The voltage-gated sodium channel gene SCN2A and idiopathic generalized epilepsy.

We tested the hypothesis that genetic variation in the human sodium channel gene SCN2A confers liability to idiopathic generalized epilepsy (IGE). We performed a systematic search for mutations in 46 familial IGE cases and detected three novel polymorphisms, however, allele frequencies did not differ significantly between patients and controls. A rare mutation (R1918H) was identified in one patient but was absent in one further affected family member. Thus, our results do not suggest a major role of SCN2A in the etiology of IGE.     

Autosomal dominant erythermalgia associated with a novel mutation in the voltage-gated sodium channel alpha subunit Nav1.7

BACKGROUND: Autosomal dominant primary erythermalgia is a rare disorder characterized by recurrent attacks of red, warm, and painful hands and/or feet. OBJECTIVE: To describe the phenotypes and molecular data of a 10-member family with 5 symptomatic living patients with erythermalgia. RESULTS: The clinical phenotype of this family was featured by episodic or continuous symmetrical red swelling, irritating warmth, and burning pain of feet and lower legs provoked or aggravated by warmth and exercise, and relief was always obtained by application of cold, such as putting feet in (ice-) cold water. The symptoms in this family were only partially controlled by analgesics and sedatives. All affected family members were heterozygous for a novel mutation (S241T) of the voltage-gated sodium channel alpha subunit Nav1.7. CONCLUSION: Primary erythermalgia may be a neuropathic disorder of the small peripheral sensory and sympathetic neurons, and may be caused by hyperexcitability of Nav1.7.     

A sodium channel gene SCN9A polymorphism that increases nociceptor excitability

Sodium channel Na_V1.7, encoded by the SCN9A gene, is preferentially expressed in nociceptive primary sensory neurons, where it amplifies small depolarizations. In studies on a family with inherited erythromelalgia associated with Na_V1.7 gain‐of‐function mutation A863P, we identified a nonsynonymous single‐nucleotide polymorphism within SCN9A in the affected proband and several unaffected family members; this polymorphism (c. 3448C&T, Single Nucleotide Polymorphisms database rs6746030, which produces the amino acid substitution R1150W in human Na_V1.7 [hNa_V1.7]) is present in 1.1 to 12.7% of control chromosomes, depending on ethnicity. In this study, we examined the effect of the R1150W substitution on function of the hNa_V1.7 channel, and on the firing of dorsal root ganglion (DRG) neurons in which this channel is normally expressed. We show that this polymorphism depolarizes activation (7.9–11mV in different assays). Current‐clamp analysis shows that the 1150W allele depolarizes (6mV) resting membrane potential and increases (～2‐fold) the firing frequency in response to depolarization in DRG neurons in which it is present. Our results suggest that polymorphisms in the Na_V1.7 channel may influence susceptibility to pain. Ann Neurol 2009;66:862–866     

Distribution of the voltage-gated sodium channel Na(v)1.7 in the rat: expression in the autonomic and endocrine systems

It is generally accepted that the voltage-gated, tetrodotoxin-sensitive sodium channel, Na(V)1.7, is selectively expressed in peripheral ganglia. However, global deletion in mice of Na(V)1.7 leads to death shortly after birth (Nassar et al. [2004] Proc. Natl. Acad. Sci. U. S. A. 101:12706-12711), suggesting that this ion channel might be more widely expressed. To understand better the potential physiological function of this ion channel, we examined Na(V)1.7 expression in the rat by in situ hybridization and immunohistochemistry. As expected, highest mRNA expression levels are found in peripheral ganglia, and the protein is expressed within these ganglion cells and on the projections of these neurons in the central nervous system. Importantly, we found that Na(V)1.7 is present in discrete rat brain regions, and the unique distribution pattern implies a central involvement in endocrine and autonomic systems as well as analgesia. In addition, Na(V)1.7 expression was detected in the pituitary and adrenal glands. These results indicate that Na(V)1.7 is not only involved in the processing of sensory information but also participates in the regulation of autonomic and endocrine systems; more specifically, it could be implicated in such vital functions as fluid homeostasis and cardiovascular control.     

No mutations in the voltage‐gated NaV1.7 sodium channel α1 subunit gene SCN9A in familial complex regional pain syndrome

Background: Mutations in the voltage‐gated Na_V1.7 Na⁺ channel α1 gene SCN9A have been linked to pain disorders, such as inherited primary erythromelalgia and paroxysmal extreme pain disorder. Both show clinical overlap with complex regional pain syndrome (CRPS), a condition that is characterized by pain in association with combinations of vasomotor, sudomotor, sensory, and motor disturbances. Therefore, we here investigated the involvement of the SCN9A gene in familial CRPS. Methods: We performed a mutation analysis of the SCN9A gene in four index cases of families with CRPS. All 26 coding exons and adjacent sequences of the SCN9A gene were analyzed for mutations using direct sequencing analysis. Results: No causal gene mutations were identified in the SCN9A gene in any of the patients. Conclusions: Despite the fact that the SCN9A gene is an excellent candidate, we did not find evidence that it plays a major role in familial CRPS.     

Molecular biology of the voltage-gated sodium channel

The integral membrane protein composing the voltage-sensitive sodium channel is a bagel-like structure (Fig. 1) with a central pore that is highly selective for sodium ions. When the electric field across a cell's membrane becomes depolarized, the pore of the channel opens to allow sodium ions to flow down their electrochemical gradient into the cell¹. Voltage-gating, a common feature of all voltage-activated channels, is an essential functional property that is only now being understood in terms of the structural features of the protein.     

Dravet综合征患者电压依赖性钠通道α1亚基基因5'-非翻译区外显子的遗传变异

目的 筛查Dravet综合征患者的电压依赖性钠通道α1亚基(voltage-gated sodium channel α1-subunit,SCN1A)基因5'-非翻译区外显子突变位点,分析并预测其致病易患性.方法 收集24例Dravet综合征患者的外周血,抽提基因组DNA,采用直接测序法进行SCN1A基因5'-非翻译区外显子突变位点的筛查;用生物信息学方法分析SCN1A基因5'-非翻译区外显子变异位点邻近序列的保守性及潜在的转录因子结合元件,推测其致病易患性.结果 发现位于外显子h2u上的突变位点166.642.520G>A,先证者1为新生突变,而先证者2的突变来自临床表型正常的母亲,该突变位点在100名健康对照者中均未发现.突变位点在哺乳动物中呈中度保守(62.5%),人与其他哺乳动物之间在突变位点邻近序列的平均同源率高达88.5%;166.642.520野生型位点的序列上预测得到一种转录因子结合元件,而突变型位点的序列上预测得到两种转录因子结合元件.结论 突变位点166.642.520G>A与Dravet综合征存在一定程度的相关性,其致病机制有待于进一步实验证实.     

Role of the sodium channel SCN9A in genetic epilepsy with febrile seizures plus and Dravet syndrome

Mutations of the SCN1A subunit of the sodium channel is a cause of genetic epilepsy with febrile seizures plus (GEFS⁺) in multiplex families and accounts for 70–80% of Dravet syndrome (DS). DS cases without SCN1A mutation inherited have predicted SCN9A susceptibility variants, which may contribute to complex inheritance for these unexplained cases of DS. Compared with controls, DS cases were significantly enriched for rare SCN9A genetic variants. None of the multiplex febrile seizure or GEFS⁺ families could be explained by highly penetrant SCN9A mutations.     

The voltage-gated sodium channel beta2-subunit gene and idiopathic generalized epilepsy

Recent identification of ion channel gene mutations in Mendelian epilepsies suggests that genetically driven neuronal hyperexcitability plays an important role in epileptogenesis. In this study, we tested the hypothesis that genetic variation in the human SCN2B gene confers liability to common subtypes of idiopathic generalized epilepsies (IGE). A systematic search for mutations was performed in 92 IGE patients. We detected a novel single nucleotide polymorphism (SNP), however, allele frequencies did not differ between IGE patients and controls (chi2 = 0.19, df = 1, p = 0.744). Furthermore, a missense mutation in codon 209 (Asn209Pro) was identified in one patient, but was found to be absent in an affected sibling of the index patient. Thus, our results do not suggest a major role of the SCN2B gene in the etiology of common IGE subtypes.     

Fibrillation potentials of denervated rat skeletal muscle are associated with expression of cardiac-type voltage-gated sodium channel isoform Nav1.5

ObjectiveThe molecular mechanisms underlying fibrillation potentials are still unclear. We hypothesised that expression of the cardiac-type voltage-gated sodium channel isoform Nav1.5 in denervated rat skeletal muscle is associated with the generation of such potentials.MethodsMuscle samples were extracted and analysed biologically from surgically denervated rat extensor digitorum longus muscle after concentric needle electromyographic recording at various time points after denervation (4 h to 6 days).ResultsBoth nav1.5 messenger RNA (mRNA) signal on northern blotting and Nav1.5 protein expression on immunohistochemistry appeared on the second day after denervation, exactly when fibrillation potentials appeared. Administration of lidocaine, which has much stronger affinity for sodium channels in cardiac muscle than for those in skeletal muscle, dramatically decreased fibrillation potentials, but had no effect on contralateral compound muscle action potentials.ConclusionsExpression of Nav1.5 participates in the generation of fibrillation potentials in denervated rat skeletal muscle.SignificanceWe proposed an altered expression of voltage-gated sodium channel isoforms as a novel mechanism to explain the occurrence of fibrillation potentials following skeletal muscle denervation.     

Pure haploinsufficiency for Dravet syndrome Na(V)1.1 (SCN1A) sodium channel truncating mutations

Purpose: Dravet syndrome (DS), a devastating epileptic encephalopathy, is mostly caused by mutations of the SCN1A gene, coding for the voltage‐gated Na⁺ channel Na_V1.1 α subunit. About 50% of SCN1A DS mutations truncate Na_V1.1, possibly causing complete loss of its function. However, it has not been investigated yet if Na_V1.1 truncated mutants are dominant negative, if they impair expression or function of wild‐type channels, as it has been shown for truncated mutants of other proteins (e.g., Ca_V channels). We studied the effect of two DS truncated Na_V1.1 mutants, R222* and R1234*, on coexpressed wild‐type Na⁺ channels. Methods: We engineered R222* or R1234* in the human cDNA of Na_V1.1 (hNa_V1.1) and studied their effect on coexpressed wild‐type hNa_V1.1, hNa_V1.2 or hNa_V1.3 cotransfecting tsA‐201 cells, and on hNa_V1.6 transfecting an human embryonic kidney (HEK) cell line stably expressing this channel. We also studied hippocampal neurons dissociated from Na_V1.1 knockout (KO) mice, an animal model of DS expressing a truncated Na_V1.1 channel. Key Findings: We found no modifications of current amplitude coexpressing the truncated mutants with hNa_V1.1, hNa_V1.2, or hNa_V1.3, but a 30% reduction coexpressing them with hNa_V1.6. However, we showed that also coexpression of functional full‐length hNa_V1.1 caused a similar reduction. Therefore, this effect should not be involved in the pathomechanism of DS. Some gating properties of hNa_V1.1, hNa_V1.3, and hNa_V1.6 were modified, but recordings of hippocampal neurons dissociated from Na_V1.1 KO mice did not show any significant modifications of these properties. Therefore, Na_V1.1 truncated mutants are not dominant negative, consistent with haploinsufficiency as the cause of DS. Significance: We have better clarified the pathomechanism of DS, pointed out an important difference between pathogenic truncated Ca_V2.1 mutants and hNa_V1.1 ones, and shown that hNa_V1.6 expression can be reduced in physiologic conditions by coexpression of hNa_V1.1. Moreover, our data may provide useful information for the development of therapeutic approaches.     

Mutations of neuronal voltage-gated Na+ channel alpha 1 subunit gene SCN1A in core severe myoclonic epilepsy in infancy (SMEI) and in borderline SMEI (SMEB)

PURPOSE: Severe myoclonic epilepsy in infancy (SMEI) is a distinct epilepsy syndrome. Patients with borderline SMEI (SMEB) are a subgroup with clinical features similar to those of core SMEI but are not necessarily consistent with the accepted diagnostic criteria for core SMEI. The aim of this study was to delineate the genetic correlation between core SMEI and SMEB and to estimate the frequency of mutations in both phenotypes. METHODS: We examined 96 healthy volunteers and 58 unrelated individuals whose clinical features were consistent with either core SMEI (n = 31) or SMEB (n = 27). We screened for genetic abnormalities within exons and their flanking introns of the genes encoding major subunits of the Na+ channels (SCN1A, SCN2A, SCN1B, and SCN2B) by using a direct sequencing method. RESULTS: In both core SMEI and SMEB, various mutations of SCN1A including nonsense and missense mutations were identified, whereas no mutations of SCN2A, SCN1B, and SCN2B were found within the regions examined. All mutations were heterozygous and not found in 192 control chromosomes. Mutations were identified in 26 (44.8%) of the 58 individuals and were more frequent (p < 0.05) in core SMEI (19 of 31) than in SMEB (seven of 27), as assessed by the continuity-adjusted chi2 test. Mutations resulting in a molecular truncation were found only in core SMEI. Among the mutations, two missense mutations were found in both core SMEI and SMEB. CONCLUSIONS: Our findings confirm that SMEB is part of the SMEI spectrum and may expand the recognition of SMEI and suggest other responsible or modifying genes.     

Localization of Nav1.5 sodium channel protein in the mouse brain

Na(v)1.5 or SCN5A is a member of the voltage-dependent family of sodium channels. The distribution of Na(v)1.5 protein was investigated in the mouse brain using immunohistochemistry. Immunostaining with a Na(v)1.5-specific antibody revealed that Na(v)1.5 protein was localized in certain distinct regions of brain including the cerebral cortex, thalamus, hypothalamus, basal ganglia, cerebellum and brain stem. Notably, we found that Na(v)1.5 protein co-localized with neurofilaments and clustered at a high density in the neuronal processes, mainly axons. These results suggest that Na(v)1.5 protein may play a role in the physiology of the central nervous system (generation and propagation of electrical signals by axons).