Reduced excitability and impaired nociception in peripheral unmyelinated fibers from Nav1.9-null mice.

The upregulation of the tetrodotoxin-resistant voltage-gated sodium channel NaV1.9 has previously been associated with inflammatory hyperalgesia. Na1.9 knockout (KO) mice, however, did not seem insensitive in conventional tests of acute nociception. Using electrophysiological, neurochemical, and behavioral techniques, we now show NaV1.9-null mice exhibit impaired mechanical and thermal sensory capacities and reduced electrical excitability of nociceptors. In single-fiber recordings from isolated skin, the electrical threshold of NaV1.9 KO C fibers was elevated by 55% and the median von Frey threshold was 32 mN in contrast to 8 mN in wild types (WTs). The prevalence of C mechano-heat-sensitive (CMH) fibers was only 25.6% in NaV1.9 KO animals compared to 75.8% in the WT group, and the heat threshold of these CMH fibers was 40.4°C in the control vs 44°C in the KO group. Compound action potential recordings from isolated sciatic nerve segments of NaV1.9 KO mice revealed lower activity-induced slowing of conduction velocity upon noxious heat stimulation: 8% vs 30% in WTs. Heat-induced calcitonin gene-related peptide release from the skin was less in the KO than in the WT group. The reduced noxious heat sensitivity was finally confirmed with the Hargreaves test using 2 rates of radiant heating of the plantar hind paws. In conclusion, NaV1.9 presumably contributes to acute thermal and mechanical nociception in mice, most likely through increasing the excitability but probably also by amplifying receptor potentials irrespective of the stimulus modality.

[Inhibitory effect of NaV1.9 gene silencing on proliferation, phagocytosis and migration in RAW264.7 cells].

OBJECTIVE: To establish the cell line with stable voltage-gated sodium channels (VGSCs/NaVs) α subunit NaV1.9 gene silencing through RNA interference (RNAi) in murine RAW264.7 macrophages, and to investigate proliferation, phagocytosis and migration in this cell line. METHODS: The stable NaV1.9-deficient cell line was generated by selection in G418 after the transfection of short hairpin (shRNA) plasmid with Lipofectamine TM2000. RNAi efficiency was qualified by RT-PCR; proliferation ability was measured by CCK-8 assay; cell cycle and phagocytic ability were analyzed by flow cytometry; and migrating ability was detected by Transwell migration assay. RESULTS: Stable NaV1.9-deficient cell line was established and the expression of NaV1.9 was reduced by 80%. CCK-8 assay and flow cytometry showed that the proliferation of the NaV1.9-deficient cell line was inhibited (P<0.05). Flow cytometry revealed that phagocytic ability was reduced in the cell line (P<0.05). Transwell migration assay demonstrated that migrating ability was depressed in the cell line (P<0.05). CONCLUSION: In the stable NaV1.9-deficient cells we successfully constructed, proliferation, phagocytosis and migration were obviously inhibited.