Title: Investigating CNS synaptogenesis at single-synapse resolution by combining reverse genetics with correlative light and electron microscopy
Authors: Urwyler, Olivier
Izadifar, Azadeh
Dascenco, Dan
Petrovic, Milan
He, Haihuai
Ayaz, Derya
Kremer, Anna
Lippens, Saskia
Baatsen, Pieter
Guérin, Christopher J
Schmucker, Dietmar # ×
Issue Date: Jan-2015
Publisher: Company of Biologists
Series Title: Development vol:142 issue:2 pages:394-405
Article number: dev.115071
Abstract: Determining direct synaptic connections of specific neurons in the central nervous system (CNS) is a major technical challenge in neuroscience. As a corollary, molecular pathways controlling developmental synaptogenesis in vivo remain difficult to address. Here, we present genetic tools for efficient and versatile labeling of organelles, cytoskeletal components and proteins at single-neuron and single-synapse resolution in Drosophila mechanosensory (ms) neurons. We extended the imaging analysis to the ultrastructural level by developing a protocol for correlative light and 3D electron microscopy (3D CLEM). We show that in ms neurons, synaptic puncta revealed by genetically encoded markers serve as a reliable indicator of individual active zones. Block-face scanning electron microscopy analysis of ms axons revealed T-bar-shaped dense bodies and other characteristic ultrastructural features of CNS synapses. For a mechanistic analysis, we directly combined the single-neuron labeling approach with cell-specific gene disruption techniques. In proof-of-principle experiments we found evidence for a highly similar requirement for the scaffolding molecule Liprin-α and its interactors Lar and DSyd-1 (RhoGAP100F) in synaptic vesicle recruitment. This suggests that these important synapse regulators might serve a shared role at presynaptic sites within the CNS. In principle, our CLEM approach is broadly applicable to the developmental and ultrastructural analysis of any cell type that can be targeted with genetically encoded markers.
ISSN: 0950-1991
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Neuronal Wiring Lab
Laboratory of Ion Channel Research
Department of Human Genetics - miscellaneous
× corresponding author
# (joint) last author

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