Calmodulin regulates current density and frequency-dependent inhibition of sodium channel Nav1.8 in DRG neurons

Sodium channel Na(v)1.8 produces a slowly inactivating, tetrodotoxin-resistant current, characterized by recovery from inactivation with fast and slow components, and contributes a substantial fraction of the current underlying the depolarizing phase of the action potential of dorsal root ganglion (DRG) neurons. Na(v)1.8 C-terminus carries a conserved calmodulin-binding isoleucine-glutamine (IQ) motif. We show here that calmodulin coimmunoprecipitates with endogenous Na(v)1.8 channels from native DRG, suggesting that the two proteins can interact in vivo. Treatment of native DRG neurons with a calmodulin-binding peptide (CBP) reduced the current density of Na(v)1.8 by nearly 65%, without changing voltage dependency of activation or steady-state inactivation. To investigate the functional role of CaM binding to the IQ motif in the Na(v)1.8 C-terminus, the IQ dipeptide was substituted by DE; we show that this impairs the binding of CaM to the IQ motif. Mutant Na(v)1.8IQ/DE channels produce currents with roughly 50% amplitude, but with unchanged voltage dependency of activation and inactivation when expressed in DRG neurons from Na(v)1.8-null mice. We also show that blocking the interaction of CaM and Na(v)1.8 using CBP or the IQ/DE substitution causes a buildup of inactivated channels and, in the case of the IQ/DE mutation, stimulation even at a low frequency of 0.1 Hz significantly enhances the frequency-dependent inhibition of the Na(v)1.8 current. This study presents, for the first time, evidence that calmodulin associates with a sodium channel, Na(v)1.8, in native neurons, and demonstrates a regulation of Na(v)1.8 currents that can significantly affect electrogenesis of DRG neurons in which Na(v)1.8 is normally expressed.