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Sci Rep. 2017 Aug 4;7(1):7351. doi: 10.1038/s41598-017-07820-6.

Red fluorescent protein-based cAMP indicator applicable to optogenetics and in vivo imaging.

Author information

1
Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, 3-8-1 Komaba, Meguro, Tokyo, 153-8902, Japan.
2
Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo, 113-0033, Japan.
3
Laboratory for Neuron-Glia Circuitry, RIKEN Brain Science Institute, Hirosawa 2-1, Wako-shi, Saitama, 351-0198, Japan.
4
Cell Signaling Group, WASEDA Bioscience Research Institute in Singapore (WABIOS), 11 Biopolis Way, #05-02 Helios, Singapore, 138667, Singapore.
5
Department of Neurophysiology and Neural Repair, Gunma University Graduate School of Medicine, Maebashi, Gunma, 371-8511, Japan.
6
Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, 3-8-1 Komaba, Meguro, Tokyo, 153-8902, Japan. takatsuboi@bio.c.u-tokyo.ac.jp.
7
Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo, 113-0033, Japan. takatsuboi@bio.c.u-tokyo.ac.jp.
8
Cell Signaling Group, WASEDA Bioscience Research Institute in Singapore (WABIOS), 11 Biopolis Way, #05-02 Helios, Singapore, 138667, Singapore. kitaguct-gfp@umin.ac.jp.
9
Comprehensive Research Organization, Waseda University, #304, Block 120-4, 513 Wasedatsurumaki-cho, Shinjuku, Tokyo, 162-0041, Japan. kitaguct-gfp@umin.ac.jp.

Abstract

cAMP is a common second messenger that is involved in various physiological processes. To expand the colour palette of available cAMP indicators, we developed a red cAMP indicator named "Pink Flamindo" (Pink Fluorescent cAMP indicator). The fluorescence intensity of Pink Flamindo increases 4.2-fold in the presence of a saturating dose of cAMP, with excitation and emission peaks at 567 nm and 590 nm, respectively. Live-cell imaging revealed that Pink Flamindo is effective for monitoring the spatio-temporal dynamics of intracellular cAMP generated by photoactivated adenylyl cyclase in response to blue light, and in dual-colour imaging studies using a green Ca2+ indicator (G-GECO). Furthermore, we successfully monitored the elevation of cAMP levels in vivo in cerebral cortical astrocytes by two-photon imaging. We propose that Pink Flamindo will facilitate future in vivo, optogenetic studies of cell signalling and cAMP dynamics.

PMID:
28779099
PMCID:
PMC5544736
DOI:
10.1038/s41598-017-07820-6
[Indexed for MEDLINE]
Free PMC Article

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