Descrizione del progetto
Far progredire l’imaging delle sinapsi neuronali
I neuroni comunicano tra loro, trasmettendo segnali elettrici e chimici attraverso giunzioni specializzate note come sinapsi. Lo studio degli organelli e degli ammassi di proteine nelle sinapsi si è rivelato impegnativo a causa delle loro piccole dimensioni e della breve distanza che li separa. Il progetto MoNaLISA, finanziato dal Consiglio europeo della ricerca, intende esplorare la microscopia a fluorescenza a super-risoluzione che offre un’analisi ad alta risoluzione e basata su singole molecole. I ricercatori supereranno i problemi legati al ritmo di registrazione grazie a un microscopio innovativo che consente di seguire gli organelli e le proteine neuronali. Questo progetto introduce un nuovo paradigma per il nano-imaging veloce e quantitativo nelle scienze della vita.
Obiettivo
Synaptic function is difficult to analyze in living neurons using conventional optics, since the synaptic organelles and protein clusters are small and tightly spaced. The solution to this problem can come from the field of super-resolution fluorescence microscopy, or nanoscopy. However, the current approaches to nanoscopy are still far from reaching this goal. Single molecule-based approaches (including STORM and PALM) provide high spatial resolution, but slow recording, insufficient for live imaging. Ensemble approaches (including SSIM and STED) are able to record faster, but with poorer resolution or with high, potentially toxic, laser powers. It is currently impossible to image the same neuron for hours and days, with both high spatial (~30 nm) and temporal (10-1000 Hz) resolution, and with minimal photodamage. My aim is to fill this gap, by developing, for the first time, a microscope that combines the advantages of both single molecule-based and ensemble approaches. I will base the microscope on RESOLFT, a low-photodamage ensemble approach that I have pioneered recently. I will use line patterns to speed up the recording and 2photon-switching for 3D ability. I will combine this with sensitive detection schemes that allow single-molecule detection and counting, relying on my previous expertise with PALM and GSDIM. The new set-up, termed molecular nanoscale long-term imaging with sequential acquisition (MoNaLISA), will track neuronal organelles and proteins on different time scales, spanning from milliseconds to days, with a resolution close to the molecular scale. To obtain the first proof-of-principle results, I will address several issues still open in the synaptic transmission field, relating to synaptic vesicle recycling, biogenesis and degradation. Overall, my project will introduce a novel paradigm to imaging in the life sciences, which will enable fast and quantitative nano-imaging of cells and tissues.
Campo scientifico
- natural sciencescomputer and information sciencessoftware
- natural sciencesphysical sciencesopticsmicroscopysuper resolution microscopy
- natural sciencesbiological sciencesbiochemistrybiomoleculesproteins
- humanitiesartsmodern and contemporary artcinematography
- natural sciencesphysical sciencesopticslaser physics
Programma(i)
Argomento(i)
Meccanismo di finanziamento
ERC-STG - Starting GrantIstituzione ospitante
100 44 Stockholm
Svezia