Multiple pathways contribute to neurodegeneration in Alzheimer’s Disease (AD). Identification of appropriate targets effective for inhibiting disease progression remains elusive. Pathways associated with synaptic plasticity are strongly implicated in the pathophysiology of AD and the associated cognitive decline. They therefore represent attractive targets for pharmacological interventions designed to restore cognitive function. Here, we propose to profile how plasticity and neurotransmitter receptor pathways malfunction in early AD at a single-cell- specific transcriptomic level, both as a function of the cell’s proximity to b-amyloid deposits and as a function of its level of activity recorded in vivo. For cell-specific transcriptomic profiling, we use a recently developed, high-throughput, multiplexed, error-robust fluorescence in situ RNA hybridization method (MERFISH), which implements a combinatorial labeling approach followed by sequential rounds of single-molecule fluorescence in situ hybridization (smFISH) to image simultaneously hundreds to thousands (up to 10000;; Xia et al., PNAS, 2019) of distinct RNA molecules at ~100nm spatial resolution. Aim 1: Develop a pipeline for characterizing Alzheimer’s-disease associated transcriptomic pathways in a topographically-resolved single-cell specific way. Map transcriptomic dysfunction in pathways associated with neural plasticity and neurotransmitter receptor expression in the 5xFAD/PS1 APP mouse model of Alzheimer’s disease. Determine how cell-specific malfunction at the transcriptomic level relates to the proximity of b-amyloid deposits early in the disease course. Neuronal hyperactivity and failure of circuit homeostasis (manifesting as hypersynchrony) confer higher risk of neurodegeneration, are associated with the presence of b-amyloid, and often precede plaque pathology. We will use chronic large-field-of-view, “Mesoscopic,” 2-photon imaging to measure cellular response functions and functional connectivity (hypersynchrony) profiles in vivo. Aligning in vitro MERFISH images with in vivo recordings allows single- cell-specific transcriptomic pathways to be characterized as a function of neuronal activity levels imaged in vivo. Aim 2: Identify functional biomarkers of early neuronal dysfunction and use MERFISH to map the transcriptomic profile of neurons and glial cells at the earliest stage of disease expression. Link transcriptomic measurements to neuronal profiles of abnormal activity obtained by in vivo 2-photon imaging and to the location of b-amyloid deposits. Our aims will 1) identify plasticity and neurotransmitter pathways differentially expressed in all neuronal and glial cell types in response to AD, 2) describe the role of b-amyloid proximity in these modifications, 3) correlate functional defects...