The gene encodes the 1A subunit of voltage-gated CaV2. In this

The gene encodes the 1A subunit of voltage-gated CaV2. In this mouse model with a cell type-specific ablation of CaV2.1 channels, we show that ablation of CaV2.1 channels restricted to Purkinje cells is sufficient to cause cerebellar ataxia. We demonstrate that spatial ablation of CaV2.1 channels may help in unraveling mechanisms of human disease. Electronic supplementary material The online version of this article (doi:10.1007/s12311-011-0302-1) contains supplementary material, which is available to authorized users. gene and are widely expressed throughout the nervous system [2, 3]. CaV2.1-1A-knockout (1KO) mice that lack CaV2.1 channels exhibit a severe phenotype of ataxia and dystonia with a strong cerebellar component that starts around postnatal day (P)12, and die, if not given very special care, around P20 [4C7]. Histological analysis in the few survivors indicated that a progressive gradual loss of Purkinje cells started between P45 and P100 [5]. Cerebellar ataxia and dystonia can also be part of the phenotype of certain naturally occurring mouse mutants. For instance, and mice, it was shown that the chronic ataxia is usually related to irregular Purkinje cell simple spike firing caused by the loss of precision in intrinsic activity of Purkinje cells and aberrant synaptic input [12C15]. In addition to the cerebellar ataxia phenotype, mice also exhibit episodic dystonia, which is usually related to transient low-frequency oscillations in the cerebellar cortex [16]. Notably, when mutants drop their Purkinje cells on a Purkinje cell degeneration mutant background, the dystonic phenotype disappears [17]. Together these findings indicate that mutations that cause a reduction of CaV2.1-mediated Ca2+-influx affect the information processing in the BIIB021 cerebellar cortex and, thereby, induce chronic ataxia and episodic dystonia. Apart from the aberrant activity in the cerebellar cortex in mutants, also the activity of cerebellar nuclear (CN) neurons is usually subject to abnormal firing patterns, as was recently shown in mice [18]. Both Purkinje cells and CN neurons receive input (direct and indirect through local interneurons and/or granule cells) from mossy fibers and rising fibers. While granule cell transmission to Purkinje cells in 1KO, and mice is usually abnormal, the response to rising fiber activation is usually seemingly normal in these mutants [15, 19, 20]. Besides the excitatory input from rising and mossy fibers, the neurons in the CN receive input from local interneurons and from the cerebellar cortex through Purkinje cells. Apart from the impact of these inputs, the firing pattern of CN neurons is usually decided by their intrinsic activities. Since Purkinje cells and CN neurons, as well as the neurons and fibers that innervate them, express CaV2.1 channels [3], the exact origin of the aberrant cerebellar activity in CaV2.1 mutants that causes cerebellar ataxia could not be resolved using the existing mouse mutants. Therefore, we used the Cre-lox system to ablate CaV2.1 channels exclusively in Purkinje cells by crossing conditional mice that carry a floxed allele [21] with ITM2B transgenic mice that express Cre recombinase under the control of the Purkinje cell-specific promoter [22]. Hereby, we were not only able to rescue the early lethality seen in 1KO BIIB021 mice, but also to demonstrate that lack of CaV2.1 channel function in Purkinje cells is sufficient to cause ataxia and that progressive Purkinje cell loss starts well after the onset of the ataxia. Materials and Methods Animals All animal experiments were performed in accordance with guidelines of the respective universities and the national legislation. Previously, we generated conditional BIIB021 mice using a gene-targeting approach [21]. The allele in these mice was floxed by introducing sites flanking exon 4 (i.at the. mice generating double heterozygous transgenic offspring. Female double transgenic mice were then crossed with (and thereby CaV2.1 channel deficient) in Purkinje cells (i.at the. Purkinje BIIB021 cell CaV2.1-1AKO (PC1KO)). The same cross also produced mice in which was not affected (wild type (WT)). In total, we used 29.