Pool-independent labelling of synaptic vesicle exocytosis with single vesicle resolution in rat hippocampal neurons.
FM dyes are an established tool to analyze synaptic vesicle pools. However, quantitative measurements using FM dyes are typically based on the re-release properties of previously labelled vesicles, which might vary depending on the experimental setup. An FM dye protocol independent of the previous labelling of vesicle membrane has not been applied for quantitative measurements of individual synaptic vesicles before. We therefore analyzed the direct staining of newly exocytosed vesicle membrane with FM dyes in cultured rat hippocampal neurons. In the presence of FM 1-43, stimulation-induced synaptic activity led to a stable fluorescence increase. The quantal release of synaptic vesicles was preserved and its amplitude correlated highly with the exocytic dye loss induced by a subsequent stimulation. Thus, the method presented here provides a tool for the pool-independent measurement of synaptic vesicle exocytosis.
Department of Psychiatry and Psychotherapy, University of Erlangen-Nuremberg, Schwabachanlage 6, 91054 Erlangen, Germany.
Oliver
O
Welzel
Carsten H
CH
Tischbirek
Johannes
J
Kornhuber
Teja W
TW
Groemer
D000818:Animals; D005089:Exocytosis / Q000502:physiology*; D005456:Fluorescent Dyes; D006624:Hippocampus / Q000378:metabolism* / Q000648:ultrastructure; D007091:Image Processing, Computer-Assisted; D009474:Neurons / Q000378:metabolism* / Q000648:ultrastructure; D051381:Rats; D017208:Rats, Wistar; D013194:Staining and Labeling / Q000379:methods*; D013572:Synaptic Vesicles / Q000378:metabolism* / Q000648:ultrastructure
D005456:Fluorescent Dyes