Generating a Mouse Model of Traumatic Brain Injury Using a Lateral Fluid Percussion Device

프로토콜

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Animal care

1. Use 5- to 6-week-old male C57BL/6J mice (20-25 g of weight) in this experiment.
2. Maintain mice on a 12 h/12 h light/dark cycle, and make sure they receive food and water ad libitum. Provide the same amounts of food and water to both the sham and TBI groups throughout the study.
3. Make all efforts to minimize the animal pain and discomfort.

2. Induction of traumatic brain injury

1. Inject Ketamine (80-100 mg/kg)/Xylazine (10 mg/kg) IP for anesthesia. Test the depth of anesthesia using an eye reflex or pain reflex. Use artificial tears or lubricant eye ointment to keep the eyes from drying.
2. After anesthesia, put the mouse in a prone position. Use a temperature-controlled heating pad to maintain the temperature at 37 °C during the surgery and for 30 min after TBI.
3. Shave the hair of the incision area.
4. Disinfect the scalp with 70% ethanol using three alternating scrubs, then incise the scalp in a sagittal plane.
5. Use forceps to retract the incision on both sides and separate the periosteum slightly.
6. Use a marker to draw a circle (3 mm diameter) on the right parietal area of the skull, 2 mm away from the midline.
7. Drill the skull with an electric drill. Ensure that this step is operated carefully to protect the dura from being damaged.
8. Remove the bone flap and expose a small bone window (3 mm in diameter).
9. Place a plastic injury cannula (internal diameter = 2.5 mm, length = 8 mm) over the craniotomy and cement the cannula to the skull using a dental acrylic.
10. Fill the cannula with sterile 0.9% NaCl (normal saline) using a syringe (5 mL) to ensure that there are no bubbles in the cannula.
11. Turn on the oscilloscope and amplifier and ensure that the high-pressure tube of the LFPI device is free of air bubbles. Test the device by delivering about 10 pulses until it gives a steady signal. Adjust the angle of the pendulum starting position to reach a pulse intensity of about 2.0 atm.
12. Connect the injury cannula to the LFPI device. Induce brain injury by pulling the trigger and releasing the pendulum. Then, obtain a pulse and transmit it to the dura through the entire closed fluid-filled tubing system.
13. Operate the mice in the sham group with the same surgical procedure. Do not perform the LFPI.

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자료

Name	Company	Catalog Number	Comments
Heating pad	Shanghai SAFE Biotech Co.	TR-200	Heating pad
Injector	The First Affiliated Hospital, School of Medicine, Zhejiang University		Injector
LFPI device	Virginia Commonwealth University	FP302	LFPI device
Micro cranial drill	RWD Life Science	78061	Micro cranial drill
Povidone Iodine	The First Affiliated Hospital, School of Medicine, Zhejiang University		Povidone Iodine