Functional Comparison of Neuronal Cells Differentiated from Human Induced Pluripotent Stem Cell–Derived Neural Stem Cells under Different Oxygen and Medium Conditions. (December 2016)

• Record Type: Journal Article
• Authors: Yamazaki, Kazuto; Fukushima, Kazuyuki; Sugawara, Michiko; Tabata, Yoshikuni; Imaizumi, Yoichi; Ishihara, Yasuharu; Ito, Masashi; Tsukahara, Kappei; Kohyama, Jun; Okano, Hideyuki
• Is Part Of: Journal of biomolecular screening. Volume 21:Number 10(2016)
• Page Start: 1054
• Page End: 1064
• Publication Date: 2016-12
• Subjects: human induced pluripotent stem cell–derived neural stem cells -- neural differentiation -- oxygen tension -- medium -- functional neurons; Drugs -- Analysis -- Periodicals; Drugs -- Testing -- Periodicals; Biomolecules -- Analysis -- Periodicals; 572.36
• DOI: 10.1177/1087057116661291
• Languages: English
• ISSNs: 1087-0571

Abstract

Because neurons are difficult to obtain from humans, generating functional neurons from human induced pluripotent stem cells (hiPSCs) is important for establishing physiological or disease-relevant screening systems for drug discovery. To examine the culture conditions leading to efficient differentiation of functional neural cells, we investigated the effects of oxygen stress (2% or 20% O2 ) and differentiation medium (DMEM/F12:Neurobasal-based [DN] or commercial [PhoenixSongs Biologicals; PS]) on the expression of genes related to neural differentiation, glutamate receptor function, and the formation of networks of neurons differentiated from hiPSCs (201B7) via long-term self-renewing neuroepithelial-like stem (lt-NES) cells. Expression of genes related to neural differentiation occurred more quickly in PS and/or 2% O2 than in DN and/or 20% O2, resulting in high responsiveness of neural cells to glutamate, N -methyl-d -aspartate (NMDA), α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA), and ( S )-3, 5-dihydroxyphenylglycine (an agonist for mGluR1/5 ), as revealed by calcium imaging assays. NMDA receptors, AMPA receptors, mGluR1, and mGluR5 were functionally validated by using the specific antagonists MK-801, NBQX, JNJ16259685, and 2-methyl-6-(phenylethynyl)-pyridine, respectively. Multielectrode array analysis showed that spontaneous firing occurred earlier in cells cultured in 2% O2 than in 20% O2 . Optimization of O2 tension and culture medium for neural differentiation of hiPSCs can efficiently generate physiologically relevant cells for screening systems.