TY - JOUR
T1 - Reconstructing neural circuits using multiresolution correlated light and electron microscopy
AU - Friedrichsen, Karl
AU - Ramakrishna, Pratyush
AU - Hsiang, Jen Chun
AU - Valkova, Katia
AU - Kerschensteiner, Daniel
AU - Morgan, Josh L.
N1 - Funding Information:
This work was supported by an unrestricted grant to the Department of Ophthalmology and Visual Sciences from Research to Prevent Blindness, by a Research to Prevent Blindness Career Development Award (JM), and by the NIH (EYE030623 to DK and JM, EY029313 to JM, EY026978, EY023341, and EY027411 to DK).
Publisher Copyright:
Copyright © 2022 Friedrichsen, Ramakrishna, Hsiang, Valkova, Kerschensteiner and Morgan.
PY - 2022/10/21
Y1 - 2022/10/21
N2 - Correlated light and electron microscopy (CLEM) can be used to combine functional and molecular characterizations of neurons with detailed anatomical maps of their synaptic organization. Here we describe a multiresolution approach to CLEM (mrCLEM) that efficiently targets electron microscopy (EM) imaging to optically characterized cells while maintaining optimal tissue preparation for high-throughput EM reconstruction. This approach hinges on the ease with which arrays of sections collected on a solid substrate can be repeatedly imaged at different scales using scanning electron microscopy. We match this multiresolution EM imaging with multiresolution confocal mapping of the aldehyde-fixed tissue. Features visible in lower resolution EM correspond well to features visible in densely labeled optical maps of fixed tissue. Iterative feature matching, starting with gross anatomical correspondences and ending with subcellular structure, can then be used to target high-resolution EM image acquisition and annotation to cells of interest. To demonstrate this technique and range of images used to link live optical imaging to EM reconstructions, we provide a walkthrough of a mouse retinal light to EM experiment as well as some examples from mouse brain slices.
AB - Correlated light and electron microscopy (CLEM) can be used to combine functional and molecular characterizations of neurons with detailed anatomical maps of their synaptic organization. Here we describe a multiresolution approach to CLEM (mrCLEM) that efficiently targets electron microscopy (EM) imaging to optically characterized cells while maintaining optimal tissue preparation for high-throughput EM reconstruction. This approach hinges on the ease with which arrays of sections collected on a solid substrate can be repeatedly imaged at different scales using scanning electron microscopy. We match this multiresolution EM imaging with multiresolution confocal mapping of the aldehyde-fixed tissue. Features visible in lower resolution EM correspond well to features visible in densely labeled optical maps of fixed tissue. Iterative feature matching, starting with gross anatomical correspondences and ending with subcellular structure, can then be used to target high-resolution EM image acquisition and annotation to cells of interest. To demonstrate this technique and range of images used to link live optical imaging to EM reconstructions, we provide a walkthrough of a mouse retinal light to EM experiment as well as some examples from mouse brain slices.
KW - confocal 3D microscopy
KW - connectomics
KW - correlated light and electron microscopy (CLEM)
KW - electron microscopy
KW - neural circuit
KW - synapse
KW - tissue mapping
UR - http://www.scopus.com/inward/record.url?scp=85141408948&partnerID=8YFLogxK
U2 - 10.3389/fncir.2022.753496
DO - 10.3389/fncir.2022.753496
M3 - Article
C2 - 36338333
AN - SCOPUS:85141408948
VL - 16
JO - Frontiers in Neural Circuits
JF - Frontiers in Neural Circuits
SN - 1662-5110
M1 - 753496
ER -