IDENTIFICATION AND DIFFERENTIAL SUBCELLULAR-LOCALIZATION OF THE NEURONAL CLASS-C AND CLASS-D L-TYPE CALCIUM CHANNEL-ALPHA-1 SUBUNITS

To identify and localize the protein products of genes encoding distin ct L-type calcium channels in central neurons, anti-peptide antibodies specific for the class C and class D alpha1 subunits were produced. A nti-CNC1 directed against class C immunoprecipitated 75% of the L-type channels solubilized from rat cerebral cortex and hippocampus. Anti-C ND1 directed against class D immunoprecipitated only 20% of the L-type calcium channels. Immunoblotting revealed two size forms of the class C L-type alpha1 subunit, L(C1) and L(C2), and two size forms of the c lass D L-type alpha1 subunit, L(D1) and L(D2). The larger isoforms had apparent molecular masses of approximately 200-210 kD while the small er isoforms were 180-190 kD, as estimated from electrophoresis in gels polymerized from 5% acrylamide. Immunocytochemical studies using CNC1 and CND1 antibodies revealed that the alpha1 subunits of both L-type calcium channel subtypes are localized mainly in neuronal cell bodies and proximal dendrites. Relatively dense labeling was observed at the base of major dendrites in many neurons. Staining in more distal dendr itic regions was faint or undetectable with CND1, while a more signifi cant level of staining of distal dendrites was observed with CNC1, par ticularly in the dentate gyrus and the CA2 and CA3 areas of the hippoc ampus. Class C calcium channels were concentrated in clusters, while c lass D calcium channels were generally distributed in the cell surface membrane of cell bodies and proximal dendrites. Our results demonstra te multiple size forms and differential localization of two subtypes o f L-type calcium channels in the cell bodies and proximal dendrites of central neurons. The differential localization and multiple size form s may allow these two channel subtypes to participate in distinct aspe cts of electrical signal integration and intracellular calcium signali ng in neuronal cell bodies. The preferential localization of these cal cium channels in cell bodies and proximal dendrites implies their invo lvement in regulation of calcium-dependent functions occurring in thos e cellular compartments such as protein phosphorylation, enzyme activi ty, and gene expression.