PROJECT SUMMARY/ABSTRACT The aim of this proposal is to provide the trainee, who has a physiology background, with training in molecular and cellular neurobiology and enable her transition to a post-doctoral position in neurodegenerative therapeutics. The rich training environment, which includes a strong mentoring team of diverse scientists, will equip the trainee with the expertise needed to successfully evaluate cell-type-specific immunometabolic mechanisms that impact tau pathology and neuroinflammation in the context of Alzheimer’s disease (AD). The objective of this proposal is to investigate the influence of cholesterol on astrocytic internalization of tau and the subsequent effect this has on neuronal health and viability. Primary astrocyte and neuron cultures from humanized apolipoprotein E (APOE) ε3 and ε4 mice will be used to test the overall hypothesis that the higher cholesterol levels inherent to APOE ε4 carriers increase tau internalization in astrocytes, evoking astrogliosis and subsequent diminished neuronal health via altered immune and metabolic function. The hypothesis of Aim 1 is that higher astrocytic cholesterol levels increase internalization of tau. To test this, astrocytes stimulated with recombinant tau monomers or aggregates will be evaluated for changes in tau internalization before and after pharmacological modulation of cholesterol levels. Phagocytic inhibitors will be used to evaluate the mechanism of internalization as well as to compare cholesterol-related effects on internalization. Astrocytes from APOE ε3 and ε4 mice will be used to compare the differences in astrocytes expressing the most common APOE isoform verses the AD risk isoform. The hypothesis of Aim 2 is that tau increases APOE ε4 astrocytic inflammatory cytokine secretion, in turn decreasing neuronal health. To test this, astrocytes stimulated with recombinant tau will be assayed by a multiplex cytokine panel to identify an inflammatory signature. Conditioned astrocytic media, as well as cytokines identified to have altered expression, will be used to stimulate primary neurons and their responses will be evaluated by metabolic, synaptic, and viability measures. Achievement of these aims will drive the field of tauopathy-related disease research forward by determining whether cholesterol levels modulate internalization of tau in astrocytes, and if secondary release of proinflammatory cytokines from astrocytes to neurons due to tau stimulation triggers diminished neuronal health. Understanding the influence of APOE isoform status on tau-related pathology will improve scientific knowledge of mechanisms of AD progression. More specifically, this knowledge will be impactful in identifying the role APOE ε4 genetic risk plays in potentiating AD pathology through loss of astrocytic support to neurons. These findings will provide rationale for identification of novel treatment targets for prevention of tau propagation, astrogliosis, and subsequent neurode...