During the development of the nervous system, neurons differentiate from progenitors into the vast array of neuronal cell types found in the mature CNS and PNS. Studies in this field have discovered an array of intrinsic and extrinsic factors that modulate this process, but there remains more to be uncovered. Independent research avenues in the two laboratories supporting this project have uncovered novel extrinsic (hormonal) and intrinsic (the signalling protein willin) regulators of neuronal differentiation. The roles played by these proteins, and the interplay between them in neurodevelopment remain to be fully elucidated.
Specific aims of the developmental aspect of this project will be:
- To determine the degree of interplay between external factors that mediate differentiation, and signalling via the willin/Hippo pathway
- To establish stable neuronal cell lines expressing fluorescently tagged signalling molecules of interest to permit image analysis of sub-cellular localisations and interactions. This can be coupled to cell painter techniques to facilitate organelle identification.
- To use high sensitivity imaging techniques to establish the nature of the neuronal networks established and to look at the effects of hormones and growth factors on the electrical activity of these networks post-differentiation.
Together these data will establish a robust evaluation of the function of newly-discovered modulators of neuronal differentiation on the establishment of a stable neuronal network. It is well known that cells in culture age more rapidly than those in the body. In a further aim, this project will capitalise upon this to use the resources from the developmental aspect to determine how differentiation-linked signalling pathways and firing properties of these cells change with age. This will be coupled to the determination of the accumulation of age-linked biomarkers (e.g. epigenetic drift, accumulation of oxidative end-products) to establish a well-characterised model of neuronal ageing in vitro. This model can be used to inform of the effects of extrinsic signals on these pathways in aged cells.
This project will use a combination of cell biological, biochemical and imaging approaches to uncover the links between extrinsic signalling during neurodevelopment and the activation of key intracellular pathways. Techniques that will be developed during this project include neuronal cell culture and transfection; molecular analysis of protein expression levels using ELISA, immunocytochemistry and western blot; the use of cell painter dyes for ultrastructural and image analysis; and the use of novel unpublished optical technologies to rapidly determine the firing properties of neurons and assessment of the development of functional neuronal networks.
The student undertaking this study will be fully trained in a number of complimentary research techniques to develop a strong technical background for their future career. Training will also be provided in the analysis and interpretation of research data. They will be expected to play a full role in preparing data for presentation both at scientific conferences and through publication in peer-reviewed journals if appropriate. Both laboratories have a strong track record in public engagement in science and training towards participation in, and development of, outreach activities will be given.