PTEN (phosphatase and tensin homologue deleted on chromosome 10) is a dual-specificity lipid and protein phosphatase. The loss of PTEN was originally discovered in numerous human cancers. PTEN inhibition by bisperoxovanadium (bpV) reduces neurological damage after ischemic brain injury. The purpose of this study was to identify the optimal neuroprotective dose of bpV when administrated after focal ischemia/reperfusion (I/R) injury in rats. Focal I/R injury was induced using the middle cerebral artery occlusion method. bpV at doses of 0.25, 0.50 and 1.0 mg/kg were injected intraperitoneally just after reperfusion, with saline serving as a vehicle control. A maximal reduction in brain injury was observed with 1.0 mg/kg bpV. This dose of bpV also significantly blocked apoptosis in the penumbral cortex of rats. This beneficial effect was associated with the increasing levels of Akt phosphorylation in the penumbral cortex. These results demonstrate that the pharmacological inhibition of PTEN protects against I/R injury in a dose-dependent manner and the protective effect might be induced through upregulation of the phosphoinositide-3 kinase/Akt pro-survival pathway, suggesting a new therapeutic strategy to combat ischemic brain injury.