Inhibition of Navβ4 Peptide-Mediated Resurgent Sodium Currents in Nav1.7 Channels by Carbamazepine, Riluzole, and Anandamide

Abstract

Paroxysmal extreme pain disorder (PEPD) and inherited erythromelalgia (IEM) are inherited pain syndromes arising from different sets of gain-of-function mutations in the sensory neuronal sodium channel isoform Nav1.7. Mutations associated with PEPD, but not IEM, result in destabilized inactivation of Nav1.7 and enhanced resurgent sodium currents. Resurgent currents arise after relief of ultra-fast open-channel block mediated by an endogenous blocking particle and are thought to influence neuronal excitability. As such, enhancement of resurgent currents may constitute a pathological mechanism contributing to sensory neuron hyperexcitability and pain hypersensitivity associated with PEPD. Furthermore, pain associated with PEPD, but not IEM, is alleviated by the sodium channel inhibitor carbamazepine. We speculated that selective attenuation of PEPD-enhanced resurgent currents might contribute to this therapeutic effect. Here we examined whether carbamazepine and two other sodium channel inhibitors, riluzole and anandamide, exhibit differential inhibition of resurgent currents. To gain further insight into the potential mechanism(s) of resurgent currents, we examined whether these inhibitors produced correlative changes in other properties of sodium channel inactivation. Using stably transfected human embryonic kidney 293 cells expressing wild-type Nav1.7 and the PEPD mutants T1464I and M1627K, we examined the effects of the three drugs on Nav␤4 peptide-mediated resurgent currents. We observed a correlation between resurgent current inhibition and a drug-mediated increase in the rate of inactivation and inhibition of persistent sodium currents. Furthermore, although carbamazepine did not selectively target resurgent currents, anandamide strongly inhibited resurgent currents with minimal effects on the peak transient current amplitude, demonstrating that resurgent currents can be selectively targeted.

Sidedness of Carbamazepine Accessibility to Voltage-Gated Sodium Channels

“…1D, n 5 4), tested with a protocol using 30-millisecond test pulses to 220 mV delivered every 3 seconds from a holding potential of 275 mV. As expected, carbamazepine block was strongly use dependent (Willow et al, 1985;Theile and Cummins, 2011), with block greatly enhanced by stimulating at 10, rather than 0.33 Hz, and was also strongly voltage dependent (Kuo et al, 1997;Kuo, 1998), with the steadystate inactivation curve shifted approximately 20 mV in the hyperpolarizing direction by 100 mM carbamazepine (data not shown).…”

The Role of Non-pore-Forming β Subunits in Physiology and Pathophysiology of Voltage-Gated Sodium Channels

“…Subsequent knockdown of β4 expression using siRNA in CGNs resulted in reduced resurgent sodium current and decreased repetitive firing, providing further evidence for β4 as the open-channel blocking particle (Bant and Raman 2010). A peptide corresponding to β4 was sufficient to invoke resurgent sodium current in HEK 293 cells (Theile and Cummins 2011), again strongly suggesting a role for β4 in the generation of resurgent sodium current in vivo. Importantly, however, co-expression of full-length β4 and Nav1.6 subunits was not sufficient to generate resurgent current , suggesting that key cellular processes, possibly including posttranslational modification, that allow β4 subunits to participate as the open-channel blocking particle may be specific to neurons.…”

CRMP2 Protein SUMOylation Modulates NaV1.7 Channel Trafficking

“…7 Cell Line in a Ubc9-dependent Manner-Because we could not pharmacologically or genetically isolate NaV1.1 and NaV1.3 currents, which account for Ͻ15% of the CAD cell sodium current, we chose to test the importance of CRMP2-SUMOylation in NaV1.7 trafficking in a HEK293 cell line expressing only NaV1.7 channels (38). Representative traces of NaV1.7 currents from these cells are illustrated in Fig.…”