Abstract 3098: Docosahexaenoic Acid Protects The Brain After Experimental Stroke By Modulating Protein Phosphorylation In The Penumbra

Abstract

Background and purpose: Docosahexaenoic acid (DHA), a member of the omega-3 essential fatty acid family, improves behavioral deficit and reduces infarct volume and brain edema after focal cerebral ischemia in rats. We hypothesize that DHA elicits neuroprotection by inducing protective cellular cascades, which in turn lead to cell survival. We used behavioral tests in conjunction with immunohistopathology and western blots to expand our understanding of the early mechanisms responsible for DHA-induced neuroprotection. Methods: Physiologically-controlled male Sprague-Dawley rats (304-370g) received 2h middle cerebral artery occlusion (MCAo) by intraluminal suture. Animals were treated with DHA (5mg/kg, iv) or saline (5ml/kg) at 3h after the onset of stroke. Behavioral function was evaluated at 1h after occlusion onset and at 24h on a grading scale of 0-12 (0=normal and 12=maximal deficit). GFAP (reactive astrocytes), ED-1 (activated microglia/microphages), and NeuN (neurons) were analyzed in the ischemic core and the penumbra regions (A, B and C) at 4 and 24h. Western blot/immunohistochemical analysis of pAKT, pGSK, and pS6 (proteins of the AKT signaling pathway) was conducted in the core and penumbra at 4 and 24h. Results: Physiological variables were stable and showed no significant differences between groups. Behavioral deficit was significantly improved by treatment with DHA compared to vehicle at 24h (7.3±0.5 vs. 10±0; p<0.001, respectively). DHA treatment significantly reduced the number of microglia cells in penumbra regions A and C (by 86% and 76%) and increased GFAP-positive cells in penumbra regions B and C (by 53% and 51%) at 24h after stroke. There were no significant differences in expression of cell markers between groups at 4h and in the ischemic core. DHA upregulated activation of AKT compared to vehicle at 4h (36% increase in p473 AKT and 79% increase in p308 AKT) and at 24h (114% increase in p473 AKT; 98% increase in p308 AKT). DHA also increased pS6 at 4h by 160% and pGSK at 24h by 61% as compared to vehicle-treated animals. DHA-treated animals had significantly higher p308 AKT cell counts in the penumbra at 4h (region A by182%, region B by 160% and region C by 103% increase). Conclusions: DHA treatment confers behavioral improvement, reduces microglial infiltration, increases astrocytosis at 24h, activates pro-survival AKT cascades as early as 4h, and has potential for the treatment of ischemic stroke.