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This Article Full Text Full Text (PDF) All Versions of this Article: 296/6/F1504    most recent 90754.2008v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by Lee, W.-K. Articles by Thévenod, F. PubMed PubMed Citation Articles by Lee, W.-K. Articles by Thévenod, F.

Organic cation transporters OCT1, 2, and 3 mediate high-affinity transport of the mutagenic vital dye ethidium in the kidney proximal tubule

¹Department of Physiology and Pathophysiology, University of Witten/Herdecke, Witten; ²Institute of Physiology, University of Regensburg, Regensburg; ³Experimentelle Nephrologie, Universitätsklinikum Münster, Medizinische Klinik und Poliklinik D, Münster; and ⁴Institute of Anatomy and Cell Biology, Bayerische Julius-Maximilians Universität, Würzburg, Germany

Submitted 17 December 2008 ; accepted in final form 1 April 2009

The positively charged fluorescent dyes ethidium (Et⁺) and propidium (Pr²⁺) are widely used as DNA and necrosis markers. Et⁺ is cytotoxic and mutagenic. The polyspecific organic cation transporters OCT1 (SLC22A1), OCT2 (SLC22A2), and OCT3 (SLC22A3) mediate electrogenic facilitated diffusion of small (500 Da) organic cations with broad specificities. In humans, OCT2 mediates basolateral uptake by kidney proximal tubules (PT), whereas in rodents OCT1/2 are involved. In mouse kidney, perfused Et⁺ accumulated predominantly in the S2/S3 segments of the PT, but not Pr²⁺. In cells stably overexpressing human OCTs (hOCTs), Et⁺ uptake was observed with K_m values of 0.8 ± 0.2 µM (hOCT1), 1.7 ± 0.5 µM (hOCT2), and 2.0 ± 0.5 µM (hOCT3), whereas Pr²⁺ was not transported. Accumulation of Et⁺ was inhibited by OCT substrates quinine, 3-methyl-4-phenylpyridinium (MPP⁺), cimetidine, and tetraethylammonium (TEA⁺). For hOCT1 and hOCT2, the IC₅₀ values for MPP⁺, TEA⁺, and cimetidine were higher than for inhibition of previously tested transported substrates. For hOCT2, the inhibition of Et⁺ uptake by MPP⁺ and cimetidine was shown to be competitive. Et⁺ also inhibited transport of 0.1 µM [³H]MPP⁺ by all hOCT isoforms with IC₅₀ values between 0.4 and 1.3 µM, and the inhibition of hOCT1-mediated uptake of MPP⁺ by Et⁺ was competitive. In Oct1/2^–/– mice, Et⁺ uptake in the PT was almost abolished. The data demonstrate that Et⁺ is taken up avidly by the PT, which is mediated by OCT1 and/or OCT2. Considering the high affinity of OCTs for Et⁺ and their strong expression in various organs, strict safety guidelines for Et⁺ handling should be reinforced.

SLC22A; cancer; epithelia; homidium; toxicity