Because alcohol activation is a property of most types of GIRK channels7,8, we reasoned that a homologous mutation in a related GIRK channel would also alter the response to alcohols. To test this idea, we investigated the effects of mutating L252 in GIRK4*. GIRK4* contains a mutation in the pore-helix (S143T) that enhances channel activity without affecting surface expression27. Substituting Ala (26 Å3), Tyr (133 Å3) or Trp (168 Å3) at L252 in GIRK4* channels did not change the basal K+ currents (Fig. 6a). Similar to mutations of L257 in GIRK2, Trp and Tyr substitutions in GIRK4* decreased EtOH, 1-PrOH and MPD activation, as compared to L252A, with 1-PrOH now inhibiting GIRK4*-L252W (Fig. 6b–f). In contrast to GIRK2, however, MPD activation of GIRK4*-L252W was not significantly different from wild-type (Fig. 6e,f). Mutating GIRK4*-L252 also significantly reduced m2R-activated GIRK currents (Fig. 6c–f). Thus, the putative hydrophobic alcohol-binding pocket in GIRK4* is important for mediating alcohol activation but GIRK4* may accommodate MPD differently than GIRK2 (see Discussion).
