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Carbon fiber technique for the investigation of single-cell mechanics in intact cardiac myocytes

Nature Protocols volume 1, pages 1453–1457 (2006)Cite this article

Abstract

This protocol describes a method for attaching single isolated cardiac myocytes to carbon fibers for mechanical manipulation and measurement. This method relies on cell-adhesive carbon fibers that attach easily to the cell membrane without causing damage, and is thus applicable to intact myocytes. To connect the carbon fiber to micromanipulators, a fiber holder with glass capillaries must first be fabricated. After connection of the fibers to the micromanipulators, firm attachment is easily established by gently pressing the fiber tip onto the cell membrane. Unlike other methods, this technique does not require vast technical expertise, and therefore greatly facilitates experiments. This method enables detection of the effect of drugs, genetic defects or the expression of exogenous proteins on both active and passive properties of cardiac myocytes. In combination with other experimental procedures, this technique can also be applied to the study of mechano-transduction. This protocol can be completed in 3.5 h.

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Figure 1: Diagram of the recommended experimental setup for the simultaneous measurement of sarcomere length (SL) and fluorescent emissions of a voltage-sensitive indicator (di-8 ANNEPS) at 560 and 620 nm.
Figure 2: Capillary-held carbon fiber.
Figure 3: Stretch experiment.

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References

  1. Brady, A.J. Mechanical properties of isolated cardiac myocytes. Physiol. Rev. 71, 413–428 (1991).

    Article  CAS  Google Scholar 

  2. Garnier, D. Attachment procedures for mechanical manipulation of isolated cardiac myocytes: a challenge. Cardiovasc. Res. 28, 1958–1964 (1994).

    Article  Google Scholar 

  3. Tarr, M., Trank, J.W., Leiffer, P. & Shepherd, N. Sarcomere length-resting tension relation in single frog atrial cardiac cells. Circ. Res. 45, 554–559 (1979).

    Article  CAS  Google Scholar 

  4. Tung, L. An ultrasensitive transducer for measurement of isometric contractile force from single heart cells. Pflugers Arch. 407, 109–115 (1986).

    Article  CAS  Google Scholar 

  5. Fabiato, A. Myoplasmic free calcium concentration reached during the twitch of an intact isolated cardiac cell and during calcium-induced release of calcium from the sarcoplasmic reticulum of a skinned cardiac cell from the adult rat or rabbit ventricle. J. Gen. Physiol. 78, 457–497 (1981).

    Article  CAS  Google Scholar 

  6. Brady, A.J., Tan, S.T. & Ricchiuti, N.V. Contractile force measured in unskinned isolated rat heart fibers. Nature 282, 728–729 (1979).

    Article  CAS  Google Scholar 

  7. Palmer, R.E., Brady, A.J. & Roos, K.P. Mechanical measurement from isolated cardiac myocytes using a pipette attachment system. Am. J. Physiol. 270, C697–C704 (1996).

    Article  CAS  Google Scholar 

  8. Hofmann, P.A. & Moss, R.L. Effects of calcium on shortening velocity in frog chemically skinned atrial myocytes and in mechanically disrupted ventricular myocardium from rat. Circ. Res. 70, 885–892 (1992).

    Article  CAS  Google Scholar 

  9. Metzger, J.M. et al. Skeletal troponin C reduces contractile sensitivity to acidosis in cardiac myocytes from transgenic mice. Proc. Natl. Acad. Sci. USA 90, 9036–9040 (1993).

    Article  CAS  Google Scholar 

  10. Bluhm, W.F., McCulloch, A.D. & Lew, W.Y. Active force in rabbit ventricular myocytes. J. Biomech. 28, 1119–1122 (1995).

    Article  CAS  Google Scholar 

  11. Le Guennec, J.-Y., Peineau, N., Argibay, J.A., Mongo, K.G. & Garnier, D. A new method of attachment of isolated mammalian ventricular myocytes for tension recording: length dependence of passive and active tension. J. Mol. Cell. Cardiol. 22, 1083–1093 (1990).

    Article  CAS  Google Scholar 

  12. Yasuda, S.I. et al. A novel method to study contraction characteristics of a single cardiac myocyte using carbon fibers. Am. J. Physiol. Heart Circ. Physiol. 281, H1442–H1446 (2001).

    Article  CAS  Google Scholar 

  13. Nishimura, S. et al. Single cell mechanics of rat cardiomyocytes under isometric, unloaded and physiologically loaded conditions. Am. J. Physiol. 287, H196–H202 (2004).

    CAS  Google Scholar 

  14. Yasuda, S. et al. Unloaded shortening increases the peak of Ca2+ transients but accelerates their decay in rat single cardiac myocytes. Am. J. Physiol. 285, H470–H475 (2003).

    CAS  Google Scholar 

  15. Yasuda, S. et al. Dystrophic heart failure blocked by membrane sealant poloxamer. Nature 463, 1025–1029 (2005).

    Article  Google Scholar 

  16. Nishimura, S. et al. Contractile dysfunction of cardiomyopathic hamster myocytes is pronounced under high load conditions. J. Mol. Cell. Cardiol. 39, 231–239 (2005).

    Article  CAS  Google Scholar 

  17. White, E., Boyett, M.R. & Orchard, C.H. The effects of mechanical loading and changes of length on single guinea-pig ventricular myocytes. J. Physiol. 482, 93–107 (1995).

    Article  CAS  Google Scholar 

  18. Nishimura, S. et al. Expression of green fluorescent protein impairs the force-generating ability of isolated rat ventricular cardiomyocytes. Mol. Cell. Biochem. 286, 59–65 (2006).

    Article  CAS  Google Scholar 

  19. Michele, D.E., Albayya, F.P. & Metzger, J.M. Direct, convergent hypersensitivity of calcium-activated force generation produced by hypertrophic cardiomyopathy mutant alpha-tropomyosins in adult cardiac myocytes. Nat. Med. 5, 1413–1417 (1999).

    Article  CAS  Google Scholar 

  20. Yoneda, M. Force exerted by a single cilium of Mytilus edulus . I. Exp. Biol. 37, 461–468 (1960).

    Google Scholar 

  21. Nishimura, S. et al. Microtubule modulates the stiffness of cardiomyocytes against shear stress. Circ. Res. 98, 81–87 (2006).

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Human and Engineered Environmental Studies, Computational Biomechanics Laboratory, Graduate School of Frontier Sciences, University of Tokyo, Bunkyo-ku, Tokyo, 113-0033, Japan

    Seiryo Sugiura

  2. Department of Cardiovascular Medicine, Graduate School of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, 113-8655, Japan

    Satoshi Nishimura, Soichiro Yasuda & Yumiko Hosoya

  3. Neuroscience Research Institute, National Institute of Advanced Industrial Science and Technology, Tsukuba, 305-8568, Ibaraki, Japan

    Kaoru Katoh

Authors
  1. Seiryo Sugiura
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  2. Satoshi Nishimura
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  3. Soichiro Yasuda
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  4. Yumiko Hosoya
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  5. Kaoru Katoh
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Correspondence to Seiryo Sugiura.

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Supplementary information

Supplementary Video

Length perturbation experiment in a single rat cardiac myocyte. The length of a cardiomyocyte was changed cyclically by applying a sinusoidal command of varying frequency (0.5 to 10 Hz) to the piezo-electric actuator connected to the carbon fiber. (MPG 3265 kb)

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Sugiura, S., Nishimura, S., Yasuda, S. et al. Carbon fiber technique for the investigation of single-cell mechanics in intact cardiac myocytes. Nat Protoc 1, 1453–1457 (2006). https://doi.org/10.1038/nprot.2006.241

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