Intracranial implantation

Each rat was anesthetized by intraperitoneal injection of a mixture of xylazine (80 mg/kg) and ketamine (10 mg/kg) and affixed in a stereotactic headframe (Model 900, David Kopf Instruments, Tujunga, California, USA). A 20-mm incision was made in the skin overlying the skull, exposing the coronal suture and bregma. A bur hole was made 3.5 mm to the right of bregma. The tumor cells were infused at a depth of 4.5 mm below the surface of the brain after the syringe (World Precision Instruments, Sarasota, Florida, USA) was advanced 5 mm to create a 0.5-mm pocket. The cell suspension was infused over 3 minutes using a UMP3-1 UltraMicroPump microinjector (World Precision Instruments, Sarasota, Florida, USA) set to a volume of 10 μL with an infusion rate of 3.33 μL/minute. The needle was withdrawn 2 minutes after the injection to minimize the backflow of the cell suspension. Bone wax was used to cover the bur hole. The skin incision was closed with interrupted sutures.