Title: Characterisation and validation of an organotypic slice culture model of Alzheimer’s disease
Funder: National Centre for the Replacement, Refinement and Reduction of Animals in Research

Abstract: Alzheimer’s disease (AD) is a progressive neurodegenerative disorder affecting around 35 million people worldwide and costing approximately £315bn annually in treatment and care. Due to an ageing population, the prevalence of AD is predicted to double every 20 years. The majority of investigations into basic mechanisms underlying AD and new therapeutic strategies to prevent disease progression use genetically modified mice to model disease. Such mice frequently develop aggressive phenotypes and display substantial neurological impairments with aging. Typically, biochemical investigations rely on analysis of brains from individual animals sacrificed at different ages. Similarly, due to the severity of invasive in vivo physiological measurements, different groups of mice are usually used in longitudinal studies of synaptic function. Here, we aim to replace, reduce and refine the number of animals used to study mechanisms underlying AD by characterising and fully validating an ex vivo model of AD. Brain slices will be prepared from 3xTg-AD mice, which recapitulate the primary neuropathological, electrophysiological and behavioural features of human disease. We plan to establish and culture organotypic slices from 3xTg-AD mice and maintain them in culture for periods of up to 6 months. We will assess the development of biochemical measures of disease severity, including tau phosphorylation and aggregation together with beta-amyloid (Aβ) production and deposition. We will also determine the degree of synaptotoxicity in this model of AD. The presence and severity of these phenotypic abnormalities will be carefully related to those observed in vivo to determine the extent to which slices mimic the development of disease in intact living mice, and thus act as a valid and sensitive replacement. Following a complete characterisation of the slice cultures, we will validate their use in drug discovery by determining the impact of disease-modifying therapies that have previously been investigated in 3xTg-AD mice. Such approaches include inhibitors of tau phosphorylation and Aβ production, anti-inflammatory agents, microtubule-stabilising drugs and NMDA receptor antagonists. Since 36 slices are obtained from an individual mouse brain, we will have the capacity to assess several different treatment conditions in a single animal. This significantly reduces both the number of animals required and also the experimental variation inherent in multiple animal testing. Furthermore, development of an AD phenotype ex vivo precludes the need for moderate to substantial severity testing of later stages of disease in living animals and the use of invasive physiological procedures, thereby significantly refining animal use for basic AD research. It is anticipated that the success of this project will lead to the wider adoption of organotypic slice cultures for investigations into a wide spectrum of neurological conditions in both academia and industry and therefore this study has high potential to generate a significant impact for the principles of the NC3Rs


Research activity codes: 2.6 RESOURCES AND INFRASTRUCTURE (AETIOLOGY);5.9 RESOURCES AND INFRASTRUCTURE (TREATMENT DEVELOPMENT)
Health categories: NEUROLOGICAL