Microglia engagement and clearance of dying neurons is regulated by CX3CR1

Neuronal cell death is a hallmark of many neurodegenerative diseases. Effective detection and clearance of cell debris is essential to prevent a degenerative cascade. Brain-resident microglia perform these functions through complex cell-cell signaling involving both “find-me” and “eat-me” cues; however, the precise in vivo mechanisms governing microglial responses to neuronal death remain incompletely understood. To examine microglial responses to tissue damage and neuronal cell death in vivo, we investigated neuron/microglia CX3CR1 signaling in mouse neocortex using intravital optical imaging combined with 2Phatal, a single-cell ablation technique that models individual neuron programmed cell death. Disruption of CX3CR1 signaling via Cx3cr1 knockout altered chemotactic responses to focal tissue damage selectively in non-inflammatory contexts. Furthermore, Cx3cr1 knockout led to delayed cell corpse clearance under both low and high neuronal death conditions, partly due to a delay in microglial engagement with dying cells. These findings uncover a precise, context-specific role for CX3CR1 signaling in regulating microglial engagement and phagocytic clearance of dying neocortical neurons in vivo, with implications for understanding microglial dysfunction in neurodegenerative disease.