Frequency-dependent kinetics and prevalence of kiss-and-run and reuse at hippocampal synapses studied with novel quenching methods.
The kinetics of exo-endocytotic recycling could restrict information transfer at central synapses if neurotransmission were entirely reliant on classical full-collapse fusion. Nonclassical fusion retrieval by kiss-and-run would be kinetically advantageous but remains controversial. We used a hydrophilic quencher, bromophenol blue (BPB), to help detect nonclassical events. Upon stimulation, extracellular BPB entered synaptic vesicles and quenched FM1-43 fluorescence, indicating retention of FM dye beyond first fusion. BPB also quenched fluorescence of VAMP (synaptobrevin-2)-EGFP, thus indicating the timing of first fusion of vesicles in the total recycling pool. Comparison with FM dye destaining revealed that kiss-and-run strongly prevailed over full-collapse fusion at low frequency, giving way to a near-even balance at high frequency. Quickening of kiss-and-run vesicle reuse was also observed at higher frequency in the average single vesicle fluorescence response. Kiss-and-run and reuse could enable hippocampal nerve terminals to conserve scarce vesicular resources when responding to widely varying input patterns.
Harata
Nobutoshi C
NC
Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, California 94305, USA.
Choi
Sukwoo
S
Pyle
Jason L
JL
Aravanis
Alexander M
AM
Tsien
Richard W
RW
D000818:Animals; D001978:Bromphenol Blue; D002478:Cells, Cultured; D004396:Coloring Agents; D004558:Electric Stimulation; D004705:Endocytosis / Q000502:physiology; D005089:Exocytosis / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006624:Hippocampus / Q000502:physiology*; D007700:Kinetics; D009474:Neurons / Q000502:physiology; D017729:Presynaptic Terminals / Q000502:physiology; D011726:Pyridinium Compounds / Q000378:metabolism; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013569:Synapses / Q000502:physiology*; D009435:Synaptic Transmission / Q000502:physiology*; D013696:Temperature
D004396:Coloring Agents; C073804:FM1 43; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds; C485184:enhanced green fluorescent protein; D001978:Bromphenol Blue; D049452:Green Fluorescent Proteins