Microglial morphology aligns with vigilance stage‐specific neuronal oscillations in a brain region‐dependent manner

Microglia, the resident immune cells in the brain, dynamically adapt their morphology based on their functional state. This study explored the relationship between microglial morphology and sleep–wake cycles in mice. Using Iba1 immunostaining to identify microglia, we quantified morphological changes in microglia at different timepoints in multiple brain regions (cortex, hippocampus, basal forebrain, hindbrain, and cerebellum) in B6 male mice using semi‐automated 3D structural analysis. Simultaneously, in a separate group, we monitored wake and sleep stage‐specific brain activity using EEG/EMG recordings. During natural sleep–wake cycles, we observed increased microglial complexity (enlarged volume, territorial coverage, and ramification) during wakefulness, characterized by high‐frequency theta (8–12 Hz) and gamma activity (30–80 Hz). Conversely, during NREM sleep, which is dominated by delta activity (0.5–4 Hz), microglia displayed reduced complexity. Notably, this pattern was absent in brain regions lacking direct functional connections to areas generating vigilance stage‐dependent thalamocortical oscillations. We then extended wakefulness to decouple circadian influence from sleep–wake‐specific neuronal activity. This procedure attenuated the decrease in microglial complexity observed during natural sleep, suggesting a crucial role for neuronal activity. Subsequent recovery sleep restored microglial features, independent of the time of day (zeitgeber time). These findings reveal a dynamic interplay between vigilance stage‐specific thalamocortical activity and microglial morphology across various brain regions. This suggests a potential role for microglia in sleep regulation and warrants further investigation to understand the underlying mechanisms. Main Points Microglia complexity ↑ during wakefulness (theta/gamma activity) & ↓ during NREM (delta activity). Effect is lost in brain areas without connection to vigilant stage‐specific activity. Extending wakefulness attenuated ↓ in microglial complexity.