CaV1.3 is an L-type voltage-gated calcium channel implicated in several functions including gene expression, pacemaking activity, and neurotransmitter release. The gene that encodes the CaVa1-pore forming subunit of CaV1.3 (Cacna1d) is a multi-exon gene that undergoes extensive alternative splicing, which provides functional versatility to this gene across tissues and cell-types. The function and expression of several Cacna1d splice variants within the C-terminus have been previously characterized. These splice variants differ in their voltage-dependence of activation, Ca 2+ -dependent inactivation, and their sensitivity to dihydropyridines. However, less is known about alternatively spliced exons in Cacna1d located downstream of domain I and upstream of the C-terminus (e11, e22a/e22, e31a/e31b/e32). Here, we performed a systematic study to determine the developmental and cell-specific expression of several Cacna1d splice variants. We found that the cassette e11 is upregulated during brain development, and in adult cortical tissue is more abundant in excitatory neurons relative to inhibitory interneurons. This exon is also upregulated upon nerve growth factor (NGF) induced differentiation of pheochromocytoma cells, PC12. At the functional level, the splice variants resulting from e11 alternative splicing (+e11-Cacna1d and De11-Cacna1d) form functional CaV1.3 channels with similar biophysical properties in expression mammalian systems. Of the pair of mutually exclusive exons, e22a and e22, the later dominates at all stages. However, we observed a slight upregulation of e22 from embryonic to adult human brain. A second pair of mutually exclusive exons, e31a and e31b, was also studied. We found that e31a increases during brain development. Finally, the cassette exon 32 is repressed in adult brain tissue. KEYWORDSCalcium channels, alternative splicing, splicing factors, brain development Parkinson's disease, primary aldosteronism and autism spectrum disorder (Chan et al., 2010;Baig et al., 2011;Kang et al., 2012;Reinbothe et al., 2013; Scholl et al., 2013; Pinggera et al., 2015). Interestingly, Cacna1d splice variants add functional versatility of CaV1.3 channels across tissues and cell-types.CaV1.3, like all CaVa1 subunits, is organized into four homologous domains (DI-IV). Each domain contains six transmembrane-spanning segments (S1-S6), with a reentrant loop between S5 and S6, which contains glutamates that are key for the selectivity filter. The amino and carboxyl termini, as well as linker sequences between the DI-II, DII-III, and DIII-IV are cytosolic (Fig. 1A). Sites of alternative splicing of sequences encoding for the N-terminus, DI, DI-II linker, DIII, DIV and C-terminus are present in Cacna1d pre-mRNAs. Alternative splicing of sequences encoding for the C-terminus in CaV1.3 have been well characterized, and results in splice isoforms with differences in calciumdependent inactivation, trafficking, protein-protein interactions, and sensitivity to dihydropyridines (DHPs) (Shen et al., 2006;Singh et al., 200...

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