Implanted imaging laboratories for deep-tissue in vivo imaging

The use of in vivo microscopic imaging is widespread for fundamental research using animal models in biomedicine, for drug discovery and tracking disease progression. Deep-tissue imaging is, however, highly invasive and so termination of the animal normally occurs after each measurement. For experiments involving testing with multiple timepoints, for example for studying disease progression or cell migration, termination of an animal for each timepoint can require the use of a large number of animals to achieve a reliable research outcome. Furthermore this provides only snapshots of phenomena, hampering understanding of cell fate and function. We will develop a multi-modal, miniaturised microscope and develop techniques for surgically implanting the microscope into an animal for multimodal microscopic imaging over extended time periods. The microscope will be sufficiently small and configured to be minimally intrusive for animal comfort. For example the microscope objective will be anchored with cement to vertebrae or bones around joints and optical image guides will transmit images to detector arrays that can be remotely located within body cavities with minimal or no discomfort to the animal. The microscope can be recovered at the end of the study.
The research project will provide the following advantages that align with the aims of NC3Rs:
-For longitudinal studies there will be a clear proportionate reduction in the number of animals sacrificed.
-The improved control provided by the use of only a single animal for each longitudinal measurement will enable reliable research outcomes to be achieved with fewer animals.
-Real-time imaging during normal animal behaviour will provide for improved quality of data for certain experiments and will reduce the animal stress associated with anaesthesia.
Taking these two first aspects into account we aim to reduce the number of animals used by a factor of ten for longitudinal studies in each of three active research programmes involving spectral imaging and fluorescence imaging of the spinal cord and of joint tissue.

In vivo microscopic imaging on mice and rats is widespread and increasing: animal models are used in fundamental science, biomedicine, for drug discovery and tracking disease progression. The number of animals experimented upon continues to rise year on year; approximately 79% of the four million animals annually being rats or mice. Deep-tissue imaging is, however, highly invasive and so termination of the animal normally occurs after each measurement. In experiments involving testing with multiple timepoints, for example, the study of disease progression or cell migration, termination of an animal at each timepoint can require the use of a large number of animals to achieve a reliable research outcome. Furthermore this provides only snapshots of phenomena, hampering understanding of cell fate and function. We propose to develop in vivo imaging technology that has the promise to reduce the number of animals terminated in these experiments by a factor of ten.

We will develop a miniaturised self-contained microscope-technology platform suitable for in vivo implantation for long-term longitudinal fluorescence imaging on single animals that will enable a step-change in the approach to animal-based research. We believe that this will be the smallest microscope system yet demonstrated. The possibility of longitudinal studies on a single animal yields the promise of an overall reduction in the number of animals required to be terminated to achieve high-quality research outcomes and refining the experiments on each animal through reduced need for anaesthesia.

As an exemplar of the possible applications, we will demonstrate the technology in fluorescence microscopy of rodent spinal-cord axonal myelination in a rat model .

Introduction
The salient impact that this project aims to develop is to enable and promote a reduction and refinement in the use of animals in research. Our vision is to generate this impact through influencing the extended research community that currently conducts research on animals using invasive deep-tissue in vivo imaging and spectroscopy, through provision of our new technology that provides the advantages highlighted in the Case for Support. We summarise below how this will be accomplished

Outcomes
The main planned beneficial outcomes associated with the research project are:
- Increased demand from the research community for implanted imaging technologies in longitudinal studies.
- An increased volume of academic research in implanted technologies for longitudinal studies to enhance the provision available.
- Increased availability of technologies from industry.
Beneficiaries
The Primary beneficiaries are
- Public benefit from improved effectiveness of the publicly-funded NC3R.
- Life-science researchers through improved research efficiency and effectiveness.
- Industry, through increased opportunities for product development and sales.
Mechanisms
The primary mechanisms through which we will achieve these outcomes are dissemination to a range of communities to generate awareness and through Knowledge Exchange with companies to produce a source of the implanted microscope technology.
Dissemination
The Pathway to Impact involves dissemination of the technology and its application benefits to:
- The life-science research community to encourage a demand-pull to use this technology.
- The academic instrumentation development community research community to catalyze further development and provision of technology solutions in the wider field of implanted research laboratories for longitudinal studies.
- Industry to manufacture microscope systems and satisfy the demand.
As described in the Communications Plan, this will be achieved through
- Publication in high-profile journals
- Coverage in the media
- Public engagement
Knowledge transfer
We will work with our collaborators Gilden Photonics and STMicroelectronics to develop the technology for commercial exploitation and availability to the research community. . We expect to submit proposals for commercial development, in collaboration with our company project partners or others, at around month 24-30 to support the commercialisation of the product to make our technology widely available to the user community. There are many excellent opportunities local to Glasgow, including the Scottish Enterprise Proof-of-Concept, Royal Society of Edinburgh Enterprise Fellowships (suitable for the PDRs working on the project) and The Innovation Centre for Sensing and Imaging Systems (CENSIS), with which Harvey and Cumming are closely involved; in addition to the many UK and European commercialisation schemes funded by for example TSB and Horizon 2020 that are appropriate to this technology.
Although we expect that such a proposal will include Gilden Photonics and STMicroelectronics, there is also the possibility to bring in alternative or additional partners.