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 mu M); non-linear analysis of the saturation curve revealed for L-DOPA a K-m of 129 mu M (114, 145) and a V-max of 30.0 +/- 0.4 nmol mg protein(-1) 6 min(-1). The uptake of L-DOPA (250 mu M) was inhibited in a concentration-dependent manner by cyanine 863, an organic cation inhibitor, with a K-i value of 638 (430, 947) mu M; 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 mu M) 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. (C) 1997 Academic Press Limited.