Functional nanoscopy of the synaptic active zone

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Fundamental insight into biological processes demands information on how structure encodes function. Despite a gradually emerging comprehensive protein catalogue, we still lack basic information describing how the nanoscopic organisation of proteins at synapses gives rise to neurotransmission. In essence, this is due to the diffraction-limited resolution of conventional light microscopy, which has hindered access to the spatial nanodomain in a physiologically relevant context.The present research proposal focuses on the synaptic active zone (AZ), the highly specialised presynaptic site of neurotransmitter release. The precise molecular architecture of AZs gives rise to different structural and functional AZ states, which shape chemical neurotransmission and fundamentally influence brain function. By engaging recent technological innovations, this project will test the hypothesis that the functional status of the AZ can be predicted by the number and the precise spatial organisation of specific molecules.To gain access to the molecular domain, the genetically accessible organism Drosophila melanogaster will be employed. In the fist project section, synaptic protein expression will be manipulated to alter the molecular organisation of AZs. Functional nanoscopy, the correlative application of electrophysiology and super-resolution fluorescence microscopy, will then focus on key AZ proteins to establish quantitative, causal relationships between the molecular ultrastructure and the function of AZs, i.e. molecular blueprints of AZ states.In the second project section, novel optogenetic tools will be utilised to trigger activity-dependent plasticity of the AZ in vivo. The molecular mechanisms underlying these physiological changes in AZ function will then be interpreted using the established blueprints.This research proposal will test how function is linked to the ultrastructure of AZs and examine how nanoscopic reorganisations of key molecules mediate distinct mechanisms of plasticity over different time scales in vivo. To this end, the synergistic combination of electrophysiology, super-resolution microscopy and optogenetics aims to identify fundamental principles of organisation underlying brain function.