Rationale: Nociceptin/orphanin FQ has been reported to inhibit capsaicin- and mechanically-provoked cough in animal models, but the mechanism of this effect has not been elucidated. Objectives: The objectives of this study were to determine if nociceptin inhibits acid-evoked cough in conscious animals and to evaluate the mechanism of this effect. Methods: We tested the effect of nociceptin on acid-induced cough in conscious guinea pigs and acid-induced nerve activation in airway-labeled vagal sensory neurons using calcium imaging techniques and the gramicidin-perforated patch clamp technique. Measurements and main results: Nociceptin (3 mg/kg, i.p.) effectively inhibited acid-evoked cough in guinea pigs by nearly 70%. Acid (pH 5) increased intracellular free calcium in acutely dissociated vagal jugular ganglionic neurons. The acid-induced increase of intracellular calcium was inhibited by a selective transient receptor potential-vanilloid-1 (TRPV1) antagonist, 5- iodo-resiniferatoxin (1 µM, ~ 80% reduction). The inhibitory effect of 5- iodo-resiniferatoxin on acid-induced increases in calcium was mimicked by nociceptin (0.1 µM). In gramicidin-perforated patch clamp recordings on airway specific capsaicin-sensitive jugular ganglion neurons, acid (pH 5) induced two distinct inward currents. A transient current was evoked that was inhibited by amiloride and a sustained current was evoked that was inhibited by 5-iodo-resiniferatoxin. Nociceptin selectively inhibited only the sustained component of acid-induced inward current. Conclusion: These results indicated that the inhibitory effect of nociceptin on acid-induced cough may result from a direct inhibitory effect on peripheral C-fiber activity caused by the selective inhibition of acid-induced TRPV1 activation.