Abstract
We have isolated a kidney-specific organic cation transporter, rat OCT2, which is distinct from rat OCT1 (Okuda M, Saito H, Urakami Y, Takano M and Inui K (1996) Biochem Biophys Res Commun224:500–507). In our study, the functional characteristics and membrane localization of OCT1 and OCT2 were investigated by uptake studies using MDCK cells transfected with rat OCT1 or OCT2 cDNA (MDCK-OCT1 or MDCK-OCT2) and immunological studies. Tetraethylammonium (TEA) uptake by both MDCK-OCT1 and MDCK-OCT2 cells was markedly elevated when TEA was added to the basolateral medium, but not to the apical medium. Efflux of TEA from MDCK-OCT1 and MDCK-OCT2 cells was not changed by extracellular pH from 5.4 to 8.4, whereas TEA uptake by both transfectants was decreased by acidification of extracellular medium. Apparent Km values for TEA uptake by MDCK-OCT1 and MDCK-OCT2 cells were 38 and 45 μM, respectively. Although various hydrophilic organic cations such as 1-methyl-4-phenylpyridinium, cimetidine, quinidine, nicotine, N1-methylnicotinamide and guanidine markedly inhibited TEA uptake by both MDCK-OCT1 and MDCK-OCT2 cells, there were no significant differences in the apparent inhibition constants (Ki ) against these organic cations between both transfectants. Furthermore, immunological studies using a polyclonal antibody against OCT1 revealed that OCT1 was expressed in the basolateral membranes but not in the brush-border membranes of the rat kidney. These results suggested that both OCT1 and OCT2 are basolateral-type organic cation transporters with broad substrate specificities, mediating tubular secretion of cationic drugs.
Footnotes
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Send reprint requests to: Professor Ken-ichi Inui, Department of Pharmacy, Kyoto University Hospital, Sakyo-ku, Kyoto 606-8507, Japan.
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↵1 This work was supported by a Grant-in-Aid for Scientific Research (B) and a Grant-in-Aid for Scientific Research on Priority Areas of “Channel-Transporter Correlation” from the Ministry of Education, Science, and Culture of Japan, and by Grants-in-Aid from the Yamada Science Foundation.
- Abbreviations:
- RT-PCR
- reverse-transcription-polymerase chain reaction
- Tris
- 2-amino-2-hydroxymethyl-1,3-propanediol
- MES
- 2-(N-morpholino)ethanesulfonic acid
- HEPES
- 2-[4-(2-hydroxyethyl)-1-piperazinyl]ethanesulfonic acid
- SDS-PAGE
- sodium dodecyl sulfate-polyacrylamide gel electrophoresis
- Received February 12, 1998.
- Accepted June 16, 1998.
- The American Society for Pharmacology and Experimental Therapeutics
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