Loss of synapses is the variable that currently appears to correlate best with the degree of dementia in Alzheimer's disease (AD). Based on the cytoskeletal disruption associated with the formation of neurofibrillary tangles (NFT), and their constituent, """"""""abnormally"""""""" phosphorylated tau, we hypothesize that neurons with NFT, rather than their adjacent tangle-free neurons, are at least partially responsible for this loss of synapses (whether this precedes neuron death remains an open question). Our focus in this proposal is to test hypotheses related the effects(s) (or lack thereof) of NFT and of tau phosphorylated at different sites (and combinations of sites) on the expression of messages related to synaptic and neuronal function: synaptophysin, GAP-43, heat shock proteins, cathepsins. Tau has 21 potential phosphorylation sites. The studies we propose are driven by a model of tau function in which the microtubule binding domain (the repeat region) is surrounded by """"""""jaws"""""""" regions which make important contributions to tau promotion of microtubule assembly. We will use in situ hybridization for synaptophysin and other selected messages to determine the molecular status of single neurons combined with double immunocytochemistry (ICC) to define classes of neurons. We will compare grain counts over single neurons from the classes of neurons revealed by antibody combinations: Three general classes of neurons will appear in AD brain: 1) with NFT (phospho-tau may also be apparent in these neurons), 2) without NFT but with phospho-tau at defined sites and combinations of sites, and 3) neither NFT nor phospho-tau. Neurons without evidence of NFT or phospho-tau will be sampled from control brain. Disease-free control and AD brain sections will be processed side-by-side on the same slides for quantitative comparison. The general hypotheses to be tested are, briefly: Hypothesis I: Neurons with NFT will have decreased message for proteins related synaptic structure and function and the growth cone. Hypothesis II: Tau phosphorylation in the """"""""jaws"""""""" region will have a greater influence on expression of synaptic and stress related messages than will tau phosphorylation at Ser 262 in the repeat, microtubule binding region of tau. Hypothesis III: Neurons with NFT or abnormally phosphorylated tau will have increased levels of message for proteins related to cellular stress. Hypothesis IV: There will be """"""""unaffected"""""""" neurons in AD brain that our measures will not distinguish from homologous neurons in control brain.
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