Calmodulin Regulates Current Density and Frequency-Dependent Inhibition of Sodium Channel Na_v1.8 in DRG Neurons

Abstract

-Hajj. Calmodulin regulates current density and frequency-dependent inhibition of sodium channel Na v 1.8 in DRG neurons. J Neurophysiol 96: 97-108, 2006. First published April 5, 2006 doi:10.1152/jn.00854.2005. 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

Aquaporin-1 Tunes Pain Perception by Interaction with Nav1.8 Na+ Channels in Dorsal Root Ganglion Neurons

“…Slow inactivation has been reported in various mammalian neuronal cells including rat hippocampal neurons (48), tetrodotoxin-resistant DRG neurons (23), and cultured neuroblastoma cells (49) and is important to firing adaptation. Several molecular partners and membrane proteins have been reported to modulate Na ϩ channel slow inactivation, including ␤ 1-4 subunits, ankyrin G (50), and calmodulin (25). Our data suggest selective AQP1 modulation of Na v 1.8 but not Na v 1.7.…”

“…Na ϩ channels Na v 1.1 to Na v 1.9 are composed of a large ␣ subunit containing the pore-and voltage-sensing machinery and several auxiliary ␤ subunits (50). Only a few other proteins have been reported to interact directly with Na v 1.8, including annexin II light chain (51), calmodulin (25), and clathrin-associated protein-1A (42). The immunoprecipitation and single molecule tracking indicated a physical interaction between AQP1 and Na v 1.8.…”

“…Use-dependence of Na ϩ channels was studied using repetitive 30-ms depolarizing pulses from Ϫ80 to 0 mV with different frequencies (5, 10, or 20 Hz). The reduction in peak I Na at the 40th pulse compared with the 1st pulse was taken as a measure of use-dependent channel inactivation (25,26).…”

A rescuable folding defective Nav1.1 (SCN1A) sodium channel mutant causes GEFS+: Common mechanism in Nav1.1 related epilepsies?

“…1C), is an important residue of the calmodulin binding IQ motif (Bahler and Rhoads, 2002), which is implicated in the modulation of several Na + channel isoforms (Young and Caldwell, 2005;Choi et al, 2006;Biswas et al, 2008), and we have shown that calmodulin can rescue hNa v 1.1-M1841T (Rusconi et al, 2007). Although R1916G should inhibit calmodulin binding, we tested the effect of cotransfection of calmodulin and found, indeed, that there was no significant modification of hNa v 1.1-R1916G current density ( Fig.…”

“…In fact, it has been shown that calmodulin can differentially modulate several Na + channel isoforms, inducing functional effects that depend on the isoform and on the cell type used for the experiments (Young and Caldwell, 2005;Choi et al, 2006;Biswas et al, 2008). However, both reduction of cell surface expression and negative shift of the inactivation curve, which are two of the effects caused by R1916G, have been observed blocking the binding of calmodulin to Na + channels (Choi et al, 2006;Biswas et al, 2008). In our experiments these effects may be induced by the inhibition of the binding of endogenous calmodulin.…”