Multiplex imaging relates quantal glutamate release to presynaptic Ca2+ homeostasis at multiple synapses in situ

Nat Commun. 2019 Mar 29;10(1):1414. doi: 10.1038/s41467-019-09216-8.


Information processing by brain circuits depends on Ca2+-dependent, stochastic release of the excitatory neurotransmitter glutamate. Whilst optical glutamate sensors have enabled detection of synaptic discharges, understanding presynaptic machinery requires simultaneous readout of glutamate release and nanomolar presynaptic Ca2+ in situ. Here, we find that the fluorescence lifetime of the red-shifted Ca2+ indicator Cal-590 is Ca2+-sensitive in the nanomolar range, and employ it in combination with green glutamate sensors to relate quantal neurotransmission to presynaptic Ca2+ kinetics. Multiplexed imaging of individual and multiple synapses in identified axonal circuits reveals that glutamate release efficacy, but not its short-term plasticity, varies with time-dependent fluctuations in presynaptic resting Ca2+ or spike-evoked Ca2+ entry. Within individual presynaptic boutons, we find no nanoscopic co-localisation of evoked presynaptic Ca2+ entry with the prevalent glutamate release site, suggesting loose coupling between the two. The approach enables a better understanding of release machinery at central synapses.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Axons / metabolism
  • Calcium / metabolism*
  • Glutamic Acid / metabolism*
  • Homeostasis*
  • Imaging, Three-Dimensional*
  • Microscopy, Fluorescence
  • Nanoparticles / chemistry
  • Neuronal Plasticity
  • Photons
  • Presynaptic Terminals / metabolism*
  • Rats, Sprague-Dawley


  • Glutamic Acid
  • Calcium