The Dynamic Embryo Imaging Platform

Understanding the biology of the cell and its component parts has always been the key goal in life science and medicine. Advances in microscopy are profoundly changing the way cells, sub-cellular structures and tissues can be studied. For a research intensive institution, such as UCL, to remain a competitive leader in biomedical sciences, it is essential to adopt transformative imaging technologies. Furthering our understanding of health and diseases requires a deep knowledge of the cell, its constituents and their development from very early stages of life. How to image intact, large-scale, live organisms at high spatial and temporal resolution with multiple fluorophores/reporters without phototoxcity or photobleaching with detectors of suitable sensitivity to capture low signal intensity remains a key challenge.
UCL's international reputation as a key contributor in bioscience/biomedical discoveries has, in part, been successful because of it's sustained commitment to be an early adopter of new, transformative technologies, by investment of capital and resources. One main path toward this goal is to image the behavior and functional properties of cells over time. This must be done with equipment that is capable of capturing images with sufficient fine detail and at very high speed so that, hitherto, unknown dynamic processes can be discovered and studied. Lightsheet imaging technology is particularly suited to the study of key events during embryogenesis, especially fast cellular events, in part because of its ability to scan through living tissue and construct three-dimensional images of developing organisms at high speed. Advanced optical techniques, such as lightsheet microscopy allow for such dynamic studies while also allowing researchers to manipulate cellular functions.

The addition of this technology to UCL core Imaging faculty will give researches access to capabilities such as:
a) Rapid feature tracking to study intracellular process, cell migration and embryo development.
b) Address optical scattering problems that are associated with other imaging techniques and allow imaging deep into thick samples.
c) Rapid image capture of cell signalling events such as calcium imaging, deep into whole embryos regions such as the CNS
d) To detect molecular interaction events within and between cells in live tissues and whole organisms such as zebra fish embryos.

The main co-applicants on this proposal are, therefore, research groups at UCL whose focus is on aspects of embryo development trying to elucidate the fundamental processes and key events/factors that underlie organismal development and in health disease. They represent some of the major research figures at UCL, whose research is extensively supported through BBSRC, MRC, CRUK and Wellcome Trust grants. In addition, the microscope will be accessible to a large number of diverse research projects addressing many outstanding questions relating to embryonic development, cell biology and neurobiology. The requested funds will enable us to obtain a one of the leading Lightsheet microscope that will significantly enhance our research capabilities and lead to new scientific discoveries.

Many researchers within the Faculty of Life Sciences require access to state-of-the-art imaging equipment for their studies. However, current core access equipment in the Centre for Cell and Molecular Dynamics (CCMD), and indeed across UCL, lacks sensitivity and the temporal resolution required for studying dynamic physiological intracellular events over extended time periods in living embryos/larvae/organ-slices or during neural development. The enhanced sensitivity and speed of acquisition in new camera technology and the geometry of the light generated by the lightsheet microscope allows a significantly reduction in laser power to the sample, and at the same time, deep penetration to image many hundreds of microns inside large samples. This eliminates the effects of phototoxicity on the living organism, which represents a major problem for long-term time-lapse imaging, particularly in developing embryos, but also in live tissue slices and 3D cultures cells. These properties provide a major advantage for understanding most cell biological processes, including the localization and redistribution of proteins, ion signalling in cells and organismal development at high temporal resolution (>100 frames/s), with the requisite spatial resolution to track individual cellular behaviours and cell fates for long periods. It is essential that our research groups have open and easy access to the equipment requested in this proposal. We aim to deliver rapid, high temporal and spatial resolution imaging deep into tissues for 3-D imaging in live zebrafish, Xenopus embryos, embryonic mouse brains, organs/organiods and 3D cell co-cultures. This will enable us to offer the UCL scientific community-
a) Rapid feature tracking to study cell migration and embryo development
b) Address optical scattering problems that are associated with other imaging modalities by using novel illumination and image reconstruction strategies
c) To implement FRET readouts of cell signalling networks

The assembled multidisciplinary team encompasses a diverse field of scientist. They will use a wide verity of model systems to investigate their particular research questions by capitalising on the unique capabilities of Lightsheet Microscopy. The areas of expertise of the groups are mainly in the field of biosciences therefore the potential Impact of these studies could be significant for a number of stake holders. The instrumentation will facilitate a wealth of on-going research projects across the whole of UCL, not only for the main applicants and will inevitably lead to more scientific discoveries with the consequential effects these discoveries will impact on human health and disease.

The outcomes of these projects will transfer knowledge and the tools generated using in vivo approaches to a wide scientific audience, both in academia and industry and may eventually impact on scientific discovery. The key objectives of the proposed projects are:
1) develop new biosensors to measure tension at tight junctions that will be useful to study the role of junctional force transduction in epithelial and endothelial cells during different physiological, pathological and developmental processes.
2) develop new 3-D models of intracellular calcium signalling for discovering novel anti-cancer therapies.
3) make new discoveries in morphogenesis and organismal/animal development.
4) apply new methods and models systems for studying brain development and cell-cell interactions that underlie neurological disorders using Xenopus embryos.
5) elucidate cell cycle events that are driven by circadian clock function in live zebrafish emryos.
6) reveal and quantify the key contributing events in cell signalling in neural circuits of visually guided behaviors.
7) elucidate common molecular pathways in cortical interneuron migration and angiogenesis that are involved in developmental brain disorders.

We expect that this research will also affect society as a whole in the long-term by influencing approaches of regenerative medicine, tissue engineering, cancer therapeutics, cognitive function and morphogenesis. It is important, however, that our impact activities also target academic audiences, as further research and application of our findings will be essential on the pathway to economic and societal impact. Moreover, the outcomes of the proposed research will not only support BBSRC research priorities but will also contribute to training and capacity building and, hence, BBSRC enabling themes.

Milestones and deliverables:
We do not anticipate any major discoveries during the 12 months timeline of the grant call. However, the equipment will continue to serve the applicants and other UCL researchers for at least 5 years and probably 10 years. We fully expect that some of the outcomes to have direct and measurable impact for the scientific community within 2 - 4 years (Objectives 1, 2, 4, 7). We anticipate that in the mid- to long-term (5 - 7years) some of the research outcomes will have societal and economic impacts (objectives 1, 2 and 7).