title: Presynaptic capacitance measurements and Ca2+ uncaging reveal submillisecond exocytosis kinetics and characterize the Ca2+ sensitivity of vesicle pool depletion at a fast CNS synapse.
abstract: The intracellular Ca2+ sensitivity of synaptic vesicle fusion is an important determinant of transmitter release probability, but it is unknown for most CNS synapses. We combined whole-cell membrane capacitance measurements and Ca2+ uncaging at the large calyx of Held nerve terminals to determine the Ca2+ sensitivity of synaptic vesicle fusion at a glutamatergic CNS synapse, independent of recording EPSCs. Capacitance increases measured 30-50 msec after elevating the intracellular Ca2+ concentration ([Ca2+]i) by Ca2+ uncaging were half-maximal at approximately 5 microm [Ca2+]i. At 10 microm [Ca2+]i, capacitance increases reached maximal values (256 +/- 125 fF; mean +/- SD), indicating the depletion of an average pool of approximately 4000 readily releasable vesicles. Vesicle pool depletion was confirmed in cross-depletion experiments, in which capacitance responses were measured after Ca2+ uncaging, or after combined stimuli of prolonged presynaptic depolarizations and Ca2+ uncaging. To analyze the Ca2+-dependent rates of vesicle pool depletion, the capacitance rise after Ca2+ uncaging was fitted with single- or double-exponential functions. The fast time constants of double-exponential fits, and the time constants of single-exponential fits were 2-3 msec at 10-15 microm [Ca2+]i and reached submillisecond values at 30 microm [Ca2+]i. These results suggest that three to five readily releasable vesicles can fuse within <1 msec at each active zone of a calyx of Held, given that [Ca2+]i rises sufficiently high. Submillisecond kinetics of exocytosis are reached at significantly lower [Ca2+]i than at ribbon-type sensory synapses previously investigated by capacitance measurements.
authors.affiliation: Arbeitsgruppe Synaptische Dynamik und Modulation and Abteilung Membranbiophysik, Max-Planck-Institut für Biophysikalische Chemie, D-37077 Göttingen, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Wölfel
authors.forename: Ralf
authors.initials: R
authors.lastname: Schneggenburger
mesh_terms: D000085:Acetates / Q000378:metabolism; D000818:Animals; D001306:Auditory Pathways / Q000502:physiology; D002118:Calcium / Q000378:metabolism*; D002490:Central Nervous System / Q000502:physiology; D002614:Chelating Agents / Q000378:metabolism; D025802:Electric Capacitance; D005029:Ethylenediamines / Q000378:metabolism; D019706:Excitatory Postsynaptic Potentials / Q000502:physiology; D005089:Exocytosis / Q000502:physiology*; D066298:In Vitro Techniques; D007700:Kinetics; D018408:Patch-Clamp Techniques; D010782:Photolysis; D017729:Presynaptic Terminals / Q000378:metabolism / Q000502:physiology*; D051381:Rats; D017208:Rats, Wistar; D013569:Synapses / Q000378:metabolism / Q000502:physiology*; D009435:Synaptic Transmission / Q000502:physiology; D013572:Synaptic Vesicles / Q000378:metabolism*; D013997:Time Factors
chemical_list: D000085:Acetates; D002614:Chelating Agents; D005029:Ethylenediamines; C057014:DM-nitrophen; D002118:Calcium