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Image-based quantitation of kainic acid-induced excitotoxicity as a model of neurodegeneration in Human iPSC-Derived Neurons

Talbot, J, Chear, S, Phipps, A ORCID: 0000-0003-4689-1198, Pebay, A, Hewitt, AW ORCID: 0000-0002-5123-5999, Vickers, JC ORCID: 0000-0001-5671-4879, King, AE ORCID: 0000-0003-1792-0965 and Cook, AL ORCID: 0000-0003-1770-7910 2021 , 'Image-based quantitation of kainic acid-induced excitotoxicity as a model of neurodegeneration in Human iPSC-Derived Neurons' , Methods in Molecular Biology , doi: 10.1007/7651_2021_421.

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Abstract

Excitotoxicity is a feature of many neurodegenerative diseases and acquired forms of neural injury that is characterized by disruption of neuronal morphology. This is typically seen as beading and fragmentation of neurites when exposed to excitotoxins such as the AMPA receptor agonist kainic acid, with the extent to which these occur used to quantitate neurodegeneration. Induced pluripotent stem cells (iPSCs) provide a means to generate human neurons in vitro for mechanistic studies and can thereby be used to investigate how cells respond to excitotoxicity and to identify or test potential neuroprotective agents. To facilitate such studies, we have optimized a protocol for human iPSC differentiation to mature neurons in a 96-well plate format that enables image-based quantitation of changes to neuron morphology when exposed to kainic acid. Our protocol assays neuron morphology across seven excitotoxin concentrations with multiple control conditions and is ideally suited to comparison of neurons generated through differentiation of two isogenic iPSC lines in a single plate. We have included detailed step-by-step protocols for neural stem cell differentiation, neuronal maturation and exposure to kainic acid treatment, as well as different approaches to image-based quantitation that involve immunofluorescence or phase-contrast microscopy.

Item Type: Article
Authors/Creators:Talbot, J and Chear, S and Phipps, A and Pebay, A and Hewitt, AW and Vickers, JC and King, AE and Cook, AL
Keywords: stem cell, neurodegeneration, disease model, image analysis
Journal or Publication Title: Methods in Molecular Biology
Publisher: Humana Press
ISSN: 1064-3745
DOI / ID Number: 10.1007/7651_2021_421
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Copyright 2021 Springer Science+Business Media, LLC

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