Improvements of spontaneous electrical activities of human induced pluripotent stem cell-derived neuronal networks, which can serve as an alternative method to study neurotoxic and pharmacological effects in vitro
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1
Leibniz-Institut für umweltmedizinische Forschung (IUF), Germany
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2
Lübecker Institut für Experimentelle Dermatologie (LIED), Germany
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3
Technische Universität München, Germany
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4
Heinrich Heine Universität Düsseldorf, Germany
The development of the central nervous system underlies strictly controlled highly complex mechanisms, which makes it particularly vulnerable for toxic chemicals. A small influence on cognitive abilities due to an exposure of the fetal brain to neurotoxic chemicals might have a huge impact on the overall of societal intelligence (Grandjean and Landrigan 2006). Therefore toxicological research for neurotoxicity is necessary and currently done by resource-intensive animal experiments. The extrapolation of animal-derived findings to the actual human hazard is problematic due to species differences (Leist and Hartung 2013). Therefore, testing strategies should be predictive for humans, affordable, sensitive and use mechanism-based methods which also allow medium-throughput screenings. However, obtaining human material from neural tissue is often not possible or bears ethical concerns. Therefore, in our in vitro studies we use human induced pluripotent stem cells (hiPSC) to overcome these restrictions. We have established hiPSC-derived neural progenitor cells, cultured as 3D neurospheres, which further differentiate into spontaneous electrically active neuronal networks consisting of neurons and astrocytes. The electrophysiological activity of in vitro generated neuronal networks is fundamental for the extrapolation of human in vitro data to the human in vivo situation. Here we use either single-well microelectrode arrays (MEA) as well as multiwell-MEA (mwMEA) to measure and analyze spontaneous electrical activities of hiPSC-derived networks after two weeks of differentiation. hiPSC-derived neuronal networks developed from single spikes to bursts indicating maturation over time. However, in contrast to neuronal networks derived from rat neurospheres, hiPSC-derived networks exhibited less electrical activity and synchronicity within 28 days of cultivation time. To improve this network maturation and its activity levels we established a new differentiation medium (CINDA medium) with neuronal maturation-supporting media supplements. This CINDA medium induces the gene and/or protein expression of neuronal receptors, pre- and postsynaptic markers and neuronal subtype-specific markers. Additionally, it increases the electrical activity and emerges synchronicity over 28 days in vitro. The network responses towards pharmacological modulators resulted in mode of action-dependent effects on the neuronal networks.
In summary, we have established hiPSC-derived neuronal networks with spontaneous electrical activity which can be detected and analyzed with the MEA technology, either single well MEA as well as mwMEA. The molecular characterization of the networks regarding neuronal subtypes and the receptor constitutions exhibited maturation over time. These establishments offer the opportunity to establish a screening assay for future toxicity and efficacy testing based on hiPSC-derived neuronal network activities.
References
References:
Grandjean, P., Landrigan, P. J., 2006. Developmental neurotoxicity of industrial chemicals. Lancet 368, 2167-2178.
Leist, M., Hartung, T., 2013. Inflammatory findings on species extrapolations: humans are definitely no 70-kg mice. Arch Toxicol 87, 563-567.
Keywords:
HiPSCs,
neuronal network,
neurotransmission,
microelectrode arrays,
Stem Cells,
in vitro
Conference:
MEA Meeting 2018 | 11th International Meeting on Substrate Integrated Microelectrode Arrays, Reutlingen, Germany, 4 Jul - 6 Jul, 2018.
Presentation Type:
Poster Presentation
Topic:
Neural Networks
Citation:
Nimtz
L,
Hofrichter
M,
Kabiri
Y,
Theiss
S,
Adjaye
J and
Fritsche
E
(2019). Improvements of spontaneous electrical activities of human induced pluripotent stem cell-derived neuronal networks, which can serve as an alternative method to study neurotoxic and pharmacological effects in vitro.
Conference Abstract:
MEA Meeting 2018 | 11th International Meeting on Substrate Integrated Microelectrode Arrays.
doi: 10.3389/conf.fncel.2018.38.00047
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Received:
05 Mar 2018;
Published Online:
17 Jan 2019.
*
Correspondence:
Ms. Laura Nimtz, Leibniz-Institut für umweltmedizinische Forschung (IUF), Dusseldorf, Germany, Laura.Nimtz@gmx.de