Analyzing Synaptic Ultrastructure with Serial Section Electron Microscopy

2016 ◽  
pp. 35-62
Author(s):  
Jennifer N. Bourne
Keyword(s):  
Electron Microscopy ◽  
Serial Section ◽  
Synaptic Ultrastructure ◽  
Section Electron ◽  
Serial Section Electron Microscopy ◽  
Section Electron Microscopy

Normal Synaptonemal Complex and Abnormal Recombination Nodules in Two Alleles of the Drosophila Meiotic Mutant mei-W68

Genetics ◽  
2003 ◽  
Vol 163 (4) ◽  
pp. 1337-1356 ◽  
Author(s):  
Adelaide T C Carpenter
Keyword(s):  
Electron Microscopy ◽  
Synaptonemal Complex ◽  
High Frequency ◽  
Serial Section ◽  
The Other ◽  
Wild Type ◽  
Recombination Nodules ◽  
Section Electron ◽  
Serial Section Electron Microscopy ◽  
Section Electron Microscopy

Abstract The meiotic phenotypes of two mutant alleles of the mei-W68 gene, 1 and L1, were studied by genetics and by serial-section electron microscopy. Despite no or reduced exchange, both mutant alleles have normal synaptonemal complex. However, neither has any early recombination nodules; instead, both exhibit high numbers of very long (up to 2 μm) structures here named “noodles.” These are hypothesized to be formed by the unchecked extension of identical but much shorter structures ephemerally seen in wild type, which may be precursors of early recombination nodules. Although the mei-W68L1 allele is identical to the mei-W681 allele in both the absence of early recombination nodules and a high frequency of noodles (i.e., it is amorphic for the noodle phene), it is hypomorphic in its effects on exchange and late recombination nodules. The differential effects of this allele on early and late recombination nodules are consistent with the hypothesis that Drosophila females have two separate recombination pathways—one for simple gene conversion, the other for exchange.

scholarly journals Reconstruction of genetically identified neurons imaged by serial-section electron microscopy

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Maximilian Joesch ◽  
David Mankus ◽  
Masahito Yamagata ◽  
Ali Shahbazi ◽  
Richard Schalek ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Serial Section ◽  
Viral Vectors ◽  
Unsupervised Segmentation ◽  
Neuronal Structure ◽  
Section Electron ◽  
Neuronal Types ◽  
Serial Section Electron Microscopy ◽  
Complete Circuit ◽  
Section Electron Microscopy

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.

scholarly journals Whole-brain serial-section electron microscopy in larval zebrafish

2017 ◽  
Author(s):  
David Grant Colburn Hildebrand ◽  
Marcelo Cicconet ◽  
Russel Miguel Torres ◽  
Woohyuk Choi ◽  
Tran Minh Quan ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
High Resolution ◽  
Serial Section ◽  
Acquisition Time ◽  
Myelinated Axons ◽  
Whole Brain ◽  
Larval Zebrafish ◽  
Section Electron ◽  
Serial Section Electron Microscopy ◽  
Section Electron Microscopy

Investigating the dense meshwork of wires and synapses that form neuronal circuits is possible with the high resolution of serial-section electron microscopy (ssEM)1. However, the imaging scale required to comprehensively reconstruct axons and dendrites is more than 10 orders of magnitude smaller than the spatial extents occupied by networks of interconnected neurons2—some of which span nearly the entire brain. The difficulties in generating and handling data for relatively large volumes at nanoscale resolution has thus restricted all studies in vertebrates to neuron fragments, thereby hindering investigations of complete circuits. These efforts were transformed by recent advances in computing, sample handling, and imaging techniques1, but examining entire brains at high resolution remains a challenge. Here we present ssEM data for a complete 5.5 days post-fertilisation larval zebrafish brain. Our approach utilizes multiple rounds of targeted imaging at different scales to reduce acquisition time and data management. The resulting dataset can be analysed to reconstruct neuronal processes, allowing us to, for example, survey all the myelinated axons (the projectome). Further, our reconstructions enabled us to investigate the precise projections of neurons and their contralateral counterparts. In particular, we observed that myelinated axons of reticulospinal and lateral line afferent neurons exhibit remarkable bilateral symmetry. Additionally, we found that fasciculated reticulospinal axons maintain the same neighbour relations throughout the extent of their projections. Furthermore, we use the dataset to set the stage for whole-brain comparisons of structure and function by co-registering functional reference atlases and in vivo two-photon fluorescence microscopy data from the same specimen. We provide the complete dataset and reconstructions as an open-access resource for neurobiologists and others interested in the ultrastructure of the larval zebrafish.

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