The long-term goal of my research is to provide a level of understanding of the cellular and molecular mechanisms of neuronal plasticity and degeneration that will allow the development of specific means of preventing and treating neurodegenerative disorders. This proposal tests the hypothesis that regulation of intraneuronal calcium levels [Ca2+]i by environmental signals (growth factors, excitatory amino acids (EAA) and amyloid precursor proteins (APP)) mediates adaptive changes in neuronal structure (plasticity), and that misregulation of these calcium signalling systems is involved in the pathological neuronal degeneration. Rat hippocampal and human cortical cell cultures will be used to examine the effects of growth factors, EAAs and amyloid proteins on the expression and function of specific calcium-regulating proteins (NMDA and AMPA receptors, voltage-dependent calcium channels, Ca2+ ATPase, Na+/Ca2+ exchanger, calcium-binding proteins). Changes in [Ca2+]i will be related to neuronal damage or neuroprotection. Several technological approaches will be employed including: fluorescence ratio (fura-2) and single wavelength (fluo-3) imaging of [Ca2+]i; confocal laser scanning microscopy; Northern and Western blots; immunocytochemistry and in situ hybridization; use of antisense oligonucleotides to selectively suppress protein levels; whole-cell patch clamp recording of ionic currents. We previously found that growth factors (NGF, bFGF, IGFs) can """"""""stabilize"""""""" [Ca2+]i and protect neurons against excitotoxic/ischemia-like insults. This proposal will explore the mechanism of the neuroprotective actions of growth factors. Preliminary data indicate that secreted forms of amyloid precursor protein (APPS) regulate [Ca2+]i in neurons. I propose that APPS play trophic/neuritotropic roles during development, and that they modulate the actions of EAAs in the mature nervous system. We will test the hypothesis that abnormal processing of APP contributes to a loss of neuronal calcium homeostasis and neurofibrillary degeneration in Alzheimer's disease. Taken together, this work will provide an integrated view of the role of systems for that regulate [Ca2+]i in the pathophysiology of stroke and Alzheimer's disease (AD), and identify potential means of preventing and treating neurodegenerative disorders based upon EAA, growth factor, and APP signalling pathways.
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