TRANSPORT OF ORGANIC CATIONS BY A RENAL EPITHELIAL-CELL LINE (OK)

The goal of this study was to determine the mechanisms involved in the transport of the organic cation, tetraethylammonium (TEA), across the apical membrane of OK cells. [C-14]TEA accumulated in OK cell monolayers reaching equilibrium in 2 h. The uptake of [C-14]TEA at equilibrium was dependent upon temperature and was inhibited by sodium azide and by various organic cations, including N1-methylnicotinamide (NMN), mepiperphenidol, and cimetidine but not by the organic anion, p-aminohippuric acid. The initial uptake of [C-14]TEA was characterized by a saturable process. The mean +/- S.D. K(m) was 27.8 +/- 2.6-mu-M and the V(max) was 414 +/- 26.5 pmol/mg protein/min. Both an accelerated efflux and influx of [C-14]TEA in the presence of a trans-gradient of unlabeled TEA and NMN was observed, whereas a deaccelerated influx and efflux was observed in the presence of a trans-gradient of mepiperphenidol. The mechanism of interaction between NMN and TEA was examined. NMN significantly increased the apparent K(m) (mean +/- S.D.) of TEA to 82.8 +/- 16.4-mu-M (p < 0.001), whereas the V(max) (mean +/- S.D.) was only slightly affected (478 +/- 72 pmol/mg protein/min) suggesting a competitive inhibition. The stimulatory effect of trans-gradients of NMN on TEA transport was due to an increase in the V(max) of TEA suggesting that NMN trans-stimulates TEA transport by increasing the turnover rate of the exchanger. In the presence of an inwardly directed proton gradient, the efflux at 30 s of [C-14]TEA from the OK cell monolayers was significantly accelerated (p < 0.05). Studies with the pH-sensitive fluorescent probe, 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein, suggested that TEA could drive the countertransport of protons. In apical membrane vesicles prepared from OK cells, the uptake of [H-3]NMN exhibited an apparent "overshoot phenomenon" in the presence of an initial outwardly directed proton gradient. Protons competitively inhibited TEA uptake suggesting that the proton/organic cation and the organic cation/organic cation self exchange mechanism are the same mechanism. This is the first report describing both TEA self-exchange and proton/TEA exchange in the apical membrane of a continuous cell line. OK cells are an excellent model for the study of organic cation transport across the apical membrane.