Format

Send to

Choose Destination
See comment in PubMed Commons below
Cell Rep. 2016 Mar 22;14(11):2718-32. doi: 10.1016/j.celrep.2016.02.057. Epub 2016 Mar 10.

Super-Resolution Mapping of Neuronal Circuitry With an Index-Optimized Clearing Agent.

Author information

1
Laboratory for Sensory Circuit Formation, RIKEN Center for Developmental Biology, Kobe 650-0047, Japan.
2
Laboratory of Developmental Neurobiology, Brain/Liver Interface Medicine Research Center, Graduate School of Medical Sciences, Kanazawa University, Kanazawa 920-8640, Japan; CREST, Japan Science and Technology Agency (JST), Saitama 332-0012, Japan.
3
Laboratory for Chromosome Segregation, RIKEN Center for Developmental Biology, Kobe 650-0047, Japan.
4
Laboratory for Chromosome Segregation, RIKEN Center for Developmental Biology, Kobe 650-0047, Japan; Graduate School of Biostudies, Kyoto University, Kyoto 606-8501, Japan.
5
Laboratory of Developmental Neurobiology, Brain/Liver Interface Medicine Research Center, Graduate School of Medical Sciences, Kanazawa University, Kanazawa 920-8640, Japan; Mathematical Neuroscience Unit, Institute for Frontier Science Initiative, Kanazawa University, Kanazawa 920-8640, Japan; CREST, Japan Science and Technology Agency (JST), Saitama 332-0012, Japan.
6
Laboratory for Sensory Circuit Formation, RIKEN Center for Developmental Biology, Kobe 650-0047, Japan; PRESTO, Japan Science and Technology Agency (JST), Saitama 332-0012, Japan; Graduate School of Biostudies, Kyoto University, Kyoto 606-8501, Japan. Electronic address: imai@cdb.riken.jp.

Abstract

Super-resolution imaging deep inside tissues has been challenging, as it is extremely sensitive to light scattering and spherical aberrations. Here, we report an optimized optical clearing agent for high-resolution fluorescence imaging (SeeDB2). SeeDB2 matches the refractive indices of fixed tissues to that of immersion oil (1.518), thus minimizing both light scattering and spherical aberrations. During the clearing process, fine morphology and fluorescent proteins were highly preserved. SeeDB2 enabled super-resolution microscopy of various tissue samples up to a depth of >100 μm, an order of magnitude deeper than previously possible under standard mounting conditions. Using this approach, we demonstrate accumulation of inhibitory synapses on spine heads in NMDA-receptor-deficient neurons. In the fly medulla, we found unexpected heterogeneity in axon bouton orientations among Mi1 neurons, a part of the motion detection circuitry. Thus, volumetric super-resolution microscopy of cleared tissues is a powerful strategy in connectomic studies at synaptic levels.

PMID:
26972009
DOI:
10.1016/j.celrep.2016.02.057
[Indexed for MEDLINE]
Free full text
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for Elsevier Science
    Loading ...
    Support Center