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CLARITY for mapping the nervous system

Nature Methods volume 10, pages 508513 (2013) | Download Citation

  • A Corrigendum to this article was published on 27 September 2013

This article has been updated

With potential relevance for brain-mapping work, hydrogel-based structures can now be built from within biological tissue to allow subsequent removal of lipids without mechanical disassembly of the tissue. This process creates a tissue-hydrogel hybrid that is physically stable, that preserves fine structure, proteins and nucleic acids, and that is permeable to both visible-spectrum photons and exogenous macromolecules. Here we highlight relevant challenges and opportunities of this approach, especially with regard to integration with complementary methodologies for brain-mapping studies.

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  • 20 June 2013

    In the version of this article initially published, several reference callouts in the text were wrong. The errors have been corrected in the HTML and PDF versions of the article.

References

  1. 1.

    et al. Structural and molecular interrogation of intact biological systems. Nature advance online publication, doi:10.1038/nature12107 (10 April 2013).

  2. 2.

    The functional organization of the barrel cortex. Neuron 56, 339–355 (2007).

  3. 3.

    et al. Visualizing an olfactory sensory map. Cell 87, 675–686 (1996).

  4. 4.

    Optogenetics and psychiatry: applications, challenges, and opportunities. Biol. Psychiatry 71, 1030–1032 (2012).

  5. 5.

    & The rise of the 'projectome'. Nat. Methods 4, 307–308 (2007).

  6. 6.

    Diffusion in brain extracellular space. Brain 6, 1277–1340 (2008).

  7. 7.

    , & A review of the optical properties of biological tissues. IEEE J. Quantum Electronics 26, 2166–2185 (1990).

  8. 8.

    & Deep tissue two-photon microscopy. Nat. Methods 2, 932–940 (2005).

  9. 9.

    Surface cryoplanning: a technique for clinical anatomical correlations. Uppsala J. Med. Sci. 91, 251–255 (1986).

  10. 10.

    , , , & Postmortem anatomy from cryosectioned whole human brain. J. Neurosci. Methods 54, 239–252 (1994).

  11. 11.

    , , , & Surface imaging microscopy, an automated method for visualizing whole embryo samples in three dimensions at high resolution. Dev. Dyn. 225, 369–375 (2002).

  12. 12.

    et al. Construction of anatomically correct models of mouse brain networks. Neurocomputing 58–60, 379–386 (2004).

  13. 13.

    et al. Micro-optical sectioning tomography to obtain a high-resolution atlas of the mouse brain. Science 330, 1404–1408 (2010).

  14. 14.

    et al. All-optical histology using ultrashort laser pulses. Neuron 39, 27–41 (2003).

  15. 15.

    et al. Serial two-photon tomography for automated ex vivo mouse brain imaging. Nat. Methods 9, 255–258 (2012).

  16. 16.

    & Serial block-face scanning electron microscopy to reconstruct three-dimensional tissue nanostructure. PLoS Biol. 2, e329 (2004).

  17. 17.

    & Array tomography: a new tool for imaging the molecular architecture and ultrastructure of neural circuits. Neuron 55, 25–36 (2007).

  18. 18.

    & Mapping brain circuitry with a light microscope. Nat. Methods 10, 515–523 (2013).

  19. 19.

    et al. Transgenic strategies for combinatorial expression of fluorescent proteins in the nervous system. Nature 450, 56–62 (2007).

  20. 20.

    et al. Correlations of neuronal and microvascular densities in murine cortex revealed by direct counting and colocalization of cell nuclei and microvessels. J. Neurosci. 18, 14553–14570 (2009).

  21. 21.

    et al. Large-scale automated histology in the pursuit of connectomes. J. Neurosci. 31, 16125–16138 (2011).

  22. 22.

    et al. Network anatomy and in vivo physiology of visual cortical neurons. Nature 471, 177–182 (2011).

  23. 23.

    , & Wiring specificity in the direction-selectivity circuit of the retina. Nature 471, 183–188 (2011).

  24. 24.

    , & Ultramicroscopy: three-dimensional visualization of neuronal networks in the whole mouse brain. Nat. Methods 4, 331–336 (2007).

  25. 25.

    , , & Three-dimensional imaging of the unsectioned adult spinal cord to assess axon regeneration and glial responses after injury. Nat. Med. 18, 166–171 (2012).

  26. 26.

    et al. Scale: a chemical approach for fluorescence imaging and reconstruction of transparent mouse brain. Nat. Neurosci. 14, 1481–1488 (2011).

  27. 27.

    et al. Fast, high-contrast imaging of animal development with scanned light sheet-based structured-illumination microscopy. Nat. Methods 7, 637–642 (2010).

  28. 28.

    , , , & Deep and fast live imaging with two-photon scanned light-sheet microscopy. Nat. Methods 8, 757–760 (2011).

  29. 29.

    , , & Quantitative high-speed imaging of entire developing embryos with simultaneous multiview light-sheet microscopy. Nat. Methods 9, 755–763 (2012).

  30. 30.

    , , , & Multiview light-sheet microscope for rapid in toto imaging. Nat. Methods 9, 730–733 (2012).

  31. 31.

    , , & Reconstruction of zebrafish early embryonic development by scanned light sheet microscopy. Science 322, 1065–1069 (2008).

  32. 32.

    et al. Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination. Nat. Methods 8, 417–423 (2011).

  33. 33.

    , , , & Superresolution imaging of chemical synapses in the brain. Neuron 68, 843–856 (2010).

  34. 34.

    , , , & Single-synapse analysis of a diverse synapse population: proteomic imaging methods and markers. Neuron 68, 639–653 (2010).

  35. 35.

    & Sequential immunofluorescence staining and image analysis for detection of large numbers of antigens in individual cell nuclei. Cytometry 47, 32–41 (2002).

  36. 36.

    et al. GFP reconstitution across synaptic partners (GRASP) defines cell contacts and synapses in living nervous systems. Neuron 57, 353–363 (2008).

  37. 37.

    & New technologies for imaging synaptic partners. Curr. Opin. Neurobiol. 22, 121–127 (2012).

  38. 38.

    & Motor control in a Drosophila taste circuit. Neuron 61, 373–384 (2009).

  39. 39.

    et al. mGRASP enables mapping mammalian synaptic connectivity with light microscopy. Nat. Methods 9, 96–102 (2012).

  40. 40.

    et al. Retrograde neuronal tracing with a deletion-mutant rabies virus. Nat. Methods 4, 47–49 (2007).

  41. 41.

    et al. Cortical representations of olfactory input by trans-synaptic tracing. Nature 472, 191–196 (2011).

  42. 42.

    et al. Optogenetic stimulation of a hippocampal engram activates fear memory recall. Nature 484, 381–385 (2012).

  43. 43.

    , , , & Permanent genetic access to transiently active neurons using targeted recombination in active populations (TRAP). Neuron (in the press).

  44. 44.

    , , & A study in developing visual systems with a new method of staining neurones and their processes in fixed tissue. Development 101, 697–713 (1987).

  45. 45.

    Cellular-resolution connectomics: challenges of dense neural circuit reconstruction. Nat. Methods 10, 501–507 (2013).

  46. 46.

    , & Functional clustering of neurons in motor cortex determined by cellular resolution imaging in awake behaving mice. J. Neurosci. 29, 13751–13760 (2009).

  47. 47.

    et al. Brain-wide neuronal dynamics during motor adaptation in zebrafish. Nature 485, 471–477 (2012).

  48. 48.

    et al. Long-term dynamics of CA1 hippocampal place codes. Nat. Neurosci. 16, 264–266 (2013).

  49. 49.

    , , , & Whole-brain functional imaging at cellular resolution using light-sheet microscopy. Nat. Methods 10, 413–420 (2013).

  50. 50.

    et al. The brain activity map. Science 339, 1284–1285 (2013).

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Acknowledgements

We acknowledge all members of the Deisseroth laboratory for discussions and support. This work was funded by a US National Institutes of Health Director's Transformative Research Award (TR01) to K.D. from the National Institute of Mental Health, as well as by the National Science Foundation, the Simons Foundation, the President and Provost of Stanford University, and the Howard Hughes Medical Institute. K.D. is also funded by the National Institute on Drug Abuse and the Defense Advanced Research Projects Agency Reorganization and Plasticity to Accelerate Injury Recovery program, and the Wiegers, Snyder, Reeves, Gatsby, and Yu Foundations. K.C. is supported by the Burroughs Wellcome Fund Career Award at the Scientific Interface.

Author information

Affiliations

  1. Department of Bioengineering, Stanford University, Stanford, California, USA.

    • Kwanghun Chung
    •  & Karl Deisseroth
  2. CNC Program, Stanford University, Stanford, California, USA.

    • Kwanghun Chung
    •  & Karl Deisseroth
  3. Department of Psychiatry and Behavioral Sciences, Stanford University, Stanford, California, USA.

    • Karl Deisseroth
  4. Howard Hughes Medical Institute, Stanford University, Stanford, California, USA.

Authors

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Competing interests

K.C. and K.D. have disclosed these findings to the Stanford Office of Technology Licensing, which is filing a patent application to ensure broad public use of the methods in microscopy systems and for studying disease mechanisms and treatments. All protocols and methods remain freely available for academic and non-profit research in perpetuity, and supported by the authors, through the CLARITY website (http://clarityresourcecenter.org/).

Corresponding authors

Correspondence to Kwanghun Chung or Karl Deisseroth.

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DOI

https://doi.org/10.1038/nmeth.2481

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