Abstract
The inward rectifier K+ channels in the Kir2 subfamily carry very small outward currents compared to the inward currents and this strong inward rectification is caused by a voltage-dependent block of the channel by intracellular cations. In this study, we show using the inside-out patch recordings that Kir2.1 channels exhibit a time- and voltage-dependent gating that depends on the internal pH in the range below 7.2. This gating was observed in the absence of polyamines, Mg2+, and EDTA in the internal (bath) solution and thus seemed to be unrelated to the mechanism of the channel block induced by cations. Substitution of the His residue (pKa = ∼6.5) situated on the wall of the cytoplasmic pore with the basic Lys residue (H226K) did not eliminate this pH-dependent gating. We then examined Kir2.1 mutant channels known to have reduced sensitivity to internal cationic blockers. Neutralization of the acidic Glu residues situated on the wall of the cytoplasmic pore (E299S and E224G) neither notably affected the pH-dependent gating. By contrast, neutralization of the acidic Asp residue on the wall of the transmembrane pore (D172N) abolished the gating. These findings suggest that the negative charges at site 172 of Kir2.1 (composed of four subunits), which are thought to form the high-affinity binding site for polyamines, are required for the pH-dependent gating mechanism. [J Physiol Sci. 2008;58 Suppl:S75]
