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The emission of light, coupled to exocytosis, can in principle be utilized to monitor the activity of a large number of individual synapses simultaneously. To illustrate this concept, fusion proteins of Cypridina luciferase and synaptotagmin-I or VAMP-2/synaptobrevin (which we term "synaptolucins") were expressed in cultured hippocampal neurons with the help of viral vectors. Synaptolucins were targeted to synaptic vesicles and, upon exocytosis, formed light-emitting complexes with their cognate luciferin, which was added to the extracellular medium. Photon emissions required a depolarizing stimulus, occurred from regions with high synaptic density as ascertained by vital staining of recycling synaptic vesicles, and were sensitive to Ca2+ depletion and clostridial neurotoxins. The method can currently detect exocytosis of the readily releasable pool of synaptic vesicles at a hippocampal synapse, corresponding to about two dozen quanta, but has the potential for greater sensitivity.

Type

Journal article

Journal

Proc Natl Acad Sci U S A

Publication Date

01/04/1997

Volume

94

Pages

3402 - 3407

Keywords

Animals, Calcium-Binding Proteins, Cyprinidae, Light, Luciferases, Membrane Glycoproteins, Membrane Proteins, Molecular Sequence Data, Nerve Net, Nerve Tissue Proteins, PC12 Cells, R-SNARE Proteins, Rats, Rats, Sprague-Dawley, Synapses, Synaptotagmin I, Synaptotagmins