Generating GABAergic interneurons through reprogramming and differentiation strategies: English

In recent years reprogramming techniques, differentiation protocols, and sequencing analysis have opened a field for generating and studying human-derived neurons with greater potential than ever before. This has generated a large number of studies with the goal of deriving bona fide neurons. Both direct reprogramming and differentiation have benefits and drawbacks, while direct reprogramming can be used to study patient-specific late-onset diseases, differentiation provides developmental information and mechanics.

Collectively this thesis provides a variety of ways to produce neurons with a special focus on GABAergic interneurons. The first part of the thesis (Paper I and II) explores reprogramming of adult human fibroblasts and human glial progenitor cells into GABAergic interneurons in 2D cultures. We provide evidence that the two cell sources can successfully convert into subtype-specific GABAergic interneurons. Following the establishment of successful conversion protocols, we then introduce a 3D model for direct conversion that replicates the cell-cell interactions pivotal for physiological relevance, leading to an accelerated induction of functional neurons (Paper III). Moreover, single-nucleus transcriptomics uncovered the need for simultaneous expression of specific transcription factors able to convert various glial subtypes into neurons, without relying on a specific subtype preference. The final part of this thesis (Paper IV) focuses on differentiation of hESCs into subtype- and subclass-specific interneurons. We further explore the contribution of various glial cells in a 3D co-culture setting with interneurons employing both functional and molecular analyses. This is aimed at developing a robust model that more accurately replicates the interactions within the human brain and holds potential for the in-depth study of interneuron development and pathology.

Together these papers encompass a collection of ways to derive human subtype- specific neurons in vitro and serve as platforms for both reprogramming and differentiation studies.