Year: 2019
Type: dissertation
Abstract: Neuroscientific research frequently utilises loss of function experiments to attribute function to a brain region. Gain of function experiments via cortical re-wiring would aid in validation of these studies and could allow the repair of damaged neural circuitry or the creation of novel neural structures. Research into neuronal re-wiring within the central nervous system demonstrates insufficient neurite extension upon implantation and highlights the need for similarity between exogenous and endogenous neurons. An aligned poly-L-lactic acid (PLLA) nanofibre scaffold was developed that induced the three dimensional aggregation of primary cortical neurons into a physiological structure of clustered soma and aligned, fasciculated neurites in a controlled way. Due to the self-assembling and physiological architecture of these 3D structures and subsequent detection of electrophysiological activities, these 3D structures were classified as “organoids”. Cerebral cortical organoids generated using the nanofibre based methodology were demonstrated to be more developmentally advanced than their two-dimensional counterparts and mechanisms were elucidated for the aggregation of the neurons and the development that occurred post-aggregation. Analysis of gene expression suggested that the organoids were also undergoing advanced developmental processes and proposed a mechanism by which this occurs. Optogenetic depolarisation of the implanted neurons and detection of downstream electrophysiological activity was selected as the means of confirming integration of exogenous neurons into endogenous circuitry post-implantation. Methods were optimised to facilitate efficient viral transfection of the optogenetic protein (Channelrhodopsin-2). Aligned PLLA nanofibres were found to significantly enhance transfection rates relative to the 2D control and the process by which this occurs was investigated. The work presented within suggests that aligned PLLA nanofibres may be used to generate a cerebral cortical organoid that is suited to implantation and cortical re-wiring and may have additional in vitro applications within diverse fields such as high throughput pharmacology, computational neuroscience and bioengineering.
Author: Nick Weir
Language: English
Subfield: Cellular and Molecular Neuroscience
Field: Neuroscience
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W3021002853