A whole-brain single-cell atlas of circadian neural activity in mice

The mammalian brain comprises numerous anatomical regions with distinct functions despite their extensive connectivity. However, how spontaneous neural activity is coordinated across regions over the circadian cycle remains elusive. We employed tissue clearing and whole-brain c-Fos immunostaining on 144 mouse brains collected over two days under constant darkness. Time-series analysis revealed brain-wide circadian rhythmicity at single-cell resolution, with 79% of the 642 anatomically defined regions oscillating in diverse circadian phases that delineate functional specializations. Voxel-wise analyses further highlighted distinct subregions, suggesting intricate spatiotemporal coordination within regions. Additionally, brain circadian time can be accurately inferred from global c-Fos patterns using omics-derived prediction methods. This whole-brain circadian atlas enhances our understanding of neural coordination and provides a resource for integrating time-of-day information into functional and pharmacological research.