Effects of fluorescent glutamate indicators on neurotransmitter diffusion and uptake

Abstract

Genetically encoded fluorescent glutamate indicators (iGluSnFRs) enable neurotransmitter release and diffusion to be visualized in intact tissue. Synaptic iGluSnFR signal time courses vary widely depending on experimental conditions, often lasting 10–100 times longer than the extracellular lifetime of synaptically released glutamate estimated with uptake measurements. iGluSnFR signals typically also decay much more slowly than the unbinding kinetics of the indicator. To resolve these discrepancies, here we have modeled synaptic glutamate diffusion, uptake and iGluSnFR activation to identify factors influencing iGluSnFR signal waveforms. Simulations suggested that iGluSnFR competes with transporters to bind synaptically released glutamate, delaying glutamate uptake. Accordingly, synaptic transporter currents recorded from iGluSnFR-expressing astrocytes in mouse cortex were slower than those in control astrocytes. Simulations also suggested that iGluSnFR reduces free glutamate levels in extrasynaptic spaces, likely limiting extrasynaptic receptor activation. iGluSnFR and lower affinity variants, nonetheless, provide linear indications of vesicle release, underscoring their value for optical quantal analysis.

Release probability increases towards distal dendrites boosting high-frequency signal transfer in the rodent hippocampus

“…This has a simple explanation. In the hippocampal neuropil, released glutamate molecules can travel for up to ~2 μm from the synapse ( Armbruster et al, 2020 ), not necessarily entering other synaptic clefts (see Discussion). Given the density of glutamatergic connections in area CA1 of ~2.0 μm −2 ( Rusakov and Kullmann, 1998 ), individual synapses have >60 neighbours within the 2 μm proximity (i.e.…”

Diverse dynamics of glutamatergic input from sensory neurons underlie heterogeneous responses of olfactory bulb outputs in vivo

“…iGluSnFR expression can prolong the decay of glutamate transients by competing with glutamate transporters 54 , and decay rates are slower (~140 ms) for the high-affinity SF-iGluSnFR.A184S 20,55 . While we did find that decay times skewed longer for ITA responses measured with the high-affinity variant, their distribution largely overlapped with that of the medium-affinity A184V.…”

The vesicular release probability sets the strength of individual Schaffer collateral synapses

“…As iGluSnFR competes with excitatory amino acids transporters (EAATs) for glutamate, it slows down the clearance of bulk glutamate from the extracellular space 37 . However, diffusion of glutamate out of the synaptic cleft occurs at much shorter time scales (<100 µs) than uptake by EAATs.…”