Abstract Body: Hyperhomocysteinemia, a metabolic disorder characterized by systemic elevation of amino acid homocysteine, is linked to deficiency in vitamins and enzymes involved in homocysteine metabolism. The incidence of hyperhomocysteinemia is high in the elderly population due to reduced micronutrient absorption and metabolism. Recent preclinical studies in animal models of stroke showed that predisposition to hyperhomocysteinemia exacerbates ischemic brain injury and worsens behavioral deficits, establishing hyperhomocysteinemia as a potential comorbidity for stroke. These studies further showed that hyperhomocysteinemia-induced stimulation of GluN2A subunit containing NMDA receptors (GluN2A-NMDARs) in conjunction with glutamate-induced stimulation of GluN2B subunit containing NMDARs (GluN2B-NMDARs) exacerbates stroke pathology. However, the precise intracellular mechanisms involved in such exacerbation of ischemic brain injury is not known. The current study sought to investigate whether under hyperhomocysteinemic condition GluN2A-NMDAR mediated early activation of the transcription factor Nuclear Factor-κB (NFκB) and subsequent expression of the pro-inflammatory chemokine Monocyte chemoattractant protein-1 (MCP-1) enhances post-ischemic inflammatory response and exacerbation of brain damage.

Adult male Wistar rats were made hyperhomocysteinemic by implantation of Alzet osmotic pumps containing L-homocysteine. The animals were hyperhomocysteinemic within 3 days with a total plasma homocysteine level of ~24 μM. Acute ischemic stroke was induced by middle cerebral artery occlusion for 60 min followed by reperfusion (6h). Brain tissues were processed for MCP-1 ELISA and RT-PCR. A significant increase in MCP-1 mRNA and protein levels were observed in the ipsilateral cortex of hyperhomocysteinemic rats compared to control rats within 6h of reperfusion. An accelerated brain damage was also observed at this time in the hyperhomocysteinemic rats. Inhibition of GluN2A-NMDAR (NVP-AAM077, i.v.) and NFκB (BMS345541, IKK inhibitor; i.p) at the onset of ischemic insult attenuated increase in MCP-1 mRNA and protein levels, as well as exacerbation of brain injury under hyperhomocysteinemic condition. The study provides compelling evidence for a novel role of GluN2A-NMDAR in promoting neuro-inflammatory response that contributes to exacerbation of stroke pathology under hyperhomocysteinemic condition.