Rea, Ruth; (2002) Ion channel dysfunction in inherited neurological disease: Mutations of potassium channels and glycine receptors. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

This thesis examines ion channel dysfunction in two inherited neurological disorders. Episodic ataxia (EA1) is linked to mutations of the KCNA1 gene encoding the Kv1.1 potassium channel subunit, and is characterised by continuous motor unit activity and intermittent cerebellar inco-ordination. Hyperekplexia is an exaggerated startle disorder linked to mutations of the GLRA1 gene, encoding the glycine receptor subunit GlyRα1. We have used two-electrode voltage clamp in vitro, molecular biology, mathematical modelling and imaging of fluorescent fusion proteins to examine the functional differences and interactions between wild-type (wt) and mutant subunits which occur in each disease. We examined the cellular mechanisms of dysfunction for each of five KCNA1 mutants found in EA1. We distinguished between wt and mutant subunits by artificially altering the sensitivity of mutant subunits to externally-applied tetraethylammonium (TEA). We examined the TEA dose-response when mutant subunits were expressed alone, with wt, and as concatemers. We then derived a quantitative description for the electrophysiological data obtained. This indicated that the assembly and trafficking of tetrameric channels were affected by some mutations. We have tested these models by introducing a fluorescent tag to hKv1.1 subunits and visualising the localisation and interactions of mutant and wt subunits in vitro. We have also co-expressed EA1 mutant subunits with wt hKv1.2. In addition, we have investigated two GLRA1 mutations underlying hyperekplexia in a family exhibiting compound heterozygosity, and the reason why inheritance of both mutations elicits the disease phenotype. In vitro, both were non-functional when expressed alone and together. Neither mutant affected wt GlyRα1 function when co-expressed, as occurs in unaffected family members. We examined fluorescent constructs and found that altered expression levels and subcellular localisation may account for the non-functionality of the mutant proteins. Thus, a complete loss of functional GlyRα1-mediated current underlies hyperekplexia in this family; a partial loss is asymptomatic.

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