The principle research focus of this grant is to gain insight into the structure and function of the intracellular Ca2+ sequestering compartments of neurons. These compartments are of great interest because recent evidence has accumulated that they play a protective role in neurons exposed to glutamate, the major excitatory neurotransmitter in the CNS and the cause of """"""""excitotoxicity"""""""". Excitotoxicity is a major cause of neuronal damage in stroke and trauma as well as a likely mediator of neuronal injury in age related neurodegenerative diseases, e.g. Huntington's, ALS.
The specific aims are: 1. To localize the specific isoforms of CaATPase and calreticulin two, by components of neuronal Ca2+ sequestering compartments in brain and cultured neurons and neuroblastoma cells. 2. To investigate the biosynthetic route(s) by which the CaATPase and calreticulin follow to reach the Ca2+ sequestering compartments. 3. To study the effects of agents which raise cytosolic Ca2+ on the mRNA and protein level of CaATPase and calreticulin using neuroblastoma cells and cultured neurons. 4. To transfect neuroblastoma cells with cDNAs for CaATPase and calreticulin to determine if these proteins are protective against toxic effects of elevated cytosolic Ca2+ and 5. To employ intracellular dyes to directly measure cytosolic Ca2+ in neurons and neuroblastoma cells. We will employ biochemical, morphological and molecular techniques in these investigations.
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