Dementia will affect 1 in 3 people in their lifetime. The inexorable decline in mental function, mood and movement ability comes about because of damage to synapses - the connections between nerve cells. Synapses come in many varieties and particular types are affected as dementia progresses. By finding and tracking these damaged synapses we can understand how the brain is damaged in dementia and help to bring on improvements in the diagnosis and treatment of these diseases. We have recently developed ground-breaking technology that enables us to examine billions of individual synapses in the human brain and discover how the are damaged. We will now drive this technology forward to discover the synapses that are damaged in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). ALS, or motor neuron disease, is a rapidly progressive, fatal condition which overlaps clinically, genetically and pathologically with frontotemporal dementia (FTD). ALS-FTD are the commonest forms of neurodegenerative disease in people under 65 years of age. Our team of scientists from the UK and Japan will use several state-of-the-art microscopy methods to examine synapses in post-mortem brain tissue obtained from individuals who have had their behavioural features examined during life. This will potentially enable us to identify the synapses that when damaged cause speech and language impairments, deterioration in mood and emotions, and movement disorders. The tools and knowledge from our program will inform on the use of brain imaging methods in the clinic and trials of therapies aimed at preventing the progression of ALS-FTD and other dementias. Our findings will also provide valuable data resources that can be exploited by the international scientific community to advance our understanding of the brain and its diseases,

Light microscopy plays an indispensable role in modern biomedical research. By providing the ability to visualise specific processes within healthy and diseased cells, often in real-time, this technique has been a driving force in increasing our knowledge of biological processes. This knowledge is, of course, essential for developing new strategies for the diagnosis and treatment of specific human diseases. Until recently, light microscopy has been hampered by the so-called "diffraction limit". This is a theoretical limit to how far two objects need to be apart before they can be resolved by the microscope, and is typically in the region of half the wavelength of visible light (~ 250 nanometers (nm), or 1/4000th of a millimetre). Many important biological structures within our cells are smaller than 250nm and therefore remain poorly characterised. These structures include the "organelles" that perform specific cellular functions and the membrane-bound carriers and filaments along which trafficking occurs, as well as infectious viruses. Several microscopy techniques have been developed in the last few years that allow the diffraction limit to be bypassed, thereby providing access to previously unappreciated processes within cells. These 'super resolution" (SR) methods-some of which allow specific molecules to be localised with a precision of up to ten times greater than conventional microscopy-promise to revolutionise biomedical research, particularly when combined with other techniques in which the UK has pre-established expertise. Two MRC-funded research organisations in Cambridge-the Laboratory of Molecular Biology (LMB) and Mitochondrial Biology Unit (MBU)-are proposing to jointly establish a multi-user centre of innovation in applied super resolution (SR) optical microscopy. Scientists at LMB and MBU have made, and continue to make, major discoveries into fundamental biological processes and have frequently translated them into commercial and therapeutic successes. There is a remarkable track record of studying biological processes at the scale of the structure of individual proteins and protein complexes, as well as at the cellular level. SR platforms would close the gap between these molecular and cellular studies, allowing multi-scale analysis of cellular processes and the relationship to disease in a world-renowned research environment. The critical importance of light microscopy to LMB and MBU is demonstrated by the presence of > 20 highly used confocal or wide-field microscopes, until recently the highest quality optical microscopes available. Ready access to SR microscopy will make a major impact on the productivity of the organisations. Indeed, 26 groups within the LMB and MBU have projects that require specific SR platforms. Their needs can only be fulfilled by providing three different, leading SR systems: structured illumination, single molecule localisation microscopy (e.g. PALM/STORM) and stimulated emission depletion (STED) microscopy. These systems have non-overlapping strengths and different projects require different systems. Substantial added value will be provided by (i) capitalising on the highly successful research programmes and complementary technological expertise within the partner organisations, (ii) building on pioneering biotechnology developed at LMB to develop superior labelling methods for SR techniques, (iii) strong collaborations with industrial partners and (iv) provision of access to other, local users to facilitate their research and foster collaborations with LMB and MBU. The project is sustainable. It builds on well-established infrastructure for management of microscopy resources and user training, and includes early access to new developments through the establishment of industrial partnerships and a significant financial commitment from LMB and MBU. There is also a strong component of cultivating the next generation of scientists by providing training in SR techniques.

Abstracts are not currently available in GtR for all funded research. This is normally because the abstract was not required at the time of proposal submission, but may be because it included sensitive information such as personal details.

The movement of cells is extremely important in many biological processes including the development of embryos, wound healing, immunity and cancer. Our research group in the School of Life Sciences at the University of Dundee is studying the way cells move, interact and signal to one another during these processes. One way to study such phenomena is to observe the cells in the microscope. Specific components of the cell are visualised by labelling them with fluorescent molecules that make them glow in the dark. In this way the role of individual components of the cell can be studied in living cells, by labelling more than one component with different fluorescent colours the interactions between the components can be studied. Cells labelled with these fluorescent markers often have a very high background fluorescence which makes it hard to see the detailed movement of fine structures within the cells. However, using a new type of optical system attached to the microscope (TIRF optics) it is possible to look only at a very thin part of the cell, at its base, where it attaches to its culture dish. The advantage this gives us is that it eliminates the fluorescent haze and allows us to see the detailed changes that occur within the living cell. We will use this type of microscope to study several different cell types and many different cell components involved in regulating cell movement. Included in our research programme are important cells of the immune system (Dendritic cells) that alert our body to invasions of bacteria and parasites. Movement of cells in developing embryos will also be studied; these movements are essential in determining the normal layout of organs in the body. A protein that is affected in colon cancer (APC) will be studied using a cell culture model of migrating cells; this mimics some aspects of cell movement in the colon. Using TIRF optics we can visualise single molecules in cells; this will be used to study how cells signal to one another from the outside to bring about reorganisations inside the cell. Finally components of the cell can be isolated, fluorescently labelled and reconstituted in a cell-free culture system; this approach will be used to study components of the cell important during cell division, again a mechanism important in cancer.

The mechanical properties of tissue, such as stiffness, have long been known to be an important indicator of the health of tissue. Indeed, manual palpation is used regularly by clinicians as part of routine diagnostic procedures. A range of techniques have been developed in order to obtain images, both in two and three dimensions, of tissue stiffness. This approach, known as elastography, is significantly more powerful than manual palpation as the clinician is able to visualise the location, and relative stiffness, of tissue within the body. Existing elastography techniques are based on ultrasonoghraphic imaging (i.e. ultrasound, US), magnetic resonance imaging (MRI) and optical imaging. US based elastography has developed to the point where it is able to be used as part of a routine breast cancer diagnosis protocols. One of the problems with these established techniques is that their ability to resolve small features, such as small tumours, degrades as these tumours are located within the body. For example, optical techniques are able to resolve features as small as 0.02mm, however, only if they are located within about 1mm of the skin. Tumours over 0.1mm in size can be detected using US elastography if they are located less than 10cm from the skin surface, and so on. We propose to make use of a recently developed X-ray imaging technique, known as phase imaging, which provides images of biological tissue which are much clearer than conventional X-ray imaging. Using this technique we believe it will be possible to obtain images of tissue stiffness, resolving features as small as 0.02mm, located deep within the body, thus breaking the limitation of existing elastography techniques. Although applicable to a range of applications, we will apply this technique to breast cancer imaging. If successful, the technique could be applied in screening, diagnosis and treatment of breast cancer. In particular, it could be integrated into a form of three-dimensional mammography known as tomosynthesis, giving the radiologist an additional form of contrast upon which to make their diagnosis. Once a suspicious lesion has been detected at the screening stage, this technique could be used in follow up imaging, again to provide the radiologist with an additional form of contrast upon which to make their diagnosis. Finally, if surgery is required, it may be possible to use this technique to check that the tumour has been completely removed at the time of surgery, rather than requiring a patient to return for follow-up surgery to remove any remaining tumour.