Functional electron microscopy (“Flash and Freeze”) of identified cortical synapses in acute brain slices

C. Borges Merjane, O. Kim, P.M. Jonas, Neuron 105 (2020) 992–1006.

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Abstract
How structural and functional properties of synapses relate to each other is a fundamental question in neuroscience. Electrophysiology has elucidated mechanisms of synaptic transmission, and electron microscopy (EM) has provided insight into morphological properties of synapses. Here we describe an enhanced method for functional EM (“flash and freeze”), combining optogenetic stimulation with high-pressure freezing. We demonstrate that the improved method can be applied to intact networks in acute brain slices and organotypic slice cultures from mice. As a proof of concept, we probed vesicle pool changes during synaptic transmission at the hippocampal mossy fiber-CA3 pyramidal neuron synapse. Our findings show overlap of the docked vesicle pool and the functionally defined readily releasable pool and provide evidence of fast endocytosis at this synapse. Functional EM with acute slices and slice cultures has the potential to reveal the structural and functional mechanisms of transmission in intact, genetically perturbed, and disease-affected synapses.
Publishing Year
Date Published
2020-03-18
Journal Title
Neuron
Acknowledgement
This project has received funding from the European Research Council (ERC) and European Commission (EC), under the European Union’s Horizon 2020 research and innovation programme (ERC grant agreement No. 692692 and Marie Sklodowska-Curie 708497) and from Fonds zur Förderung der Wissenschaftlichen Forschung (Z 312-B27 Wittgenstein award and DK W1205-B09). We thank Johann Danzl and Ryuichi Shigemoto for critically reading the manuscript; Walter Kaufmann, Daniel Gutl, and Vanessa Zheden for extensive EM training, advice, and experimental assistance; Benjamin Suter for substantial help with light stimulation, ImageJ plugins for analysis, and manuscript editing; Florian Marr and Christina Altmutter for technical support; Eleftheria Kralli-Beller for manuscript editing; Julia König and Paul Wurzinger (Leica Microsystems) for helpful technical discussions; and Taija Makinen for providing the Prox1-CreERT2 mouse line.
Volume
105
Page
992-1006
ISSN
IST-REx-ID

Cite this

Borges Merjane C, Kim O, Jonas PM. Functional electron microscopy (“Flash and Freeze”) of identified cortical synapses in acute brain slices. Neuron. 2020;105:992-1006. doi:10.1016/j.neuron.2019.12.022
Borges Merjane, C., Kim, O., & Jonas, P. M. (2020). Functional electron microscopy (“Flash and Freeze”) of identified cortical synapses in acute brain slices. Neuron. Elsevier. https://doi.org/10.1016/j.neuron.2019.12.022
Borges Merjane, Carolina, Olena Kim, and Peter M Jonas. “Functional Electron Microscopy (‘Flash and Freeze’) of Identified Cortical Synapses in Acute Brain Slices.” Neuron. Elsevier, 2020. https://doi.org/10.1016/j.neuron.2019.12.022.
C. Borges Merjane, O. Kim, and P. M. Jonas, “Functional electron microscopy (‘Flash and Freeze’) of identified cortical synapses in acute brain slices,” Neuron, vol. 105. Elsevier, pp. 992–1006, 2020.
Borges Merjane C, Kim O, Jonas PM. 2020. Functional electron microscopy (“Flash and Freeze”) of identified cortical synapses in acute brain slices. Neuron. 105, 992–1006.
Borges Merjane, Carolina, et al. “Functional Electron Microscopy (‘Flash and Freeze’) of Identified Cortical Synapses in Acute Brain Slices.” Neuron, vol. 105, Elsevier, 2020, pp. 992–1006, doi:10.1016/j.neuron.2019.12.022.
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