Expansion microscopy: A powerful nanoscale imaging tool for neuroscientists

One of the biggest unsolved questions in neuroscience is how molecules and neuronal circuitry create behaviors, and how their misregulation or dysfunction results in neurological disease. Light microscopy is a vital tool for the study of neural molecules and circuits. However, the fundamental optical diffraction limit precludes the use of conventional light microscopy for sufficient characterization of critical signaling compartments and nanoscopic organizations of synapse-associated molecules. We have witnessed rapid development of super-resolution microscopy methods that circumvent the resolution limit by controlling the number of emitting molecules in specific imaging volumes and allow highly resolved imaging in the 10?100 nm range. Most recently, Expansion Microscopy (ExM) emerged as an alternative solution to overcome the diffraction limit by physically magnifying biological specimens, including nervous systems. Here, we discuss how ExM works in general and currently available ExM methods. We then review ExM imaging in a wide range of nervous systems, including Caenorhabditis elegans, Drosophila, zebrafish, mouse, and human, and their applications to synaptic imaging, neuronal tracing, and the study of neurological disease. Finally, we provide our prospects for expansion microscopy as a powerful nanoscale imaging tool in the neurosciences.

Automated summary

One of the biggest challenges in neuroscience is understanding how molecules and neuronal circuitry give rise to behaviors, and how their dysfunction leads to neurological diseases. While light microscopy is a crucial tool for studying neural molecules and circuits, the diffraction limit hinders detailed characterization of subcellular structures and nanoscopic organizations. Super-resolution microscopy methods have been developed to bypass this limitation, allowing highly resolved imaging in the nanometer range. Expansion Microscopy (ExM) has emerged as a promising solution to surpass the diffraction limit by physically enlarging biological specimens, providing a powerful tool for studying neural systems at the nanoscale.