PROJECT SUMMARY/ABSTRACT Defining how we form, store, and retrieve memories is one of neuroscience’s most researched areas. The hippocampus is a well-characterized forebrain structure with a crucial role in the formation and retrieval of episodic memory, and is known to be vulnerable to age-related memory dysfunction. Cognitive decline in age and neurodegenerative disease is an increasing burden on healthcare systems and society at large. To treat the causes of memory decline in age and age-related disease, the mechanisms by which hippocampal cell populations form and maintain individual memories over time must be characterized. Decades of research have shown that new learning results in strengthened synaptic connections between networks of hippocampal neurons. These distributed connections between cells are believed to make up the physical basis for memories, often defined as an engram. Significant progress has been made in finding the engram in the brain with the application of activity-dependent genetic strategies, which isolate populations of cells expressing immediate- early genes (IEGs), to identify the neurons activated by learning. Subpopulations of neurons expressing the IEG c-Fos are activated during learning and reactivated during memory recall, so they are often referred to as engram neurons. Previous studies demonstrated the crucial role of hippocampal engram neurons in memory recall behavior, and artificial reactivation of these cells in mouse models of aging and Alzheimer’s disease rescued memory retrieval deficits, pointing to engram cells as a promising target for future interventions to treat memory deficits. However, the in vivo mechanisms by which these cells store associations, and how their reactivation drives memory retrieval, have yet to be explored. To address this gap in knowledge, this project will utilize novel two-photon imaging to combine an inducible c-Fos tagging strategy with large-scale calcium imaging, to investigate how these cell populations contribute to memory formation. Aim 1 of this project is to characterize the dynamics of engram cell populations across learning in order to inform our understanding of circuit mechanisms underlying memory formation in healthy states. This mechanistic understanding will then be utilized in Aim 2 to define how these processes are negatively impacted by aging. Overall, the aims of this proposal will contribute to the advancement of our understanding of the circuit mechanisms of hippocampal memory formation in health and disease, with the potential to inform current treatment strategies for cognitive decline.