Abstract
We combined cryo-preservation of intact Drosophila larvae and electron tomography with comprehensive segmentation of key features to reconstruct the complete ultrastructure of a model glutamatergic synapse in a near-to-native state. Presynaptically, we detail a complex network of filaments that connects and organizes synaptic vesicles. We link the complexity of this synaptic vesicle network to proximity to the active zone cytomatrix, consistent with the model that these protein structures function together to regulate synaptic vesicle pools. We identify a net-shaped network of electron-dense filaments spanning the synaptic cleft that suggests conserved organization of trans-synaptic adhesion complexes at excitatory synapses. Postsynaptically, we characterize a regular pattern of macromolecules that yields structural insights into the scaffolding of neurotransmitter receptors. Together, these analyses reveal an unexpected level of conservation in the nanoscale organization of diverse glutamatergic synapses and provide a structural foundation for understanding the molecular machines that regulate synaptic communication at a powerful model synapse.
Acknowledgements
We thank Ben August, Alex Kvit, the UW Medical School Electron Microscope Facility and the UW-Madison Materials Science Center for excellent technical support. We are grateful to Desiree Benefield, Janice Pennington, Marisa Otegui, Paul Ahlquist and Bill Hickey for generously sharing equipment and expertise. Janet Richmond, Szi-Chieh Yu, and Shigeki Watanabe provided helpful advice on optimizing use of HPF/FS techniques in intact animals. Finally, we thank members of the O’Connor-Giles lab for helpful discussions and comments on the manuscript.