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Histochemistry

, Volume 100, Issue 3, pp 193–202 | Cite as

Remodelling of cardiomyocyte cytoarchitecture visualized by three-dimensional (3D) confocal microscopy

  • J. Marius Messerli
  • Monika E. Eppenberger-Eberhardt
  • Barbara M. Rutishauser
  • Patrick Schwarb
  • P. von Arx
  • S. Koch-Schneidemann
  • Hans M. Eppenberger
  • Jean-Claude Perriard
Originals

Abstract

The break-down and reassembly of myofibrils in long-term cultures of adult rat cardiomyocytes was investigated by a novel combination of confocal laser scanning microscopy and three-dimensional image reconstruction, referred to as FTCS, to visualize the morphological changes these cells undergo in culture. FTCS is discussed as an alternative imaging mode to low-magnification scanning electron microscopy. The three-dimensional shape of the cells are correlated with the assembly state of myofibrils in different stages. Based on immunofluorescence and confocal laser scanning microscopy it was shown that myofibrils are degraded within a few days after plating and that newly assembled myofibrils are predominantly confined to the continuous area in the perinuclear region close to the membrane in contact with the substratum. The localization of myofibrils along the cell's vertical axis has been investigated both by optical sectioning using confocal light microscopy and by physical sectioning following by transmission electron microscopy. Based on the distribution of myofibrillar proteins we propose a model of myofibrillar growth locating the putative assembly sites to a region concentric around the nuclei. We provide evidence that the cell shape is dominated by the myofibrillar apparatus.

Keywords

Confocal Laser Scanning Microscopy Image Reconstruction Assembly State Optical Section Myofibrillar Protein 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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References

  1. Antin PB, Tokunaka S, Nachmias VT, Holtzer H (1986) Role of stress fiber-like structures in assembling nascent myofibrils in myosheets recovering from exposure to ethyl methanesulfonate. J Cell Biol 102:1464–1479Google Scholar
  2. Atherton BT, Meyer DM, Simpson DG (1986) Assembly and remodelling of myofibrils and intercalated discs in cultured neonatal rat heart cells. J Cell Sci 86:233–248Google Scholar
  3. Bähler M, Moser H, Eppenberger HM, Wallimann T (1985) Heart C-protein is transiently expressed during skeletal muscle development in the embryo, but persists in cultured myogenic cells. Dev Biol 112:345–352Google Scholar
  4. Borg TK, Rubin K, Lundgren E, Borg K, Obrink B (1984) Recognition of extracellular matrix components by neonatal and adult cardiac myocytes. Dev Biol 104:86–96Google Scholar
  5. Borisov AB, Goncharova EI, Pinaev GP, Rumiantsev PP (1989) Changes in alpha-actinin localization and myofibrillogenesis in rat cardiomyocytes under cultivation. Tsitologiia 31:642–646Google Scholar
  6. Bouvagnet P, Léger J, Pons F, Dechesne CA, Léger JJ (1984) Fiber types and myosin in human atrial and ventricular myocardium. Circ Res 55:794–804Google Scholar
  7. Claycomb WC, Palazzo MC (1980) Culture of terminally differentiated adult cardiac muscle cells. A light and scanning electron microscope study. Dev Biol 80:466–482Google Scholar
  8. Danowski BA, Imanakayoshida K, Sanger JM, Sanger JW (1992) Costameres are sites of force transmission to the substratum in adult rat cardiomyocytes. J Cell Biol 118:1411–1420Google Scholar
  9. De Clerck NM, Clases VA, Brutsaert DL (1984) Uniform sarcomere behavior during twitch of intact single cardiac cells. J Mol Cell Cardiol 16:735–745Google Scholar
  10. Decker ML, Behnke-Barclay M, Cook MG, LaPres JJ, Clark WA, Decker RS (1991) Cell shape and organization of the contractile apparatus in cultured adult cardiac myocytes. J Mol Cell Cardiol 23:817–832Google Scholar
  11. Dlugosz AA, Antin PB, Nachmias VT, Holtzer H (1984) The relationship between stress fiber-like structures and nascent myofibrils in cultured cardiac myocytes. J Cell Biol 99:2268–2278Google Scholar
  12. Eppenberger ME, Hauser I, Baechi T, Schaub MC, Brunner UT, Dechesne CA, Eppenberger HM (1988) Immunocytochemical analysis of the regeneration of myofibrils in long-term cultures of adult cardiomyocytes of the rat. Dev Biol 130:1–15Google Scholar
  13. Eppenberger-Eberhardt ME, Flamme I, Kurer V, Eppenberger HM (1990) Reexpression of α-smooth muscle actin isoform in cultured adult rat cardiomyocytes. Dev Biol 139:269–278Google Scholar
  14. Eppenberger-Eberhardt M, Riesinger I, Messerli JM, Schwarb P, Müller M, Eppenberger HM, Wallimann T (1991) Adult rat cardiomyocytes cultured in creatine-deficient medium display large mitochondria with paracrystalline inclusions enriched for creatine kinase. J Cell Biol 113:289–302Google Scholar
  15. Gerdes AM (1992) Isolated cardiac myocytes from adult rat heart. Trends Cardiovasc Med 2:152–155Google Scholar
  16. Goncharova EJ, Kam Z, Geiger B (1992) The involvement of adherens junction components in myofibrillogenesis in cultured cardiac myocytes. Development 114:173Google Scholar
  17. Grove BK, Kurer V, Lehner C, Doetschman TC, Perriard JC, Eppenberger HM (1984) Monoclonal antibodies detect new 185000 dalton muscle M-line protein. J Cell Biol 98:518–524Google Scholar
  18. Guo JX, Jacobson SL, Brown DL (1986) Rearrangement of tubulin, actin, and myosin in cultured ventricular cardiomyocytes of the adult rat. Cell Motil Cytoskel 6:291–304Google Scholar
  19. Hilenski LL, Terracio L, Sawyer R, Borg TK (1989) Effects of extracellular matrix on cytoskeletal and myofibrillar organization in vitro. Scanning Microsc 3:535–548Google Scholar
  20. Hilenski LL, Ma XH, Vinson N, Terracio L, Borg TK (1992) The role of beta 1 integrin in spreading and myofibrillogenesis in neonatal rat cardiomyocytes in vitro. Cell Motil Cytoskel 21:87–100Google Scholar
  21. Hill CS, Duran S, Lin ZX, Weber K and Holtzer H (1986) Titin and myosin, but not desmin, are linked during myofibrillogenesis in postmitotic mononucleated myoblasts. J Cell Biol 103:2185–2196Google Scholar
  22. LoRusso SM, Imanakayoshida K, Shuman H, Sanger JM, Sanger JW (1992) Incorporation of fluorescently labeled contractile proteins into freshly isolated living adult cardiac myocytes. Cell Motil Cytoskel 21:111–122Google Scholar
  23. Lu MH, Dilullo C, Schultheiss T, Holtzer S, Murray JM, Choi J, Fischman DA, Holtzer H (1992) The vinculin/sarcomeric-alpha-actinin/alpha-actin nexus in cultured cardiac myocytes. J Cell Biol 117:1007–1022Google Scholar
  24. Luft JJ (1960) Improvements in epoxy resin embedding methods. J Biophys Biochem Cytol 9:409–414Google Scholar
  25. Lundgren E, Terracio L, Mardh S, Borg TK (1985) Extracellular matrix components influence the survival of adult cardiac myocytes in vivo. Exp Cell Res 158:371–381Google Scholar
  26. Messerli JM, van der Voort HTM, Rungger-Brändle E, Perriard J-C (1993) Three-dimensional visualization of multi-channel voxel data. Cytometry Vol. 14, Nr. 7:725–735Google Scholar
  27. Minsky M (1957). Microscopy Apparaturs. U.S. patent. 3013467.Google Scholar
  28. Nag AC, Cheng M, Fischman DA, Zak R (1983) Long-term cell culture of adult mammalian cardiac myocytes: electron microscopic and immunofluorescent analysis of myofibrillar structure. J Mol Cell Cardiol 15:301–317Google Scholar
  29. Rappaport L, Samuel JL (1988) Microtubules in cardiac myocytes. Int Rev Cytol 113:101–143Google Scholar
  30. Reynolds ES (1963) The use of lead citrate at high pH as an electron opaque stain in electron microscopy. J Cell Biol 17:208–212Google Scholar
  31. Sheppard CJR (1987) Scanning optical microscopy, 10. Academic Press, LondonGoogle Scholar
  32. Skalli O, Ropraz P, Trzeciak A, Benzonana DG, Gabbiani G (1986) A monoclonal antibody against α-smooth muscle actin: a new probe for smooth muscle differentiation. J Cell Biol 193:2787–2796Google Scholar
  33. Soldati T, Perriard J-C (1991) Intracompartmental sorting of essential myosin light chains: molecular dissection and in vivo monitoring by epitope tagging. Cell 66:277–289Google Scholar
  34. Stemmer P, Wisler PL, Watanabe AM (1992) Isolated myocytes in experimental cardiology. In: Fozzard HA (ed) The heart and cardiovascular system. Raven Press, New York, pp 387–404Google Scholar
  35. Terracio L, Simpson DG, Hilenski L, Carver W, Decker RS, Vinson N, Borg TK (1990) Distribution of vinculin in the z-disk of striated muscle, analysis by laser scanning confocal microscopy. J Cell Physiol 145:78–87Google Scholar
  36. Terracio L, Rubin K, Gullberg D, Balog E, Carver W, Jyring R, Borg TK (1991) Expression of collagen binding integrins during cardiac development and hypertrophy. Circ Res 68:734–744Google Scholar
  37. van der Voort HTM, Brakenhoff GJ (1990) 3-D image formation in high-aperture fluorescence confocal microscopy: a numerical analysis. J Microsc 158:43–54Google Scholar
  38. van der Voort HTM, Messerli JM, Noordmans HJ, Smeulders AWM (1993) Volume visualization for interactive microscopic image analysis. Bioimaging 1:20–29Google Scholar
  39. Wang SM, Greaser ML, Schultz E, Bulinski JC, Lin JJ, Lessard JL (1988) Studies on cardiac myofibrillogenesis with antibodies to titin, actin, tropomyosin, and myosin. J Cell Biol 107:1075–1083Google Scholar
  40. Wijnaends van Resandt RW, Marsman HJB, Kaplan R, Davoust J, Stelzer EKH, Stricker R (1985) Optical fluorescence microscopy in 3 dimensions: microtomoscopy. J Microsc 138:29–34Google Scholar

Copyright information

© Springer-Verlag 1993

Authors and Affiliations

  • J. Marius Messerli
    • 1
  • Monika E. Eppenberger-Eberhardt
    • 1
  • Barbara M. Rutishauser
    • 1
  • Patrick Schwarb
    • 1
  • P. von Arx
    • 1
  • S. Koch-Schneidemann
    • 1
  • Hans M. Eppenberger
    • 1
  • Jean-Claude Perriard
    • 1
  1. 1.Institute for Cell BiologySwiss Federal Institute of TechnologyZürichSwitzerland

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