title: Imaging Exocytosis of Single Synaptic Vesicles at a Fast CNS Presynaptic Terminal.
abstract: Synaptic vesicles are tethered to the active zone where they are docked/primed so that they can fuse rapidly upon Ca(2+) influx. To directly study these steps at a CNS presynaptic terminal, we used total internal reflection fluorescence (TIRF) microscopy at the live isolated calyx of Held terminal and measured the movements of single synaptic vesicle just beneath the plasma membrane. Only a subset of vesicles within the TIRF field underwent exocytosis. Following exocytosis, new vesicles (newcomers) approached the membrane and refilled the release sites slowly with a time constant of several seconds. Uniform elevation of the intracellular Ca(2+) using flash photolysis elicited an exocytotic burst followed by the sustained component, representing release of the readily releasable vesicles and vesicle replenishment, respectively. Surprisingly, newcomers were not released within a second of high Ca(2+). Instead, already-tethered vesicles became release-ready and mediated the replenishment. Our results reveal an important feature of conventional synapses.
authors.affiliation: Graduate School of Brain Science, Doshisha University, Kyoto 6100394, Japan. Electronic address: mmidorik@mail.doshisha.ac.jp.
authors.forename: Mitsuharu
authors.initials: M
authors.lastname: Midorikawa
authors.affiliation: Graduate School of Brain Science, Doshisha University, Kyoto 6100394, Japan. Electronic address: tsakaba@mail.doshisha.ac.jp.
authors.forename: Takeshi
authors.initials: T
authors.lastname: Sakaba
mesh_terms: D000818:Animals; D002490:Central Nervous System / Q000378:metabolism / Q000648:ultrastructure; D005089:Exocytosis / Q000502:physiology*; D005260:Female; D008297:Male; D008856:Microscopy, Fluorescence / Q000379:methods; D057054:Molecular Imaging / Q000379:methods*; D017729:Presynaptic Terminals / Q000378:metabolism* / Q000648:ultrastructure*; D051381:Rats; D017208:Rats, Wistar; D009435:Synaptic Transmission / Q000502:physiology; D013572:Synaptic Vesicles / Q000378:metabolism* / Q000648:ultrastructure*; D013997:Time Factors