Organelle optogenetics: Direct manipulation of intracellular Ca2+ dynamics by light

Toshifumi Asano, Hiroyuki Igarashi, Toru Ishizuka, Hiromu Yawo

Research output: Contribution to journalArticlepeer-review

6 Citations (Scopus)

Abstract

As one of the ubiquitous second messengers, the intracellular Ca2+, has been revealed to be a pivotal regulator of various cellular functions. Two major sources are involved in the initiation of Ca2+-dependent signals: influx from the extracellular space and release from the intracellular Ca2+ stores such as the endoplasmic/sarcoplasmic reticulum (ER/SR). To manipulate the Ca2+ release from the stores under high spatiotemporal precision, we established a new method termed "organelle optogenetics." That is, one of the light-sensitive cation channels (channelrhodopsin-green receiver, ChRGR), which is Ca2+-permeable, was specifically targeted to the ER/SR. The expression specificity as well as the functional operation of the ER/SR-targeted ChRGR (ChRGRER) was evaluated using mouse skeletal myoblasts (C2C12): (1) the ChRGRER co-localized with the ER-marker KDEL; (2) no membrane current was generated by light under whole-cell clamp of cells expressing ChRGRER; (3) an increase of fluorometric Ca2+ was evoked by the optical stimulation (OS) in the cells expressing ChRGRER in a manner independent on the extracellular Ca2+ concentration ([Ca2+]o); (4) the ΔF/F0 was sensitive to the inhibitor of sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) and (5) the store-operated Ca2+ entry (SOCE) was induced by the OS in the ChRGRER-expressing cells. Our organelle optogenetics effectively manipulated the ER/SR to release Ca2+ from intracellular stores. The use of organelle optogenetics would reveal the neuroscientific significance of intracellular Ca2+ dynamics under spatiotemporal precision.

Original languageEnglish
Article number561
JournalFrontiers in Neuroscience
Volume12
Issue numberAUG
DOIs
Publication statusPublished - 2018 Aug 17

Keywords

  • C2C12
  • Channelrhodopsin
  • ER/SR
  • Endoplasmic reticulum
  • Muscle
  • Sarcoplasmic reticulum
  • Store-operated Ca entry (SOCE)
  • Super-resolution microscopy

ASJC Scopus subject areas

  • Neuroscience(all)

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