The effects of removing extracellular Ca2+ and Mg2+ on the membrane potential, membrane current and intracellular Na+ activity (aiNa) were investigated in guinea-pig and rat ventricular myocytes. Membrane potential was recorded with a patch pipette and whole-cell membrane currents using a single-electrode voltage clamp. Both guinea-pig and rat cells depolarize when the bathing Ca2+ and Mg2+ are removed and the steady-state aiNa increases rapidly from a resting value of 6.4+/- 0.6 mM to 33+/-3.8 mM in guinea-pig (n=9) and from 8.9+/-0.8 mM to 29.3+/-3.0 mM (n=5) in rat ventricular myocytes. Guinea-pig myocytes partially repolarized when, in addition to removal of the bathing Ca2+ and Mg2+, K+ was also removed, however rat cells remained depolarized. A large diltiazem-sensitive inward current was recorded in guinea-pig and rat myocytes, voltage-clamped at -20 mV, when the bathing divalent cations were removed. When the bathing K+ was removed after Ca2+ and Mg2+ depletion, a large outward K+ current developed in guinea-pig, but not in rat myocytes. This current had a reversal potential of -80+/-0.7 mV and was not inhibited by high Mg2+ or glybenclamide indicating that it is not due to activation of non-selective cation or adenosine triphosphate (ATP)-sensitive K channels. The current was not activated when Li+ replaced the bathing Na+ and was blocked by R-56865, suggesting that it was due to the activation of KNa channels.