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Methods Cell Biol,
1995]
Although Caenorhabditis elegans was originally chosen as a model organism for cell biology with serial section electron microscopy (EM) methods in mind, these methods have remained a daunting challenge. There is an apocryphal story that Nichol Thomson originally advised Sydney Brenner that C. elegans was unsuitable for electron microscopy and that Brenner should choose another species. Other experienced microscopists have probably shared similar dark thoughts from time to time. Nonetheless, the worm's very small size, simple organization, and cablelike nervous system have permitted Brenner's colleagues to characterize every cell and cell contact in the wild-type animal, potentiating the genetic characterization of cellular development in remarkable detail. We attempt to provide an adequate background for anyone to initiate EM studies of C. elegans. Two decades ago, as the first of Brenner's postdoctoral fellows left his laboratory to establish new worm laboratories, it was standard practice to include an EM component in their studies. Their combined efforts to characterize the adult animal's cell types and the essential steps in its development helped to erect a lovely scaffold of key manuscripts, capped by the description of the "Mind of the Worm" in some 600 micrographs and 175 drawings. Many of these works required technical heroics or suffered long delays before publication. Most people later chose to leave electron microscopy behind in pursuit of molecular quarry. The fruits of their molecular and genetic studies should soon stimulate a renewed flowering of electron microscopy. We hope to smooth your entry or reentry into these techniques. We also summarize our methods for three-dimensional (3D) image reconstruction, based largely on film techniques introduced by John White and Randle Ware. Digital imaging techniques seem poised to make 3D reconstruction more accessible, and may simplify the exchange of morphological data between laboratories. We discuss several computer systems that the C. elegans community could adopt for high-resolution studies of structure and function. In addition, we briefly cover several specialized specimen preparation techniques for electron microscopy, including freeze fracture and electron microscopic immunocytochemistry.
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J Cell Sci,
2018]
The mitotic spindle is a complex three-dimensional (3D) apparatus that functions to ensure the faithful segregation of chromosomes during cell division. Our current understanding of spindle architecture is mainly based on a plethora of information derived from light microscopy with rather few insights about spindle ultrastructure obtained from electron microscopy. In this Review, we will provide insights into the history of imaging of mitotic spindles and highlight recent technological advances in electron tomography and data processing, which have delivered detailed 3D reconstructions of mitotic spindles in the early embryo of the nematode Caenorhabditis elegans Tomographic reconstructions provide novel views on spindles and will enable us to revisit and address long-standing questions in the field of mitosis.
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Methods Cell Biol,
2010]
We describe a method for automatically finding the location and conformations of microtubules in tomograms of high-pressure frozen, freeze substituted cells. Our approach uses two steps: a preprocessing step that finds locations in the tomograms that are likely to lie inside microtubules and a tracking step that uses the preprocessed data to identify the trajectories of individual microtubules. We test this method on a reconstruction of a Caenorhabditis elegans mitotic spindle and we compare our results with those obtained by a human expert who manually segmented the same data. At present, the method could be used to assist the analysis of large-scale tomography reconstructions. With further improvements, it may be possible to reliably segment cellular tomograms without human intervention.
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J Dev Biol,
2019]
The complete structure and connectivity of the <i>Caenorhabditis elegans</i> nervous system ("mind of a worm") was first published in 1986, representing a critical milestone in the field of connectomics. The reconstruction of the nervous system (connectome) at the level of synapses provided a unique perspective of understanding how behavior can be coded within the nervous system. The following decades have seen the development of technologies that help understand how neural activity patterns are connected to behavior and modulated by sensory input. Investigations on the developmental origins of the connectome highlight the importance of role of neuronal cell lineages in the final connectivity matrix of the nervous system. Computational modeling of neuronal dynamics not only helps reconstruct the biophysical properties of individual neurons but also allows for subsequent reconstruction of whole-organism neuronal network models. Hence, combining experimental datasets with theoretical modeling of neurons generates a better understanding of organismal behavior. This review discusses some recent technological advances used to analyze and perturb whole-organism neuronal function along with developments in computational modeling, which allows for interrogation of both local and global neural circuits, leading to different behaviors. Combining these approaches will shed light into how neural networks process sensory information to generate the appropriate behavioral output, providing a complete understanding of the worm nervous system.
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Science,
1994]
In 1967, Sydney Brenner isolated the first behavioral mutants of the nematode Caenorhabditis elegans, and in 1970, John White began the systematic reconstruction of its nervous system. This dual approach of genetics coupled with detailed morphological analysis, now enhanced by the tools of molecular biology and electrophysiology, still dominates the study of the function and development of the C. elegans nervous system. Although Brenner's vision of a comprehensive understanding of this simple animal has taken time to mature, findings of the past few years indicate that the tree is bearing fruit.
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Curr Opin Struct Biol,
2019]
The organization of microtubules in spindles is complex and not fully understood. Here we report on current advances in generating 3D reconstructions of staged spindles by serial-section electron tomography, exemplified by the first mitotic spindle in early Caenorhabditis elegans embryo. We then review how advances in correlative light microscopy and quantitative electron tomography enable the development of theory and stochastic simulations, which describe how the microtubule organization in spindles emerges from their dynamics. We show how theory and simulations can be used to address long-standing questions in cell division research, advancing the field beyond a pure structural description of microtubules in spindles.
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Curr Opin Neurobiol,
2014]
With a fully reconstructed and extensively characterized neural circuit, the nematode Caenorhabditis elegans is a promising model system for integrating our understanding of neuronal, circuit and whole-animal dynamics. Fundamental to addressing this challenge is the need to consider the tight neuronal-environmental coupling that allows the animal to survive and adapt to changing conditions. Locomotion behaviors are affected by environmental variables both at the biomechanical level and via adaptive sensory responses that drive and modulate premotor and motor circuits. Here we review significant advances in our understanding of proprioceptive control of locomotion, and more abstract models of spatial orientation and navigation. The growing evidence of the complexity of the underlying circuits suggests that the intuition gained is but the first step in elucidating the secrets of neural computation in this relatively simple system.
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Trends in Genetics,
2000]
How far down the phylogenic tree should we look for the origins of innate immunity? We know that mammalian cells respond to microbes using Toll-like signalling systems that are remarkably similar in arthropods. Prototypes of these pathways might have arisen in more primitive phyla (initially, perhaps, to regulate development) and their identification would help us to reconstruct the evolution of this facet of immunity. Elements of Toll pathways exist in plants. Does this mean that the last common ancestor of plants, chordates and arthropods, which was a unicellular eukaryote, expressed a Toll-like pathway, or is a similar developmental logic at work in all multicellular life-forms? To address some of these issues we decided to seek a 'worm' Toll pathway, concentrating on the simple and versatile metazoan, Caenorhabditis elegans.
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Proc. Fifth International Congress on Parasitology,
1982]
During the past ten years the nematode Caenorhabditis elegans has been subjected to intensive anatomical, genetic, developmental and behavioral analysis. More than 2500 mutants have been isolated; the complete developmental lineages of all embryonic and post embryonic cells have been determined; and the complete wiring diagram of its 300 neurons has been reconstructed by serial electron microscopy. Although C. elegans is a nonparasitic bacteria eating soil nematode and thus is not a proper subject for a parasitology congress, so much has been learned about this worm that it was elevated to honorary parasite status for this meeting. I will review some examples of how the genetic analysis of this helminth has helped established the function of parts of the sensory nervous system. Since the neuroanatomy of nematodes is so highly conserved these results should apply to parasitic nematodes as well.
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Sci China Life Sci,
2015]
Since Caenorhabditis elegans was chosen as a model organism by Sydney Brenner in 1960's, genetic studies in this organism have been instrumental in discovering the function of genes and in deciphering molecular signaling network. The small size of the organism and the simple nervous system enable the complete reconstruction of the first connectome. The stereotypic developmental program and the anatomical reproducibility of synaptic connections provide a blueprint to dissect the mechanisms underlying synapse formation. Recent technological innovation using laser surgery of single axons and in vivo imaging has also made C. elegans a new model for axon regeneration. Importantly, genes regulating synaptogenesis and axon regeneration are highly conserved in function across animal phyla. This mini-review will summarize the main approaches and the key findings in understanding the mechanisms underlying the development and maintenance of the nervous system. The impact of such findings underscores the awesome power of C. elegans genetics.