Background and Purpose
Alpha-linolenic acid (ALA) and riluzole (RLZ), potent activators of two-pore domain K+ channel TREK-1 and TRAAK are known to be neuroprotective against forebrain global ischemia and epileptic seizures. Recent studies have revealed that TREK-1 plays a major role in neuronal excitability control and ALA-induced neuroprotection. We investigated the potential protective effect of ALA and riluzole in a model of focal ischemia clinically relevant to stroke.
Methods
Mice were subjected to transient middle cerebral artery occlusion (MCAO, 1 hour) and reperfusion by use of intraluminal filament model. To determine the therapeutic window, mice received a single dose of 4 mg/kg RLZ or 500 nmol/kg ALA at different times of reperfusion. Then, the efficiency of a single dose of 4 mg/kg RLZ or 500 nmol/kg ALA injected 1 (RLZ) or 2 hours (ALA) after reperfusion was compared to a dose of 2 mg/kg RLZ and 250 nmol/kg ALA given 1–2, 48 and 72 hours after reperfusion or 1–2 hours after reperfusion and once each week during the next two weeks of reperfusion. A combined treatment with 2 mg/kg RLZ + 250 nmol/kg ALA injected 2 hours after reperfusion was also tested. RLZ and ALA were injected as a bolus directly in the jugular vein. Ischemic neuronal injury was evaluated 24 hours (Infarct volume, neurological deficit, counting of TUNEL-positive cells, Bax expression) and 28 days (neuronal counting on cresyl-violet sections, GFAP expression) after ischemia. The same protocol was also performed with administration of palmitic acid, a saturated fatty acid that did not activate TREK-1 and TRAAK channels.
Results
A single dose of RLZ (4 mg/kg) or ALA (500 nmol/kg) up to 3 hours after reperfusion reduced the stroke volume by 75% and 86 % (P<0. 001), respectively and improved the neurological scoring. ALA- and RLZ-treatment was associated with a reduction in cytopathological features of cell injury, including cell shrinkage, DNA-fragmentation, Bax and GFAP expression both in the cortex and the caudate-putamen. In term of survival rate observed in a 28-day time, the best protection was obtained with the injection of 250 nmol/kg ALA given 2 hours after reperfusion and once each week during the next two weeks of reperfusion or with a single dose of the combined treatment (2 mg/kg RLZ + 250 nmol/kg ALA). The palmitic acid failed to induce a neuroprotective effect against focal ischemia.
Conclusion
These results provide further evidence for a therapeutical value of a riluzole and alpha-linolenic acid treatment in brain injury resulting from focal ischemia/reperfusion.