Timing Matters: How Daily Rhythms Affect Remote Ischemic Postconditioning Therapy for Stroke

BACKGROUND: Despite promising preclinical results, remote limb ischemic postconditioning efficacy in human stroke treatment remains unclear, with mixed clinical trial outcomes. A potential reason for translational difficulties could be differences in circadian rhythms between nocturnal rodent models and diurnal humans. METHODS: Male C57BL/6J mice were subjected to transient focal cerebral ischemia and then exposed to remote postconditioning during their active or inactive phase and euthanized at 24 hours and 3 days. Infarct volumes were measured using tetrazolium staining. Behavioral outcomes were assessed using the modified Neurological Severity Score, foot-fault test, and adhesive removal test. Brain (striatum ipsilateral) protein levels of PER2 (period circadian regulator 2) and nNOS (neuronal NO synthase) were analyzed by Western blot at 24 hours and 3 days postischemia. 7-nitroindazole, an nNOS inhibitor, was injected intracerebroventricular, and PER2 protein levels were measured during the inactive phase. RESULTS: Remote postconditioning reduced infarct volumes and improved behavioral outcomes in mice when strokes occurred during the inactive phase (Zeitgeber 1–5) but had no effect during the active phase (Zeitgeber 13–17) 24 hours and 3 days postischemia. During the inactive phase, remote postconditioning reduced PER2 expression and increased nNOS levels; however, these changes were not observed during the active phase. Treatment with the nNOS inhibitor abolished the difference in PER2 expression between the transient middle cerebral artery occlusion and remote limb ischemic postconditioning groups during the inactive phase. CONCLUSIONS: Circadian rhythms influence the neuroprotective effects of remote postconditioning, with greater efficacy observed during the rodent inactive phase up to 3 days. The underlying mechanism may involve nNOS-dependent regulation of PER2 during remote limb ischemic postconditioning. These findings suggest that circadian timing should be considered in remote postconditioning stroke research to improve the translation of neuroprotective strategies from animals to humans.