Voltage-gated sodium channel-associated proteins and alternative mechanisms of inactivation and block

Abstract

Voltage-gated sodium channels mediate inward current of action potentials upon membrane depolarization of excitable cells. The initial transient sodium current is restricted to milliseconds through three distinct channel-inactivating and blocking mechanisms. All pore-forming alpha subunits of sodium channels possess structural elements mediating fast inactivation upon depolarization and recovery within milliseconds upon membrane repolarization. Accessory subunits modulate fast inactivation dynamics, but these proteins can also limit current by contributing distinct inactivation and blocking particles. A-type isoforms of fibroblast growth factor homologous factors (FHFs) bear a particle that induces long-term channel inactivation, while sodium channel subunit Navβ4 employs a blocking particle that rapidly dissociates upon membrane repolarization to generate resurgent current. Despite their different physiological functions, the FHF and Navβ4 particles have similarity in amino acid composition and mechanisms for docking within sodium channels. The three competing channel-inactivating and blocking processes functionally interact to regulate a neuron's intrinsic excitability.

Fig. 1

Topology and interactions of voltage-gated sodium channel alpha subunit, beta subunit, and FHF. Nav α The alpha subunit of each sodium channel (Nav1.1–Nav1.9) consists of four pseudo-homologous domains (D), each bearing six transmembrane helical segments (S) and partial reentrant loops between S5 and S6 that form the sodium ion selectivity filter. The S4 segments (blue) have periodic cationic residues that constitute the voltage sensors, while the S6 segments (red) define the inner walls of the channel beneath the ion selectivity filter. The channel has cytoplasmic amino (N) and carboxyl (C) tails, and the four domains are connected by cytoplasmic loops (not to scale). The small DIII/DIV loop includes a short α-helical region and the IFM triad required for fast inactivation. The C-tail critically modulates fast inactivation and is the site for physical interactions with channel β subunits and FHFs (arrows). Nav β Beta subunits (Nav2.1–Nav2.4) are single-pass transmembrane proteins with an extracellular immunoglobulin-like domain and a short cytoplasmic tail that mediates binding to Nav α. Navβ4 bears a unique motif in its C-tail that serves as a channel-blocking particle responsible for resurgent current. FHF FHFs (FHF1–FHF4, each with multiple isoforms bearing different N-termini) are small cytoplasmic proteins that assume a β-trefoil fold bearing the surface for interaction with the Nav α C-tail. The A-type isoforms of FHFs have N-terminal motifs that serve as long-term inactivation particles. Navβ4 and A-type FHF particles terminate transient sodium current by competing with the Nav α fast inactivation machinery

Fig. 2

Sodium channel gating. a Schematic flow diagram of fast sodium channel dynamics. Voltage-driven conformation transitions (thick arrow) lead to channel activation upon depolarization (rightward, unfilled) and deactivation at more negative membrane potential (leftward, solid). Activating transitions enable intrinsic fast inactivation (downward arrows) from closed or open states, from which channels recover upon sufficient deactivating transitions. A-type FHFs enable alternative long-term inactivation (upward arrows) from near-open or open states. Navβ4 mediates channel block (diagonal arrow) at positive membrane potential, and expulsion of the blocking particle as membrane potential trends negative channels generates transient resurgent current. b Markov state modeling of activation, inactivation, and block. Voltage-driven channel transitions are along the horizontal axis, while voltage-independent transitions are shown on orthogonal axes. For voltage-independent transitions, relative sizes of arrowheads indicate favored direction of transition. Black Depolarizing membrane potential drives channels through closed states (C) towards an open conducting state (O), with concomitant increasing rates for transition to inactivated states (I) [11]. Sufficiently strong depolarization thereby generates transient sodium current before inactivation. Upon return to more negative potentials, inactivated channels deactivate (I6 → I5 → etc.) before recovering from inactivation (I → C), so that channels do not conduct as they reset. Red A-type FHFs mediate an alternative competing mechanism for transitions to long-term inactivated states (L) [45]. Upon return to more negative potentials, deactivation rates (L6 → L5 → etc.) are far slower than following fast inactivation, increasing the time until channels are again available. Green Positive membrane potentials approaching the Na⁺ reversal potential limits sodium flux through open channels, generating a permissive state (Op) for block by Navβ4 (OpB). Subsequent return to negative potentials allows inward sodium permeation (OB), causing expulsion of the blocking particle (OB → O) and consequent resurgent current [15, 68]. Blue Additional states added to reflect the ability of open channels to transit to any of the three non-conducting states (Ip6, Lp6, OpB)