Acute cerebral infarction (ACI) is the most common type of acute cerebrovascular diseases resulting in high rate of death and disability. Numerous evidences show that inflammation is the leading cause of ischemic brain injury, thus anti-inflammatory therapy is an attractive candidate for ischemic brain damage. Eicosapentaenoic acid (EPA) exerts anti-inflammatory activity in lots of human inflammatory diseases, whereas its effect in ACI is left to elucidate. Nlpr3-/- mice, Gpr40-/-; Gpr120-/- mice and mice with right middle cerebral artery occlusion (MCAO) were used to detect NLR family pyrin domain containing 3 (NLRP3) inflammasome activation by Western Blot and the release of proinflammatory cytokines by ELISA. To estimate the acute ischemic condition in vitro, oxygen-glucose deprivation (OGD) was induced in BV2 microglia cells. Transfection of the shRNA targeting GPR40 and GPR120 mRNA into BV2 cells was also assessed. Apoptosis in ischemic cerebral tissues and BV2 cells was detected by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay and flow cytometry. Here we show that EPA suppresses ACI-induced inflammatory responses through blocking NLRP3 inflammasome activation. In addition, EPA inhibits NLRP3 inflammasome activation through G protein-coupled receptor 40 (GPR40) and GPR120. Importantly, EPA ameliorates ACI-induced apoptosis. EPA exerts beneficial effect on ACI-induced inflammation through blocking NLRP3 inflammasome activation by GPR40 and GPR120. Our findings suggest the potential clinical use of EPA in ACI. Copyright 2019 The Author(s). Published by Elsevier Inc. All rights reserved.