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Project grant

Applying the 3Rs to elucidate the mechanisms of tau pathology using DRG neurons in culture

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At a glance

Completed
Award date
January 2014 - December 2016
Grant amount
£301,421
Principal investigator
Professor Maria Grazia Spillantini

Co-investigator(s)

Institute
University of Cambridge

R

  • Reduction
  • Replacement
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Contents

Overview

Aims

This research aims to develop and characterise an in vitro model of neuronal cell death in response to accumulation of the tau protein, reducing and replacing the use of live animal models in the study of the molecular basis of neurodegeneration and dementia.

Background

In Alzheimer's disease the neuronal protein, tau, becomes misfolded, forming neurofibrillary tangles (NFT), thought to be critical for neurodegeneration in the disease, making tau of interest as a therapeutic target. However, how tau causes neurons to die is still an open question, and dying cells in the brain are not easily accessible for study. Current cellular models of ‘tauopathy’ fail to exhibit NFT formation and subsequent neuronal death, meaning animal models are the only currently viable model for study.

This research aims to develop and characterise a cell culture system where neurons develop pathological forms of tau and go on to die, allowing the molecular mechanisms underlying this and potential therapies to be investigated in vitro, minimising the use of animals and associated suffering with advanced disease pathology.

Preliminary data suggests that dorsal root ganglia (DRGs) neurons from adult transgenic P301S human mutant tau mice can be cultured for at least eight weeks during which they develop pathological forms of tau and go on to die. Up to 12 replicate cultures can be obtained from a single mouse prior to the stage when the animals develop severe pathology.

Research details and methods

Long lived DRG neuronal cultures will be prepared from a small number of adult transgenic mice at different stages of tau pathology. These cultures will be characterised to elucidate the mechanisms by which tau positive neurons die and to probe the causes of this cell death by studying the function of cellular organelles, such as mitochondria, and structural proteins called microtubules.

Whether inflammatory cytokines or immune cells, such as macrophages and microglia, exacerbate the pathology of the disease will be investigated. Finally, neuronal cell cultures will be developed as a platform for testing drugs that potentially decrease various stages of tau pathology.

This research could offer a new option, with distinct scientific advantages such as allowing neuronal death related to tau pathology to be visualised, probed and monitored. Furthermore, it would allow for much more rapid screening of potential therapies which impact tau and NFT formation, preventing neuronal death, hastening the drug development process. Alongside the scientific advantages, it is estimate that the in vitro model could reduce animal use around eight-fold. 

Publications

  1. Tolkovsky AM and Spillantini MG (2021). Tau aggregation and its relation to selected forms of neuronal cell death. Essays in Biochemistry 65(7): 847–857. doi: 10.1042/EBC20210030
  2. Brelstaff J et al. (2018). Living Neurons with Tau Filaments Aberrantly Expose Phosphatidylserine and Are Phagocytosed by Microglia. Cell Reports 24(8):1939-1948.e4. doi: 10.1016/j.celrep.2018.07.072
  3. Tolkovsky AM and Brelstaff J (2018). Sensory Neurons from Tau Transgenic Mice and Their Utility in Drug Screening. In: Skaper S. (eds) Neurotrophic Factors. Methods in Molecular Biology, vol 1727. Humana Press, New York, NY doi: 10.1007/978-1-4939-7571-6_7
  4. Sidoryk-Wegrzynowicz M et al. (2017). Astrocytes in mouse models of tauopathies acquire early deficits and lose neurosupportive functions. Acta Neuropathologica Communications 5:89. doi :10.1186/s40478-017-0478-9
  5. Brelstaff J et al. (2015). The fluorescent pentameric oligothiophene pFTAA identifies filamentous tau in live neurons cultured from adult P301S tau mice. Frontiers in Neuroscience 9:184. doi: 10.3389/fnins.2015.00184
  6. Brelstaff J et al. (2015). pFTAA - a high affinity oligothiophene probe that detects filamentous tau in vivo and in cultured neurons. Neural Regeneration Research 10(11):1746-7. doi: 10.4103/1673-5374.165298
  7. Goedert M et al. (2015). A Brief History of Tau. Clinical Chemistry 61(11):1417-8. doi: 10.1373/clinchem.2015.245142