Goryu Fukuma

A missense mutation of the Na+ channel αII subunit gene Nav1.2 in a patient with febrile and afebrile seizures causes channel dysfunction

Generalized epilepsy with febrile seizures plus (GEFS+), a clinical subset of febrile seizures (FS), is characterized by frequent episodes beyond 6 years of age (FS+) and various types of subsequent epilepsy. Mutations in β1 and αI-subunit genes of voltage-gated Na+ channels have been associated with GEFS+1 and 2, respectively. Here, we report a mutation resulting in an amino acid exchange (R187W) in the gene encoding the α-subunit of neuronal voltage-gated Na+ channel type II (Nav1.2) in a patient with FS associated with afebrile seizures. The mutation R187W occurring on Arg187, a highly conserved residue among voltage-gated Na+ channels, was not found in 224 alleles of unaffected individuals. Whole-cell patch clamp recordings on human embryonic kidney (HEK) cells expressing a rat wild-type (rNav1.2) and the corresponding mutant channels showed that the mutant channel inactivated more slowly than wild-type whereas the Na+ channel conductance was not affected. Prolonged residence in the open state of the R187W mutant channel may augment Na+ influx and thereby underlie the neuronal hyperexcitability that induces seizure activity. Even though a small pedigree could not show clear cosegregation with the disease phenotype, these findings strongly suggest the involvement of Nav1.2 in a human disease and propose the R187W mutation as the genetic defect responsible for febrile seizures associated with afebrile seizures.

Mutations of Neuronal Voltage-gated Na+ Channel α1 Subunit Gene SCN1A in Core Severe Myoclonic Epilepsy in Infancy (SMEI) and in Borderline SMEI (SMEB)

Summary:  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.

Nav1.1 mutations cause febrile seizures associated with afebrile partial seizures

Recent evidence has suggested that the neuronal voltage-gated sodium channel α1-subunit gene (Nav1.1: SCN1A) is responsible for generalized epilepsy with febrile seizures plus (GEFS+2). Here the authors report two novel disease mutations of Nav1.1 in patients with febrile seizures associated with afebrile partial seizures. One is a Val1428Ala substitution in the pore-forming region, and the other is Ala1685Val in the transmembrane helix. These results support the previous findings and contribute to the reliable diagnosis of epilepsy.

Effect of localization of missense mutations in SCN1A on epilepsy phenotype severity

Background and Methods: Many missense mutations in the voltage-gated sodium channel subunit gene SCN1A were identified in patients with generalized epilepsy with febrile seizures plus (GEFS+) and severe myoclonic epilepsy of infancy (SMEI), although GEFS+ is distinct from SMEI in terms of clinical symptoms, severity, prognosis, and responses to antiepileptic drugs. The authors analyzed the localization of missense mutations in SCN1A identified in patients with GEFS+ and SMEI to clarify the phenotype-genotype relationships. Results: Mutations in SMEI occurred more frequently in the “pore” regions of SCN1A than did those in GEFS+. These SMEI mutations in the “pore” regions were more strongly associated than mutations in other regions with the presence of ataxia and tendency to early onset of disease. The possibility of participation of ion selectivity dysfunction of the channel in the pathogenesis of SMEI was suggested by a mutation in the pore region (R946C) identified in a SMEI patient. Conclusions: There was a significant phenotype-genotype relationship in generalized epilepsy with febrile seizures plus and severe myoclonic epilepsy of infancy with SCN1A missense mutations. More severe sodium channel dysfunctions including abnormal ion selectivity that are caused by mutations in the pore regions may be involved in the pathogenesis of SMEI.

A novel mutation of KCNQ3 (c.925T→C) in a Japanese family with benign familial neonatal convulsions

At present, only one mutation of KCNQ3, a KCNQ potassium channel gene, has been identified as a cause of benign familial neonatal convulsions type 2 (BFNC2). We found a T to C substitution (c.925T→C) on one allele of affected individuals in a Japanese family with BFNC but not on 200 alleles from healthy subjects. c.925T→C replaced Trp309, a conserved residue within the P‐loop of the KCNQ potassium channel family that holds the channel pore open, with an Arg (W309R). We report c.925T→C as the second mutation of KCNQ3 responsible for BFNC2. Ann Neurol 2000;47:822–826

Autosomal dominant epilepsy with febrile seizures plus with missense mutations of the (Na^+)-channel α1 subunit gene, SCN1A

Evidence that febrile seizures have a strong genetic predisposition has been well documented. In families of probands with multiple febrile convulsions, an autosomal dominant inheritance with reduced penetrance is suspected. Four candidate loci for febrile seizures have been suggested to date; FEB1 on 8q13-q21, FEB2 on 19p, FEB3 on 2q23-q24, and FEB4 on 5q14-15. A missense mutation was identified in the voltage-gated sodium (Na(+))-channel beta 1 subunit gene, SCN1B at chromosome 19p13.1 in generalized epilepsy with the febrile seizures plus type 1 (GEFS+1) family. Several missense mutations of the (Na(+))-channel alpha 1 subunit (Nav1.1) gene, SCN1A were also identified in GEFS+2 families at chromosome 2q23-q24.3. The aim of this report is precisely to describe the phenotypes of Japanese patients with novel SCN1A mutations and to reevaluate the entity of GEFS+. Four family members over three generations and one isolated (phenotypically sporadic) case with SCN1A mutations were clinically investigated. The common seizure type in these patients was febrile and afebrile generalized tonic-clonic seizures (FS+). In addition to FS+, partial epilepsy phenotypes were suspected in all affected family members and electroencephalographically confirmed in three patients of two families. GEFS+ is genetically and clinically heterogeneous, and associated with generalized epilepsy and partial epilepsy as well. The spectrum of GEFS+ should be expanded to include partial epilepsies and better to be termed autosomal dominant epilepsy with febrile seizures plus (ADEFS+).

Rats Harboring S284L Chrna4 Mutation Show Attenuation of Synaptic and Extrasynaptic GABAergic Transmission and Exhibit the Nocturnal Frontal Lobe Epilepsy Phenotype

Mutations of genes encoding α4, β2, or α2 subunits (CHRNA4, CHRNB2, or CHRNA2, respectively) of nAChR [neuronal nicotinic ACh (acetylcholine) receptor] cause nocturnal frontal lobe epilepsy (NFLE) in human. NFLE-related seizures are seen exclusively during sleep and are characterized by three distinct seizure phenotypes: “paroxysmal arousals,” “paroxysmal dystonia,” and “episodic wandering.” We generated transgenic rat strains that harbor a missense mutation S284L, which had been identified in CHRNA4 in NFLE. The transgenic rats were free of biological abnormalities, such as dysmorphology in the CNS, and behavioral abnormalities. The mRNA level of the transgene (mutant Chrna4) was similar to the wild type, and no distorted expression was detected in the brain. However, the transgenic rats showed epileptic seizure phenotypes during slow-wave sleep (SWS) similar to those in NFLE exhibiting three characteristic seizure phenotypes and thus fulfilled the diagnostic criteria of human NFLE. The therapeutic response of these rats to conventional antiepileptic drugs also resembled that of NFLE patients with the S284L mutation. The rats exhibited two major abnormalities in neurotransmission: (1) attenuation of synaptic and extrasynaptic GABAergic transmission and (2) abnormal glutamate release during SWS. The currently available genetically engineered animal models of epilepsy are limited to mice; thus, our transgenic rats offer another dimension to the epilepsy research field.

Mutation (Ser284Leu) of neuronal nicotinic acetylcholine receptor α4 subunit associated with frontal lobe epilepsy causes faster desensitization of the rat receptor expressed in oocytes

To date five mutations in two major constituents of neuronal nicotinic acetylcholine receptor (nAChR) in the brain, i.e. alpha4 and beta2 subunits, have been identified to be associated with autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE). Among them, only Ser284Leu, a point mutation in alpha4 subunit identified in ADNFLE as well as in a sporadic case with nocturnal frontal lobe epilepsy, remains to be characterized electrophysiologically. We examined the properties of rat nAChR harboring Ser284Leu reconstituted on Xenopus oocytes. Currents elicited in response to application of acetylcholine to oocytes expressing wild type or mutant nAChR were measured by a standard two-microelectrode voltage clamp method. Compared with wild-type nAChR, the mutant nAChR had a comparable EC(50) value for acetylcholine whereas it showed faster desensitization and lower Cs(+)/Na(+) permeability ratio. Ser284Phe, a putative mutation constructed for comparison, exhibited similar properties. These findings indicate that Ser(284) plays an important role in gating of nAChR along with Thr(276) and Ser(280), and suggest that mutation at Ser(284) could reduce nAChR activity similar to other mutations of alpha4 subunit found in ADNFLE.

A de novo KCNQ2 mutation detected in non-familial benign neonatal convulsions

BACKGROUND The underlying genetic abnormalities of rare familial idiopathic epilepsy have been identified, such as mutation in KCNQ2, a K(+) channel gene. Yet, few genetic abnormalities have been reported for commoner epilepsy, i.e., sporadic idiopathic epilepsy, which share a phenotype similar to those of familial epilepsy. OBJECTIVE To search for the genetic cause of seizures in a girl with the diagnosis of non-familial benign neonatal convulsions, and define the consequence of the genetic abnormality identified. METHODS Genetic abnormality was explored within candidate genes for benign familial neonatal and infantile convulsions, such as KCNQ2, 3, 5, KCNE2, SCN1A and SCN2A. The electrophysiological properties of the channels harboring the identified mutation were examined. Western blotting and immunostaining were employed to characterize the expression and intracellular localization of the mutant channel molecules. RESULTS A novel heterozygous mutation (c.910-2delTTC or TTT, Phe304del) of KCNQ2 was identified in the patient. The mutation was de novo verified by parentage analysis. The mutation was associated with impaired functions of KCNQ K(+) channel. The mutant channels were expressed on the cell surface. CONCLUSION The mutant Phe304del of KCNQ2 leads to null function of the KCNQ K(+) channel but the mutation does not alter proper channel sorting onto the cell membrane. Our findings indicate that the genes responsible for rare inherited forms of idiopathic epilepsy could be also involved in sporadic forms of idiopathic epilepsy and expand our notion of the involvement of molecular mechanisms in the more common forms of idiopathic epilepsy.

Phenotypes and genotypes in epilepsy with febrile seizures plus

In the last several years, mutations of sodium channel genes, SCN1A, SCN2A, and SCN1B, and GABA(A) receptor gene, GABRG2 were identified as causes of some febrile seizures related epilepsies. In 19 unrelated Japanese families whose probands had febrile seizures plus or epilepsy following febrile seizures plus, we identified 2 missense mutations of SCN1A to be responsible for the seizure phenotypes in two FS+ families and another mutation of SCN2A in one family. The combined frequency of SCN1A, SCN2A, SCN1B, SCN2B, and GABRG2 mutations in Japanese patients with FS+ was 15.8%. One family, which had R188W mutation in SCN2A, showed digenic inheritance, and another modifier gene was thought to take part in the seizure phenotype. The phenotypes of probands were FS+ in 5, FS+ and partial epilepsy in 10, FS+ and generalized epilepsy in 3, and FS+ and unclassified epilepsy in 1. We proposed the term epilepsy with febrile seizures plus (EFS+), because autosomal-dominant inheritance in EFS+ might be rare, and most of EFS+ display a complex pattern of inheritance, even when it appears to be an autosomal-dominant inheritance. There is a possibility of simultaneous involvement of multiple genes for seizure phenotypes.