We developed an improved method for suspending and fixing rats with rat clothes in an acupuncture experiment. This protocol aims to provide detailed descriptions of its procedural steps, including important precautions, offering a reference for researchers conducting fundamental experiments in the field of acupuncture and moxibustion. In previously reported studies, researchers have often used anesthesia fixation, high platform fixation, binding fixation, or various other self-made fixators while performing acupuncture or electroacupuncture operations on rats.
Given the side effects of anesthesia on the body and the real clinical conditions, fixation in our weakened stage offer the better alternative. However, in the non-anesthetized stage, the mentioned fixations also possess limitations, such as restricted acupoint exposure, requirement of multiple labels, nonadjustable fixator sites, or forced posture. This method utilizes the rat's preference for burrowing to create rat jackets that fit their body shape.
It offers several advantages over other techniques. It is cost-effective, user-friendly, manpower and space-saving, accommodates rats of varying weights, and can also be used for needle operation on most acupoints. In subsequent studies, we will focus on investigating the impact of the restraints and suspensions of rat clothing on physiological indicators and behavioral aspects related to emotional cognition in rats.
This will enable us to cognitively assess its potential influence on research and refine the fixation method. Wear a pair of gloves before proceeding to fit and secure the rat in the custom-made rat jacket. Then, using one hand, hold the rat jacket's middle and back portion on the table.
Using the other hand, gently hold the rat by its tail to pick it up from the feeding cage and place it into the jacket. As the rat moves forward into the jacket and its head reaches the headgear point, using both hands, carefully pull out its left and right upper limbs from the holes in the jacket. Securely wrap the tying ropes from the opening end of the jacket around the rat's trunk, and tie them tightly at the opposite end to adequately restrain its upper limbs.
Tie the rope loose enough to prevent hypoxia or local limb ischemia, and tight enough to prevent the rat's escape. In a similar manner, secure its lower limbs. Before beginning the procedure of electropuncture on the rat, dress the rat in the custom-made rat jacket, then disinfect the local skin of acupoints using alcohol swabs.
After locating the acupoint accurately, insert the sterile disposable acupuncture needle into the spot at the appropriate depth. Gently suspend the animal from the horizontal iron rod with the help of the S-shaped hooks attached to the rat jacket. Wrap the electrode wire around the horizontal iron rod before securely attaching the small clip at the end of the electrode wire to the needle handle.
Then, after switching on the power, select the appropriate frequency, current, and time. Once the parameters are adjusted, commence the electroacupuncture treatment process. Using the S-shaped hooks, suspend the rats in the same group together on the iron rod for simultaneous electroacupuncture.
When the treatment is over, remove the acupuncture needle along with the clamp from the animal. Then, carefully take the needle out of the clamp and put it in the box. After removing the animal from the suspension hooks, loosen the straps, lift the head cover of the rat's jacket, and position the rat above the cage.
The rat will then be free to release itself from the jacket and enter the cage independently. While performing a 15 minute electroacupuncture experiment on 12 rats, only one rat exhibited intense struggle during the needle retention period, resulting in needle drop. While other rats maintained proper needle fixation throughout the treatment.
When compared to the other fixation methods reported in the literature, the demonstrated method exhibited the combined benefits of stability and secure fixation, efficiency in terms of time and number of personnel required, affordability, as well as relative comfort for the rats. Additionally, in terms of the exposed acupoint range, the specially designed rat jackets in this protocol met the requirements for exposing most acupoints on the body.
