Combining array tomography with electron tomography provides insights into leakiness of the blood-brain barrier in mouse cortex

Research output: Contribution to journalResearch articleContributedpeer-review

Contributors

  • Georg Kislinger - , Technical University of Munich, German Center for Neurodegenerative Diseases (DZNE) (Author)
  • Gunar Fabig - , Core Facility Cellular Imaging, Experimental Center of the Faculty of Medicine (Author)
  • Antonia Wehn - , Hospital of the Ludwig-Maximilians-University (LMU) Munich (Author)
  • Lucia Rodriguez - , German Center for Neurodegenerative Diseases (DZNE) (Author)
  • Hanyi Jiang - , Greifswald University Hospital (Author)
  • Cornelia Niemann - , German Center for Neurodegenerative Diseases (DZNE) (Author)
  • Andrey S Klymchenko - , University of Strasbourg (Author)
  • Nikolaus Plesnila - , Ludwig Maximilian University of Munich (Author)
  • Thomas Misgeld - , Ludwig Maximilian University of Munich (Author)
  • Thomas Müller-Reichert - , Core Facility Cellular Imaging, Experimental Center of the Faculty of Medicine (Author)
  • Igor Khalin - , Université de Caen, Normandie Université (Author)
  • Martina Schifferer - , Ludwig Maximilian University of Munich (Author)

Abstract

Like other volume electron microscopy approaches, automated tape-collecting ultramicrotomy (ATUM) enables imaging of serial sections deposited on thick plastic tapes by scanning electron microscopy (SEM). ATUM is unique in enabling hierarchical imaging and thus efficient screening for target structures, as needed for correlative light and electron microscopy. However, SEM of sections on tape can only access the section surface, thereby limiting the axial resolution to the typical size of cellular vesicles with an order of magnitude lower than the acquired xy resolution. In contrast, serial-section electron tomography (ET), a transmission electron microscopy-based approach, yields isotropic voxels at full EM resolution, but requires deposition of sections on electron-stable thin and fragile films, thus making screening of large section libraries difficult and prone to section loss. To combine the strength of both approaches, we developed 'ATUM-Tomo, a hybrid method, where sections are first reversibly attached to plastic tape via a dissolvable coating, and after screening detached and transferred to the ET-compatible thin films. As a proof-of-principle, we applied correlative ATUM-Tomo to study ultrastructural features of blood-brain barrier (BBB) leakiness around microthrombi in a mouse model of traumatic brain injury. Microthrombi and associated sites of BBB leakiness were identified by confocal imaging of injected fluorescent and electron-dense nanoparticles, then relocalized by ATUM-SEM, and finally interrogated by correlative ATUM-Tomo. Overall, our new ATUM-Tomo approach will substantially advance ultrastructural analysis of biological phenomena that require cell- and tissue-level contextualization of the finest subcellular textures.

Details

Original languageEnglish
Article numberRP90565
Number of pages21
JournaleLife
Volume12 (2024)
Publication statusPublished - 5 Aug 2024
Peer-reviewedYes

External IDs

PubMedCentral PMC11299977
Scopus 85200530784
ORCID /0000-0003-3017-0978/work/166325970

Keywords

Keywords

  • Animals, Blood-Brain Barrier/ultrastructure, Cerebral Cortex/diagnostic imaging, Electron Microscope Tomography/methods, Male, Mice, Mice, Inbred C57BL, Microscopy, Electron, Scanning/methods, Microtomy