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Rapid Neuronal Differentiation of Induced Pluripotent Stem Cells for Measuring Network Activity on Micro-electrode Arrays
Neurons derived from human induced Pluripotent Stem Cells (hiPSCs) provide a promising new tool for studying neurological disorders. In the past decade, many protocols for differentiating hiPSCs into neurons have been developed. However, these protocols are often slow with high variability, low repr...
Autores principales: | , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MyJove Corporation
2017
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5407693/ https://www.ncbi.nlm.nih.gov/pubmed/28117798 http://dx.doi.org/10.3791/54900 |
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author | Frega, Monica van Gestel, Sebastianus H. C. Linda, Katrin van der Raadt, Jori Keller, Jason Van Rhijn, Jon-Ruben Schubert, Dirk Albers, Cornelis A. Nadif Kasri, Nael |
author_facet | Frega, Monica van Gestel, Sebastianus H. C. Linda, Katrin van der Raadt, Jori Keller, Jason Van Rhijn, Jon-Ruben Schubert, Dirk Albers, Cornelis A. Nadif Kasri, Nael |
author_sort | Frega, Monica |
collection | PubMed |
description | Neurons derived from human induced Pluripotent Stem Cells (hiPSCs) provide a promising new tool for studying neurological disorders. In the past decade, many protocols for differentiating hiPSCs into neurons have been developed. However, these protocols are often slow with high variability, low reproducibility, and low efficiency. In addition, the neurons obtained with these protocols are often immature and lack adequate functional activity both at the single-cell and network levels unless the neurons are cultured for several months. Partially due to these limitations, the functional properties of hiPSC-derived neuronal networks are still not well characterized. Here, we adapt a recently published protocol that describes production of human neurons from hiPSCs by forced expression of the transcription factor neurogenin-2(12). This protocol is rapid (yielding mature neurons within 3 weeks) and efficient, with nearly 100% conversion efficiency of transduced cells (>95% of DAPI-positive cells are MAP2 positive). Furthermore, the protocol yields a homogeneous population of excitatory neurons that would allow the investigation of cell-type specific contributions to neurological disorders. We modified the original protocol by generating stably transduced hiPSC cells, giving us explicit control over the total number of neurons. These cells are then used to generate hiPSC-derived neuronal networks on micro-electrode arrays. In this way, the spontaneous electrophysiological activity of hiPSC-derived neuronal networks can be measured and characterized, while retaining interexperimental consistency in terms of cell density. The presented protocol is broadly applicable, especially for mechanistic and pharmacological studies on human neuronal networks. |
format | Online Article Text |
id | pubmed-5407693 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | MyJove Corporation |
record_format | MEDLINE/PubMed |
spelling | pubmed-54076932017-05-12 Rapid Neuronal Differentiation of Induced Pluripotent Stem Cells for Measuring Network Activity on Micro-electrode Arrays Frega, Monica van Gestel, Sebastianus H. C. Linda, Katrin van der Raadt, Jori Keller, Jason Van Rhijn, Jon-Ruben Schubert, Dirk Albers, Cornelis A. Nadif Kasri, Nael J Vis Exp Developmental Biology Neurons derived from human induced Pluripotent Stem Cells (hiPSCs) provide a promising new tool for studying neurological disorders. In the past decade, many protocols for differentiating hiPSCs into neurons have been developed. However, these protocols are often slow with high variability, low reproducibility, and low efficiency. In addition, the neurons obtained with these protocols are often immature and lack adequate functional activity both at the single-cell and network levels unless the neurons are cultured for several months. Partially due to these limitations, the functional properties of hiPSC-derived neuronal networks are still not well characterized. Here, we adapt a recently published protocol that describes production of human neurons from hiPSCs by forced expression of the transcription factor neurogenin-2(12). This protocol is rapid (yielding mature neurons within 3 weeks) and efficient, with nearly 100% conversion efficiency of transduced cells (>95% of DAPI-positive cells are MAP2 positive). Furthermore, the protocol yields a homogeneous population of excitatory neurons that would allow the investigation of cell-type specific contributions to neurological disorders. We modified the original protocol by generating stably transduced hiPSC cells, giving us explicit control over the total number of neurons. These cells are then used to generate hiPSC-derived neuronal networks on micro-electrode arrays. In this way, the spontaneous electrophysiological activity of hiPSC-derived neuronal networks can be measured and characterized, while retaining interexperimental consistency in terms of cell density. The presented protocol is broadly applicable, especially for mechanistic and pharmacological studies on human neuronal networks. MyJove Corporation 2017-01-08 /pmc/articles/PMC5407693/ /pubmed/28117798 http://dx.doi.org/10.3791/54900 Text en Copyright © 2017, Journal of Visualized Experiments http://creativecommons.org/licenses/by-nc-nd/3.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License. To view a copy of this license, visithttp://creativecommons.org/licenses/by-nc-nd/3.0/ |
spellingShingle | Developmental Biology Frega, Monica van Gestel, Sebastianus H. C. Linda, Katrin van der Raadt, Jori Keller, Jason Van Rhijn, Jon-Ruben Schubert, Dirk Albers, Cornelis A. Nadif Kasri, Nael Rapid Neuronal Differentiation of Induced Pluripotent Stem Cells for Measuring Network Activity on Micro-electrode Arrays |
title | Rapid Neuronal Differentiation of Induced Pluripotent Stem Cells for Measuring Network Activity on Micro-electrode Arrays |
title_full | Rapid Neuronal Differentiation of Induced Pluripotent Stem Cells for Measuring Network Activity on Micro-electrode Arrays |
title_fullStr | Rapid Neuronal Differentiation of Induced Pluripotent Stem Cells for Measuring Network Activity on Micro-electrode Arrays |
title_full_unstemmed | Rapid Neuronal Differentiation of Induced Pluripotent Stem Cells for Measuring Network Activity on Micro-electrode Arrays |
title_short | Rapid Neuronal Differentiation of Induced Pluripotent Stem Cells for Measuring Network Activity on Micro-electrode Arrays |
title_sort | rapid neuronal differentiation of induced pluripotent stem cells for measuring network activity on micro-electrode arrays |
topic | Developmental Biology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5407693/ https://www.ncbi.nlm.nih.gov/pubmed/28117798 http://dx.doi.org/10.3791/54900 |
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