Background and Purpose-Time of ischemia onset is the most critical factor for patient selection for available drug treatment strategies. The purpose of this study was to evaluate the abilities of the absolute longitudinal rotating frame (T 1 ) and transverse (T 2 ) MR relaxation times to estimate the onset time of ischemia in rats. Methods-Permanent middle cerebral artery occlusion in rats was used to induce focal cerebral ischemia and animals were imaged with multiparametric MRI at several time points up to 7 hours postischemia. Ischemic parenchyma was defined as tissue with apparent diffusion coefficient of water Ͻ70% from that in the contralateral nonischemic brain. Results-The difference in the absolute T 1 and T 2 between ischemic and contralateral nonischemic striatum increased linearly within the first 6 hours of middle cerebral artery occlusion. The slopes for T 1 and T 2 fits for both tissue types were similar; however, the time offsets were significantly longer for both MR parameters in the cortex than in the striatum. Conclusions-T 1 and T 2 MRI provide estimates for the onset time of cerebral ischemia requiring regional calibration curves from ischemic brain. Assuming that patients with suspected ischemic stroke are scanned by MRI within this timeframe, these MRI techniques may constitute unbiased tools for stroke onset time evaluation potentially aiding the decision-making for drug treatment strategies. (Stroke. 2010;41:2335-2340.)Key Words: acute stroke Ⅲ animal models Ⅲ brain imaging Ⅲ brain ischemia Ⅲ MRI C ollapse of adenosine 5Ј-triphosphate due to acute ischemia is associated with subtle changes in both ionic and water homeostasis in the brain. Altered homeostasis in the brain is probed by MR techniques to follow ionic and water signals during the evolution of ischemia. 1,2 Diffusion and T 2 MRI form the cornerstones for imaging diagnosis of stroke as supplemented by hemodynamic sensitive perfusion MRI techniques. 3 Combined use of these MRI techniques with T 2 * MRI allows accurate diagnosis of acute stroke as well as separation of ischemic and hemorrhagic stroke.A major challenge for imaging of acute stroke remains to provide an accurate estimate for the duration of ischemia. The duration of ischemia is one of the key factors determining the choice of patient management at hospital admission, including application of currently available drug treatment strategies. Thrombolysis must be initiated within 3 to 4.5 hours of onset of symptoms to render beneficial effect on patient outcome, yet with acceptable adverse effects. 4,5 Similarly, other treatment methods such as hypothermia provide a beneficial outcome when applied within a time window of 3 to 6 hours after the onset. 6 Several neuroprotective drugs have shown an effect in preclinical settings only when administered within a narrow time window from stroke onset. 7 It is well established by electrophysiological and tissue ion analysis techniques that brain sodium increases in a linear fashion after ischemia. 8 Based on this fact, it has be...