Organelle Optogenetics: Direct Manipulation of Intracellular Ca(2+) Dynamics by Light

As one of the ubiquitous second messengers, the intracellular Ca(2+), has been revealed to be a pivotal regulator of various cellular functions. Two major sources are involved in the initiation of Ca(2+)-dependent signals: influx from the extracellular space and release from the intracellular Ca(2+) stores such as the endoplasmic/sarcoplasmic reticulum (ER/SR). To manipulate the Ca(2+) 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 Ca(2+)-permeable, was specifically targeted to the ER/SR. The expression specificity as well as the functional operation of the ER/SR-targeted ChRGR (ChRGR(ER)) was evaluated using mouse skeletal myoblasts (C2C12): (1) the ChRGR(ER) co-localized with the ER-marker KDEL; (2) no membrane current was generated by light under whole-cell clamp of cells expressing ChRGR(ER); (3) an increase of fluorometric Ca(2+) was evoked by the optical stimulation (OS) in the cells expressing ChRGR(ER) in a manner independent on the extracellular Ca(2+) concentration ([Ca(2+)](o)); (4) the ΔF/F(0) was sensitive to the inhibitor of sarco/endoplasmic reticulum Ca(2+)-ATPase (SERCA) and (5) the store-operated Ca(2+) entry (SOCE) was induced by the OS in the ChRGR(ER)-expressing cells. Our organelle optogenetics effectively manipulated the ER/SR to release Ca(2+) from intracellular stores. The use of organelle optogenetics would reveal the neuroscientific significance of intracellular Ca(2+) dynamics under spatiotemporal precision.