We have previously shown that melatonin reduces infarct volumes and enhances neurobehavioral and electrophysiological recoveries following transient middle cerebral artery (MCA) occlusion in rats. In the study, we examined whether melatonin would display neuroprotection against neuronal, axonal and oligodendrocyte pathology after 24 hr of reperfusion following 1 hr of MCA occlusion in mice. Melatonin (5 mg/kg) or vehicle was given intraperitoneally at the commencement of reperfusion. Neurological deficits were assessed 24 hr after ischemia. Gray matter damage was evaluated by quantitative histopathology. Axonal damage was determined with amyloid precursor protein and microtubule-associated protein tau-1 immunohistochemistry to identify postischemic disrupted axonal flow and oligodendrocyte pathology, respectively. Oxidative damage was assessed by 8-hydroxy-2'-deoxyguanosine (8-OHdG) and 4-hydroxynonenal (4-HNE) immunohistochemistry. Relative to controls, melatonin-treated animals not only had a significantly reduced volume of gray matter infarction by 42% (P<0.001), but also exhibited a decreased score of axonal damage by 42% (P<0.001) and a reduction in the volume of oligodendrocyte pathology by 58% (P<0.005). Melatonin-treated animals also had significantly reduced immunopositive reactions for 8-OHdG and 4-HNE by 53% (P<0.001) and 49% (P<0.001), respectively. In addition, melatonin improved sensory and motor neurobehavioral outcomes by 47 and 30%, respectively (P<0.01). Thus, delayed (1 hr) treatment with melatonin reduced both gray and white matter damage and improved neurobehavioral outcomes following transient focal cerebral ischemia in mice. The finding of reduced oxidative damage observed with melatonin suggests that its major mechanisms of action are mediated through its antioxidant and radical scavenging activity.

We have previously shown that exogenous melatonin improves the preservation of the blood-brain barrier (BBB) and neurovascular unit following cerebral ischemia-reperfusion. Recent evidence indicates that postischemic microglial activation exaggerates the damage to the BBB. Herein, we explored whether melatonin mitigates the cellular inflammatory response after transient focal cerebral ischemia for 90 min in rats. Melatonin (5 mg/kg) or vehicle was given intravenously at reperfusion onset. Immunohistochemistry and flow cytometric analysis were used to evaluate the cellular inflammatory response at 48 hr after reperfusion. Relative to controls, melatonin-treated animals did not have significantly changed systemic cellular inflammatory responses in the bloodstream (P > 0.05). Melatonin, however, significantly decreased the cellular inflammatory response by 41% (P < 0.001) in the ischemic hemisphere. Specifically, melatonin effectively decreased the extent of neutrophil emigration (Ly6G-positive/CD45-positive) and macrophage/activated microglial infiltration (CD11b-positive/CD45-positive) by 51% (P < 0.01) and 66% (P < 0.01), respectively, but did not significantly alter the population composition of T lymphocyte (CD3-positive/CD45-positive; P > 0.05). This melatonin-mediated decrease in the cellular inflammatory response was accompanied by both reduced brain infarction and improved neurobehavioral outcome by 43% (P < 0.001) and 50% (P < 0.001), respectively. Thus, intravenous administration of melatonin upon reperfusion effectively decreased the emigration of circulatory neutrophils and macrophages/monocytes into the injured brain and inhibited focal microglial activation following cerebral ischemia-reperfusion. The finding demonstrates melatonin's inhibitory ability against the cellular inflammatory response after cerebral ischemia-reperfusion, and further supports its pleuripotent neuroprotective actions suited either as a monotherapy or an add-on to the thrombolytic therapy for ischemic stroke patients.