Transcranial Electrical Stimulation of the Motor Cortex in an Awake Rat

Begin with an anesthetized rat with screws embedded in its skull.

The rat has a chest-implanted counter electrode with a subcutaneously tunneled cable secured around one of the skull-embedded screws.

Solder the cable to a connector.

Glue a transcranial electrical brain stimulation or tES electrode socket onto the skull above the motor cortex.

Seal the assembly with glue, then secure it with dental cement.

Reposition the skin and allow the rat to recover.

Fill the socket with saline.

Screw the tES electrode cap containing the stimulation electrode.

Connect a stimulation device to the connectors.

Place the rat in a cage that allows free movement.

Initiate anodal stimulation to generate an electric field in the motor cortex, enabling voltage-gated ion channel opening, positive ion influx, and neuron depolarization.

This enhances synaptic activity, triggering ATP release, which activates microglia, crucial for immune responses.

Neuronal and microglial activation highlights the neurotherapeutic potential of tES.

Begin by scraping off the periosteum, the connective tissue on the skull, to the sides with the scalpel, and thoroughly wipe off with cotton swabs. Fixate the connective tissue at the four corners of the cut with bulldog clamps, and let them hang laterally to keep the surgery field open.

Next, apply 0.9% saline to clean the bone surface and tissue with cotton swabs. Then clean the bone surface with 3% hydrogen peroxide. Use a cotton swab to remove any residuals of the periosteum as they become visible. As fixation screws will improve setup adherence, choose a drill bit fitting the screw size. Place two burr holes on two different bone plates by pre-drilling with a hand drill, and then by slight vertical pressure with the bone drill.

In case of an implanted counter electrode, as shown in this video, burr a third hole located in the right posterior parietal bone for future fixation of the tunneled cable. Place the plastic screws in the burr holes and screw until the first friction is felt. Then perform three additional 180-degree screw turns.

Use forceps to check for stability of the screw and add 1 more turn, if not tight enough. Turn on the soldering iron and preheat for approximately 5 minutes. Wind the cable exiting the tissue tunnel occipitally around the right parietal screw and then cut it, leaving approximately 1 centimeter of cable behind the winding.

Carefully strip the insulation at the end of the cable with a scalpel, then fix the winded cable to the screw and bone with cyanoacrylic glue. Next, apply a small amount of the lead-free tin solder to the connector and to the bare wires of the counter electrode cable, and connect both by briefly pressing both pre-sorted parts together, while touching the soldering tip until the tin solder melts.

Remove the soldering tip immediately to avoid excessive metal heating of the cable with subsequent tissue damage. For stimulation of the motor cortex, use a 4-millimeter diameter socket. Pick up the custom made tES electrode socket with bent serrated tip forceps, and apply a thin layer of cyanoacrylic glue to the bottom rim of the socket.

The inner medial border of the socket should end directly at the sagittal suture, and the caudal inner border should end at the height of bregma. Then press the socket briefly onto the bone until the glue hardens. And use a light to ensure that the bone within the area of the socket does not have any reflective glue on it.

After the socket is in place and the future stimulation area is free of glue, seal the lateral border of the socket to the neighboring tissue with a small drop of cyanoacrylic glue to avoid a fluid bridge that could lead to shunting of current at this location. Then cover all screws with cyanoacrylic glue.

Next, mix the two component dental acrylic cement in a small silicon tube or glass. As soon as it becomes viscous, apply it with a dental spatula to seal the remaining borders of the socket to the bone. Finally, cover the whole skull screws, counter electrode cable, and up to one-third of the socket with dental acrylic cement. After all of the bone is covered and the cement has hardened, remove the bulldog clamps and reposition the skin.

Begin by filling the tES electrode socket half with 0.9% saline and remove air bubbles. Before cathodal tDCS sessions, check chlorination. And if needed, re-chlorinate the silver-silver chloride electrode. Before anodal tDCS sessions, remove possible excess silver chloride deposits from previous stimulations with sandpaper to allow for good conductivity during stimulation. Then screw in the tES electrode cap.

Connect the cables to the two connectors on the head. Then, place the rodent into the experimental cage with the cables connected to a swivel above the cage that allows for free movement. Turn on the stimulator and adjust the stimulation parameters. Finally, after the end of the stimulation, disconnect the cables, unscrew the electrode cap on the head, and clean and dry the socket with a cotton swab.