Abstract
We report a water-based optical clearing agent, SeeDB, which clears fixed brain samples in a few days without quenching many types of fluorescent dyes, including fluorescent proteins and lipophilic neuronal tracers. Our method maintained a constant sample volume during the clearing procedure, an important factor for keeping cellular morphology intact, and facilitated the quantitative reconstruction of neuronal circuits. Combined with two-photon microscopy and an optimized objective lens, we were able to image the mouse brain from the dorsal to the ventral side. We used SeeDB to describe the near-complete wiring diagram of sister mitral cells associated with a common glomerulus in the mouse olfactory bulb. We found the diversity of dendrite wiring patterns among sister mitral cells, and our results provide an anatomical basis for non-redundant odor coding by these neurons. Our simple and efficient method is useful for imaging intact morphological architecture at large scales in both the adult and developing brains.
Publication types
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Comparative Study
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Evaluation Study
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Benzoates / pharmacology
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Benzyl Alcohol / pharmacology
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Brain / drug effects
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Brain / embryology
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Brain / ultrastructure*
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Dendrites / ultrastructure
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Electroporation
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Female
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Fixatives / pharmacology
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Fluorescent Dyes / analysis*
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Fluorescent Dyes / chemistry
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Formaldehyde / pharmacology
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Fructose / chemistry
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Fructose / pharmacology*
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Gene Knock-In Techniques
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Glycerol / analogs & derivatives*
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Glycerol / chemistry
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Glycerol / pharmacology
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Histocytological Preparation Techniques*
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Light
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Mice
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Mice, Inbred C57BL
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Mice, Inbred ICR
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Mice, Transgenic
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Microscopy / methods
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Nerve Net / ultrastructure*
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Neurons / ultrastructure
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Odorants
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Olfactory Bulb / cytology
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Olfactory Bulb / drug effects
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Olfactory Perception / physiology
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Phenyl Ethers / pharmacology
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Polymers / pharmacology
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Pregnancy
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Scattering, Radiation
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Solutions / chemistry
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Solutions / pharmacology*
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Sucrose / pharmacology
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Tissue Fixation / methods
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Urea / pharmacology
Substances
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Benzoates
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Fixatives
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Fluorescent Dyes
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Phenyl Ethers
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Polymers
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ScaleA2 solution
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SeeDB solution
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Solutions
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Formaldehyde
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dibenzyl ether
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Fructose
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Sucrose
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Urea
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Benzyl Alcohol
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benzyl benzoate
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Glycerol
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paraform