title: Alkali cation binding and permeation in the rat organic cation transporter rOCT2.
abstract: Organic cation transporters of the OCT family mediate downhill transport of organic cations, compatible with carrier, pore, or gate-lumen-gate mechanisms. We studied rat OCT2 expressed in Xenopus oocytes by the two-electrode voltage-clamp technique, including membrane capacitance (C(m)) monitoring. Choline, a transported cationic substrate, elicited the expected inward currents but also elicited decreases of C(m). Similar C(m) decreases were caused by the non-transported inhibitors tetrabutylammonium (a cation) and corticosterone (uncharged). Effects on C(m) were voltage-dependent, with a maximum at -140 mV. These findings suggest that the empty rOCT2 protein can undergo an electrogenic conformation change, with one conformation highly favored at physiological voltage. Moreover, alkali cations elicited considerable inward currents and inhibited uptake of [(14)C]tetraethylammonium with a sequence Cs(+) > Rb(+) > K(+) > Na(+) approximately Li(+). Cs(+) affected current and capacitance with similar affinity (K(0.5) approximately 50 mm). Tetraethylammonium inhibited Cs(+) currents in a concentration-dependent manner. Conversely, Cs(+) inhibited tetraethylammonium uptake by a competitive mechanism. Activation energy of the currents estimated from measurements between 12 degrees C and 32 degrees C was approximately 81 kJ/mol for Cs(+) and 39 kJ/mol for tetramethylammonium, compatible with permeation of Cs(+) through rOCT2 along the same path as organic substrates and by a mechanism different from simple electrodiffusion. Rationalization of Cs(+) selectivity in terms of a pore pointed to a pore diameter of approximately 4 A. Intriguingly, that value matches the known selectivity of rOCT2 for organic compounds. Our data show that selective permeability of rOCT2 is not determined by ligand affinity but might rather be understood in terms of the ion channel concept of a distinct "selectivity filter."
authors.affiliation: Department of Anatomy and Cell Biology, University of Würzburg, Koellikerstrasse 6, 97070 Würzburg, Germany. bernhard.schmitt@mail.uni-wuerzburg.de
authors.forename: Bernhard M
authors.initials: BM
authors.lastname: Schmitt
authors.forename: Hermann
authors.initials: H
authors.lastname: Koepsell
mesh_terms: D000305:Adrenal Cortex Hormones / Q000737:chemistry; D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D002412:Cations; D002586:Cesium / Q000737:chemistry / Q000494:pharmacology; D002794:Choline / Q000737:chemistry*; D004058:Diffusion; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D007477:Ions; D008024:Ligands; D026901:Membrane Transport Proteins / Q000737:chemistry; D008954:Models, Biological; D009865:Oocytes / Q000378:metabolism; D027701:Organic Cation Transport Proteins / Q000737:chemistry* / Q000378:metabolism*; D000075102:Organic Cation Transporter 2; D018408:Patch-Clamp Techniques; D011485:Protein Binding; D051381:Rats; D013696:Temperature; D019789:Tetraethylammonium / Q000737:chemistry; D013997:Time Factors; D014982:Xenopus laevis
chemical_list: D000305:Adrenal Cortex Hormones; D002412:Cations; D007477:Ions; D008024:Ligands; D026901:Membrane Transport Proteins; D027701:Organic Cation Transport Proteins; D000075102:Organic Cation Transporter 2; C100590:Slc22a2 protein, rat; D002586:Cesium; D019789:Tetraethylammonium; D002794:Choline; D002118:Calcium