Characterization of glutamate transport in isolated adult rat heart cells

Abstract

Uptake of L-glutamate was studied in isolated adult rat heart myocytes in the presence and absence of oxygen. Normoxic incubation of cardiomyocytes with glutamate concentrations varying from 0.0625 to 0.33 mM led to an apparent K_m-value for glutamate of 0.18 mM. However, incubation of cardiomyocytes with glutamate concentrations up to 10 mM did not saturate glutamate uptake. We therefore assume that the high affinity glutamate carrier is saturated and further nonspecific transport mechanisms are responsible for the non-saturation kinetics. The initial glutamate uptake rate from the incubation medium, under normoxia, amounted to 24.3 ±1.6 pmol min-¹ mg-¹ protein and increased under anoxia to 38.1 ± 3.8 pmol min-¹ mg-¹ protein. Even this increased glutamate uptake could not prevent a depletion of intracellular glutamate in anaerobic cardiomyocytes. Under reoxygenation glutamate increased again to control values with a rate of 930 pmol min-¹ mg-¹ protein within 15 min. Glutamate uptake from the incubation medium after reoxygenation amounted to 31.9 ± 2.5 pmol min-¹ mg-¹ protein. Therefore the glutamate uptake from extracellular space cannot be responsible for the rapid increase in intracellular glutamate concentration after reoxygenation. Cardiomyocytes must replenish glutamate by other mechanisms, e.g. intracellular proteolysis.