PT - JOURNAL ARTICLE AU - Miao Jing AU - Peng Zhang AU - Guangfu Wang AU - Huoqing Jiang AU - Lukas Mesik AU - Jiesi Feng AU - Jianzhi Zeng AU - Shaohua Wang AU - Jess Looby AU - Nick A. Guagliardo AU - Linda W. Langma AU - Ju Lu AU - Yi Zuo AU - David A. Talmage AU - Lorna W. Role AU - Paula Q. Barrett AU - Li I. Zhang AU - Minmin Luo AU - Yan Song AU - J. Julius Zhu AU - Yulong Li TI - A genetically-encoded fluorescent acetylcholine indicator AID - 10.1101/311126 DP - 2018 Jan 01 TA - bioRxiv PG - 311126 4099 - http://biorxiv.org/content/early/2018/05/01/311126.short 4100 - http://biorxiv.org/content/early/2018/05/01/311126.full AB - Acetylcholine (ACh) regulates a diverse array of physiological processes throughout the body, yet cholinergic transmission in the majority of tissues/organs remains poorly understood due primarily to the limitations of available ACh-monitoring techniques. We developed a family of G-protein-coupled receptor activation-based ACh sensors (GACh) with sensitivity, specificity, signal-to-noise ratio, kinetics and photostability suitable for monitoring ACh signals in vitro and in vivo. GACh sensors were validated with transfection, viral and/or transgenic expression in a dozen types of neuronal and non-neuronal cells prepared from several animal species. In all preparations, GACh sensors selectively responded to exogenous and/or endogenous ACh with robust fluorescence signals that were captured by epifluorescent, confocal and/or two-photon microscopy. Moreover, analysis of endogenous ACh release revealed firing pattern-dependent release and restricted volume transmission, resolving two long-standing questions about central cholinergic transmission. Thus, GACh sensors provide a user-friendly, broadly applicable toolbox for monitoring cholinergic transmission underlying diverse biological processes.