L-cysteine and S-(1,2-dichlorovinyl)-L-cysteine transport in rat liver canalicular membrane vesicles: potential reabsorption mechanisms for biliary metabolites of glutathione and its S-conjugates

Abstract

Transport of L-cysteine and a cysteine S-conjugate, S-(1,2-dichlorovinyl)-L-cysteine (DCVC) was investigated in rat liver canalicular plasma membrane (cLPM) vesicles. Cysteine uptake into an osmotically active intravesicular space was temperature sensitive and further enhanced by an inwardly directed Na+ gradient. Na(+)-dependent and -independent L-cysteine uptake exhibited saturation kinetics with apparent Km of 53 +/- 0.7 and 1300 +/- 300 microM and Vmax of 95 +/- 21 and 1600 +/- 200 pmol.mg protein-1.10 sec-1 for the Na(+)-dependent components, and an apparent Km of 207 +/- 48 microM and a Vmax of 355 +/- 71 pmol.mg protein-1.10 sec-1 for the Na(+)-independent component. Na(+)-dependent uptake was inhibited by L-alanine, glycine, L-phenylalanine and L-leucine, whereas Na(+)-independent uptake was inhibited by L-phenylalanine, L-leucine and 2-amino-2-norbornanecarboxylic acid. Both Na(+)-dependent and -independent L-cysteine transport processes were inhibited by several cysteine S-conjugates, with DCVC having the strongest effect. Inhibition of [35S]L-cysteine uptake by DCVC was noncompetitive with a Ki of 1.2 +/- 0.1 mM. On the other hand, uptake of [35S]DCVC by the rat cLPM vesicles was not stimulated by a Na(+)-gradient, but was inhibited by several other amino acids, including L-cysteine. Further investigation of [35S]DCVC uptake in rat cLPM vesicles indicated a saturable Na(+)-independent process with an apparent Km of 155 +/- 42 microM, and a Vmax of 393 +/- 53 pmol.mg protein-1.5 sec-1.2+.