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

Abstract

Local anaesthetics such as lidocaine (lignocaine) interact with sodium channels in a manner that is exquisitely sensitive to the voltage‐dependent conformational state of the ion channel. When depolarized in the presence of lidocaine, sodium channels assume a long‐lived quiescent state. Although studies over the last decade have localized the lidocaine receptor to the inner aspect of the aqueous pore, the mechanistic basis of depolarization‐induced ‘use‐dependent’ lidocaine block remains uncertain. Recent studies have shown that lowering the extracellular Na+ concentration ([Na+]o) and mutations in the sodium channel outer P‐loop modulate occupancy of a quiescent ‘slow’ inactivated state with intermediate kinetics (termed IM) that involves structural rearrangements in the outer pore. Site‐directed mutagenesis and ion‐replacement experiments were performed using voltage‐clamped Xenopus oocytes and cultured (HEK‐293) cells expressing wild‐type and mutant rat skeletal muscle (μ1) sodium channels. Our results show that lowering [Na+]o potentiates use‐dependent lidocaine block. The effect of [Na+]o is maintained despite a III‐IV linker mutation that partially disrupts fast inactivation (F1304Q). In contrast, the effect of lowering [Na+]o on lidocaine block is reduced by a P‐loop mutation (W402A) that limits occupancy of IM. 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.

Ion conduction and conformational flexibility of a bacterial voltage-gated sodium channel

“…Fenestration size is modulated by S6 bending, which we have proposed may be associated with a collapse into an inactivated state. These observations suggest that the binding of drugs at this crucial site could modulate conversion to a slowinactivated state, as suggested experimentally (28,31,32,46). These simulations have therefore proven to be important for exploring the roles of protein flexibility in ion conduction mechanisms and have given us clues as to the molecular-level processes that underscore inactivation and drug inhibition of Na v channels.…”

“…Mutation of the helix-weakening T206 (T220 in NaChBac) prevents slow inactivation (43), whereas the mutation of the equivalent of residue I202 in Nav1.4 domain IV dramatically affects slow inactivation (44). In addition, mutation of Phe at that same level (F203 in Na v Ab) strongly inhibits slow inactivation in mammalian Na v (28,31), reinforcing the significance of the S6 bend around this residue. It is also of much interest that changes in gateforming helices are correlated with changes in the SF, as suggested by NaChBac studies (10), and akin to C-type (slow) inactivation in K + channels (27), although apparently via a different mechanism.…”

“…It has been suggested that drugs can enter the cavity (48) and physically block the channel (49), or bind at the level of the opening via π-π stacking of aromatics (47) and inhibit the current by favoring a nonconductive inactivated state (46). Moreover, it is known that slow inactivation is affected by local anesthetics or antiepileptics drugs (30)(31)(32) and by the mutation of the same Phe involved in drug binding (28). Evidence suggests that drug binding affinity varies depending on the conducting state of the channels (46), corroborating our own observations that accessibility of both the Phe and cavity openings are codependent and correlated with changes in the S6 helix and gate collapse, suggesting a transition to the inactivated state.…”

“…Bacterial channels display slow inactivation reminiscent of C-type inactivation in K v channels (27), involving its PD (28,29), and is known to be modulated by local anesthetics (8). Interestingly, there is recent evidence that mammalian channels are also affected by this process (30)(31)(32), despite a historical focus on fast inactivation (33). Our multimicrosecond simulations reveal conformational fluctuations in common with proposed inactivated crystal structures (34), which allow us to begin to describe the molecular mechanisms of inactivation and its relation to Na v drug inhibition.…”

Voltage-Gated Sodium Channels

“…The open fenestrations have a maximum ~4 Å width, feasible for the entry of lipophilic drugs (i.e. local anesthetics), suggesting binding at this site may influence slow channel inactivation propagated through this region from the selectivity filter (Carboni et al, 2005;Z. Chen et al, 2000;Li et al, 1999).…”