Inhalational anaesthetics and n‐alcohols share a site of action in the neuronal Shaw2 Kv channel

Background and purpose: Neuronal ion channels are key targets of general anaesthetics and alcohol, and binding of these drugs to pre‐existing and relatively specific sites is thought to alter channel gating. However, the underlying molecular mechanisms of this action are still poorly understood. Here, we investigated the neuronal Shaw2 voltage‐gated K+ (Kv) channel to ask whether the inhalational anaesthetic halothane and n‐alcohols share a binding site near the activation gate of the channel. Experimental approach: Focusing on activation gate mutations that affect channel modulation by n‐alcohols, we investigated n‐alcohol‐sensitive and n‐alcohol‐resistant Kv channels heterologously expressed in Xenopus oocytes to probe the functional modulation by externally applied halothane using two‐electrode voltage clamping and a gas‐tight perfusion system. Key results: Shaw2 Kv channels are reversibly inhibited by halothane in a dose‐dependent and saturable manner (K0.5= 400 µM; nH= 1.2). Also, discrete mutations in the channel's S4S5 linker are sufficient to reduce or confer inhibition by halothane (Shaw2‐T330L and Kv3.4‐G371I/T378A respectively). Furthermore, a point mutation in the S6 segment of Shaw2 (P410A) converted the halothane‐induced inhibition into halothane‐induced potentiation. Lastly, the inhibition resulting from the co‐application of n‐butanol and halothane is consistent with the presence of overlapping binding sites for these drugs and weak binding cooperativity. Conclusions and implications: These observations strongly support a molecular model of a general anaesthetic binding site in the Shaw2 Kv channel. This site may involve the amphiphilic interface between the S4S5 linker and the S6 segment, which plays a pivotal role in Kv channel activation.