Lidocaine induces a slow inactivated state in rat skeletal muscle sodium channels

Lidocaine (lignocaine) and its analogues, compounds that suppress Na+ current (INa), are used as local anaesthetics and as therapies for cardiac arrhythmias and epilepsy. When depolarized, Na+ channels only transiently conduct ions due to rapidly developing (fast) inactivation (Hodgkin & Huxley, 1952), a conformational change involving amino acid residues in the III-IV interdomain linker (Stuhmer et al. 1989; West et al. 1992) and vicinal sites near the inner surface of the channel (McPhee et al. 1994, 1995, 1998; Smith & Goldin, 1997; Lerche et al. 1997; Filatov et al. 1998; Tang et al. 1998). While drug-free Na+ channels quickly recover from fast inactivation between depolarizing stimuli, lidocaine delays such recovery, inducing a stable non-conducting state that results in cumulative reduction in INa with brief, repetitive stimuli (so-called ‘use-dependent block’) (Courtney, 1975). Lidocaine binds to amino acids positioned on the intracellular side of the putative selectivity filter (Ragsdale et al. 1994, 1996; Sunami et al. 1997), near (and perhaps overlapping) the fast-inactivation gating structures (Bennett et al. 1995). A high-affinity binding interaction between lidocaine and the fast inactivated conformational state is a structural hypothesis for use-dependent local anaesthetic block supported by compelling, but circumstantial, experimental evidence (Hille, 1977; Hondeghem & Katzung, 1977; Cahalan, 1978; Yeh, 1978; Bean et al. 1983; Balser et al. 1996b). Surprisingly, a recent study found that amino acid residues intimately involved in fast inactivation recover to their pre-inactivated conformation rapidly, even while conduction is fully suppressed by lidocaine (Vedantham & Cannon, 1999), casting doubt on the prevailing notion that lidocaine arrests Na+ channels in the fast inactivated conformational state. In rare circumstances such as prolonged depolarization, Na+ channels may assume slow inactivated conformations from which recovery is considerably delayed (Adelman & Palti, 1969; Chandler & Meves, 1970; Rudy, 1978). One such slow inactivated state with ‘intermediate’ kinetics (Cummins & Sigworth, 1996; Hayward et al. 1996) is influenced by externally positioned residues in the outer pore region (μ1-W402) (Balser et al. 1996a; Kambouris et al. 1998; Benitah et al. 1999). Earlier studies (Cahalan & Almers, 1979) suggested that external Na+ opposed the use-dependent action of charged (quaternary) lidocaine derivatives through a competitive ‘knock-off’ mechanism. However, recent studies have shown that external Na+ also prevents Na+ channels from entering slow inactivated states (Townsend & Horn, 1997), including an inactivated state of ‘intermediate’ stability (denoted IM) (Benitah et al. 1999). Moreover, earlier studies proposed that local anaesthetic action involves a gating conformational change linked to a stable inactivated state (Khodorov et al. 1976; Zilberter et al. 1991), and we recently showed that an outer pore (P-loop) mutation that opposes IM also reduces use-dependent lidocaine action (Kambouris et al. 1998). Here we examine the possibility that this gating effect of external Na+, inhibition of IM, plays a role in Na+ antagonism of use-dependent lidocaine action. Our data show that augmenting slow inactivation by lowering the extracellular Na+ concentration ([Na+]o) sensitizes the Na+ channel to use-dependent lidocaine action, even when fast inactivation is disabled (μ1-F1304Q). Moreover, the effects of lowering [Na+]o on slow inactivation and lidocaine action are decreased by the P-loop mutation that opposes IM (W402A). Our findings are consistent with a simple allosteric model where lidocaine binding induces channels to occupy a native slow inactivated state that is inhibited by [Na+]o.