Adenosine 5’ triphosphate (ATP) works as an extracellular signaling molecule in the brain, and is involved in regulation of neuronal activity and blood flow. However, the spatiotemporal dynamics of extracellular ATP are not fully understood due to lack of appropriate techniques, and their effects on cellular activity in brain disorders remain elusive. To explore the roles of ATP signaling, we developed a fluorescent ATP sensor, which was designed for detection of extracellular ATP. We applied the ATP sensor to the cerebral cortex of living mice, and succeeded in visualizing extracellular ATP dynamics in vivo. Using the ATP imaging technique, we visualized a wave of extracellular ATP release propagating across the cortex during pathological neuronal activity associated with brain ischemia. We compared the dynamics of the wave-like ATP release with that of vascular diameter changes during the neuronal activity, and found that the elevation of extracellular ATP concentration is accompanied by contractile response of pial arteries. These results suggest that the wave-like ATP release is involved in the regulation of vascular tone in brain ischemia. Thus, extracellular ATP imaging will contribute to elucidation of pathophysiological mechanisms and to development of therapies for brain disorders.