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. 1998 Sep 1;334(Pt 2):431–435. doi: 10.1042/bj3340431

Luminal Ca2+ regulates passive Ca2+ efflux from the intracellular stores of hepatocytes.

M D Beecroft 1, C W Taylor 1
PMCID: PMC1219706  PMID: 9716502

Abstract

Ca2+ uptake into the intracellular stores of permeabilized hepatocytes was entirely dependent on ATP and substantially inhibited by either ionomycin or thapsigargin, although both were required for complete inhibition. Unidirectional efflux of 45Ca2+ after removal of ATP from cells loaded to steady state (1.60+/-0.12 nmol/10(6) cells) was monoexponential and occurred with a half-time of 103+/-10 s. However, the 45Ca2+ content of the stores did not return to their pre-ATP level, but reached a plateau at 0.12+/-0.04 nmol/10(6) cells. A similar amount of Ca2+ was trapped within the stores when Ca2+ uptake was prevented by thapsigargin and chelation of Ca2+; at all temperatures between 2 degreesC and 37 degreesC; and after stores had first been loaded with unlabelled Ca2+. Simultaneous addition of inositol 1,4,5-trisphosphate (InsP3) and inhibition of Ca2+ uptake reduced the amount of trapped Ca2+ to a level consistent with InsP3 rapidly and more completely emptying a fraction of the stores that could be only partially emptied by the passive leak. After dilution of the specific activity of the 45Ca2+ under conditions that maintained the steady-state activities of the pumps and leaks, the stores rapidly lost their entire 45Ca2+ content. We conclude that the channel responsible for mediating the leak of Ca2+ abruptly closes when the luminal [Ca2+] of the intracellular stores falls below a critical threshold corresponding to about 7% of their steady-state loading. Whereas InsP3 is capable of completely emptying a fraction of the stores, regulation of the passive leak by luminal [Ca2+] is likely to prevent it from completely emptying them; such regulation may ensure that the many other functions of Ca2+ within the endoplasmic reticulum are not compromised.

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Selected References

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  1. Beecroft M. D., Taylor C. W. Incremental Ca2+ mobilization by inositol trisphosphate receptors is unlikely to be mediated by their desensitization or regulation by luminal or cytosolic Ca2+. Biochem J. 1997 Aug 15;326(Pt 1):215–220. doi: 10.1042/bj3260215. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Berridge M. J. Capacitative calcium entry. Biochem J. 1995 Nov 15;312(Pt 1):1–11. doi: 10.1042/bj3120001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Berridge M. J. Elementary and global aspects of calcium signalling. J Physiol. 1997 Mar 1;499(Pt 2):291–306. doi: 10.1113/jphysiol.1997.sp021927. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Berridge M. J. Inositol trisphosphate and calcium signalling. Nature. 1993 Jan 28;361(6410):315–325. doi: 10.1038/361315a0. [DOI] [PubMed] [Google Scholar]
  5. Du G. G., Ashley C. C., Lea T. J. Ca2+ effluxes from the sarcoplasmic reticulum vesicles of frog muscle: effects of cyclopiazonic acid and thapsigargin. Cell Calcium. 1996 Oct;20(4):355–359. doi: 10.1016/s0143-4160(96)90041-x. [DOI] [PubMed] [Google Scholar]
  6. Fasolato C., Zottini M., Clementi E., Zacchetti D., Meldolesi J., Pozzan T. Intracellular Ca2+ pools in PC12 cells. Three intracellular pools are distinguished by their turnover and mechanisms of Ca2+ accumulation, storage, and release. J Biol Chem. 1991 Oct 25;266(30):20159–20167. [PubMed] [Google Scholar]
  7. Favre C. J., Schrenzel J., Jacquet J., Lew D. P., Krause K. H. Highly supralinear feedback inhibition of Ca2+ uptake by the Ca2+ load of intracellular stores. J Biol Chem. 1996 Jun 21;271(25):14925–14930. doi: 10.1074/jbc.271.25.14925. [DOI] [PubMed] [Google Scholar]
  8. Gaut J. R., Hendershot L. M. The modification and assembly of proteins in the endoplasmic reticulum. Curr Opin Cell Biol. 1993 Aug;5(4):589–595. doi: 10.1016/0955-0674(93)90127-c. [DOI] [PubMed] [Google Scholar]
  9. Golovina V. A., Blaustein M. P. Spatially and functionally distinct Ca2+ stores in sarcoplasmic and endoplasmic reticulum. Science. 1997 Mar 14;275(5306):1643–1648. doi: 10.1126/science.275.5306.1643. [DOI] [PubMed] [Google Scholar]
  10. Hofer A. M., Curci S., Machen T. E., Schulz I. ATP regulates calcium leak from agonist-sensitive internal calcium stores. FASEB J. 1996 Feb;10(2):302–308. doi: 10.1096/fasebj.10.2.8641563. [DOI] [PubMed] [Google Scholar]
  11. Hofer A. M., Fasolato C., Pozzan T. Capacitative Ca2+ entry is closely linked to the filling state of internal Ca2+ stores: a study using simultaneous measurements of ICRAC and intraluminal [Ca2+]. J Cell Biol. 1998 Jan 26;140(2):325–334. doi: 10.1083/jcb.140.2.325. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Marshall I. C., Taylor C. W. Biphasic effects of cytosolic Ca2+ on Ins(1,4,5)P3-stimulated Ca2+ mobilization in hepatocytes. J Biol Chem. 1993 Jun 25;268(18):13214–13220. [PubMed] [Google Scholar]
  13. Meldolesi J., Pozzan T. The endoplasmic reticulum Ca2+ store: a view from the lumen. Trends Biochem Sci. 1998 Jan;23(1):10–14. doi: 10.1016/s0968-0004(97)01143-2. [DOI] [PubMed] [Google Scholar]
  14. Mikoshiba K. The InsP3 receptor and intracellular Ca2+ signaling. Curr Opin Neurobiol. 1997 Jun;7(3):339–345. doi: 10.1016/s0959-4388(97)80061-x. [DOI] [PubMed] [Google Scholar]
  15. Missiaen L., De Smedt H., Parys J. B., Raeymaekers L., Droogmans G., Van Den Bosch L., Casteels R. Kinetics of the non-specific calcium leak from non-mitochondrial calcium stores in permeabilized A7r5 cells. Biochem J. 1996 Aug 1;317(Pt 3):849–853. doi: 10.1042/bj3170849. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Nunn D. L., Taylor C. W. Luminal Ca2+ increases the sensitivity of Ca2+ stores to inositol 1,4,5-trisphosphate. Mol Pharmacol. 1992 Jan;41(1):115–119. [PubMed] [Google Scholar]
  17. Oldershaw K. A., Nunn D. L., Taylor C. W. Quantal Ca2+ mobilization stimulated by inositol 1,4,5-trisphosphate in permeabilized hepatocytes. Biochem J. 1991 Sep 15;278(Pt 3):705–708. doi: 10.1042/bj2780705. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Parekh A. B., Fleig A., Penner R. The store-operated calcium current I(CRAC): nonlinear activation by InsP3 and dissociation from calcium release. Cell. 1997 Jun 13;89(6):973–980. doi: 10.1016/s0092-8674(00)80282-2. [DOI] [PubMed] [Google Scholar]
  19. Pozzan T., Rizzuto R., Volpe P., Meldolesi J. Molecular and cellular physiology of intracellular calcium stores. Physiol Rev. 1994 Jul;74(3):595–636. doi: 10.1152/physrev.1994.74.3.595. [DOI] [PubMed] [Google Scholar]
  20. Putney J. W., Jr Capacitative calcium entry revisited. Cell Calcium. 1990 Nov-Dec;11(10):611–624. doi: 10.1016/0143-4160(90)90016-n. [DOI] [PubMed] [Google Scholar]
  21. Renard-Rooney D. C., Hajnóczky G., Seitz M. B., Schneider T. G., Thomas A. P. Imaging of inositol 1,4,5-trisphosphate-induced Ca2+ fluxes in single permeabilized hepatocytes. Demonstration of both quantal and nonquantal patterns of Ca2+ release. J Biol Chem. 1993 Nov 5;268(31):23601–23610. [PubMed] [Google Scholar]
  22. Sagara Y., Inesi G. Inhibition of the sarcoplasmic reticulum Ca2+ transport ATPase by thapsigargin at subnanomolar concentrations. J Biol Chem. 1991 Jul 25;266(21):13503–13506. [PubMed] [Google Scholar]
  23. Short A. D., Bian J., Ghosh T. K., Waldron R. T., Rybak S. L., Gill D. L. Intracellular Ca2+ pool content is linked to control of cell growth. Proc Natl Acad Sci U S A. 1993 Jun 1;90(11):4986–4990. doi: 10.1073/pnas.90.11.4986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Smith P. M., Gallacher D. V. 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. 1994 Apr 1;299(Pt 1):37–40. doi: 10.1042/bj2990037. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Steenbergen J. M., Fay F. S. The quantal nature of calcium release to caffeine in single smooth muscle cells results from activation of the sarcoplasmic reticulum Ca(2+)-ATPase. J Biol Chem. 1996 Jan 26;271(4):1821–1824. doi: 10.1074/jbc.271.4.1821. [DOI] [PubMed] [Google Scholar]
  26. Taylor C. W., Potter B. V. The size of inositol 1,4,5-trisphosphate-sensitive Ca2+ stores depends on inositol 1,4,5-trisphosphate concentration. Biochem J. 1990 Feb 15;266(1):189–194. doi: 10.1042/bj2660189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. van de Put F. H., De Pont J. J., Willems P. H. Heterogeneity between intracellular Ca2+ stores as the underlying principle of quantal Ca2+ release by inositol 1,4,5-trisphosphate in permeabilized pancreatic acinar cells. J Biol Chem. 1994 Apr 29;269(17):12438–12443. [PubMed] [Google Scholar]

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