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The voltage-gated sodium channel Nav1.7 is encoded by SCN9A gene and plays a critical role in pain sensitivity. Several SCN9A gain-of-function (GOF) mutations have been found in patients with small fiber neuropathy (SFN) having chronic pain, including the R185H mutation. However, for most of these variants, their involvement in pain phenotype still needs to be experimentally elucidated. In order to delineate the impact of R185H mutation on pain sensitivity, we have established the Scn9aR185H mutant mouse model using the CRISPR/Cas9 technology. The Scn9aR185H mutant mice show no cellular alteration in the dorsal root ganglia (DRG) containing cell bodies of sensory neurons and no alteration of growth or global health state. Heterozygous and homozygous animals of both sexes were investigated for pain sensitivity. The mutant mice were more sensitive than the wild-type mice in the tail flick and hot plate tests, acetone, and von Frey tests for sensitivity to heat, cold, and touch, respectively, although with sexual dimorphic effects. The newly developed bioinformatic pipeline, Gdaphen is based on general linear model (GLM) and random forest (RF) classifiers as well as a multifactor analysis of mixed data and shows the qualitative and quantitative variables contributing the most to the pain phenotype. Using Gdaphen, tail flick, Hargreaves, hot plate, acetone, cold plate, and von Frey tests, sex and genotype were found to be contributing most to the pain phenotype. Importantly, the mutant animals displayed spontaneous pain as assessed in the conditioned place preference (CPP) assay. Altogether, our results indicate that Scn9aR185H mice show a pain phenotype, suggesting that the SCN9AR185H mutation identified in patients with SFN having chronic pain contributes to their symptoms. Therefore, we provide genetic evidence for the fact that this mutation in Nav1.7 channel plays an important role in nociception and in the pain experienced by patients with SFN who have this mutation. These findings should aid in exploring further pain treatments based on the Nav1.7 channel.

The voltage-gated sodium channel NAV1.8 is expressed in primary nociceptive neurons and is involved in pain transmission. Mutations in the SCN10A gene (encoding NAV1.8 channel) have been identified in patients with idiopathic painful small fiber neuropathy (SFN) including the SCN10AG1662S gain-of-function mutation. However, the role of this mutation in pain sensation remains unknown. We have generated the first mouse model for the G1662S mutation by using homologous recombination in embryonic stem cells. The corresponding Scn10aG1663S mouse line has been analyzed for Scn10a expression, intraepidermal nerve fiber density (IENFD), and nociception using behavioral tests for thermal and mechanical sensitivity. The Scn10aG1663S mutants had a similar Scn10a expression level in dorsal root ganglia (DRG) to their wild-type littermates and showed normal IENFD in hindpaw skin. Mutant mice were more sensitive to touch than wild types in the von Frey test. In addition, sexual dimorphism was observed for several pain tests, pointing to the relevance of performing the phenotypical assessment in both sexes. Female homozygous mutants tended to be more sensitive to cooling stimuli in the acetone test. For heat sensitivity, male homozygous mutants showed shorter latencies to radiant heat in the Hargreaves test while homozygous females had longer latencies in the tail flick test. In addition, mutant males displayed a shorter reaction latency on the 54°C hot plate. Collectively, Scn10aG1663S mutant mice show a moderate but consistent increased sensitivity in behavioral tests of nociception. This altered nociception found in Scn10aG1663S mice demonstrates that the corresponding G1662 mutation of SCN10A found in SFN patients with pain contributes to their pain symptoms.