Organelle Optogenetics: Direct Manipulation of Intracellular Ca2+ Dynamics by Light
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+ store...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2018-08-01
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| Series: | Frontiers in Neuroscience |
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| Online Access: | https://www.frontiersin.org/article/10.3389/fnins.2018.00561/full |
| _version_ | 1811321532504342528 |
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| author | Toshifumi Asano Hiroyuki Igarashi Toru Ishizuka Hiromu Yawo Hiromu Yawo |
| author_facet | Toshifumi Asano Hiroyuki Igarashi Toru Ishizuka Hiromu Yawo Hiromu Yawo |
| author_sort | Toshifumi Asano |
| collection | DOAJ |
| description | 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. |
| first_indexed | 2024-04-13T13:19:43Z |
| format | Article |
| id | doaj.art-54eb7eea66f046a892b3fab199ace694 |
| institution | Directory Open Access Journal |
| issn | 1662-453X |
| language | English |
| last_indexed | 2024-04-13T13:19:43Z |
| publishDate | 2018-08-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Neuroscience |
| spelling | doaj.art-54eb7eea66f046a892b3fab199ace6942022-12-22T02:45:22ZengFrontiers Media S.A.Frontiers in Neuroscience1662-453X2018-08-011210.3389/fnins.2018.00561368221Organelle Optogenetics: Direct Manipulation of Intracellular Ca2+ Dynamics by LightToshifumi Asano0Hiroyuki Igarashi1Toru Ishizuka2Hiromu Yawo3Hiromu Yawo4Department of Cell Biology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, JapanDepartment of Physiology and Pharmacology, Tohoku University Graduate School of Medicine, Sendai, JapanDepartment of Developmental Biology and Neuroscience, Tohoku University Graduate School of Life Sciences, Sendai, JapanDepartment of Physiology and Pharmacology, Tohoku University Graduate School of Medicine, Sendai, JapanDepartment of Developmental Biology and Neuroscience, Tohoku University Graduate School of Life Sciences, Sendai, JapanAs 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.https://www.frontiersin.org/article/10.3389/fnins.2018.00561/fullchannelrhodopsinendoplasmic reticulumsarcoplasmic reticulumER/SRmuscleC2C12 |
| spellingShingle | Toshifumi Asano Hiroyuki Igarashi Toru Ishizuka Hiromu Yawo Hiromu Yawo Organelle Optogenetics: Direct Manipulation of Intracellular Ca2+ Dynamics by Light Frontiers in Neuroscience channelrhodopsin endoplasmic reticulum sarcoplasmic reticulum ER/SR muscle C2C12 |
| title | Organelle Optogenetics: Direct Manipulation of Intracellular Ca2+ Dynamics by Light |
| title_full | Organelle Optogenetics: Direct Manipulation of Intracellular Ca2+ Dynamics by Light |
| title_fullStr | Organelle Optogenetics: Direct Manipulation of Intracellular Ca2+ Dynamics by Light |
| title_full_unstemmed | Organelle Optogenetics: Direct Manipulation of Intracellular Ca2+ Dynamics by Light |
| title_short | Organelle Optogenetics: Direct Manipulation of Intracellular Ca2+ Dynamics by Light |
| title_sort | organelle optogenetics direct manipulation of intracellular ca2 dynamics by light |
| topic | channelrhodopsin endoplasmic reticulum sarcoplasmic reticulum ER/SR muscle C2C12 |
| url | https://www.frontiersin.org/article/10.3389/fnins.2018.00561/full |
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