Transport of the glutathione-methylmercury complex across liver canalicular membranes on reduced glutathione carriers.
Methylmercury transport across liver canalicular membranes into bile, a major route of excretion of this toxic compound, is dependent upon intracellular GSH, and a glutathione-methylmercury complex (CH3Hg.SG) has been detected in liver tissue and bile. To examine whether the CH3Hg.SG complex is itself transported across the canalicular membrane and to identify the transport system involved, studies were performed in isolated rat liver canalicular plasma membrane vesicles. Uptake of CH3(203)Hg.SG (10 microM) into an osmotically active space was temperature-sensitive and unaffected by either ATP (5 mM) or an inwardly directed Na+ gradient (100 mM); however, CH3Hg.SG uptake was enhanced by a valinomycin-induced K+ diffusion potential (inside-positive) indicating that its transport was electrogenic. Transport of CH3Hg.SG exhibited saturation kinetics with both high affinity (Km = 12 +/- 2 microM, Vmax = 0.23 +/- 0.02 nmol.mg-1.20 s-1) and low affinity (Km = 1.47 +/- 0.22 mM, Vmax = 1.23 +/- 0.14 nmol.mg-1.20 s-1) components. Uptake of this complex was inhibited by GSH, the GSH analog ophthalmic acid, S-methyl, S-ethyl, S-butyl, S-hexyl, S-octyl, and S-dinitrophenyl glutathione, but not by GSSG, bile acids, amino acids, and P-glycoprotein inhibitors. Furthermore, GSH competitively inhibited (Ki = 83 microM) and trans-stimulated CH3Hg.SG uptake into the canalicular vesicles. These studies provide the first kinetic characterization of a transport system for glutathione-mercaptides and indicate that CH3Hg.SG is not a substrate for the ATP-dependent, canalicular GSSG or glutathione S-conjugate carriers, but appears to be a substrate for canalicular carriers that also transport GSH. Because efflux systems for GSH are found in all mammalian cells, transport of glutathione-metal complexes by such carriers may be a common mechanism for the removal of methylmercury and possibly other metals from cells.
Dutczak
W J
WJ
Department of Environmental Medicine, University of Rochester School of Medicine, New York 14642.
Ballatori
N
N
D000818:Animals; D001648:Bile Canaliculi / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D005978:Glutathione / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000494:pharmacology; D007700:Kinetics; D008094:Lithium / Q000494:pharmacology; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008767:Methylmercury Compounds / Q000378:metabolism*; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012964:Sodium / Q000494:pharmacology; D013379:Substrate Specificity; D013696:Temperature; D014634:Valinomycin / Q000494:pharmacology
D002352:Carrier Proteins; D008767:Methylmercury Compounds; D014634:Valinomycin; C030588:methylmercury glutathione; D008094:Lithium; D012964:Sodium; D005978:Glutathione; D011188:Potassium