Function and regeneration of dopaminergic neurons in the brain of zebrafish

Lead Research Organisation: University of Edinburgh
Department Name: Centre for Discovery Brain Sciences

Abstract

Adult zebrafish have an amazing capacity to repair injuries to their brain and spinal cord. In particular, there are so-called progenitor cells in the brain that can divide and thus generate new nerve cells. In mammals, including humans, such regeneration usually fails. Nerve cells that produce the chemical dopamine, an important modulator of brain activity, are particularly vulnerable to influences of toxins, aging and disease, such as Parkinson's disease which leads to life-long disability. We plan to determine whether dopamine producing nerve cells are regenerated in the adult zebrafish and whether this repairs leads to recovery of lost functions. This would show that new nerve cells make useful connections in the brain network. Importantly, we will elucidate the genes that are activated during regeneration and which molecular signals induce progenitor cells to generate dopamine producing nerve cells in zebrafish. In particular, we will develop pharmacological and genetic tools to follow regeneration of these cells over time. This will help us determine at a fundamental level how it is possible to replace lost nerve cells in the adult and fully wired nervous system. Moreover, many of the molecular signals and progenitor cells are also present in the brain of mammals, such that our project will also shed light on the specific mechanisms missing in mammals to achieve successful replacement of degenerating nerve cells.

Technical Summary

In contrast to mammals, adult zebrafish regenerate neurons and axons in the CNS. The dopaminergic (DA) system is evolutionarily conserved, with neuronal somata situated mostly in the forebrain and axons reaching through the CNS. Thus the DA system in zebrafish offers an excellent opportunity to study replacement of a specific, highly connected cell type in the adult CNS in a genetically accessible model vertebrate. Moreover, the zebrafish is being developed as a model for neurodegeneration of DA neurons. However, surprisingly little is known about regeneration of this cell type in fish. We aim to determine the regenerative capacity of DA neurons in the CNS of zebrafish after selective ablation of these cells by genetic and pharmacological means. We will analyse changes in gene expression, functional recovery and signals that promote regeneration of DA neurons. In this context, we will explore the role of dopamine itself, which has been described as a negative regulator of DA neuron regeneration in salamanders (Berg et al., 2011, Cell Stem Cell, 8, 426-33), but has been found by us to augment neuronal regeneration from spinal progenitor cells in zebrafish (Reimer et al., 2013, Dev Cell 25, 478-91). Our preliminary results indicate that toxin-induced ablation of specific populations of DA neurons leads to microglia activation, a loss of the entire DA projection to the spinal cord and to robust deficits in shoaling behaviour, but not in basic swimming capacity. Within weeks, numbers of DA neurons are restored. Elucidating the mechanisms and signals of dopaminergic cell regeneration in the CNS of adult zebrafish will help us to understand the inability of the mammalian brain to regenerate this type of neurons.

Planned Impact

Exploitation and Application: In this project, we aim to better understand fundamental mechanisms of neuronal replacement in the fully differentiated adult vertebrate CNS. This fundamental research may inform (therapeutic) cell replacement approaches in the long term. No immediate commercial output is foreseen, however, we will liaise with "Edinburgh Research and Innovation" (ERI), which ensures early identification of commercial potential. Our collaboration with ERI has led to a patent application and IP development funding for our group.
Dopaminergic neurons in the striatum are lost in Parkinson's Disease. We will liaise with Parkingson's UK's CSO Dr. Kieran Breen and attend/present at their meeting to maximise exchange between their interests and our findings.

Collaboration: Our project will strengthen scientific collaborations with Prof Douglas Armstrong and Dr. Dirk Sieger in Edinburgh, expanding the range of phenomena we can investigate in the adult fish by making full use of the computational tools Prof Armstrong has developed/is developing for analyses of complex in vivo behaviours and the tools to analyse microglia activation developed by Dr. Sieger. These computational and transgenic tools will be useful to the research community and will be available through us.

Communications and Engagement: We will ensure the optimal dissemination of our results to an expert audience by presentations at the Edinburgh Neuroscience Open Day (CB was a speaker in 2009, DS in 2013), meetings of the Scottish Neuroscience Group (CB spoke in 2008 and 2010), national (BNA - the Becker group's research will be featured in their Spring 2014 bulletin) and international neuroscience (e.g. former PhD students, Jolanda Münzel and Jochen Ohnmacht spoke at SFN, Tatyana Dias spoke at a Keystone Meeting) meetings, local, regional, national and international meetings of the zebrafish community (CB and TB were co-organizer of the 2011 international zebrafish meeting held in Edinburgh an of the "Zebrafish in Neuroscience" meeting held in Salamanca in 2012 at which DS also spoke). Furthermore, CB is the current secretary of EuFishBioMed, the The European Society for Fish Models in Biology and Medicine, and will use their meetings and newletters for dissemination of this project's findings.

We are currently in discussions with Matt Sharp, Director of Edinburgh's Transgenic Services to develop a regular contribution to workshops at the meetings of the International Society for Transgenic Technologies to disseminate our experience in zebrafish transgenesis to the ~350 delegates.

To ensure efficient communication of our findings to a wider audience, we collaborate with the University press office and Dr Jane Haley, the Edinburgh Neuroscience co-ordinator. Our work has been regularly covered in press releases, received media coverage (e.g. BBC radio, BBC website, The Herald) and has been covered by various charity newsletters and blogs (i.e. the Packard Center; MNDA). We will also update the "Research Briefings" held on the School's site with pertinent findings from the present grant. News pertaining to our findings will be added to the Centre's and School's web sites.

Developmental Biology is a particularly attractive topic to engage pupils in science. We will participate in Science and Career Fair activities at local primary and high schools and use work from the current grant to underscore the importance of developmental biology and the responsible use of animals in research.

Capability: If this project is funded we will train 1 Post-doc, 1 PhD student, 2 MSc students and 1 RA in regeneration research using widely applicable cutting edge anatomical, molecular and behavioural techniques, and we will present data at national and international meetings. We will engage the public in events organised by Edinburgh Neuroscience and strive to use our findings for Edinburgh Science Festival Activities and charity events suggested by Dr Breen.
 
Description We have found that upon application of a neurotoxin to the fish brain, we can selectively ablate neuronal populations that produce dopamine. This stimulates proliferation of progenitor (stem) cells in the brain, which leads to the replacement of some (but not all) neuronal populations. Stimulation of the immune system increases the progenitor proliferation, but not the number of fully mature neurons.
We find that the loss of these neurons leads to a specific movement deficit - fish swim further apart, which also reduces reproductive success. This loss is not repaired, since axons, the connections between the new neurons and the spinal cord, are not regrown.
After over a year of additional work, the manuscript was rejected from Nature Communications and is now under revision at The Journal of Neuroscience. We have deposited the manuscript in BioRxiv to make the data freely available.
Update: the paper has now been accepted and has appeared in the Journal of Neuroscience.
Exploitation Route We are exploring a collaboration with an EU colleague who has a different model of Parkinson's Disease in fishes to see whether there are common mechanisms of de- and regeneration.
Sectors Education,Pharmaceuticals and Medical Biotechnology,Other

 
Description Our group is part of the Edinburgh Parkinson's researchers, which has presented to stakeholders from Parkinson's UK. We are now (since 2017) also members of the Dundee-Edinburgh Parkinson's Research Initiative (inaugural meeting April 2018)
Sector Education
 
Description 1ST SYMPOSIUM OF THE DUNDEE-EDINBURGH PARKINSON'S RESEARCH INITIATIVE (April 2018) 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Other audiences
Results and Impact Launch of a new partnership between UoE and University of Dundee aimed at Parkinson's Research, attended by researchers, funders, donors, patients and families
Year(s) Of Engagement Activity 2018
 
Description VIP visit 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Policymakers/politicians
Results and Impact Ruth Davidson, leader of the Scottish Conservatives, visited the facility with a small delegation. They learned about the capacity of the facility and we also talked about our Dopamine-related work, since Ms Davidson was very interested in Parkinson's Disease.
Year(s) Of Engagement Activity 2018