Use of FM1‐43 and Other Derivatives to Investigate Neuronal Function

Michael A. Cousin1

1 University of Edinburgh, Edinburgh, Scotland
Publication Name:  Current Protocols in Neuroscience
Unit Number:  Unit 2.6
DOI:  10.1002/0471142301.ns0206s43
Online Posting Date:  April, 2008
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Abstract

The fluorescent dye FM1‐43 and its derivatives can be used to monitor the physiology of synaptic vesicle turnover in central nerve terminals. They do so by their ability to reversibly partition into membranes, a process that results in a huge increase in fluorescence in comparison to their quantum yield in solution. This unit provides protocols for quantifying total synaptic vesicle turnover, the kinetics and extent of synaptic vesicle exocytosis, and the kinetics and mode of synaptic vesicle endocytosis. Descriptions of other ways these protocols have been used to derive information about the life cycle of the synaptic vesicle are also provided. Curr. Protoc. Neurosci. 43:2.6.1‐2.6.12. © 2008 by John Wiley & Sons, Inc.

Keywords: FM1‐43; synaptic vesicle; exocytosis; endocytosis; fluorescence; FM2‐10; FM4‐64

     
 
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Table of Contents

  • Introduction
  • Basic Protocol 1: Visualization and Quantification of Synaptic Vesicle Exocytosis
  • Alternate Protocol 1: Visualization and Quantification of Synaptic Vesicle Endocytosis
  • Reagents and Solutions
  • Commentary
  • Literature Cited
  • Figures
  • Tables
     
 
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Materials

Basic Protocol 1: Visualization and Quantification of Synaptic Vesicle Exocytosis

  Materials
  • Cultured neurons grown on glass coverslips (poly‐lysine‐coated or otherwise; e.g., Chapter 3)
  • Saline+ solution (see recipe), room temperature
  • 10 µM FM1‐43 (Molecular Probes) in saline+ solution (see recipe for saline+ solution)
  • 10 µM FM1‐43 in saline+ solution (see recipe for saline+ solution) supplemented with 50 to 100 mM KCl (reduce NaCl accordingly to maintain osmolarity)
  • Perfusion chamber with parallel platinum electrodes (RC‐21 BRFS, Warner Instrument) and perfusion apparatus (VC‐66CS, Warner Instrument)
  • Inverted epi‐fluorescence microscope with attached imaging system: light source (monochromator or filter wheel), cooled CCD camera, computer, and imaging software (see unit 2.1)
  • Stimulator (D330‐Multistim System, Digitimer Ltd.)
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Figures

Videos

Literature Cited

Literature Cited
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