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Monitoring hippocampal glycine with the computationally designed optical sensor GlyFS

Nature Chemical Biology volume 14, pages 861–869 (2018)Cite this article

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Abstract

Fluorescent sensors are an essential part of the experimental toolbox of the life sciences, where they are used ubiquitously to visualize intra- and extracellular signaling. In the brain, optical neurotransmitter sensors can shed light on temporal and spatial aspects of signal transmission by directly observing, for instance, neurotransmitter release and spread. Here we report the development and application of the first optical sensor for the amino acid glycine, which is both an inhibitory neurotransmitter and a co-agonist of the N-methyl-d-aspartate receptors (NMDARs) involved in synaptic plasticity. Computational design of a glycine-specific binding protein allowed us to produce the optical glycine FRET sensor (GlyFS), which can be used with single and two-photon excitation fluorescence microscopy. We took advantage of this newly developed sensor to test predictions about the uneven spatial distribution of glycine in extracellular space and to demonstrate that extracellular glycine levels are controlled by plasticity-inducing stimuli.

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Fig. 1: Design of the optical glycine FRET sensor (GlyFS).
Fig. 2: Characterization of GlyFS using two-photon excitation (2PE) fluorescence microscopy (λ2PE = 800 nm).
Fig. 3: Measuring extracellular glycine levels using GlyFS in hippocampal tissue.
Fig. 4: Extracellular glycine levels reported by GlyFS are lower at dendritic spines.
Fig. 5: GlyFS identifies activity patterns that control extracellular glycine levels.

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Acknowledgements

We thank Dr. O'Mara (Australian National University) for helpful discussions. Research was funded by the Human Frontiers Science Program Young Investigator Award (HFSP to H.J., C.H., and C.J.J.; grant number: RGY0084/2012), German Academic Exchange Service (DAAD-Go8) Travel Fellowship (to C.H. and C.J.J.), NRW-Rückkehrerprogramm (to C.H.), the European Union (ITN EU-GliaPhD) and German Research Foundation (DFG, SFB1089 B03, SPP1757 HE6949/1 and HE6949/3, to C.H.).

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  1. These authors contributed equally: William H. Zhang, Michel K. Herde.

Authors and Affiliations

  1. Research School of Chemistry, Australian National University, Canberra, Australia

    William H. Zhang, Joshua A. Mitchell, Jason H. Whitfield, Vanessa Vongsouthi & Colin J. Jackson

  2. Institute of Cellular Neurosciences, University of Bonn Medical School, Bonn, Germany

    Michel K. Herde, Andreas B. Wulff, Daniel Minge, Björn Breithausen & Christian Henneberger

  3. Institute of Science and Technology Austria (IST Austria), Klosterneuburg, Austria

    Inmaculada Sanchez-Romero & Harald Janovjak

  4. Institute of Neuropathology, University of Bonn Medical School, Bonn, Germany

    Polina E. Gulakova & Susanne Schoch

  5. Australian Regenerative Medicine Institute (ARMI), Faculty of Medicine, Nursing and Health Sciences, Monash University, Melbourne, Australia

    Harald Janovjak

  6. European Molecular Biology Laboratory Australia (EMBL Australia), Monash University, Melbourne, Australia

    Harald Janovjak

  7. German Center for Neurodegenerative Diseases (DZNE), Bonn, Germany

    Christian Henneberger

  8. UCL Institute of Neurology, London, UK

    Christian Henneberger

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Contributions

W.H.Z., J.A.M., V.V., J.H.W. and C.J.J. designed, produced and analyzed the sensor. M.K.H., J.H.W., A.B.W., W.H.Z., D.M., B.B. and C.H. performed and analyzed all experiments using two-photon excitation and electrophysiology in acute brain slices. M.K.H., J.H.W., I.S.-R., P.E.G., H.J., S.S. and A.B.W. performed studies on GlyFS expressed by cells. C.H., C.J.J. and H.J. designed the study. C.H., C.J.J. and W.H.Z. wrote the initial manuscript, to which all authors subsequently contributed.

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Correspondence to Colin J. Jackson or Christian Henneberger.

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Zhang, W.H., Herde, M.K., Mitchell, J.A. et al. Monitoring hippocampal glycine with the computationally designed optical sensor GlyFS. Nat Chem Biol 14, 861–869 (2018). https://doi.org/10.1038/s41589-018-0108-2

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