enzymes catalase (CAT), Mn SOD, and Cu/Zn SOD are present in enteric neurons of the opossum esophagus, and that O2-. alters esophageal motor function. Immunostaining demonstrated CAT, Mn SOD, and Cu/Zn SOD immunoreactivity in interganglionic nerve bundles and ganglia of the myenteric and submucosal plexuses. Western blot analysis confirmed the presence of these enzymes in homogenates of esophageal muscularis propria, and enzyme assays demonstrated Cu/Zn SOD and Mn SOD activities of 262 and 73 U/mg protein, respectively. Both diethyldithiocarbamic acid, an inhibitor of Cu/Zn SOD, and xanthine (X) with xanthine oxidase (XO), which generate O2-., shortened the latency of the nerve-mediated contraction of circular esophageal muscle, the off response, by 20.2 and 23.4%, respectively. SOD alone did not affect the latency, but it inhibited the effect of X with XO on the latency. Antioxidant enzymes found in intramural esophageal nerves may play a role in regulating NO-mediated neuromuscular communication in the esophagus.