A novel ion conducting route besides the central pore in an inherited mutant of G‐protein‐gated inwardly rectifying K + channel

G‐protein‐gated inwardly rectifying K + (GIRK; Kir3.x) channels play important physiological roles in various organs. Some of the disease‐associated mutations of GIRK channels are known to induce loss of K + selectivity but their structural changes remain unclear. In this study, we investigated the mechanisms underlying the abnormal ion selectivity of inherited GIRK mutants. By the two‐electrode voltage‐clamp analysis of GIRK mutants heterologously expressed in Xenopus oocytes, we observed that Kir3.2 G156S permeates Li + better than Rb + , while T154del or L173R of Kir3.2 and T158A of Kir3.4 permeate Rb + better than Li + , suggesting a unique conformational change in the G156S mutant. Applications of blockers of the selectivity filter (SF) pathway, Ba 2+ or Tertiapin‐Q (TPN‐Q), remarkably increased the Li + ‐selectivity of Kir3.2 G156S but did not alter those of the other mutants. In single‐channel recordings of Kir3.2 G156S expressed in mouse fibroblasts, two types of events were observed, one attributable to a TPN‐Q‐sensitive K + current and the second a TPN‐Q‐resistant Li + current. The results show that a novel Li + ‐permeable and blocker‐resistant pathway exists in G156S in addition to the SF pathway. Mutations in the pore helix, S148F and T151A also induced high Li + permeation. Our results demonstrate that the mechanism underlying the loss of K + selectivity of Kir3.2 G156S involves formation of a novel ion permeation pathway besides the SF pathway, which allows permeation of various species of cations. Key points G‐protein‐gated inwardly rectifying K + (GIRK; Kir3.x) channels play important roles in controlling excitation of cells in various organs, such as the brain and the heart. Some of the disease‐associated mutations of GIRK channels are known to induce loss of K + selectivity but their structural changes remain unclear. In this study, we investigated the mechanisms underlying the abnormal ion selectivity of inherited mutants of Kir3.2 and Kir3.4. Here we show that a novel Na + , Li + ‐permeable and blocker‐resistant pathway exists in an inherited mutant, Kir3.2 G156S, in addition to the conventional ion conducting pathway formed by the selectivity filter (SF). Our results demonstrate that the mechanism underlying the loss of K + selectivity of Kir3.2 G156S involves formation of a novel ion permeation pathway besides the SF pathway, which allows permeation of various species of cations.