Permanent middle cerebral artery occlusion

Adult male mice (aged 10 to 14 weeks, weighing 24 to 27 g) were exposed to permanent middle cerebral artery occlusion (pMCAO) with an intraluminal filament²⁷. Mice were anesthetized by intraperitoneal injection of ketamine (80 mg/kg) and xylazine (10 mg/kg). In anesthetized mice, the right side of the common carotid artery was exposed and isolated. The middle cerebral artery (MCA) was occluded by inserting a 6–0 surgical monofilament nylon suture with a heat-rounded tip into the internal carotid artery, which was advanced further until it closed the origin of the MCA. During the experiments, MCA blood flow was monitored using a single fiber optic probe (MoorVMS-LDF, Moor Instruments Ltd, UK) adhered onto the skull surface of the core area supplied by the MCA (5 mm lateral and 2 mm posterior from the bregma) before and after clamping the right MCA. Mice that showed a decrease in MCA blood flow of less than 70–80% of the initial value were excluded. Rectal temperature was maintained at 37 ± 0.5 °C by means of a heating blanket and heating lamp throughout the surgery. Sham-operated control mice received the same surgical procedure without inserting a filament.