Permeant Ion-Dependent Changes in Gating of Kir2.1 Inward Rectifier Potassium Channels

We studied the effect of monovalent thallium ion (Tl⁺) on the gating of single Kir2.1 channels, which open and close spontaneously at a constant membrane potential. In cell-attached recordings of single-channel inward current, changing the external permeant ion from K⁺ to Tl⁺ decreases the mean open-time by ∼20-fold. Furthermore, the channel resides predominantly at a subconductance level, which results from a slow decay (τ = 2.7 ms at −100 mV) from the fully open level immediately following channel opening. Mutation of a pore-lining cysteine (C169) to valine abolishes the slow decay and subconductance level, and single-channel recordings from channels formed by tandem tetramers containing one to three C169V mutant subunits indicate that Tl⁺ must interact with at least three C169 residues to induce these effects. However, the C169V mutation does not alter the single-channel closing kinetics of Tl⁺ current. These results suggest that Tl⁺ ions change the conformation of the ion conduction pathway during permeation and alter gating by two distinct mechanisms. First, they interact with the thiolate groups of C169 lining the cavity to induce conformational changes of the ion passageway, and thereby produce a slow decay of single-channel current and a dominant subconductance state. Second, they interact more strongly than K⁺ with the main chain carbonyl oxygens lining the selectivity filter to destabilize the open state of the channel and, thus, alter the open/close kinetics of gating. In addition to altering gating, Tl⁺ greatly diminishes Ba²⁺ block. The unblocking rate of Ba²⁺ is increased by >22-fold when the external permeant ion is switched from K⁺ to Tl⁺ regardless of the direction of Ba²⁺ exit. This effect cannot be explained solely by ion–ion interactions, but is consistent with the notion that Tl⁺ induces conformational changes in the selectivity filter.