For ischemic injury, published protocols39 for middle cerebral artery occlusion (MCAO) were modified as follows for P5 mice. Pups were anesthetized and maintained with 2–3% isoflurane in O2 on a rectal thermometer feedback heat pad at 37 °C. The animal’s heads were shaved, and cleaned with chlorhexidine, then sterile saline. Mice were injected subcutaneously with antibiotic (25 mg/kg cefazolin) and analgesic (0.1 mg/kg buprenorphine). A 4 mm horizontal incision and a 4 mm vertical incision were made to create a skin flap over the temporalis muscle. The temporalis muscle was also incised in a similar manner so that the skull was exposed. A micro drill was used to create a 2 mm diameter hole directly over the middle cerebral artery, the meninges were removed, and the middle cerebral artery was cauterized. The brain surface was rinsed with saline, and the temporalis muscle was folded back in to place and the skin was sealed with surgical glue. Animals were placed in a warm cage to recover from anesthesia until awake and ambulatory. Animals were left for 24 hours, at which time
