Optogenetic stimulation promotes Schwann cell proliferation, differentiation, and myelination in vitro

Kyuhwan Jung,Noo Li Jeon,Sung-Yon Kim,Ji Hye Park,Sung-Rae Cho,Sujin Hyung

doi:10.1038/s41598-019-40173-w

Kyuhwan Jung, Noo Li Jeon + Show 4 more

Open Access

https://doi.org/10.1038/s41598-019-40173-w

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Abstract

Schwann cells (SCs) constitute a crucial element of the peripheral nervous system, by structurally supporting the formation of myelin and conveying vital trophic factors to the nervous system. However, the functions of SCs in developmental and regenerative stages remain unclear. Here, we investigated how optogenetic stimulation (OS) of SCs regulates their development. In SC monoculture, OS substantially enhanced SC proliferation and the number of BrdU+-S100ß+-SCs over time. In addition, OS also markedly promoted the expression of both Krox20 and myelin basic protein (MBP) in SC culture medium containing dBcAMP/NRG1, which induced differentiation. We found that the effects of OS are dependent on the intracellular Ca2+ level. OS induces elevated intracellular Ca2+ levels through the T-type voltage-gated calcium channel (VGCC) and mobilization of Ca2+ from both inositol 1,4,5-trisphosphate (IP3)-sensitive stores and caffeine/ryanodine-sensitive stores. Furthermore, we confirmed that OS significantly increased expression levels of both Krox20 and MBP in SC-motor neuron (MN) coculture, which was notably prevented by pharmacological intervention with Ca2+. Taken together, our results demonstrate that OS of SCs increases the intracellular Ca2+ level and can regulate proliferation, differentiation, and myelination, suggesting that OS of SCs may offer a new approach to the treatment of neurodegenerative disorders.

Highlights

Axon regeneration and remyelination after injury are limited in the central nervous system (CNS) of adult mammals, but recovery capacity is considerably greater in the peripheral nervous system (PNS)
To more accurately measure Schwann cells (SCs) proliferation in the SC monoculture, we evaluated SC proliferation using 5-bromo-2′-deoxyuridine (BrdU), a thymidine analog that is incorporated into the DNA of proliferating SCs when they are in the DNA replication phase (i.e., S-phase)
In the SC monoculture, SC proliferation was substantially enhanced by optogenetic stimulation (OS), and the number of BrdU+-S100ß+-SCs increased over time

Summary

Introduction

Axon regeneration and remyelination after injury are limited in the central nervous system (CNS) of adult mammals, but recovery capacity is considerably greater in the peripheral nervous system (PNS). Optogenetics has gained attention as a promising biological technology, in which genetically modified cells within complex neural tissues are regulated using light of a specific wavelength after introducing light-sensitive microbial opsins, such as channelrhodopsin-2 (ChR-2), halorhodopsin (NpHR), and archaerhodopsin[10,11,12,13]. This revolutionary technique can control the electrical processes of specific types of cells, including cell signaling, modulation, etc.; in particular, it affords unprecedented control of neural activity with high spatiotemporal precision in many types of nervous system investigations. We discuss below how OS affects the differentiation and myelination of SCs in a coculture model of SC and motor neuron (MN) cells

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