Tracking quantum dot-tagged calcium channels at vertebrate photoreceptor synapses: Retinal slices and dissociated Cells

Aaron J. Mercer, Wallace B Thoreson

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

At synapses in the central nervous system, precisely localized assemblies of presynaptic proteins, neurotransmitter-filled vesicles, and postsynaptic receptors are required to communicate messages between neurons. Our understanding of synaptic function has been significantly advanced using electrophysiological methods, but the dynamic spatial behavior and real-time organization of synapses remains poorly understood. In this unit, we describe a method for labeling individual presynaptic calcium channels with photostable quantum dots for single-particle tracking analysis.We have used this technique to examine the mobility of L-type calcium channels in the presynaptic membrane of rod and cone photoreceptors in the retina. These channels control release of glutamate-filled synaptic vesicles at the ribbon synapses in photoreceptor terminals. This technique offers the advantage of providing a real-time biophysical readout of ion channel mobility and can be manipulated by pharmacological or electrophysiological methods. For example, the combination of electrophysiological and single-particle tracking experiments has revealed that fusion of nearby vesicles influences calcium channel mobility and changes in channel mobility can influence release. These approaches can also be readily adapted to examine membrane proteins in other systems.

Original languageEnglish (US)
Article number2.18
JournalCurrent Protocols in Neuroscience
Issue numberSUPPL.62
DOIs
StatePublished - Jan 1 2013

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Quantum Dots
Vertebrate Photoreceptor Cells
Calcium Channels
Synapses
Spatial Behavior
Retinal Cone Photoreceptor Cells
Retinal Rod Photoreceptor Cells
L-Type Calcium Channels
Synaptic Vesicles
Ion Channels
Neurotransmitter Agents
Retina
Glutamic Acid
Membrane Proteins
Central Nervous System
Pharmacology
Neurons
Membranes
Proteins

Keywords

  • L-type calcium channels
  • Membrane diffusion
  • Ribbon synapse
  • Single particle tracking

ASJC Scopus subject areas

  • Neuroscience(all)

Cite this

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abstract = "At synapses in the central nervous system, precisely localized assemblies of presynaptic proteins, neurotransmitter-filled vesicles, and postsynaptic receptors are required to communicate messages between neurons. Our understanding of synaptic function has been significantly advanced using electrophysiological methods, but the dynamic spatial behavior and real-time organization of synapses remains poorly understood. In this unit, we describe a method for labeling individual presynaptic calcium channels with photostable quantum dots for single-particle tracking analysis.We have used this technique to examine the mobility of L-type calcium channels in the presynaptic membrane of rod and cone photoreceptors in the retina. These channels control release of glutamate-filled synaptic vesicles at the ribbon synapses in photoreceptor terminals. This technique offers the advantage of providing a real-time biophysical readout of ion channel mobility and can be manipulated by pharmacological or electrophysiological methods. For example, the combination of electrophysiological and single-particle tracking experiments has revealed that fusion of nearby vesicles influences calcium channel mobility and changes in channel mobility can influence release. These approaches can also be readily adapted to examine membrane proteins in other systems.",
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