Stroke is a leading cause of mortality and chronic neurologic disability. Yet, the successful treatment remains limited. In this study, we investigated the efficacy and the mechanism of a novel treatment, microRNA-210 (miR-210) inhibition, in protecting acute ischemic brain injury in adult mice. Focal cerebral ischemia was induced by middle cerebral artery occlusion (MCAO) in adult male C57BL/6 mice. MiR-210-LNA (miR-210 inhibitor) or the negative control was administered via intracerebroventricular injection 24 h prior or 4 h after MCAO. Cerebral infarction volume and behavioral deficits were determined 48 h after MCAO. The expression of inflammation-related genes and infiltration/activation of various immune cells in the brain were assessed by RT-qPCR, flow cytometry, and immunohistochemistry. Acute ischemic stroke significantly increased miR-210 levels in the brain, which was abolished by miR-210-LNA administered prior to MCAO. Pre- and post-MCAO treatments with miR-210-LNA significantly decreased cerebral infarction and ameliorated behavioral deficits induced by MCAO. Long-term behavioral recovery was also improved by miR-210-LNA post-treatment. At the same time, inhibition of miR-210 significantly reduced the expression of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) and chemokines (CCL2 and CCL3), but had no significant effect on anti-inflammatory factors (TGF-β and IL-10). In addition, MCAO-induced macrophage infiltration and microglial activation in the brain were inhibited by the miR-210-LNA treatment. In summary, inhibition of miR-210 suppresses pro-inflammatory response and reduces brain damage in the acute phase of ischemic stroke, providing new insight in molecular basis of a novel therapeutic strategy of miR-210 inhibition in the treatment of acute ischemic stroke.