The Extracellular K+ Concentration Dependence of Outward Currents through Kir2.1 Channels Is Regulated by Extracellular Na+ and Ca2

It has been known for more than three decades that outward Kir currents (IK1) increase with increasing extracellular K+ concentration ([K+]o). Although this increase in IK1 can have significant impacts under pathophysiological cardiac conditions, where [K+]o can be as high as 18 mm and thus predispose the heart to re-entrant ventricular arrhythmias, the underlying mechanism has remained unclear. Here, we show that the steep [K+]o dependence of Kir2.1-mediated outward IK1 was due to [K+]o-dependent inhibition of outward IK1 by extracellular Na+ and Ca2+. This could be accounted for by Na+/Ca2+ inhibition of IK1 through screening of local negative surface charges. Consistent with this, extracellular Na+ and Ca2+ reduced the outward single-channel current and did not increase open-state noise or decrease the mean open time. In addition, neutralizing negative surface charges with a carboxylate esterifying agent inhibited outward IK1 in a similar [K+]o-dependent manner as Na+/Ca2+. Site-directed mutagenesis studies identified Asp114 and Glu153 as the source of surface charges. Reducing K+ activation and surface electrostatic effects in an R148Y mutant mimicked the action of extracellular Na+ and Ca2+, suggesting that in addition to exerting a surface electrostatic effect, Na+ and Ca2+ might inhibit outward IK1 by inhibiting K+ activation. This study identified interactions of K+ with Na+ and Ca2+ that are important for the [K+]o dependence of Kir2.1-mediated outward IK1.