Endocytosis is a fundamental biological process that couples exocytosis to maintain the homeostasis of the plasma membrane and sustained neurotransmission.Super-resolution microscopy enables optical imaging of exocyto...Endocytosis is a fundamental biological process that couples exocytosis to maintain the homeostasis of the plasma membrane and sustained neurotransmission.Super-resolution microscopy enables optical imaging of exocytosis and endocytosis in live cells and makes an essential contribution to understanding molecular mechanisms of endocytosis in neuronal somata and other types of cells.However,visualization of exo-endocytic events at the single vesicular level in a synapse with optical imaging remains a great challenge to reveal mechanisms governing the synaptic exo-endocytotic coupling.In this protocol,we describe the technical details of stimulated emission depletion(STED)imaging of synaptic endocytosis at the single-vesicle level,from sample preparation and microscopy calibration to data acquisition and analysis.展开更多
基金National Natural Science Foundation of China(32171233,81901308,81974203,32300819,31670843,32000704,21790390,and 21790394)Key Research and Development Program of Shaanxi Province of China(2023-ZDLSF-23)+5 种基金Natural Science Foundation of Shaanxi Province of China(2019JC-07,2020JQ-029,and 2023-JC-QN-0236)Natural Science Foundation of Sichuan Province of China(2020YJ0337 and 2020YJ0378)Sichuan Science and Technology Program(2022YFS0615)China Postdoctoral Science Foundation(2018M640972)Innovation Capability Support Program of Shaanxi Province,China(2021TD-37)The Shaanxi Postdoc Funding(2023BSHTBZZ15,2023BSHYDZZ39 and 2023BSHEDZZ67).
文摘Endocytosis is a fundamental biological process that couples exocytosis to maintain the homeostasis of the plasma membrane and sustained neurotransmission.Super-resolution microscopy enables optical imaging of exocytosis and endocytosis in live cells and makes an essential contribution to understanding molecular mechanisms of endocytosis in neuronal somata and other types of cells.However,visualization of exo-endocytic events at the single vesicular level in a synapse with optical imaging remains a great challenge to reveal mechanisms governing the synaptic exo-endocytotic coupling.In this protocol,we describe the technical details of stimulated emission depletion(STED)imaging of synaptic endocytosis at the single-vesicle level,from sample preparation and microscopy calibration to data acquisition and analysis.
