Early steps in ventral nerve cord development in chelicerates and myriapods and formation of brain compartments in spiders

The central point of this work is the investigation of neurogenesis in chelicerates and myriapods. By comparing decisive mechanisms in neurogenesis in the four arthropod groups (Chelicerata, Crustacea, Insecta, Myriapoda) I was able to show which of these mechanisms are conserved and which developmental modules have diverged. Thereby two processes of embryonic development of the central nervous system were brought into focus. On the one hand I studied early neurogenesis in the ventral nerve cord of the spiders Cupiennius salei and Achaearanea tepidariorum and the millipede Glomeris marginata and on the other hand the development of the brain in Cupiennius salei.rnWhile the nervous system of insects and crustaceans is formed by the progeny of single neural stem cells (neuroblasts), in chelicerates and myriapods whole groups of cells adopt the neural cell fate and give rise to the ventral nerve cord after their invagination. The detailed comparison of the positions and the number of the neural precursor groups within the neuromeres in chelicerates and myriapods showed that the pattern is almost identical which suggests that the neural precursors groups in these arthropod groups are homologous. This pattern is also very similar to the neuroblast pattern in insects. This raises the question if the mechanisms that confer regional identity to the neural precursors is conserved in arthropods although the mode of neural precursor formation is different. The analysis of the functions and expression patterns of genes which are known to be involved in this mechanism in Drosophila melanogaster showed that neural patterning is highly conserved in arthropods. But I also discovered differences in early neurogenesis which reflect modifications and adaptations in the development of the nervous systems in the different arthropod groups.rnThe embryonic development of the brain in chelicerates which was investigated for the first time in this work shows similarities but also some modifications to insects. In vertebrates and arthropods the adult brain is composed of distinct centres with different functions. Investigating how these centres, which are organised in smaller compartments, develop during embryogenesis was part of this work. By tracing the morphogenetic movements and analysing marker gene expressions I could show the formation of the visual brain centres from the single-layered precheliceral neuroectoderm. The optic ganglia, the mushroom bodies and the arcuate body (central body) are formed by large invaginations in the peripheral precheliceral neuroectoderm. This epithelium itself contains neural precursor groups which are assigned to the respective centres and thereby build the three-dimensional optical centres. The single neural precursor groups are distinguishable during this process leading to the assumption that they carry positional information which might subdivide the individual brain centres into smaller functional compartments.rn

The Role of Antennapedia in Anteroposterior Patterning of the Drosophila Melanogaster Ventral Nervous System

A major unresolved question in developmental neurobiology is how the nervous system is adapted to the specific needs of the organism at different life stages. In the holometabolous insect Drosophila melanogaster, the larval ventral nervous system (VNS) is comprised of similar repeating segments, as opposed to the adult VNS, which varies greatly from segment to segment both in number and types of neurons. The adult-specific neurons of each segment are generated by 25 distinct types of neuronal progenitor cells called neuroblasts (NBs) that appear in a stereotyped array (Truman et al., 2004). Each NB divides repeatedly to produce a distinct set of daughter cells termed a lineage, which is bilaterally symmetric but present to varying degrees in each segment. These daughter cells can be distinguished by their position within the nervous system as well as by their axonal projections. Each of the 25 NBs produces neurons; if both daughter cells are present in a lineage then both sibling populations survived, whereas if only one projection is seen cell death occurred, leaving a hemilineage (half lineage). In some lineages, the same sibling type survives in all segments in which the lineage appears, but in others, the surviving sibling type varies across segments, resulting in a different morphology for the same lineage in different segments. How are these differences in survival and morphology controlled? The Hox genes provide positional information for developing structures along the anterior-posterior (AP) axis of animals. They encode transcription factors, thereby controlling the activity of genes down stream. In the postembryonic VNS, each NB lineage features its own characteristic expression pattern of Hox genes Antp and Ubx, which can vary from segment-to-segment, and can thereby cause variation in the number of neural cells and axonal projections that survive. This study defines the wild-type expression pattern of Antp and elucidates the role of Antp in gain of function studies. These studies are possible due to the MARCM (Mosaic Analysis with a Repressible Cell Marker) method, which allows the genetically manipulated cells to be specifically labeled in an otherwise normal, unlabeled organism. The results indicate that Antp is expressed in a segment-, lineage-, and hemilineage-specific manner. Antp is sufficient for both anterior and posterior transformations of particular lineages, including promotion of cell death and/or survival as well as axon guidance.

Microsurgical and laser ablation analysis of leaf positioning and dorsoventral patterning in tomato

Leaves are arranged according to regular patterns, a phenomenon referred to as phyllotaxis. Important determinants of phyllotaxis are the divergence angle between successive leaves, and the size of the leaves relative to the shoot axis. Young leaf primordia are thought to provide positional information to the meristem, thereby influencing the positioning of new primordia and hence the divergence angle. On the contrary, the meristem signals to the primordia to establish their dorsoventral polarity, which is a prerequisite for the formation of a leaf blade. These concepts originate from classical microsurgical studies carried out between the 1920s and the 1970s. Even though these techniques have been abandoned in favor of genetic analysis, the resulting insights remain a cornerstone of plant developmental biology. Here, we employ new microsurgical techniques to reassess and extend the classical studies on phyllotaxis and leaf polarity. Previous experiments have indicated that the isolation of an incipient primordium by a tangential incision caused a change of divergence angle between the two subsequent primordia, indicating that pre-existing primordia influence further phyllotaxis. Here.. we repeat these experiments and compare them with the results of laser ablation of incipient primordia. Furthermore. we explore to what extent the different pre-existing primordia influence the size and position of new organs. and hence phyllotaxis. We propose that the two youngest primordia (P-1 and P-2) are sufficient for the approximate positioning of the incipient primordium (I-1), and therefore for the perpetuation of the generative spiral, whereas the direct contact neighbours of I-1 (P-2 and P-3) control its delimitation and hence its exact size and position. Finally. we report L I specific cell ablation experiments suggesting that the meristem L-1 layer is essential for the dorsoventral patterning of leaf primordia.

Measuring the gravitational free-fall of antihydrogen

Antihydrogen holds the promise to test, for the first time, the universality of freefall with a system composed entirely of antiparticles. The AEgIS experiment at CERN’s antiproton decelerator aims to measure the gravitational interaction between matter and antimatter by measuring the deflection of a beam of antihydrogen in the Earths gravitational field (g). The principle of the experiment is as follows: cold antihydrogen atoms are synthesized in a Penning-Malberg trap and are Stark accelerated towards a moir´e deflectometer, the classical counterpart of an atom interferometer, and annihilate on a position sensitive detector. Crucial to the success of the experiment is the spatial precision of the position sensitive detector.We propose a novel free-fall detector based on a hybrid of two technologies: emulsion detectors, which have an intrinsic spatial resolution of 50 nm but no temporal information, and a silicon strip / scintillating fiber tracker to provide timing and positional information. In 2012 we tested emulsion films in vacuum with antiprotons from CERN’s antiproton decelerator. The annihilation vertices could be observed directly on the emulsion surface using the microscope facility available at the University of Bern. The annihilation vertices were successfully reconstructed with a resolution of 1–2 μmon the impact parameter. If such a precision can be realized in the final detector, Monte Carlo simulations suggest of order 500 antihydrogen annihilations will be sufficient to determine gwith a 1 % accuracy. This paper presents current research towards the development of this technology for use in the AEgIS apparatus and prospects for the realization of the final detector.

Morphological analysis of the mammalian postcranium: A developmental perspective

The past two decades have greatly improved our knowledge of vertebrate skeletal morphogenesis. It is now clear that bony morphology lacks individual descriptive specification and instead results from an interplay between positional information assigned during early limb bud deployment and its “execution” by highly conserved cellular response programs of derived connective tissue cells (e.g., chondroblasts and osteoblasts). Selection must therefore act on positional information and its apportionment, rather than on more individuated aspects of presumptive adult morphology. We suggest a trait classification system that can help integrate these findings in both functional and phylogenetic examinations of fossil mammals and provide examples from the human fossil record.

Segregation of Two Spectrin Isoforms: Polarized Membrane-binding Sites Direct Polarized Membrane Skeleton Assembly

Spectrin isoforms are often segregated within specialized plasma membrane subdomains where they are thought to contribute to the development of cell surface polarity. It was previously shown that ankyrin and β spectrin are recruited to sites of cell–cell contact in Drosophila S2 cells expressing the homophilic adhesion molecule neuroglian. Here, we show that neuroglian has no apparent effect on a second spectrin isoform (αβH), which is constitutively associated with the plasma membrane in S2 cells. Another membrane marker, the Na,K-ATPase, codistributes with ankyrin and αβ spectrin at sites of neuroglian-mediated contact. The distributions of these markers in epithelial cells in vivo are consistent with the order of events observed in S2 cells. Neuroglian, ankyrin, αβ spectrin, and the Na,K-ATPase colocalize at the lateral domain of salivary gland cells. In contrast, αβH spectrin is sorted to the apical domain of salivary gland and somatic follicle cells. Thus, the two spectrin isoforms respond independently to positional cues at the cell surface: in one case an apically sorted receptor and in the other case a locally activated cell–cell adhesion molecule. The results support a model in which the membrane skeleton behaves as a transducer of positional information within cells.

GenMapDB: a database of mapped human BAC clones

GenMapDB (http://genomics.med.upenn.edu/genmapdb) is a repository of human bacterial artificial chromosome (BAC) clones mapped by our laboratory to sequence-tagged site markers. Currently, GenMapDB contains over 3000 mapped clones that span 19 chromosomes, chromosomes 2, 4, 5, 9–22, X and Y. This database provides positional information about human BAC clones from the RPCI-11 human male BAC library. It also contains restriction fragment analysis data and end sequences of the clones. GenMapDB is freely available to the public. The main purpose of GenMapDB is to organize the mapping data and to allow the research community to search for mapped BAC clones that can be used in gene mapping studies and chromosomal mutation analysis projects.

Pax9 and Jagged1 act downstream of Gli3 in vertabrate limb development

From early in limb development the transcription factor Gli3 acts to define boundaries of gene expression along the anterior-posterior (AP) axis, establishing asymmetric patterns required to provide positional information. As limb development proceeds, posterior mesenchyme expression of Sonic hedgehog (Shh) regulates Gli3 transcription and post-translational processing to specify digit number and identity. The molecular cascades dependent on Gli3 at later stages of limb development, which link early patterning events with final digit morphogenesis, remain poorly characterised. By analysing the transcriptional consequences of loss of Gli3 in the anterior margin of the E11.5 and E12.5 limb bud in the polydactylous mouse mutant extra-toes (Gli3(Xt/Xt)), we have identified a number of known and novel transcripts dependent on Gli3 in the limb. In particular, we demonstrated that the genes encoding the paired box transcription factor Pax9, the Notch ligand Jagged1 and the cell surface receptor Cdo are dependent on Gli3 for correct expression in the anterior limb mesenchyme. Analysis of expression in compound Shh;Gli3 mutant mouse embryos and in both in vitro and in vivo Shh signaling assays, further defined the importance of Shh regulated processing of Gli3 in controlling gene expression. In particular Pax9 regulation by Shh and Gli3 was shown to be context dependent, with major differences between the limb and somite revealed by Shh bead implantation experiments in the chick. Jagged1 was shown to be induced by Shh in the chick limb and in a C3H10T1/2 cell based signaling assay, with Shh;Gli3 mutant analysis indicating that expression is dependent on Gli3 derepression. Our data have also revealed that perturbation of early patterning events within the Gli3(Xt/Xt), limb culminates in a specific delay of anterior chondrogenesis which is subsequently realised as extra digits. (c) 2005 Elsevier Ireland Ltd. All rights reserved.

Reduced attentional capacity, but normal processing speed and shifting of attention in developmental dyslexia: evidence from a serial task

We report the performance of a group of adult dyslexics and matched controls in an array-matching task where two strings of either consonants or symbols are presented side by side and have to be judged to be the same or different. The arrays may differ either in the order or identity of two adjacent characters. This task does not require naming – which has been argued to be the cause of dyslexics’ difficulty in processing visual arrays – but, instead, has a strong serial component as demonstrated by the fact that, in both groups, Reaction times (RTs) increase monotonically with position of a mismatch. The dyslexics are clearly impaired in all conditions and performance in the identity conditions predicts performance across orthographic tasks even after age, performance IQ and phonology are partialled out. Moreover, the shapes of serial position curves are revealing of the underlying impairment. In the dyslexics, RTs increase with position at the same rate as in the controls (lines are parallel) ruling out reduced processing speed or difficulties in shifting attention. Instead, error rates show a catastrophic increase for positions which are either searched later or more subject to interference. These results are consistent with a reduction in the attentional capacity needed in a serial task to bind together identity and positional information. This capacity is best seen as a reduction in the number of spotlights into which attention can be split to process information at different locations rather than as a more generic reduction of resources which would also affect processing the details of single objects.

Clinical translation of a hand-held optical imager for breast cancer diagnostics: In vitro and in vivo tomography studies

Optical imaging is an emerging technology towards non-invasive breast cancer diagnostics. In recent years, portable and patient comfortable hand-held optical imagers are developed towards two-dimensional (2D) tumor detections. However, these imagers are not capable of three-dimensional (3D) tomography because they cannot register the positional information of the hand-held probe onto the imaged tissue. A hand-held optical imager has been developed in our Optical Imaging Laboratory with 3D tomography capabilities, as demonstrated from tissue phantom studies. The overall goal of my dissertation is towards the translation of our imager to the clinical setting for 3D tomographic imaging in human breast tissues. A systematic experimental approach was designed and executed as follows: (i) fast 2D imaging, (ii) coregistered imaging, and (iii) 3D tomographic imaging studies. (i) Fast 2D imaging was initially demonstrated in tissue phantoms (1% Liposyn solution) and in vitro (minced chicken breast and 1% Liposyn). A 0.45 cm3 fluorescent target at 1:0 contrast ratio was detectable up to 2.5 cm deep. Fast 2D imaging experiments performed in vivo with healthy female subjects also detected a 0.45 cm3 fluorescent target superficially placed ∼2.5 cm under the breast tissue. (ii) Coregistered imaging was automated and validated in phantoms with ∼0.19 cm error in the probe’s positional information. Coregistration also improved the target depth detection to 3.5 cm, from multi-location imaging approach. Coregistered imaging was further validated in-vivo , although the error in probe’s positional information increased to ∼0.9 cm (subject to soft tissue deformation and movement). (iii) Three-dimensional tomography studies were successfully demonstrated in vitro using 0.45 cm3 fluorescence targets. The feasibility of 3D tomography was demonstrated for the first time in breast tissues using the hand-held optical imager, wherein a 0.45 cm3 fluorescent target (superficially placed) was recovered along with artifacts. Diffuse optical imaging studies were performed in two breast cancer patients with invasive ductal carcinoma. The images showed greater absorption at the tumor cites (as observed from x-ray mammography, ultrasound, and/or MRI). In summary, my dissertation demonstrated the potential of a hand-held optical imager towards 2D breast tumor detection and 3D breast tomography, holding a promise for extensive clinical translational efforts.

MIMU sensors for joint kinematic estimation: anatomical landmarking and frame correction

The ability to capture human motion allows researchers to evaluate an individual’s gait. Gait can be measured in different ways, from camera-based systems to Magnetic and Inertial Measurement Units (MIMU). The former uses cameras to track positional information of photo-reflective markers, while the latter uses accelerometers, gyroscopes, and magnetometers to measure segment orientation. Both systems can be used to measure joint kinematics, but the results vary because of their differences in anatomical calibrations. The objective of this thesis was to study potential solutions for reducing joint angle discrepancies between MIMU and camera-based systems. The first study worked to correct the anatomical frame differences between MIMU and camera-based systems via the joint angles of both systems. This study looked at full lower body correction versus correcting a single joint. Single joint correction showed slightly better alignment of both systems, but does not take into account that body segments are generally affected by more than one joint. The second study explores the possibility of anatomical landmarking using a single camera and a pointer apparatus. Results showed anatomical landmark position could be determined using a single camera, as the anatomical landmarks found from this study and a camera-based system showed similar results. This thesis worked on providing a novel way for obtaining anatomical landmarks with a single point-and-shoot camera, as well aligning anatomical frames between MIMUs and camera-based systems using joint angles.

Axis Mapping: The Estimation of Surface Orientations and its Applications in Vehicle Localization and Structural Geology

The map representation of an environment should be selected based on its intended application. For example, a geometrically accurate map describing the Euclidean space of an environment is not necessarily the best choice if only a small subset its features are required. One possible subset is the orientations of the flat surfaces in the environment, represented by a special parameterization of normal vectors called axes. Devoid of positional information, the entries of an axis map form a non-injective relationship with the flat surfaces in the environment, which results in physically distinct flat surfaces being represented by a single axis. This drastically reduces the complexity of the map, but retains important information about the environment that can be used in meaningful applications in both two and three dimensions. This thesis presents axis mapping, which is an algorithm that accurately and automatically estimates an axis map of an environment based on sensor measurements collected by a mobile platform. Furthermore, two major applications of axis maps are developed and implemented. First, the LiDAR compass is a heading estimation algorithm that compares measurements of axes with an axis map of the environment. Pairing the LiDAR compass with simple translation measurements forms the basis for an accurate two-dimensional localization algorithm. It is shown that this algorithm eliminates the growth of heading error in both indoor and outdoor environments, resulting in accurate localization over long distances. Second, in the context of geotechnical engineering, a three-dimensional axis map is called a stereonet, which is used as a tool to examine the strength and stability of a rock face. Axis mapping provides a novel approach to create accurate stereonets safely, rapidly, and inexpensively compared to established methods. The non-injective property of axis maps is leveraged to probabilistically describe the relationships between non-sequential measurements of the rock face. The automatic estimation of stereonets was tested in three separate outdoor environments. It is shown that axis mapping can accurately estimate stereonets while improving safety, requiring significantly less time and effort, and lowering costs compared to traditional and current state-of-the-art approaches.

Shaping the Dicot Fruit: Molecular and Genomic Approaches to Fruit Development

The fruit is one of the most complex and important structures produced by flowering plants, and understanding the development and maturation process of fruits in different angiosperm species with diverse fruit structures is of immense interest. In the work presented here, molecular genetics and genomic analysis are used to explore the processes that form the fruit in two species: The model organism Arabidopsis and the diploid strawberry Fragaria vesca. One important basic question concerns the molecular genetic basis of fruit patterning. A long-standing model of Arabidopsis fruit (the gynoecium) patterning holds that auxin produced at the apex diffuses downward, forming a gradient that provides apical-basal positional information to specify different tissue types along the gynoecium’s length. The proposed gradient, however, has never been observed and the model appears inconsistent with a number of observations. I present a new, alternative model, wherein auxin acts to establish the adaxial-abaxial domains of the carpel primordia, which then ensures proper development of the final gynoecium. A second project utilizes genomics to identify genes that regulate fruit color by analyzing the genome sequences of Fragaria vesca, a species of wild strawberry. Shared and distinct SNPs among three F. vesca accessions were identified, providing a foundation for locating candidate mutations underlying phenotypic variations among different F. vesca accessions. Through systematic analysis of relevant SNP variants, a candidate SNP in FveMYB10 was identified that may underlie the fruit color in the yellow-fruited accessions, which was subsequently confirmed by functional assays. Our lab has previously generated extensive RNA-sequencing data that depict genome-scale gene expression profiles in F. vesca fruit and flower tissues at different developmental stages. To enhance the accessibility of this dataset, the web-based eFP software was adapted for this dataset, allowing visualization of gene expression in any tissues by user-initiated queries. Together, this thesis work proposes a well-supported new model of fruit patterning in Arabidopsis and provides further resources for F. vesca, including genome-wide variant lists and the ability to visualize gene expression. This work will facilitate future work linking traits of economic importance to specific genes and gaining novel insights into fruit patterning and development.

Eulerian-Lagrangian definition of coarse bed-load transport: Theory and verification with low-cost inertial measurement units

Fluvial sediment transport is controlled by hydraulics, sediment properties and arrangement, and flow history across a range of time scales. This physical complexity has led to ambiguous definition of the reference frame (Lagrangian or Eulerian) in which sediment transport is analysed. A general Eulerian-Lagrangian approach accounts for inertial characteristics of particles in a Lagrangian (particle fixed) frame, and for the hydrodynamics in an independent Eulerian frame. The necessary Eulerian-Lagrangian transformations are simplified under the assumption of an ideal Inertial Measurement Unit (IMU), rigidly attached at the centre of the mass of a sediment particle. Real, commercially available IMU sensors can provide high frequency data on accelerations and angular velocities (hence forces and energy) experienced by grains during entrainment and motion, if adequately customized. IMUs are subjected to significant error accu- mulation but they can be used for statistical parametrisation of an Eulerian-Lagrangian model, for coarse sediment particles and over the temporal scale of individual entrainment events. In this thesis an Eulerian-Lagrangian model is introduced and evaluated experimentally. Absolute inertial accelerations were recorded at a 4 Hz frequency from a spherical instrumented particle (111 mm diameter and 2383 kg/m3 density) in a series of entrainment threshold experiments on a fixed idealised bed. The grain-top inertial acceleration entrainment threshold was approximated at 44 and 51 mg for slopes 0.026 and 0.037 respectively. The saddle inertial acceleration entrainment threshold was at 32 and 25 mg for slopes 0.044 and 0.057 respectively. For the evaluation of the complete Eulerian-Lagrangian model two prototype sensors are presented: an idealised (spherical) with a diameter of 90 mm and an ellipsoidal with axes 100, 70 and 30 mm. Both are instrumented with a complete IMU, capable of sampling 3D inertial accelerations and 3D angular velocities at 50 Hz. After signal analysis, the results can be used to parametrize sediment movement but they do not contain positional information. The two sensors (spherical and ellipsoidal) were tested in a series of entrainment experiments, similar to the evaluation of the 111 mm prototype, for a slope of 0.02. The spherical sensor entrained at discharges of 24.8 ± 1.8 l/s while the same threshold for the ellipsoidal sensor was 45.2 ± 2.2 l/s. Kinetic energy calculations were used to quantify the particle-bed energy exchange under fluvial (discharge at 30 l/s) and non-fluvial conditions. All the experiments suggest that the effect of the inertial characteristics of coarse sediments on their motion is comparable to the effect hydrodynamic forces. The coupling of IMU sensors with advanced telemetric systems can lead to the tracking of Lagrangian particle trajectories, at a frequency and accuracy that will permit the testing of diffusion/dispersion models across the range of particle diameters.

Information or context : what accounts for positional proximity between the European Commission and lobbyists?

Peer reviewed