Driving Neurogenesis in Neural Stem Cells with High Sensitivity Optogenetics

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.