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Biophys. J. BioFAST: First Published August 19, 2005. doi:10.1529/biophysj.105.059295
© 2005 by the Biophysical Society.

A more recent version of this article appeared on November 1, 2005.
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CELL BIOPHYSICS

Probing Vesicle Dynamics in Single Hippocampal Synapses

Matthew Shtrahman 1, Chuck Yeung 2, David W. Nauen 3, Guo-qiang Bi 3 and Xiao-lun Wu 1*

1 University of Pittsburgh
2 The Pennsylvania State University at Erie
3 University of Pittsburgh School of Medicine

* To whom correspondence should be addressed. E-mail: xlwu{at}pitt.edu.

Submitted on January 7, 2005
Revised on February 22, 2005
Accepted on 9 August 2005


  Abstract
We use fluorescence correlation spectroscopy (FCS) and fluorescence recovery after photobleaching (FRAP) to study vesicle dynamics inside the synapses of cultured hippocampal neurons labeled with the fluorescent vesicle marker FM 1-43. These studies show that when the cell is electrically at rest, only a small population of vesicles is mobile, taking seconds to traverse the synapse. Applying the phosphatase inhibitor okadaic acid (OA) causes vesicles to diffuse freely, moving 30 times faster than vesicles in control synapses. These results suggest that vesicles move sluggishly due to binding to elements of the synaptic cytomatrix, and that this binding is altered by phosphorylation. Motivated by these results, a model is constructed consisting of diffusing vesicles that bind reversibly to the cytomatrix. This stick-and-diffuse model accounts for the FCS and FRAP data, and also predicts the well-known exponential refilling of the readily releasable pool. Our measurements suggest that the movement of vesicles to the active zone is the rate limiting step in this process.

Key Words: fluorescence correlation spectroscopy (FCS), fluorescence recovery after photobleaching (FRAP), readily releasable pool, refilling, synapsin, synaptic depression




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Copyright © 2005 by the Biophysical Society.