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In vivo Imaging of Retrogradely Transported Synaptic Vesicle Proteins in Caenorhabditis elegans Neurons
Author(s) -
Murthy Kausalya,
Bhat Jaffar M.,
Koushika Sandhya P.
Publication year - 2011
Publication title -
traffic
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.677
H-Index - 130
eISSN - 1600-0854
pISSN - 1398-9219
DOI - 10.1111/j.1600-0854.2010.01127.x
Subject(s) - biology , vesicle fusion , synaptobrevin , synaptic vesicle , microbiology and biotechnology , green fluorescent protein , synaptotagmin 1 , snap25 , caenorhabditis elegans , rab , kiss and run fusion , exocytosis , vesicle , live cell imaging , synaptic vesicle recycling , synapse , axoplasmic transport , neuroscience , biochemistry , cell , secretion , gtpase , membrane , gene
Axonal transport is an essential process that carries cargoes in the anterograde direction to the synapse and in the retrograde direction back to the cell body. We have developed a novel in vivo method to exclusively mark and dynamically track retrogradely moving compartments carrying specific endogenous synaptic vesicle proteins in the Caenorhabditis elegans model. Our method is based on the uptake of a fluorescently labeled anti‐green fluorescent protein (GFP) antibody delivered in an animal expressing the synaptic vesicle protein synaptobrevin‐1::GFP in neurons. We show that this method largely labels retrogradely moving compartments. Very little labeling is observed upon blocking vesicle exocytosis or if the synapse is physically separated from the cell body. The extent of labeling is also dependent on the dyenin–dynactin complex. These data support the interpretation that the labeling of synaptobrevin‐1::GFP largely occurs after vesicle fusion and the major labeling likely takes place at the synapse. Further, we observe that the retrograde compartment carrying synaptobrevin contains synaptotagmin but lacks the endosomal marker RAB‐5. This labeling method is very general and can be readily adapted to any transmembrane protein on synaptic vesicles with a GFP tag inside the vesicle and can also be extended to other model systems.