To investigate how circadian rhythm influences hematoma clearance after intracerebral hemorrhage (ICH) through modulation of melatonin signaling and key components of the brain clearance system.

Hematoma accumulation drives secondary injury after ICH. The endogenous brain clearance system—including the glymphatic pathway, blood–brain barrier (BBB) clearance, and phagocytic receptor-mediated scavenging—plays a crucial role in removing blood and metabolites. Melatonin, under circadian control, regulates aquaporin-4 (AQP4) polarization, BBB permeability, and microglial activity. Disruption of circadian rhythm may therefore impair hematoma clearance.

Male mice underwent collagenase-induced ICH and received circadian disruption (Clock knockout or 48 h sleep deprivation), light intervention (daytime vs. nighttime blue-light exposure), or pharmacological modulation (melatonin + agonist agomelatine vs. receptor antagonist luzindole). Neurological deficits, hematoma volume, BBB permeability, oxidative stress, and neuronal injury were assessed. Expression of AQP4, low-density lipoprotein receptor–related protein 1 (LRP1), occludin (OCLN), and melatonin receptor (MT1) was measured by qPCR, Western blot, and immunofluorescence.

Circadian disruption exacerbated neurological deficits, hematoma enlargement, BBB leakage, ROS accumulation, neuronal degeneration, and iron deposition. It reduced MT1, AQP4, LRP1, and OCLN expression and impaired glymphatic drainage. Conversely, melatonin + agomelatine treatment or daytime rhythm restoration enhanced melatonin signaling, improved BBB integrity, reduced oxidative and neuronal injury, and promoted hematoma clearance with increased AQP4/LRP1/OCLN expression and tracer drainage to cervical lymph nodes.

Circadian rhythm promotes hematoma resolution after ICH by upregulating melatonin-dependent AQP4/LRP1/OCLN expression and enhancing glymphatic and vascular clearance pathways. Strengthening circadian rhythm or melatonin signaling may represent a novel therapeutic strategy for improving outcomes after hemorrhagic stroke.