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Characterising the molecular heterogeneity within human inhibitory interneurons – at single-cell resolution

Keefe, Francesca 2019. Characterising the molecular heterogeneity within human inhibitory interneurons – at single-cell resolution. PhD Thesis, Cardiff University.
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Inhibitory interneurons are a heterogeneous class of GABAergic neurons, classified into discrete subtypes based on the expression of molecular markers, physiology, morphology and connectivity. However, the mechanism of interneuron diversification is poorly understood. hPSC-derived interneurons may help solve the mystery, by providing an unrestricted trajectory of interneuron differentiation, which is not confounded by species differences. Although, how well the model recapitulates native development remains under scrutiny. Moreover, the full extent of interneuron subtype diversity that can be achieved in vitro is unclear. Here an evaluation of hPSC model of interneuron differentiation has been presented. To tease apart the molecular heterogeneity, longitudinal single-cell RNA sequencing was conducted. The analysis revealed a heterogeneous MGE-like enriched progenitor population that differentiated into a diverse population of interneuron-like cells. On examining the authenticity of the model, using a novel cross-comparison to the native human MGE (10WPC-15WPC), encouraging similarities were found, especially between post-mitotic hPSC-derived interneurons and 15WPC MGE. However, several shortcomings of the model were also indicated. First, under-representation of specific genes encoding for transcription factors. Second, absent or low expression of late MGE markers and subtypes. Enforced expression of the under-expressed transcription factors may provide a means to enrich the interneuron-like population with these subtypes. Furthermore, the molecular resemblance to foetal-derived interneurons was reflected in the immature intrinsic and evoked neuronal responses of hPSC-derived interneurons, recorded using whole-cell patch clamp. Nevertheless, monosynaptic viral tracing demonstrated the capability of the interneuron-like cells to make synaptic connections, which (although not extensive) may be enhanced in a co-culture system. Overall, the interneuron differentiation of hPSC holds promise as a neurodevelopmental model. The novel characterisation of hPSC-derived interneurons and human MGE, at single-cell resolution, has simultaneously v revealed the limitations and refinements necessary to generate more authentic and defined interneuron subtypes in vitro. Thereby, widening the scope and validity of the model.

Item Type: Thesis (PhD)
Date Type: Completion
Status: Unpublished
Schools: Biosciences
Subjects: Q Science > Q Science (General)
Date of First Compliant Deposit: 26 February 2020
Last Modified: 30 Oct 2021 01:06

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