| Abstract |
Microglia, the brain's innate immune cells, possess complex, highly motile branched processes. These act independently, enabling individual processes to carry out entirely distinct functions in parallel. Intracellular Ca2+ signaling is implicated in many of these distinct microglial functions. However, it has been difficult to quantify how such Ca2+ activity is compartmentalized in space and time to prevent unwanted cross-talk between signaling pathways. Previous studies have typically relied on manually drawn regions-of-interest (ROIs), which averages fluorescence within predefined compartments and therefore cannot resolve the fine-scale spatio-temporal propagation patterns that may be functionally relevant. To address this, we adopt an unbiased non-ROI-based analytical approach to comprehensively characterize the temporal, spatial and spatio-temporal dimensions of microglial Ca2+ activity in vivo. We find that microglial Ca2+ activity predominantly occurs in processes, tends to remain localized at its site of origin, and, when it propagates, often follows a well-defined direction (either toward or away from the soma) rather than spreading isotropically as would be expected under purely passive diffusion. The tendency of microglial Ca2+ activity to spread between intracellular regions does not correlate with peak amplitude, but appears to be limited by the branching points of the microglial processes. Finally, we show that Ca2+ activity can differ between the microglial soma and its processes in response to various pharmacological stimuli. These results suggest that Ca2+ signals are actively compartmentalized within microglia in a context dependent manner, rather than being synchronized across the entire cell.
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