FeCl3-induced common carotid artery thrombosis

Arterial thrombosis was induced in the common carotid artery (CCA) using FeCl₃.^22,23 Briefly, anesthesia was induced with 5% isoflurane in a mixture of 70% N₂O and 30% O₂ and maintained with 2% isoflurane. Body temperature was continuously monitored and maintained at 37.0±0.5℃ using a homeothermic blanket control unit and heating pad with a rectal probe (Harvard Apparatus, Holliston, MA, USA) during the operative procedures. Under a surgical microscope, a midline incision was made at the cervical area in the supine position, and the left CCA was exposed and isolated. Blood flow was measured in the midportion of the CCA using an ultrasonic Doppler flow probe (MA0.7PSB; Transonic Instruments, Ithaca, NY, USA) and an iWorx IX-304T data acquisition system (iWorx Systems, Inc., Dover, NH, USA). After measurement of baseline blood flow for 5 min, arterial thrombosis was induced using FeCl₃ (F2877; Sigma-Aldrich Inc., St. Louis, MO, USA). A piece of filter paper (F2877; Sigma-Aldrich, 0.5×0.7 mm) was soaked in 2 µL of 50% FeCl₃ and placed on the midportion of the CCA for 5 min. The filter paper was then removed, the CCA was washed with normal saline, and blood flow was checked again. Subsequently, the probe was carefully positioned in the upper intact CCA, and blood flow was monitored for 10 min. The success of arterial occlusion with thrombus formation was defined as a decrease in blood flow to zero. The blood flow data were analyzed using iWorx LabScribe software (version 4.01).²³ After blood flow recording for 10 min, the CCA was washed and excised.