Expansion-assisted selective plane illumination microscopy for nanoscale imaging of centimeter-scale tissues.

Recent advances in tissue processing, labeling, and fluorescence microscopy are providing unprecedented views of the structure of cells and tissues at sub-diffraction resolutions and near single molecule sensitivity, driving discoveries in diverse fields of biology, including neuroscience. Biological tissue is organized over scales of nanometers to centimeters. Harnessing molecular imaging across three-dimensional samples on this scale requires new types of microscopes with larger fields of view and working distance, as well as higher imaging throughput. We present a new expansion-assisted selective plane illumination microscope (ExA-SPIM) with diffraction-limited and aberration-free performance over a large field of view (85 mm 2 ) and working distance (35 mm). Combined with new tissue clearing and expansion methods, the microscope allows nanoscale imaging of centimeter-scale samples, including entire mouse brains, with diffraction-limited resolutions and high contrast without sectioning. We illustrate ExA-SPIM by reconstructing individual neurons across the mouse brain, imaging cortico-spinal neurons in the macaque motor cortex, and tracing axons in human white matter.

SNT: a unifying toolbox for quantification of neuronal anatomy.

SNT is an end-to-end framework for neuronal morphometry and whole-brain connectomics that supports tracing, proof-editing, visualization, quantification and modeling of neuroanatomy. With an open architecture, a large user base, community-based documentation, support for complex imagery and several model organisms, SNT is a flexible resource for the broad neuroscience community. SNT is both a desktop application and multi-language scripting library, and it is available through the Fiji distribution of ImageJ.

Reconstruction of 1,000 Projection Neurons Reveals New Cell Types and Organization of Long-Range Connectivity in the Mouse Brain.

Neuronal cell types are the nodes of neural circuits that determine the flow of information within the brain. Neuronal morphology, especially the shape of the axonal arbor, provides an essential descriptor of cell type and reveals how individual neurons route their output across the brain. Despite the importance of morphology, few projection neurons in the mouse brain have been reconstructed in their entirety. Here we present a robust and efficient platform for imaging and reconstructing complete neuronal morphologies, including axonal arbors that span substantial portions of the brain. We used this platform to reconstruct more than 1,000 projection neurons in the motor cortex, thalamus, subiculum, and hypothalamus. Together, the reconstructed neurons constitute more than 85 meters of axonal length and are available in a searchable online database. Axonal shapes revealed previously unknown subtypes of projection neurons and suggest organizational principles of long-range connectivity.