The long-term goal of this project is to understand the molecular bases for the diverse structures and functions of brain voltage-sensitive calcium channels (VSCCs) using biochemical and molecular genetics techniques. Multiple types of VSCCs, (i.e., T, N, L, and P-type) are present in both excitable and nonexcitable cells and are important for excitation-contraction coupling and secretion of neurotransmitters and hormones. cDNAs corresponding to the alphal, alpha2,and beta subunits of the brain L-type dihydropyridine (DHP)-sensitive Ca2 + channels have been cloned and the cDNA clones encoding the alpha1 and beta subunits of N- and P-types of neuronal Ca2+ channels have been isolated. This molecular cloning has indicated that a heterogeneous family of voltage-sensitive Ca2 + channels are expressed in the mammalian brain, providing structural bases for the functional diversity of neuronal Ca2 + channels. As a first step toward understanding the genetic bases for the diversity of brain Ca2 + channels, genes encoding the alpha1, alpha2, and beta subunits of the VSCCs have been mapped in human and mouse genomes. With these cDNA clones as probes, the following areas are actively being pursued to elucidate molecular and genetic bases for the diverse VSCC forms and functions: 1)functional expression of VSCC by introducing full-length cDNAs for alpha1, alpha2, and beta subunits into neuronal cells; 2) in situ hybridization histochemistry technique is being utilized to determine the spatiotemporal expression of VSCC subunit mRNAs in brain; and 3) analyses of various cDNA and genomic clones to investigate a possible role(s) of alternative splicing in generating diverse forms of VSCC present in brain. In addition, the human genomic clones will be used to identify and characterize regulatory elements in the promoter region of Ca2 + channel subunit genes.
