title: Tools and limitations to study the molecular composition of synapses by fluorescence microscopy.
abstract: The synapse is densely packed with proteins involved in various highly regulated processes. Synaptic protein copy numbers and their stoichiometric distribution have a drastic influence on neuronal integrity and function. Therefore, the molecular analysis of synapses is a key element to understand their architecture and function. The overall structure of the synapse has been revealed with an exquisite amount of details by electron microscopy. However, the molecular composition and the localization of proteins are more easily addressed with fluorescence imaging, especially with the improved resolution achieved by super-resolution microscopy techniques. Notably, the fast improvement of imaging instruments has not been reflected in the optimization of biological sample preparation. During recent years, large efforts have been made to generate affinity probes smaller than conventional antibodies adapted for fluorescent super-resolution imaging. In this review, we briefly discuss the current views on synaptic organization and necessary key technologies to progress in the understanding of synaptic physiology. We also highlight the challenges faced by current fluorescent super-resolution methods, and we describe the prerequisites for an ideal study of synaptic organization.
authors.affiliation: Department of Neuro- and Sensory Physiology, University of Göttingen Medical Center, Göttingen, Germany Center for Biostructural Imaging of Neurodegeneration, University of Göttingen Medical Center, Göttingen, Germany.
authors.forename: Manuel
authors.initials: M
authors.lastname: Maidorn
authors.affiliation: Department of Neuro- and Sensory Physiology, University of Göttingen Medical Center, Göttingen, Germany Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, Göttingen, Germany.
authors.forename: Silvio O
authors.initials: SO
authors.lastname: Rizzoli
authors.affiliation: Department of Neuro- and Sensory Physiology, University of Göttingen Medical Center, Göttingen, Germany Center for Biostructural Imaging of Neurodegeneration, University of Göttingen Medical Center, Göttingen, Germany Cluster of Excellence Nanoscale Microscopy and Molecular Physiology of the Brain, Göttingen, Germany.
authors.forename: Felipe
authors.initials: F
authors.lastname: Opazo
mesh_terms: D000818:Animals; D005456:Fluorescent Dyes; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence / Q000379:methods*; D008954:Models, Biological; D013569:Synapses / Q000378:metabolism* / Q000648:ultrastructure*
chemical_list: D005456:Fluorescent Dyes; D008565:Membrane Proteins
keywords: VHH; affinity probes; nanobody; super-resolution; synapses