Enhancer-AAVs to monitor and restore cell-type selective inhibitory deficits and circuit dysfunction in humanized Alzheimer's disease models - SUMMARY Alzheimer’s disease (AD) is a progressive neurodegenerative disorder that results in cognitive decline and altered brain network activity, including neural hyperactivity and changes in oscillatory rhythms. These brain network alterations correlate with AD-related pathological changes, including Ab and tau accumulation, and cognitive dysfunction in early and late stages of AD. Inhibitory interneuron impairment, including parvalbumin (Pvalb) and somatostatin (Sst) cells, has been suggested to underlie AD-related network dysfunction, as they play a critical role organizing information flow and timing neuronal firing required for cognitive processing. However, the contribution of Pvalb and Sst cells and their interactions to brain network and behavioral deficits in AD remains poorly understood due to current methodological limitations in monitoring and manipulating multiple cell types independently in vivo, as well as by the constraints of conventional behavioral approaches in assessing cell-type-specific manipulations. To overcome these methodological barriers, we propose an innovative approach that includes (1) the use of dual cell-type-specific enhancer-AAVs to monitor and manipulate two interneuron cell populations independently in freely moving mice, and (2) the implementation of our recently developed machine learning (ML) approach, ML-VAME, to assess AD-dependent behavioral changes and evaluate the efficacy of the proposed cell-type-specific manipulations. Specifically, we propose to monitor with Ca2+ indicators (GCaMP8s and jRGECO) and to manipulate with excitatory opsins (ChR2 and Chrimson) inhibitory Pvalb and Sst cell-types in wildtype (WT) and AD-knock in (KI) mouse models to assess their contribution to AD-related brain network dysfunction, including oscillatory deficits, network hypersynchrony, and behavioral deficits. In Phase 1, we propose to validate dual enhancer-AAV vectors expressing opsins or Ca2+ sensors in Pvalb and Sst cells in young (1 month) and aged (12 months) WT mice (Aim 1) and assess their suitability to monitor and manipulate these cell populations in vivo (Aim 2). We will validate the selected approach in aged 12-month-old AppSAA/ApoE4 mice (Aim 3). Our phase 1 milestone is to successfully demonstrate cell- specific single and dual expression of enhancer-AAVs (Aim 1) and their suitability for monitoring Ca2+ dynamics and for eliciting optogenetic manipulations (Aim 2) in freely moving WT mice, and validate the selected approach in aged AppSAA/ApoE4 mice (Aim 3). In Phase 2, we will define Pvalb and Sst cell type deficits through dual Ca2+ imaging (Aim 4) and optogenetic tagging (Aim 5a) in behaving 12-month-old AppSAA/ApoE4 and AppSAA/MAPTN279K mice, and determine whether optogenetic stimulation of these interneuron cell types restores network and behavioral functions, as well as transcriptomic and pathological changes (Aim 5b). Overall, we aim to develop and deploy dual enhancer-AAV vectors to identify cell-type-selective interneuron deficits and to restore circuit, molecular, and cognitive alterations in humanized AD KI mouse models.
