Alternative splicing generates a smaller assortment of CaV2.1 transcripts in cerebellar Purkinje cells than in the cerebellum. Physiol Genomics 24: 86 -96, 2006. First published November 8, 2005 doi:10.1152/physiolgenomics.00149.2005.-P/Qtype calcium channels control many calcium-driven functions in the brain. The CACNA1A gene encoding the pore-forming CaV2.1 (␣1A) subunit of P/Q-type channels undergoes alternative splicing at multiple loci. This results in channel variants with different phenotypes. However, the combinatorial patterns of alternative splice events at two or more loci, and hence the diversity of CaV2.1 transcripts, are incompletely defined for specific brain regions and types of brain neurons. Using RT-PCR and splice variant-specific primers, we have identified multiple CaV2.1 transcript variants defined by different pairs of splice events in the cerebellum of adult rat. We have uncovered new splice variations between exons 28 and 34 (some of which predict a premature stop codon) and a new variation in exon 47 (which predicts a novel extended COOH-terminus). Single cell RT-PCR reveals that each individual cerebellar Purkinje neuron also expresses multiple alternative CaV2.1 transcripts, but the assortment is smaller than in the cerebellum. Two of these variants encode different extended COOH-termini which are not the same as those previously reported in Purkinje cells of the mouse. Our patch-clamp recordings show that calcium channel currents in the soma and dendrites of Purkinje cells are largely inhibited by a concentration of -agatoxin IVA selective for P-type over Q-type channels, suggesting that the different transcripts may form phenotypic variants of P-type calcium channels in Purkinje cells. These results expand the known diversity of Ca V2.1 transcripts in cerebellar Purkinje cells, and propose the selective expression of distinct assortments of Ca V2.1 transcripts in different brain neurons and species. splice variants; P type; calcium channels; Purkinje neurons 0 VOLTAGE-GATED calcium (Ca 2ϩ ) channels regulate numerous cellular functions, from neuronal electrical activity to intracellular signaling pathways. Diversity of Ca 2ϩ channels in the brain is apparent from the many roles that Ca 2ϩ channels play in different cell types and subcellular compartments (3). The diversity reflects, in part, the existence of nine classes of the pore-forming Ca V (␣1) subunit in the brain and alternative pre-mRNA splicing of the gene encoding each class of Ca V subunit (12, 17). P/Q-type Ca 2ϩ channels control many functions throughout the brain and contain a Ca V 2.1 subunit (also known as ␣ 1A ). However, information about splicing of the CACNA1A gene encoding Ca V 2.1 in different brain regions and neurons is incomplete (2,5,16,30,33,35,37). Augmentation of what is currently known about Ca V 2.1 splicing in the cerebellum and cerebellar Purkinje cells is of particular interest, because P-type currents in Purkinje cells are the prototypical P-type currents against which putative P-type and Q-...

To determine if the properties of Ca2+ channels in cerebellar Purkinje cells change during postnatal development, we recorded Ca 2+ channel currents from Purkinje cells in cerebellar slices of mature (postnatal days (P) 40-50) and immature (P13-20) rats. We found that at P40-50, the somatic Ca 2+ channel current was inhibited by ω-agatoxin IVA at concentrations selective for P-type Ca 2+ channels (∼85%; IC 50 , <1 nM) and by the dihydropyridine ( Voltage-gated Ca 2+ channels regulate electrical activity and many intracellular processes (Catterall et al. 2005). The classification of Ca 2+ channels as T-, N-, L-, P-, Q-or R-type is based on the electrophysiological and pharmacological properties of Ca 2+ channel currents in different cell types (Catterall et al. 2005). A finer subdivision of this classification follows from the identification of 10 classes of the pore-forming Ca V (α1) Ca 2+ channel subunit and alternative pre-mRNA splicing of the gene encoding each Ca V class, as well as different classes and splice variants of the accessory α 2 δ, β and γ subunits (Catterall et al. 2005).Currents mediated by native L-type channels or by recombinant channels consisting of one of the four classes of the Ca V 1 family (Ca V 1.1, Ca V 1.2, Ca V 1.3 and Ca V

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