A Spauschus

Human epilepsy associated with dysfunction of the brain P/Q-type calcium channel

BACKGROUND The genetic basis of most common forms of human paroxysmal disorders of the central nervous system, such as epilepsy, remains unidentified. Several animal models of absence epilepsy, commonly accompanied by ataxia, are caused by mutations in the brain P/Q-type voltage-gated calcium (Ca(2+)) channel. We aimed to determine whether the P/Q-type Ca(2+) channel is associated with both epilepsy and episodic ataxia type 2 in human beings. METHODS We identified an 11-year-old boy with a complex phenotype comprising primary generalised epilepsy, episodic and progressive ataxia, and mild learning difficulties. We sequenced the entire coding region of the gene encoding the voltage-gated P/Q-type Ca(2+) channel (CACNA1A) on chromosome 19. We then introduced the newly identified heterozygous mutation into the full-length rabbit cDNA and did detailed electrophysiological expression studies of mutant and wild type Ca(2+) channels. FINDINGS We identified a previously undescribed heterozygous point mutation (C5733T) in CACNA1A. This mutation introduces a premature stop codon (R1820stop) resulting in complete loss of the C terminal region of the pore-forming subunit of this Ca(2+) channel. Expression studies provided direct evidence that this mutation impairs Ca(2+) channel function. Mutant/wild-type co-expression studies indicated a dominant negative effect. INTERPRETATION Human absence epilepsy can be associated with dysfunction of the brain P/Q-type voltage-gated Ca(2+) channel. The phenotype in this patient has striking parallels with the mouse absence epilepsy models.

Clinical, genetic, and expression studies of mutations in the potassium channel gene KCNA1 reveal new phenotypic variability.

Episodic ataxia type 1 (EA1) is an autosomal dominant central nervous system potassium channelopathy characterized by brief attacks of cerebellar ataxia and continuous interictal myokymia. Point mutations in the voltage‐gated potassium channel gene KCNA1 on chromosome 12p associate with EA1. We have studied 4 families and identified three new and one previously reported heterozygous point mutations in this gene. Affected members in Family A (KCNA1 G724C) exhibit partial epilepsy and myokymia but no ataxic episodes, supporting the suggestion that there is an association between mutations of KCNA1 and epilepsy. Affected members in Family B (KCNA1 C731A) exhibit myokymia alone, suggesting a new phenotype of isolated myokymia. Family C harbors the first truncation to be reported in KCNA1 (C1249T) and exhibits remarkably drug‐resistant EA1. Affected members in Family D (KCNA1 G1210A) exhibit attacks typical of EA1. This mutation has recently been reported in an apparently unrelated family, although no functional studies were attempted. Heterologous expression of the proteins encoded by the mutant KCNA1 genes suggest that the four point mutations impair delayed‐rectifier type potassium currents by different mechanisms. Increased neuronal excitability is likely to be the common pathophysiological basis for the disease in these families. The degree and nature of the potassium channel dysfunction may be relevant to the new phenotypic observations reported in this study. Ann Neurol 2000;48:647–656

Variable K+ channel subunit dysfunction in inherited mutations of KCNA1

Mutations of KCNA1, which codes for the K+ channel subunit hKv1.1, are associated with the human autosomal dominant disease episodic ataxia type 1 (EA1). Five recently described mutations are associated with a broad range of phenotypes: neuromyotonia alone or with seizures, EA1 with seizures, or very drug‐resistant EA1. Here we investigated the consequences of each mutation for channel assembly, trafficking, gating and permeation. We related data obtained from co‐expression of mutant and wild‐type hKv1.1 to the results of expressing mutant‐wild‐type fusion proteins, and combined electrophysiological recordings in Xenopus oocytes with a pharmacological discrimination of the contribution of mutant and wild‐type subunits to channels expressed at the membrane. We also applied confocal laser scanning microscopy to measure the level of expression of either wild‐type or mutant subunits tagged with green fluorescent protein (GFP). R417stop truncates most of the C‐terminus and is associated with severe drug‐resistant EA1. Electrophysiological and pharmacological measurements indicated that the mutation impairs both tetramerisation of R417stop with wild‐type subunits, and membrane targeting of heterotetramers. This conclusion was supported by confocal laser scanning imaging of enhanced GFP (EGFP)‐tagged hKv1.1 subunits. Co‐expression of R417stop with wild‐type hKv1.2 subunits yielded similar results to co‐expression with wild‐type hKv1.1. Mutations associated with typical EA1 (V404I) or with neuromyotonia alone (P244H) significantly affected neither tetramerisation nor trafficking, and only altered channel kinetics. Two other mutations associated with a severe phenotype (T226R, A242P) yielded an intermediate result. The phenotypic variability of KCNA1 mutations is reflected in a wide range of disorders of channel assembly, trafficking and kinetics.

The molecular biology of the autosomal-dominant cerebellar ataxias

Autosomal‐dominant cerebellar ataxias (ADCA) may present as progressive or paroxysmal disorders. While the progressive ataxias have been named spinocerebellar ataxias (SCA), the paroxysmal disorders are designated episodic ataxias (EA). Until now, three different mutational mechanisms resulting in distinctive pathogenesis have been identified. The first type of mutation present in SCA1, SCA2, SCA3, and SCA7 is an expanded CAG repeat in genes of unknown function that are translated into proteins with expanded polyglutamine tracts. A common ultrastructural feature of these disorders is the formation of neuronal intranuclear inclusions (NII) harboring the expanded disease proteins and a variety of other proteins. The pathogenic role of these inclusions has yet to be clarified. A second group of disorders is the result of mutations in genes that code for ion channels. In EA‐1, a disorder characterized by episodes of ataxia provoked by movement and startle, missense mutations in a potassium channel gene, KCNA1, have been found. Patients with EA‐2, another form of paroxysmal ataxia, carry nonsense mutations of the gene encoding the α1A voltage‐dependent calcium channel subunit, CACNA1A, that are predicted to result in truncated channel proteins. In SCA6, a progressive ataxia, an expanded CAG repeat in the 3` translated region of the CACNA1A gene, has been found. The third type of mutation is an untranslated CTG expansion resembling the mutation found in myotonic dystrophy. It is associated with a progressive ataxia, SCA8.

Functional Characterization of a Novel Mutation in KCNA1 in Episodic Ataxia Type 1 Associated with Epilepsy

pisodic ataxia type 1 (EA1) is a rare autosomal dominant neurological disorder in which patients develop sudden episodes of ataxia precipitated by physical or emotional stress. These can last seconds to minutes, and between attacks patients often have spontaneous, repetitive muscle activity (myokymia), which is not always clinically apparent. Missense point mutations in KCNA1, the gene encoding the human orthologue of Kv1.1 on chromosome 12p13, have been linked to EA1, and the physiological properties of some mutations have been studied. In a large kindred with EA1 where two out of five affected family members have epilepsy we recently detected a previously undescribed point mutation in the second membrane-spanning domain of KCNA1. This mutation results in a change of threonine at amino acid position 226 to arginine (T226R). Further linkage study using mismatch primer PCR revealed that patients were heterozygous for T226R, while none of the unaffected family members nor 100 control individuals carried the mutation. T226 is highly conserved in the Kv1 subfamily through different species, and the replacement by an arginine side chain is a radical exchange. We therefore expressed wild-type and mutant hKv1.1 subunits in Xenopus laevis oocytes to investigate the functional implications of T226R.

The Inherited Episodic Ataxias: How Well Do We Understand the Disease Mechanisms?

The past few years have seen the elucidation of several neurological diseases caused by inherited mutations of ion channels. In contrast to many other types of genetic disorders, the “channelopathies” can be studied with high precision by applying electrophysiological methods. This review evaluates the success of this approach in explaining the mechanisms of two forms of episodic ataxia that are known to be caused by mutations of ion channels: episodic ataxia type 1 (EA1, caused by K+ channel mutations) and episodic ataxia type 2 (EA2, caused by Ca2+ channel mutations). Although both of these disorders are rare, they raise many important questions about the roles of identified channels in brain function. Indeed, a resolution of the mechanisms by which both diseases occur will represent a major milestone in understanding diseases of the CNS, in addition to opening the way to novel possible treatments.