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This book provides protocols on the preparation of synaptic vesicles from different model systems and provides instructions for functional investigations of their transport and formation, scaffolding, filling and priming, release, recycling, and degradation. Chapters guide readers through computational approaches, simulating different aspects of synaptic vesicle anatomy and function. Written in the format of the highly successful Methods in Molecular Biology series, each chapter includes an introduction to the topic, lists necessary materials and reagents, includes tips on troubleshooting and known pitfalls, and step-by-step, readily reproducible protocols.
Authoritative and cutting-edge, Synaptic Vesicles: Methods and Protocols aims to be a useful practical guide to researchers to help further their study in this field.
List of contents
Real-time quantitative PCR and fluorescence in situ hybridization for subcellular localization of miRNAs in neurons.- Live-imaging of axonal cargoes in Drosophila brain explants using confocal microscopy.- Whole Endosome Recording of Vesicular Neurotransmitter Transporter Currents.- Quantitative analysis of presynaptic vesicle luminal pH in cultured neurons.- Investigation of synaptic vesicle proteins in rat brain tissue using real-time qPCR.- Mass Synaptometry: Applying Mass Cytometry to Single Synapse Analysis.- A Guide to Analysis of Relative Synaptic Protein Abundance by Quantitative Fluorescent Western Blotting.- Synaptosomes and metamodulation of receptors.- A novel method to monitor GABA loading into synaptic vesicles by combining patch pipette perfusion and intracellular, caged-GABA photolysis in brain slice preparations.- Rapid Isolation of Functional Synaptic Vesicles from Tissues through Cryo-grinding, Ultracentrifugation and Size Exclusion Chromatography.- Isolation of synaptic vesicles from mammalian brain.- Building and Using a Two-Photon Fluorescence Crosscorrelation Spectroscopy Setup including Fluorescence Lifetime Analysis.- Fluorescence Lifetime and Cross-correlation Spectroscopy for Observing Membrane Fusion of Liposome Models Containing Synaptic Proteins.- Synaptic vesicle pool monitoring with synaptopH-Iuorin.- Imaging neuropeptide release at Drosophila neuromuscular junction with a genetically engineered neuropeptide release reporter.- Imaging synaptic glutamate release with two-photon microscopy in organotypic slice cultures.- Dynole 34-2 and Acrylo-Dyn 2-30, novel dynamin GTPase chemical biology probes.- Synthesis of Pthaladyn-29 and Napthalimide-10, GTP site directed dynamin GTPase inhibitors.
