Neuronal model systems for examining toxicity

Lead Research Organisation: University of Cambridge

Abstract

In neurodegenerative diseases certain classes of highly sensitive neuronal populations die. This is also the case in Parkinson’s disease, where only dopamine neurons that are located in a region called the substantia nigra (SN) degenerate. Dopamine neurons that are located in the neighbouring ventral tegmental area (VTA) are almost not affected. Parkinson’s disease can be caused by a mutation in a gene that results in the production of a toxic protein or by neurotoxins that are present in the environment. However, it is not so clear why the dopamine neurons located in the SN are much more sensitive to environmental toxicity than VTA neurons. In our research we will investigate why only SN neurons are affected by these toxic insults. The use of human embryonic stem (ES) and induced pluripotent stem (iPS) cell technologies will allow us to culture SN and VTA dopaminergic neurons in a dish. Human ES and iPS cells are pluripotent, meaning that they have the ability to generate different cell types upon differentiation. The iPS cells that we will use are derived from patients with Parkinson’s disease Enriched cultures of SN neurons will allow us to examine specifically the neurons that degenerate in Parkinson’s disease in vitro. We will analyse the response of these neurons to toxic insults and compare them to other dopamine neurons that do not degenerate in the disease. This will give us novel insights into the selective sensitivity of SN neurons and will reveal the potential risks of environmental compounds in developing Parkinson’s disease.

Technical Summary

Neurodegenerative diseases are caused by a combination of genetic and environmental factors selectively affecting vulnerable populations of neurons. In most cases the symptoms only occur when most of the neurons have already died and there is currently no cure. Therefore, it is important to understand early processes leading to disease onset and mechanisms of toxicity caused by exposure to environmental chemicals. Midbrain dopaminergic (mDA) neurons can be broadly subdivided into two major anatomically and functionally groups, which form the substantia nigra (SN) and ventral tegmental area (VTA). Despite commonalities in gene expression and developmental origin, only SN dopaminergic neurons degenerate in Parkinson’s disease. There are several indications that environmental compounds may cause neurodegeneration in Parkinson’s disease, but in many cases the cellular events leading to toxicity are not understood. In addition, it is unclear why SN neurons are in particular susceptible to toxic insults and the neighbouring VTA neurons are relatively spared. Progress in understanding the mechanisms of toxicity and disease onset has been hampered by the lack of human relevant neuronal model systems. Human pluripotent stem (PS) cells present new opportunities in this regard, however it is important that pure populations of the relevant neuronal cell types are generated.

Our aim is to provide mechanistic insight into cellular events leading to the selective degeneration of SN dopaminergic neurons in Parkinson’s disease. We will explore mechanisms of toxicity for several classes of environmental chemicals in human ES cell derived SN dopaminergic neurons. Furthermore, we will identify cell intrinsic differences between these different populations of dopaminergic neurons that underlie their selective vulnerability.

In this proposed study we will use a PS cell based model system for exploring toxicity and disease onset in highly vulnerable populations of SN dopaminergic neurons. Our human ES and iPS cell based platform can be used to examine mechanisms of toxicity of several environmental agents in highly vulnerable neuronal populations. The enriched cultures of human ES and iPS cell derived SN and VTA dopaminergic neurons enable system biological approaches to identify pathways and proteins that mediate the cellular response to the toxic insult. In addition, this approach will give insight in cellular response pathways that are contributing to disease onset. Altogether, we have developed an ex vivo model of healthy and diseased tissue that can be used to analyse mechanisms of toxicity.

Publications

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Title established novel human ES cell differentiation protocol 
Description We have developed a novel human ES/iPS cell differentiation protocol that results in the selection of several distinct dopaminergic subpopulations 
Type Of Material Model of mechanisms or symptoms - human 
Year Produced 2018 
Provided To Others? No  
Impact Better methodology to model PD in vitro Amendable for drug discovery and toxicology research 
 
Title mouse ES cell differentiation 
Description use of extrinsic factors to direct differentiation of mouse ES cells into specific dopaminergic subpopulations 
Type Of Material Model of mechanisms or symptoms - in vitro 
Provided To Others? No  
Impact Knowledge about factors required for direction of mouse ES cells into specific dopaminergic populations can be applied to iPS and human ES cell differentiations 
 
Description collaboration with Brian Brooks 
Organisation Bethesda University
PI Contribution We have provided research data to the pending manuscript from the lab of Brian Brooks
Collaborator Contribution Brian Brooks lab has given us a mouse line and send us embryonic tissue for our analysis
Impact Discovery of skeletal phenotype in Nolz1 mutant embryos which will result in contribution to manuscript Analysis of dopaminergic axon guidance phenotype. Data has already been presented at several international conferences. Manuscript is pending
Start Year 2015
 
Description collaboration with Flaviano Giorgini 
Organisation University of Leicester
Department Department of Genetics
Country United Kingdom 
Sector Academic/University 
PI Contribution We have provided cultures of ES cell derived dopaminergic neurons that they could use for their research purposes
Collaborator Contribution We have set up the neuronal differentiation and helped with interpretation of results. In addition we have written a part of the manuscript
Impact We have submitted a manuscript
Start Year 2016
 
Description MRC festival 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact To inform public about research done at our Unit. We organised several hands-on activities
Year(s) Of Engagement Activity 2018