Super-resolution STED microscopy for investigating the brain extracellular space

by Jan Tønnesen, Achucarro Basque Center for Neuroscience, Leioa, Spain

will take place on Tuesday, April 25th, 2023 from 16:00 to 17:00 hours in the CBBM Building, Ground Floor, Seminar Room Levi-Montalcini.

Host: Prof. Dr. Markus Schwaninger
Institute for Experimental and Clinical Pharmacology and Toxicology, University of Lübeck

The brain extracellular space (ECS) is a reticular continuum of interconnected channels and reservoirs. The ranges of ECS channel widths, lengths, and shapes easily exceed the morphological complexity of individual neural cells, so that on one hand the ECS is comparatively vast in extent, while at the same time its even more geometrically complex. The combination of spatial continuity and a fine mesh-like structure makes the ECS geometry extremely difficult to visualize in live tissue, because conventional light-microscopy simply blurs out structural details beyond cell somata and large dendrites. Current data on ECS structure are therefore based largely on volume-averaging imaging techniques or on electron microscopy in fixed tissue. Accordingly, our knowledge about ECS structure and putative dynamics, not to mention functional roles, is rudimentary.

In a step forward, we recently introduced super-resolution shadow imaging (SUSHI) as a new way to image the extracellular space in live brain slices by combining fluorescent perfusion-labeling of the interstitial fluid with STED microscopy. This approach reveals the complex geometry and dynamics of the ECS, while simultaneously outlining all cellular structures in the field of view as shadows at around 50 nm resolution.

In this talk I will introduce STED microscopy for imaging in live organotypic mouse brain slices, and how we went from imaging individual neuronal synapses to also imaging the extracellular space. I will show preliminary results from a computational diffusion model we are establishing based on SUSHI images of the ECS, which may help us understand functional aspects of extrasynaptic signaling and metabolite clearance from dense parenchyma.

Biosketch: Dr. Jan Tønnesen (Danish) is a group leader at the University of the Basque country and the Achucarro Basque Center for Neuroscience in Bilbao (Spain). Tønnesen obtained his BSc and MSc in Biology from the university of Copenhagen (Denmark). In 2010 he earned his PhD from Lund University (Sweden), working with then new optogenetic technologies in settings of epilepsy and Parkinson's disease under the supervision of Prof. M Kokaia. Hereafter, starting as a postdoc and later working as a staff researcher in the group of Prof. Nagerl at the University of Bordeaux, Tønnesen developed and applied STED microscopy for imaging dendritic spines and the brain extracellular space in live brain slices. Since 2016 he is a group leader in Bilbao, where he continues to combine his expertise with disease models and STED microscopy, with a main focus on revealing new structure-function relationships of the extracellular space. Lab website: