Abeta-mediated toxicity in Alzheimer's disease: delineating mechanisms of internalisation, cell-cell transmission and synaptic dysfunction

Alzheimer's disease (AD) is a devastating illness that affects 1% of the population. Although some familial forms of AD exist, most Alzheimer's patients suffer from sporadic, late-onset AD, associated with ageing. The causes of Alzheimer's disease remain unclear, although it is now well-established that in brain cells (neurons), abnormal folding of small proteins (called Abeta and tau) represents a pivotal step in disease pathology. Our work aims to understand how these proteins can lead to the deterioration of neuronal function underlying the degenerative symptoms seen in Alzheimer's sufferers. To examine this we will use both artificially-grown and native networks of living neurons and recapitulate the changes occurring at the onset of AD as the target proteins change and become toxic. We will utilise state-of-the-art imaging methods to directly visualise the effects on basic neuronal function. Our research will also examine how the spread of toxic proteins between neurons might underlie the progression of the disease. A specific consequence of AD is the gradual dysfunction of synapses - the sites for neuron-to-neuron signalling - that are essential for the brain's ability to process information. We will use specialised fluorescence markers to examine synaptic signalling events and characterise the specific mechanisms which become disrupted. Finally, we will combine our fluorescence measurements with novel state-of-the-art microscopy experiments, allowing us to examine the detailed structural changes in neurons caused by misfolding proteins at high-resolution. Using this approach, we anticipate major new insights into understanding the critical cellular and molecular changes associated with Alzheimer's disease. With this knowledge, the rational design of therapeutic agents that can target these processes and alleviate or prevent disease symptoms becomes a realistic prospect.

Alzheimer's disease is characterised by the abnormal association of Abeta (Ab) peptide to form oligomeric species, with catastrophic consequences for neuronal function and survival. However, clear understanding of the mechanisms by which Ab exerts toxicity remains elusive. Specifically, it is not known how Ab is transmitted around neural networks, how it internalises/accumulates in neurons, the nature of its cellular/molecular targets, or the mechanisms which underlie its disruption of critical neuronal synaptic processes. This proposal will exploit established and novel biochemical, imaging and ultrastructural approaches in cultured and native hippocampal neurons to examine these issues, providing fundamental new insights into the toxic mechanisms of Ab action.

A key aim is to translate our recent findings from neuroblastoma cells, defining a pathway of Ab internalisation, into primary neurons. Ab will be tracked alongside cellular/organelle markers to examine endocytic processes and targeting to specific compartments, and followed by ultrastructural investigation. Ab transmission around neuronal networks will be assayed with genetically-encoded fusion constructs providing insight into the spreading of Ab pathology in Alzheimer's patients. We will also examine Ab consequences on presynaptic function - specifically dynamic properties of vesicle pools. Our pilot data indicates striking Ab-induced effects on functional pool partitioning, lateral vesicle trafficking and vesicle endocytic efficiency. Alongside culture experiments we will use an established transgenic model to explore synapse-specific presynaptic deficits in acute slice. This approach also permits a function-ultrastructure readout of recycling vesicles in native tissue, offering unique insights into Ab-induced vesicle pool re-organisation. Our findings will shed new light on pivotal steps in the pathway of Ab action, providing important potential targets for the rational development of therapeutics.

This proposal aims to understand the molecular mechanisms underlying the toxicity of Abeta in Alzheimer's disease. The prevalence of Alzheimer's is growing as the ageing population increases. Therapies are available, but these slow disease progress rather than halting it. The need for more fundamental research into underlying mechanisms is highlighted by the recent withdrawal from further development of a promising therapeutic agent, Bapineuzumab. Therapeutic design is hampered by a lack of clear insight into the disease process. In this work, we aim to provide a better understanding of the underlying disease mechanisms and our long term goal is to identify key targets for rational drug design that will lead to an improvement both for society and for the individual patient.

1. Who will benefit from this research?
This research will provide a clearer understanding of the key processes involved in Alzheimer's disease. This will impact in two major ways: First, it will provide the scientific basis for potential targets for drug development; Second, it will provide a clearer knowledge regarding early changes associated with disease development and therefore contribute to the possibility of early diagnosis. If Alzheimer's disease is to be combated, it is essential that any therapeutic is provided prior to irreversible neuronal cell death. This research will provide information for pharmaceutical companies developing therapeutics agents for Alzheimer's disease and in the longer term this will impact on the health service and wider public. In the past few years, clear statements have been made regarding the priority for research into neurodegenerative diseases from the UK Government and the EU, as well as the USA. This research project is likely to inform policy makers in terms of specific investment in research into the most important targets for Alzheimer's disease.

2. How will they benefit?
In the long term, this research will contribute to the enhancement of public health by adding to understanding aimed at improving treatment for Alzheimer's disease. This research will be disseminated in peer-review journals, at international meetings and also communicated to the public via Café Scientifique and the media. The research group has links with Pharma via collaboration with the translational drug discovery group recently established at the University of Sussex. Pharmaceutical companies will benefit from the clearer understanding of the disease process and potential targets on which to base design and development of drugs for Alzheimer's disease. Clinicians working with Alzheimer's patients will have a foundation for researching early diagnosis, since our findings will report on initial changes that occur in response to elevated toxic Abeta. The broader public will gain longer term benefits from improvements in treatments for Alzheimer's disease and this in turn will impact on the economy and wealth and health of the nation. The research may be used to produce therapeutics that slow or halt the disease progression, providing enhanced quality of life for those suffering from Alzheimer's disease and those caring for patients. This is a three year grant from which we would aim to provide the basis for targets for the development of therapeutics. The realistic expectation for the impact is that in the next 10 years, drugs may be developed based on our results.

Professional development
The work will deliver and train highly-skilled researchers, with expertise in organization, analysis, communication and scientific writing, relevant to many employment sectors. The PDRA will undertake a broad range of techniques including novel state-of-the art function-ultrastructure approaches developed in the Staras lab - skills of benefit to the wider academic and industrial communities. Both laboratories regularly provide projects for MSc and undergraduate internships and in this way training will be disseminated to the broader community.