title: Synaptotagmin-7 Is Essential for Ca2+-Triggered Delayed Asynchronous Release But Not for Ca2+-Dependent Vesicle Priming in Retinal Ribbon Synapses.
abstract: Most synapses release neurotransmitters in two phases: (1) a fast synchronous phase lasting a few milliseconds; and (2) a delayed "asynchronous" phase lasting hundreds of milliseconds. Ca(2+) triggers fast synchronous neurotransmitter release by binding to synaptotagmin-1, synaptotagmin-2, or synaptotagmin-9, but how Ca(2+) triggers delayed asynchronous release has long remained enigmatic. Recent results suggested that consistent with the Ca(2+)-sensor function of synaptotagmin-7 in neuroendocrine exocytosis, synaptotagmin-7 also functions as a Ca(2+) sensor for synaptic vesicle exocytosis but operates during delayed asynchronous release. Puzzlingly, a subsequent study postulated that synaptotagmin-7 is not a Ca(2+) sensor for release but mediates Ca(2+)-dependent vesicle repriming after intense stimulation. To address these issues, we here analyzed synaptic transmission at rod bipolar neuron-AII amacrine cell synapses in acute mouse retina slices as a model system. Using paired recordings, we show that knock-out of synaptotagmin-7 selectively impairs delayed asynchronous release but not fast synchronous release. Delayed asynchronous release was blocked in wild-type synapses by intracellular addition of high concentrations of the slow Ca(2+)-chelator EGTA, but EGTA had no effect in synaptotagmin-7 knock-out neurons because delayed asynchronous release was already impaired. Moreover, direct measurements of vesicle repriming failed to uncover an effect of the synaptotagmin-7 knock-out on vesicle repriming. Our data demonstrate that synaptotagmin-7 is selectively essential for Ca(2+)-dependent delayed asynchronous release in retinal rod bipolar cell synapses, that its function can be blocked by simply introducing a slow Ca(2+) buffer into the cells, and that synaptotagmin-7 is not required for normal vesicle repriming.
  
  
  CONCLUSIONS
  How Ca(2+) triggers delayed asynchronous release has long remained enigmatic. Synaptotagmin-7 has been implicated recently as Ca(2+) sensor in mediating delayed asynchronous release, or vesicle repriming, in cultured neurons. To test the precise function of synaptotagmin-7 in a physiologically important synapse in situ, we have used pair recordings to study the synaptic transmission between retinal rod bipolar cells and AII amacrine cells. Our data demonstrate that the knock-out of synaptotagmin-7 selectively impaired delayed asynchronous release but not synchronous release. In contrast, the readily releasable vesicles after depletion recover normally in knock-out mice. Therefore, our findings extend our knowledge of synaptotagmins as Ca(2+) sensors in vesicle fusion and support the idea that synapses are governed universally by different synaptotagmin Ca(2+) sensors mediating distinct release.
authors.affiliation: Department of Molecular and Cellular Physiology and Howard Hughes Medical Institute, Stanford University, Stanford, California 94304-5453 fluo@stanford.edu tcs1@stanford.edu.
authors.forename: Fujun
authors.initials: F
authors.lastname: Luo
authors.affiliation: Department of Molecular and Cellular Physiology and Howard Hughes Medical Institute, Stanford University, Stanford, California 94304-5453.
authors.forename: Taulant
authors.initials: T
authors.lastname: Bacaj
authors.affiliation: Department of Molecular and Cellular Physiology and Howard Hughes Medical Institute, Stanford University, Stanford, California 94304-5453 fluo@stanford.edu tcs1@stanford.edu.
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Südhof
mesh_terms: D025042:Amacrine Cells / Q000166:cytology / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D012160:Retina / Q000166:cytology / Q000378:metabolism; D051245:Retinal Bipolar Cells / Q000166:cytology / Q000378:metabolism*; D013569:Synapses / Q000235:genetics / Q000378:metabolism*; D009435:Synaptic Transmission / Q000502:physiology; D013572:Synaptic Vesicles / Q000378:metabolism*; D050857:Synaptotagmins / Q000235:genetics / Q000378:metabolism*
chemical_list: D050857:Synaptotagmins; D002118:Calcium
keywords: active zone; calcium channel; ribbon synapse; vesicle fusion