Differential sensitivity of TREK ‐1, TREK ‐2 and TRAAK background potassium channels to the polycationic dye ruthenium red

Background and Purpose Pharmacological separation of the background potassium currents of closely related K 2P channels is a challenging problem. We previously demonstrated that ruthenium red ( RR ) inhibits TASK ‐3 ( K 2 P 9.1), but not TASK ‐1 ( K 2 P 3.1) channels. RR has been extensively used to distinguish between TASK currents in native cells. In the present study, we systematically investigate the RR sensitivity of a more comprehensive set of K 2 P channels. Experimental Approach K + currents were measured by two‐electrode voltage clamp in X enopus oocytes and by whole‐cell patch clamp in mouse dorsal root ganglion ( DRG ) neurons. Key Results RR differentiates between two closely related members of the TREK subfamily. TREK ‐2 ( K 2 P 10.1) proved to be highly sensitive to RR (IC 50 = 0.2 μM), whereas TREK ‐1 ( K 2 P 2.1) was not affected by the compound. We identified aspartate 135 ( D 135) as the target of the inhibitor in mouse TREK ‐2c. D 135 lines the wall of the extracellular ion pathway ( EIP ), a tunnel structure through the extracellular cap characteristic for K 2 P channels. TREK ‐1 contains isoleucine in the corresponding position. The mutation of this isoleucine ( I 110 D ) rendered TREK ‐1 sensitive to RR . The third member of the TREK subfamily, TRAAK ( K 2 P 4.1) was more potently inhibited by ruthenium violet, a contaminant in some RR preparations, than by RR . DRG neurons predominantly express TREK ‐2 and RR ‐resistant TREK ‐1 and TRESK ( K 2 P 18.1) background K + channels. We detected the RR ‐sensitive leak K + current component in DRG neurons. Conclusions and Implications We propose that RR may be useful for distinguishing TREK ‐2 (K 2P 10.1) from TREK ‐1 (K 2P 2.1) and other RR ‐resistant K 2 P channels in native cells.