Voltage-gated Sodium Channel Subtype Nav1 Research Articles

Therapeutic targeting of voltage-gated sodium channel NaV1.7 for cancer metastasis

This review focuses on the expression and function of voltage-gated sodium channel subtype NaV1.7 in various cancers and explores its impact on the metastasis driving cell functions such as proliferation, migration, and invasiveness. An overview of its structural characteristics, drug binding sites, inhibitors and their likely mechanisms of action are presented. Despite the lack of clarity on the precise mechanism by which NaV1.7 contributes to cancer progression and metastasis; many studies have suggested a connection between NaV1.7 and proteins involved in multiple signaling pathways such as PKA and EGF/EGFR-ERK1/2. Moreover, the functional activity of NaV1.7 appears to elevate the expression levels of MACC1 and NHE-1, which are controlled by p38 MAPK activity, HGF/c-MET signaling and c-Jun activity. This cascade potentially enhances the secretion of extracellular matrix proteases, such as MMPs which play critical roles in cell migration and invasion activities. Furthermore, the NaV1.7 activity may indirectly upregulate Rho GTPases Rac activity, which is critical for cytoskeleton reorganization, cell adhesion, and actin polymerization. The relationship between NaV1.7 and cancer progression has prompted researchers to investigate the therapeutic potential of targeting NaV1.7 using inhibitors. The positive outcome of such studies resulted in the discovery of several inhibitors with the ability to reduce cancer cell migration, invasion, and tumor growth underscoring the significance of NaV1.7 as a promising pharmacological target for attenuating cancer cell proliferation and metastasis. The research findings summarized in this review suggest that the regulation of NaV1.7 expression and function by small molecules and/or by genetic engineering is a viable approach to discover novel therapeutics for the prevention and treatment of metastasis of cancers with elevated NaV1.7 expression.

Therapeutic targeting of voltage-gated sodium channel NaV1.7 for cancer metastasis.

This review focuses on the expression and function of voltage-gated sodium channel subtype NaV1.7 in various cancers and explores its impact on the metastasis driving cell functions such as proliferation, migration, and invasiveness. An overview of its structural characteristics, drug binding sites, inhibitors and their likely mechanisms of action are presented. Despite the lack of clarity on the precise mechanism by which NaV1.7 contributes to cancer progression and metastasis; many studies have suggested a connection between NaV1.7 and proteins involved in multiple signaling pathways such as PKA and EGF/EGFR-ERK1/2. Moreover, the functional activity of NaV1.7 appears to elevate the expression levels of MACC1 and NHE-1, which are controlled by p38 MAPK activity, HGF/c-MET signaling and c-Jun activity. This cascade potentially enhances the secretion of extracellular matrix proteases, such as MMPs which play critical roles in cell migration and invasion activities. Furthermore, the NaV1.7 activity may indirectly upregulate Rho GTPases Rac activity, which is critical for cytoskeleton reorganization, cell adhesion, and actin polymerization. The relationship between NaV1.7 and cancer progression has prompted researchers to investigate the therapeutic potential of targeting NaV1.7 using inhibitors. The positive outcome of such studies resulted in the discovery of several inhibitors with the ability to reduce cancer cell migration, invasion, and tumor growth underscoring the significance of NaV1.7 as a promising pharmacological target for attenuating cancer cell proliferation and metastasis. The research findings summarized in this review suggest that the regulation of NaV1.7 expression and function by small molecules and/or by genetic engineering is a viable approach to discover novel therapeutics for the prevention and treatment of metastasis of cancers with elevated NaV1.7 expression.

MAPK-ERK-CREB signaling pathway upregulates Nav1.6 in oxaliplatin-induced neuropathic pain in the rat

The voltage-gated sodium channel subtype Nav1.6 is involved in the electrophysiological changes of primary sensory neurons that occur in oxaliplatin-induced neuropathic pain, but its regulatory mechanism remains unclear. In this study, Western blot, RT-qPCR, immunofluorescence staining, chromatin immunoprecipitation were used to prove the mechanism of MAPK-ERK-CREB signaling pathway participating in oxaliplatin-induced neuropathic pain by regulating Nav1.6. The results showed that p-Raf1 and p-ERK, key molecules in MAPK/ERK pathway, and Nav1.6 were significantly increased in DRGs of oxaliplatin-induced neuropathic pain rats. Inhibition of p-Raf1 and p-ERK respectively not only reduced the expression of Nav1.6 protein in DRGs of OXA rats, but also caused a decrease in Nav1.6 mRNA, which led us to further explore the transcription factor CREB regulated by MAPK/ERK pathway. Results showed that CREB was co-distributed with Nav1.6. Inhibition of CREB resulted in decreased mRNA and protein expression of Nav1.6, and alleviated oxaliplatin-induced neuropathic pain. A chromatin immunoprecipitation experiment proved that OXA caused p-CREB to directly bind to the promoter region of Scn8A, which is the encoding gene for Nav1.6, and promote the transcription of Scn8A. In summary, in this study, we found that oxaliplatin can activate the MAPK/ERK pathway, which promotes the expression and activation of CREB and leads to an increase in Scn8A transcription, and then leads to an increase in Nav1.6 protein expression to enhance neuronal excitability and cause pain. This study provides an experimental basis for the molecular mechanism of sodium channel regulation in oxaliplatin-induced neuropathic pain.

Changes in Potency and Subtype Selectivity of Bivalent NaV Toxins are Knot-Specific.

Disulfide-rich peptide toxins have long been studied for their ability to inhibit voltage-gated sodium channel subtype NaV1.7, a validated target for the treatment of pain. In this study, we sought to combine the pore blocking activity of conotoxins with the gating modifier activity of spider toxins to design new bivalent inhibitors of NaV1.7 with improved potency and selectivity. To do this, we created an array of heterodimeric toxins designed to target human NaV1.7 by ligating a conotoxin to a spider toxin and assessed the potency and selectivity of the resulting bivalent toxins. A series of spider-derived gating modifier toxins (GpTx-1, ProTx-II, gHwTx-IV, JzTx-V, CcoTx-1, and Pn3a) and two pore-blocker μ-conotoxins, SxIIIC and KIIIA, were used for this study. We employed either enzymatic ligation with sortase A for C- to N-terminal ligation or click chemistry for N- to N-terminal ligation. The bivalent peptide resulting from ligation of ProTx-II and SxIIIC (Pro[LPATG6]Sx) was shown to be the best combination as native ProTx-II potency at hNaV1.7 was conserved following ligation. At hNaV1.4, a synergistic effect between the pore blocker and gating modifier toxin moieties was observed, resulting in altered sodium channel subtype selectivity compared to the parent peptides. Further studies including mutant bivalent peptides and mutant hNaV1.7 channels suggested that gating modifier toxins have a greater contribution to the potency of the bivalent peptides than pore blockers. This study delineated potential benefits and drawbacks of designing pharmacological hybrid peptides targeting hNaV1.7.

µ-Conotoxins Targeting the Human Voltage-Gated Sodium Channel Subtype NaV1.7

µ-Conotoxins are small, potent, peptide voltage-gated sodium (NaV) channel inhibitors characterised by a conserved cysteine framework. Despite promising in vivo studies indicating analgesic potential of these compounds, selectivity towards the therapeutically relevant subtype NaV1.7 has so far been limited. We recently identified a novel µ-conotoxin, SxIIIC, which potently inhibits human NaV1.7 (hNaV1.7). SxIIIC has high sequence homology with other µ-conotoxins, including SmIIIA and KIIIA, yet shows different NaV channel selectivity for mammalian subtypes. Here, we evaluated and compared the inhibitory potency of µ-conotoxins SxIIIC, SmIIIA and KIIIA at hNaV channels by whole-cell patch-clamp electrophysiology and discovered that these three closely related µ-conotoxins display unique selectivity profiles with significant variations in inhibitory potency at hNaV1.7. Analysis of other µ-conotoxins at hNaV1.7 shows that only a limited number are capable of inhibition at this subtype and that differences between the number of residues in loop 3 appear to influence the ability of µ-conotoxins to inhibit hNaV1.7. Through mutagenesis studies, we confirmed that charged residues in this region also affect the selectivity for hNaV1.4. Comparison of µ-conotoxin NMR solution structures identified differences that may contribute to the variance in hNaV1.7 inhibition and validated the role of the loop 1 extension in SxIIIC for improving potency at hNaV1.7, when compared to KIIIA. This work could assist in designing µ-conotoxin derivatives specific for hNaV1.7.

Bimodal Imaging of Mouse Peripheral Nerves with Chlorin Tracers.

Almost 17 million Americans have a history of cancer, a number expected to reach over 22 million by 2030. Cancer patients often undergo chemotherapy in the form of antineoplastic agents such as cis-platin and paclitaxel. Though effective, these agents can induce debilitating side effects; the most common neurotoxic effect, chemotherapy-induced peripheral neuropathy (CIPN), can endure long after treatment ends. Despite the widespread and chronic nature of the dysfunction, no tools exist to quantitatively measure chemotherapy-induced peripheral neuropathy. Such a tool would not only benefit patients but their stratification could also save significant financial and social costs associated with neuropathic pain. In our first step toward addressing this unmet clinical need, we explored a novel dual approach to localize peripheral nerves: Cerenkov luminescence imaging (CLI) and fluorescence imaging (FI). Our approach revolves around the targeting and imaging of voltage-gated sodium channel subtype NaV1.7, highly expressed in peripheral nerves from both harvested human and mouse tissues. For the first time, we show that Hsp1a, a radiolabeled NaV1.7-selective peptide isolated from Homoeomma spec. Peru, can serve as a targeted vector for delivering a radioactive sensor to the peripheral nervous system. In situ, we observe high signal-to-noise ratios in the sciatic nerves of animals injected with fluorescently labeled Hsp1a and radiolabeled Hsp1a. Moreover, confocal microscopy on fresh nerve tissue shows the same high ratios of fluorescence, corroborating our in vivo results. This study indicates that fluorescently labeled and radiolabeled Hsp1a tracers could be used to identify and demarcate nerves in a clinical setting.

Re-expression of voltage-gated sodium channel subtype Nav1.3 in the substantia nigra after dopamine depletion

Abnormal synchronized oscillatory bursts occurring in the basal ganglia (BG) are suggested to be correlated with motor symptoms in Parkinson’s disease (PD) patients and animal models of PD. Voltage-gated sodium channels (VGSCs) have been demonstrated to play an important role in the abnormal electrical activity of neurons in the BG. Nav1.3, a VGSCs subtype, is predominantly expressed in embryonic and neonatal nervous system but barely detected in the normal adult nervous system in rodents. Here we investigated the expression patterns of Nav1.3 in the BG of 6-OHDA lesioned Sprague Dawley rats. The results showed that Nav1.3 at mRNA and protein levels was abundantly re-expressed in the ipsilateral and contralateral SN at 49 days postlesion, but was rarely detected in the other nuclei of the BG in the 6-OHDA lesioned rats. Furthermore, Nav1.3 was not only expressed in TH-positive dopaminergic neurons of the ipsilateral and contralateral SN, but also in nestin-positive neural progenitor cells surrounding the ipsilateral SN and the midline region adjacent to the ipsilateral SN in the midbrain at 49 days postlesion. These results suggested that the re-expression of Nav1.3 may influence the electrical activity of dopaminergic neurons in the SN in 6-OHDA lesioned rats.

Assessing the Structural Basis of μ-Conotoxin KIIIA Inhibition of the Voltage-Gated Sodium Channel Nav1.7

The voltage-gated sodium channel subtype Nav1.7 has been shown to play a critical role in pain signaling, making it an important drug target. A number of peptide toxins from cone snails (conotoxins) inhibit sodium conduction through Nav1.7 with high affinity, and make potential scaffolds for selective channel inhibitors. However, the atomistic details of protein-protein interactions between conotoxins and Nav channels necessary for rational design of such inhibitors remain unclear. Our study aims to further define the molecular determinants of μ-conotoxin KIIIA action against the human Nav1.7 channel (hNav1.7). We have previously used Rosetta computational modeling software to generate preliminary molecular models of the hNav1.7 pore-forming domain based on the NavAb crystal structure (PDB ID: 3RVY). To capitalize on recent advances, we have updated our hNav1.7 homology model using the recent cryo-EM structure of the electric eel Nav1.4 channel (PDB ID: 5XSY) as a template. To test predicted pairwise contacts between toxin and channel, point mutations of hNav1.7 and conotoxin KIIIA in the putative binding site have been generated. We have identified several residues that alter the kinetics of toxin binding, notably acidic residues on the DII pore-helix. Our experimental and modeling results may be useful for rational design of novel peptides to inhibit hNav1.7 with high affinity and selectivity.

Activity and connectivity changes of central projection areas revealed by functional magnetic resonance imaging in NaV1.8-deficient mice upon cold signaling

The voltage-gated sodium channel subtype NaV1.8 is expressed in the peripheral nervous system in primary afferent nociceptive C-fibers and is essential for noxious cold signaling. We utilized functional magnetic resonance imaging on NaV1.8-deficient (NaV1.8−/−) compared with wildtype (WT) mice to identify brain structures decoding noxious cold and/or heat signals. In NaV1.8−/− mice functional activity patterns, activated volumes and BOLD signal amplitudes are significantly reduced upon noxious cold stimulation whereas differences of noxious heat processing are less pronounced. Graph-theoretical analysis of the functional connectivity also shows dramatic alterations in noxious cold sensation in NaV1.8−/− mice and clearly reduced interactions between certain brain structures. In contrast, upon heat stimulation qualitatively quite the same functional connectivity pattern and consequently less prominent connectivity differences were observed between NaV1.8−/− and WT mice. Thus, the fact that NaV1.8−/− mice do not perceive nociceptive aspects of strong cooling in contrast to their WT littermates seems not only to be a pure peripheral phenomenon with diminished peripheral transmission, but also consists of upstream effects leading to altered subsequent nociceptive processing in the central nervous system and consequently altered connectivity between pain-relevant brain structures.

Preliminary study on the role of voltage-gated sodium channel subtype Nav1.5 in lymph node metastasis of oral squamous cell carcinoma

Objective: To investigate the relationship between the expression of voltage-gated sodium channel subtype Nav1.5 in oral squamous cell carcinoma (OSCC) and the occurrence and lymph node metastasis of OSCC. Methods: Totally 10 samples of normal oral mucosa tissue as control group, 26 samples of OSCC as the experimental group was divided into non-metastatic group (n=16) and metastatic group (n=10) according to the presence or absence of lymph node metastasis. Quantitative real-time PCR (qPCR), Western blotting, immunohistochemistry, and enzyme-linked immunosorbent assay (ELISA) were used to detect the expression of Nav1.5 in control group and experimental groups at mRNA and protein levels. The data were analyzed by one-way analysis. Results: The expression of Nav1.5 mRNA in the experimental group (non-metastatic group: 2.311±0.134, metastatic group: 4.462±0.362) was higher than those in the control group (1.054±0.162) (P=0.037; P=0.029), and the metastasis group was significantly higher than the non-metastasis group (P=0.031). Western blotting showed the expression of Nav1.5 in experimental groups (non-metastatic: 0.143±0.005, metastatic: 0.253±0.015) was up-regulated significantly compared with control group (0.080±0.010) (P=0.034, P=0.026), and the metastasis group was significantly higher than the non-metastasis group (P=0.033). The immunohistochemistry show the positive expression rates of Nav1.5 in normal and OSCC tissues were 1/10 and 92% (24/26). The differences were statistically significant (P=0.016), and the metastasis group was significantly higher than the non-metastasis group (P=0.028). The ELISA results revealed that the level of Nav1.5 in control was control group (0.834±0.103) μg/L, in non-metastasis group was (1.578±0.167) μg/L, in metastasis group was (3.882±0.422) μg/L (P=0.041; P=0.032), and the metastasis group was significantly higher than the non-metastasis group (P=0.030). Conclusions: Nav1.5 was highly expressed in poorly differentiated OSCC and the expression was significantly different with or without lymph node metastasis. Nav1.5 may be involved in the occurrence and metastasis of OSCC.

CNV1014802 Rescues the Paroxysmal Extreme Pain Disorders Nav1.7 Mutants by Restoring Impaired Inactivation

Voltage-gated sodium channel subtype Nav1.7 plays a critical role in pain reception and represents a promising target for pain relief. CNV1014802, a selective Nav1.7 inhibitor developed by Convergence Pharmaceuticals, has recently completed its phase II clinical trial in the treatment of trigeminal neuralgia. However, the inhibitory features and other potential applications of CNV1014802 remain elusive. Firstly, the effects of CNV on kinetics of Nav1.7 were characterized using whole-cell patch clamp. We find that holding potentials predominantly determines its inhibition potency on Nav1.7 channels. The IC50 values are 1.77 ± 0.28 µM and 71.66 ± 11.93 µM, respectively, when holding potentials are set at −70 mV or −120 mV. Moreover, CNV1014802 significantly hyperpolarizes channel inactivation whereas does not affect the channel activation. In addition, CNV1014802 accelerates the inactivation onset and delays the recovery from inactivation. Finally, CNV1014802 is capable of rescuing Nav1.7 mutations casual of paroxysmal extreme pain disorders (PEPD) by restoring their impaired inactivation to that of wild-type channels. Our study indicates that CNV1014802 may benefit hereditary painful disorder patients with Nav1.7 mutations impairing inactivation.Keywords: Nav1.7, CNV1014802, Inactivation, Paroxysmal extreme pain disorders

L’érythromélalgie : approche diagnostique et thérapeutique actuelle

Erythromelalgia is a rare intermittent vascular acrosyndrome characterized by the combination of recurrent burning pain, warmth and redness of the extremities. It is considered in its primary form as an autosomal dominant neuropathy related to mutations of SCN9A, the encoding gene of a voltage-gated sodium channel subtype Nav1.7. Secondary erythromelalgia is associated with myeloproliferative disorders, drugs (bromocriptine, calcium channel blockers), or clinical conditions such as rheumatic diseases or viral infection. Primary familial erythromelalgia include genetics and sporadic forms associated with small fibers neuropathy. Aspirin is a useful treatment of erythromelagia associated with myeloproliferative disorders. Treatment of primary erythromelalgia is difficult, individualized, with sodium channel blockers such as lidocaine, carbamazepine and mexiletine.

Primary erythromelalgia: a review.

Primary erythromelalgia (PE ORPHA90026) is a rare autosomal dominant neuropathy characterized by the combination of recurrent burning pain, warmth and redness of the extremities. The incidence rate of PE ranges from 0.36 to 1.1 per 100,000 persons. Gender ratio differs according to different studies and no evidence showed a gender preference. Clinical onset of PE is often in the first decade of life. Burning pain is the most predominant symptom and is usually caused and precipitated by warmth and physical activities. Reported cases of PE contain both inherited and sporadic forms. Genetic etiology of PE is mutations on SCN9A, the encoding gene of a voltage-gated sodium channel subtype Nav1.7. Diagnosis of PE is made upon clinical manifestations and screening for mutations on SCN9A. Exclusion of several other treatable diseases/secondary erythromelalgia is also necessary because of the lack of biomarkers specifically for PE. Differential diagnoses can include Fabry disease, cellulites, Raynaud phenomenon, vasculitis and so on. Diagnostic methods often involve complete blood count, imaging studies and thermograph. Treatment for PE is unsatisfactory and highly individualized. Frequently used pain relieving drugs involve sodium channel blockers such as lidocaine, carbamazepine and mexiletine. Novel drugs such as PF-05089771 and TV-45070 could be promising in ameliorating pain symptoms due to their Nav1.7 selectivity. Patients’ symptoms often worsen over time and many patients develop ulcerations and gangrenes caused by excessive exposure to low temperature in order to relieve pain. This review mainly focuses on PE and the causative gene SCN9A -- its mutations and their effects on Nav1.7 channels’ electrophysiological properties. We propose a genotype-channelopathy-phenotype correlation network underlying PE etiology which could provide guidance for future therapeutics.

The role of Nav1.9 channel in the development of neuropathic orofacial pain associated with trigeminal neuralgia.

BackgroundTrigeminal neuralgia is accompanied by severe mechanical, thermal and chemical hypersensitivity of the orofacial area innervated by neurons of trigeminal ganglion (TG). We examined the role of the voltage-gated sodium channel subtype Nav1.9 in the development of trigeminal neuralgia.ResultsWe found that Nav1.9 is required for the development of both thermal and mechanical hypersensitivity induced by constriction of the infraorbital nerve (CION). The CION model does not induce change on Nav1.9 mRNA expression in the ipsilateral TG neurons when evaluated 9 days after surgery.ConclusionsThese results demonstrate that Nav1.9 channels play a critical role in the development of orofacial neuropathic pain. New routes for the treatment of orofacial neuropathic pain focussing on regulation of the voltage-gated Nav1.9 sodium channel activity should be investigated.

Molecular surface of JZTX-V (β-Theraphotoxin-Cj2a) interacting with voltage-gated sodium channel subtype NaV1.4.

Voltage-gated sodium channels (VGSCs; NaV1.1–NaV1.9) have been proven to be critical in controlling the function of excitable cells, and human genetic evidence shows that aberrant function of these channels causes channelopathies, including epilepsy, arrhythmia, paralytic myotonia, and pain. The effects of peptide toxins, especially those isolated from spider venom, have shed light on the structure–function relationship of these channels. However, most of these toxins have not been analyzed in detail. In particular, the bioactive faces of these toxins have not been determined. Jingzhaotoxin (JZTX)-V (also known as β-theraphotoxin-Cj2a) is a 29-amino acid peptide toxin isolated from the venom of the spider Chilobrachys jingzhao. JZTX-V adopts an inhibitory cysteine knot (ICK) motif and has an inhibitory effect on voltage-gated sodium and potassium channels. Previous experiments have shown that JZTX-V has an inhibitory effect on TTX-S and TTX-R sodium currents on rat DRG cells with IC50 values of 27.6 and 30.2 nM, respectively, and is able to shift the activation and inactivation curves to the depolarizing and the hyperpolarizing direction, respectively. Here, we show that JZTX-V has a much stronger inhibitory effect on NaV1.4, the isoform of voltage-gated sodium channels predominantly expressed in skeletal muscle cells, with an IC50 value of 5.12 nM, compared with IC50 values of 61.7–2700 nM for other heterologously expressed NaV1 subtypes. Furthermore, we investigated the bioactive surface of JZTX-V by alanine-scanning the effect of toxin on NaV1.4 and demonstrate that the bioactive face of JZTX-V is composed of three hydrophobic (W5, M6, and W7) and two cationic (R20 and K22) residues. Our results establish that, consistent with previous assumptions, JZTX-V is a Janus-faced toxin which may be a useful tool for the further investigation of the structure and function of sodium channels.

Multiple roles for NaV1.9 in the activation of visceral afferents by noxious inflammatory, mechanical, and human disease–derived stimuli

Chronic visceral pain affects millions of individuals worldwide and remains poorly understood, with current therapeutic options constrained by gastrointestinal adverse effects. Visceral pain is strongly associated with inflammation and distension of the gut. Here we report that the voltage-gated sodium channel subtype NaV1.9 is expressed in half of gut-projecting rodent dorsal root ganglia sensory neurons. We show that NaV1.9 is required for normal mechanosensation, for direct excitation and for sensitization of mouse colonic afferents by mediators from inflammatory bowel disease tissues, and by noxious inflammatory mediators individually. Excitatory responses to ATP or PGE2 were substantially reduced in NaV1.9−/− mice. Deletion of NaV1.9 substantially attenuates excitation and subsequent mechanical hypersensitivity after application of inflammatory soup (IS) (bradykinin, ATP, histamine, PGE2, and 5HT) to visceral nociceptors located in the serosa and mesentery. Responses to mechanical stimulation of mesenteric afferents were also reduced by loss of NaV1.9, and there was a rightward shift in stimulus–response function to ramp colonic distension. By contrast, responses to rapid, high-intensity phasic distension of the colon are initially unaffected; however, run-down of responses to repeat phasic distension were exacerbated in NaV1.9−/− afferents. Finally colonic afferent activation by supernatants derived from inflamed human tissue was greatly reduced in NaV1.9−/− mice. These results demonstrate that NaV1.9 is required for persistence of responses to intense mechanical stimulation, contributes to inflammatory mechanical hypersensitivity, and is essential for activation by noxious inflammatory mediators, including those from diseased human bowel. These observations indicate that NaV1.9 represents a high-value target for development of visceral analgesics.

A Nav1.7 channel mutation associated with hereditary erythromelalgia contributes to neuronal hyperexcitability and displays reduced lidocaine sensitivity

Mutations in the TTX-sensitive voltage-gated sodium channel subtype Nav1.7 have been implicated in the painful inherited neuropathy, hereditary erythromelalgia. Hereditary erythromelalgia can be difficult to treat and, although sodium channels are targeted by local anaesthetics such as lidocaine (lignocaine), some patients do not respond to treatment with local anaesthetics. This study examined electrophysiological differences in Nav1.7 caused by a hereditary erythromelalgia mutation (N395K) that lies within the local anaesthetic binding site of the channel. The N395K mutation produced a hyperpolarized voltage dependence of activation, slower kinetics of deactivation, and impaired steady-state slow inactivation. Computer simulations indicate that the shift in activation is the major determinant of the hyperexcitability induced by erythromelalgia mutations in sensory neurons, but that changes in slow inactivation can modulate the overall impact on excitability. This study also investigated lidocaine inhibition of the Nav1.7-N395K channel. We show that the N395K mutation attenuates the inhibitory effects of lidocaine on both resting and inactivated Nav1.7. The IC50 for lidocaine was estimated at 500 microM for inactivated wild-type Nav1.7 and 2.8 mM for inactivated Nav1.7-N395K. The N395K mutation also significantly reduced use-dependent inhibition of lidocaine on Nav1.7 current. In contrast, a different hereditary erythromelalgia mutation (F216S), not located in the local anaesthetic binding site, had no effect on lidocaine inhibition of Nav1.7 current. Our observation of reduced lidocaine inhibition on Nav1.7-N395K shows that the residue N395 is critical for lidocaine binding to Nav1.7 and suggests that the response of individuals with hereditary erythromelalgia to lidocaine treatment may be determined, at least in part, by their specific genotype.