title: Reconstruction of genetically identified neurons imaged by serial-section electron microscopy.
abstract: Resolving patterns of synaptic connectivity in neural circuits currently requires serial section electron microscopy. However, complete circuit reconstruction is prohibitively slow and may not be necessary for many purposes such as comparing neuronal structure and connectivity among multiple animals. Here, we present an alternative strategy, targeted reconstruction of specific neuronal types. We used viral vectors to deliver peroxidase derivatives, which catalyze production of an electron-dense tracer, to genetically identify neurons, and developed a protocol that enhances the electron-density of the labeled cells while retaining the quality of the ultrastructure. The high contrast of the marked neurons enabled two innovations that speed data acquisition: targeted high-resolution reimaging of regions selected from rapidly-acquired lower resolution reconstruction, and an unsupervised segmentation algorithm. This pipeline reduces imaging and reconstruction times by two orders of magnitude, facilitating directed inquiry of circuit motifs.
authors.affiliation: Center for Brain Science, Harvard University, Cambridge, United States.|Department of Molecular and Cellular Biology, Harvard University, Cambridge, United States.
authors.forename: Maximilian
authors.initials: M
authors.lastname: Joesch
authors.affiliation: Center for Brain Science, Harvard University, Cambridge, United States.|Department of Molecular and Cellular Biology, Harvard University, Cambridge, United States.
authors.forename: David
authors.initials: D
authors.lastname: Mankus
authors.affiliation: Center for Brain Science, Harvard University, Cambridge, United States.|Department of Molecular and Cellular Biology, Harvard University, Cambridge, United States.
authors.forename: Masahito
authors.identifier: 0000-0001-8193-2931
authors.initials: M
authors.lastname: Yamagata
authors.affiliation: University of Notre Dame, Notre Dame, United States.
authors.forename: Ali
authors.initials: A
authors.lastname: Shahbazi
authors.affiliation: Center for Brain Science, Harvard University, Cambridge, United States.|Department of Molecular and Cellular Biology, Harvard University, Cambridge, United States.
authors.forename: Richard
authors.initials: R
authors.lastname: Schalek
authors.affiliation: School of Engineering and Applied Sciences, Harvard University, Cambridge, United States.
authors.forename: Adi
authors.initials: A
authors.lastname: Suissa-Peleg
authors.affiliation: Division of Biology, California Institute of Technology, Pasadena, United States.
authors.forename: Markus
authors.identifier: 0000-0003-2136-6506
authors.initials: M
authors.lastname: Meister
authors.affiliation: Center for Brain Science, Harvard University, Cambridge, United States.|Department of Molecular and Cellular Biology, Harvard University, Cambridge, United States.
authors.forename: Jeff W
authors.initials: JW
authors.lastname: Lichtman
authors.affiliation: University of Notre Dame, Notre Dame, United States.
authors.forename: Walter J
authors.initials: WJ
authors.lastname: Scheirer
authors.affiliation: Center for Brain Science, Harvard University, Cambridge, United States.|Department of Molecular and Cellular Biology, Harvard University, Cambridge, United States.
authors.forename: Joshua R
authors.identifier: 0000-0001-8926-8836
authors.initials: JR
authors.lastname: Sanes
mesh_terms: D000818:Animals; D005260:Female; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D008297:Male; D008810:Mice, Inbred C57BL; D008854:Microscopy, Electron / Q000379:methods*; D008867:Microtomy / Q000379:methods*; D009415:Nerve Net / Q000033:anatomy & histology*; D009434:Neural Pathways / Q000033:anatomy & histology*; D012160:Retina / Q000166:cytology*; D013194:Staining and Labeling / Q000379:methods*
keywords: D. melanogaster; cell biology; connectomics; electron microscopy; mouse; neuroscience; peroxidase; reconstruction