The effect of H+-K+ ATPase inhibitors on airway smooth muscle tone was investigated in vitro. Four H+-K+ ATPase inhibitors, SCH 28080 (2-methyl-8-(phenylmethoxy)-imidazo[1,2-a] pyridine-3-acetonitrile), SK&F 96067 (3-butyryl-4-(2-methylphenylamino)-8-methoxy-quinoline), omaprezole (5-methoxy-2-(((4-methoxy-3,5-dimethyl-2-pyridinyl)-methyl)-sulfinyl)-1H -benzimidazole) and NC-1300-B (2-(2-dimethylaminobenzylsulfiny)-5-methoxybenzimidazole), induced concentration-dependent relaxation of guinea pig trachea with spontaneous tone, with IC50 values of 5.9, 7.1, 155 and 79 microM, respectively, SCH 28080 and omeprazole also relaxed airways precontracted with carbachol or histamine in the presence of indomethacin. Relaxation was similar in intact and epithelium-denuded tracheal preparations, suggesting that the airway epithelium neither mediates or modulates relaxation induced by H+-K+ ATPase inhibitors. SCH 28080-induced relaxation was not influenced by tetrodotoxin, suggesting that it is not neurogenically mediated. Bafilomycin A1 and concanamycin A had no effect on guinea pig tracheal spontaneous tone, suggesting that relaxation induced by H+-K+ ATPase inhibitors is not due to an interaction with a vacuolar H+ ATPase. SCH 28080 also induced concentration-dependent relaxation of human bronchi precontracted with histamine. These results demonstrate that several structurally and/or mechanistically distinct H+-K+ ATPase inhibitors cause relaxation of airway smooth muscle in vitro, and suggest that a H+-K+ ATPase or similar pathway may play a role in the maintenance of airway smooth muscle tone.