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Assessing the Nanoscale Organization of Excitatory and Inhibitory Synapses Using Recombinant Probes to Visualize Endogenous Synaptic Proteins

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Translational Research Methods in Neurodevelopmental Disorders

Part of the book series: Neuromethods ((NM,volume 185))

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Abstract

Synapses are highly dynamic and complex structures responsible for neuronal communication. The recent progresses in super-resolution imaging technologies unraveled an inherently complex nanorganization of synaptic components. At the single synapse level, nanodomains of key proteins of pre- and postsynaptic terminals are transcellularly aligned in nanocolumns, which turned out to have a significant impact on synaptic function. Perturbations of this dynamic organization might also be involved in neurodevelopmental disorders, such as autism and intellectual disability. Therefore, it is crucial to understand the organization principles of synaptic connections and their dynamic regulation with nanometer precision. Here, we present a detailed protocol that enables to visualize endogenous synaptic proteins both in vitro and in vivo. This method combines the use of transcriptionally regulated fibronectin intrabodies generated with mRNA display (FingRs) against PSD95 and Gephyrin, two major scaffolding proteins of excitatory and inhibitory synapses, with in utero electroporation (IUE), confocal, and stimulated emission depletion (STED) microscopy. This approach may be considered as a method of choice to assess a number of morphometric parameters of synaptic connections up to their nanoscale organization. We believe that this strategy might help to progress our understanding of the molecular mechanisms underlying brain function and dysfunction.

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Acknowledgments

This work was supported by the Italian Ministry of Health (GR-2018-12366478 to M.F.) and the European Union’s Horizon 2020 research and innovation program (Marie Sklodowska-Curie grant agreement 845466 to A.F.).

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Correspondence to Matteo Fossati or Alessandra Folci .

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Fossati, M., Erreni, M., Biagioni, M., Folci, A. (2022). Assessing the Nanoscale Organization of Excitatory and Inhibitory Synapses Using Recombinant Probes to Visualize Endogenous Synaptic Proteins. In: Martin, S., Laumonnier, F. (eds) Translational Research Methods in Neurodevelopmental Disorders. Neuromethods, vol 185. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2569-9_3

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  • DOI: https://doi.org/10.1007/978-1-0716-2569-9_3

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2568-2

  • Online ISBN: 978-1-0716-2569-9

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