Neurodegenerative diseases represent an ever-increasing societal and economic burden with WHO estimates indicating that they will replace cancer as the second leading cause of death by 2040. Degenerative ataxias are a common form of neurodegeneration affecting cerebellar Purkinje cells (PCs) and other neurons in the CNS. Spinocerebellar ataxia type 2 (SCA2) is an autosomal dominant form of ataxia caused by expansion of a coding CAG repeat and belongs to the group of polyglutamine (polyQ) diseases. Mutations in the SCA2 (ATXN2) gene can also cause or contribute to the development of Parkinson disease and ALS, two common forms of neurodegeneration. Neither symptomatic nor neuroprotective agents have been identified for the treatment of human ataxias. In the previous funding period we developed a multidisciplinary approach to characterize several SCA2 mouse models combining morphologic, biochemical, physiologic, and behavioral techniques. We used the cerebellar slice preparation to show that a decrease in PC firing frequency closely mirrored the decline in motor function. Importantly, significant transcriptome and neurophysiological changes occurred before any dendritic or cell loss was apparent in the cerebellum. We discovered that mutant ATXN2 induced abnormal calcium release from the ER via enhanced interaction with the inositol-triphosphate receptor (ITPR1). Based on these findings, two specific aims are proposed.
In Aim 1, we will test the hypothesis that the mGluR1-ITPR1 axis is hyperactive in cerebella from SCA2 mice expressing polyQ expanded ATXN2. We will use the cerebellar slice preparation to evaluate mGluR1 signaling and the relationship between PC firing and intracellular calcium. We will also test the mGluR1-ITPR1 axis in vivo by genetic interaction with mGluR1 haploinsufficient mouse lines.
In aim 2, we will use transcriptome profiling at presymptomatic time points to identify novel genes involved in SCA2 pathogenesis. To increase sensitivity and the ability to identify changes in less abundantly PC-expressed genes we have established laser-capture microdissection (LCM) of cerebellar regions. Transcriptomes will be analyzed by WGCNA, GO and KEGG pathway analysis with a particular emphasis on mGluR1-ERK downstream targets. Changes in key genes will be verified by qPCR and ICC/ western blot analysis. We hypothesize that some expression changes will be homeostatic, but others will contribute to pathogenesis. We will differentiate between these alternatives by normalizing expression of the respective gene using AAV- transduction in vivo. Five genes will be chosen based on presence of early changes and presence of changes in multiple SCA2 mouse models. Mice will be evaluated by behavioral testing as well as analysis in the cerebellar slice. The proposed experiments address two significant questions relating to SCAs: the importance of intracellular calcium levels on PC function and survival and the identification of novel path- ways that are shared across multiple SCA2 rodent models. Answers to these questions will help in the identification of new avenues towards treatments of SCA2 and other degenerative ataxias.
Neurodegenerative diseases are imposing an increasing health and financial burden given the rapid increase of the aging segment of the population. The spinocerebellar ataxias are inherited neurodegenerative disorders affecting brain circuits involved in movement coordination. This grant will use a multidisciplinary approach to identify novel targets for therapy of ataxias. We will focus on normalizing disturbed calcium balance within cells of the cerebellum using mouse models of the human disease. Our approaches will likely have a broad impact on other neurodegenerative diseases as well.