Dopamine : a versatile player in development, regeneration and disease

The dopamine neurotransmitter is present in all multicellular organisms. In the brain, the dopaminergic system orchestrates reward-motivation pathways and is involved in the control of voluntary movements and endocrine hormone secretion. Dysfunction of dopamine signalling may lead to pathological conditions such as Parkinson’s disease, where dopaminergic neurons of the midbrain degenerate. Moreover, modulation of dopamine receptor signalling influences tumour growth. The aim of this work was to explore the regeneration capacity of the dopaminergic system in the vertebrate brain and to test whether dopamine may control the growth of brain tumours. To this end we performed two sets of studies. Initially, we investigated the development and regeneration of the dopaminergic system in newts, which are aquatic salamanders capable of complete regeneration of the dopaminergic system in the brain. Thereafter, we investigated how dopaminergic ligands impinge specifically on brain tumour cells.

In paper I, we screened a library of dopaminergic ligands for their ability to stimulate or to inhibit glioblastoma cell growth and survival. We identified the dopamine receptor 2 antagonist, trifluoperazine, as an inhibitor of glioblastoma growth. We also showed that susceptibility to trifluoperazine correlates with the dopamine receptor expression profile of the investigated glioblastoma cell lines. We concluded that dopamine receptor signalling pathways are promising targets for pharmacological interventions to inhibit glioblastoma growth.

In paper II, we characterized the cellular basis of brain development and stereotyped behaviour in two regeneration model salamander species. These data provide insight into the maturation of neural stem cells that are found in the adult salamander brain. Furthermore, we showed how lesioning of the dopaminergic innervation affects neurogenesis in the forebrain and behavioural performance. This study provides a new evolutionary perspective on the genesis and dynamics of brain cells in the salamander brain, including dopaminergic cells.

In paper III, we developed a tissue clearing method, CUBICe, to extend our study of dopaminergic neurite outgrowth during development as well as regeneration. We demonstrated that CUBICe is compatible for high resolution imaging of whole salamander brains. It is also a faster and more robust method, which allows to maintain a better sample integrity of embryonic brains in general, compared to Advanced CUBIC and Advanced CLARITY. In addition, using CUBICe we achieved tracing of genetically marked cells with neurite outgrowth of over 3600 µm. Ultimately, we showed that our method is ideal for tracing genetically marked dopaminergic cells in the salamander brain and for quantifying dopaminergic neurite density and regeneration in whole brain regions.

In summary, this thesis provides insight into the versatile role of dopamine in both normal and pathological conditions of the vertebrate brain, as well as offers innovative tools for studying the regeneration of the dopaminergic system.

List of scientific papers

I. A chemical screen identifies trifluoperazine as an inhibitor of glioblastoma growth. Tiago Pinheiro, Magdalena Otrocka, Brinton Seashore-Ludlow, Vilma Rraklli, Johan Holmberg, Karin Forsberg-Nilsson, András Simon and Matthew Kirkham. Biochemical and Biophysical Research Communications. 2017;494(3-4):477-483.
https://doi.org/10.1016/j.bbrc.2017.10.106

II. Cellular basis of brain maturation and acquisition of complex behaviors in salamanders. Alberto Joven, Heng Wang, Tiago Pinheiro, L. Shahul Hameed, Laure Belnoue and András Simon. Development. 2018;145:1.
https://doi.org/10.1242/dev.160051

III. CUBICe: an optimized clearing method for cell tracing and evaluation of neurite density in the salamander brain. Tiago Pinheiro, Steven Edwards, Ivanna Mayorenko, Alberto Joven, Christina Kantzer, András Simon and Matthew Kirkham. [Manuscript]