A mutation in the local anaesthetic binding site abolishes toluene effects in sodium channels

Toluene is a solvent of abuse that inhibits cardiac sodium channels in a manner that resembles the action of local anaesthetics. The purpose of this work was to analyze toluene effects on skeletal muscle sodium channels with and without β1 subunit (Nav1.4 + β1 and Nav1.4 − β1, respectively) expressed in Xenopus laevis oocytes and to compare them with those produced in the F1579A mutant channel lacking a local anaesthetic binding site. Toluene inhibited Nav1.4 sodium currents (IC50 = 2.7 mM in Nav1.4 + β1 and 2.2 mM in Nav1.4 − β1) in a concentration dependent way. Toluene (3 mM) blocked sodium currents in Nav1.4 channels proportionally throughout the entire current-voltage relationship producing inactivation at more negative potentials. Minimal inhibition was produced by 3 mM toluene in F1579A mutant channels. Recovery from inactivation was slower both in Nav1.4 and F1579A channels in the presence of 3 mM toluene. The solvent blocked sodium currents in a use-dependent and frequency-dependent manner in Nav1.4 channels. A single mutation in the local anaesthetic binding site of Nav1.4 channels almost abolished toluene effects. These results suggest that this site is important for toluene action.