Neuropeptide Y (NPY) inhibits the release of noradrenaline (NA) or acetylcholine (ACh) from nerve terminals in the heart. Thus, we differentiated the blocking effects of NPY on the negative and positive (atrial chronotropic and inotropic) responses to simultaneous stimulation of the intracardiac parasympathetic and sympathetic nerves in isolated, blood-perfused dog atria. NPY (0.1-3 nmol) injected into the sinus node artery decreased atrial contractile force slightly, but not sinus rate by itself. NPY inhibited the negative atrial responses to intracardiac autonomic nerve stimulation in a dose-dependent manner, whereas it rather augmented the positive responses. The inhibition on the negative chronotropic effect was more prolonged than that on the inotropic one. NPY also inhibited the negative responses to intracardiac parasympathetic nerve stimulation alone. However, NPY (3 nmol) did not change the positive responses to intracardiac sympathetic stimulation after atropine was given. NPY did not affect the atrial responses to exogenous ACh or NA. These results suggest that, under activation of autonomic nerves, NPY released from sympathetic nerves attenuates the negative chronotropic and inotropic responses to ACh released from parasympathetic nerves at the prejunctional sites more effectively than the positive responses to NA released from sympathetic nerves, and modifies the sympathetic-parasympathetic interactions in the dog heart.