Spinocerebellar ataxia-1 (SCA-1) belongs to a group of dominantly inherited neurodegenerative diseases caused by a mutant expansion of a polyglutamine-repeated sequence within the affected gene product ataxin-1. One of the major cell types affected by ataxin-1 is the cerebellar Purkinje cell. The mechanism by which ataxin-1 causes Purkinje cell degeneration in SCA-1 is not known, however, ataxin-1 down regulates Purkinje cell specific proteins involved in calcium homeostasis and signaling in patients with SCA-1, and in presymptomatic SCA-1 transgenic mice. Therefore, the present proposal is designed to determine if targeted deprivation of one of the specific proteins involved in calcium homeostasis will further enhance ataxin-1 toxicity and if trophic upregulation of this protein will rescue SCA-1 Purkinje cells from degeneration. The long-term goal of this project is to understand the role of calcium signaling pathways in neuronal degeneration in order to design therapeutic approaches in clinical management of SCA-1 and other dominantly inherited cerebellar ataxias. To determine if decreased expression of calcium binding protein calbindin-D 28k (CAB) will increase the toxic effects of ataxin-1 on Purkinje cells, double mutant mice will be generated by mating CaB null mice with SCA-1 transgenic mice. To determine if overexpression of insulin-like growth factor - I (IGF-I) will rescue SCA-1 Purkinje cells from degeneration, double mutant mice will be generated by mating mice overexpressing IGF-I with SCA-1 transgenic mice. The changes in Purkinje cells will be assessed by behavioral, biochemical, immunochemical and immunohistochemical methods, focusing on the alterations in the expression of Purkinje cell markers, calcium binding proteins, and proteins involved in calcium signaling. Purkinje cells cultured from 0-1 day old SCA-1 transgenic mice will be used to determine if cultured Purkinje cells expressing mutant ataxin-1 show (i) altered expression of calcium binding proteins and proteins involved in calcium signaling (ii) sensitivity to increased Ca 2+-influx (iii) altered response to the inhibitors of calcium-dependent proteases and (iv) if treatment with IGF-I can reverse ataxin-1 mediated pathological changes. Purkinje cell cultures from non-transgenic mice will be used as controls. Complementary analysis to compare changes in cultured Purkinje cells with those cultured from CaB null and transgenics with Huntington's disease will also be performed. Further, changes observed in SCA-1 patients and in transgenic mice will be compared with that in Machado-Joseph disease/SCA-3, other cerebellar ataxias and normal controls.
