Inhibition of wild-type and mutant neuronal nicotinic acetylcholine receptors by local anesthetics.

Inhibition of neuronal nicotinic receptors can be regulated by the presence of specific amino acids in the beta subunit second transmembrane domain (TM2) domain. We show that the incorporation of a mutant beta4 subunit, which contains sequence from the muscle beta subunit at the TM2 6' and 10' positions of the neuronal beta4 subunit, greatly reduces the sensitivity of receptors to the local anesthetic [2-(triethylamino)-N-(2,6-dimethylphenyl)acetamide] (QX-314). Although differing in potency, the inhibition of both wild-type alpha3beta4 receptors and alpha3beta4(6'F10'T) receptors by QX-314 is voltage-dependent and noncompetitive. Interestingly, the potency of the local anesthetic tetracaine for the inhibition of alpha3beta4 and alpha3beta4(6'F10'T) receptors seems unchanged when measured at -50 mV. However, whereas the onset of inhibition of wild-type alpha3beta4 receptors is voltage-dependent and noncompetitive, the onset of inhibition of alpha3beta4(6'F10'T) receptors by tetracaine is unaffected by membrane voltage, and at concentrations < or = 30 microM seems to be competitive with acetylcholine. This may be due to either direct effects of tetracaine at the acetylcholine binding site or preferential block of closed rather than open channels in the mutant receptors. Further analysis of receptors containing the 6' mutation alone suggests that although the 6' mutation is adequate to alter the voltage dependence of tetracaine inhibition, both point mutations are required to produce the apparent competitive effects.