This work was supported by grants from ANR (Programme Blanc Neurosciences), FRC (F&#233;d&#233;ration pour la Recherche sur le Cerveau), City of Paris (Programme Emergence), INSERM and La Piti&#233; Salp&#234;tri&#232;re Hospital (Translational research contract) to N.R., from Neurop&#244;le de Recherche Francilien (NeRF) to E.D., from the Universit&#233; Pierre et Marie Curie UPMC (Programme Convergence) and from the Institut du Cerveau et e la Moelle epiniere (Paris) to G.H.&#160;

NOTE: The protocol here described follows the guidelines of the French &#8220;Comit&#233; Consultatif National d&#8217;Ethique&#8221; and is declared as a collection activity by INSERM.
1. Resection of Human Epileptic Tissue
<ol>
	<li>Patients
	<ol>
		<li>Obtain brain tissue from patients suffering from intractable epilepsy. Obtain a written informed consent before surgery.</li>
		<li>For all operated epileptic patients perform an extensive presurgical workup including neurological examination, neuropsychological assessment, surface EEG, and neuroimaging (MRI and sometimes 18FDG-PET scan; Figure 1A), followed by post-surgical histological examination of resected tissue (Figure 1B). NOTE: Surgery is indicated when the diverse explorations localize similarly the seizure onset zone and when the benefit of its removal exceeds the surgical risks.</li>
	</ol>
	</li>
	<li>Surgery
	<ol>
		<li>Maintain antiepileptic drugs pre-operatively. Perform surgery under general anesthesia: induction with inhaled sevoflurane (6% delivered ratio and 100% inhaled fraction of&#160;O2), intra venous sufentanyl (0.3 &#181;g/kg) and antracurium (0.5 mg/kg), followed by maintenance with intra venous sufentanyl (0.5 &#181;g/kg/hr) and inhaled sevoflurane (1 MAC).</li>
		<li>Use a neuronavigation system to allow a precise localization of the lesioned cortex (Figure 1C). After skin incision, perform a burr hole in the bone&#160;followed by a craniotomy with a high spill drill. Gently elevate the bone flap, and open the dura with scissors (Figure 1D).</li>
		<li>Use intra-operative ultra sound to facilitate the identification of the abnormal cortex, and microsurgical instruments and an ultrasonic aspirator under the operating microscope.</li>
		<li>Coagulate and cut the pio-arachnoidal plane according to the&#160;sulcal limits. Use subpial dissection to release the cortex from the pia around the lesioned area.</li>
		<li>Cut the white matter under the abnormal cortex to perform one piece &#8220;en bloc&#8221; resection, sparing as much as possible the vasculature. Avoid traumatic manipulation of the cortex.</li>
		<li>Cut a 1 cm thick slice out of the resected tissue along a direction orthogonal to the pial surface, including the sulcus bottom and transitional cortex.</li>
	</ol>
	</li>
</ol>
2. Artificial Cerebrospinal Fluid (ACSF) for Slice Preparation, Incubation and Recording
<ol>
	<li>ACSF for slice preparation
	<ol>
		<li>Prepare 1 L of sucrose-based ACSF7,8, containing (in mM): 250 sucrose, 3 KCl, 25 NaHCO3, 10 D-Glucose, 1 CaCl2, 10 MgCl2; dissolve these salts in deionized water and oxygenate the solution with carbogen for at least 10 min.</li>
		<li>Put ~700 ml of sucrose ACSF at -80 &#176;C for 50-60 min (or -150 &#176;C for 15-20 min). Keep the rest of the solution under oxygenation in ice. NOTE: The ACSF for slice preparation can be stored at 4 &#176;C; in this case, add CaCl2 and MgCl2 the day of the experiment, before cooling it.</li>
	</ol>
	</li>
	<li>ACSF for slice incubation and recording
	<ol>
		<li>Prepare at least 3 L of recording ACSF7,8, containing (in mM): 124 NaCl, 3 KCl, 26 NaHCO3, 10 D-Glucose, 1.6 CaCl2, 1.3 MgCl2; dissolve these salts in deionized water and oxygenate the solution with carbogen. Before adding MgCl2, keep some ACSF to perform recordings in 0 Mg2+ ACSF. NOTE: The incubation/recording ACSF can be stored at 4 &#176;C; in this case, add CaCl2 and MgCl2 the day of the experiment.</li>
	</ol>
	</li>
</ol>
3. Equipment for Slice Incubation and Recording
<ol>
	<li>Interface chamber for slice incubation
	<ol>
		<li>Use a brain slice chamber to maintain living slices ex vivo in interface mode. NOTE: The chamber is composed of a lower section, which contains the heating and the sensor elements and a ceramic bubbler and is filled with distilled water, and an upper part, where the slices rest on a sheet of lens tissue in the flat area in the center of the chamber (Figure 2A-B).</li>
		<li>Use two separate fine PTFE tubes, through which the preoxygenated ACSF enters the chamber. NOTE: The tubes spiral in the heated water and reach the upper part of the chamber; then the solution exits by means of capillary action with lens tissue, that conveys the solution into exit wells.</li>
		<li>Connect the PTFE tubes of the interface chamber to a peristaltic pump, to allow continuous perfusion of the slices with the preoxygenated medium.</li>
		<li>Maintain the high oxygen tension by oxygenating with carbogen (95% O2 and 5% CO2) through the ceramic bubbler. NOTE: This warmed and moistened gas mixture reaches the slices in the upper part of the chamber through appropriate holes.</li>
		<li>Maintain the temperature in the upper part of the chamber by connecting the chamber to a temperature controller, through a special double-ended cable, having a connector for the chamber heating element and an external temperature sensor at one end. Place the sensor near the slices to check the temperature throughout the incubation (Figure 2C).</li>
	</ol>
	</li>
	<li>Multi-electrode array recordings
	<ol>
		<li>Use planar multi-electrode arrays with 120 titanium nitride electrodes (30 &#181;m diameter) insulated with silicon nitride and internal reference electrode, arranged in a 12 x 12 matrix with 200 &#181;m center-to-center spacing. NOTE: Other configurations with different electrode diameter and spacing are possible. In particular, the MEA chip can be widened to sample larger slices of human tissues.</li>
		<li>Acquire electrical signals at 10 kHz using a MEA2100-120 system (pre- and filter amplifier with integrated data acquisition and analog-digital converter, 16 bit data resolution, input voltage range and bandwidth adjustable via software) through a dedicated software (MC_Rack). NOTE: The MEA2100 headstage is endowed of a metal plate for fixing of heatable perfusion element through magnetic holders, to continuously perfuse the slices throughout the recording session.</li>
		<li>Hold the tissue down on the MEA by a small homemade platinum anchor, which ensures a good electrical coupling between the slice and the electrodes.</li>
	</ol>
	</li>
</ol>
4. Preparation of Interface Chamber and Tools for Dissection and Slicing
<ol>
	<li>Interface chamber
	<ol>
		<li>Fill the lower section of the interface chamber with distilled water. Ensure that the level of water completely immerses the heating element and is 2 to 4 mm below the junction between the lower and upper section of the chamber. Check this level daily before switching on the heating, in order to avoid damage of the heating element.</li>
		<li>Connect the chamber to a carbogen source, endowed with a secondary flow regulator for fine adjustments of gas pressure.</li>
		<li>Cut a sheet of lens tissue in order to fit the flat area of the slice chamber, and position it close to input solution tubes, to let any bubble escape from the input line before reaching the slices.</li>
		<li>Cut two pieces of stranded cotton thread as long as the piece of lens tissue, and position them along the main sides of the flat area of the chamber. NOTE: This helps to slightly raise the perfusion solution level in the chamber.</li>
		<li>Set the flow rate of the peristaltic pump at 1 ml/min and activate the pump.</li>
		<li>Start oxygenation of the water in the bottom part of the chamber.</li>
		<li>Set the temperature of the temperature controller at 37 &#176;C and turn it on. Cover the upper part of the interface chamber with a piece of parafilm and wait at least 30 min for the temperature to increase and stabilize.</li>
	</ol>
	</li>
	<li>Tools for dissection and slicing
	<ol>
		<li>Prepare a dissection kit consisting of two fine forceps, a spatula, a small spoon and a blade; two glass petri dishes, two brushes as well as cyanoacrylate glue.</li>
		<li>Set the parameters of the vibratome: thickness, 400 &#181;m; frequency, 70 - 90 Hz; amplitude, 0.9 - 1 mm, speed: 0.1 mm/sec.</li>
	</ol>
	</li>
</ol>
5. Human Cortical Slice Preparation
<ol>
	<li>Remove the sucrose-based ACSF from -80 &#176;C freezer and mix it in order to obtain a sort of slush. Oxygenate it with carbogen, put ~250 ml in a glass bottle and keep it in a Dewar bottle filled with ice, while going to the surgery room of the hospital to get the tissue. In the meantime, keep the rest at -20 &#176;C. 
	NOTE: During the surgery, the neurosurgeon dissects a small block of cortical tissue. Immediately place the sample in ice-cold sucrose ACSF for transfer to the laboratory.</li>
	<li>Keep the time needed for the transfer from the hospital to the laboratory to a maximum of 30 min. In the laboratory, take the sucrose-based oxygenated ACSF out of -20 &#176;C freezer, mix it to have a slush, fill a petri dish and the vibratome buffer tray and start oxygenating both with carbogen. Carefully take out the piece of human tissue from the bottle with a spoon and put it in the petri dish (Figure 2D).</li>
	<li>Using the forceps, carefully remove blood clots, vessels and meninges to avoid resistance during slicing. Remove the meninges, consisting in pia-matter, starting from the sides of the tissue block, paying attention not to damage the cortical surface. With a blade, cut a side of the tissue to obtain a flat surface, which will be glued on the vibratome specimen plate. NOTE: The meninges, consisting in pia-matter, are removed starting from the sides of the tissue block, paying attention not to damage the cortical surface.</li>
	<li>If the block of tissue is bigger than the MEA chamber, cut a piece of appropriate dimensions before gluing it on the specimen plate (Figure 2E).</li>
	<li>Glue the tissue in order to obtain transverse cortical slices, put it into the buffer tray and cut 400 &#181;m thick slices at low speed (0.1 mm/sec) (Figure 2F).</li>
	<li>Cut pieces of lens tissue with slice dimensions; with a spatula and a brush, transfer the slices on these lens tissues and incubate them in the interface chamber at 37 &#176;C for at least 1 hr before starting recordings (Figure 2G-I). Continuously store the slices in the same conditions until use. NOTE: Placing slices on lens paper is an important step to facilitate both the perfusion of bottom side of slices and the removal of slices from the interface chamber before recording.
	<ol>
		<li>While transferring the slices on the lens tissue, manage them very carefully, avoiding to touch the cortical layer area with the brush.</li>
		<li>Prepare the slices one at a time and immediately transfer each of them to the interface chamber, in order to ensure proper oxygen supply and temperature; slice the block of resected tissue as quickly as possible.</li>
	</ol>
	</li>
</ol>
6. Preparation of MEA Setup for Recordings
<ol>
	<li>Plug the perfusion system to a peristaltic pump and the MEA system to a computer. Oxygenate the recording ACSF. Place an empty MEA chip inside the amplifier.</li>
	<li>Set the temperature controller for heating cannula element to reach 37 &#176;C in the MEA chamber and start the perfusion at 5-6 ml/min. NOTE: It is usually necessary to set the temperature 3-4 &#176;C higher than the one desired, depending on the room temperature and the flow rate.
	<ol>
		<li>Check the temperature in the MEA chamber with a fine thermometer, placing it the closest possible to the center, without touching the electrode area.</li>
	</ol>
	</li>
	<li>Start recording software and check that all the MEA electrodes are well connected and that there is no noise due to perfusion.</li>
	<li>Start MEA_Monitor to check the functioning of the camera table.</li>
</ol>
7. Transfer of a Slice From Interface Chamber to MEA Chip for Recording
<ol>
	<li>Pour some warmed recording ACSF in a glass Petri dish; with a forceps, take a slice from the interface chamber, by grabbing it through the lens tissue, and put it in the petri dish. Carefully detach the slice from the tissue. If the level of the solution in the interface chamber is carefully adjusted during incubation time, the slice will detach from the lens tissue only by submerging it in the solution; otherwise, use a small brush to facilitate the detachment.</li>
	<li>Fill a MEA chip with some ACSF and with a spatula, transfer the slice into it. With a plastic Pasteur pipette, gently remove the solution to make the slice adhere on the electrode area and keep it in place with a platinum anchor. Add 1 ml of recording ACSF, place the MEA chip into the amplifier and start perfusion at 5-6 ml/min (Figure 2J-L).</li>
	<li>Check the slice position on the MEA recording area by using MEA_Monitor, adjust it if necessary in order to record from the cortical layers and take a picture. Start the recording.</li>
</ol>

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Sponsorship by Multichannel Systems was provided for the production of the video and open access publication.

Pharmacoresistant epilepsy is a rare condition, which can be explored in human tissue in vitro. This allows studying epileptic human cortex, which displays specific defects that are only partially reproduced in animal models. The method here described allows preparing and recording human postoperative tissues ex vivo with preserved cellular viability and networks so that they spontaneously produce epileptic activities. Retaining similar activities than those recorded in vivo is crucial to study mechanisms of genesis of pathological activities. Further, such methods allow exploring human tissues and avoid non perfect animal models of the disease. However, studying human tissue requires synchronization between neurosurgeons and the experimental laboratory. Tissue transportation requires specific care not to be traumatic. In addition, both the quantity of specimen and tissue is limited. Finally, access to proper control tissues is the main concern. With this preparation, the post operative tissues spontaneously produce interictal-like discharges in normal ACSF7,8. Ictal-like events can also be elicited in modified, proconvulsive ACSF so that the mechanisms of seizure initiation and transition from the interictal state to seizures can be investigated.
Human tissue can be kept viable for up to 10 hr in interface conditions. Slices were considered viable when multi-unit activity or field potentials were observed spontaneously and when such activities were evoked by increasing excitability through extracellular potassium increases and/or magnesium decrease. Although variability in activity occurs, likely reflecting differences in pathologies and cortical areas, we explored epileptogenic characteristics of a tissue only in healthy slices showing spontaneous interictal and evoked ictal discharges. In order to preserve tissue vitality and activity, an interface chamber is used to store the slices at 36-37 &#176;C for recovery before recording with the MEA system. Indeed, several groups have clearly demonstrated the advantages of interface based storage system compared to standard beaker storage and the importance of temperature for the preservation of network activity such as spontaneous sharp wave-ripples or cholinergic-induced oscillations9,10. Interface storage of slices has been previously used to record epileptic activity from human hippocampal and subicular slices1,11. With the current MEA technique, after the recovery period from slicing in interface conditions, the network activity is recorded in submerged conditions, in presence of high flow rate (5-6 ml/min) at 37 &#176;C, with the MEA system. The reduced diameter (1.8 cm) of MEA chip, which delimits a small volume chamber (&#60;1.5 ml), together with the increased flow rate, enhances oxygen supply of the slice, which has been demonstrated to be a critical factor for spontaneous and pharmacologically-induced network activities9,10. Further, the reduced amount of circulating ACSF facilitates pharmacological testing.
The slicing procedure is however a traumatism for the tissues12. Both neuronal architecture and chloride homeostasis appear to be perturbed at the tissue surface (50 &#181;m). The origin of the activities recorded by MEA chips, which sample the tissue mostly superficially without deep penetration, may arise from traumatized areas. However, our data show that the extracellular field potentials detected locally are recorded in most MEA electrodes and previous work show that they are integrated over a 100 to 200 &#181;m distance from the recording site13, suggesting that seizures recorded in our preparation are unlikely to be produced by traumatized areas. Further, in studies performed with tungsten electrodes enabling deep penetration, the epileptic activities recorded in human tissues are similar to the ones observed in epileptic patients1,7,8.
Another limit of ex vivo tissue recording is the disruption of the connections between the different brain areas, thus restricting dynamic neuromodulations. This may explain why in such tissue no ictal-like event is spontaneously recorded, but requires to be triggered by ionic manipulation or pharmacological stimulation enhancing excitability. Accordingly, in this protocol, seizure-like events are induced by combining a change in extracellular K+ from 3 to 6 mM and a reduction of external Mg2+ from 1.3 mM to Mg2+-free ACSF, in order to increase tissue excitability and remove Mg2+-dependent NMDA receptor block. Indeed, it has previously been demonstrated that epileptiform activity induced in human neocortical and hippocampal slices using Mg2+-free ACSF resembles the electrographic seizures recorded in vivo14. Besides, it has been shown that epileptiform discharges obtained in temporal lobe slices become resistant to clinically used anticonvulsants after prolonged exposure to Mg2+-free ACSF15,16, thus providing a model for investigating pharmacoresistant seizure-like events in vitro.
MEAs allow recording of both, field potentials and multi-unit activities consisting in neuronal action potentials ex vivo. Thus, MEAs are a powerful electrophysiological tool compared to EEGs, which explore field potentials generated by synchronous activities of neuronal ensembles in vivo, but do not give access to single neuron behaviors17. Although more recent microelectrodes can record in vivo multi-unit activities consisting in neuronal action potentials, they are invasive, so their use is mostly restricted to research purpose during intracranial recordings. In particular, MEAs recordings represent a technique of choice to study the spatio-temporal patterns of epileptic events, the mechanisms controlling seizure onset and propagation and the action of classic and new antiepileptic drugs. Remarkably, to unravel the cell types and the signaling basis of epileptic discharges, spike sorting techniques and pharmacological testing should be combined with MEA techniques. Although MEAs can give access to individual spikes, they do not provide information about synaptic and biophysical properties. In the future, other techniques should be coupled to MEA recordings in order to better sample cellular behavior, network activities and synaptic signaling. For instance, fluorescence imaging to unravel neurons or glial cells behavior, as well as ion dynamics, may be combined with MEAs recordings. Further post hoc histological analysis can also reveal specific alterations of cell types, proteins or receptors so that the location of the epileptic activities can be correlated with specific rearrangements of the nervous structure. The MEAs system can also be embedded in a patch-clamp set-up to correlate single cells or conductances with population activities. In the future, optogenetic tools could also be used, provided that human slices can be cultured in the long-term, as performed for organotypic slices, so that transfection or infection of specific cell types can be performed.

Epilepsy is a chronic disorder in which epileptic seizures, which are intermittent patterned discharges lasting several seconds to tens of seconds on electroencephalographic (EEG) recordings associated with clinical manifestations, interrupt an interictal state, characterized by the presence of synchronous neuronal discharges lasting tens of milliseconds and called interictal events1. It affects approximately 1% world&#8217;s population and although seizures are controlled in the majority of the patients, approximately one third of people with epilepsy do not show adequate response to antiepileptic drugs2. In this condition, called pharmacoresistant epilepsy and whose mechanisms still have to be clearly identified, the surgical resection of the specific part of the brain identified as the seizure-onset zone remains the only alternative treatment giving a positive outcome to patients. Thus, the resected specimens from surgery yield the opportunity to study the mechanisms of interictal discharges and seizure generation and propagation, as well as pharmacoresistance on viable human central nervous tissue ex vivo.
Multi-electrode arrays (MEAs), consisting of an arrangement of spatially distributed microelectrodes, allow the simultaneous stimulation and recording of electrophysiological activity from several sites of the tissue, thus providing an excellent approach to study the spatio-temporal patterns of spontaneous and evoked activity. This technique, first applied to monitor the developmental changes of neuronal cell culture activity3 and then adapted for acute and organotypic brain and spinal cord slices4&#8211;6, is presently considered a valuable electrophysiological tool.
The current protocol describes how to prepare human cortical slices from surgically resected tissue and to record with MEAs reliable ex vivo interictal and seizure-like events from these slices. This technique thus provides a way to address the basic mechanisms underlying epileptic activities initiation, propagation and effects of antiepileptic drugs at both the cellular and network levels. All the procedures used to obtain, prepare, maintain and record human slices are here described in detail.

<table><tbody><tr><td>Brain Slice Chamber-2: Interface</td><td>AutoMate Scientific, Inc.</td><td>S-BSC2</td><td>It requires separate temperature controller</td></tr><tr><td>2-channel Temperature Controller</td><td>Multichannel Systems, Germany</td><td>TC02</td><td>It allows to plug in and use 2 interface chambers.&amp;nbsp;</td></tr><tr><td>Special cable for connection of TC01 to S-BSC2 with external reference PT100</td><td>Multichannel Systems, Germany</td><td>CA3</td><td>The reference PT100 is placed near the slices&amp;nbsp;</td></tr><tr><td>6pin plug-connector with PT100</td><td>Multichannel Systems, Germany</td><td>TS-PT100</td><td>External reference for CA3 cable</td></tr><tr><td>MEA workstation for recording data from 120-electrode MEAs</td><td>Multichannel Systems, Germany</td><td>MEA2100-120</td><td>It includes MEA2100-120 headstage and MEA2100 interface board. www.multichannelsystems.com</td></tr><tr><td>Microelectrode array for MEA2100-120</td><td>Multichannel Systems, Germany</td><td>120MEA200/30</td><td>Electrode spacing: 200 &amp;micro;m; electrode diameter: 30 &amp;micro;m; glass ring: 6 mm high. Different configurations (spacing, diameter, ring) possible.&amp;nbsp;</td></tr><tr><td>Video Microscope Table</td><td>Multichannel Systems, Germany</td><td>MEA-VMT-1</td><td>Table with a video microscope underneath to image the electrode field of the MEAs in an amplifier placed on top of the table and transfer the image to a computer.</td></tr><tr><td>Perfusion cannula</td><td>Multichannel Systems, Germany</td><td>PH01</td><td>Heatable perfusion cannula with temperature sensor; temperature can be programmed with TC02 controller</td></tr><tr><td>MC_Rack</td><td>Multichannel Systems, Germany</td><td></td><td>Software for data acquisition and recordings</td></tr><tr><td>Magnetic Perfusion Holder</td><td>Multichannel Systems, Germany</td><td>MPH</td><td>Magnetic perfusion holder for PH01 element to fix the perfusion cannula and connect the perfusion system to the amplifier&#039;s ground</td></tr><tr><td>Neuroexplorer</td><td>Nex Technologies</td><td></td><td>Software for data analysis; info@neuroexplorer.com</td></tr><tr><td>Peristaltic pump (drive unit)</td><td>Gilson</td><td>F155001</td><td>0,01 to 48 rp</td></tr><tr><td>Peristaltic pump (pump head)</td><td>Gilson</td><td>F117800</td><td>R2 two channel</td></tr><tr><td>Ultrasonic aspirator</td><td>Integra Life sciences, USA</td><td>Cusa Excel +</td><td>It allows blunt subpial dissection of the cortex</td></tr><tr><td>Neuronavigation</td><td>Isis Solutions, France</td><td>Surgiscope</td><td>It allows real time identification of the brain structure on the preoperative MRI</td></tr><tr><td>Vibratome HM 650 V</td><td>Microm</td><td></td><td>Block slicing into 400 &amp;mu;m thick slices</td></tr></tbody></table>

multi-electrode array recordings of human epileptic postoperative cortical tissue

Epilepsy, affecting about 1% of the population, comprises a group of neurological disorders characterized by the periodic occurrence of seizures, which disrupt normal brain function. Despite treatment with currently available antiepileptic drugs targeting neuronal functions, one third of patients with epilepsy are pharmacoresistant. In this condition, surgical resection of the brain area generating seizures remains the only alternative treatment. Studying human epileptic tissues has contributed to understand new epileptogenic mechanisms during the last 10 years. Indeed, these tissues generate spontaneous interictal epileptic discharges as well as pharmacologically-induced ictal events which can be recorded with classical electrophysiology techniques. Remarkably, multi-electrode arrays (MEAs), which are microfabricated devices embedding an array of spatially arranged microelectrodes, provide the unique opportunity to simultaneously stimulate and record field potentials, as well as action potentials of multiple neurons from different areas of the tissue. Thus MEAs recordings offer an excellent approach to study the spatio-temporal patterns of spontaneous interictal and evoked seizure-like events and the mechanisms underlying seizure onset and propagation. Here we describe how to prepare human cortical slices from surgically resected tissue and to record with MEAs interictal and ictal-like events ex vivo.

The recording of human cortical slice activity starts while perfusing the tissue with normal recording ACSF (as previously described)7,8. In this condition, when the tissue is well preserved during surgery and later on during tissue transfer from the hospital and slice preparation, one can observe spontaneous activity, in the form of small interictal-like events, detected from small clusters of MEA electrodes. Interictal-like discharges consisted in a field potential, usually biphasic, comprising an initial sharp negative deflection, reaching tens of microvolts, followed by a longer opposite wave lasting several tens of milliseconds. Fast multi-unit activities are usually embedded in the field potential mainly in the initial part. A representative example of interictal-like activity is illustrated in Figure 3Aii, where the trace recorded by a MEA electrode is shown.
To record epileptic seizures from in vitro human cortical slices, it is necessary to perfuse them with an &#8220;epileptogenic&#8221; ACSF, in which Mg2+ is absent and K+ is raised to 6 mM, to boost tissue excitability1 (the increase of K+ from 3 to 6 mM alone is not enough to induce seizures; data not shown). Once the perfusion with 0 Mg2+ 6 mM K+ ACSF has started, the activity of cortical slices gradually increases and the first seizure appears within 15 to 20 min (15.4 &#177; 1.7 min, n = 10, from 5 patients; Figure 3 Ai). We evoked ictal-like activity, as shown in Figure 3, in 75% of patients tested and in 66.7% of all recorded slices.
Epileptic activities are recorded from adjacent electrodes of the MEA chip, and comparison of the onset dynamics of field potential events allows studying their site of genesis and propagation. A representative seizure recorded from a slice of human cerebral cortex with the MEA system is shown in Figure 3B (a representative trace of a seizure recorded by a single MEA electrode is shown at higher temporal resolution in Figure 3 Aiii). Note that the seizure is recorded from almost all the electrodes of the MEA chip, thus indicating that it is able to propagate to a large cortical area. Furthermore, looking at the single traces recorded from each electrode, it is possible to observe the progressive propagation of the seizure across the tissue: indeed, it starts first in the cortical area covering the right half of the electrode array, and it gradually spreads to the area covering the left half (i. e., compare traces from electrode L6 and B6; Figure 3B).
<img alt="Figure 1" fo:content-width="3in" src="/files/ftp_upload/51870/51870fig1highres.jpg" width="300" /> 
Figure 1: A cortical dysplasia and its surgical removal. A Taylor type focal cortical dysplasia (white arrow) embedded in a left frontal lobe sulcus is visualized on the preoperative MRI (A), and later confirmed by histological examination (B) MAP kinase labeling; scale bar: 200 &#956;m) showing a cortical dyslamination, abnormal neurons and a track on neurons with an incomplete migration in the lower white matter. During surgery, the dysplasia is localized according to the MRI data with a neuronavigation system (C). Visualization of the gyrus containing the dysplasia during surgery (D).
<img alt="Figure 2" fo:content-width="4in" src="/files/ftp_upload/51870/51870fig2highres.jpg" width="400" /> 
Figure 2: Equipment and procedure for human cortical slice preparation. An interface chamber is used to maintain human cortical slices ex vivo (A); the slices rest on a sheet of lens tissue in the flat area in the upper part of the chamber (B), where a temperature sensor (C, left), connected to a temperature controller (C, right, up) through a double-ended cable (C, right, down), is placed to maintain 37 &#176;C throughout slice incubation time. Once obtained, place the resected block of tissue in a petri dish filled with ice-cold sucrose-based ACSF (D) and remove blood clots, vessels and meninges, to facilitate the slicing. If the tissue block is bigger than MEA chambers, isolate a piece of appropriate dimensions with a blade (E) and glue it on the vibratome specimen plate (F). Cut 400 &#181;m thick slices and with the help of a spatula (G) transfer them on pieces of lens tissue (H) and incubate them in the interface chamber (I). Before recording, remove the lens tissue by submerging the slice in a petri dish filled with ACSF and transfer it to an ACSF filled MEA chip with a spatula (J). Remove the solution to let the slice adhere on the MEA (K) and keep it in position by using a platinum anchor. Place the MEA in the amplifier (L), start perfusion and recording.
<img alt="Figure 3" fo:content-width="3in" src="/files/ftp_upload/51870/51870fig3highres.jpg" width="300" /> 
Figure 3: MEA recordings of epileptic seizures in human cortical slices. (A) A representative trace of human cortical slice activity recorded by a MEA electrode is shown in panel i. In presence of normal ACSF, it is possible to observe interictal-like spontaneous activity (small grey rectangle, zoomed in panel ii); then, when the perfusion with 0 Mg2+ 6 mM K+ ACSF has started, the first seizure appears after a ~15 min delay and is followed by other events at 2-3 min interval. Panel iii shows the seizure in the big grey rectangle in panel i with a zoomed scale. The blue trace illustrates a zoom of the activity, as indicated by the blue line and arrow. Panel iv) shows the data illustrated in panel iii) high pass filtered at 250 Hz, which reveal multi-unit activity when zoomed (blue trace). (B) Representative recording of a seizure in presence of 0 Mg2+ 6 mM K+ ACSF from all MEA electrodes. Each square represents an electrode of 12 x 12 MEA array and shows an 80 sec time window; the dotted line indicates the position of the cortical surface relatively to MEA array.

Watch this Scientific Journal Video about Multi-electrode Array Recordings of Human Epileptic Postoperative Cortical Tissue at JoVE.com

Multi-electrode Array Recordings of Human Epileptic Postoperative Cortical Tissue

We here describe how to perform multi-electrode array recordings of human epileptic cortical tissue. Epileptic tissue resection, slice preparation and multi-electrode array recordings of interictal and ictal events are demonstrated in detail.

Epilepsy, affecting about 1% of the population, comprises a group of neurological disorders characterized by the periodic occurrence of seizures, which ...

multi-electrode-array-recordings-human-epileptic-postoperative

Nanyang Technological University

Research

JoVE Journal

Medicine

20.6K Views.  CNRS UMR 7241, INSERM U1050, Collège de France.  We beschrijven hier hoe je multi-electrode matrix recordings van menselijke epileptische corticale weefsel uit te voeren. Epileptische weefsel resectie, slice voorbereiding en multi-electrode matrix recordings van interictale en ictal gebeurtenissen worden gedemonstreerd in detail. 

Video: Multi-electrode Array Recordings of Human Epileptic Postoperative Cortical Tissue

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), ...

Neuroscience

Simultaneous Eye Tracking and Single-Neuron Recordings in Human Epilepsy Patients

Intracranial recordings from patients with intractable epilepsy provide a unique opportunity to study the activity of individual human neurons during active behavior. An important tool for quantifying behavior is eye tracking, which is an indispensable tool for studying visual attention. However, eye tracking is challenging to use concurrently with invasive electrophysiology and this approach has consequently been little used. Here, we present a proven experimental protocol to conduct single-neuron recordings with simultaneous eye tracking in humans. We describe how the systems are connected and the optimal settings to record neurons and eye movements. To illustrate the utility of this method, we summarize results that were made possible by this setup. This data shows how using eye tracking in a memory-guided visual search task allowed us to describe a new class of neurons called target neurons, whose response was reflective of top-down attention to the current search target. Lastly, we discuss the significance and solutions to potential problems of this setup. Together, our protocol and results suggest that single-neuron recordings with simultaneous eye tracking in humans are an effective method to study human brain function. It provides a key missing link between animal neurophysiology and human cognitive neuroscience.

We describe a method to conduct single-neuron recordings with simultaneous eye tracking in humans. We demonstrate the utility of this method and illustrate how we used this approach to obtain neurons in the human medial temporal lobe that encode targets of a visual search.

Intracranial recordings from patients with intractable epilepsy provide a unique opportunity to study the activity of individual human neurons during ...

Behavior

Investigating the Function of Deep Cortical and Subcortical Structures Using Stereotactic Electroencephalography: Lessons from the Anterior Cingulate Cortex

Stereotactic Electroencephalography (SEEG) is a technique used to localize seizure foci in patients with medically intractable epilepsy. This procedure involves the chronic placement of multiple depth electrodes into regions of the brain typically inaccessible via subdural grid electrode placement. SEEG thus provides a unique opportunity to investigate brain function. In this paper we demonstrate how SEEG can be used to investigate the role of the dorsal anterior cingulate cortex (dACC) in cognitive control. We include a description of the SEEG procedure, demonstrating the surgical placement of the electrodes. We describe the components and process required to record local field potential (LFP) data from consenting subjects while they are engaged in a behavioral task. In the example provided, subjects play a cognitive interference task, and we demonstrate how signals are recorded and analyzed from electrodes in the dorsal anterior cingulate cortex, an area intimately involved in decision-making. We conclude with further suggestions of ways in which this method can be used for investigating human cognitive processes.

Stereotactic Electroencephalography (SEEG) is an operative technique used in epilepsy surgery to help localize seizure foci. It also affords a unique opportunity to investigate brain function. Here we describe how SEEG can be used to investigate cognitive processes in human subjects.

Stereotactic Electroencephalography (SEEG) is a technique used to localize seizure foci in patients with medically intractable epilepsy. This procedure ...

Fabrication of Ti3C2 MXene Microelectrode Arrays for In Vivo Neural Recording

Implantable microelectrode technologies have been widely used to elucidate neural dynamics at the microscale to gain a deeper understanding of the neural underpinnings of brain disease and injury. As electrodes are miniaturized to the scale of individual cells, a corresponding rise in the interface impedance limits the quality of recorded signals. Additionally, conventional electrode materials are stiff, resulting in a significant mechanical mismatch between the electrode and the surrounding brain tissue, which elicits an inflammatory response that eventually leads to a degradation of the device performance. To address these challenges, we have developed a process to fabricate flexible microelectrodes based on Ti3C2 MXene, a recently discovered nanomaterial that possesses remarkably high volumetric capacitance, electrical conductivity, surface functionality, and processability in aqueous dispersions. Flexible arrays of Ti3C2 MXene microelectrodes have remarkably low impedance due to the high conductivity and high specific surface area of the Ti3C2 MXene films, and they have proven to be exquisitely sensitive for recording neuronal activity. In this protocol, we describe a novel method for micropatterning Ti3C2 MXene into microelectrode arrays on flexible polymeric substrates and outline their use for in vivo micro-electrocorticography recording. This method can easily be extended to create MXene electrode arrays of arbitrary size or geometry for a range of other applications in bioelectronics and it can also be adapted for use with other conductive inks besides Ti3C2 MXene. This protocol enables simple and scalable fabrication of microelectrodes from solution-based conductive inks, and specifically allows harnessing the unique properties of hydrophilic Ti3C2 MXene to overcome many of the barriers that have long hindered the widespread adoption of carbon-based nanomaterials for high-fidelity neural microelectrodes.

Fabrication of Ti3C2 MXene Microelectrode Arrays for In Vivo Neural Recording

We describe here a method for fabricating Ti3C2 MXene microelectrode arrays and utilizing them for in vivo neural recording.

Implantable microelectrode technologies have been widely used to elucidate neural dynamics at the microscale to gain a deeper understanding of the neural ...

Bioengineering

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 ...

Recording and Modulation of Epileptiform Activity in Rodent Brain Slices Coupled to Microelectrode Arrays

Temporal lobe epilepsy (TLE) is the most common partial complex epileptic syndrome and the least responsive to medications. Deep brain stimulation (DBS) is a promising approach when pharmacological treatment fails or neurosurgery is not recommended. Acute brain slices coupled to microelectrode arrays (MEAs) represent a valuable tool to study neuronal network interactions and their modulation by electrical stimulation. As compared to conventional extracellular recording techniques, they provide the added advantages of a greater number of observation points and a known inter-electrode distance, which allow studying the propagation path and speed of electrophysiological signals. However, tissue oxygenation may be greatly impaired during MEA recording, requiring a high perfusion rate, which comes at the cost of decreased signal-to-noise ratio and higher oscillations in the experimental temperature. Electrical stimulation further stresses the brain tissue, making it difficult to pursue prolonged recording/stimulation epochs. Moreover, electrical modulation of brain slice activity needs to target specific structures/pathways within the brain slice, requiring that electrode mapping be easily and quickly performed live during the experiment. Here, we illustrate how to perform the recording and electrical modulation of 4-aminopyridine (4AP)-induced epileptiform activity in rodent brain slices using planar MEAs. We show that the brain tissue obtained from mice outperforms rat brain tissue and is thus better suited for MEA experiments. This protocol guarantees the generation and maintenance of a stable epileptiform pattern that faithfully reproduces the electrophysiological features observed with conventional field potential recording, persists for several hours, and outlasts sustained electrical stimulation for prolonged epochs. Tissue viability throughout the experiment is achieved thanks to the use of a small-volume custom recording chamber allowing for laminar flow and quick solution exchange even at low (1 mL/min) perfusion rates. Quick MEA mapping for real-time monitoring and selection of stimulating electrodes is performed by a custom graphic user interface (GUI).

We illustrate how to perform recording and electrical modulation of 4-aminopyridine-induced epileptiform activity in rodent brain slices using microelectrode arrays. A custom recording chamber maintains tissue viability throughout prolonged experimental sessions. Live electrode mapping and selection of stimulating pairs are performed by a custom graphical user interface.

Temporal lobe epilepsy (TLE) is the most common partial complex epileptic syndrome and the least responsive to medications. Deep brain stimulation (DBS) ...

Surgical Training for the Implantation of Neocortical Microelectrode Arrays Using a Formaldehyde-fixed Human Cadaver Model

This protocol describes a procedure to assist surgeons in training for the implantation of microelectrode arrays into the neocortex of the human brain. Recent technological progress has enabled the fabrication of microelectrode arrays that allow recording the activity of multiple individual neurons in the neocortex of the human brain. These arrays have the potential to bring unique insight onto the neuronal correlates of cerebral function in health and disease. Furthermore, the identification and decoding of volitional neuronal activity opens the possibility to establish brain-computer interfaces, and thus might help restore lost neurological functions. The implantation of neocortical microelectrode arrays is an invasive procedure requiring a supra-centimetric craniotomy and the exposure of the cortical surface; thus, the procedure must be performed by an adequately trained neurosurgeon. In order to provide an opportunity for surgical training, we designed a procedure based on a human cadaver model. The use of a formaldehyde-fixed human cadaver bypasses the practical, ethical and financial difficulties of surgical practice on animals (especially non-human primates) while preserving the macroscopic structure of the head, skull, meninges and cerebral surface and allowing realistic, operating room-like positioning and instrumentation. Furthermore, the use of a human cadaver is closer to clinical daily practice than any non-human model. The major drawbacks of the cadaveric simulation are the absence of cerebral pulsation and of blood and cerebrospinal fluid circulation. We suggest that a formaldehyde-fixed human cadaver model is an adequate, practical and cost-effective approach to ensure proper surgical training before implanting microelectrode arrays in the living human neocortex.

We designed a procedure in which a formaldehyde-fixed human cadaver is used to assist neurosurgeons in training for the implantation of microelectrode arrays into the neocortex of the human brain.

This protocol describes a procedure to assist surgeons in training for the implantation of microelectrode arrays into the neocortex of the human brain. ...

Chronic Implantation of Whole-cortical Electrocorticographic Array in the Common Marmoset

Electrocorticography (ECoG) allows the monitoring of electrical field potentials from the cerebral cortex with high spatiotemporal resolution. Recent development of thin, flexible ECoG electrodes has enabled conduction of stable recordings of large-scale cortical activity. We have developed a whole-cortical ECoG array for the common marmoset. The array continuously covers almost the entire lateral surface of cortical hemisphere, from the occipital pole to the temporal and frontal poles, and it captures whole-cortical neural activity in one shot. This protocol describes a chronic implantation procedure of the array in the epidural space of the marmoset brain. Marmosets have two advantages regarding ECoG recordings, one being the homologous organization of anatomical structures in humans and macaques, including frontal, parietal, and temporal complexes. The other advantage is that the marmoset brain is lissencephalic and contains a large number of complexes, which are more difficult to access in macaques with ECoG, that are exposed to the brain surface.These features allow direct access to most cortical areas beneath the surface of the brain. This system provides an opportunity to investigate global cortical information processing with high resolutions at a sub-millisecond order in time and millimeter order in space.

We have developed a whole-cortical electrocorticographic array for the common marmoset that continuously covers almost the entire lateral surface of cortex, from the occipital pole to the temporal and frontal poles. This protocol describes a chronic implantation procedure of the array in the epidural space of the marmoset brain.

Electrocorticography (ECoG) allows the monitoring of electrical field potentials from the cerebral cortex with high spatiotemporal resolution. Recent ...

Subdural Soft Electrocorticography (ECoG) Array Implantation and Long-Term Cortical Recording in Minipigs

Neurological impairments and diseases can be diagnosed or treated using electrocorticography (ECoG) arrays. In drug-resistant epilepsy, these help delineate the epileptic region to resect. In long-term applications such as brain-computer interfaces, these epicortical electrodes are used to record the movement intention of the brain, to control the robotic limbs of paralyzed patients. However, current stiff electrode grids do not answer the need for high-resolution brain recordings and long-term biointegration. Recently, conformable electrode arrays have been proposed to achieve long-term implant stability with high performance. However, preclinical studies for these new implant technologies are needed to validate their long-term functionality and safety profile for their translation to human patients. In this context, porcine models are routinely employed in developing medical devices due to their large organ sizes and easy animal handling. However, only a few brain applications are described in the literature, mostly due to surgery limitations and integration of the implant system on a living animal.
Here, we report the method for long-term implantation (6 months) and evaluation of soft ECoG arrays in the minipig model. The study first presents the implant system, consisting of a soft microfabricated electrode array integrated with a magnetic resonance imaging (MRI)-compatible polymeric transdermal port that houses instrumentation connectors for electrophysiology recordings. Then, the study describes the surgical procedure, from subdural implantation to animal recovery. We focus on the auditory cortex as an example target area where evoked potentials are induced by acoustic stimulation. We finally describe a data acquisition sequence that includes MRI of the whole brain, implant electrochemical characterization, intraoperative and freely moving electrophysiology, and immunohistochemistry staining of the extracted brains.
This model can be used to investigate the safety and function of novel design of cortical prostheses; mandatory preclinical study to envision translation to human patients.

Subdural Soft Electrocorticography ECoG Array Implantation and Long-Term Cortical Recording in Minipigs

Here, we present a method for the long-term performance and safety assessment of soft subdural electrode arrays in a minipig model, describing surgical method and tools, postoperative magnetic resonance imaging, electrophysiology of the auditory cortex, electrochemical properties of the implant, and postmortem immunochemistry.

Neurological impairments and diseases can be diagnosed or treated using electrocorticography (ECoG) arrays. In drug-resistant epilepsy, these help ...

Recording Network Activity in Brain Assembloids: Multi-Electrode Array Approach

A Multi-Electrode Array Platform for Modeling Epilepsy Using Human Pluripotent Stem Cell-Derived Brain Assembloids

Human brain organoids are three-dimensional (3D) structures derived from human pluripotent stem cells (hPSCs) that recapitulate&#160;aspects of fetal brain development. The fusion of dorsal with ventral regionally specified brain organoids in vitro generates assembloids, which have functionally integrated microcircuits with excitatory and inhibitory neurons. Due to their structural complexity and diverse population of neurons, assembloids have become a useful in vitro tool for studying aberrant network activity. Multi-electrode array (MEA) recordings serve as a method for capturing electrical field potentials, spikes, and longitudinal network dynamics from a population of neurons without compromising cell membrane integrity. However, adhering assembloids onto the electrodes for long-term recordings can be challenging due to their large size and limited contact surface area with the electrodes. Here, we demonstrate a method to plate assembloids onto MEA plates for recording electrophysiological activity over a 2-month span. Although the current protocol utilizes human cortical organoids, it can be broadly adapted to organoids differentiated to model other brain regions. This protocol establishes a robust, longitudinal, electrophysiological assay for studying the development of a neuronal network, and this platform has the potential to be used in drug screening for therapeutic development in epilepsy.

Author Spotlight: Advancing Genetic Epilepsy Studies with Multi-Electrode Array-Based Long-Term Electrophysiological Monitoring of Human Brain Assembloids

This protocol aims to stably plate dorsal-ventral fused assembloids on multi-electrode arrays for modeling epilepsy in vitro.

Human brain organoids are three-dimensional (3D) structures derived from human pluripotent stem cells (hPSCs) that recapitulate&#160;aspects of fetal ...

Multi-electrode Array Recordings of Human Epileptic Postoperative Cortical Tissue

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