Advertisement

Pflügers Archiv - European Journal of Physiology

, Volume 468, Issue 10, pp 1779–1792 | Cite as

Regulation by L channels of Ca2+-evoked secretory responses in ouabain-treated chromaffin cells

  • Ricardo De Pascual
  • Inés Colmena
  • Lucía Ruiz-Pascual
  • Andrés Mateo Baraibar
  • Javier Egea
  • Luis Gandía
  • Antonio G. GarcíaEmail author
Signaling and cell physiology

Abstract

It is known that the sustained depolarisation of adrenal medullary bovine chromaffin cells (BCCs) with high K+ concentrations produces an initial sharp catecholamine release that subsequently fades off in spite depolarisation persists. Here, we have recreated a sustained depolarisation condition of BCCs by treating them with the Na+/K+ ATPase blocker ouabain; in doing so, we searched experimental conditions that permitted the development of a sustained long-term catecholamine release response that could be relevant during prolonged stress. BCCs were perifused with nominal 0Ca2+ solution, and secretion responses were elicited by intermittent application of short 2Ca2+ pulses (Krebs-HEPES containing 2 mM Ca2+). These pulses elicited a biphasic secretory pattern with an initial 30-min period with secretory responses of increasing amplitude and a second 30-min period with steady-state, non-inactivating responses. The initial phase was not due to gradual depolarisation neither to gradual increases of the cytosolic calcium transients ([Ca2+]c) elicited by 2Ca2+ pulses in BBCs exposed to ouabain; both parameters increased soon after ouabain addition. Νifedipine blocked these responses, and FPL64176 potentiated them, suggesting that they were triggered by Ca2+ entry through non-inactivating L-type calcium channels. This was corroborated by nifedipine-evoked blockade of the L-type Ca2+ channel current and the [Ca2+]c transients elicited by 2Ca2+ pulses. Furthermore, the plasmalemmal Na+/Ca2+ exchanger (NCX) blocker SEA0400 caused a mild inhibition followed by a large rebound increase of the steady-state secretory responses. We conclude that these two phases of secretion are mostly contributed by Ca2+ entry through L calcium channels, with a minor contribution of Ca2+ entry through the reverse mode of the NCX.

Keywords

Ouabain Chromaffin cell L-type Ca2+ channels N-type Ca2+ channels P/Q-type Ca2+ channels NCX Catecholamine release Amperometry 

Notes

Acknowledgments

Supported by a grant from MINECO (SAF 2013-44108-P). Also by CABYCIC, UAM/Bioiberica, Spain. We thank the continued support of Fundación Teófilo Hernando, Madrid, Spain.

References

  1. 1.
    Albillos A, Carbone E, Gandia L, Garcia AG, Pollo A (1996) Opioid inhibition of Ca2+ channel subtypes in bovine chromaffin cells: selectivity of action and voltage-dependence. Eur J Neurosci 8:1561–1570CrossRefPubMedGoogle Scholar
  2. 2.
    Albillos A, Garcia AG, Gandia L (1993) omega-Agatoxin-IVA-sensitive calcium channels in bovine chromaffin cells. FEBS Lett 336:259–262CrossRefPubMedGoogle Scholar
  3. 3.
    Albillos A, Garcia AG, Olivera B, Gandia L (1996) Re-evaluation of the P/Q Ca2+ channel components of Ba2+ currents in bovine chromaffin cells superfused with solutions containing low and high Ba2+ concentrations. Pflugers Arch 432:1030–1038CrossRefPubMedGoogle Scholar
  4. 4.
    Alonso MT, Barrero MJ, Michelena P, Carnicero E, Cuchillo I, Garcia AG, Garcia-Sancho J, Montero M, Alvarez J (1999) Ca2+-induced Ca2+ release in chromaffin cells seen from inside the ER with targeted aequorin. J Cell Biol 144:241–254CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Arnáiz-Cot JJ, de Diego AM, Hernández-Guijo JM, Gandía L, García AG (2008) A two-step model for acetylcholine control of exocytosis via nicotinic receptors. Biochem Biophys Res Commun 365:413–419CrossRefPubMedGoogle Scholar
  6. 6.
    Aunis D, Garcia AG (1981) Correlation between catecholamine secretion from bovine isolated chromaffin cells and [3H]-ouabain binding to plasma membranes. Br J Pharmacol 72:31–40CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Baker PF, Rink TJ (1975) Catecholamine release from bovine adrenal medulla in response to maintained depolarization. J Physiol 253:593–620CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Bauer N, Muller-Ehmsen J, Kramer U, Hambarchian N, Zobel C, Schwinger RH, Neu H, Kirch U, Grunbaum EG, Schoner W (2005) Ouabain-like compound changes rapidly on physical exercise in humans and dogs: effects of beta-blockade and angiotensin-converting enzyme inhibition. Hypertension 45:1024–1028CrossRefPubMedGoogle Scholar
  9. 9.
    Bittner MA, Holz RW (1992) Kinetic analysis of secretion from permeabilized adrenal chromaffin cells reveals distinct components. J Biol Chem 267:16219–16225PubMedGoogle Scholar
  10. 10.
    Blaustein MP (1993) Physiological effects of endogenous ouabain: control of intracellular Ca2+ stores and cell responsiveness. Am J Phys 264:C1367–C1387Google Scholar
  11. 11.
    Borges R, Sala F, Garcia AG (1986) Continuous monitoring of catecholamine release from perfused cat adrenals. J Neurosci Methods 16:289–300CrossRefPubMedGoogle Scholar
  12. 12.
    Chan SA, Polo-Parada L, Smith C (2005) Action potential stimulation reveals an increased role for P/Q-calcium channel-dependent exocytosis in mouse adrenal tissue slices. Arch Biochem Biophys 435:65–73CrossRefPubMedGoogle Scholar
  13. 13.
    de Diego AM, Tapia L, Alvarez RM, Mosquera M, Cortes L, Lopez I, Gutierrez LM, Gandia L, Garcia AG (2008) A low nicotine concentration augments vesicle motion and exocytosis triggered by K(+) depolarisation of chromaffin cells. Eur J Pharmacol 598:81–86CrossRefPubMedGoogle Scholar
  14. 14.
    Douglas WW (1968) Stimulus-secretion coupling: the concept and clues from chromaffin and other cells. Br J Pharmacol 34:451–474CrossRefPubMedGoogle Scholar
  15. 15.
    Douglas WW, Poisner AM (1962) On the mode of action of acetylcholine in evoking adrenal medullary secretion: increased uptake of calcium during the secretory response. J Physiol 162:385–392CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Douglas WW, Rubin RP (1961) Mechanism of nicotinic action at the adrenal medulla: calcium as a link in stimulus-secretion coupling. Nature 192:1087–1089CrossRefPubMedGoogle Scholar
  17. 17.
    Douglas WW, Rubin RP (1961) The role of calcium in the secretory response of the adrenal medulla to acetylcholine. J Physiol Paris 159:40–57CrossRefGoogle Scholar
  18. 18.
    Fenwick EM, Marty A, Neher E (1982) A patch-clamp study of bovine chromaffin cells and of their sensitivity to acetylcholine. J Physiol 331:577–597CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Fenwick EM, Marty A, Neher E (1982) Sodium and calcium channels in bovine chromaffin cells. J Physiol 331:599–635CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Gandia L, Albillos A, Garcia AG (1993) Bovine chromaffin cells possess FTX-sensitive calcium channels. Biochem Biophys Res Commun 194:671–676CrossRefPubMedGoogle Scholar
  21. 21.
    Gandia L, Garcia AG, Morad M (1993) ATP modulation of calcium channels in chromaffin cells. J Physiol 470:55–72CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Gandia L, Lara B, Imperial JS, Villarroya M, Albillos A, Maroto R, Garcia AG, Olivera BM (1997) Analogies and differences between omega-conotoxins MVIIC and MVIID: binding sites and functions in bovine chromaffin cells. Pflugers Arch 435:55–64CrossRefPubMedGoogle Scholar
  23. 23.
    García AG, García-De-Diego AM, Gandía L, Borges R, García-Sancho J (2006) Calcium signaling and exocytosis in adrenal chromaffin cells. Physiol Rev 86:1093–1131CrossRefPubMedGoogle Scholar
  24. 24.
    Garcia AG, Hernandez M, Horga JF, Sanchez-Garcia P (1980) On the release of catecholamines and dopamine-beta-hydroxylase evoked by ouabain in the perfused cat adrenal gland. Br J Pharmacol 68:571–583CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Gil A, Viniegra S, Gutierrez LM (2001) Temperature and PMA affect different phases of exocytosis in bovine chromaffin cells. Eur J Neurosci 13:1380–1386CrossRefPubMedGoogle Scholar
  26. 26.
    Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ (1981) Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflugers Arch 391:85–100CrossRefPubMedGoogle Scholar
  27. 27.
    Haynes CL, Siff LN, Wightman RM (2007) Temperature-dependent differences between readily releasable and reserve pool vesicles in chromaffin cells. Biochim Biophys Acta 1773:728–735CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Hernández-Guijo JM, Carabelli V, Gandía L, García AG, Carbone E (1999) Voltage-independent autocrine modulation of L-type channels mediated by ATP, opioids and catecholamines in rat chromaffin cells. Eur J Neurosci 11:3574–3584CrossRefPubMedGoogle Scholar
  29. 29.
    Hernandez-Guijo JM, Gandia L, de Pascual R, Garcia AG (1997) Differential effects of the neuroprotectant lubeluzole on bovine and mouse chromaffin cell calcium channel subtypes. Br J Pharmacol 122:275–285CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Hernández-Guijo JM, Maneu-Flores VE, Ruiz-Nuno A, Villarroya M, García AG, Gandía L (2001) Calcium-dependent inhibition of L, N, and P/Q Ca2+ channels in chromaffin cells: role of mitochondria. J Neurosci 21:2553–2560PubMedGoogle Scholar
  31. 31.
    Inagami T, Tamura M (1988) Purification and characterization of specific endogenous ouabainlike substance from bovine adrenal. Am J Med Sci 295:400–405CrossRefPubMedGoogle Scholar
  32. 32.
    Kao LS, Westhead EW (1984) Temperature dependence of catecholamine secretion from cultured bovine chromaffin cells. J Neurochem 43:590–592CrossRefPubMedGoogle Scholar
  33. 33.
    Lara B, Gandia L, Martinez-Sierra R, Torres A, Garcia AG (1998) Q-type Ca2+ channels are located closer to secretory sites than L-type channels: functional evidence in chromaffin cells. Pflugers Arch 435:472–478CrossRefPubMedGoogle Scholar
  34. 34.
    Li S, Eim C, Kirch U, Lang RE, Schoner W (1998) Bovine adrenals and hypothalamus are a major source of proscillaridin A- and ouabain-immunoreactivities. Life Sci 62:1023–1033CrossRefPubMedGoogle Scholar
  35. 35.
    Livett BG (1984) Adrenal medullary chromaffin cells in vitro. Physiol Rev 64:1103–1161PubMedGoogle Scholar
  36. 36.
    Lopez MG, Albillos A, de la Fuente MT, Borges R, Gandia L, Carbone E, Garcia AG, Artalejo AR (1994) Localized L-type calcium channels control exocytosis in cat chromaffin cells. Pflugers Arch 427:348–354CrossRefPubMedGoogle Scholar
  37. 37.
    Mahapatra S, Calorio C, Vandael DH, Marcantoni A, Carabelli V, Carbone E (2012) Calcium channel types contributing to chromaffin cell excitability, exocytosis and endocytosis. Cell Calcium 51:321–330CrossRefPubMedGoogle Scholar
  38. 38.
    Matsuda T, Arakawa N, Takuma K, Kishida Y, Kawasaki Y, Sakaue M, Takahashi K, Takahashi T, Suzuki T, Ota T, Hamano-Takahashi A, Onishi M, Tanaka Y, Kameo K, Baba A (2001) SEA0400, a novel and selective inhibitor of the Na + −Ca2+ exchanger, attenuates reperfusion injury in the in vitro and in vivo cerebral ischemic models. J Pharmacol Exp Ther 298:249–256PubMedGoogle Scholar
  39. 39.
    McKechnie K, Killingback PG, Naya I, Ò Conner SE, Smith GW, Wattam DG, Wells E, Whitehead YM, Williams GE (1989) Calcium channel activator properties in a novel non-dihydropyridine, FPL 64176. Br J Pharmacol 96:673Google Scholar
  40. 40.
    Milla J, Montesinos MS, Machado JD, Borges R, Alonso E, Moreno-Ortega AJ, Cano-Abad MF, Garcia AG, Ruiz-Nuno A (2011) Ouabain enhances exocytosis through the regulation of calcium handling by the endoplasmic reticulum of chromaffin cells. Cell Calcium 50:332–342CrossRefPubMedGoogle Scholar
  41. 41.
    Moro MA, López MG, Gandía L, Michelena P, García AG (1990) Separation and culture of living adrenaline- and noradrenaline-containing cells from bovine adrenal medullae. Anal Biochem 185:243–248CrossRefPubMedGoogle Scholar
  42. 42.
    Neher E (1998) Vesicle pools and Ca2+ microdomains: new tools for understanding their roles in neurotransmitter release. Neuron 20:389–399CrossRefPubMedGoogle Scholar
  43. 43.
    Nicholls MG, Lewis LK, Yandle TG, Lord G, McKinnon W, Hilton PJ (2009) Ouabain, a circulating hormone secreted by the adrenals, is pivotal in cardiovascular disease. Fact or fantasy? J Hypertens 27:3–8CrossRefPubMedGoogle Scholar
  44. 44.
    Olivera BM, Miljanich GP, Ramachandran J, Adams ME (1994) Calcium channel diversity and neurotransmitter release: the omega-conotoxins and omega-agatoxins. Annu Rev Biochem 63:823–867CrossRefPubMedGoogle Scholar
  45. 45.
    Orozco C, Garcia-de-Diego AM, Arias E, Hernandez-Guijo JM, Garcia AG, Villarroya M, Lopez MG (2006) Depolarization preconditioning produces cytoprotection against veratridine-induced chromaffin cell death. Eur J Pharmacol 553:28–38CrossRefPubMedGoogle Scholar
  46. 46.
    Padín JF, Fernández-Morales JC, Olivares R, Vestring S, Arranz-Tagarro JA, Calvo-Gallardo E, de Pascual R, Gandía L, Garcia AG (2013) Plasmalemmal sodium-calcium exchanger shapes the calcium and exocytotic signals of chromaffin cells at physiological temperature. Am J Physiol Cell Physiol 305:C160–C172CrossRefPubMedGoogle Scholar
  47. 47.
    Park YB, Herrington J, Babcock DF, Hille B (1996) Ca2+ clearance mechanisms in isolated rat adrenal chromaffin cells. J Physiol 492(Pt 2):329–346CrossRefPubMedPubMedCentralGoogle Scholar
  48. 48.
    Pintado AJ, Herrero CJ, Garcia AG, Montiel C (2000) The novel Na(+)/Ca(2+) exchange inhibitor KB-R7943 also blocks native and expressed neuronal nicotinic receptors. Br J Pharmacol 130:1893–1902CrossRefPubMedPubMedCentralGoogle Scholar
  49. 49.
    Rosa JM, de Diego AM, Gandia L, Garcia AG (2007) L-type calcium channels are preferentially coupled to endocytosis in bovine chromaffin cells. Biochem Biophys Res Commun 357:834–839CrossRefPubMedGoogle Scholar
  50. 50.
    Rosa JM, Gandia L, Garcia AG (2009) Inhibition of N and PQ calcium channels by calcium entry through L channels in chromaffin cells. Pflugers Arch 458:795–807CrossRefPubMedGoogle Scholar
  51. 51.
    Rosa JM, Nanclares C, Orozco A, Colmena I, de Pascual R, Garcia AG, Gandia L (2012) Regulation by L-type calcium channels of endocytosis: an overview. J Mol Neurosci 48:360–367CrossRefPubMedGoogle Scholar
  52. 52.
    Schneider R, Wray V, Nimtz M, Lehmann WD, Kirch U, Antolovic R, Schoner W (1998) Bovine adrenals contain, in addition to ouabain, a second inhibitor of the sodium pump. J Biol Chem 273:784–792CrossRefPubMedGoogle Scholar
  53. 53.
    Skou JC (1957) The influence of some cations on an adenosine triphosphatase from peripheral nerves. Biochim Biophys Acta 23:394–401CrossRefPubMedGoogle Scholar
  54. 54.
    Sorensen JB (2004) Formation, stabilisation and fusion of the readily releasable pool of secretory vesicles. Pflugers Arch 448:347–362CrossRefPubMedGoogle Scholar
  55. 55.
    Villalobos C, Nunez L, Montero M, Garcia AG, Alonso MT, Chamero P, Alvarez J, Garcia-Sancho J (2002) Redistribution of Ca2+ among cytosol and organella during stimulation of bovine chromaffin cells. FASEB J 16:343–353CrossRefPubMedGoogle Scholar
  56. 56.
    Villarroya M, De la Fuente MT, Lopez MG, Gandia L, Garcia AG (1997) Distinct effects of omega-toxins and various groups of Ca(2+)-entry inhibitors on nicotinic acetylcholine receptor and Ca2+ channels of chromaffin cells. Eur J Pharmacol 320:249–257CrossRefPubMedGoogle Scholar
  57. 57.
    Villarroya M, Olivares R, Ruiz A, Cano-Abad MF, de Pascual R, Lomax RB, Lopez MG, Mayorgas I, Gandia L, Garcia AG (1999) Voltage inactivation of Ca2+ entry and secretion associated with N- and P/Q-type but not L-type Ca2+ channels of bovine chromaffin cells. J Physiol 516(Pt 2):421–432CrossRefPubMedPubMedCentralGoogle Scholar
  58. 58.
    von Ruden L, Neher E (1993) A Ca-dependent early step in the release of catecholamines from adrenal chromaffin cells. Science 262:1061–1065CrossRefGoogle Scholar
  59. 59.
    Walker A, Glavinovic MI, Trifaro J (1996) Temperature dependence of release of vesicular content in bovine chromaffin cells. Pflugers Arch 432:885–892CrossRefPubMedGoogle Scholar
  60. 60.
    Watano T, Kimura J, Morita T, Nakanishi H (1996) A novel antagonist, no. 7943, of the Na+/Ca2+ exchange current in Guinea-pig cardiac ventricular cells. Br J Pharmacol 119:555–563CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Ricardo De Pascual
    • 1
    • 2
  • Inés Colmena
    • 1
    • 2
  • Lucía Ruiz-Pascual
    • 1
  • Andrés Mateo Baraibar
    • 1
  • Javier Egea
    • 1
  • Luis Gandía
    • 1
    • 2
  • Antonio G. García
    • 1
    • 2
    • 3
    Email author
  1. 1.Instituto Teófilo Hernando, Facultad de MedicinaUniversidad Autónoma de MadridMadridSpain
  2. 2.Departamento de Farmacología y Terapéutica, Facultad de MedicinaUniversidad Autónoma de MadridMadridSpain
  3. 3.Servicio de Farmacología Clínica, Instituto de Investigación SanitariaHospital Universitario de la Princesa, Universidad Autónoma de MadridMadridSpain

Personalised recommendations