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Nat Methods. 2015 Jan;12(1):64-70. doi: 10.1038/nmeth.3185. Epub 2014 Nov 24.

Rational design of a high-affinity, fast, red calcium indicator R-CaMP2.

Author information

1
1] Department of Neurochemistry, Graduate School of Medicine, University of Tokyo, Tokyo, Japan. [2] Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency, Tokyo, Japan.
2
Department of Neurophysiology, Graduate School of Medicine, University of Tokyo, Tokyo, Japan.
3
Department of Neurochemistry, Graduate School of Medicine, University of Tokyo, Tokyo, Japan.
4
Brain Science Institute, Saitama University, Saitama, Japan.
5
1] Department of Neurochemistry, Graduate School of Medicine, University of Tokyo, Tokyo, Japan. [2] Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency, Saitama, Japan.
6
1] Department of Neurophysiology, Graduate School of Medicine, University of Tokyo, Tokyo, Japan. [2] Precursory Research for Embryonic Science and Technology (PRESTO), Japan Science and Technology Agency, Saitama, Japan.

Abstract

Fluorescent Ca(2+) reporters are widely used as readouts of neuronal activities. Here we designed R-CaMP2, a high-affinity red genetically encoded calcium indicator (GECI) with a Hill coefficient near 1. Use of the calmodulin-binding sequence of CaMKK-α and CaMKK-β in lieu of an M13 sequence resulted in threefold faster rise and decay times of Ca(2+) transients than R-CaMP1.07. These features allowed resolving single action potentials (APs) and recording fast AP trains up to 20-40 Hz in cortical slices. Somatic and synaptic activities of a cortical neuronal ensemble in vivo were imaged with similar efficacy as with previously reported sensitive green GECIs. Combining green and red GECIs, we successfully achieved dual-color monitoring of neuronal activities of distinct cell types, both in the mouse cortex and in freely moving Caenorhabditis elegans. Dual imaging using R-CaMP2 and green GECIs provides a powerful means to interrogate orthogonal and hierarchical neuronal ensembles in vivo.

PMID:
25419959
DOI:
10.1038/nmeth.3185
[Indexed for MEDLINE]

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