Name: A Multi-Electrode Array Platform for Modeling Epilepsy Using Human Pluripotent Stem Cell-Derived Brain Assembloids (Video) | JoVE Journal
Uploaded: 2024-09-27T14:57:00.000+00:00
Duration: 6 min 30 s
Description: Learn multi-electrode array techniques for modeling epilepsy using human stem cell-derived brain assembloids. This method enables electrophysiological recordings to study genetic epilepsy mechanisms and drug responses in vitro.

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This manuscript was supported by R01NS127829 NIH/NINDS (LTD). Figure 1 has been generated using biorender.com.

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

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The authors have no relevant disclosures.

MEA-based methods for electrophysiological recordings of network activity in iPSC-derived assembloids have been used for in vitro modeling of epilepsy22,23. This proposed platform integrating excitatory and inhibitory synaptic connections has the potential to address mechanisms of neuronal hyperexcitability and the role of cortical interneurons in the process of epileptogenesis. Additionally, this platform allows for the stabilization of assembloids and collection of longitudinal electrophysiological data for about 50 days in vitro, even in the presence of pharmacological perturbations, thereby providing an approach for testing compounds with potential therapeutic targets.
Stabilizing mature, relatively large assembloids on a hydrophobic surface for a long period in vitro has been technically challenging. To overcome this obstacle, the twice-coating strategy is applied with conversely charged chemicals to facilitate short- and long-term adherence of organoids to the electrode surface. This protocol optimizes the previously used method24 by switching the plate-coating reagents from poly-L-ornithine (PLO)/laminin to PEI/BMM, achieving solid attachment with high success rates and low toxicity. Furthermore, this stabilized long-term culture system has the potential to yield high-throughput data. For example, pre-labeling with dual viruses can complement spike sorting by individual electrodes to localize and compare activity from different regions. The proposed platform also provides a stable tool for recording local field potentials, which is highly relevant to interneuron function without disrupting the intact structure of assembloids25. The major limitation of this protocol is that assembloids are flattened after plating, which raises concerns about whether the initial 3D structures are preserved or not. Growing organoids around the electrodes using more advanced MEA platforms that envelope an organoid with electrodes26 could potentially overcome the limitation. 
One of the main obstacles of MEA-based methods for electrophysiological recording of organoids is the variability and reproducibility of experimental findings across differentiation protocols and even batches of differentiation. Previous studies show that the network activity of neurons measured by MEA is dependent upon the neuronal maturation status of both dorsal and ventral organoids27. Recording from assembloids at a later timepoint with relatively mature neurons and circuits is one way to improve consistency. Furthermore, the choice of multi-well MEA systems generating high throughput data will allow statistical comparisons in light of experimental variability. If the electrophysiological activity of a culture is inconsistent with what is expected at a given time point, collecting the intact culture from the functional platform for quality control measures, including immunocytochemical and transcriptomic analysis, will be important and helpful.
Paramount to the proposed platform is to functionally integrate regionally specified excitatory cortical neurons on the dorsal side and inhibitory interneurons on the ventral side. Protocols for the generation of brain region-specific organoids found in the literature differ extensively in relation to media conditions, the timing of cultures, yield, and maturation profiles, among other factors28. While this protocol is cortex- and ganglionic eminence-specific in its ability to generate excitatory neurons and GABAergic interneurons, well-established differentiation techniques can be used to generate iPSC-derived neurons displaying other identities, such as motor neurons29 and Purkinje cells30. Thus, the proposed platform has the potential to model neural circuits between the cerebral cortex and the spinal cord/ cerebellum. After some optimization related to the timing of plating and visualization of cell types with viral labeling, this method can be translated to other disease models aside from epilepsy, like Amyotrophic Lateral Sclerosis (ALS)17.

Human pluripotent stem cell (hPSC)-derived brain organoids are spatially self-organized 3D structures mirroring the tissue architecture and developmental trajectories in vivo. They are composed of multiple cell types, including progenitors (neuroepithelial cells, radial glia, neuronal progenitors, glial progenitors), neurons (cortical-like excitatory neurons and inhibitory interneurons), and glial cells (astrocytes and oligodendrocytes)1,2. Assembloids represent the next generation of brain organoids, capable of integrating multiple brain regions and/or cell lineages within a 3D culture. They provide a useful tool for modeling connections between various brain regions resembling the in vivo counterparts, capturing interactions between neurons and astrocytes to better mimic mature and complex neural networks, and to study the assembly of neural circuits. Therefore, assembloids are becoming a widely used tool to recapitulate hallmarks of epilepsy pathophysiology, in which functional measures are necessary to interrogate aberrant neural networks that may underlie the cause of the disease3,4,5,6.
To model interactions between cortical glutamatergic neurons and GABAergic interneurons, several groups developed separate organoids resembling the dorsal and ventral brain and then fused them together into a multi-region assembloid7,8,9,10. Here, a previously described assembloid culture protocol with regionally specific neural subtypes was applied9. However, a significant barrier is the lack of reproducible functional assays to monitor neural network activity during neurodevelopment. Many functional tests of the networks in organoids yield results that have high variability among batches of differentiations and cell lines. Techniques that involve slicing or dissociating organoids alter their inherent networks by severing synaptic connectivity8.
Multi-electrode arrays (MEA) provide a large-scale view of network activity over time with high temporal resolution to characterize electrophysiological properties of organoids without disrupting culture conditions or cell membrane integrity11. Compared to patch clamp electrophysiology, MEA enables high-throughput data acquisition based on large populations of neurons rather than single cells. MEA platforms vary in their electrode density, catering to different needs in brain organoid research12. The widely used systems, as shown in this protocol, record from 8 to 64 electrodes per well13,14,15. High-density MEAs with up to 26,400 electrodes per well enable increased spatial and temporal resolution, quantification of action potential propagation speed, and combination with optogenetic stimulation14,16,17. Therefore, MEA serves as a powerful tool for modeling epilepsy in vitro and a translational paradigm for anti-seizure medication screening.
One major challenge is to stabilize the large-sized assembloids on a hydrophobic metal surface for long-term recordings. This protocol outlines a detailed methodology for plating intact assembloids on MEA plates for long-term longitudinal recording together with pharmacological assays. Unique advantages of the protocol include stable attachment of assembloids to the electrode surface without losing electrical activity, use of commercially available neurophysiological basal media to accelerate functional network maturation after plating, feasibility to conduct downstream functional assays like drug treatment, and wide application to organoids generated with other region-specific protocols.
The goal is to provide a functional assay with high temporal resolution to investigate network activity, examine disease-specific changes, and test drugs with therapeutic potential in epilepsy. Video instructions are provided for the most challenging steps of this protocol, showing the techniques for plating assembloids on MEA plates, as well as representative recordings from these cultures.

<table><tbody><tr><td>10 cm Corning Non TC-treated culture dishes</td><td>Corning</td><td>08-772-32</td><td>For suspension culture on the shaker</td></tr><tr><td>100 mL Beaker&amp;nbsp;</td><td>Fisher Scientific</td><td>FB100100</td><td></td></tr><tr><td>100% Ethanol</td><td>Fisher Scientific</td><td>BP28184-4L</td><td></td></tr><tr><td>2-Mercaptoethanol (&amp;beta;-ME)</td><td>Thermo Fisher</td><td>21985023</td><td>Working concentration 100 &amp;mu;M</td></tr><tr><td>48-well cell culture plate</td><td>Fisher Scientific</td><td>50-202-140</td><td></td></tr><tr><td>6-well cell culture plate</td><td>Fisher Scientific</td><td>07-200-83</td><td></td></tr><tr><td>Aggrewell 800</td><td>Fisher Scientific</td><td>501974754</td><td></td></tr><tr><td>Allegra X-14R Refrigerated Centrifuge</td><td>Beckman Coulter&amp;nbsp;</td><td>BE-AX14R</td><td></td></tr><tr><td>Allopregnanolone</td><td>Cayman</td><td>16930</td><td>Suspended 5mg into DMSO to get 1 mM stock solution. Aliquot and freeze at &amp;minus;80 &amp;deg;C. Dilute at 1:10,000 for use. Working concentration 100 nM.</td></tr><tr><td>Automated cell counter</td><td>Thermo Fisher</td><td>AMQAX2000</td><td></td></tr><tr><td>Axion CytoView MEA 6-well plates&amp;nbsp;</td><td>Axion Biosystems</td><td>M384-tMEA-6B</td><td></td></tr><tr><td>Axion Maestro MEA platform</td><td>Axion Biosystems</td><td>Maestro</td><td>With temp environmental control</td></tr><tr><td>B-27 supplement (regular, with Vitamin A)</td><td>Thermo Fisher</td><td>21985023</td><td></td></tr><tr><td>B-27 supplement (without Vitamin A)</td><td>Thermo Fisher</td><td>12587010</td><td></td></tr><tr><td>Basement membrane matrix- Geltrex</td><td>Thermo Fisher</td><td>A1569601</td><td></td></tr><tr><td>Bead bath</td><td>Fisher Scientific</td><td>10-876-001</td><td>Isotemp</td></tr><tr><td>Benchtop inverted microscope</td><td>Olympus</td><td>CKX53</td><td>Kept in laminar flow clean bench</td></tr><tr><td>Bicuculline</td><td>Sigma-Aldrich</td><td>14340</td><td>Working concentration 10 &amp;mu;M</td></tr><tr><td>Bleach</td><td>CLOROX</td><td>67619-26</td><td></td></tr><tr><td>Borate buffer 20x</td><td>Thermo Fisher</td><td>28341</td><td>Working concentration at 1x</td></tr><tr><td>BrainPhys media</td><td>StemCell Technologies</td><td>5790</td><td></td></tr><tr><td>Cell dissociation reagent (StemPro Accutase)</td><td>Thermo Fisher</td><td>A1110501</td><td></td></tr><tr><td>Celltron orbital shaker</td><td>HT-Infors</td><td>&amp;nbsp;I69222&amp;nbsp;</td><td></td></tr><tr><td>Detergent/enzyme (Terg-A-Zyme)</td><td>Sigma-Aldrich</td><td>Z273287</td><td>Working concentration 1% m/v</td></tr><tr><td>DMEM/F12 + HEPES/L-Glutamine</td><td>Thermo Fisher</td><td>113300</td><td></td></tr><tr><td>DMSO</td><td>Sigma-Aldrich</td><td>67685</td><td></td></tr><tr><td>Dorsomorphin</td><td>Sigma-Aldrich</td><td>P5499</td><td>Dissolve 5mg into DMSO to get 10 mM stock solution. Aliquot and freeze at &amp;minus;20 &amp;deg;C. Dilute at 1:2000 for use. (working concentration 5 &amp;mu;M)</td></tr><tr><td>D-PBS w/o calcium or magnesium</td><td>Thermo Fisher</td><td>14190144</td><td></td></tr><tr><td>Glial cell line-derived neurotrophic (GDNF)</td><td>Peprotech</td><td>450-10</td><td>Dissolve 100 &amp;mu;g in 1mL of PBS to 100 &amp;mu;g/mL. Aliquot and freeze at &amp;minus;20 &amp;deg;C. Dilute at 1:5000 for use. (working concentration 20 ng/mL).</td></tr><tr><td>GlutaMAX supplement</td><td>Thermo Fisher</td><td>35050061</td><td></td></tr><tr><td>Hemacytometer</td><td>Election Microscopy Sciences</td><td>63510-20</td><td></td></tr><tr><td>HEPES</td><td>Thermo Fisher</td><td>15630080</td><td></td></tr><tr><td>Heraguard ECO Clean Bench</td><td>Thermo Fisher</td><td>51029692</td><td></td></tr><tr><td>Humidity controlled cell culture incubator</td><td>Thermo Fisher</td><td>370</td><td>set to 37 &amp;deg;C, 5% CO2</td></tr><tr><td>IWP-2</td><td>Selleckchem</td><td>S7085</td><td>Aliquot and freeze at &amp;minus;80 &amp;deg;C. It will precipitate if thawed at room temp. Frozen aliquots should be placed directly into 37 &amp;deg;C before use.</td></tr><tr><td>Knockout serum replacement (KOSR)</td><td>Thermo Fisher</td><td>10828010</td><td></td></tr><tr><td>mTeSRplus (medium + supplements)</td><td>StemCell Technologies</td><td>100-0276</td><td>cGMP, stabilized feeder-free medium for human iPSC cells</td></tr><tr><td>N2 supplement</td><td>Thermo Fisher</td><td>17502048</td><td></td></tr><tr><td>Neurobasal A</td><td>Thermo Fisher</td><td>21103049</td><td></td></tr><tr><td>Non-essential amino acids (NEAA)</td><td>Thermo Fisher</td><td>11140050</td><td></td></tr><tr><td>NT3</td><td>Peprotech</td><td>450-03</td><td>Dissolve 100 &amp;mu;g in 1mL of PBS to 100 &amp;mu;g/mL. Aliquot and freeze at &amp;minus;20 &amp;deg;C. Dilute at 1:5000 for use. (working concentration 20 ng/mL).</td></tr><tr><td>NuFF Human neonatal foreskin fibroblasts</td><td>MTI-GlobalStem</td><td>GSC-3002</td><td></td></tr><tr><td>Parafilm</td><td>PARAFILM</td><td>P7793</td><td></td></tr><tr><td>Penicilin/Streptomycin</td><td>Thermo Fisher</td><td>15140122</td><td></td></tr><tr><td>Pipette (P10, P200, P1000)</td><td>Eppendorf</td><td>EP4926000034</td><td>Autoclaved cut P1000 tips for organoid collection</td></tr><tr><td>Poly (Ethyleneimine) (PEI)</td><td>Sigma-Aldrich</td><td>P3143</td><td>Dilute stock in sterile borate buffer. Working concentration 0.07%. See details in manuscript.</td></tr><tr><td>Recombinant human epidermal growth factor (EGF)</td><td>R&amp;amp;D Systems</td><td>236-EG-200</td><td>Suspended in PBS. Aliquot and freeze at -20 &amp;deg;C.</td></tr><tr><td>Recombinant Human fibroblast growth factor (FGF)-basic&amp;nbsp;</td><td>Peprotech</td><td>100-18B</td><td>Suspended in PBS. Aliquot and freeze at -20 &amp;deg;C.</td></tr><tr><td>Recombinant human-brain-derived neurotrophic factor (BDNF)</td><td>Peprotech</td><td>450-02</td><td>Centrifuge briefly before reconstitution. Dissolve 100 &amp;mu;g in 1 mL of PBS to 100 &amp;mu;g/mL. Aliquot and freeze at &amp;minus;20 &amp;deg;C. Dilute at 1:5000 for use. (working concentration 20 ng/mL).</td></tr><tr><td>Retinoic acid (RA)</td><td>Sigma-Aldrich</td><td>R2625</td><td>Dissolve 100 mg into 3.3 mL of DMSO to get 100 mM stock solution. Aliquot the stock 100 &amp;mu;L/tube and freeze at &amp;minus;80 &amp;deg;C. Take 200 &amp;mu;L of 100 mM stock and dilute 10x (add 1.8 mL of DMSO) to make 10 mM stock. Aliquot 50 &amp;mu;L/tube and store at &amp;minus;80 &amp;deg;C. Dilute at 1:100,000 for use. (working concentration 100 nM).</td></tr><tr><td>ROCK inhibitor Y-27632</td><td>Tocris</td><td>1254</td><td>1:200 from 10 mM stock</td></tr><tr><td>SAG (smoothened agonist)</td><td>Selleckchem</td><td>S7779</td><td>Aliquot and freeze at &amp;minus;80 &amp;deg;C. Stock concentration 1mM. Use at 1:10,000 dilution (working concentration 100 nM).</td></tr><tr><td>SB-431542</td><td>Tocris</td><td>1614</td><td>Dissolve 5mg into 1.3 mL of DMSO to get 10 mM stock solution. Aliquot and freeze at &amp;minus;80 &amp;deg;C. Dilute at 1:1000 for use. (working concentration 10 &amp;mu;M)</td></tr><tr><td>Serological pipette filler</td><td>Fisher Scientific</td><td>14-387-166</td><td></td></tr><tr><td>Steriflip vacuum tube top filter</td><td>Sigma-Aldrich</td><td>SE1M179M6</td><td></td></tr><tr><td>Sterile cell culture hoods</td><td>Baker Company</td><td>SG-600</td><td></td></tr><tr><td>Trypan blue solution (0.4%)</td><td>Thermo Fisher</td><td>15250061</td><td></td></tr><tr><td>Trypsin-EDTA (0.25%)</td><td>Thermo Fisher</td><td>25200056</td><td></td></tr><tr><td>Zoom stereomicroscope</td><td>Olympus</td><td>SZ61/SZ51</td><td>Kept in laminar flow clean bench</td></tr></tbody></table>

a multi-electrode array platform for modeling epilepsy using human pluripotent stem cell-derived brain assembloids

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.

Human dorsal-ventral assembloids were plated on a 6-well MEA plate (n = 6 per differentiation, 3 differentiations), with each well containing 64 electrodes (Figure 2A). Nine out of ten assembloids planned for longitudinal recording were firmly attached to the electrodes for over 50 days in vitro (Figure 2D). 6 out of 8 assembloids designated for pharmacological assays were successfully settled through the wash-out phase with development stage-appropriate network activity (Figure 2E). Phase contrast and fluorescence images from one representative well are shown (Figure 2B), in which the AAV-CAG-tdTomato-labeled dorsal and AAV-mDLX-GFP-labeled ventral organoids are easily distinguished7,9.
Activity of neurons on an MEA can be analyzed to assess overall cellular excitability and network activity, through a variety of parameters. Overall firing rates across the culture provide information on cellular and network excitability, while the frequency of synchronized bursts across multiple electrodes indicates the range of synaptically active networks of neurons. Multiple repeated spikes occurring across spatially distinct electrodes over a short period of time (network bursting) can serve as a biological correlate to seizure-like activity, although assembloids on MEAs remain limited compared to a whole organism that can have bona fide seizures. Figure 2C illustrates the network bursting patterns of assembloids cultured in commercially available neurophysiological basal media on Days 84 and 91, with increased bursting duration at the later timepoint likely due to functional maturation of the neurons. In this case, cellular and network activity gradually rises from the day when assembloids are plated, usually peaking on Day 92, and almost fading away on Day 125 (Figure 2D). The increased maturation at the later stage likely contributes to the extended synchronized burst duration at the later recording time point.
Bicuculline, a GABAA receptor antagonist, enhances the synchronized network activity of the assembloids, as shown by significantly higher firing rates and (network) burst frequencies after adding the drug on Day 108 (Figure 2E). This is similar to what has been previously demonstrated in single-cell electrophysiology recordings21. A power analysis for the pharmacological assay was conducted using G*Power3 base on an effect size of 0.94, and an alpha of 0.05. A total sample of 15 assembloids with three equal-sized groups of n = 5 was required to achieve a power of 0.8 (Figure 2E). A vehicle control is included to verify that the response is a true signaling event and not merely a change in signal intensity that may arise from the addition of a small amount of media (Figure 2E).
<img alt="Neural differentiation process timeline diagram; cell culture, medium changes, viral labeling, MEA recording." class="xfigimg" src="/files/ftp_upload/67396/67396fig01.jpg" data-altupdatedat="1788304762000" data-alt="Figure 1"> 
Figure 1: Overview of the study. Protocol for the generation and plating of dual labeled assembloids on MEA timeline of the differentiation protocol detailing critical stages and corresponding small-molecule supplements (for additional information on the composition, see Table 1 and Table of Materials). <a href="https://www.jove.com/files/ftp_upload/67396/67396fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Neural activity measurement diagrams and graphs showing mean firing rates; microelectrode array setup." class="xfigimg" src="/files/ftp_upload/67396/67396fig02.jpg" data-altupdatedat="1788304762000" data-alt="Figure 2"> 
Figure 2: MEA recording of assembloids. (A) Heat map of mean firing activity from a single MEA plate (n = 6 wells) at Day 82, i.e., post-plating day 10. All wells represent cultures of the same condition and genotype. (B) Representative phase-contrast and confocal images of the assembloid, with dorsal assembloid labeled with red and ventral with green. White arrowheads indicate migrated GABAergic interneurons from the ventral to the dorsal side. Scale bar = 500 µm. (C) Representative raster plots over 120 s of recording time from a single well. Network bursts are highlighted with a purple box. Two separate timepoints, Day 78 (1 week after plating, starting day of recording) and Day 92 (3 weeks after plating, also when the activity peaks), are demonstrated. (D) Quantification of mean firing rates, burst frequency, and network bursting frequency from day 78 to day 127 (n = 3 differentiations, 3 assembloids/ differentiation). Each curve indicates one individual assembloid, compiled from 3 differentiations (color-coded). The red curve demonstrates the mean value. (E) Quantification of bicuculline assay results (n = 6 assembloids/ condition). The bar graph was displayed as mean ± SEM. Each dot represents one assembloid, compiled from 3 batches of differentiation (color-coded). A nonparametric Friedman test was performed. **p &lt; 0.01; ns, not significant. MEA, multi-electrode arrays. Heat map and raster plots are generated using the analytic software Neural Metric Tool. <a href="https://www.jove.com/files/ftp_upload/67396/67396fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Table 1: Components of media at various differentiation stages. <a href="https://www.jove.com/files/ftp_upload/67396/67396_Table%201.xlsx" target="_blank">Please click here to download this Table.</a>

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 ...

a-multi-electrode-array-platform-for-modeling-epilepsy-using-human

Research

JoVE Journal

Neuroscience

The video demonstrates a method for generating excitatory cortical neurons from human-induced pluripotent stem cells (hiPSCs). Upon doxycycline induction, the hiPSCs overexpress neurogenin-2, a neural-specific transcription factor. By using specific media and co-culturing with astrocytes, this process facilitates the hiPSCs' differentiation into neurons with synaptic connections.

Differentiating Human Induced Pluripotent Stem Cells into Neurons on Micro-Electrode Arrays

This video demonstrates the procedure of using the multielectrode array (MEA) to measure synaptic physiology in 3D coculture spheres. This method allows to understand the neural network dynamics and synaptic network burst activity within the 3D spheres.

Utilizing Multielectrode Arrays to Measure Synaptic Physiology in 3D Coculture Spheres

This protocol involves using a microelectrode array (MEA) to record electrical activity in a brain slice exposed to a seizure-inducing drug. The procedure includes positioning the brain slice in an MEA recording chamber, applying electrical stimulation to induce artificial seizures, and analyzing the resulting signals.

Monitoring Seizure-Induced Neural Electrical Activity in Brain Slices Using Microelectrode Arrays

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.

Multi-electrode Array Recordings of Human Epileptic Postoperative Cortical Tissue

Electrophysiological characterization of cardiomyocytes derived from human Pluripotent Stem Cells (hPSC-CMs) is crucial for cardiac disease modeling and for determining drug responses. This protocol provides the necessary information to dissociate and plate hPSC-CMs on multi-electrode arrays, measure their field potential, and a method for analyzing QT and RR intervals.

Electrophysiological Analysis of human Pluripotent Stem Cell-derived Cardiomyocytes (hPSC-CMs) Using Multi-electrode Arrays (MEAs)

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.

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

In this protocol, we aim to describe a reproducible method for combining dissociated human pluripotent stem cell derived neurons and astrocytes together into 3D sphere cocultures, maintaining these spheres in free floating conditions, and subsequently measuring synaptic circuit activity of the spheres with immunoanalysis and multielectrode array recordings.

Synaptic Microcircuit Modeling with 3D Cocultures of Astrocytes and Neurons from Human Pluripotent Stem Cells

Here, we describe the preparation of rhinal cortex-hippocampus organotypic slices. Under a gradual and controlled deprivation of serum, these slices depict evolving epileptic-like events and can be considered an ex vivo model of epileptogenesis. This system represents an excellent tool for monitoring the dynamics of spontaneous activity, as well as for assessing the progression of neuroinflammatory features throughout the course of epileptogenesis.

A Model of Epileptogenesis in Rhinal Cortex-Hippocampus Organotypic Slice Cultures

We describe a method for differentiating spinal cord human induced pluripotent-derived astrocytes and neurons and their co-culture for electrophysiological recording.

Establishment of an Electrophysiological Platform for Modeling ALS with Regionally-Specific Human Pluripotent Stem Cell-Derived Astrocytes and Neurons

The amygdala plays a key role in temporal lobe epilepsy, which originates in and propagates from this structure. This article provides a detailed description of the fabrication of deep brain electrodes with both recording and stimulating functions. It introduces a model of medial temporal lobe epilepsy originating from the amygdala.

Using a Bipolar Electrode to Create a Temporal Lobe Epilepsy Mouse Model by Electrical Kindling of the Amygdala

Here, we employ HD-MEA to delve into computational dynamics of large-scale neuronal ensembles, particularly in hippocampal, olfactory bulb circuits, and human neuronal networks. Capturing spatiotemporal activity, combined with computational tools, provides insights into neuronal ensemble complexity. The method enhances understanding of brain functions, potentially identifying biomarkers and treatments for neurological disorders.

Recording and Analyzing Multimodal Large-Scale Neuronal Ensemble Dynamics on CMOS-Integrated High-Density Microelectrode Array

Learn multi-electrode array techniques for modeling epilepsy using human stem cell-derived brain assembloids. This method enables electrophysiological recordings to study genetic epilepsy mechanisms and drug responses in vitro.

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

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