This study demonstrates a surgical procedure for injecting growth factors and amyloid-beta peptides into the rat brain to model Alzheimer's disease. The methodology includes precise drilling and cannula implantation to facilitate targeted injections.
Begin with an immobilized, anesthetized rat with its skull exposed.
Locate the bregma, mark three reference points, and carefully drill the holes. Clean the area.
Through one of the holes, inject a growth factor into the anterodorsal thalamic nucleus near the hippocampus.
Secure two screws in two of the holes, insert a guide cannula into the third hole, and fix them with dental cement. Cover the guide cannula with a dummy cannula.
The next day, insert the internal cannula into the guide cannula and connect it to the injection system.
Regularly inject amyloid-beta peptides and aluminum trichloride into the brain's lateral ventricle to facilitate their diffusion in the hippocampus.
Later, remove the cannula implantation system. Disinfect the area, fill the holes with bone cement, and suture the incision.
Over time, the amyloid-beta peptides aggregate into toxic plaques.
The injected growth factor and aluminum trichloride also enhance plaque formation around the hippocampal neurons, which are crucial for memory.
This leads to neuronal death, causing memory impairment similar to Alzheimer's disease.
On an anesthetized rat, begin by snipping the fur on the vertex of the head with surgical scissors and disinfect with iodophor.
Next, make an incision on the head skin along the median longitudinal calvaria with surgical bistouries and scissors.
Separate the subcutaneous tissue and fascia. Wipe the skull calvarium with 0.3% hydrogen peroxide, and mark the bregma with a marker pen. Next, mark three points: the anterodorsal thalamic nucleus for injecting RHTGF-β1 and fixing one screw, the lateral ventricle or LV area for injecting Aβ25-35 and AICI3, and lastly the second screw fixing place.
Gently, drill three 1-millimeter diameter holes with a flexible bone drill at the three marked points on the skull. Stop the bleeding and clean the skull surface repeatedly with sterile dry cotton.
Next, insert a needle linked to the microinjection pump to the brain at 4.2 millimeters depth, and gently inject 1 microliter RHTGF-β1 into the ad area. Keep the microinjection two minutes after injection, then slowly pull it out.
Fix the two screws into the skull, designated in the previous marked points with a small screwdriver. Following screw placement, assemble the cannula implantation system by first inserting the dummy cannula into the guide cannula after disinfection with 75% alcohol immersion for 24 hours. Then, insert the stainless steel tubing guide cannula to the brain at 4.6 millimeters into LV area through the skull hole, with the help of a cannula holder on the rat's stereotaxic apparatus.
Next, mix the denture base material with denture base water, at a ratio of 1.5 grams per 1 milliliters. Put the paste to cover the guide cannula plastic pedestal and two screws for immobilizing the guide cannula. Cover the whole skin incision to avoid skin infection.
The next day, draw out the dummy cannula and insert the internal cannula into the guide cannula. Screw the fixing screw to immobilize the internal cannula.
Set the polyethylene pipe that links the microinjection pump to the internal cannula, and regulate the injection speed to 1 microliter per minute. Microinject the Aβ25-35 to the LV. Wait five minutes after finishing the injection, and gently draw out the internal cannula. Then, insert the dummy cannula again into the guide cannula.
On day 15, post-surgery, dismantle the cannula implantation system by removing the denture base material solid with surgical scissors and forceps, and disinfecting the wound with iodophor.
Finally, fill in the hole of the skull with bone cement and suture the skin with a simple interrupted suture method.
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