Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

Minimally invasive high-speed imaging of sarcomere contractile dynamics in mice and humans

Nature volume 454, pages 784–788 (2008)Cite this article

Abstract

Sarcomeres are the basic contractile units of striated muscle. Our knowledge about sarcomere dynamics has primarily come from in vitro studies of muscle fibres1 and analysis of optical diffraction patterns obtained from living muscles2,3. Both approaches involve highly invasive procedures and neither allows examination of individual sarcomeres in live subjects. Here we report direct visualization of individual sarcomeres and their dynamical length variations using minimally invasive optical microendoscopy4 to observe second-harmonic frequencies of light generated in the muscle fibres5,6 of live mice and humans. Using microendoscopes as small as 350 μm in diameter, we imaged individual sarcomeres in both passive and activated muscle. Our measurements permit in vivo characterization of sarcomere length changes that occur with alterations in body posture and visualization of local variations in sarcomere length not apparent in aggregate length determinations. High-speed data acquisition enabled observation of sarcomere contractile dynamics with millisecond-scale resolution. These experiments point the way to in vivo imaging studies demonstrating how sarcomere performance varies with physical conditioning and physiological state, as well as imaging diagnostics revealing how neuromuscular diseases affect contractile dynamics.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Minimally invasive microendoscopy system.
Figure 2: Static imaging of individual sarcomeres.
Figure 3: Dynamic imaging of mouse sarcomeres.
Figure 4: Dynamic imaging of human sarcomeres.

Similar content being viewed by others

References

  1. Telley, I. A. & Denoth, J. Sarcomere dynamics during muscular contraction and their implications to muscle function. J. Muscle Res. Cell Motil. 28, 89–104 (2007)

    Article  Google Scholar 

  2. Lieber, R. L., Loren, G. J. & Friden, J. In vivo measurement of human wrist extensor muscle sarcomere length changes. J. Neurophysiol. 71, 874–881 (1994)

    Article  CAS  Google Scholar 

  3. Yeh, Y., Baskin, R. J., Lieber, R. L. & Roos, K. P. Theory of light diffraction by single skeletal muscle fibers. Biophys. J. 29, 509–522 (1980)

    Article  CAS  Google Scholar 

  4. Jung, J. C. & Schnitzer, M. J. Multiphoton endoscopy. Opt. Lett. 28, 902–904 (2003)

    Article  ADS  Google Scholar 

  5. Campagnola, P. J. & Loew, L. M. Second-harmonic imaging microscopy for visualizing biomolecular arrays in cells, tissues and organisms. Nature Biotechnol. 21, 1356–1360 (2003)

    Article  CAS  Google Scholar 

  6. Zipfel, W. R. et al. Live tissue intrinsic emission microscopy using multiphoton-excited native fluorescence and second harmonic generation. Proc. Natl Acad. Sci. USA 100, 7075–7080 (2003)

    Article  ADS  CAS  Google Scholar 

  7. Huxley, A. F. & Niedergerke, R. Structural changes in muscle during contraction; interference microscopy of living muscle fibres. Nature 173, 971–973 (1954)

    Article  ADS  CAS  Google Scholar 

  8. Huxley, H. & Hanson, J. Changes in the cross-striations of muscle during contraction and stretch and their structural interpretation. Nature 173, 973–976 (1954)

    Article  ADS  CAS  Google Scholar 

  9. Laing, N. G. & Nowak, K. J. When contractile proteins go bad: the sarcomere and skeletal muscle disease. Bioessays 27, 809–822 (2005)

    Article  CAS  Google Scholar 

  10. Delp, S. L. et al. OpenSim: open-source software to create and analyze dynamic simulations of movement. IEEE Trans. Biomed. Eng. 54, 1940–1950 (2007)

    Article  Google Scholar 

  11. Cutts, A. The range of sarcomere lengths in the muscles of the human lower limb. J. Anat. 160, 79–88 (1988)

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Goulding, D., Bullard, B. & Gautel, M. A survey of in situ sarcomere extension in mouse skeletal muscle. J. Muscle Res. Cell Motil. 18, 465–472 (1997)

    Article  CAS  Google Scholar 

  13. Jung, J. C., Mehta, A. D., Aksay, E., Stepnoski, R. & Schnitzer, M. J. In vivo mammalian brain imaging using one- and two-photon fluorescence microendoscopy. J. Neurophysiol. 92, 3121–3133 (2004)

    Article  Google Scholar 

  14. Levene, M. J., Dombeck, D. A., Kasischke, K. A., Molloy, R. P. & Webb, W. W. In vivo multiphoton microscopy of deep brain tissue. J. Neurophysiol. 91, 1908–1912 (2004)

    Article  Google Scholar 

  15. Rothstein, E. C., Carroll, S., Combs, C. A., Jobsis, P. D. & Balaban, R. S. Skeletal muscle NAD(P)H two-photon fluorescence microscopy in vivo: topology and optical inner filters. Biophys. J. 88, 2165–2176 (2005)

    Article  CAS  Google Scholar 

  16. Hoppeler, H. & Fluck, M. Plasticity of skeletal muscle mitochondria: structure and function. Med. Sci. Sports Exerc. 35, 95–104 (2003)

    Article  CAS  Google Scholar 

  17. Plotnikov, S. V., Millard, A. C., Campagnola, P. J. & Mohler, W. A. Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres. Biophys. J. 90, 693–703 (2006)

    Article  ADS  CAS  Google Scholar 

  18. Schurmann, S., Weber, C., Fink, R. H. & Vogel, M. in Multiphoton Microscopy in the Biomedical Sciences VII Vol. 6442 (eds Ammasi, P. & Peter, T. C. S.) 64421U (SPIE, Bellingham, WA, 2007)

    Book  Google Scholar 

  19. Chu, S. W. et al. Studies of chi(2)/chi(3) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy. Biophys. J. 86, 3914–3922 (2004)

    Article  ADS  CAS  Google Scholar 

  20. Moreaux, L., Sandre, O., Charpak, S., Blanchard-Desce, M. & Mertz, J. Coherent scattering in multi-harmonic light microscopy. Biophys. J. 80, 1568–1574 (2001)

    Article  CAS  Google Scholar 

  21. Zoumi, A., Yeh, A. & Tromberg, B. J. Imaging cells and extracellular matrix in vivo by using second-harmonic generation and two-photon excited fluorescence. Proc. Natl Acad. Sci. USA 99, 11014–11019 (2002)

    Article  ADS  CAS  Google Scholar 

  22. Perreault, E. J., Day, S. J., Hulliger, M., Heckman, C. J. & Sandercock, T. G. Summation of forces from multiple motor units in the cat soleus muscle. J. Neurophysiol. 89, 738–744 (2003)

    Article  Google Scholar 

  23. Campagnola, P. J. et al. Three-dimensional high-resolution second-harmonic generation imaging of endogenous structural proteins in biological tissues. Biophys. J. 82, 493–508 (2002)

    Article  ADS  CAS  Google Scholar 

  24. Delp, S. L. et al. An interactive graphics-based model of the lower extremity to study orthopaedic surgical procedures. IEEE Trans. Biomed. Eng. 37, 757–767 (1990)

    Article  CAS  Google Scholar 

  25. Pellegrino, M. A. et al. Orthologous myosin isoforms and scaling of shortening velocity with body size in mouse, rat, rabbit and human muscles. J. Physiol. (Lond.) 546, 677–689 (2003)

    Article  CAS  Google Scholar 

  26. Clarkson, E., Costa, C. F. & Machesky, L. M. Congenital myopathies: diseases of the actin cytoskeleton. J. Pathol. 204, 407–417 (2004)

    Article  CAS  Google Scholar 

  27. Plotnikov, S. V. et al. Measurement of muscle disease by quantitative second-harmonic generation imaging. J. Biomed. Opt. (in the press)

  28. Ponten, E., Gantelius, S. & Lieber, R. L. Intraoperative muscle measurements reveal a relationship between contracture formation and muscle remodeling. Muscle Nerve 36, 47–54 (2007)

    Article  Google Scholar 

  29. Lieber, R. L., Murray, W. M., Clark, D. L., Hentz, V. R. & Friden, J. Biomechanical properties of the brachioradialis muscle: implications for surgical tendon transfer. J. Hand Surg. (Am.) 30, 273–282 (2005)

    Article  Google Scholar 

  30. Manal, K., Gonzalez, R. V., Lloyd, D. G. & Buchanan, T. S. A real-time EMG-driven virtual arm. Comput. Biol. Med. 32, 25–36 (2002)

    Article  Google Scholar 

Download references

Acknowledgements

We thank M. Cromie and B. Flusberg for technical assistance, M. Cromie and A. Lewis for assistance with the human subject protocol, D. Profitt for expert machining, and Y. Goldman, R. Lieber, F. Zajac and T. Sanger for discussions. This work was supported by the Coulter Foundation (S.L.D. and M.J.S.), a Stanford Bio-X Interdisciplinary Initiatives award (S.L.D. and M.J.S.), NINDS R01NS050533 (M.J.S.), the Stanford-NIH Medical Scientist Training Program (M.E.L.), the Stanford-NIH Biophysics Training Grant (R.P.J.B.), and an equipment donation from Prairie Technologies, Inc. (M.J.S.).

Author Contributions All authors designed the experiments and interpreted data. M.E.L. and R.P.J.B collected data. M.E.L. performed the analysis. All authors discussed the results and contributed to the text.

Author information

Authors and Affiliations

  1. Bio-X Program, James H. Clark Center for Biomedical Engineering & Sciences, Stanford University, Stanford, California 94305, USA ,

    Michael E. Llewellyn, Robert P. J. Barretto, Scott L. Delp & Mark J. Schnitzer

Authors
  1. Michael E. Llewellyn
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Robert P. J. Barretto
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Scott L. Delp
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Mark J. Schnitzer
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Mark J. Schnitzer.

Ethics declarations

Competing interests

This research involved the use of equipment that was a gift to Stanford University from Prairie Technologies. Stanford University has filed patent applications based on part of the work documented in this manuscript.

Supplementary information

Supplementary Information 1

This file contains Supplementary Figure 1 and Legend, Supplementary Video 1 Legend, and Supplementary Methods with additional references. (PDF 766 kb)

Supplementary Video

This file contains Supplementary Video 1 which presents a three-dimensional stack of images acquired by second-harmonic microendoscopy from the lateral gastrocnemius muscle of a living mouse. (MOV 5779 kb)

PowerPoint slides

Rights and permissions

Reprints and permissions

About this article

Cite this article

Llewellyn, M., Barretto, R., Delp, S. et al. Minimally invasive high-speed imaging of sarcomere contractile dynamics in mice and humans. Nature 454, 784–788 (2008). https://doi.org/10.1038/nature07104

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature07104

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing