The aim of the present study was to determine the mechanism of the positive inotropic effect of carbachol on ventricular myocardium. Carbachol produced a concentration-dependent (0.1 to 300 mumol/l) increase in contraction force on the catecholamine-depleted papillary muscle of the guinea pig without affecting the normal action potential or the slow action potential evoked in 24 mmol/l K+. Since atropine prevented the inotropic effect of carbachol, muscarinic receptors were involved. Carbachol (300 mumol/l) produced an increase in intracellular Na+-ion-activity, aiNa, by about 3 mmol/l in the quiescent muscle, and the time course of the aiNa change corresponded with the development of the positive inotropic effect as determined in the stimulated preparation (0.2 Hz). The effect of carbachol on force of contraction and on aiNa was diminished by reducing [Ca2+]0. The positive inotropic effect of carbachol was dependent on repetitive activity and was markedly enhanced in the presence of dihydro-ouabain. The results are consistent with the hypothesis, that carbachol increases the Na+ permeability of the sarcolemma via muscarinic receptors, and enhances force of contraction by stimulating the Na+-Ca2+-exchange.