Dataset Information

Organotypic brain slice cultures of adult transgenic P301S mice--a model for tauopathy studies.

ABSTRACT:

Background

Organotypic brain slice cultures represent an excellent compromise between single cell cultures and complete animal studies, in this way replacing and reducing the number of animal experiments. Organotypic brain slices are widely applied to model neuronal development and regeneration as well as neuronal pathology concerning stroke, epilepsy and Alzheimer's disease (AD). AD is characterized by two protein alterations, namely tau hyperphosphorylation and excessive amyloid β deposition, both causing microglia and astrocyte activation. Deposits of hyperphosphorylated tau, called neurofibrillary tangles (NFTs), surrounded by activated glia are modeled in transgenic mice, e.g. the tauopathy model P301S.

Methodology/principal findings

In this study we explore the benefits and limitations of organotypic brain slice cultures made of mature adult transgenic mice as a potential model system for the multifactorial phenotype of AD. First, neonatal (P1) and adult organotypic brain slice cultures from 7- to 10-month-old transgenic P301S mice have been compared with regard to vitality, which was monitored with the lactate dehydrogenase (LDH)- and the MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays over 15 days. Neonatal slices displayed a constant high vitality level, while the vitality of adult slice cultures decreased significantly upon cultivation. Various preparation and cultivation conditions were tested to augment the vitality of adult slices and improvements were achieved with a reduced slice thickness, a mild hypothermic cultivation temperature and a cultivation CO(2) concentration of 5%. Furthermore, we present a substantial immunohistochemical characterization analyzing the morphology of neurons, astrocytes and microglia in comparison to neonatal tissue.

Conclusion/significance

Until now only adolescent animals with a maximum age of two months have been used to prepare organotypic brain slices. The current study provides evidence that adult organotypic brain slice cultures from 7- to 10-month-old mice independently of the transgenic modification undergo slow programmed cell death, caused by a dysfunction of the neuronal repair systems.

SUBMITTER: Mewes A

PROVIDER: S-EPMC3439393 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Alzheimer's disease, the most common cause of dementia, is a progressive neurodegenerative disorder characterised by amyloid-beta deposits in extracellular plaques, intracellular neurofibrillary tangles of aggregated tau, synaptic dysfunction and neuronal death. Transgenic rodent models to study Alzheimer's mimic features of human disease such as age-dependent accumulation of abnormal beta-amyloid and tau, synaptic dysfunction, cognitive deficits and neurodegeneration. These models have proven vital for improving our understanding of the molecular mechanisms underlying AD and for identifying promising therapeutic approaches. However, modelling neurodegenerative disease in animals commonly involves aging animals until they develop harmful phenotypes, often coupled with invasive procedures. We have developed a novel organotypic brain slice culture model to study Alzheimer's disease using 3xTg-AD mice which brings the potential of substantially reducing the number of rodents used in dementia research from an estimated 20,000 per year. Using a McIllwain tissue chopper, we obtain 36 x 350 micron slices from each P8-P9 mouse pup for culture between 2 weeks and 6 months on semi-permeable 0.4 micron pore membranes, considerably reducing the numbers of animals required to investigate multiple stages of disease. This tractable model also allows the opportunity to modulate multiple pathways in tissues from a single animal. We believe that this model will most benefit dementia researchers in the academic and drug discovery sectors. We validated the slice culture model against aged mice, showing that the molecular phenotype closely mimics that displayed in vivo, albeit in an accelerated timescale. We showed beneficial outcomes following treatment of slices with agents previously shown to have therapeutic effects in vivo, and we also identified new mechanisms of action of other compounds. Thus, organotypic brain slice cultures from transgenic mouse models expressing Alzheimer's disease-related genes may provide a valid and sensitive replacement for in vivo studies that do not involve behavioural analysis.