A transfected cell model for the renal toxin transporter, rOCT2

A cDNA for the organic cation transporter (rOCT2) of the rat kidney was ins erted into the retroviral plasmid pLXSN. This plasmid was used to stably tr ansfect NIH3T3 cells. The transfected cell line exhibited an enhanced rate of tetraethylammonium (TEA) uptake and efflux compared to wild-type NIH3T3 cells. Uptake of TEA by the transfected cells was markedly reduced upon inc ubation at 4 degrees C. When the extracellular pH was lowered from 8.1 to 5 .9, uptake was also reduced, suggesting inhibition of rOCT2 by extracellula r protons. The apparent K-m for TEA in the transfected cells was 141 mu M. The classical organic cation transport inhibitors, cyanine 863 and cimetidi ne, produced noncompetitive inhibition with apparent K-i values of 0.81 and 198 mu M, respectively. Daunomycin, vinblastine, and the deoxyadenosine an alogs, 2'-deoxytubercidin and 2-chlorodeoxyadenosine, did not appear to be substrates for rOCT2. However, the anticancer drug, cisplatin, competitivel y inhibited TEA uptake by rOCT2 with an apparent K-i value of 925 mu M, sug gesting that rOCT2 may play a role in its renal secretion. In summary, tran sfected NIH3T3 cells provide a facile system by which this and other organi c ion transporters can be studied.