Opossum kidney cells take up L-DOPA through an organic cation potential-dependent and proton-independent transporter

The present work was aimed at studying the kinetics and nature of the L-DOPA transporter in opossum kidney (OK) cells. Saturation experiments were performed in OK cells incubated for 6 min with increasing concentrations of L-DOPA (10 to 2500 microM); non-linear analysis of the saturation curve revealed for L-DOPA a K(m) of 129 microM (114, 145) and Vmax of 30.0 +/- 0.4 nmol mg protein-1 6 min-1. The uptake of L-DOPA (250 microM) was inhibited in a concentration-dependent manner by cyanine 863, an organic cation inhibitor, with a Ki value of 638 (430, 947) microM; the organic anion inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulphonic acid (DIDS), was devoid of effect upon the uptake of L-DOPA. The uptake of L-DOPA (250 microM) was significantly (P < 0.02) decreased (25% reduction) when cells were incubated in the presence of 137 mM K+ plus 5 mM Na+, when compared with the control condition (137 mM Na+ plus 5 mM K+); substitution of NaCl by choline chloride (137 mM) did not affect L-DOPA uptake. Similarly, inwardly or outwardly directed proton gradients of 0.5 pH units (7.9, 7.4, 6.9, 6.4 and 5.9) were found not to change L-DOPA uptake. In conclusion, the L-DOPA uptake system in OK cells has the characteristics of an organic cation potential-dependent and proton-independent transporter.