The Journal of Membrane Biology

, Volume 211, Issue 2, pp 127–137

Calcium-Sensing Receptor Stimulation Induces Nonselective Cation Channel Activation in Breast Cancer Cells

  • Yassine El Hiani
  • Ahmed Ahidouch
  • Morad Roudbaraki
  • Stéphanie Guenin
  • Gérard Brûlé
  • Halima Ouadid-Ahidouch


The calcium-sensing receptor (CaR) is expressed in epithelial ducts of both normal human breast and breast cancer tissue, as well as in the MCF-7 cell line as assessed by immunohistochemistry and Western blot analysis. However, to date, there are no data regarding the transduction pathways of CaR in breast cancer cells. In this study, we show that a CaR agonist, spermine, and increased extracellular Ca2+ ([Ca2+]o) sequentially activate two inward currents at –80 mV. The first was highly permeable to Ca2+ and inhibited by 2-aminophenyl borate (2-APB). In contrast, the second was more sensitive to Na+ and Li+ than to Ca2+ and insensitive to 2-APB. Furthermore, intracellular dialysis with high Mg2+, flufenamic acid or amiloride perfusion was without any effect on the second current. Both currents were inhibited by La3+. Calcium imaging recordings showed that both [Ca2+]o and spermine induced an increase in intracellular calcium ([Ca2+]i) and that removal of extracellular Ca2+ or perfusion of 2-APB caused a decline in [Ca2+]i. It is well known that stimulation of CaR by an increase in [Ca2+]o or with spermine is associated with activation of phospholipase C (PLC). Inhibition of PLC reduced the [Ca2+]o-stimulated [Ca2+]i increase. Lastly, reverse-transcriptase polymerase chain reaction showed that MCF-7 cells expressed canonical transient receptor potential (TRPCs) channels. Our results suggest that, in MCF-7 cells, CaR is functionally coupled to Ca2+-permeable cationic TRPCs, for which TRPC1 and TRPC6 are the most likely candidates for the highly selective Ca2+ current. Moreover, the pharmacology of the second Na+ current excludes the involvement of the more selective Na+ transient receptor potential melastatin (TRPM4 and TRPM5) and the classical epithelial Na+ channels.


Calcium-sensing receptor Cationic current Transient receptor potential channel Breast cancer cell 


  1. Beech D.J. 2005. TRPC1: Store-operated channel and more. Pfluegers Arch. 451:53–60CrossRefGoogle Scholar
  2. Boring C.C., Squire T.S., Tong T., Montgomery S. 1994. Cancer statistics. CA Cancer J. Clin. 44:7–26PubMedGoogle Scholar
  3. Broad L.M., Braun F.J., Lievremont J.P., Bird G.S., Kurosaki T., Putney J.W., Jr. 2001. Role of the phospholipase C-inositol 1,4,5-trisphosphate pathway in calcium release-activated calcium current and capacitative calcium entry. J. Biol. Chem. 276:15945–15952PubMedCrossRefGoogle Scholar
  4. Brown E.M., Enyedi P., LeBoff M., Rotberg J., Preston J., Chen C. 1987. High extracellular Ca2+ and Mg2+ stimulate accumulation of inositol phosphates in bovine parathyroid cells. FEBS Lett. 218:113–138PubMedCrossRefGoogle Scholar
  5. Brown E.M., Gamba G., Riccardi D., Lombardi M., Butters R., Kifor O., Sun A., Hediger M.A., Lytton J., Hebert S.C. 1993. Cloning and characterization of an extracellular Ca2+-sensing receptor from bovine parathyroid. Nature 366:575–580PubMedCrossRefGoogle Scholar
  6. Brown E.M., MacLeod R.J. 2001. Extracellular calcium sensing and extracellular calcium signalling. Physiol. Rev. 81:239–297PubMedGoogle Scholar
  7. Buchs N., Manen D., Bonjour J.P., Rizzoli R. 2000. Calcium stimulates parathyroid hormone-related protein production in Leydig tumor cells through a putative cation-sensing mechanism. Eur. J. Endocrinol. 142:500–505PubMedCrossRefGoogle Scholar
  8. Chattopadyay N., Evliyaoglu C., Heese O., Carroll R., Sanders J., Black P., Brown E.M. 2000. Regulation of secretion of PTHrP by Ca2+-sensing receptor in human astrocytes, astrocytomas, and meningiomas. Am. J. Physiol. 279:C691-C699Google Scholar
  9. Cheng I., Klingensmith M.E., Chattopadhyay N., Kifor O., Butters R.R., Soybel D.I., Brown E.M. 1998. Identification and localization of the extracellular calcium-sensing receptor in human breast. J. Clin. Endocrinol. Metab. 83:703–707PubMedCrossRefGoogle Scholar
  10. Falzon M., Du P. 2000. Enhanced growth of MCF-7 breast cancer cells overexpressing parathyroid hormone-related peptide. Endocrinology 141:1882–1892PubMedCrossRefGoogle Scholar
  11. Fatherazi S., Belton C.M., Cai S., Zarif S., Goodwin P.C., Lamont R.J., Izutsu K.T. 2004. Calcium receptor message, expression and function decrease in differentiating keratinocytes. Pfluegers Arch. 448:93–104CrossRefGoogle Scholar
  12. Guise T.A., Yin T.A., Taylor S.D., Kumagai Y., Dallas M., Boyce B.F., Yoneda T., Mundy G.R. 1996. Evidence for a causal role of parathyroid hormone-related protein in the pathogenesis of human breast cancer-mediated osteolysis. J. Clin. Invest. 98:1544–1549PubMedCrossRefGoogle Scholar
  13. Huang C., Handlogten M.E., Miller R.T. 2002. Parallel activation of phosphotidylinositol 4-kinase and phospholipase C by the extracellular calcium-sensing receptor. J. Biol. Chem. 277:20293–20300PubMedCrossRefGoogle Scholar
  14. Jiang Y.F., Zhang Z., Kifor O., Lane C.R., Suinn S.J., Bai M. 2002. Protein kinase C (PKC) phosphorylation of the Ca2+-sensing receptor (CaR) modulates functional interaction of G proteins with the CaR cytoplasmic tail. J. Biol. Chem. 277:50543–50549PubMedCrossRefGoogle Scholar
  15. Journe F., Dumon J.C., Kheddoumi N., Fox J., Laios I., Leclercq G., Body J.J. 2004. Extracellular calcium downregulates estrogen receptor alpha and increases its transcriptional activity through calcium-sensing receptor in breast cancer cells. Bone 35:479–488PubMedCrossRefGoogle Scholar
  16. Kraft R., Harteneck C. 2005. The mammalian melastatin-related transient receptor potential cation channels: An overview. Pfluegers Arch. 451:204–211CrossRefGoogle Scholar
  17. Li S., Huang S., Peng S.B. 2005. Overexpression of G protein-coupled receptors in cancer cells: Involvement in tumor progression. Int. J. Oncol. 27:1329–1338PubMedGoogle Scholar
  18. Liapis H., Crouch E.C., Grosso L.E., Kitazawa S., Wick M.R. 1993. Expression of parathyroidlike protein in normal, proliferative, and neoplastic human breast tissues. Am. J. Pathol. 174:1169–1178Google Scholar
  19. Mundy G.R. 1997. Mechanisms of bone metastasis. Cancer 80:1546–1556PubMedCrossRefGoogle Scholar
  20. Nemeth E.F., Scarpa A. 1987. Rapid mobilization of cellular Ca2+ in bovine parathyroid cells evoked by extracellular divalent cations. Evidence for a cell surface calcium receptor. J. Biol. Chem. 262:5188–5196PubMedGoogle Scholar
  21. Ouadid-Ahidouch H., Roudbaraki M., Delcourt P., Ahidouch A., Joury N., Prevarskaya N. 2004. Functional and molecular identification of intermediate-conductance Ca2+-activated K+ channels in breast cancer cells: Association with cell cycle progression. Am. J. Physiol. 287:C125-C134CrossRefGoogle Scholar
  22. Parekh A.B., Putney J.W., Jr. 2005. Store-operated calcium channels. Physiol. Rev. 85:757–810PubMedCrossRefGoogle Scholar
  23. Parkash J., Chaudhry M.A., Rhoten W.B. 2004. Calbindin-D28k and calcium sensing receptor cooperate in MCF-7 human breast cancer cells. Int. J. Oncol. 24:1111–1119PubMedGoogle Scholar
  24. Pedersen S.F., Owsianik G., Nilius B. 2005. TRP channels: An overview. Cell Calcium 38:233–252PubMedCrossRefGoogle Scholar
  25. Rodland K.D. 2004. The role of the calcium-sensing receptor in cancer. Cell Calcium 35:291–295PubMedCrossRefGoogle Scholar
  26. Sanders J.L., Chattopadhyay N., Kifor O., Yamaguchi T., Butter R.R., Brown E.M. 2000. Extracellular calcium-sensing receptor expression and its potential role in regulating parathyroid hormone-related peptide secretion in human breast cancer cell lines. Endocrinology 141:4357–4364PubMedCrossRefGoogle Scholar
  27. Sanders J.L., Chattopadhyay N., Kifor O., Yamaguchi T., Butter R.R., Brown E.M. 2001. Ca2+-sensing receptor expression and PTHrP secretion in PC-3 human prostate cancer cells. Am. J. Physiol. 281:E1267-E1274Google Scholar
  28. Shen X., Qian L., Falzon M. 2004. PTH-related protein enhances MCF-7 breast cancer cell adhesion, migration, and invasion via an intracrine pathway. Exp. Cell. Res. 294:420–433PubMedCrossRefGoogle Scholar
  29. Silver I.A., Murrills R.J., Etherington D.J. 1988. Microelectrode studies on the acid microenvironment beneath adherent macrophages and osteoclast. Exp. Cell. Res. 175:266–276PubMedCrossRefGoogle Scholar
  30. Ullrich N.D., Voets T., Prenen J., Vennekens R., Talavera K., Droogmans G., Nilius B. 2005. Comparison of functional properties of the Ca2+-activated cation channels TRPM4 and TRPM5 from mice. Cell Calcium 37:267–278PubMedCrossRefGoogle Scholar
  31. VanHouten J.N. 2005. Calcium sensing by the mammary gland. Neoplasia 10:129–139Google Scholar
  32. VanHouten J.N., Danna P., McGeoch G., Brown E.M., Krapcho K., Neville M., Wysolmerski J.J. 2004. The calcium-sensing receptor regulates mammary gland parathyroid hormone-related protein production and calcium transport. J. Clin. Invest. 113:598–608PubMedCrossRefGoogle Scholar
  33. Yamaguchi T., Ye C., Chattopadhyay N., Sanders J.L., Vassilev P.M., Brown E.M. 2000. Enhanced expression of extracellular calcium sensing receptor in monocyte-differentiated versus undifferentiated HL-60 cells: Potential role in regulation of a nonselective cation channel. Calcif. Tissue Int. 66:375–382PubMedCrossRefGoogle Scholar
  34. Ye C.P., Kanazirskia M., Quinn S., Brown E.M., Vassilev P.M. 1996a. Modulation by polycationic Ca2+-sensing receptor agonists of nonselective cation channels in rat hippocampal neurons. Biochem. Biophys. Res. Commun. 224:271–280CrossRefGoogle Scholar
  35. Ye C.P., Rogers K., Bai M., Quinn S., Seidman C.E., Seidman J.G., Brown E.M., Vassilev P.M. 1996b. Agonists of the Ca2+-sensing receptor (CaR) activate nonselective cation channels in HEK293 cells stably transfected with the human CaR. Biochem. Biophys. Res. Commun. 226:272–279CrossRefGoogle Scholar
  36. Ye C.P., Ho-Pao C.L., Kanazirskia M., Quinn S., Rogers K., Seidman C.E., Seidman J.G., Brown E.M., Vassilev P.M. 1997. Amyloid-beta proteins activate Ca2+-permeable channels through calcium-sensing receptors. J. Neurosci. Res. 47:547–554PubMedCrossRefGoogle Scholar
  37. Zitt C., Halaszovich C.R., Luckhoff A. 2002. The TRP family of cation channels: Probing and advancing the concepts on receptor-activated calcium entry. Prog. Neurobiol. 66:243–264PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, Inc. 2006

Authors and Affiliations

  • Yassine El Hiani
    • 1
  • Ahmed Ahidouch
    • 1
    • 2
  • Morad Roudbaraki
    • 3
  • Stéphanie Guenin
    • 4
  • Gérard Brûlé
    • 1
  • Halima Ouadid-Ahidouch
    • 1
  1. 1.Laboratoire de Physiologie Cellulaire et Moléculaire, EA 2086, Faculté des SciencesUniversité de Picardie Jules VerneFrance
  2. 2.Laboratoire de Physiologie Animale, Faculté des SciencesUniversité Ibn-ZohrAgadirMorocco
  3. 3.INSERM EMI 0228Université de LilleVilleneuve d’AscqFrance
  4. 4.Centre de Ressources Régionales en Biologie MoléculaireUniversité de Picardie Jules VerneCedex 1France

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