Summary
We have measured Ca2+ uptake and Ca2+ release in isolated permeabilized pancreatic acinar cells and in isolated membrane vesicles of endoplasmic reticulum prepared from these cells. Ca2+ uptake into cells was monitored with a Ca2+ electrode, whereas Ca2+ uptake into membrane vesicles was measured with45Ca2+. Using inhibitors of known action, such as the H+ ATPase inhibitors NBD-Cl and NEM, the Ca2+ ATPase inhibitor vanadate as well as the second messenger inositol 1,4,5-trisphosphate (IP3) and its analog inositol 1,4,5-trisphosphorothioate (IPS3), we could functionally differentiate two non-mitochondrial Ca2+ pools. Ca2+ uptake into the IP3-sensitive Ca2+ pool (IsCaP) occurs by a MgATP-dependent Ca2+ uptake mechanism that exchanges Ca2+ for H+ ions. In the absence of ATP Ca2+ uptake can occur to some extent at the expense of an H+ gradient that is established by a vacuolar-type MgATP-dependent H+ pump present in the same organelle. The other Ca2+ pool takes up Ca2+ by a vanadate-sensitive Ca2+ ATPase and is insensitive to IP3 (IisCaP). The IsCaP is filled at “higher” Ca2+ concentrations (∼10−6 mol/liter) which may occur during stimulation. The low steady-state [Ca2+] of ∼10−7 mol/liter is adjusted by the IisCaP.
It is speculated that both Ca2+ pools can communicate with each other, the possible mechanism of which, however, is at present unknown.
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References
Al-Awqati, Q. 1986. Proton-translocating ATPases.Annu. Rev. Cell Biol. 2:179–199
Amsterdam, A., Jamieson, J.D. 1972. Structural and functional characterization of isolated pancreatic exocrine cells.Proc. Natl. Acad. Sci. USA 69:3028–3032
Bais, R. 1975. A rapid and sensitive radiometric assay for adenosine triphosphatase activity using cerenkov radiation.Anal. Biochem. 63:271–273
Bayerdörffer, E., Streb, H., Eckhardt, L., Haase, W., Schulz, I. 1984. Characterization of calcium uptake into rough endoplasmic reticulum of rat pancreas.J. Membrane Biol. 81:69–82
Berridge, M.J., Irvine, R.F. 1984. Inositol trisphosphate, a novel second messenger in cellular signal transduction.Nature (London) 312:315–321
Bradford, M.M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.Anal. Biochem. 72:248–254
Breemen, C., van Cauvin, C., Johns, A., Leijten, P., Yamamoto, H. 1986. Ca2+ regulation of vascular smooth muscle.Fed. Proc. 45:2746–2751
Burgess, G.M., Irvine, R.F., Berridge, M.J., McKinney, J.S., Putney, J.W., Jr. 1984. Actions of inositol phosphates on Ca2+ pools in guinea-pig hepatocytes.Biochem. J. 224:41–746
Carafoli, E. 1987. Intracellular calcium homeostasis.Annu. Rev. Biochem. 56:395–433
Cooke, A.M., Gigg, R., Potter, B.V.L. 1987. Myo-inositol 1,4,5-tris-phosphorothioate.: A novel analogue of a biological second messenger.J. Chem. Soc. Commun. 698:1525–1526
Cooke, A.M., Nahorski, S.R., Potter, B.V.L. 1989.FEBS Lett. (in press)
Haynes, D.H. 1983. Mechanism of Ca2+ transport by Ca2+−Mg2+-ATPase pump: Analysis of major states and pathways.Am. J. Physiol. 244:G3-G12
Imamura, K., Schulz, I. 1985. Phosphorylated intermediate of (Ca2++K+)-stimulated Mg2+-dependent transport ATPase in endoplasmic reticulum from rat pancreatic acinar cells.J. Biol. Chem. 260:11339–11347
Irvine, R.F., Moor, R.M., Pollock, W.K., Smith, P.M., Wreggett, K.A. 1988. Inostiol phosphates: Proliferation, metabolism and function.Phil. Trans. R. Soc. (London) B 320:281–298
Kemmer, T.P., Bayerdörffer, E., Will, H., Schulz, I. 1987. Anion dependence of Ca2+ transport and (Ca2++K+)-stimulated Mg2+-dependent transport ATPase in rat pancreatic endoplasmic reticulum.J. Biol. Chem. 262:13758–13764
Mellman, I., Fuchs, R., Helenius, A. 1986. Acidification of The endocytotic and exocytotic pathways.Annu. Rev. Biochem. 55:663–700
Mullaney, J.M., Chueh, S.-H., Ghosh, T.K., Gill, D.L. 1987. Intracellular calcium uptake activated by GTP.J. Biol. Chem. 262:13865–13872
Mullaney, J.M., Yu, M., Ghosh, T.K., Gill, D.L. 1988. Calcium entry into the inositol 1,4,5-trisphosphate-releasable calcium pool is mediated by a GTP-regulatory mechanism.Proc. Natl. Acad. Sci. USA 85:2499–2503
Ochs, D.L., Korenbrot, J.I., Williams, J.A. 1983. Intracellular free calcium concentrations in isolated pancreatic acini: Effects of secretagogues.Biochem. Biophys. Res. Commun. 117:122–128
Rudnick, G. 1986. ATP-driven H+ pumping into intracellular organelles.Annu. Rev. Physiol. 48:403–413
Schäfer, R., Christian, A.-L., Schulz, I. 1988. Photoaffinity labeling with GTP-γ-azidoanilide of a cholera toxin-sensitive 40 kDa protein from pancretic acinar cells.Biochem. Biophys. Res. Commun. 155:1051–1059
Schnefel, S., Banfić, H., Eckhardt, L., Schultz, G., Schulz, I. 1988. Acetylcholine and cholecystokinin receptors functionally couple by different G-proteins to phospholipase C in pancreatic acinar cells.FEBS Lett. 230:125–130
Schumaker, K.S., Sze, H. 1985. A Ca2+/H+ antiport system driven by the proton electrochemical gradient of a tonoplast H+-ATPase from oat roots.Plant Physiol. 1111–1117
Schumaker, K.S., Sze, H. 1987. Inositol 1,4,5-trisphosphate releases Ca2+ from vacuolar membrane vesicles of oat roots.J. Biol. Chem. 262:3944–3946
Streb, H., Bayerdörffer, E., Haase, W., Irvine, R.F., Schulz, I. 1984. Effcct of inositol-1,4,5-trisphosphate on isolated subcellular fractions of rat pancreas.J. Membrane Biol. 81:241–253
Streb, H., Irvine, R.F., Berridge, M.J., Schulz, I. 1983. Release of Ca2+ from a nonmitochondrial intracellular store in pancreatic acinar cells by inositol-1,4,5-trisphosphate.Nature (London) 306:67–69
Streb, H., Schulz, I. 1983. Regulation of cytosolic free Ca2+ concentration in acinar cells of rat pancreas.Am. J. Physiol. 245:G347-G357
Taylor, C.W., Berridge, M.J., Brown, K.D., Cooke, A.M., Potter, B.V.L. 1988.Myo-inositol 1,4,5-trisphosphorothioate mobilizes intracellular calcium in Swiss 3T3 cells andXenopus oocytes.Biochem. Biophys. Res. Commun. 150:626–632
Taylor, C.W., Berridge, M.J., Cooke, A.M., Potter, B.V.L. 1989. Inositol 1,4,5-trisphosphorothioate: A stable analogue of inositol 1,4,5-trisphosphate which mobilizes intracellular calcium.Biochem. J. (in press)
Thévenod, F., Kemmer, T.P., Christian, A.L., Schulz, I. 1989. Characterization of MgATP-driven H+ uptake into a microsomal vesicle fraction from rat pancreatic acinar cells.J. Membrane Biol. 107:263–275
Thévenod, F., Schulz, I. 1988. H+-ion dependent calcium uptake into an inositol 1,4,5-trisphosphate sensitive calcium pool from rat parotid gland.Am. J. Physiol. 255:G429-G440
Volpe, P., Krause, K.-H., Hashimoto, S., Zorzato, F., Pozzan, T., Meldolesi, J., Lew, D.P. 1988. “Calciosome,” a cytoplasmic organelle: The inositol 1,4,5-trisphosphate-sensitive Ca2+ store of nonmuscle cells?Proc. Natl. Acad. Sci USA 85:1091–1095
Willcocks, A.L., Potter, B.V.L., Cooke, A.M., Nahorski, S.R. 1988.Myo-inositol 1,4,5-trisphosphorothioate binds to specific3H-inosiiol 1,4,5-trisphosphate sites in rat cerebellum and is resistant to 5-phosphatase.Eur. J. Pharmacol. 155:181–183
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Thévenod, F., Dehlinger-Kremer, M., Kemmer, T.P. et al. Characterization of inositol 1,4,5-trisphosphate-sensitive (IsCaP) and-insensitive (IisCaP) nonmitochondrial Ca2+ pools in rat pancreatic acinar cells. J. Membrain Biol. 109, 173–186 (1989). https://doi.org/10.1007/BF01870856
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DOI: https://doi.org/10.1007/BF01870856