Application of in vivo ESR spectroscopy to measurement of cerebrovascular ROS generation in stroke

This study used an in vivo ESR spectroscopy/spin probe technique to measure directly the generation of reactive oxygen species (ROS) in the brain after cerebral ischemia-reperfusion. Transient middle cerebral artery occlusion (MCAO) was induced in rats by inserting a nylon thread into the internal carotid artery for 1 h. The in vivo generation of ROS and its location in the brain were analyzed from the enhanced ESR signal decay data of three intra-arterially injected spin probes with different membrane permeabilities. The ESR signal decay of the probe with intermediate permeability was significantly enhanced 30 min after reperfusion following MCAO, whereas no enhancement was observed with the other probes or in the control group. The enhanced in vivo signal decay was significantly suppressed by superoxide dismutase (SOD). Brain damage was barely discernible until 3 h of reperfusion, and was clearly suppressed with the probe of intermediate permeability. The antioxidant MCI-186 completely suppressed the enhanced in vivo signal decay after transient MCAO. These results clearly demonstrate that ROS are generated at the interface of the cerebrovascular cell membrane when reperfusion follows MCAO in rats, and that the ROS generated during the initial stages of transient MCAO cause brain injury.