In electrically stimulated myocytes loaded with the fluorescent Ca2+ indicator indo 1-acetoxymethyl ester, spermine induced a dose-dependent (100-500 microM) negative inotropic effect, which was associated with a decrease in the magnitude of the cytosolic Ca2+ concentration ([Ca2+]i) transient but not with changes in myofilament responsiveness to Ca2+. Spermidine induced a less pronounced negative inotropic effect, whereas putrescine did not modify myocyte contraction. In the unstimulated state, spermine did not alter resting [Ca2+]i. Superfusion of the cardiac myocytes with 10 mM alpha-difluoromethylornithine, an inhibitor of polyamine synthesis, did not modify cellular responses to isoproterenol (10(-9)-10(-7) M). beta-Adrenergic stimulation did not affect either ornithine decarboxylase activity or intracellular polyamine levels within a 10-s to 15-min period of treatment. In summary, only exogenously administered polyamines were able to influence myocyte contractility. Their negative inotropic effect resulted from changes in [Ca2+]i homeostasis and required cellular depolarization.