L-Arginine (L-Arg) is a basic amino acid that plays a central role in the biosynthesis of nitric oxide, creatine, agmantine, polyamines, proline, and glutamate. Most tissues, including myocardium, must import L-Arg from the circulation to ensure adequate intracellular levels of this amino acid. This study reports novel L-Arg-activated inward currents in whole-cell voltage-clamped rat ventricular cardiomyocytes. Ion-substitution experiments identified extracellular L-Arg as the charge-carrying cationic species responsible for these currents, which, thus, represent L-Arg import into cardiac myocytes. This result was independently confirmed by an increase in myocyte nitric oxide production upon extracellular application of L-Arg. The inward movement of Arg molecules was found to be passive and independent of Na+, K+, Ca2+, and Mg2+. The process displayed saturation and membrane potential (Vm)-dependent kinetics, with a K0.5 for L-Arg that increased from 5 mM at hyperpolarizing Vm to 20 mM at +40 mV. L-lysine and L-ornithine but not D-Arg produced currents with characteristics similar to that activated by L-Arg indicating that the transport process is stereospecific for cationic L-amino acids. L-Arg current was fully blocked after brief incubation with 0.2 mM N-ethylmaleimide. These features suggest the activity of the low-affinity, high-capacity CAT-2A member of the y+ family of transporters as responsible for L-Arg currents in acutely-isolated cardiomyocytes. Regardless of the mechanism, we hypothesize that a low-affinity arginine transport process in heart, by ensuring substrate availability for sustained NO production, might play a cardio-protective role during catabolic states known to increase Arg plasma levels several fold.