A genetically engineered fluorescent protein that changes its fluorescence when bound to the neurotransmitter glutamate. [1] Glutamate-sensitive fluorescent reporters (iGluSnFR, colloquially pronounced 'glue sniffer') are used to monitor the activity of presynaptic terminals by fluorescence microscopy. GluSnFRs are a class of optogenetic sensors used in neuroscience research. [2] In brain tissue, two-photon microscopy is typically used to monitor GluSnFR fluorescence.
The widely used iGluSnFR consists of a circularly permuted enhanced green fluorescent protein (cpEGFP) fused to a glutamate binding protein (GluBP) from a bacterium. [3] When GluBP binds a glutamate molecule, it changes its shape, pulling the EGFP barrel together, increasing fluorescence. A specific peptide segment (PDGFR) is included to bring the sensor to the outside of the cell membrane. [4] In the more recent version by Aggarwal et al. (2022), [1] researchers introduced iGluSnFR to two additional anchoring domains, a glycosylphostidylinositol (GPI) anchor, and a modified form of the cytosolic -cterminal domain of Stargazin with a PDZ ligand.
The first genetically encoded fluorescent glutamate sensors (FLIPE, GluSnFR and SuperGluSnFR) were constructed by attaching cyan-fluorescent protein (CFP) and yellow-fluorescent protein (YFP) to a bacterial glutamate binding protein (GluBP). [5] [6] Glutamate binding changed the distance between CFP and YFP, changing the efficiency of energy transfer (FRET) between the two fluorophores. [7] [8] A breakthrough in visualizing glutamate release was achieved with iGluSnFR, a single-fluorophore glutamate sensor based on EGFP producing a ~5‑fold increase in fluorescence. [3] To measure synaptic transmission at high frequencies, novel iGluSnFR variants with accelerated kinetics have recently been developed. [9] [10]