title: Presynaptic architecture of the larval zebrafish neuromuscular junction.
abstract: This article shows the ultrastructural architecture of larval zebrafish (Danio rerio) neuromuscular junctions in three dimensions. We compare classical electron microscopy fixation techniques with high-pressure freezing followed by freeze substitution (HPF/FS) in combination with electron tomography. Furthermore, we compare the structure of neuromuscular junctions in 4- and 8-dpf zebrafish larvae with HPF/FS because this allows for close-to-native ultrastructural preservation. We discovered that synaptic vesicles of 4-dpf zebrafish larvae are larger than those of 8-dpf larvae. Furthermore, we describe two types of dense-core vesicles and quantify a filamentous network of small filaments interconnecting synaptic vesicles as well as tethers connecting synaptic vesicles to the presynaptic cell membrane. In the center of active zones, we found elaborate electron-dense projections physically connecting vesicles of the synaptic vesicle pool to the presynaptic membrane. Overall this study establishes the basis for systematic comparisons of synaptic architecture at high resolution in three dimensions of an intact vertebrate in a close-to-native state. Furthermore, we provide quantitative information that builds the basis for diverse systems biology approaches in neuroscience, from comparative anatomy to cellular simulations.
authors.affiliation: Division of Electron Microscopy, Biocenter, University of Würzburg, 97074, Würzburg, Germany.
authors.forename: Frederik
authors.initials: F
authors.lastname: Helmprobst
authors.affiliation: Division of Electron Microscopy, Biocenter, University of Würzburg, 97074, Würzburg, Germany.
authors.forename: Miriam
authors.initials: M
authors.lastname: Frank
authors.affiliation: Division of Electron Microscopy, Biocenter, University of Würzburg, 97074, Würzburg, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Stigloher
mesh_terms: D000367:Age Factors; D000818:Animals; D005614:Freeze Fracturing; D021621:Imaging, Three-Dimensional; D007814:Larva / Q000166:cytology* / Q000648:ultrastructure; D008854:Microscopy, Electron; D008953:Models, Anatomic; D009469:Neuromuscular Junction / Q000254:growth & development / Q000378:metabolism / Q000648:ultrastructure*; D017729:Presynaptic Terminals / Q000378:metabolism / Q000648:ultrastructure*; D022142:Secretory Vesicles / Q000648:ultrastructure; D015027:Zebrafish
keywords: electron microscopy; electron tomography; freeze substitution; high-pressure freezing; synapse