Quantitative Nanoscopy of Synaptic Sites
von Clara-Marie Gürth
Datum der mündl. Prüfung:2022-08-25
Erschienen:2023-08-23
Betreuer:Prof. Dr. Stefan Hell
Gutachter:Prof. Dr. Stefan Hell
Gutachter:Prof. Dr. Silvio O. Rizzoli
Zum Verlinken/Zitieren:
http://dx.doi.org/10.53846/goediss-10073
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Name:Dissertation_Clara-Marie_Guerth.pdf
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Zusammenfassung
Englisch
Synapses are the basic information processing unit of the brain. Neuronal activity and memory formation thereby require a vastly dynamic and rapidly changing synaptic environment and composition. In order to function, these changes need to be finely regulated and tailored to specific needs of individual synapses. How synaptic composition and function are connected and regulated is so far not fully understood. New optical imaging techniques such as STED and MINFLUX nanoscopy have enabled to investigate the synaptic environment with molecular specificity at so far unprecedented detail. While qualitative measures become increasingly available, quantitative analyses in nanoscopic imaging data are still challenging. Precise quantitative measures are essential to shed light into vastly heterogenous populations such as synapses as well as complex dynamic processes such as synaptic plasticity. To this aim different aspects of the synaptic composition and environment were quantitatively characterized within the context of synaptic activity combining different nanoscopy methods and implementing different labelling strategies. Firstly, the presence and distribution of secretory pathway elements in synaptic proximity was quantified within the context of markers of synaptic strength and activity. This analysis revealed the tools single synapses potentially utilize for local protein synthesis or delivery and how these depend on both increased strength and activity. Secondly, the content of cytoskeletal neurofilaments was quantified within postsynaptic spines with respect to the synaptic status. Here it was shown how different neurofilament isoforms depend differentially on both strength and activity of the individual synapse. Thirdly, the regulation of the postsynaptic scaffolding protein PSD-95 was characterised at the single synapse level. Here gene editing tools combined with pulse-chase labelling approaches added temporal information to imaging data, enabling to visualise protein turnover at the nanoscale, while imaging with three-dimensional molecular resolution, revealed the structural rearrangements during synaptic plasticity. Taken together this work combines different advanced imaging techniques, implements labelling tools and image analysis workflows to quantify how activity impacts the organization and composition of postsynaptic sites to better understand mechanisms of synaptic plasticity.
Keywords: Synapses; STED; MINFLUX; Nanoscopy; Super-Resolution; Spines; Plasticity
