Skip to main content
Log in

The Role of F-Actin in Hypo-Osmotically Induced Cell Volume Change and Calcium Signaling in Anulus Fibrosus Cells

  • Published:
Annals of Biomedical Engineering Aims and scope Submit manuscript

Abstract

Loading of the spine induces dynamic changes in the osmotic environment of the intervertebral disc (IVD) due to the exudation and recovery of tissue water. Cells from the anulus fibrosus (AF) respond to osmotic stress with altered biosynthesis through a pathway that may involve calcium (Ca2+) as a second messenger. We examined the hypothesis that AF cells respond to hypo-osmotic stress by swelling and initiating regulatory volume decrease (RVD). Further, the role of F-actin disruption and transient increases in intracellular calcium concentration ([Ca2+] i ) in volume adaptation were studied. In response to hypo-osmotic stress, AF cells swelled, disrupted F-actin, and exhibited [Ca2+] i transients in proportion to the magnitude of the stress. The transient disruption of F-actin was dependent on the presence of extracellular Ca2+. After swelling, AF cells underwent RVD at all magnitudes of hypo-osmotic stress. The extent of RVD was diminished significantly by F-actin breakdown using cytochalasin D or by inhibition of swelling-induced F-actin disruption by removing extracellular Ca2+. Swelling-induced disruption of F-actin facilitated RVD, as evidenced by a more rapid volume recovery with increased F-actin breakdown. In conclusion, our findings suggest that the F-actin network plays an important role in the response of AF cells to osmotic stress.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  • 1Alexopoulos, L. G., G. R. Erickson, and F. Guilak. A method for quantifying cell size from differential interference contrast images: Validation and application to osmotically stressed chondrocytes. J. Microsc. 205:125-135, 2002.

    Google Scholar 

  • 2Antoniou, J., T. Steffen, F. Nelson, N. Winterbottom, A. P. Hollander, R. A. Poole, M. Aebi, and M. Alini. The human lumbar intervertebral disc: Evidence for changes in the biosynthesis and denaturation of the extracellular matrix with growth, maturation, ageing, and degeneration. J. Clin. Invest. 98:996-1003, 1996.

    Google Scholar 

  • 3Asada, S., K. Fukuda, F. Nishisaka, M. Matsukawa, and C. Hamanisi. Hydrogen peroxide induces apoptosis of chondrocytes; involvement of calcium ion and extracellular signal-regulated protein kinase. Inflamm. Res. 50:19-23, 2001.

    Google Scholar 

  • 4Baer, A. E., T. A. Laursen, F. Guilak, and L. A. Setton. The micromechanical environment of intervertebral disc cells determined by a finite deformation, anisotropic, and biphasic finite element model. J. Biomech. Eng. 125:1-11, 2003.

    Google Scholar 

  • 5Baer, A. E., and L. A. Setton. The micromechanical environment of intervertebral disc cells: Effect of matrix anisotropy and cell geometry predicted by a linear model. J. Biomech. Eng. 122:245-251, 2000.

    Google Scholar 

  • 6Baer, A. E., J. Y. Wang, V. B. Kraus, and L. A. Setton. Collagen gene expression and mechanical properties of intervertebral disc cell-alginate cultures. J. Orthop. Res. 19:2-10, 2001.

    Google Scholar 

  • 7Buckwalter, J. A. Aging and degeneration of the human intervertebral disc. Spine 20:1307-1314, 1995.

    Google Scholar 

  • 8Cantiello, H. F. Role of the actin cytoskeleton on epithelial Na+ channel regulation. Kidney Int. 48:970-984, 1995.

    Google Scholar 

  • 9Cantiello, H. F. Role of actin filament organization in cell volume and ion channel regulation. J. Exp. Zool. 279:425-435, 1997.

    Google Scholar 

  • 10Chen, J., A. E. Baer, P. Y. Paik, W. Yan, and L. A. Setton. Matrix protein gene expression in intervertebral disc cells subjected to altered osmolarity. Biochem. Biophys. Res. Commun. 293:932-938, 2002.

    Google Scholar 

  • 11Colliou, H. F. Matrix disorganization, apoptosis and gene expression in the intervertebral disc are modulated by compressive loading: A mouse model for disc degeneration. Trans. Orthop. Res. Soc. 23:189, 1998.

    Google Scholar 

  • 12Cornet, M., Y. Isobe, and L. F. Lemanski. Effects of anisosmotic conditions on the cytoskeletal architecture of cultured PC12 cells. J. Morphol. 222:269-286, 1994.

    Google Scholar 

  • 13Dascalu, A., R. Korenstein, Y. Oron, and Z. Nevo. A hyperosmotic stimulus regulates intracellular pH, calcium, and S-100 protein levels in avian chondrocytes. Biochem. Biophys. Res. Commun. 227:368-373, 1996.

    Google Scholar 

  • 14Dascalu, A., A. Matithyou, Y. Oron, and R. Korenstein. A hyperosmotic stimulus elevates intracellular calcium and inhibits proliferation of a human keratinocyte cell line. J. Invest. Dermatol. 115:714-718, 2000.

    Google Scholar 

  • 15Erickson, G. R., L. G. Alexopoulos, and F. Guilak. Hyper-osmotic stress induces volume change and calcium transients in chondrocytes by transmembrane, phospholipid, and G-protein pathways. J. Biomech. 34:1527-1535, 2001.

    Google Scholar 

  • 16Erickson, G. R., D. L. Northrup, and F. Guilak. Hypo-osmotic stress induces calcium-dependent actin reorganization in articular chondrocytes. Osteoarthritis Cartilage 11:187-197, 2003.

    Google Scholar 

  • 17Gual, P., S. Shigematsu, M. Kanzaki, T. Gremeaux, T. Gonzalez, J. E. Pessin, Y. LeMarchand-Brustel, and J. F. Tanti. A Crk-II/TC10 signaling pathway is required for osmotic shock-stimulated glucose transport. J. Biol. Chem. 277:43980-43986, 2002.

    Google Scholar 

  • 18Guilak, F., G. R. Erickson, and H. P. Ting-Beall. The effects of osmotic stress on the viscoelastic and physical properties of articular chondrocytes. Biophys. J. 82:720-727, 2002.

    Google Scholar 

  • 19Guilak, F., H. P. Ting-Beall, A. E. Baer, W. R. Trickey, G. R. Erickson, and L. A. Setton. Viscoelastic properties of intervertebral disc cells. Identification of two biomechanically distinct cell populations. Spine 24:2475-2483, 1999.

    Google Scholar 

  • 20Guizouarn, H., R. Motais, F. Garcia-Romeu, and F. Borgese. Cell volume regulation: The role of taurine loss in maintaining membrane potential and cell pH. J. Physiol. 523(Pt. 1):147-154, 2000.

    Google Scholar 

  • 21Hallows, K. R., F. Y. Law, C. H. Packman, and P. A. Knauf. Changes in cytoskeletal actin content, F-actin distribution, and surface morphology during HL-60 cell volume regulation. J. Cell Physiol. 167:60-71, 1996.

    Google Scholar 

  • 22Hardingham, G. E., and H. Bading. Calcium as a versatile second messenger in the control of gene expression. Microsc. Res. Tech. 46:348-355, 1999.

    Google Scholar 

  • 23Henson, J. H. Relationships between the actin cytoskeleton and cell volume regulation. Microsc. Res. Tech. 47:155-162, 1999.

    Google Scholar 

  • 24Hildebrandt, J. P., and A. Prowald. Ca(2+) and p38 MAP kinase regulate mAChR-mediated c-Fos expression in avian exocrine cells. Am. J. Physiol. Cell. Physiol. 278:C879-884, 2000.

    Google Scholar 

  • 25Hutton, W. C., T. M. Ganey, W. A. Elmer, E. Kozlowska, J. L. Ugbo, E. S. Doh, and T. E. Whitesides. Does long-term compressive loading on the intervertebral disc cause degeneration? Spine 25:2993-3004, 2000.

    Google Scholar 

  • 26Hutton, W. C., Y. Toribatake, W. A. Elmer, T. M. Ganey, K. Tomita, and T. E. Whitesides. The effect of compressive force applied to the intervertebral disc in vivo. A study of proteoglycans and collagen. Spine 23:2524-2537, 1998.

    Google Scholar 

  • 27Iatridis, J. C., P. L. Mente, I. A. Stokes, D. D. Aronsson, and M. Alini. Compression-induced changes in intervertebral disc properties in a rat tail model. Spine 24:996-1002, 1999.

    Google Scholar 

  • 28Ingber, D. E., L. Dike, L. Hansen, S. Karp, H. Liley, A. Maniotis, H. McNamee, D. Mooney, G. Plopper, and J. Sims. Cellular tensegrity: Exploring how mechanical changes in the cytoskeleton regulate cell growth, migration, and tissue pattern during morphogenesis. Int. Rev. Cytol. 150:173-224, 1994.

    Google Scholar 

  • 29Ishihara, H., K. Warensjo, S. Roberts, and J. P. Urban. Proteoglycan synthesis in the intervertebral disk nucleus:The role of extracellular osmolality. Am. J. Physiol. 272:C1499-C1506, 1997.

    Google Scholar 

  • 30Janmey, P. A. The cytoskeleton and cell signaling: Component localization and mechanical coupling. Physiol. Rev. 78:763-781, 1998.

    Google Scholar 

  • 31Kajstura, J., and K. Reiss. F-actin organization influences the osmotic reactions of animal cells. Folia Histochem. Cytobiol. 27:201-208, 1989.

    Google Scholar 

  • 32Kanzaki, M., and J. E. Pessin. Insulin-stimulated GLUT4 translocation in adipocytes is dependent upon cortical actin remodeling. J. Biol. Chem. 276:42436-42444, 2001.

    Google Scholar 

  • 33Kinosian, H. J., L. A. Selden, J. E. Estes, and L. C. Gershman. Actin filament annealing in the presence of ATP and phalloidin. Biochemistry 32:12353-12357, 1993.

    Google Scholar 

  • 34Lang, F., G. L. Busch, and H. Volkl. The diversity of volume regulatory mechanisms. Cell Physiol. Biochem. 8:1-45, 1998.

    Google Scholar 

  • 35Lange, K. Microvillar Ca++ signaling: A new view of an old problem. J. Cell. Physiol. 180:19-34, 1999.

    Google Scholar 

  • 36Lange, K. Regulation of cell volume via microvillar ion channels. J. Cell. Physiol. 185:21-35, 2000.

    Google Scholar 

  • 37Le Bihan, T., and C. Gicquaud. Stabilization of actin by phalloidin: A differential scanning calorimetric study. Biochem. Biophys. Res. Commun. 181:542-547, 1991.

    Google Scholar 

  • 38Maldonado, B. A., and T. R. Oegema. Initial characterization of the metabolism of intervertebral disc cells encapsulated in microspheres. J. Orthop. Res. 10:677-690, 1992.

    Google Scholar 

  • 39Malko, J. A., W. C. Hutton, and W. A. Fajman. An in vivo magnetic resonance imaging study of changes in the volume (and fluid content) of the lumbar intervertebral discs during a simulated diurnal load cycle. Spine 24:1015-1022, 1999.

    Google Scholar 

  • 40McCarty, N. A., and R. G. O'Neil. Calcium signaling in cell volume regulation. Physiol. Rev. 72:1037-1061, 1992.

    Google Scholar 

  • 41McMillan, D. W., G. Garbutt, and M. A. Adams. Effect of sustained loading on the water content of intervertebral discs: Implications for disc metabolism. Ann. Rheum. Dis. 55:880-887, 1996.

    Google Scholar 

  • 42Mills, J. W., E. M. Schwiebert, and B. A. Stanton. The cytoskeleton and membrane transport. Curr. Opin. Nephrol. Hypertens. 3:529-534, 1994.

    Google Scholar 

  • 43Moran, J., M. Sabanero, I. Meza, and H. Pasantes-Morales. Changes of actin cytoskeleton during swelling and regulatory volume decrease in cultured astrocytes. Am. J. Physiol. 271:C1901-C1907, 1996.

    Google Scholar 

  • 44Oegema, T. R. Biochemistry of the intervertebral disc. Clin. Sports Med. 12:419-439, 1993.

    Google Scholar 

  • 45Ohshima, H., H. Tsuji, N. Hirano, H. Ishihara, Y. Katoh, and H. Yamada. Water diffusion pathway, swelling pressure, and biomechanical properties of the intervertebral disc during compression load. Spine 14:1234-1244, 1989.

    Google Scholar 

  • 46Ohshima, H., J. P. Urban, and D. H. Bergel. Effect of static load on matrix synthesis rates in the intervertebral disc measured in vitro by a new perfusion technique. J. Orthop. Res. 13:22-29, 1995.

    Google Scholar 

  • 47O'Neill, W. C. Physiological significance of volume-regulatory transporters. Am. J. Physiol. 276:C995-C1011, 1999.

    Google Scholar 

  • 48Paajanen, H., I. Lehto, A. Alanen, M. Erkintalo, and M. Komu. Diurnal fluid changes of lumbar discs measured indirectly by magnetic resonance imaging. J. Orthop. Res. 12:509-514, 1994.

    Google Scholar 

  • 49Perlman, D. F., and L. Goldstein. Organic osmolyte channels in cell volume regulation in vertebrates. J. Exp. Zool. 283:725-733, 1999.

    Google Scholar 

  • 50Petrov, A. G., and P. N. Usherwood. Mechanosensitivity of cell membranes. Ion channels, lipid matrix and cytoskeleton. Eur. Biophys. J. 23:1-19, 1994.

    Google Scholar 

  • 51Pritchard, S., G. R. Erickson, and F. Guilak. Hyperosmotically induced volume change and calcium signaling in intervertebral disk cells: The role of the actin cytoskeleton. Biophys. J. 83:2502-2510, 2002.

    Google Scholar 

  • 52Rivero, F., B. Koppel, B. Peracino, S. Bozzaro, F. Siegert, C. J. Weijer, M. Schleicher, R. Albrecht, and A. A. Noegel. The role of the cortical cytoskeleton: F-actin crosslinking proteins protect against osmotic stress, ensure cell size, cell shape and motility, and contribute to phagocytosis and development. J. Cell. Sci. 109(Pt. 11):2679-2691, 1996.

    Google Scholar 

  • 53Schliess, F., R. Sinning, R. Fischer, C. Schmalenbach, and D. Haussinger. Calcium-dependent activation of Erk-1 and Erk-2 after hypo-osmotic astrocyte swelling. Biochem. J. 320(Pt. 1):167-171, 1996.

    Google Scholar 

  • 54Schliwa, M. Action of cytochalasin D on cytoskeletal networks. J. Cell. Biol. 92:79-91, 1982.

    Google Scholar 

  • 55Sheikh, S., W. B. Gratzer, J. C. Pinder, and G. B. Nash. Actin polymerisation regulates integrin-mediated adhesion as well as rigidity of neutrophils. Biochem. Biophys. Res. Commun. 238:910-915, 1997.

    Google Scholar 

  • 56Sullivan, E., E. M. Tucker, and I. L. Dale. Measurement of [Ca2+] using the Fluorometric Imaging Plate Reader (FLIPR). Methods Mol. Biol. 114:125-133, 1999.

    Google Scholar 

  • 57Trickey, W. R., T. Vail, T. Wright, and F. Guilak. The role of the cytoskeleton in the viscoelastic properties of osteoarthritic human chondrocytes. Trans. Orthop. Res. Soc. 25:930, 2000.

    Google Scholar 

  • 58Tseng, Y., E. Fedorov, J. M. McCaffery, S. C. Almo, and D. Wirtz. Micromechanics and ultrastructure of actin filament networks crosslinked by human fascin: A comparison with alpha-actinin. J. Mol. Biol. 310:351-366, 2001.

    Google Scholar 

  • 59Urban, J. P., H. Ishihara, and M. J. Mouat. Differential metabolic responses of the nucleus and outer anulus to changes in extracellular physical factors. Trans. Orthop. Res. Soc. 19:134, 1994.

    Google Scholar 

  • 60Verkman, A. S., A. N. van Hoek, T. Ma, A. Frigeri, W. R. Skach, A. Mitra, B. K. Tamarappoo, and J. Farinas. Water transport across mammalian cell membranes. Am. J. Physiol. 270:C12-C30, 1996.

    Google Scholar 

  • 61Waldegger, S., S. Steuer, T. Risler, A. Heidland, G. Capasso, S. Massry, and F. Lang. Mechanisms and clinical significance of cell volume regulation. Nephrol. Dial. Transplant. 13:867-874, 1998.

    Google Scholar 

  • 62Wang, N. Mechanical interactions among cytoskeletal filaments. Hypertension 32:162-165, 1998.

    Google Scholar 

  • 63Wendel, H., and P. Dancker. Influence of phalloidin on both the nucleation and the elongation phase of actin polymerization. Biochim. Biophys. Acta 915:199-204, 1987.

    Google Scholar 

  • 64Yin, H. L., J. H. Albrecht, and A. Fattoum. Identification of gelsolin, a Ca2+-dependent regulatory protein of actin gel–sol transformation, and its intracellular distribution in a variety of cells and tissues. J. Cell. Biol. 91:901-906, 1981.

    Google Scholar 

  • 65Yin, H. L., and T. P. Stossel. Control of cytoplasmic actin gel–sol transformation by gelsolin, a calcium-dependent regulatory protein. Nature 281:583-586, 1979.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Pritchard, S., Guilak, F. The Role of F-Actin in Hypo-Osmotically Induced Cell Volume Change and Calcium Signaling in Anulus Fibrosus Cells. Annals of Biomedical Engineering 32, 103–111 (2004). https://doi.org/10.1023/B:ABME.0000007795.69001.35

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/B:ABME.0000007795.69001.35

Navigation