This work was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) under the Emmy-Noether program, BL 1591/1-1 - Project ID 411333557. We thank Suong Nguyen, Manuela J&#228;ger, and Maria Stollmann for their assistance in filming the video. We thank Joanna Scanlon for the video voiceover.

The general procedure used in this protocol was approved by the ethics board of the University of Oldenburg. The participant provided written informed consent prior to their participation.
NOTE: The c-grids should only be used on undamaged skin and with participants that do not have an allergy to the adhesive used. It has two sides.&#160;There is black text on the outside. The conductive surfaces of the electrodes are on the inside, and they face the skin of the participant during the recording. Importantly, handle these grids with care. Do not touch the conductive surfaces, do not fold the grids, do not excessively bend them, and avoid pulling on them.
1. Testing
NOTE: If handled with care, c-grids can be reused several times. To ensure optimal functioning, check that all the electrodes are working properly before the next recording. Perform the same procedure for new grids to identify potential problems (e.g., due to problems in the manufacturing process) before the recording starts. There are several options to quickly check for problems (e.g., a broken electrode).
<ol>
	<li>Option 1: Multimeter.
	<ol>
		<li>Set a multimeter to measure resistance.</li>
		<li>Attach one pin of the multimeter to the electrode and the other pin to the corresponding contact on the connector end.</li>
		<li>Check if a low resistance (&#60;10 k&#8486;) can be measured for each electrode.</li>
	</ol>
	</li>
	<li>Option 2: Electrode gel
	<ol>
		<li>Use electrode gel to bridge all the electrodes. Make sure that there are no gaps between the electrodes.</li>
		<li>Attach the grid to the connector of an amplifier. To see a signal, attach the grid to the side with the reference and ground electrodes according to the connector layout that is used.</li>
		<li>Use the impedance check of the amplifier. Check the impedance of the reference electrode and all eight recording electrodes (10 electrodes in total minus the ground and reference electrodes); they must all have a low impedance (&#60;10 k&#8486;). Afterward, wipe off the gel.</li>
	</ol>
	</li>
	<li>Option 3: Water 
	NOTE: Use this option with caution in order to not cause any water damage to the equipment.
	<ol>
		<li>Submerge all the electrodes in a glass of water, but make sure to keep the tail of the grid dry. Alternatively, place the c-grid in a plate filled with water (with the electrodes facing the plate).</li>
		<li>Attach the grid to the connector of the amplifier.</li>
		<li>Use the impedance check of the amplifier. Check the impedance of the reference electrode and all eight recording electrodes (10 electrodes in total minus the ground and reference electrodes); they must all have a low impedance (&#60;10 k&#8486;). Afterward, dry the c-grid with a tissue.</li>
	</ol>
	</li>
</ol>
2. Preparing the participant
NOTE: For high-quality recordings, the participant should have clean and dry hair, should not have used any hair products (e.g., styling products) or skin products, and should not be wearing make-up. If possible, the participants should wash their hair directly before the recording with a mild and neutral shampoo and also wash the areas around the ears. Ask the participants to indicate if any of the preparatory steps are uncomfortable for them.
<ol>
	<li>To prepare the participant, the experimenter needs access to the area behind and around the ear. For participants with longer hair, use hair clips for easier access.</li>
	<li>Place a c-grid around the participant&#39;s ear to see how it fits. Additionally, check if it can be positioned around the ear without touching the ear. Ensure it does not touch the back of the ear or the ear lobe, as this can be uncomfortable after some time. This pre-fitting also gives an indication of the area that will be covered and, thus, needs to be cleaned. 
	NOTE: These grids come in one size and do not fit all ear sizes. For larger ears, cut some of the plastic around the electrodes on the inside of the C with a small pair of scissors. Pay special attention to not cut into the electrodes or the conductive path.</li>
	<li>Apply a small drop of abrasive electrode gel onto a tissue. Use the gel to clean the skin around the participant&#39;s ear with some pressure, but make sure it remains comfortable for the participant. Make sure to generously clean the whole area that will be covered.</li>
	<li>Dip a tissue in some alcohol, and clean the area behind the ear with this tissue.</li>
	<li>Dry the cleaned area with a clean towel.</li>
	<li>For higher levels of comfort, optionally place a small piece of tape on the back side of the ear.</li>
	<li>Repeat all of the above steps (steps 2.1-2.5) for the other ear.</li>
</ol>
3. Preparing and fitting the grids
NOTE: There are different ways of attaching the c-grid using double-sided tape. Presented here are two options: c-shaped stickers (provided by the manufacturer) that cover the whole surface and small circular stickers that are placed individually around the electrodes (e.g., when reusing).
<ol>
	<li>Attach double-sided adhesive stickers (either the c-shaped or individual stickers) around each electrode. Make sure that the stickers do not cover the conductive surface of the electrodes.</li>
	<li>Put small drops (lentil-sized) of electrode gel on each electrode. Avoid using too much gel, as this might spill onto the adhesive material and reduce the adhesion to the skin. Too much gel could also create bridges between the electrodes.</li>
	<li>Remove the cover of the adhesive sticker(s). Reapply the gel in case it was removed during this step. Alternatively, remove the first cover, and apply the gel then; however, this requires a very steady hand so the gel does not spill accidently onto the adhesive.</li>
	<li>Ask the participant to hold their hair away from the ear so that it does not obstruct the fitting. Move any hair out of the way as much as possible so that the stickers touch the skin directly. Depending on the hairline, this is not always possible (e.g., when there is hair directly above the ear).</li>
	<li>Position the grid around the ear, and when in place, press it into the skin. Make sure to not place it too close to the ear, as this may become uncomfortable for the participant. Leave some space (1 mm to 2 mm) between the grid and the back side of the ear. Additionally, ask the participant to press onto the electrodes.</li>
	<li>Repeat all of the above steps (steps 3.1-3.5) for the other ear.</li>
	<li>Remove any hair clips. Carefully place glasses or the strips of facial masks on the ears if necessary.</li>
</ol>
4. Connecting
<ol>
	<li>Connect the connector to the amplifier. During this step, avoid excessively bending or pulling on the c-grid.</li>
	<li>Plug the contacts into the connector. Make sure that the contacts are plugged in on the correct side. Ensure that the exposed contacts on the inside of the c-grid face the contacts in the connector. 
	NOTE: It is important to know the layout of the connector being used (including the position of the ground and the reference electrodes). Depending on the system being used, the layout may differ. To build a connector, visit&#160;https://uol.de/psychologie/abteilungen/ceegrid. With the right connector, c-grids can be connected to any amplifier.</li>
	<li>To hold the amplifier in place, use a headband, for example, to fixate it on the head. 
	&#8203;NOTE: The Oldenburg lab uses an amplifier that is built into a neck speaker called nEEGlace. The nEEGlace makes the setup more comfortable and faster.</li>
</ol>
5. Check the impedance and data
<ol>
	<li>Connect the amplifier to a smartphone (optionally: a laptop) via Bluetooth.</li>
	<li>Check the impedance of the electrodes with the impedance check of the amplifier. The impedance usually improves over time (5 min to 10 min) and does not have to be below 10 k&#8486; for each electrode in the beginning. Do not try to put more gel underneath electrodes with a high impedance.</li>
	<li>Check the EEG signal. Ask the participant to clench their jaws, to blink, and to close their eyes (alpha activity). Observe the corresponding artifacts and alpha activity in the signal. Make sure every electrode provides a good signal. If the resulting EEG signal is poor, remove the grid, wipe off any residual gel around the ear of the participant, and fit a new one.</li>
	<li>Begin recording.</li>
</ol>
6. Removing and cleaning up
<ol>
	<li>After finishing the data recording, disconnect the phone (or laptop) from the amplifier. Detach the grids from the amplifier, and remove the amplifier from the participant. Gently remove the c-grids from the participant. Make sure to neither bend the c-grid too much nor to pull out the hair of the participant. Allow the participants to clean themselves with tissues or a towel.</li>
	<li>Soak the grids in water for some minutes. They can be submerged completely.</li>
	<li>Carefully detach the stickers to avoid damage. Rinse off any remaining gel. Air-dry the grids. Do not rub over the conductive surface of the electrodes.</li>
	<li>Store the c-grids safely in a dark and dry place.</li>
</ol>

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The authors report no conflicts of interest.

Provided here is a protocol for ear-EEG recordings with the c-grids. Following the steps of this protocol ensures high-quality recordings. In the following paragraphs, a comparison is made with cap-EEG, the most critical steps in the protocol along with some best practice recommendations are discussed, and some modifications are discussed.
Comparison of c-grids to cap-EEG and in-ear EEG 
The c-grid allows the unobtrusive recording of brain activity in everyday life settings and is well suited for longer recordings. It has several advantages compared to cap-EEG. First, due to its weight, comfort, and low visibility, it barely restricts participants in their everyday activities1. Second, it can be worn for extended periods of time - more than 11 hours in one study6- without the electrodes falling off1,3,6, since they are sealed by the adhesive stickers. On the downside, the c-grid covers only a fraction of the surface of cap-EEG and, thus, cannot replace cap-EEG for all purposes. However, in cases when a lightweight, unobtrusive, quick-to-setup, minimally restricting solution is necessary (e.g., in the workplace), c-grids can provide relevant neural information.
The comparison of results across participants is potentially more difficult for c-grids compared to cap-EEG. For cap-EEG, often the international 10-20 system is used to facilitate the comparison of results across studies and across participants with different head sizes. In this system, the electrodes are positioned relative to specific anatomical landmarks (i.e., the nasion and inion for front to back and the ears for left to right). In practice, different cap sizes are used to account for different head sizes and thereby approximate the optimal electrode positioning. The c-grid cannot be easily integrated into that system for two reasons. First, these are currently available in one size and, thus, cover more or less space depending on the head size. Second, the shape of the ear influences the positioning of the grids. In general, the two top-most electrodes will be directly above the ear, but depending on the ear shape, they might be tilted more to the front or to the back. We are not aware of any study that has investigated whether these shifts in electrode positions are large enough to be of relevance.
Another approach to measuring&#160;ear-EEG is to place the electrodes inside the ear, for instance, in the outer ear canal or the concha23,24,25. Such an approach offers even lower visibility than the c-grid but leads to recording signals with lower amplitudes due to the small distances between the electrodes26.
Most critical steps 
EEG in general, and especially mobile ear-centered EEG, remains a challenging technology. Therefore, the careful preparation of the participant and placement of the grids is essential to ensure good data quality over time. The preparation starts with the hair and the skin of the participants. The hair and skin around the ear should be washed and dried. In addition to that, the experimenter needs to carefully clean the area around the ear with abrasive gel and alcohol and ensure that the grids are firmly attached with the adhesive stickers. These steps are important and should be performed carefully to ensure good electrode-skin adhesion and a low impedance for longer periods. The skin cleaning especially can make the difference between a successful and an unsuccessful recording.
Even with proper care, however, the impedance for individual electrodes may still be poor directly after the placement of the electrodes. In general, the electrode-skin interface stabilizes over time, and we often observe that the impedance decreases within 5 min to 15 min. If the signal quality remains poor, it is recommended to completely remove the grids, wipe off any residual gel around the participant&#39;s ear, and fit a new one. It is faster to fit a new one as opposed to cleaning and preparing the previously removed grid. It is not recommended to add electrode gel to individual electrodes once the grid is fitted as this can compromise the adhesion strength of the stickers and can even lead to the bridging of the neighboring electrodes.
After the grid has been placed and when the impedance of the electrodes is low, data recording can begin. For longer recordings (&#62;1 h), a brief data quality check should be conducted at the beginning. For instance, a 3 min auditory oddball task is exemplified in this study, which can be conducted and analyzed quickly to ensure a good signal quality.
In some cases, recording with the c-grid might not be possible at all, such as when the grid is too small for the ear (even after cutting) or when the hairline is too close to the ear, meaning that the grid does not stick to the skin. If the grid &#34;hovers&#34; over some hair, researchers cannot expect high-quality data.
Troubleshooting 
Bad impedance and/or signal 
To avoid these problems, it is imperative that the skin is carefully cleaned before fitting. Additionally, one should make sure to test the functionality of each electrode before the fitting. For example, one should check that the grid is plugged in correctly into the connector and that each electrode has firm contact with the skin and then wait for a few minutes until the impedance and the signal improve. To further check the functionality after fitting, the individual electrodes should be pressed, and the resulting signal should be checked. If the corresponding signal of each electrode shows a response, the electrode is functional in principle. If all of the above steps do not help, one should remove the grid, wipe off the residual gel around the participant&#39;s ear, and fit a new one.
Situations with no signal 
Firstly, one should make sure the grid is connected properly to the amplifier, as well as ensuring that the grid connector is not upside down. There will be a signal only if the ground and reference electrodes are connected; whether the ground and reference will be on the left, the right, or on both sides depends on the connector.
Signal getting worse during the recording 
There may be several reasons for this issue that need to be addressed. First, some of the electrodes might have become detached from the skin. This can happen when the adhesive is compromised by the remains of the electrode gel, by hair under the electrodes, or due to interference from the participant (e.g., scratching around the ear or adjusting glasses). Second, there might be problems with the connection between the grid and the amplifier (i.e., the grid might have been pulled out of the amplifier, or its position might have shifted). Finally, the grid might have sustained damage during usage. This can happen if the tail of the c-grid is bent too strongly.
Channels showing identical signals 
In this case, the electrodes are bridged. One should remove the grid, wipe off the residual gel around the participant&#39;s ear, and fit a new one. One should also make sure to only use lentil-sized drops of electrode gel on each electrode to avoid bridging.
Participants reporting that the placement is uncomfortable 
The most common reason for decreased comfort is that the grid is placed too close to the backside of the ear. One should make sure to leave 1 mm to 2 mm between the c-grid and the back side of the ear. A small piece of tape attached behind the ear helps to increase comfort.
Modifications of the method 
The c-grid comes in one size. However, it allows for some flexibility regarding its size. By cutting the plastic of the inner side, the size may be reduced to fit larger ears. One should pay special attention in order to not cut into the electrodes or the conductive paths.
Depending on the amplifier used and the recording scenario, there are different ways to place the amplifier on the body. The fixed length of the tail of the grid and the fact that it points horizontally away from the ear limits the possible locations for placing the connector of the amplifier. Different manufacturers provide adapter cables that connect the grid to a specific amplifier (either mobile or lab-based). Different solutions have been proposed for placing the amplifier; some researchers use a headband3, whereas others integrate it into a basecap27. For shorter experiments, a headband is suitable. For longer experiments, the amplifier can be taped to the clothes6 or body2, stored in custom-made straps, taped to headphones worn around the neck1, or taped to a neck protector commonly used for mountain biking. We have developed a prototype that combines a neck speaker (for presenting auditory stimuli) with a mobile EEG amplifier and connectors to the c-grid (building instructions can be found here:&#160;https://github.com/mgbleichner/nEEGlace). We have used this approach successfully in a recent study (in preparation) in which we recorded ear-EEG for 4 h while participants worked in an office.
Future applications 
The c-grid is a promising tool for long-term recordings in everyday life. For instance, one can use it to investigate sound processing in everyday life1. With long-term recordings, circadian variations in cognition and auditory function can also be investigated28,29. For diagnostic purposes, the grid can be used for the long-term monitoring of epileptic seizures2, sleep staging6, or for measuring attention for hearing devices7,11.
Conclusion 
This protocol comprehensively equips researchers for experimenting with these c-grids in and beyond the lab. If researchers follow this protocol and carefully perform the steps, including the most important ones, such as skin cleaning and fitting the c-grid, they can expect high-quality data for their ear-EEG experiments.

With mobile ear-electroencephalography (EEG), brain activity can be recorded in everyday life, and new insights into neural processing beyond the lab can be gained1. To be suitable for everyday life, a mobile ear-EEG system should be transparent, unobtrusive, easy to use, motion-tolerant, and comfortable to wear even for several hours2. The c-grid (sold under the name cEEGrid), a c-shaped ear-EEG system, aims to meet these requirements to minimize interference with natural behavior. The grid consists of 10 Ag/AgCl electrodes printed on Flexprint material3. Combined with a miniaturized, mobile amplifier and a smartphone for data acquisition4,5, these grids can be used to collect ear-EEG data for more than 8 hours1,6.
Several studies conducted in the lab have shown the potential of c-grids for studying auditory and other cognitive processes. C-grids have been successfully used for auditory attention decoding with accuracies above chance level7,8,9,10,11. Segaert et al.12 used these arrays to quantify language impairment in patients with mild cognitive impairment. Garrett et al.13 showed that these arrays can capture auditory brain potentials originating from the brain stem. Apart from the research focusing on the auditory domain, Knierim et al.14 used the grids to investigate flow experiences (i.e., the sensation of total involvement in a task), as measured by changes in alpha power. Finally, Pacharra et al.15 used these grids for a visual task. All of these lab-based studies showcase the various cognitive processes that can be captured with these grids.
These grids can also be used for EEG recordings beyond the lab, as illustrated by several studies. For example, these arrays have been used to evaluate mental load in a driving simulator16,17 and to study inattentional deafness, the non-perception of critical alarm sounds, in a flight simulator18. The grids are especially promising for long-term recordings, such as the long-term monitoring of epileptic seizures2 and sleep staging6. H&#246;lle et al.1 used these grids to measure auditory attention during an office day for 6 hours. In sum, all of these studies highlight their potential to investigate various brain processes in and outside of the lab.
Every EEG recording requires careful preparation to obtain valid results. This is especially important for mobile applications where more artifacts can be expected than in the lab due to the movement of the participant. To ensure optimal results, specific preparation steps are necessary. We state the critical steps in preparing the grids, preparing the participant for data collection, and fitting and connecting the grids for EEG recordings. We point out potential mistakes and show examples of poor data quality when the attachment is not proper. Finally, representative results of a piano-played&#160;oddball task are shown.

<table><tbody><tr><td>Abrasive gel: Abralyt HiCl</td><td>easycap GmbH, Germany</td><td></td><td></td></tr><tr><td>AFEx app</td><td>University of Oldenburg, Germany</td><td></td><td>for our exemplary data recording, open-source: https://zenodo.org/record/5814670#.Y0AavXZByUk</td></tr><tr><td>Alcohol</td><td>Carl Roth GmbH + Co. KG, Germany</td><td></td><td>70% isopropanol, 30% destilled water</td></tr><tr><td>c-grid: cEEGrid</td><td>TSMI, Oldenzaal, The Netherlands</td><td></td><td></td></tr><tr><td>cEEGrid connector</td><td>University of Oldenburg, Germany</td><td></td><td>costum build</td></tr><tr><td>EEG acquisition app: Smarting</td><td>mBrainTrain, Serbia</td><td></td><td></td></tr><tr><td>Matlab</td><td>The MathWorks, Inc., USA</td><td></td><td>used for data analyses and creating the figures</td></tr><tr><td>Medical tape: Leukosilk</td><td>BSN medical GmbH, Germany</td><td></td><td></td></tr><tr><td>mobile EEG amplifier: Smarting MOBI</td><td>mBrainTrain, Serbia</td><td></td><td></td></tr><tr><td>Multimeter</td><td>PeakTech Pr&amp;uuml;f- und Messtechnik GmbH, Germany</td><td></td><td>optional device to check functionality of electrodes</td></tr><tr><td>nEEGlace</td><td>University of Oldenburg, Germany</td><td></td><td>costumized neckspeaker with integrated EEG amplifier (Smarting, mBrainTrain, Serbia) and cEEGrid connectors</td></tr><tr><td>Paper wipes&amp;nbsp;</td><td>-</td><td></td><td></td></tr><tr><td>Record-a app</td><td>University of Oldenburg, Germany</td><td></td><td>for our exemplary data recording, open-source: https://github.com/NeuropsyOL/Pocketable-Labs</td></tr><tr><td>Smartphone: Google Pixel 3a&amp;nbsp;</td><td>Google LLC, USA</td><td></td><td></td></tr><tr><td>Yahama Digital Piano P-35</td><td>Hamamatsu, Japan</td><td></td><td>for our exemplary data recording</td></tr></tbody></table>

recording brain activity with ear-electroencephalography

The c-grid (ear-electroencephalography, sold under the name cEEGrid) is an unobtrusive and comfortable electrode array that can be used for investigating brain activity after affixing around the ear. The c-grid is suitable for use outside of the laboratory for long durations, even for the whole day. Various cognitive processes can be studied using these grids, as shown by previous research, including research beyond the lab. To record high-quality ear-EEG data, careful preparation is necessary. In this protocol, we explain the steps needed for its successful implementation. First, how to test the functionality of the grid prior to a recording is shown. Second, a description is provided on how to prepare the participant and how to fit the c-grid, which is the most important step for recording high-quality data. Third, an outline is provided on how to connect the grids to an amplifier and how to check the signal quality. In this protocol, we list best practice recommendations and tips that make c-grid recordings successful. If researchers follow this protocol, they are comprehensively equipped for experimenting with the c-grid both in and beyond the lab.

When following this protocol, the impedance of each electrode is usually below 10 k&#937; or approaches this value a few minutes after placing the grid (Figure 1), indicating a good electrode-skin contact. Of note, the impedance may still improve within 2 h after fitting it.
Figure 2 illustrates different unprocessed EEG signals. Figure 2A illustrates how the data looks when no gel is used. A conductive gel is required, and the grid does not work properly without using gel. If too much gel is used, the electrodes might be bridged. The data for this scenario are shown in Figure 2B. Bridged electrodes show exactly the same signal. When the preparation and fitting are carried out carefully, one can expect high-quality data, as displayed in Figure 2C.
Figure 3 illustrates the procedure and data from an exemplary event-related potential (ERP) paradigm (oddball task) with one participant. Figure 3A illustrates the paradigm. Specifically, the experimenter played a predefined sequence of two different notes on the piano (middle C and middle G). Middle C was played frequently (328 times), and middle G was played infrequently (78 times); the participant had to count the infrequent notes. The open-source AFEx app recorded the tone onsets, loudness (RMS), and spectral content (PSD) for all the tones. The Record-A app concurrently recorded the acoustic features and EEG4. In the analyses, infrequent and frequent tones were differentiated based on the power spectral density (PSD; see H&#246;lle et al.19 for details). The EEG data were high-pass filtered at 0.1 Hz and low-pass filtered at 25 Hz. A spatial filter was computed by using generalized eigenvector decomposition, which maximizes the signal of interest20. In Figure 3B,C, the resulting ERP with typical components of auditory processing can be observed, such as the N1 for both tones and the P3 for the infrequent tone that had to be counted. These results are consistent with previous oddball studies both with c-grids1, 3 and with cap-EEG21,22.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/64897/64897fig01.jpg" /> 
Figure 1: Example of good impedance. All values are in kiloohms (k&#937;). <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/64897/64897fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/64897/64897fig02.jpg" /> 
Figure 2: Illustration of unprocessed signals with different qualities. (A) Example of 10 s of data when no electrode gel is used at all. (B) Example of 10 s of data when the electrodes are bridged. (C) Example of 10 s of good data acquired in the lab. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/64897/64897fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/64897/64897fig03.jpg" /> 
Figure 3: Results from an event-related potential (ERP) paradigm (oddball task) with one participant. (A) Overview of the paradigm. The participant listened to a sequence of tones played on a piano and had to count the infrequent one. The smartphone concurrently recorded the EEG and acoustic features (B) ERPs of all the c-grid channels. Abbreviations: REF = reference electrode; DRL = ground electrode. (C) ERP based on the spatial filter displayed in the top-left corner. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/64897/64897fig03large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Recording Brain Activity with Ear-Electroencephalography at JoVE.com

Recording Brain Activity with Ear-Electroencephalography

Presented here is the procedure for using the c-grid (ear-electroencephalography, sold under the name cEEGrid) for recording brain activity in and beyond the lab for extended durations. This protocol describes how to set up these arrays and how to record brain activity using them.

The c-grid (ear-electroencephalography, sold under the name cEEGrid) is an unobtrusive and comfortable electrode array that can be used for investigating ...

recording-brain-activity-with-ear-electroencephalography

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Neuroscience

2.7K Views.  University of Oldenburg. Presented here is the procedure for using the c-grid (ear-electroencephalography, sold under the name cEEGrid) for recording brain activity in and beyond the lab for extended durations. This protocol describes how to set up these arrays and how to record brain activity using them.

Video: Recording Brain Activity with Ear-Electroencephalography

Performing Behavioral Tasks in Subjects with Intracranial Electrodes

Patients having stereo-electroencephalography (SEEG) electrode, subdural grid or depth electrode implants have a multitude of electrodes implanted in different areas of their brain for the localization of their seizure focus and eloquent areas. After implantation, the patient must remain in the hospital until the pathological area of brain is found and possibly resected. During this time, these patients offer a unique opportunity to the research community because any number of behavioral paradigms can be performed to uncover the neural correlates that guide behavior. Here we present a method for recording brain activity from intracranial implants as subjects perform a behavioral task designed to assess decision-making and reward encoding. All electrophysiological data from the intracranial electrodes are recorded during the behavioral task, allowing for the examination of the many brain areas involved in a single function at time scales relevant to behavior. Moreover, and unlike animal studies, human patients can learn a wide variety of behavioral tasks quickly, allowing for the ability to perform more than one task in the same subject or for performing controls. Despite the many advantages of this technique for understanding human brain function, there are also methodological limitations that we discuss, including environmental factors, analgesic effects, time constraints and recordings from diseased tissue. This method may be easily implemented by any institution that performs intracranial assessments; providing the opportunity to directly examine human brain function during behavior.

Patients implanted with intracranial electrodes provide a unique opportunity to record neurological data from multiple areas of the brain while the patient performs behavioral tasks. Here, we present a method of recording from implanted patients that can be reproducible at other institutions with access to this patient population.

Patients having stereo-electroencephalography (SEEG) electrode, subdural grid or depth electrode implants have a multitude of electrodes implanted in ...

Behavior

Recording Human Electrocorticographic (ECoG) Signals for Neuroscientific Research and Real-time Functional Cortical Mapping

Neuroimaging studies of human cognitive, sensory, and motor processes are usually based on noninvasive techniques such as electroencephalography (EEG), magnetoencephalography or functional magnetic-resonance imaging. These techniques have either inherently low temporal or low spatial resolution, and suffer from low signal-to-noise ratio and/or poor high-frequency sensitivity. Thus, they are suboptimal for exploring the short-lived spatio-temporal dynamics of many of the underlying brain processes. In contrast, the invasive technique of electrocorticography (ECoG) provides brain signals that have an exceptionally high signal-to-noise ratio, less susceptibility to artifacts than EEG, and a high spatial and temporal resolution (i.e., &lt;1 cm/&lt;1 millisecond, respectively). ECoG involves measurement of electrical brain signals using electrodes that are implanted subdurally on the surface of the brain. Recent studies have shown that ECoG amplitudes in certain frequency bands carry substantial information about task-related activity, such as motor execution and planning1, auditory processing2 and visual-spatial attention3. Most of this information is captured in the high gamma range (around 70-110 Hz). Thus, gamma activity has been proposed as a robust and general indicator of local cortical function1-5. ECoG can also reveal functional connectivity and resolve finer task-related spatial-temporal dynamics, thereby advancing our understanding of large-scale cortical processes. It has especially proven useful for advancing brain-computer interfacing (BCI) technology for decoding a user's intentions to enhance or improve communication6 and control7. Nevertheless, human ECoG data are often hard to obtain because of the risks and limitations of the invasive procedures involved, and the need to record within the constraints of clinical settings. Still, clinical monitoring to localize epileptic foci offers a unique and valuable opportunity to collect human ECoG data. We describe our methods for collecting recording ECoG, and demonstrate how to use these signals for important real-time applications such as clinical mapping and brain-computer interfacing. Our example uses the BCI2000 software platform8,9 and the SIGFRIED10 method, an application for real-time mapping of brain functions. This procedure yields information that clinicians can subsequently use to guide the complex and laborious process of functional mapping by electrical stimulation.
Prerequisites and Planning:
Patients with drug-resistant partial epilepsy may be candidates for resective surgery of an epileptic focus to minimize the frequency of seizures. Prior to resection, the patients undergo monitoring using subdural electrodes for two purposes: first, to localize the epileptic focus, and second, to identify nearby critical brain areas (i.e., eloquent cortex) where resection could result in long-term functional deficits. To implant electrodes, a craniotomy is performed to open the skull. Then, electrode grids and/or strips are placed on the cortex, usually beneath the dura. A typical grid has a set of 8 x 8 platinum-iridium electrodes of 4 mm diameter (2.3 mm exposed surface) embedded in silicon with an inter-electrode distance of 1cm. A strip typically contains 4 or 6 such electrodes in a single line. The locations for these grids/strips are planned by a team of neurologists and neurosurgeons, and are based on previous EEG monitoring, on a structural MRI of the patient's brain, and on relevant factors of the patient's history. Continuous recording over a period of 5-12 days serves to localize epileptic foci, and electrical stimulation via the implanted electrodes allows clinicians to map eloquent cortex. At the end of the monitoring period, explantation of the electrodes and therapeutic resection are performed together in one procedure.
In addition to its primary clinical purpose, invasive monitoring also provides a unique opportunity to acquire human ECoG data for neuroscientific research. The decision to include a prospective patient in the research is based on the planned location of their electrodes, on the patient's performance scores on neuropsychological assessments, and on their informed consent, which is predicated on their understanding that participation in research is optional and is not related to their treatment. As with all research involving human subjects, the research protocol must be approved by the hospital's institutional review board. The decision to perform individual experimental tasks is made day-by-day, and is contingent on the patient's endurance and willingness to participate. Some or all of the experiments may be prevented by problems with the clinical state of the patient, such as post-operative facial swelling, temporary aphasia, frequent seizures, post-ictal fatigue and confusion, and more general pain or discomfort.
At the Epilepsy Monitoring Unit at Albany Medical Center in Albany, New York, clinical monitoring is implemented around the clock using a 192-channel Nihon-Kohden Neurofax monitoring system. Research recordings are made in collaboration with the Wadsworth Center of the New York State Department of Health in Albany. Signals from the ECoG electrodes are fed simultaneously to the research and the clinical systems via splitter connectors. To ensure that the clinical and research systems do not interfere with each other, the two systems typically use separate grounds. In fact, an epidural strip of electrodes is sometimes implanted to provide a ground for the clinical system. Whether research or clinical recording system, the grounding electrode is chosen to be distant from the predicted epileptic focus and from cortical areas of interest for the research. Our research system consists of eight synchronized 16-channel g.USBamp amplifier/digitizer units (g.tec, Graz, Austria). These were chosen because they are safety-rated and FDA-approved for invasive recordings, they have a very low noise-floor in the high-frequency range in which the signals of interest are found, and they come with an SDK that allows them to be integrated with custom-written research software. In order to capture the high-gamma signal accurately, we acquire signals at 1200Hz sampling rate-considerably higher than that of the typical EEG experiment or that of many clinical monitoring systems. A built-in low-pass filter automatically prevents aliasing of signals higher than the digitizer can capture. The patient's eye gaze is tracked using a monitor with a built-in Tobii T-60 eye-tracking system (Tobii Tech., Stockholm, Sweden). Additional accessories such as joystick, bluetooth Wiimote (Nintendo Co.), data-glove (5th Dimension Technologies), keyboard, microphone, headphones, or video camera are connected depending on the requirements of the particular experiment.
Data collection, stimulus presentation, synchronization with the different input/output accessories, and real-time analysis and visualization are accomplished using our BCI2000 software8,9. BCI2000 is a freely available general-purpose software system for real-time biosignal data acquisition, processing and feedback. It includes an array of pre-built modules that can be flexibly configured for many different purposes, and that can be extended by researchers' own code in C++, MATLAB or Python. BCI2000 consists of four modules that communicate with each other via a network-capable protocol: a Source module that handles the acquisition of brain signals from one of 19 different hardware systems from different manufacturers; a Signal Processing module that extracts relevant ECoG features and translates them into output signals; an Application module that delivers stimuli and feedback to the subject; and the Operator module that provides a graphical interface to the investigator.
A number of different experiments may be conducted with any given patient. The priority of experiments will be determined by the location of the particular patient's electrodes. However, we usually begin our experimentation using the SIGFRIED (SIGnal modeling For Realtime Identification and Event Detection) mapping method, which detects and displays significant task-related activity in real time. The resulting functional map allows us to further tailor subsequent experimental protocols and may also prove as a useful starting point for traditional mapping by electrocortical stimulation (ECS).
Although ECS mapping remains the gold standard for predicting the clinical outcome of resection, the process of ECS mapping is time consuming and also has other problems, such as after-discharges or seizures. Thus, a passive functional mapping technique may prove valuable in providing an initial estimate of the locus of eloquent cortex, which may then be confirmed and refined by ECS. The results from our passive SIGFRIED mapping technique have been shown to exhibit substantial concurrence with the results derived using ECS mapping10.
The protocol described in this paper establishes a general methodology for gathering human ECoG data, before proceeding to illustrate how experiments can be initiated using the BCI2000 software platform. Finally, as a specific example, we describe how to perform passive functional mapping using the BCI2000-based SIGFRIED system.

Recording Human Electrocorticographic ECoG Signals for Neuroscientific Research and Real-time Functional Cortical Mapping

We present a method for collecting electrocorticographic signals for research purposes from humans who are undergoing invasive epilepsy monitoring. We show how to use the BCI2000 software platform for data collection, signal processing and stimulus presentation. Specifically, we demonstrate SIGFRIED, a BCI2000-based tool for real-time functional brain mapping.

Neuroimaging studies of human cognitive, sensory, and motor processes are usually based on noninvasive techniques such as electroencephalography (EEG), ...

High-density Electroencephalographic Acquisition in a Rodent Model Using Low-cost and Open-source Resources

Advanced electroencephalographic analysis techniques requiring high spatial resolution, including electrical source imaging and measures of network connectivity, are applicable to an expanding variety of questions in neuroscience. Performing these kinds of analyses in a rodent model requires higher electrode density than traditional screw electrodes can accomplish. While higher-density electroencephalographic montages for rodents exist, they are of limited availability to most researchers, are not robust enough for repeated experiments over an extended period of time, or are limited to use in anesthetized rodents.1-3 A proposed low-cost method for constructing a durable, high-count, transcranial electrode array, consisting of bilaterally implantable headpieces is investigated as a means to perform advanced electroencephalogram analyses in mice or rats.

Procedures for headpiece fabrication and surgical implantation necessary to produce high signal to noise, low-impedance electroencephalographic and electromyographic signals are presented. While the methodology is useful in both rats and mice, this manuscript focuses on the more challenging implementation for the smaller mouse skull. Freely moving mice are only tethered to cables via a common adapter during recording. One version of this electrode system that includes 26 electroencephalographic channels and 4 electromyographic channels is described below.

Instructions for the low-cost construction and surgical implantation of a chronic transcranial high-density electroencephalographic montage into mice are provided. Signal recording, extraction, and processing techniques are also described.

Advanced electroencephalographic analysis techniques requiring high spatial resolution, including electrical source imaging and measures of network ...

Concurrent Recording of Co-localized Electroencephalography and Local Field Potential in Rodent

Although electroencephalography (EEG) is widely used as a non-invasive technique for recording neural activities of the brain, our understanding of the neurogenesis of EEG is still very limited. Local field potentials (LFPs) recorded via a multi-laminar microelectrode can provide a more detailed account of simultaneous neural activity across different cortical layers in the neocortex, but the technique is invasive. Combining EEG and LFP measurements in a pre-clinical model can greatly enhance understanding of the neural mechanisms involved in the generation of EEG signals, and facilitate the derivation of a more realistic and biologically accurate mathematical model of EEG. A simple procedure for acquiring concurrent and co-localized EEG and multi-laminar LFP signals in the anesthetized rodent is presented here. We also investigated whether EEG signals were significantly affected by a burr hole drilled in the skull for the insertion of a microelectrode. Our results suggest that the burr hole has a negligible impact on EEG recordings.

This protocol describes a simple method for concurrent recording of co-localized electroencephalography (EEG) and multi-laminar local field potential in an anesthetized rat. A burr hole drilled in the skull for the insertion of a microelectrode is shown to produce negligible distortion of the EEG signal.

Although electroencephalography (EEG) is widely used as a non-invasive technique for recording neural activities of the brain, our understanding of the ...

Simultaneous Recordings of Cortical Local Field Potentials, Electrocardiogram, Electromyogram, and Breathing Rhythm from a Freely Moving Rat

Monitoring the physiological dynamics of the brain and peripheral tissues is necessary for addressing a number of questions about how the brain controls body functions and internal organ rhythms when animals are exposed to emotional challenges and changes in their living environments. In general experiments, signals from different organs, such as the brain and the heart, are recorded by independent recording systems that require multiple recording devices and different procedures for processing the data files. This study describes a new method that can simultaneously monitor electrical biosignals, including tens of local field potentials in multiple brain regions, electrocardiograms that represent the cardiac rhythm, electromyograms that represent awake/sleep-related muscle contraction, and breathing signals, in a freely moving rat. The recording configuration of this method is based on a conventional micro-drive array for cortical local field potential recordings in which tens of electrodes are accommodated, and the signals obtained from these electrodes are integrated into a single electrical board mounted on the animal's head. Here, this recording system was improved so that signals from the peripheral organs are also transferred to an electrical interface board. In a single surgery, electrodes are first separately implanted into the appropriate body parts and the target brain areas. The open ends of all of these electrodes are then soldered to individual channels of the electrical board above the animal's head so that all of the signals can be integrated into the single electrical board. Connecting this board to a recording device allows for the collection of all of the signals into a single device, which reduces experimental costs and simplifies data processing, because all data can be handled in the same data file. This technique will aid the understanding of the neurophysiological correlates of the associations between central and peripheral organs.

This study introduces a method for the simultaneous recording of local field potentials in the brain, electrocardiograms, electromyograms, and breathing signals of a freely moving rat. This technique, which reduces experimental costs and simplifies data analysis, will contribute to the understanding of the interactions between the brain and peripheral organs.

Monitoring the physiological dynamics of the brain and peripheral tissues is necessary for addressing a number of questions about how the brain controls ...

Data Acquisition and Analysis In Brainstem Evoked Response Audiometry In Mice

Brainstem evoked response audiometry (BERA) is of central relevance in the clinical neurophysiology. As other evoked potential (EP) techniques, such as visually evoked potentials (VEPs) or somatosensory evoked potentials (SEPs), the auditory evoked potentials (AEPs) are triggered by the repetitive presentation of identical stimuli, the electroencephalographic (EEG) response of which is subsequently averaged resulting in distinct positive (p) and negative (n) deflections. In humans, both the amplitude and the latency of individual peaks can be used to characterize alterations in synchronization and conduction velocity in the underlying neuronal circuitries. Importantly, AEPs are also applied in basic and preclinical science to identify and characterize the auditory function in pharmacological and genetic animal models. Even more, animal models in combination with pharmacological testing are utilized to investigate for potential benefits in the treatment of sensorineural hearing loss (e.g., age- or noise-induced hearing deficits). Here we provide a detailed and integrative description of how to record auditory brainstem-evoked responses (ABRs) in mice using click and tone-burst application. A specific focus of this protocol is on pre-experimental animal housing, anesthesia, ABR recording, ABR filtering processes, automated wavelet-based amplitude growth function analysis, and latency detection.

Brainstem evoked response audiometry is an important tool in clinical neurophysiology. Nowadays, brainstem evoked response audiometry is also applied in the basic science and preclinical studies involving both pharmacological and genetic animal models. Here we provide a detailed description of how auditory brainstem responses can be successfully recorded and analyzed in mice.

Brainstem evoked response audiometry (BERA) is of central relevance in the clinical neurophysiology. As other evoked potential (EP) techniques, such as ...

Stimulating Mouse Brain with a Low-Cost EEG and Millimeter-Sized Coil

Low-Cost Electroencephalographic Recording System Combined with a Millimeter-Sized Coil to Transcranially Stimulate the Mouse Brain In Vivo

A low-cost electroencephalographic (EEG) recording system is proposed here to drive transcranial magnetic stimulation (TMS) of the mouse brain in vivo, utilizing a millimeter-sized coil. Using conventional screw electrodes combined with a custom-made, flexible, multielectrode array substrate, multi-site recording can be carried out from the mouse brain. In addition, we explain how a millimeter-sized coil is produced using low-cost equipment usually found in laboratories. Practical procedures for fabricating the flexible multielectrode array substrate and the surgical implantation technique for screw electrodes are also presented, which are necessary to produce low-noise EEG signals. Although the methodology is useful for recording from the brain of any small animal, the present report focuses on electrode implementation in an anesthetized mouse skull. Furthermore, this method can be easily extended to an awake small animal that is connected with tethered cables via a common adapter and fixed with a TMS device to the head during recording.The present version of the EEG-TMS system, which can include a maximum of 32 EEG channels (a device with 16 channels is presented as an example with fewer channels) and one TMS channel device, is described. Additionally, typical results obtained by the application of the EEG-TMS system to anesthetized mice are briefly reported.

Author Spotlight: Low-Cost Electroencephalographic Recording System Combined with a Millimeter-Sized Coil to Transcranially Stimulate the Mouse Brain In Vivo  

A low-cost electroencephalographic recording system combined with a millimeter-sized coil is proposed to drive transcranial magnetic stimulation of the mouse brain in vivo. Using conventional screw electrodes with a custom-made, flexible, multielectrode array substrate, multi-site recording can be carried out from the mouse brain in response to transcranial magnetic stimulation.

A low-cost electroencephalographic (EEG) recording system is proposed here to drive transcranial magnetic stimulation (TMS) of the mouse brain in vivo, ...

Electro-encephalography (EEG)

EEG is a non-invasive technique that can measure brain activity. The neural activity generates electrical signals that are recorded by EEG electrodes placed on the scalp. When an individual is engaged in performing a cognitive task, brain activity changes and these changes can be recorded on the EEG graph. Therefore, it is a powerful tool for cognitive scientist aiming to better understand the neural correlates associated with different aspects of cognition, which will ultimately help them devise improved treatments for patients with cognitive deficits.
Here, JoVE presents a brief overview of EEG and its applications in cognitive research. First, we discuss where and how EEG signals are generated. Then, we explain the use of EEG in studying cognition along with a detailed step-by-step protocol to perform an EEG experiment. Lastly, the video reviews some specific cognitive experiments that use EEG in combination with other techniques such as functional Magnetic Resonance imaging (fMRI) or transcranial direct current stimulation (tDCS).

EEG: EEG Signal Origin, Even Related Potentials &amp; Cognition

EEG is a non-invasive technique that can measure brain activity.  The neural activity generates electrical signals that are recorded by EEG electrodes ...

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Recording Cortical and Subcortical Neuronal Activity Using Electrode Systems

Source: Trenado, C., et al. <a href="https://app-jove-com.remotexs.ntu.edu.sg/v/53466">Combined Invasive Subcortical and Non-invasive Surface Neurophysiological Recordings for the Assessment of Cognitive and Emotional Functions in Humans.</a> J. Vis. Exp. (2016).
This video demonstrates the process of recording cortical and subcortical neuronal activity using electroencephalography (EEG) and deep brain stimulation (DBS) electrodes during an attention-based visual stimulation test.

All procedures involving human participants have been performed in compliance with the institutional, national, and international guidelines for human ...

JoVE Encyclopedia of Experiments

Neurophysiology

Stimulation and Modulation Techniques

Concurrent Electroencephalographic and Local Field Potential Recordings in an Anesthetized Rat

Source: Kang, S., et al. <a href="https://app-jove-com.remotexs.ntu.edu.sg/v/56447">Concurrent Recording of Co-localized Electroencephalography and Local Field Potential in Rodent.</a> J. Vis. Exp. (2017).
This video demonstrates the procedure for concurrent recordings of co-localized electroencephalography (EEG) and local field potentials (LFPs) in an anesthetized rat. The process involves preparing the rat and attaching stimulating electrodes to its whisker pads. An EEG spider electrode is placed on the rat&#39;s skull for surface EEG measurements. At the same time, a multi-channel microelectrode is inserted into a specific brain region through a burr hole in the skull to record neural activity from a localized brain area.

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