The n-alcohol site in the nicotinic receptor pore is a hydrophobic patch

Alcohols and volatile anesthetics inhibit peripheral nicotinic acetylcholin e receptors noncompetitively, primarily via an open-channel block mechanism . Analysis of hydrophobic mutations near the middle of the pore-forming M2 domains suggested that alcohols interact with the pore in this vicinity. To establish the extent of this inhibitory site, we created a series of hydro phobicity-altering mutations scanning most of the alpha subunit M2 domain. Using both single-channel and rapid patch perfusion electrophysiology, we m easured how these mutations affect nAChR sensitivity to ethanol and hexanol . We find a near-contiguous series of amino acids in alpha -M2, extending f rom alpha L250 (8') to alpha V255 (13'), where mutagenesis strongly influen ces inhibition by alcohols. These results support the existence of a large inhibitory patch in the nAChR pore lining where interactions with alcohols an primarily due to hydrophobic forces. Ethanol appears to interact with de eper regions of this site than does hexanol. Because alcohols apparently ac t as open-channel blockers, we infer from our results that most of the resi dues between alpha L250 and alpha V255 are exposed to the aqueous environme nt of the pore when the channel is open. The location and extent of this si te can explain why small alcohols occupy the nAChR pore at the same time as larger alcohols or charged blockers, while two large alcohols bind in a mu tually exclusive manner.