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High-speed imaging of glutamate release with genetically encoded sensors

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High-speed imaging of glutamate release with genetically encoded sensors. / Dürst, Céline D; Wiegert, J Simon; Helassa, Nordine; Kerruth, Silke; Coates, Catherine; Schulze, Christian; Geeves, Michael A; Török, Katalin; Oertner, Thomas G.

In: NAT PROTOC, Vol. 14, No. 5, 05.2019, p. 1401-1424.

Research output: SCORING: Contribution to journal › SCORING: Journal article › Research › peer-review

Harvard

Dürst, CD, Wiegert, JS, Helassa, N, Kerruth, S, Coates, C, Schulze, C, Geeves, MA, Török, K & Oertner, TG 2019, 'High-speed imaging of glutamate release with genetically encoded sensors', NAT PROTOC, vol. 14, no. 5, pp. 1401-1424. https://doi.org/10.1038/s41596-019-0143-9

APA

Dürst, C. D., Wiegert, J. S., Helassa, N., Kerruth, S., Coates, C., Schulze, C., Geeves, M. A., Török, K., & Oertner, T. G. (2019). High-speed imaging of glutamate release with genetically encoded sensors. NAT PROTOC, 14(5), 1401-1424. https://doi.org/10.1038/s41596-019-0143-9

Vancouver

Dürst CD, Wiegert JS, Helassa N, Kerruth S, Coates C, Schulze C et al. High-speed imaging of glutamate release with genetically encoded sensors. NAT PROTOC. 2019 May;14(5):1401-1424. https://doi.org/10.1038/s41596-019-0143-9

Bibtex

@article{25e980dc93da4ca4b4d574d76748e02b,
title = "High-speed imaging of glutamate release with genetically encoded sensors",
abstract = "The strength of an excitatory synapse depends on its ability to release glutamate and on the density of postsynaptic receptors. Genetically encoded glutamate indicators (GEGIs) allow eavesdropping on synaptic transmission at the level of cleft glutamate to investigate properties of the release machinery in detail. Based on the sensor iGluSnFR, we recently developed accelerated versions of GEGIs that allow investigation of synaptic release during 100-Hz trains. Here, we describe the detailed procedures for design and characterization of fast iGluSnFR variants in vitro, transfection of pyramidal cells in organotypic hippocampal cultures, and imaging of evoked glutamate transients with two-photon laser-scanning microscopy. As the released glutamate spreads from a point source-the fusing vesicle-it is possible to localize the vesicle fusion site with a precision exceeding the optical resolution of the microscope. By using a spiral scan path, the temporal resolution can be increased to 1 kHz to capture the peak amplitude of fast iGluSnFR transients. The typical time frame for these experiments is 30 min per synapse.",
keywords = "Biosensing Techniques/methods, CA3 Region, Hippocampal/cytology, Cells, Cultured, Glutamic Acid/analysis, Humans, Microscopy, Confocal, Molecular Probes/analysis, Optical Imaging, Synaptic Transmission/genetics, Transfection",
author = "D{\"u}rst, {C{\'e}line D} and Wiegert, {J Simon} and Nordine Helassa and Silke Kerruth and Catherine Coates and Christian Schulze and Geeves, {Michael A} and Katalin T{\"o}r{\"o}k and Oertner, {Thomas G}",
year = "2019",
month = may,
doi = "10.1038/s41596-019-0143-9",
language = "English",
volume = "14",
pages = "1401--1424",
journal = "NAT PROTOC",
issn = "1754-2189",
publisher = "NATURE PUBLISHING GROUP",
number = "5",

}

RIS

TY - JOUR

T1 - High-speed imaging of glutamate release with genetically encoded sensors

AU - Dürst, Céline D

AU - Wiegert, J Simon

AU - Helassa, Nordine

AU - Kerruth, Silke

AU - Coates, Catherine

AU - Schulze, Christian

AU - Geeves, Michael A

AU - Török, Katalin

AU - Oertner, Thomas G

PY - 2019/5

Y1 - 2019/5

N2 - The strength of an excitatory synapse depends on its ability to release glutamate and on the density of postsynaptic receptors. Genetically encoded glutamate indicators (GEGIs) allow eavesdropping on synaptic transmission at the level of cleft glutamate to investigate properties of the release machinery in detail. Based on the sensor iGluSnFR, we recently developed accelerated versions of GEGIs that allow investigation of synaptic release during 100-Hz trains. Here, we describe the detailed procedures for design and characterization of fast iGluSnFR variants in vitro, transfection of pyramidal cells in organotypic hippocampal cultures, and imaging of evoked glutamate transients with two-photon laser-scanning microscopy. As the released glutamate spreads from a point source-the fusing vesicle-it is possible to localize the vesicle fusion site with a precision exceeding the optical resolution of the microscope. By using a spiral scan path, the temporal resolution can be increased to 1 kHz to capture the peak amplitude of fast iGluSnFR transients. The typical time frame for these experiments is 30 min per synapse.

AB - The strength of an excitatory synapse depends on its ability to release glutamate and on the density of postsynaptic receptors. Genetically encoded glutamate indicators (GEGIs) allow eavesdropping on synaptic transmission at the level of cleft glutamate to investigate properties of the release machinery in detail. Based on the sensor iGluSnFR, we recently developed accelerated versions of GEGIs that allow investigation of synaptic release during 100-Hz trains. Here, we describe the detailed procedures for design and characterization of fast iGluSnFR variants in vitro, transfection of pyramidal cells in organotypic hippocampal cultures, and imaging of evoked glutamate transients with two-photon laser-scanning microscopy. As the released glutamate spreads from a point source-the fusing vesicle-it is possible to localize the vesicle fusion site with a precision exceeding the optical resolution of the microscope. By using a spiral scan path, the temporal resolution can be increased to 1 kHz to capture the peak amplitude of fast iGluSnFR transients. The typical time frame for these experiments is 30 min per synapse.

KW - Biosensing Techniques/methods

KW - CA3 Region, Hippocampal/cytology

KW - Cells, Cultured

KW - Glutamic Acid/analysis

KW - Humans

KW - Microscopy, Confocal

KW - Molecular Probes/analysis

KW - Optical Imaging

KW - Synaptic Transmission/genetics

KW - Transfection

U2 - 10.1038/s41596-019-0143-9

DO - 10.1038/s41596-019-0143-9

M3 - SCORING: Journal article

C2 - 30988508

VL - 14

SP - 1401

EP - 1424

JO - NAT PROTOC

JF - NAT PROTOC

SN - 1754-2189

IS - 5

ER -