Small interfering RNA-mediated selective knockdown of NaV1.8 tetrodotoxin-resistant sodium channel reverses mechanical allodynia in neuropathic rats

Abstract

The biophysical properties of a tetrodotoxin resistant (TTXr) sodium channel, NaV1.8, and its restricted expression to the peripheral sensory neurons suggest that blocking this channel might have therapeutic potential in various pain states and may offer improved tolerability compared with existing sodium channel blockers. However, the role of NaV1.8 in nociception cannot be tested using a traditional pharmacological approach with small molecules because currently available sodium channel blockers do not distinguish between sodium channel subtypes. We sought to determine whether small interfering RNAs (siRNAs) might be capable of achieving the desired selectivity. Using Northern blot analysis and membrane potential measurement, several siRNAs were identified that were capable of a highly-selective attenuation of NaV1.8 message as well as functional expression in clonal ND7/23 cells which were stably transfected with the rat NaV1.8 gene. Functional knockdown of the channel was confirmed using whole-cell voltage-clamp electrophysiology. One of the siRNA probes showing a robust knockdown of NaV1.8 current was evaluated for in vivo efficacy in reversing an established tactile allodynia in the rat chronic constriction nerve-injury (CCI) model. The siRNA, which was delivered to lumbar dorsal root ganglia (DRG) via an indwelling epidural cannula, caused a significant reduction of NaV1.8 mRNA expression in lumbar 4 and 5 (L4–L5) DRG neurons and consequently reversed mechanical allodynia in CCI rats. We conclude that silencing of NaV1.8 channel using a siRNA approach is capable of producing pain relief in the CCI model and further support a role for NaV1.8 in pathological sensory dysfunction.