, Volume 37, Issue 4, pp 296–302 | Cite as

Mechanisms Underlying Leakage of Calcium from the Endoplasmic Reticulum of Acinar Cells of the Submandibular Salivary Gland

  • O. V. Kopach
  • I. A. Kruglikov
  • P. G. Kostyuk
  • N. V. Voitenko
  • N. V. Fedirko


In the resting state, the Ca2+ concentration in agonist-sensitive intracellular stores reflects the balance between active uptake of Ca2+, which is mediated by Ca2+-ATPase (SERCA), and passive leakage of Ca2+. The mechanisms underlying such a leakage in cells of the submaxillary salivary gland were not studied. In our experiments, we examined possible pathways of passive leakage of Ca2+ from the endoplasmic reticulum (ER) of acinar cells obtained from the rat submaxillary salivary gland; direct measurements of the concentration of Ca2+ in the ER ([Ca2+]ER) using a low-affinity calcium-sensitive dye, mag-fura 2/AM, were performed. The cellular membrane was permeabilized with the help of β-escin (40 μg/ml); the Ca2+ concentration in the cytoplasm ([Ca2+] i ) was clamped at its level typical of the resting state (∼100 nM) using an EGTA/Ca2+ buffer. Incubation of permeabilized acinar cells in a calcium-free intracellular milieu, as well as application of thapsigargin, resulted in complete inhibition of the uptake of Ca2+ with the involvement of SERCA. This effect was observed 1 min after the beginning of superfusion of the cells with the corresponding solutions and was accompanied by the leakage of Ca2+ from the ER; this is confirmed by a gradual drop in the [Ca2+]ER. Such a leakage of Ca2+ remained unchanged in the presence of thapsigargin, heparin, and ruthenium red; therefore, it is not mediated by SERCA, inositol 1,4,5-trisphosphate-sensitive receptors (InsP3R), or ryanodine receptors (RyRs). At the same time, an antibiotic, puromycin (0.1 to 1.0 mM), which disconnects polypeptides from the ER-ribosome translocon complex, caused intensification of passive leakage of Ca2+ from the ER. This effect did not depend on the functioning of SERCA, InsP3R, or RyR. Therefore, passive leakage of Ca2+ from the ER in acinar cells of the submaxillary salivary gland is realized through pores of the translocon complex of the ER membrane.


intracellular calcium concentration endoplasmic reticulum acinar cells Ca2+/Mg2+-ATPases InsP3 receptor ryanodine receptor 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    W. Paschen and A. Frandsen, “Endoplasmic reticulum dysfunction a common denominator for cell injury in acute and degenerative diseases of the brain?” J. Neurochem., 79, No. 4, 719–725 (2001).CrossRefPubMedGoogle Scholar
  2. 2.
    M. J. Berridge, “The endoplasmic reticulum: a multifunctional signaling organelle,” Cell Calcium, 32, Nos. 5/6, 235–249 (2002).PubMedGoogle Scholar
  3. 3.
    I. S. Ambudkar, “Regulation of calcium in salivary gland secretion,” Cr. Rev. Oral Biol. Med., 11, No. 1, 4–25 (2000).Google Scholar
  4. 4.
    O. H. Petersen, “Stimulus-secretion coupling: cytoplasmic calcium signals and the control of ion channels in exocrine acinar cells,” J. Physiol., 448, 1–51 (1992).PubMedGoogle Scholar
  5. 5.
    P. L. Pedersen and E. Carafoli, “Ion motive ATPases. Ubiquity, properties, and significance for cell function,” Trends Biochem. Sci., 12, 146–150 (1987).Google Scholar
  6. 6.
    A. M. Hofer, S. Curci, T. E. Machen, and I. Schulz, “ATP regulates calcium leak from agonist-sensitive internal calcium stores,” FASEB J., 2, 302–308 (1996).Google Scholar
  7. 7.
    M. Endo and M. Iino, “Measurement of Ca2+ release in skinned fibers from skeletal muscle,” Meth. Enzymol., 157, 12–26 (1988).PubMedGoogle Scholar
  8. 8.
    R. Sitsapesan and A. J. Williams, “Mechanisms of caffeine activation of single calcium-release channels of sheep cardiac sarcoplasmic reticulum,” J. Physiol., 423, 425–439 (1990).PubMedGoogle Scholar
  9. 9.
    P. M. Smith and D. V. Gallacher, “Thapsigargin-induced Ca2+ mobilization in acutely isolated mouse lacrimal acinar cells is dependent on a basal level of Ins(1,4,5)P3 and is inhibited by heparin,” Biochem. J., 299, No. 1, 37–40 (1994).PubMedGoogle Scholar
  10. 10.
    C. J. Favre, D. P. Lew, and K. H. Krause, “Rapid heparin-sensitive Ca2+ release following Ca2+-ATPase inhibition in intact HL-60 granulocytes. Evidence for Ins(1,4,5)P3-dependent Ca2+ cycling across the membrane of Ca2+ stores,” Biochem. J., 302, No. 1, 155–162 (1994).PubMedGoogle Scholar
  11. 11.
    R. B. Lomax, C. Camello, F. Van Coppenolle, et al., “Basal and physiological Ca2+ leak from the endoplasmic reticulum of pancreatic acinar cells. Second messenger-activated channels and translocons,” J. Biol. Chem., 277, No. 29, 26479–26485 (2002).CrossRefPubMedGoogle Scholar
  12. 12.
    S. M. Simon and G. Blobel, “A protein-conducting channel in the endoplasmic reticulum,” Cell, 65, No. 3, 371–380 (1991).CrossRefPubMedGoogle Scholar
  13. 13.
    D. Heritage and W. F. Wonderlin, “Translocon pores in the endoplasmic reticulum are permeable to a neutral, polar molecule,” J. Biol. Chem., 276, No. 25, 22655–22662 (2001).CrossRefPubMedGoogle Scholar
  14. 14.
    J. A. Vats, N. V. Fedirko, M. Y. Klevets, et al., “Role of SH groups in the functioning of Ca2+-transporting ATPases regulating Ca2+ homeostasis and exocytosis,” Neurophysiology, 34, No. 1, 5–12 (2002).CrossRefGoogle Scholar
  15. 15.
    S. Kobayashi, T. Kitazawa, A. V. Somlyo, and A. P. Somlyo, “Cytosolic heparin inhibits muscarinic and alpha-adrenergic Ca2+ release in smooth muscle. Physiological role of inositol 1,4,5-trisphosphate in pharmacomechanical coupling,” J. Biol. Chem., 264, No. 30, 17997–8004 (1989).PubMedGoogle Scholar
  16. 16.
    A. M. Hofer and T. E. Machen, “Technique for in situ measurement of calcium in intracellular inositol 1,4,5-trisphosphate-sensitive stores using the fluorescent indicator mag-fura-2,” Proc. Natl. Acad. Sci. USA, 90, No. 7, 2598–2602 (1993).PubMedGoogle Scholar
  17. 17.
    O. V. Kopach, I. A. Kruglikov, N. V. Voitenko, et al., “Permeable cells of salivary glands as a model for examination of calcium-transport systems of the membrane of the endoplasmic reticulum,” Fiziol. Zh., 49, No. 5, 31–42 (2003).PubMedGoogle Scholar
  18. 18.
    M. G. Lee, X. Xu, W. Zeng, et al., “Polarized expression of Ca2+ pumps in pancreatic and salivary gland cells. Role in initiation and propagation of [Ca2+]i waves,” J. Biol. Chem., 272, No. 25, 15771–15776 (1997).PubMedGoogle Scholar
  19. 19.
    P. Palade, “Drug-induced Ca2+ release from isolated sarcoplasmic reticulum. I. Use of pyrophosphate to study caffeine-induced Ca2+ release,” J. Biol. Chem., 262, 6135–6141 (1987).PubMedGoogle Scholar
  20. 20.
    D. R. Wetmore and K. D. Hardman, “Roles of the propeptide and metal ions in the folding and stability of the catalytic domain of stromelysin (matrix metalloproteinase 3),” Biochemistry, 35, 6549–6558 (1996).CrossRefPubMedGoogle Scholar
  21. 21.
    C. R. Prostko, J. N. Dholakia, M. A. Bromstrom, and C. O. Bromstrom, “Activation of the double-stranded RNA-regulated protein kinase by depletion of endoplasmic reticular calcium stores,” J. Biol. Chem., 270, 6211–6215 (1995).CrossRefPubMedGoogle Scholar
  22. 22.
    J. Jeffery, J. M. Kendall, and A. K. Campbell, “Apoaequorin monitors degradation of endoplasmic reticulum (ER) proteins initiated by loss of ER Ca2+,” Biochem. Biophys. Res. Commun., 268, 711–715 (2000).CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, Inc. 2005

Authors and Affiliations

  1. 1.Franko National UniversityL'vovUkraine
  2. 2.Bogomolets Institute of PhysiologyNational Academy of Sciences of UkraineKyivUkraine

Personalised recommendations