The pore-forming Ca_V2.1 subunit of P/Q-type Ca channels is encoded by the Cacna1a gene. Human, rat and mouse cDNA and genomic analysis have determined that this gene undergoes alternative splicing at multiple loci. However, it is unclear how Ca_V2.1 splicing differs between brain regions or cell-types or species. For instance, the P-type Ca channel current described in rat cerebellar Purkinje cells is the prototypical current against which putative P-type currents are compared, and yet the full range of Ca_V2.1 splice variants in rat cerebellar Purkinje cells is not known. We used RT-PCR and single-cell RT-multiplex PCR to examine Ca_V2.1 alternative splicing in adult rat cerebellum and in single Purkinje neurons. Splice sites in rodent Ca_V2.1 genes were identified by aligning rat and mouse Ca_V2.1 cDNA sequences. PCR primers were designed to detect the following splicing events: use of different acceptor/donor pairs at the 5′ end of exon (e) 10, inclusion or exclusion (-) of e31a or e43 or e44, coincident exclusion of e33, e36 and e37 (-e33/-e36/-e37) or e43 and e44 (-e43/-e44), mutually-exclusive expression of e37a and e37b, extension or shortening of the 5′ end of e47, and deletion of ~150 nucleotides from e47. RT-PCR was carried out with mRNA extracted from the cerebellar vermis of mature male Wistar rats (6-7 weeks) or on cell contents harvested into a patch-pipette during whole-cell voltage-clamp of Purkinje cells located in a cerebellar slice. Donor rats were humanely killed. Analysis of PCR products confirmed that the adult rat cerebellum expresses multiple Ca_V2.1 transcript variants. However, not all splicing events investigated were detected. These include -e33/-e36/-e37, which occur in a rat pancreatic variant (Ligon et al., 1998) and -e43 or -e43/-e44, which occur in human cerebellar variants (Soong et al. 2002). Furthermore, some cerebellar transcripts are generated by combinations of splicing events not previously described for rat Ca_V2.1, such as e31a/e37a, -e31a/e37b, -e31a/e37a, e37a/-e44, -e31a/-e44 (detected in 70-100% of 8-16 reactions). We found that individual rat cerebellar Purkinje cells also express multiple Ca_V2.1 transcript variants, but the range is smaller than in the cerebellum. For example, the combinations -e31a/e37b, e31a/e37a, e31a/e37b were absent from Purkinje cells (detected in 0-12% of 15-20 cells). Furthermore, we did not detect two of the e47 splicing events previously reported in mouse cerebellar Purkinje cells (Tsunemi et al. 2002). These data show that the cerebellar Purkinje cell expresses multiple Ca_V2.1 transcripts and suggest that cell-specific regulation of alternative splicing results in a smaller assortment of transcripts in Purkinje cells than in the cerebellum. The data also suggest species-specific control of Ca_V2.1 splicing.