-
[
West Coast Worm Meeting,
2002]
We have recently launched the prototype of Wormatlas (www.wormatlas.org). This atlas is designed to serve the scientific community with the main goal of bringing all the anatomical information pertinent to C. elegans within one readily accessible and easy to use web site. By creating extensive links to the WormBase as well as the C. elegans WWW server, we are aiming to provide users with seamless links between these databases. We hope to create the most comprehensive and complete online anatomy atlas for any genetic model organism.
-
[
International Worm Meeting,
2013]
WormGUIDES (Global Understanding in Dynamic Embryonic Systems) is a novel resource that aims to create and share the first 4D atlas with single cell resolution of embryogenesis and neurodevelopment for any animal. The first goal of WormGUIDES is production of an interactive atlas of nuclear positions from zygote until hatching. A simple navigation program for computers or hand-held devices facilitates the use of WormGUIDES as a reference tool in cell identification, quantification of developmental processes and visualization of nascent patterns and symmetries in the embryo. This program and data will be demonstrated at the meeting. WormGUIDES will also incorporate a complementary atlas of neurodevelopmental processes-neurite outgrowth and synaptogenesis-that will facilitate a dynamic understanding of the connectome emerging throughout development. Building on the C. elegans community's open-sharing of systems-level knowledge and resources, WormGUIDES will further enhance the value of C. elegans as a model organism.
-
Breimann, L., Bahry, E., Preibisch, S., Wang, Y., Myers, E., Sun, M.
[
International Worm Meeting,
2017]
We are creating a high-resolution nuclear atlas of the C. elegans dauer from a nanometer-resolved transmission electron microscopy (TEM) reconstruction of an entire dauer larva. Onto this atlas, we map fixed and live light microscopy data exploiting the fixed lineage of C. elegans. This high-resolution atlas can therefore serve as template for any type of microscopy acquisition containing a nuclear marker. This atlas will contribute to C. elegans dauer research as a resource. It will shed light on the dauer lineage, it will allow to directly compare microscopy acquisitions from different dauer larvae at single cell resolution and thus pave the way for studying the dauer diapause systematically. We acquired a TEM dataset of an entire Dauer C. elegans (
daf-2 e1370 mutant) consisting of 25,000 high-resolution images, covering 561 sections, and totaling a size of approximately one terabyte. Following non-rigid alignment using TrakEM2, we developed a semi-automatic approach based on machine learning and dynamic programming to extract all nuclei including their location, size, shape, and specifics of nuclear architecture. We therefore manually annotated a small subset (~10%) of the TEM sections and used them as a training set for machine learning based classification that was subsequently applied to all non-annotated sections to retrieve the probability for each pixel being inside a nucleus. The final outline of each nucleus in each section was then identified by computing an optimal contour that balances its distance to the most probable outline as defined by the machine-learning classification, its smoothness, and its similarity to neighboring sections. To verify the resulting dauer nuclei atlas from the TEM dataset and to assess variability, we acquired confocal images of fixed dauer larvae expressing LMN-1::GFP and stained with DAPI to highlight nuclei locations and outlines. The nuclei were segmented using a combination of local thresholding and machine-learning approaches, while unique geometric constellations of nuclei facilitated registration of the confocal images to the TEM-based atlas. We aim for the dauer atlas to be an accessible web-based open source and open access resource for the community that will ultimately allow integration of a wide range of different types of imaging data into a common model.
-
[
International Worm Meeting,
2007]
Despite detailed knowledge of C. elegans anatomy, the lack of a digital atlas of the spatial location of nuclei impedes automatic high-content analysis of cellular information. We have used confocal images of newly hatched first larval stage hermaphrodites of C. elegans to build such a 3D digital nuclei atlas. We used DAPI to stain all 558 cells in an L1 larva and GFP to label the 81 body wall muscle cells and 1 depressor muscle cell. To build the atlas, we first developed an automatic algorithm to straighten the curved worm body into a standard rod shape. We then developed an automatic approach to segment each individual nucleus. The method has achieved over 95% accuracy in our bench tests. We also developed a 3D annotation tool to label each segmented nucleus, correct segmentation errors, and quantitatively evaluate segmentation accuracy. Finally, we generated the atlas by mapping the nuclei from 14 stacks into a canonical framework defined by linear transformations with respect to anterior/posterior, left/right, and dorsal/ventral (A/P, L/R, and D/V) landmarks. The nuclei atlas is represented using statistical metrics, including mean and standard deviations of cell locations, spacing, and co-variations in the A/P, L/R, and D/V dimensions. We also derived the invariant nuclei location relationship along each dimension represented as graphs. Our analysis shows that the locations of the majority of nuclei are highly stereotyped across different individuals. The several nuclei that have big variations have also been identified reliably, consistent with earlier literature. The atlas shows that different types of cells exhibit specific location patterns and cell spacing patterns, and that the locations of specific groups of cells tend to co-vary. Such spatial synergy patterns may indicate functional links between the respective cells. Using this atlas, we developed an automatic cell recognition method to name the cells in any C.elegans L1 image. On the 181 cells that are easily identifiable by human eye, we achieved an average recognition accuracy of 98.31%. Moreover for the 82 GFP-marked muscle cells we achieved an accuracy of 99.30%. This technique thus enables high throughput analysis of cellular information such as gene expression in the L1 larvae using wCherry to mark a target gene (see the abstract by Liu, Long, Peng, Myers and Kim). Our techniques can also be used to build digital atlases for other larvae stages and adult worms.
-
Moyle, Mark, Kovacevic, Ismar, Mohler, William, Colon-Ramos, Daniel, Gutierrez, Natasha, Shroff, Hari, Marquina, Javier, Bao, Zhirong, Christensen, Ryan, Santella, Anthony, Kumar, Abhishek, Shah, Pavak, Wu, Yicong
[
International Worm Meeting,
2015]
How neuronal networks develop during embryogenesis to generate a functional nervous system is a fundamental question in developmental biology and neuroscience. In C. elegans the entire wiring diagram of the adult nervous system ("the connectome") has been mapped. However, it is unclear how the nervous system is organized during embryogenesis to correctly form the connectome. Through a multidisciplinary collaboration, we have established an imaging and analysis pipeline to generate and share a 4D atlas of C. elegans neurodevelopment throughout embryogenesis. To this end, we created a light-sheet microscope capable of prolonged time-lapse imaging with subcellular resolution. The dual-view selective plane illumination microscope (diSPIM) is capable of constant volumetric 350nm isotropic imaging at speeds 10-1000x faster than conventional 4D imaging techniques. Also, we have collected existing and are generating novel C. elegans strains that label sparse subsets of neurons to facilitate segmentation and are pioneering strategies for heat-shocking/photoactivation to allow for efficient use of pan-neuronal promoters. Furthermore, we developed novel image analysis software (StarryNite) capable of systematically tracking and identifying every developing cell, as well as software for segmenting individual neurons in 4D, and linearizing the twisted C. elegans embryo. Finally, to allow for widespread dissemination of these large data sets, we created iOS and Android applications, and an Internet-based viewing program called CytoSHOW, which allows the scientific community access to all the neurodevelopmental data. We envision a dynamic workspace where any researcher can visualize, integrate, and interrelate the diverse scope of C. elegans neurological and developmental data.
-
Christensen, Ryan, Shroff, Hari, Colon-Ramos, Daniel, Harvey, Brandon, Guo, Min, Santella, Anthony, Xu, Stephen, Del Toro-Pedrosa, Daniel, Mohler, William, Wu, Yicong, Bokinsky, Alexandra, Moyle, Mark, Duncan, Leighton, Levin, Michael, Ardiel, Evan, Schwartz, Gabi, Lauziere, Andrew, Karaj, Nensi, McCreedy, Evan, Bao, Zhirong, Vazquez Martinez, Nabor
[
International Worm Meeting,
2021]
The limited number of cells and invariant cell lineage of the Caenorhabditis elegans embryo make it an excellent system for examining complex developmental events, such as tissue movement and neurodevelopment. Prior work from the WormGUIDES project has digitized the position of all nuclei for the first half of embryogenesis, creating a computational map of the embryo that can be used to overlay developmentally relevant information like gene expression or neurite outgrowth. Creating a similar map for the second half of embryogenesis is difficult due to embryo elongation and movement. We have developed software to computationally untwist the moving embryo, allowing for analysis of cell position during this period of development, and have begun expanding our computational map into the second half of embryogenesis. Our current map includes 202 nuclei across the embryo, including 32 neuronal nuclei, 81 body wall muscle nuclei, 20 intestinal, and 20 seam cell nuclei. We also include a tract-based model of the nerve ring, showing how it is positioned relative to neuronal and body wall muscle nuclei as the embryo elongates. In addition to our partial nuclear atlas, we describe improvements to our untwisting and tracking workflow, including a deep-learning image restoration capability which improves image quality during rapid embryo movements, and a semi-automated tracking upgrade to our untwisting software which improves tracking throughput. As we continue to add nuclei and neuronal morphology to the atlas, we plan to integrate our post twitching model with previous pre-twitching work to develop a digital atlas spanning the entirety of embryogenesis.
-
[
International Worm Meeting,
2017]
Cell identification is a ubiquitous task in C. elegans research, allowing researchers to exploit the power of the stereotyped cell lineage uniquely available in the worm. However, cell identification from still or video microscopy images is a notoriously difficult, time consuming, and error prone process, more an art than a science and best done by experienced experts in the field. We introduce an automated, community-driven, machine learning based cellular identification system we term a probabilistic cell atlas and demonstrate its use on C. elegans adult hermaphrodite pan-neuronal fluorescent calcium imaging recordings. We demonstrate use of the system on both stills of static fluorescent GFP and mCherry markers and videos of dynamic GCaMP markers, and combinations of these imaging modalities. The distinguishing features of our approach vis a vis prior approaches include: (1) generation of probabilistic guesses at cell ID rather than single best-guesses for each cell found in the test data, (2) flexible/optional utilization of multi-modal training data, including cell position, morphology, color, and, notably, activity through time, and (3) iterative Bayesian improvement of the probabilistic model of cell features (i.e. the probabilistic atlas). Because of this design, the accuracy, confidence, and input flexibility of the system progressively improve with each dataset uploaded by users. The system is free for use, open source and available online. We encourage fellow worm researchers to upload their datasets they wish to ID and thereby contribute to the collective value of the system. We hope this system becomes another long-term shared resource for the worm community.
-
[
International Worm Meeting,
2021]
Interactions between neurons and glia are essential for all normal nervous system functions. C. elegans glia have similar types of function as mammalian glia and are now established as a powerful genetic model to study glial biology. The adult hermaphrodite nervous system contains 302 neurons and 56 glia, with at least some of the glia shown to profoundly impact associated neuron shape and associated animal behaviors. Adult males have an additional 36 glia and 89 neurons, many of which control male mating-related behaviors. A major limitation in studying C. elegans glia functions is the lack of cell-type specific markers for each glial subtype in the adult worm, and a global comparative assessment of glial functional and molecular heterogeneity. Further, each C. elegans glia is born of an invariant developmental lineage and makes invariant neuron-contact. This not only makes the nematode a powerful model to study glia-neuron interactions, but also suggests a unique advantage of this model to examine glia at single-cell and molecular detail. Addressing this gap, we present our single nuclear RNAseq (snRNAseq) studies on adult C. elegans glia across the entire nervous system. We have done these studies in three settings, (1) young adult hermaphrodites, (2) young adult males, and (3) aged hermaphrodites. These data will be compiled and shared as a searchable three-dimensional online atlas of adult glial gene expression as a community resource. Our datasets allow us to now create a gene expression and marker atlas of glia at single cell resolution. Our preliminary data reveal 30 clusters, hinting already at interesting biological insights into glial heterogeneity. Further, studies in our lab and others have found that multiple glial cues within a single glia, the amphid sheath (AMsh), regulate neuron morphology and sensory behaviors, with age-related effects. Therefore, in addition to describing glia-specific and glial subclass markers, we are currently validating our datasets functionally in similar assays, including neural aging. The aim of our studies is to expand our understanding of how different glia execute these regulatory neural functions across cell-type, sex and animal aging.
-
Wang, C, Aghayeva, U, Sun, H, Glenwinkel, L, Vidal Iglesias, B, Hobert, O, Bayer, E
[
International Worm Meeting,
2017]
One goal of modern day neuroscience is the establishment of molecular maps that assign unique molecular, and hence, functional features to individual neuron types. Such maps provide important starting points for a number of downstream applications. We describe here nervous system-wide maps of the expression of 250 members of the largest gene family in the C .elegans genome, rhodopsin-like (class A) GPCR receptors, composed of more than 1300 genes. We synthesize our expression patterns with those previously described and we have now expression information for about 25% of the gene family. As previously anticipated, 90% of the genes are expressed in sensory neurons, although not always exclusively. GPCR expression is significantly enriched in amphid and phasmid neurons, which in some cases express over 50 different receptors. Remarkably, the neurons with most GPCRs expressed are all involved in nociceptive behaviors. Independent of its "classic" sensory modality, we find that every sensory neuron, except URY, expresses at least one sensory-type GPCR. Moreover, around 20% are expressed in interneurons and motorneurons, implying that they have other functions beyond being sensory receptors. Including sensory, inter- and motorneurons we have so far revealed 25 new neuron types showing sensory-type GPCR expression. We have identified 6 new GPCRs asymmetrically expressed in the AWC olfactory neuron, which we found to be regulated by known mechanisms establishing AWC asymmetry. The fact that we did not find asymmetric GPCR expression in any new neuron type suggests that C. elegans might not use functional lateralization as a widespread mechanism to increase sensory discrimination. After examining a subset of our reporters at different larval stages including dauer, we found two GPCRs that show different expression in L1 versus adult and 16 GPCRs that change their expression pattern during dauer diapause. Thus, sensory-type GPCRs seem to be very dynamically regulated in dauer, when the animal is exposed to a dramatically different sensory environment. Our analysis constitutes a resource that provides multitude of starting points for multi-level downstream analysis, including the functional analysis of GPCRs in specific neuron types or the bioinformatic and experimental analysis of cis-regulatory control mechanisms and their evolution. Reporter transgenes are also invaluable neuronal identity markers that will help shed light on neuronal differentiation processes.
-
Wu, Y., Guo, M., Moyle, M., Ardiel, E., Shroff, H., Bokinsky, A., Santella, A., McCreedy, E., Colon-Ramos, D., Karaj, N., Duncan, L., Bao, Z., Levin, M., Mohler, W., Lauziere, A., Harvey, B., Christensen, R.P.
[
International Worm Meeting,
2019]
The Caenorhabditis elegans embryo represents an excellent model system in which to study tissue formation. However, the onset of twitching and elongation makes data analysis during the second half of embryogenesis difficult. Previously, we developed software to enable computational untwisting of the C. elegans embryo, removing the effects of embryo movement and placing embryo images in a common reference frame for analysis. We have now improved our software suite to incorporate more user-friendly positional tracking and better segmentation, reducing clipping of images around the edges of the embryo. We also apply deep learning to segment nuclei in a semi-automated fashion. We apply our software to generate a map showing the position of 158 nuclei in the post-twitching worm embryo, as a partial step in the generation of a complete embryonic nuclear atlas. Tracked nuclei include 16 neurons and 81 body wall muscles. Our improved tools, combined with pre-twitching work from our collaborators on the WormGUIDES project, allow us to pursue the goal of developing a complete nuclear and neurite outgrowth atlas for the nematode embryo from the two cell stage until hatching.