CA1 Engram Cell Dynamics Before and After Learning

A fundamental question in neuroscience is how memory formation shapes brain activity at the level of neuronal populations. Recent studies of hippocampal 'engram' cells-neurons identified by learning-induced immediate early gene (IEG) expression-propose that these populations form the cellular substrate for memory. Previous experimental work suggests that cells are recruited into engrams via elevated intrinsic excitability and that learning drives coactivity among these cells to support retrieval. Despite this, an understanding of how engram dynamics evolve across learning and recall remains incomplete. Here, we combined activity-dependent genetic tagging with longitudinal two-photon calcium imaging to track CA1 engram population dynamics before and after fear conditioning. Our results reveal that engram activity is modulated by intrinsic dynamics, behavioral state, and stimulus-cued reactivation. Consistent with the idea that intrinsic dynamics bias engram allocation, spontaneous activity during rest predicted future engram membership-up to two days prior to Fos expression. In parallel, we found sequential activity during locomotion recruited both engram and non-engram cells, but that engram cells were less modulated by velocity. Furthermore, after fear conditioning, within the engram population, we identified a subset of cells that increased their correlations after fear learning, specifically during quiet rest. We also used a trace fear conditioning paradigm to show that conditioned stimulus presentation drove elevated activity and stable correlations amongst engram cells, demonstrating learning-dependent reactivation. Finally, computational modeling of CA3-CA1 circuit dynamics demonstrated that a network with strong excitatory-inhibitory balance, capable of CA3-driven reactivation, is consistent with our experimental results. Together, these results show that engram population dynamics are shaped by spontaneous states, behavior, and memory.