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Targeted sensors for glutamatergic neurotransmission
Yuchen Hao1,2, Estelle Toulmé3, Benjamin König1,2
1Institute of Biology, Cellular Biophysics, Humboldt-Universität zu Berlin, Berlin, Germany.
Elife
|January 9, 2023
Summary
Researchers developed new fluorescent sensors (SnFR-γ2 and SnFR-γ8) that precisely visualize excitatory neurotransmitter release at synapses. Postsynaptic targeting improved sensor function and revealed deficits in synaptic transmission.
Area of Science:
- Neuroscience
- Molecular Biology
- Biochemistry
Background:
- Optical reporting of neurotransmitter release is crucial for visualizing excitatory synaptic transmission.
- Genetically-encoded fluorescent glutamate reporters exist but often lack synaptic specificity.
- Targeting reporters to specific subcellular locations, like synapses, enhances signal accuracy.
Purpose of the Study:
- To develop and validate novel fluorescent glutamate reporters targeted to postsynaptic sites.
- To improve the specificity and functional properties of glutamate sensors for studying synaptic transmission.
- To investigate the role of Stargazin in synaptic function and neurotransmission.
Main Methods:
- Fusion of the iGluSnFR reporter with glutamate receptor auxiliary proteins (Stargazin, gamma-8) to create SnFR-γ2 and SnFR-γ8.
- Expression and validation of these chimeras in rat hippocampal cells and mouse neurons.
- Simultaneous optical recording of neurotransmitter release and electrical recording of synaptic currents.
- Analysis of synaptic function under varying calcium concentrations and Stargazin overexpression conditions.
Main Results:
- SnFR-γ2 and SnFR-γ8 were successfully targeted and enriched at postsynaptic sites, reporting spontaneous glutamate release with high spatial precision.
- Quantitative detection of neurotransmitter release at multiple synapses was achieved in autaptic neurons.
- Stargazin overexpression caused a postsynaptic deficit, impairing responses despite normal neurotransmitter release, which was reversible.
- SnFR-γ2 demonstrated linear reporting of quantal parameters and synaptic plasticity, unlike the original iGluSnFR, with significantly reduced signal rundown.
Conclusions:
- Postsynaptic targeting of glutamate reporters like iGluSnFR significantly enhances their specificity and functional performance.
- The novel SnFR-γ2 reporter offers improved linearity and stability for quantifying synaptic transmission parameters.
- Subcellular targeting is critical for the effective development and application of optogenetic tools.
- This study provides an open-source analysis suite for extracting quantal parameters from fluorescence data.
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