The long term objectives of this proposal are to examine the mechanisms underlie the cytoskeletal alterations associated with Alzheimer's disease (AD) neuropathology. Specifically, we propose to examine i) the ability that sprouting contributes to the cytoskeletal alterations characteristic of AD, and ii) the association of a fetal or 'early' microtubule-associated protein, MAP5, with AD pathology. We hypothesize that there is a fine balance between plasticity and pathology and that sprouting-related alterations may be an early event in the development of AD pathology. The specific questions which this proposal aims to answer include: 1) Is MAP5 present in neurofibrillary tangles, neuritic plaques, or neuropil threads and is MAP5 a component of paired helical filaments? 2) Does the shift in the cellular compartmentation of MAP5 in AD represent an early event in the development of pathology? 3) Are the immunocytochemical alterations accompanied by corresponding changes in mRNA expression? 4) Do similar alterations in protein and mRNA expression occur in the rat brain in association with lesion-induced sprouting? 5) Do components of the neuritic plaque, such as Beta-amyloid and basic FGF, influence the immunoreactivity or phosphorylation of MAP5, tau, and early tubulin isotypes, or alter the expression of their encoding mRNAs? Aims 1-3 will be examined in human brain tissues, obtained at autopsy, from individuals with a clinical diagnosis of 'probable AD' and from neurologically normal controls. Many of these individuals will have been clinically evaluated at the Sanders-Brown Center on Aging.
Aim 4 will examine rats in which either the entorhinal cortex or fimbria-fornix has been lesioned.
Aim 5 will utilize cultured rat hippocampal neurons and represents a collaboration with Dr. Mark Mattson. A variety of experimental methodologies will be used. These include immunocytochemistry, Western blots, in situ hybridization, ribonuclease protection assay, subcellular fractionation, preparation of PHF-enriched fractions, and confocal and immunoelectron microscopy. The immunocytochemistry experiments will employ a panel of antibodies against MAP5. Where possible, the various probes and methods will be used on tissues obtained from the same individual to allow comparison of the results. Results obtained from these experiments will enable us to compare sprouting-related alterations in cytoskeletal proteins with the alterations observed in AD. This is essential for evaluation of the possible role of sprouting in the pathogenesis of Alzheimer's disease.
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