Abstract 723: A Nitric Oxide-donor Based Vasodilatory Therapy Mitigates Hypoxia-induced Brain Injury on a Zebrafish Model of Hypoxic Ischemic Encephalopathy

Abstract

Hypoxic ischemic encephalopathy (HIE), a brain injury caused by cerebral hypoxia due to perinatal asphyxia, has high mortality, high morbidity and severe sequelae, but therapeutic options remain limited. Cerebral vasoconstriction has been known to contribute to neuronal damage of HIE. Here we report a zebrafish model of brain injury induced by hypoxia and reoxygenation to mimic HIE, and explore the therapeutic potential of a vasodilatory agent to mitigate neuronal damage. Larval zebrafish (6 dpf) were immersed in a hypoxic medium (dissolved O2 below 0.2 ppm) for 15 min to induce brain injury, and then a normoxic medium to mimic resuscitation. The vascular width, blood flow and cerebral cell death were assessed with confocal imaging on transgenic fish that express fluorescent proteins in endothelium, red blood cells (RBCs) and cranial motor neurons, respectively. Neurological function was determined with a modified scoring system based on the movement, coordination and response of zebrafish to stimuli. Larvae survived from the hypoxic assault were categorized according to the severity of impaired cerebral hemodynamics. The serious group was featured with greater cerebral vasoconstriction (32% vs. 6%) and lower blood flow (82 vs. 728 RBCs/min) relative to the moderate group. These two groups showed notably different outcome measured 2 d post hypoxia with the serious group having a significantly higher cerebral cell death (34% vs. 4%), a lower neurological score (2.5 vs. 6) and survival rate (60% vs. 89%) relative to the moderate group. The strong correlation between the post-hypoxia cerebral hemodynamics and the outcome indicates that a vasodilatory therapy might be a strategy to ameliorate hypoxia induced brain injury. A treatment of s-nitrosoglutathione (GSNO, 15 μM), a nitric oxide donor, effectively alleviated vasoconstriction (7.4% vs. 14.3%), improved the neurological score (3.5 vs. 2.5) and increased the survival rate (63% vs. 54%) relative to the untreated control. In conclusion, a zebrafish model of HIE has been developed. We show that cerebral vasoconstriction may contribute the brain injury associated with HIE, and vasodilatory therapies possess a potential for the remedy of HIE.