Mouse model of transient focal cerebral ischemia

Male mice (8 to 12 weeks, 23 to 28 g) were anesthetized with isoflurane (3% for induction and 1.5% for maintenance) in mixed O₂ and N₂O. Focal cerebral ischemia was induced by introducing a 7-0 monofilament suture (Doccol Corporation) in the left common carotid artery, advanced to the origin of the middle cerebral artery for 1 hour as described previously (19, 49). Body temperature was measured with a rectal thermometer and maintained at 37.0 ± 0.5°C by a temperature-controlled heating pad during the ischemic period. Only those mice with CBF reduced to less than 25% of baseline were considered to be successfully MCA occluded (MCAo). Reperfusion was performed by removing the suture after 1-hour occlusion. Animals that did not show reperfusion of CBF over 60% of baseline were excluded. Regional CBF was measured using a laser speckle imager (Perimed PeriCam PSI HR) at 15 min before MCAo surgery, 15 min after occlusion, and 15 min after the onset of reperfusion. Two identical elliptical regions of interest were selected. The relative CBF was calculated as the ratio of the ipsilateral and contralateral hemispheres, which was then normalized to the mean value of pre-surgery CBF baseline level for each animal. Animals were allowed 24- or 48-hour reperfusion before being sacrificed according to the design of the experiment.