title: Dynamics of volume-averaged intracellular Ca2+in a rat CNS nerve terminal during single and repetitive voltage-clamp depolarizations.
abstract: The intracellular concentration of free calcium ions ([Ca2+]i) in a nerve terminal controls both transmitter release and synaptic plasticity. The rapid triggering of transmitter release depends on the local micro- or nanodomain of highly elevated [Ca2+]iin the vicinity of open voltage-gated Ca2+channels, whereas short-term synaptic plasticity is often controlled by global changes in residual [Ca2+]i, averaged over the whole nerve terminal volume. Here we describe dynamic changes of such global [Ca2+]iin the calyx of Held - a giant mammalian glutamatergic nerve terminal, which is particularly suited for biophysical studies. We provide quantitative data on Ca2+inflow, Ca2+buffering and Ca2+clearance. These data allow us to predict changes in [Ca2+]iin the nerve terminal in response to a wide range of stimulus protocols at high temporal resolution and provide a basis for the modelling of short-term plasticity of glutamatergic synapses.
  
  
  
  Many aspects of short-term synaptic plasticity (STP) are controlled by relatively slow changes in the presynaptic intracellular concentration of free calcium ions ([Ca2+]i) that occur in the time range of a few milliseconds to several seconds. In nerve terminals, [Ca2+]iequilibrates diffusionally during such slow changes, such that the globally measured, residual [Ca2+]ithat persists after the collapse of local domains is often the appropriate parameter governing STP. Here, we study activity-dependent dynamic changes in global [Ca2+]iat the rat calyx of Held nerve terminal in acute brainstem slices using patch-clamp and microfluorimetry. We use low concentrations of a low-affinity Ca2+indicator dye (100 μm Fura-6F) in order not to overwhelm endogenous Ca2+buffers. We first study voltage-clamped terminals, dialysed with pipette solutions containing minimal amounts of Ca2+buffers, to determine Ca2+binding properties of endogenous fixed buffers as well as the mechanisms of Ca2+clearance. Subsequently, we use pipette solutions including 500 μm EGTA to determine the Ca2+binding kinetics of this chelator. We provide a formalism and parameters that allow us to predict [Ca2+]ichanges in calyx nerve terminals in response to a wide range of stimulus protocols. Unexpectedly, the Ca2+affinity of EGTA under the conditions of our measurements was substantially lower (KD= 543 ± 51 nm) than measured in vitro, mainly as a consequence of a higher than previously assumed dissociation rate constant (2.38 ± 0.20 s-1), which we need to postulate in order to model the measured presynaptic [Ca2+]itransients.
authors.affiliation: Emeritus Group Membrane Biophysics, Max Planck Institute for Biophysical Chemistry, 37077, Göttingen, Germany.
authors.forename: Kun-Han
authors.initials: KH
authors.lastname: Lin
authors.affiliation: Emeritus Group Membrane Biophysics, Max Planck Institute for Biophysical Chemistry, 37077, Göttingen, Germany.|Department of Molecular Neurobiology, Max Planck Institute of Experimental Medicine, 37075, Göttingen, Germany.|DFG-Research Centre for Nanoscale Microscopy and Molecular Physiology of the Brain (CNMPB), 37073, Göttingen, Germany.
authors.forename: Holger
authors.initials: H
authors.lastname: Taschenberger
authors.affiliation: Emeritus Group Membrane Biophysics, Max Planck Institute for Biophysical Chemistry, 37077, Göttingen, Germany.|DFG-Research Centre for Nanoscale Microscopy and Molecular Physiology of the Brain (CNMPB), 37073, Göttingen, Germany.
authors.forename: Erwin
authors.identifier: 0000-0002-9758-7922
authors.initials: E
authors.lastname: Neher
mesh_terms: D000818:Animals; D001933:Brain Stem / Q000502:physiology*; D002118:Calcium / Q000502:physiology*; D005260:Female; D066298:In Vitro Techniques; D008297:Male; D009411:Nerve Endings / Q000502:physiology*; D018408:Patch-Clamp Techniques; D017208:Rats, Wistar
chemical_list: D002118:Calcium
keywords: Calyx of Held; EGTA; calcium buffer; calcium current; modelling