Driving Neurogenesis in Neural Stem Cells with High Sensitivity Optogenetics

Abstract

Optogenetic stimulation of neural stem cells (NSCs) enables their activity-dependent photo-modulation. This provides a spatio-temporal tool for studying activity-dependent neurogenesis and for regulating the differentiation of the transplanted NSCs. Currently, this is mainly driven by viral transfection of channelrhodopsin-2 (ChR2) gene, which requires high irradiance and complex in vivo/vitro stimulation systems. Additionally, despite the extensive application of optogenetics in neuroscience, the transcriptome-level changes induced by optogenetic stimulation of NSCs have not been elucidated yet. Here, we made transformed NSCs (SFO-NSCs) stably expressing one of the step-function opsin (SFO)-variants of chimeric channelrhodopsins, ChRFR(C167A), which is more sensitive to blue light than native ChR2, via a non-viral transfection system using piggyBac transposon. We set up a simple low-irradiance optical stimulation (OS)-incubation system that induced c-fos mRNA expression, which is activity-dependent, in differentiating SFO-NSCs. More neuron-like SFO-NCSs, which had more elongated axons, were differentiated with daily OS than control cells without OS. This was accompanied by positive/negative changes in the transcriptome involved in axonal remodeling, synaptic plasticity, and microenvironment modulation with the up-regulation of several genes involved in the Ca2+-related functions. Our approach could be applied for stem cell transplantation studies in tissue with two strengths: lower carcinogenicity and less irradiance needed for tissue penetration.

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Acknowledgements

This work is supported by R-175-000-137-112 (Singapore Ministry of Education Tier 1), R-175-000-136-305 (Singapore Agency for Science, Technology and Research A*STAR - Japan Science and Technology Agency JST), Grants-in-Aid for Scientific Research (KAKENHI) from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan (25670103 to HY), and JST, Strategic International Collaborative Research Program, SICORP (15657509A to HY). We thank Jun Takahashi for his generous gift of PZ5 cells, Lee Shu Ying from the Confocal Microscopy Unit, Yong Hui Yee for assistance in microscopy and transcriptome analysis, and Brent Bell for language assistance.

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Experiment design: DT, HY, TI. Supervision: HY, TI, NT, LX, AA, LW. Conducted experiment: DT, AP, WJ, NZ, YY, YW, HS, LW. Analyzed Data: DT, LW, TI, HY. Manuscript writing and revision: All authors.

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Correspondence to Daniel Boon Loong Teh or Hiromu Yawo or Angelo All.

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R-175-000-137-112 (Singapore Ministry of Education Tier 1) and R-175-000-136-305 (Singapore Agency for Science, Technology and Research A*STAR - Japan Science and Technology Agency JST). Other authors declare that they have no conflicts of interest.

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Teh, D.B.L., Prasad, A., Jiang, W. et al. Driving Neurogenesis in Neural Stem Cells with High Sensitivity Optogenetics. Neuromol Med 22, 139–149 (2020). https://doi.org/10.1007/s12017-019-08573-3

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Keywords

  • Neural stem cells
  • Optogenetics stimulation
  • Neurogenesis
  • Microarray genomic