Direct effects of neuropeptide Y were studied in left ventricular myocytes isolated from guinea pigs. Contraction was measured as the change in unloaded cell length using a photodiode array. Action potentials were elicited at 1 Hz in current-clamp mode, and membrane currents were measured using a switch-clamp amplifier with 2 M-KCl microelectrodes. At concentrations of 10(-6) M and above, neuropeptide Y reduced contraction in a concentration-dependent fashion. The reduction in contraction by the peptide was proportionately greater in the presence of isoproterenol, and the increase in contraction caused by isoproterenol was completely inhibited by 10(-6) M neuropeptide Y. In response to neuropeptide Y, action potential duration was shortened, and the time course of the shortening was similar to that of the reduction in contraction. Under voltage clamp, 1 x 10(-5) M neuropeptide Y reduced peak L-type calcium current by 32% and shifted the myocyte current-voltage relation during a slow ramp in a manner that suggested a reduction in the background rectifier K+ current. The effects of the peptide on membrane currents were greatly attenuated by preincubation of the cells with pertussis toxin (100 ng/ml). We conclude that neuropeptide Y reduces developed shortening, action potential duration, L-type calcium current, and background rectifier current in single guinea pig ventricular myocytes and that these effects are mediated, at least in part, via membrane G proteins.