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Real-time visualization of neuronal activity in zebrafish and C. elegans

›’†ˆä ~ˆê / Junichi Nakai:1@‘å‘q ³“¹ / Masamichi Ohkura:1,2@‰i‘º ‚䂤Žq / Yuko Kagawa-Nagamura:1,2@•“¡ Ê / Akira Muto:3@ˆäã ¹r / Masatoshi Inoue:4@”ö“¡ °•F / Haruhiko Bito:4@ìã _ˆê / Koichi Kawakami:3@ˆÀ“¡ ŒbŽq / Keiko Gengyo-Ando:1,2@ 1:é‹Ê‘嗝HŒ¤ / Grad Sch Sci Eng, Saitama Univ, Saitama Japan@2:é‹Ê‘å”]ƒZ / Saitama Univ Brain Sci Inst, Saitama Univ, Saitama, Japan@3:ˆâ“`Œ¤‰Šú”­¶ / Div Molec Dev Biol, NIG, Mishima, Japan@4:“Œ‘åˆã_Œo¶‰» / Dept Neurochem, Grad Sch Med, Univ of Tokyo, Tokyo, Japan

To monitor neuronal activities calcium imaging is one of the promising methods. We have been developing genetically encoded calcium indicators (GECIs) called G-CaMPs (green GECIs) and R-CaMPs (red GECIs). Because G-CaMPs and R-CaMPs are genetically encoded and their expression can be spatially and temporally controlled, G-CaMPs and R-CaMPs are particularly useful to monitor neuron- and glial-activities in vivo. To monitor neural activity in zebrafish, we expressed G-CaMP7a in the tectum, which is the visual center in the zebrafish brain, and performed calcium imaging under a fluorescent microscope. When a prey paramecium swam around a zebrafish larva expressing G-CaMP7a in the tectum, we could detect fluorescent increases in cell bodies and dendrites. We could functionally map the tectal-neuron activities according to the position of the paramecium. Recently we developed new R-CaMP called R-CaMP2. Using the calmodulin-binding sequence of CaMKK-ƒ¿ and CaMKK-ƒÀ in lieu of an M13 sequence resulted in high affinity and threefold-faster kinetics of Ca2+ transients than the parental probe R-CaMP1.07. We expressed R-CaMP2 in C. elegans neurons and/or the body wall muscles (BWMs) and imaged their activities with a confocal microscope. We could monitor neuronal or musclular activities from freely-moving C. elegans. Next we generated transgenic C. elegans that expressed channelrhodopsin 2 and G-CaMP6 in GABA neurons and R-CaMP2 in the BMWs. We successfully achieved dual-color monitoring of neuronal and musclular activities in response to photostimulation.