Characterization of L‐carnitine transport into rat skeletal muscle plasma membrane vesicles

Abstract

Transport ofl‐carnitine into skeletal muscle was investigated using rat sarcolemmal membrane vesicles. In the presence of an inwardly directed sodium chloride gradient,l‐carnitine transport showed a clear overshoot. The uptake ofl‐carnitine was increased, when vesicles were preloaded with potassium. When sodium was replaced by lithium or cesium, and chloride by nitrate or thiocyanate, transport activities were not different from in the presence of sodium chloride. However,l‐carnitine transport was clearly lower in the presence of sulfate or gluconate, suggesting potential‐dependent transport. An osmolarity plot revealed a positive slope and a significant intercept, indicating transport ofl‐carnitine into the vesicle lumen and binding to the vesicle membrane. Displacement experiments revealed that approximately 30% of thel‐carnitine associated with the vesicles was bound to the outer and 30% to the inner surface of the vesicle membrane, whereas 40% was unbound inside the vesicle. Saturable transport could be described by Michaelis–Menten kinetics with an apparentKmof 13.1 µmand aVmaxof 2.1 pmol·(mg protein−1)·s−1.l‐Carnitine transport could betrans‐stimulated by preloading the vesicles withl‐carnitine but not with the carnitine precursor butyrobetaine, and wascis‐inhibited byl‐palmitoylcarnitine,l‐isovalerylcarnitine, and glycinebetaine. On comparing carnitine transport into rat kidney brush‐border membrane vesicles and OCTN2, a sodium‐dependent high‐affinity human carnitine transporter, cloned recently from human kidney also expressed in muscle, theKmvalues are similar but driving forces, pattern of inhibition and stereospecificity are different. This suggests the existence of more than one carnitine carrier in skeletal muscle.