An optimized fluorescent probe for visualizing glutamate neurotransmission

Nat Methods. 2013 Feb;10(2):162-70. doi: 10.1038/nmeth.2333. Epub 2013 Jan 13.


We describe an intensity-based glutamate-sensing fluorescent reporter (iGluSnFR) with signal-to-noise ratio and kinetics appropriate for in vivo imaging. We engineered iGluSnFR in vitro to maximize its fluorescence change, and we validated its utility for visualizing glutamate release by neurons and astrocytes in increasingly intact neurological systems. In hippocampal culture, iGluSnFR detected single field stimulus-evoked glutamate release events. In pyramidal neurons in acute brain slices, glutamate uncaging at single spines showed that iGluSnFR responds robustly and specifically to glutamate in situ, and responses correlate with voltage changes. In mouse retina, iGluSnFR-expressing neurons showed intact light-evoked excitatory currents, and the sensor revealed tonic glutamate signaling in response to light stimuli. In worms, glutamate signals preceded and predicted postsynaptic calcium transients. In zebrafish, iGluSnFR revealed spatial organization of direction-selective synaptic activity in the optic tectum. Finally, in mouse forelimb motor cortex, iGluSnFR expression in layer V pyramidal neurons revealed task-dependent single-spine activity during running.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Astrocytes / metabolism
  • Biosensing Techniques
  • Caenorhabditis elegans
  • Calcium Signaling / physiology
  • Escherichia coli Proteins* / chemical synthesis
  • Excitatory Postsynaptic Potentials / physiology
  • Fluorescent Dyes* / chemical synthesis
  • Fluorescent Dyes* / metabolism
  • Glutamic Acid / metabolism*
  • Green Fluorescent Proteins* / chemical synthesis
  • Hippocampus / metabolism
  • Mice
  • Motor Cortex / metabolism
  • Neurons / metabolism
  • Photic Stimulation
  • Pyramidal Cells / metabolism
  • Recombinant Fusion Proteins* / chemical synthesis
  • Retina / physiology
  • Signal-To-Noise Ratio
  • Synaptic Transmission / physiology*
  • Zebrafish


  • Escherichia coli Proteins
  • Fluorescent Dyes
  • Recombinant Fusion Proteins
  • iGluSnFR protein
  • Green Fluorescent Proteins
  • Glutamic Acid