Sodium ions are required for the active transport of amino acids such as alpha-aminoisobutyric acid (AIB) into skeletal muscle. To examine the role of Na+-K+-ATPase in this phenomenon, studies were carried out using the isolated perfused rat hindquarter preparation. Perfusion for 30 min with ouabain at a dose sufficient to inhibit the Na+-K+ pump (10(-4) M) inhibited the basal rate of AIB uptake in all muscles studied by up to 80%. However, it failed to inhibit the stimulation of AIB uptake, either by insulin (200 microU/ml) or electrically-induced muscle contractions. The increase in K+ release by the hindquarter in the presence of ouabain was the same under all conditions suggesting comparable inhibition of the Na+-K+ pump. These studies suggest that the basal, but not insulin or exercise-stimulated AIB transport into muscle is acutely dependent on a functional Na+-K+ pump. They also suggest that stimulated and basal uptake of AIB involve different mechanisms.