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Revealing ultrastructural changes that determine the development of a CNS synapse

dc.contributor.advisorWichmann, Carolin Prof. Dr.
dc.contributor.authorHintze, Anika
dc.format.extentXXX Seitende
dc.titleRevealing ultrastructural changes that determine the development of a CNS synapsede
dc.contributor.refereeWichmann, Carolin Prof. Dr.
dc.description.abstractengEndbulbs of Held are synaptic contacts between auditory nerve fibers and bushy cells in the cochlear nucleus (CN) and represent the first central synapses of the auditory pathway. Endbulbs undergo functional and structural development that allows fast and reliable neurotransmission with high temporal precision. Its large terminal size and number of individual active zones (AZs) as well as its good accessibility make the endbulb of Held an ideal synapse to study structure-function relationships. While morphogenesis from small to large, highly branched terminals needs several weeks, functional maturation is completed shortly after hearing onset that occurs on postnatal day (P) 12 in mice. The developmental process is modulated by auditory nerve activity. Loss of this activity by depletion of otoferlin, an indispensable protein for synaptic transmission at inner hair cell ribbon synapses, results in smaller endbulbs. Synaptic transmission at endbulbs is further mediated by a plethora of proteins. Development from pre-hearing to adult stages in wild type (wt) mice, the lack of auditory nerve activity and the lack of presynaptic proteins, such as Rab-interacting molecule (RIM)-binding protein 2 (BP2) and Mover are presumably reflected in structural changes at endbulb AZs. In my thesis, I addressed the ultrastructural consequences of these functional changes by using high-pressure freezing and freeze substitution prior to electron tomography of freshly sliced CNs. With this methodology, the morphological details of endbulb terminals like synaptic vesicle (SV) distribution and the structure of mitochondria can be examined with highest z resolution. I found that SV pools were unchanged upon maturation from pre-hearing (P10) to hearing (P21) in wt but increased mitochondrial volume and crista membrane density suggest higher mitochondrial performance after the hearing onset. Endbulb AZs of six-months-old (6M) mice exhibited larger SV pools and a clustering of SVs within 5 15 nm from the AZ membrane compared to P10. SV densities at endbulb AZs were comparable between mice lacking otoferlin (Otof-/-) and wt at P10 and P21/22. However, AZs of 6M Otof-/- mice contained a considerably reduced number of SV compared to wt. This decrease also affected morphologically docked (0-2 nm) and membrane-proximal SVs (2-40 nm away from the AZ membrane) that were analyzed separately. While SVs of Otof-/- mice were larger at P10, the docked SVs at 6M were significantly smaller than at wt. Upon RIM-BP2 disruption, docked SVs and SVs in membrane proximity (2 20 nm) were decreased at endbulb AZs compared to wt. The lack of Mover resulted in reduced membrane-proximal SVs at endbulb AZs of adult mice and elevated numbers of docked SVs at inhibitory synapses. These results suggest that the maturation from pre-hearing to hearing is accompanied by a higher energy demand in wt. Increased SV pool sizes upon further development towards adulthood might be required to support the temporally precise neurotransmission at endbulbs. The elevated SVs within 5-15 nm from the AZ membrane might thereby contribute to an efficient SV recruitment. At 6M Otof-/- endbulb AZs, reliability of synaptic transmission might be reduced due to reduced numbers of docked and membrane-proximal SVs. Finally, Mover and RIM-BP2 seem to be involved in recruiting SVs to membrane proximity at endbulb of Held AZs. RIM-BP2 is also important for SV docking at endbulbs, while Mover seems to prevent SV docking at inhibitory
dc.contributor.coRefereeDresbach, Thomas Prof. Dr.
dc.subject.engEndbulb of Heldde
dc.subject.engPresynaptic Mitochondriade
dc.subject.engElectron Tomographyde
dc.subject.engSynaptic Vesicle Poolsde
dc.affiliation.instituteBiologische Fakultät für Biologie und Psychologiede
dc.subject.gokfullBiologie (PPN619462639)de
dc.notes.confirmationsentConfirmation sent 2023-04-17T06:15:01de

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