Open-channel block by the cytoplasmic tail of sodium channel β4 as a mechanism for resurgent sodium current

Voltage-gated sodium channels with "resurgent" kinetics are specialized for high-frequency firing. The α subunits interact with a blocking protein that binds open channels upon depolarization and unbinds upon repolarization, producing resurgent sodium current. By limiting classical inactivation, the cycle of block and unblock shortens refractory periods. To characterize the blocker in Purkinje neurons, we briefly exposed inside-out patches to substrate-specific proteases. Trypsin and chymotrypsin each removed resurgent current, consistent with established roles for positively charged and hydrophobic/aromatic groups in blocking sodium channels. In Purkinje cells, the only known sodium channel-associated subunit that has a cytoplasmic sequence with several positive charges and clustered hydrophobic/aromatic residues is β4 (KKLITFILKKTREK; β4_154-167). After enzymatic removal of block, β4_154-167 fully reconstituted resurgent current, whereas scrambled or point-mutated peptides were ineffective. In CA3 pyramidal neurons, which lack β4 and endogenous block, β4_154-167 generated resurgent current. Thus, β4 may be the endogenous open-channel blocker responsible for resurgent kinetics.