In Vitro Techniques for Assessing Neurotoxicity Using Human iPSC-Derived Neuronal Models

  • Anke M. Tukker
  • Fiona M. J. Wijnolts
  • Aart de Groot
  • Richard W. Wubbolts
  • Remco H. S. WesterinkEmail author
Part of the Neuromethods book series (NM, volume 145)


The central nervous system consists of a multitude of different neurons and supporting cells that form networks for transmitting neuronal signals. Proper function of the nervous system depends critically on a wide range of highly regulated processes including intracellular calcium homeostasis, neurotransmitter release, and electrical activity. Due to the diversity of cell types and complexity of signaling processes, the (central) nervous system is very vulnerable to toxic insults.

Nowadays, a broad range of approaches and cell models is available to study neurotoxicity. In this chapter we show the applicability of human induced pluripotent stem cell (hiPSC)-derived neuronal co-cultures for in vitro neurotoxicity testing. We demonstrate that immunocytochemistry can be used to visualize networks of cultured cells and to differentiate between different cell types. Live cell imaging and electrophysiology techniques demonstrate that the neuronal networks develop spontaneous activity, including synchronized calcium oscillations that coincide with spontaneous changes in membrane potential as well as spontaneous electrical activity with defined (network) bursting. Importantly, as shown in this chapter, spontaneously active human iPSC-derived neuronal co-cultures are suitable for in vitro neurotoxicity assessment. Future application of live imaging and electrophysiological techniques on hiPSC from different donors and/or patients differentiated in different cell types holds great promise for personalized neurotoxicity assessment and safety screening.

Key words

In vitro neurotoxicity screening Human induced pluripotent stem cell-derived neuronal models Single-cell fluorescent microscopy Calcium homeostasis Membrane potential Spontaneous neuronal activity Immunocytochemistry Multi-well microelectrode array 



We gratefully acknowledge members of the Neurotoxicology Research Group for helpful discussions. This work was funded by a grant from the National Centre for the Replacement, Refinement and Reduction of Animals in Research (NC3Rs; project number 50308-372160), the Netherlands Organisation for Health Research and Development (ZonMW; InnoSysTox project number 114027001), and the Faculty of Veterinary Medicine (Utrecht University, The Netherlands).


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Anke M. Tukker
    • 1
  • Fiona M. J. Wijnolts
    • 1
  • Aart de Groot
    • 1
  • Richard W. Wubbolts
    • 2
  • Remco H. S. Westerink
    • 1
    Email author
  1. 1.Neurotoxicology Research Group, Toxicology and Pharmacology Division, Institute for Risk Assessment Sciences (IRAS), Faculty of Veterinary MedicineUtrecht UniversityUtrechtThe Netherlands
  2. 2.Centre for Cell Imaging (CCI), Department of Biochemistry and Cell Biology, Faculty of Veterinary MedicineUtrecht UniversityUtrechtThe Netherlands

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