Brain Mapping Using a Graphene Electrode Array

The cortical map is a set of local patches representing response properties to sensory motor stimuli in the cerebral cortex. It can discover the spatial formation of neural networks which can offer a prediction for perception and cognition. Hence, cortical maps are used to study neural responses from external stimuli and the processing of sensory motor information To create cortical maps are invasive and non-invasive methods.

Using intracortical electrodes is one of the most common invasive methods used for mapping the cerebral cortex. As it can damage the brain for high resolution, it prevents further measurements from being made. Alternative brain mapping tools such as EEG, PET, MEG, fMRI, are non-invasive methods that allow for whole brain mapping and repeated sampling.

However, several disadvantages are low spatial temporal resolution, temporal lag, errors due to unspecified modulatory inputs and prohibitive costs. Optical techniques such as calcium imaging and optogenetic fMRI provide large scale brain mapping but are not clinically beneficial due to indicator toxicity and low spatial temporal resolution. The Graphene electrode array exhibits long-term biocompatibility and mechanical flexibility which provides stable recordings of the convoluted brain.

Recently, our group has developed a graphene electrode array that provides high resolution recordings for neural signals on the cortical surface. This protocol uses a graphene electrode array to record the SEPs from an SD rat's forepaw, forelimb, hind paw, hind limb, trunk, and whisker to create a cortical map. Administer an intraperitoneal injection of ketamine and xylazine cocktail using a dosage of 90 milligram per kilogram for ketamine and 10 milligram per kilogram for xylazine.

To maintain a desired depth of anesthesia throughout the surgery, inject a 45 milligram per kilogram ketamine, and five milligram per kilogram xylazine cocktail if the rat shows signs of awakening. Pinch the toe to confirm the depth of anesthesia. If the rat trembles, wait for additional minutes until it shows no response.

Shave the fur on the rat's head from the space between the eyes to the back of the ears using a trimmer. Then apply an ophthalmic ointment onto the eyes. Using ear bars and stereotaxic adaptors, fix the rat's head onto the stereotaxic apparatus.

Make sure that the rat's head is fixed properly. Sterilize a shaved area with povidone-soaked cotton swap. Scrub the shaved area with alcohol.

Repeat the sterilization process three times. Then inject 0.1 milliliters of 2%lidocaine into the scalp to induce local anesthesia. Creates a two to three-centimeter midline incision and laterally pull apart the scalp To expose the skull.

Clamp the scalp to expose the skull with mosquito forceps. Start by removing the periosteum by scratching the surface of the skull with forceps. In a straight line towards the lambda, locate the sternum magna by tearing the muscle behind the occipital bone on the posterior edge.

Use a microscope to get a close view of the cisterna magna. Incise the cisterna magna gently. After tearing the cisterna magna, cerebral spinal fluids flow out.

Drain the cerebral spinal fluid with the coiled sterile gauze to sink down the brain. Mark where the graphene electrode will be placed based on the predefined stereotaxic coordinates according to the position of the bregma. The somatosensory cortex is located three millimeters in the anterior posterior axis and six millimeters in the right lateral direction from the bregma of the right hemisphere skull.

Drill the area marked according to the stereotaxic coordinate. Remove the skull with the bone rongeur. Stick a cotton swab into a 26 garish needle.

Bend the needle to 90 degrees. To remove the dura mater, create a hole using a bent needle. Lift the dura mater upward, insert forceps into that hole and proceed to cut the dura mater with scissors or tear it with forceps.

Attach the sterile gauze wet with saline on the somatosensory cortex to prevent the cortex from drying out. Detach the graphene multielectrode array without causing any damage by applying saline solution. Remove the outer covering of the reference and ground wires from the connector.

To set up the graphene multi electrode array for mapping, connect the head stage with the connector. Connect the graphene electrode array with the connector. Plug the interface cable into the recording controller.

Connect the graphene multielectrode array and connector complex to an interface cable. Then secure the graphene multielectrode array complex onto the stereotaxic arm. Place the graphene electrode array onto the somatosensory cortex based on the predefined stereotaxic coordinates.

Place the reference wire in the tissue behind the occipital bone. Then connect the ground wire to the grounded optical table. Open the recording software and set the sampling rate at 30 kilohertz and click the OK button.

Set the 60 hertz notch filter to remove the noise from power. Bend the whisker with a fine stick. Record neural signals through the data acquisition system for the desired time.

These signals are neural signals obtained through stimuli for whiskers. Other body parts also stimulate the same way to record SEPs. Open the MATLAB code named read_Intan_RHS2000 file.

Click the Run button. Choose the recording file and click that file and then wait for the file to be read. Enter the command plot to create a 2D line plot.

Select the representative peak signals that were in response to the stimulus, and calculate the amplitudes of the SEPs by measuring the maximum and minimum values. Obtain the topographic map by coloring the grade with the different shade of color according to the SEP's amplitude. The graphene electrode array is on the left.

There are through holes of the substrate between all electrodes. These holes help the electrodes maintain strong contact with the cortex. On the right, the image shows the electrode array on the cortex.

On the left, the ratio of each location-dependent neural response caused by stimulations to different parts of the rat's body is displayed in the shape of a rat. Each body part is represented with a different color. This configures a somatosensory cortex map.

On the right, the following figure shows a stimuli specific responses gained through the multichannel graphene electrodes that were placed on the surface of the cortex. Each colored box represents the responses that different body parts had on the 30 channels. The figure above represents the LFPs detected on the surface of the somatosensory cortex by using graphene electrode arrays.

The color bar shows that a larger amplitude is represented by a darker shade of color and each color represents a different body part. The figure flex the amplitude based on the shades in the color bar. This homunculus of of the rodent was created using the results that were just explained.

The larger the amplitude a certain body parts had, the larger it would be illustrated on the homonculus. The figure below represents a separate data of LFPs collected from each body part. It represents the varying amplitudes of LFPs detected based on the color bar.

There are several precautions for this procedure. When removing the cerebral spinal fluid, the experimenter should be careful not to touch the brainstem or a spinal cord. This step takes practice.

The rodents whisker is developed enough to sense a deflection direction, stimulus intensity and location of the whisker that was stimulated. That is why the whiskers should be stimulated in a constant direction and intensity for this protocol. Other body parts should also be stimulated consistently.

The limitation of this protocol is that signals evoked in deep brain structures can't be recorded as a graphene electrode array is mounted on the cortical surface. Thus, the experimenter can't identify how the columnar network is hierarchically organized concerning the neural responses. Nonetheless, this protocol can be further applied if the experimenter changes the multiple areas where the electrode arrays are placed.

For instance, if the experimenter places the electrode on the auditory or visual cortex to extract auditory and visual information, they can acquire the auditory or visual maps. Also, this protocol can be extended to the chronic implantation of the graphene multielectrode array.