biostudies-literatureKagiampaki ZSwiss National Science FoundationEuropean Research CouncilNINDS NIH HHS1426-1436https://www.ebi.ac.uk/biostudies/studies/S-EPMC7615053biostudies-literatureUnknown20(9)Genetically encoded indicators engineered from G-protein-coupled receptors are important tools that enable high-resolution in vivo neuromodulator imaging. Here, we introduce a family of sensitive multicolor norepinephrine (NE) indicators, which includes nLightG (green) and nLightR (red). These tools report endogenous NE release in vitro, ex vivo and in vivo with improved sensitivity, ligand selectivity and kinetics, as well as a distinct pharmacological profile compared with previous state-of-the-art GRAB_NE indicators. Using in vivo multisite fiber photometry recordings of nLightG, we could simultaneously monitor optogenetically evoked NE release in the mouse locus coeruleus and hippocampus. Two-photon imaging of nLightG revealed locomotion and reward-related NE transients in thNature methodsSensitive multicolor indicators for monitoring norepinephrine in vivo.PMC7615053U19 NS107464891959310030196455Curreli SWiegert JSBhat MASonmez LReiner ADieter ADernic JRavotto LWasielewski LMHarada MZhou XFellin TRohner VWeber BDuss SNKiss CZiebarth TBohacek JWilhelm MBenke DPatriarchi TKagiampaki ZfalseSensitive multicolor indicators for monitoring norepinephrine in vivo.Genetically encoded indicators engineered from G-protein-coupled receptors are important tools that enable high-resolution in vivo neuromodulator imaging. Here, we introduce a family of sensitive multicolor norepinephrine (NE) indicators, which includes nLightG (green) and nLightR (red). These tools report endogenous NE release in vitro, ex vivo and in vivo with improved sensitivity, ligand selectivity and kinetics, as well as a distinct pharmacological profile compared with previous state-of-the-art GRAB_NE indicators. Using in vivo multisite fiber photometry recordings of nLightG, we could simultaneously monitor optogenetically evoked NE release in the mouse locus coeruleus and hippocampus. Two-photon imaging of nLightG revealed locomotion and reward-related NE transients in th2023-01-01T00:00:00Z2023 Sep2025-04-04T02:26:29.159Z2025-02-19T01:42:07.143ZS-EPMC76150533747480710.1038/s41592-023-01959-z