Vol. 60, Issue 6, 1365-1374, December 2001

Inhibition of Wild-Type and Mutant Neuronal Nicotinic Acetylcholine Receptors by Local Anesthetics

Roger L. Papke, Benjamin A. Horenstein, and Andon N. Placzek

Departments of Pharmacology & Therapeutics (R.L.P., A.N.P.) and Chemistry (B.A.H.), University of Florida, Gainesville, Florida

Inhibition of neuronal nicotinic receptors can be regulated by the presence of specific amino acids in the  subunit second transmembrane domain (TM2) domain. We show that the incorporation of a mutant 4 subunit, which contains sequence from the muscle  subunit at the TM2 6' and 10' positions of the neuronal 4 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 34 receptors and 34(6'F10'T) receptors by QX-314 is voltage-dependent and noncompetitive. Interestingly, the potency of the local anesthetic tetracaine for the inhibition of 34 and 34(6'F10'T) receptors seems unchanged when measured at 50 mV. However, whereas the onset of inhibition of wild-type 34 receptors is voltage-dependent and noncompetitive, the onset of inhibition of 34(6'F10'T) receptors by tetracaine is unaffected by membrane voltage, and at concentrations 30 µM 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.