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Live FRET-FLIM Imaging to Study Metabotropic Signaling via the NMDA Receptor

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New Technologies for Glutamate Interaction

Part of the book series: Neuromethods ((NM,volume 207))

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Abstract

Until recently, NMDA receptor (NMDAR) functions have been attributed to its ability to conduct calcium ions. However, growing evidence demonstrates that the NMDAR can induce synaptic depression without ion flux, suggesting that it has a metabotropic function. Our results show that glutamate binding or elevated amounts of beta-amyloid can trigger a conformational change in the NMDAR c-terminal domain. We have shown previously that this movement affects interactions between the NMDAR and signaling molecules, which results in synaptic depression. Here, we describe in detail how to monitor conformational movement in the NMDAR and its interactions with associated signaling molecules using FRET-FLIM live imaging in primary hippocampal neurons. A method to selectively block the NMDAR metabotropic function will also be explained. These approaches could be directly used to test the effect of novel NMDAR binding compounds on the NMDAR intracellular conformation or to study signaling proteins implicated in ion-flux-independent synaptic depression. Moreover, one could adapt these procedures to study any kind of protein-protein interaction and its dynamics in living cells.

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Manikkoth, M., Dore, K. (2024). Live FRET-FLIM Imaging to Study Metabotropic Signaling via the NMDA Receptor. In: Kukley, M. (eds) New Technologies for Glutamate Interaction. Neuromethods, vol 207. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3742-5_4

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  • DOI: https://doi.org/10.1007/978-1-0716-3742-5_4

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3741-8

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