MOLECULAR STUDIES OF SODIUM CHANNELS IN SKELETAL MUSCLE

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The voltage-gated sodium channel is responsible for the generation and propagation of action potential in electrically excitable cells. Sodium channels exhibit different pharmacological and functional properties and are encoded by a multigene family. Three highly homologous sodium channel alpha subunits have been identified in rat brain by cDNA nucleotide sequence analysis. The sponsor's lab has cloned and characterized a full- length cDNA encoding a unique rat sodium channel cDNA in skeletal muscle. A goal of the proposed research is to elucidate the primary structure of a second novel rat skeletal muscle sodium channel gene and to generate probes to determine the structural basis for differences in biological properties of the sodium channels in nerve and skeletal muscle. The specific goals are: 1) to determine how many distinct sodium channel subunits are present in rat skeletal muscle by nucleotide sequence analysis of isolate full-length cDNAs and authentication of their functional properties using the Xenopus oocyte expression system; 2) to determine the cellular and subcellular localization of the rat skeletal muscle sodium channel alpha subunits by immuno-cytochemical and ultra-structural techniques, using antibodies raised against unique portions of the alpha subunit peptides; 3) to determine the molecular basis of tetrodotoxin (TTX)-sensitivity and -insensitivity utilizing sodium channel mRNA analysis in TTX-sensitive adult muscle and relatively TTX-resistant embryonic and denervated muscle. In mammals, spontaneous mutations of sodium channel genes that overtly disturb normal nervous system and skeletal muscle function are anticipated to occur, based on the key role of the sodium channel in signalling and information-processing, and the large number of genes involved in sodium channel protein production in may tissues. These planned studies, in conjunction with my experience in genetics, will provide the framework for addressing the role of putative muscle sodium channel gene mutations in disease of skeletal muscle in man.