Autosomal dominant nocturnal frontal lobe epilepsy with a mutation in the CHRNB2 gene

Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE; MIM 600513) has been associated with mutations in the genes coding for the alfa-4 (CHRNA4), beta-2 (CHRNB2), and alpha-2 (CHRNA2) subunits of the neuronal nicotinic acetylcholine receptor (nAChR) and for the corticotropin-releasing hormone (CRH). A four-generation ADNFLE family with six affected members was identified. All affected members presented the clinical characteristics of ADNFLE. Interictal awake and sleep EEG recordings showed no epileptiform abnormalities. Ictal video-EEG recordings showed focal seizures with frontal lobe semiology. Mutation analysis of the CHRNB2 gene revealed a c.859G>A transition (Val287Met) within the second transmembrane domain, identical to that previously described in a Scottish ADNFLE family. To our knowledge, this is the third family reported presenting a mutation in CHRNB2. The clinical phenotype appears similar to that described with mutations in CHRNA4, suggesting that mutations in these two subunits lead to similar functional alterations of the nAChR.     

Further evidence of genetic heterogeneity in families with autosomal dominant nocturnal frontal lobe epilepsy

PURPOSE: Mutations in the genes encoding the alfa(2), alfa(4) and beta(2) subunits of the neuronal nicotinic acetylcholine receptor (nAChR) play a causative role in autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE). Moreover, variations in the promoter of the corticotropic-releasing hormone gene (CRH) were also associated with ADNFLE. Here, we investigated whether nine brain-expressed genes (CHRNA2, CHRNA3, CHRNA4, CHRNA5, CHRNA6, CHRNA7, CHRNB2, CHRNB3, CHRNB4), encoding distinct nAChR subunits, and CRH are associated with the disease in three distinct ADNFLE families from Southern Italy. METHODS: There were 14 living affected individuals (9 women), ranging in age from 14 to 57 years, pertaining to three unrelated families. Age at onset of seizures clustered around 9 years of age (range from 7 and 16 years, mean: 9.1 years+/-3.8). All affected individuals manifested nocturnal partial seizures of frontal lobe origin, which were well controlled by medications. Exon 5 of CHRNA4 and CHRNB2 genes, harboring all the known mutations, was sequenced in the probands. Then, we performed a linkage study on 13 affected and 26 non-affected individuals belonging to the three families with microsatellite markers and an intragenic polymorphisms encompassing the chromosome localization of the nAChR subunit genes and of the CRH gene. RESULTS: Mutational and linkage analyses allowed us to exclude the involvement of all known nAChR subunit genes and of the CRH gene in ADNFLE in our families. CONCLUSION: Our results further illustrate the considerable genetic heterogeneity for such a syndrome, despite the quite homogeneous clinical picture. It is therefore reasonable to hypothesize that at least another gene not belonging to the nAChR gene family, in addition to CRH, is involved in the pathogenesis of ADNFLE.     

Autosomal dominant nocturnal frontal lobe epilepsy

Abstract. Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is an idiopathic epilepsy, with a spectrum of clinical manifestations, ranging from brief, stereotyped, sudden arousals to more complex dystonic–dyskinetic seizures. Video–polysomnography allows a correct differential diagnosis. There is no difference between sporadic nocturnal frontal lobe epilepsy (NFLE) and ADNFLE in the clinical and neurophysiological findings. ADNFLE is the first idiopathic epilepsy for which a genetic basis has been identified. Mutations have been found in two genes (CHRNA4 and CHRNB2) coding for neuronal nicotinic receptor subunits (4 and 2, respectively). Contrasting data have been reported on the effect of these mutations on the functionality of the receptor.Moreover, the incomplete data on the neuronal network/s in which this receptor is involved, make difficult the understanding of the genotype–phenotype correlation. This is an overview on the clinical and genetic aspects of ADNFLE including a discussion of some open questions on the role of the neuronal nicotinic receptor subunit mutations in the pathogenesis of this form of epilepsy.     

Mutational analysis of nicotinic acetylcholine receptor beta2 subunit gene (CHRNB2) in a representative cohort of Italian probands affected by autosomal dominant nocturnal frontal lobe epilepsy

Twenty-four autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) probands were analyzed for the presence of V287L and V287M mutations in the CHRNB2 gene, which have been recently associated with the disease. In all patients, the involvement of the two additional loci reported as being associated with ADNFLE (CHRNA4 gene and chromosome 15q24 region) had been previously excluded. Mutational screening was performed by sequencing a polymerase chain reaction-amplified CHRNB2 DNA fragment, spanning the whole exon 5, which contains the V287L and V287M mutations and codes for approximately 65% of the mature protein. In none of the patients were mutations in the analyzed region of CHRNB2 found. These data, obtained in the largest ADNFLE cohort so far analyzed, demonstrate the rarity of the identified CHRNB2 mutations in ADNFLE patients.     

The identification of a novel mutation of nicotinic acetylcholine receptor gene CHRNB2 in a Chinese patient: Its possible implication in non-familial nocturnal frontal lobe epilepsy

Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is partly caused by mutations in the nicotinic acetylcholine receptor (nAChR) genes CHRNA4, CHRNB2, and CHRNA2. Cases of non-familial nocturnal frontal lobe epilepsy (NFLE) are more common than the familial type and the phenotypes of the two are similar. CHRNA4 mutations have been found in sporadic NFLE, but no mutation in CHRNB2 or CHRNA2 have been reported. To analyze the genetic features of sporadic NFLE, we designed mutation screening of exon 5 of CHRNA4, exon 5 of CHRNB2, and exon 6 of CHRNA2, mutations in which are associated with ADFLE. We screened a group of 105 Chinese sporadic NFLE cases and identified a novel CHRNB2 mutation, V337G, in an evolutionary conserved region of the intracellular loop between transmembrane domains M3 and M4 in one patient. This mutation was not observed in the control group of 200 subjects. Bioinformatics analysis indicated that the mutation altered the hydrophobicity and secondary structure of the protein. To the best of our knowledge, this study established for the first time that CHRNB2 is potentially associated with non-familial NFLE patient. No mutations in CHRNA4 or CHRNA2 were revealed by our screening method.     

Refined mapping of CHRNA3/A5/B4 gene cluster and its implications in ADNFLE

The chromosome 15q24 region, containing the CHRNA3/A5/B4 gene cluster, coding for the alpha3, alpha5 and beta4 subunits of neuronal nicotinic acetylcholine receptors, has been reported to be linked to autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) in one family. However, nor the gene nor the mutation involved have been identified. We report the refined mapping of CHRNA3/A5/B4 cluster. Segregation analyses of CHRNA3/A5/B4 polymorphisms in families showing recombinations for 15q24 G?en?ethon STR markers allowed to position the cluster in a 0.6 cM interval, between STRs D15S1027 and D15S1005. This location is external to the 15q24-ADNFLE-linked region, therefore excluding the involvement of this cluster in the pathogenesis of ADNFLE in the 15q24-linked family. Moreover, these data provide more precise information for further linkage studies.     

Evidence for a fourth locus for autosomal dominant nocturnal frontal lobe epilepsy

Mutations responsible for autosomal dominant nocturnal frontal lobe epilepsy have been identified in two members of the neuronal nicotinic acetylcholine receptor gene family: CHRNA4(ENFL1 locus) and CHRNB2 (ENFL3 locus) coding for alpha4 and beta2 subunit, respectively. However, mutations in these genes account for only a minority (less than 10%) of cases. For a third ADNFLE locus (ENFL2) on chromosome 15q24 the gene was not identified. The involvement of the three loci in the pathogenesis of ADNFLE was investigated in 12 unrelated Italian families, selected on the basis of anamnestic and video-polysomnographic data. Compliant family members were typed for polymorphic markers spanning the analyzed chromosome regions. Linkage analyses excluded association of all chromosome regions with ADNFLE in 72% of cases. In two, four and one families it was impossible to ascertain or exclude association with ENFL1, ENFL2, or ENFL3, respectively, however, no mutations have been detected in the nicotinic receptor genes located in these regions. These data strongly suggest that ENFL1, ENFL2 and ENFL3 are minor loci for the disease and point to the existence of at least a fourth locus for ADNFLE.     

Mutations in familial nocturnal frontal lobe epilepsy might be associated with distinct neurological phenotypes

Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is a rare familial seizure disorder caused by mutations in at least two different subunit genes of the neuronal nicotinic acetylcholine receptor (nAChR), CHRNA4 and CHRNB2. ADNFLE was initially described as a "pure" seizure disorder with a mostly benign course. We have analysed the clinical features of 19 ADNFLE families from 12 countries with a total of 150 patients and grouped them with respect to their nAChR mutations. These data suggest that certain nAChR mutations might be associated with an increased risk for major neurological symptoms such as mental retardation, schizophrenia-like symptoms or marked cognitive deficits, but the risk for these disorders seems to be low for most other ADNFLE mutations. The functional data confirm that the mutations differ from each other with respect to the size of their gain-of function effects and other biopharmacological characteristics although these functional changes are not predictive for the severity of the clinical phenotype.     

A new locus for autosomal dominant nocturnal frontal lobe epilepsy maps to chromosome 1

BACKGROUND: Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is caused by mutations in the alpha4 subunit of the neuronal nicotinic acetylcholine receptor (CHRNA4) gene, mapping on chromosome 20q13.2. A second ADNFLE locus was mapped on chromosome 15q24. OBJECTIVE: To report a new third ADNFLE locus on chromosome 1 in a large Italian family. METHODS: The authors performed a clinical and genetic study in a large, three-generation ADNFLE family from southern Italy, including eight affected individuals and three obligate carriers. RESULTS: The age at onset of seizures was around 9 years of age and all affected individuals manifested nocturnal partial seizures of frontal lobe origin. Interictal awake and sleep EEG recordings showed no definite epileptiform abnormalities in most patients. Ictal video-EEG showed that the attacks were partial seizures with a frontal lobe semiology. Intellectual and neurologic examinations, and brain CT or MRI results were always normal. Carbamazepine was effective in all treated patients. Exclusion mapping of the known loci linked to ADNFLE-ENFL1, and ENFL2, on chromosomes 20q13.2 and 15q24-was performed on the pedigree before starting the genome-wide linkage analysis. The whole genome scan mapping allowed the identification of a new ADNFLE locus spanning the pericentromeric region of chromosome 1. CONCLUSIONS: The authors provided evidence for a third locus associated to autosomal dominant nocturnal frontal lobe epilepsy on chromosome 1. Among the known genes mapping within this critical region, the ss2 subunit of the nicotinic receptor (CHRNB2) represents the most obvious candidate.     

Thursday July 6, 200611:30–13:30Hall 5CDiscussion Group SessionNon–age-related familial focal epilepsies: genotype-phenotype correlations

¹ A. Gambardella (   ¹ Università Magna Graecia, Italy )
Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is a recently identified partial epilepsy, which is characterized by brief frontal-lobe motor seizures occurring mostly during light sleep. The age of onset is usually infancy and adolescence, although seizures may start in adult life. Inheritance is autosomal dominant with 70% penetrance. The clinical picture of ADNFLE is relatively homogeneous, even if a broad range of severity has been observed even among affected members of the same pedigree. The interictal EEG is usually normal but ictal recordings show that these events are epileptic and appear to arise from the frontal lobes. Misdiagnosis as nightmares, night terrors, other parasomnias or even hysteria is common if clinicians are unaware of ADNFLE.
So far, ADNFLE has been associated with mutations affecting two genes coding for alfa4 and beta2 subunits of the neuronal nicotinic acetylcholine receptor (nAChR), which are located on chromosome 20q and chromosome 1 respectively. Moreover, although the gene has not yet been identified, another ADNFLE locus has been mapped to chromosome 15q24. More recently, there has been evidence that variations in the promoter of the corticotropic-releasing hormone gene may be also associated with ADNFLE. Nonetheless, the underlying gene has not yet been found in most ADNFLE families. Overall, these data support the pathogenic role of the cholinergic system in ADNFLE, even if its etiology appears to be the result of a variety of molecular defects despite the relative homogeneity of the clinical manifestations.     

The 1674+11C>T polymorphism of CHRNA4 is associated with juvenile myoclonic epilepsy

The α4 subunit gene (CHRNA4) of the neuronal nicotinic acetylcholine receptor (nAChR), linked to an idiopathic partial epilepsy, autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE), may also play a key role in the development of the idiopathic generalized epilepsy syndrome (IGE), juvenile myoclonic epilepsy (JME). This study was designed to explore an association of four polymorphisms of the CHRNA4 with JME in Polish children and young patients. The study included 92 JME patients and 222 unrelated healthy individuals. In each group the frequencies of the CHRNA4 c.555C>T, c.594C>T, 1674⁺¹¹C>T, and 1674⁺¹⁴A>G polymorphisms were determined using PCR-RFLP analyses. An association between the 1674⁺¹¹C>T polymorphism of the CHRNA4 and JME was evidenced. Allele T (the risk factor) appeared with a significantly higher frequency in the JME patients than in the controls (p = 0.0299). The patients harboring the 1674⁺¹¹CT+TT genotypes showed an increased risk of JME (CT+TT versus CC: OR = 1.925; 95% CI = 1.021–3.629; p = 0.0408). No association was found for the other CHRNA4 polymorphisms tested. The CHRNA4 1674⁺¹¹C>T polymorphism may be a susceptibility factor for epilepsy, and its higher frequency in patients with juvenile myoclonic epilepsy suggests that the CHRNA4 may be one of the candidate genes for this epileptic syndrome.     

CHRNA2 mutations are rare in the NFLE population: evaluation of a large cohort of Italian patients

ObjectiveForty-six nocturnal frontal lobe epilepsy (NFLE) patients (in which the involvement of the CHRNA4 and CHRNB2 genes coding for neuronal nicotinic acetylcholine receptor (nAChRs) subunits associated to the disease were previously excluded) were analyzed for the presence of mutations in the CHRNA2 gene coding for the alpha2 subunit of the same receptor, which has been recently associated with the disease.MethodsMutational screening was performed by sequencing two polymerase chain reaction-amplified CHRNA2 DNA fragments, spanning the whole exon 6 and exon 7, respectively, which code for ～75% of the mature protein and contain all four transmembrane domains contributing to the ion pore.ResultsNo mutations were identified in the analyzed region of CHRNA2.ConclusionsThese data demonstrate the rarity of the identified CHRNA2 mutations in NFLE patients, supporting the recently reported hypothesis of a restricted role for this gene in the disease.     

Genetic association analysis of tagging SNPs in alpha4 and beta2 subunits of neuronal nicotinic acetylcholine receptor genes (<Emphasis Type="Italic">CHRNA4</Emphasis> and <Emphasis Type="Italic">CHRNB2</Emphasis>) with schizophrenia in the Japanese population

Several lines of evidence suggest that nicotinic cholinergic dysfunction may contribute to the cognitive impairments in schizophrenia. The majority of high affinity nicotine binding sites in the human brain have been implicated in heteropentameric alpha4 and beta2 subunits of neuronal nicotinic acetylcholine receptors; therefore, these two neuronal nicotinic acetylcholine receptors genes (CHRNA4 and CHRNB2) are considered to be attractive candidate genes for the pathophysiology of schizophrenia. To represent these two genes in a gene-wide manner, we first evaluated the linkage disequilibrium structure using our own control samples. Thirteen SNPs (7 SNPs for CHRNA4 and 5 SNPs for CHRNB2) were selected as tagging SNPs. Using these tagging SNPs, we then conducted genetic association analysis of case-control samples (738 schizophrenia and 753 controls) in the Japanese population. No significant association was detected in the allele/genotype-wise or haplotype-wise analysis. Our results suggest that CHRNA4 and CHRNB2 do not play a major role in Japanese schizophrenia.     

The CHRNB2 mutation I312M is associated with epilepsy and distinct memory deficits

Mutations in nAChRs are found in a rare form of nocturnal frontal lobe epilepsy (ADNFLE). Previously, some nAChR mutations have been described that are associated with additional neurological features such as psychiatric disorders or cognitive defects. Here, we report a new CHRNB2 mutation located in transmembrane region 3 (M3), outside the known ADNFLE mutation cluster. The CHRNB2 mutation I312M, which occurred de novo in twins, markedly increases the receptor's sensitivity to acetylcholine. Phenotypically, the mutation is associated not only with typical ADNFLE, but also with distinct deficits in memory. The cognitive problems are most obvious in tasks requiring the organization and storage of verbal information.     

Severe autosomal dominant nocturnal frontal lobe epilepsy associated with psychiatric disorders and intellectual disability

Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is a relatively benign epilepsy syndrome with few comorbidities. Here we describe two families with unusually severe ADNFLE, with associated psychiatric, behavioral, and cognitive features. Detailed clinical data on 17 affected individuals were obtained, and genotyping of microsatellite markers, linkage analysis, and sequencing of candidate genes was performed. The severe ADNFLE phenotype in these families was often refractory to treatment, with status epilepticus occurring in 24% of subjects. Psychiatric or behavioral disorders occurred in 53%, with intellectual disability in 24%, and developmental regression in two individuals. No mutations were identified in alpha4, alpha2, or beta2 nAChR subunits. In one family there was evidence of linkage to a region of 15q24 without nAChR subunit genes. In conclusion, severe ADNFLE has significant medical, psychiatric, and intellectual morbidity. The molecular basis of severe ADNFLE is unknown but may involve non-nAChR-related mechanisms.     

Impact of our understanding of the genetic aetiology of epilepsy

A genetic contribution to aetiology is estimated to be present in up to 40% of patients with epilepsy. It is useful to categorise genetic epilepsies according to the mechanisms of inheritance into Mendelian disorders, non-mendelian or ‘complex’ disorders, and chromosomal disorders. Over 200 Mendelian diseases include epilepsy as part of the phenotype, and the genes for a number of these have been identified recently. These include autosomal recessive progressive myoclonic epilepsies such as Unverricht-Lundborg disease, Lafora disease and the neuronal ceroid lipofuscinoses, and three autosomal dominant idiopathic epilepsies. The last named have been shown to arise from mutations in ion channel genes. Autosomal dominant nocturnal frontal lobe epilepsy is caused by mutations in CHRNA4, benign familial neonatal convulsions by mutations in KCNQ2 and KCNQ3, and generalised epilepsy with febrile seizures plus by mutations in SCN1B. ‘Complex’, familial epilepsies are more difficult to analyse, but evidence has been obtained for loci predisposing to juvenile myoclonic epilepsy on chromosome 6p and 15q. Lastly, the genes underlying several spike-wave epilepsies in mice have been cloned, and three of these encode sub-units of voltage-gated calcium channels. Received: 29 September 1999/Accepted: 7 December 1999     

Two new putative susceptibility loci for ADNFLE

Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) has been up to now considered a simple Mendelian trait caused by mutations in neuronal nicotinic acetylcholine receptor (nAChR) subunit genes. We previously demonstrated that in a three-generation Italian family the disease was unlinked to all known ADNFLE loci as well as to all known brain-expressed nAChR subunits. The genome-wide linkage analysis here presented performed on this family points to the existence of two new putative ADNFLE loci on chromosomes 3p22-p24 and 8q11.2-q21.1. These findings, together with several ADNFLE characteristics, suggest that this epilepsy could be, at least in the above family, a complex disorder. In particular, we propose and discuss the hypothesis of a digenic transmission of the disease.     

6-hydroxydopamine lesion of rat nigrostriatal dopaminergic neurons differentially affects nicotinic acetylcholine receptor subunit mRNA expression

Nicotinic acetylcholine receptor (nAChR) subunit mRNA expression in the rat substantia nigra (SN) was assayed by semiquantitative RT-PCR following 6-hydroxydopamine (6-OHDA) lesion of nigrostriatal dopaminergic neurons. Six months after unilateral injection of 6-OHDA or saline into the SN, total RNA was isolated from ipsilateral and contralateral tissue samples. RT-PCR amplifications were performed with template titration using primers specific for sequences encoding