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Acute reversible SERCA blockade facilitates or blocks exocytosis, respectively in mouse or bovine chromaffin cells

  • Signaling and cell physiology
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Abstract

Pre-blockade of the sarco-endoplasmic reticulum (ER) calcium ATPase (SERCA) with irreversible thapsigargin depresses exocytosis in adrenal bovine chromaffin cells (BCCs). Distinct expression of voltage-dependent Ca2+-channel subtypes and of the Ca2+-induced Ca2+ release (CICR) mechanism in BCCs versus mouse chromaffin cells (MCCs) has been described. We present a parallel study on the effects of the acute SERCA blockade with reversible cyclopizonic acid (CPA), to repeated pulsing with acetylcholine (ACh) at short (15 s) and long intervals (60 s) at 37 °C, allowing the monitoring of the initial size of a ready-release vesicle pool (RRP) and its depletion and recovery in subsequent stimuli. We found (i) strong depression of exocytosis upon ACh pulsing at 15-s intervals and slower depression at 60-s intervals in both cell types; (ii) facilitation of exocytosis upon acute SERCA inhibition, with back to depression upon CPA washout in MCCs; (iii) blockade of exocytosis upon acute SERCA inhibition and pronounced rebound of exocytosis upon CPA washout in BCCs; (iv) basal [Ca2+]c elevation upon stimulation with ACh at short intervals (but not at long intervals) in both cell types; and (v) augmentation of basal [Ca2+]c and inhibition of peak [Ca2+]c amplitude upon CPA treatment in both cell types, with milder effects upon stimulation at 60-s intervals. These results are compatible with the view that while in MCCs the uptake of Ca2+ via SERCA contributes to the mitigation of physiological ACh triggered secretion, in BCCs the uptake of Ca2+ into the ER facilitates such responses likely potentiating a Ca2+-induced Ca2+ release mechanism. These drastic differences in the regulation of ACh-triggered secretion at 37 °C may help to understand different patterns of the regulation of exocytosis by the circulation of Ca2+ at a functional ER Ca2+ store.

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References

  1. Alés E, Fuentealba J, García AG, López MG (2005) Depolarization evokes different patterns of calcium signals and exocytosis in bovine and mouse chromaffin cells: the role of mitochondria. Eur J Neurosci 21:142–150. https://doi.org/10.1111/j.1460-9568.2004.03861.x

    Article  PubMed  Google Scholar 

  2. Alonso MT, Barrero MJ, Michelena P, Carnicero E, Cuchillo I, García AG, García-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 144:241–254. https://doi.org/10.1083/jcb.144.2.241

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Berridge M, Lipp P, Bootman M (1999) Calcium signalling. Curr Biol 9:R157–R159. https://doi.org/10.1016/s0960-9822(99)80101-8

    Article  CAS  PubMed  Google Scholar 

  4. Calvo-Gallardo E, López-Gil Á, Méndez-López I, Martínez-Ramírez C, Padín JF, García AG (2016) Faster kinetics of quantal catecholamine release in mouse chromaffin cells stimulated with acetylcholine, compared with other secretagogues. J Neurochem 139:722–736. https://doi.org/10.1111/jnc.13849

    Article  CAS  PubMed  Google Scholar 

  5. Calvo-Gallardo E, de Pascual R, Fernández-Morales J-C, Arranz-Tagarro J-A, Maroto M, Nanclares C, Gandía L, de Diego AMG, Padín J-F, García AG (2015) Depressed excitability and ion currents linked to slow exocytotic fusion pore in chromaffin cells of the SOD1(G93A) mouse model of amyotrophic lateral sclerosis. Am J Physiol Cell Physiol 308:C1-19. doi: https://doi.org/10.1152/ajpcell.00272.2014

  6. Carbone E, Borges R, Eiden LE, García AG, Hernández-Cruz A (2019) Chromaffin cells of the adrenal medulla: physiology, pharmacology, and disease. Compr Physiol 9:1443–1502. https://doi.org/10.1002/cphy.c190003

    Article  PubMed  Google Scholar 

  7. Chow RH, von Rüden L, Neher E (1992) Delay in vesicle fusion revealed by electrochemical monitoring of single secretory events in adrenal chromaffin cells. Nature 356:60–63. https://doi.org/10.1038/356060a0

    Article  CAS  PubMed  Google Scholar 

  8. Cuchillo-Ibáñez I, Olivares R, Aldea M, Villarroya M, Arroyo G, Fuentealba J, García AG, Albillos A (2002) Acetylcholine and potassium elicit different patterns of exocytosis in chromaffin cells when the intracellular calcium handling is disturbed. Pflugers Arch 444:133–142. https://doi.org/10.1007/s00424-002-0810-4

    Article  CAS  PubMed  Google Scholar 

  9. Demaurex N, Lew DP, Krause KH (1992) Cyclopiazonic acid depletes intracellular Ca2+ stores and activates an influx pathway for divalent cations in HL-60 cells. J Biol Chem 267:2318–2324

    Article  CAS  Google Scholar 

  10. Dinkelacker V, Voets T, Neher E, Moser T (2000) The readily releasable pool of vesicles in chromaffin cells is replenished in a temperature-dependent manner and transiently overfills at 37 degrees C. J Neurosci 20:8377–8383

    Article  CAS  Google Scholar 

  11. Douglas WW, Poisner AM (1961) Stimulation of uptake of calcium-45 in the adrenal gland by acetylcholine. Nature 192:1299. https://doi.org/10.1038/1921299a0

    Article  CAS  PubMed  Google Scholar 

  12. Douglas WW, Rubin RP (1961) The role of calcium in the secretory response of the adrenal medulla to acetylcholine. J Physiol 159:40–57. https://doi.org/10.1113/jphysiol.1961.sp006791

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Duchen MR (1999) Contributions of mitochondria to animal physiology: from homeostatic sensor to calcium signalling and cell death. J Physiol 516(Pt 1):1–17. https://doi.org/10.1111/j.1469-7793.1999.001aa.x

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Fenwick EM, Marty A, Neher E (1982) Sodium and calcium channels in bovine chromaffin cells. J Physiol 331:599–635. https://doi.org/10.1113/jphysiol.1982.sp014394

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. García AG, García-De-Diego AM, Gandía L, Borges R, García-Sancho J (2006) Calcium signaling and exocytosis in adrenal chromaffin cells. Physiol Rev 86:1093–1131. https://doi.org/10.1152/physrev.00039.2005

    Article  CAS  PubMed  Google Scholar 

  16. García AG, Padín F, Fernández-Morales JC, Maroto M, García-Sancho J (2012) Cytosolic organelles shape calcium signals and exo-endocytotic responses of chromaffin cells. Cell Calcium 51:309–320. https://doi.org/10.1016/j.ceca.2011.12.004

    Article  CAS  PubMed  Google Scholar 

  17. García-Sancho J, de Diego AMG, García AG (2012) Mitochondria and chromaffin cell function. Pflugers Arch 464:33–41. https://doi.org/10.1007/s00424-012-1074-2

    Article  CAS  PubMed  Google Scholar 

  18. Heinemann C, von Rüden L, Chow RH, Neher E (1993) A two-step model of secretion control in neuroendocrine cells. Pflugers Arch 424:105–112. https://doi.org/10.1007/BF00374600

    Article  CAS  PubMed  Google Scholar 

  19. Herrington J, Park YB, Babcock DF, Hille B (1996) Dominant role of mitochondria in clearance of large Ca2+ loads from rat adrenal chromaffin cells. Neuron 16:219–228. https://doi.org/10.1016/s0896-6273(00)80038-0

    Article  CAS  PubMed  Google Scholar 

  20. Kawagoe KT, Zimmerman JB, Wightman RM (1993) Principles of voltammetry and microelectrode surface states. J Neurosci Methods 48:225–240. https://doi.org/10.1016/0165-0270(93)90094-8

    Article  CAS  PubMed  Google Scholar 

  21. Lara B, López MG, Villarroya M, Gandía L, Cleeman L, Morad M, García AG (1997) A caffeine-sensitive Ca2+ store modulates K+-evoked secretion in chromaffin cells. Am J Phys 272:C1211–C1221. https://doi.org/10.1152/ajpcell.1997.272.4.C1211

    Article  CAS  Google Scholar 

  22. López-Gil A, Nanclares C, Méndez-López I, Martínez-Ramírez C, de Los RC, Padín-Nogueira JF, Montero M, Gandía L, García AG (2017) The quantal catecholamine release from mouse chromaffin cells challenged with repeated ACh pulses is regulated by the mitochondrial Na+ /Ca2+ exchanger. J Physiol 595:2129–2146. https://doi.org/10.1113/JP273339

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Mahapatra S, Calorio C, Vandael DHF, Marcantoni A, Carabelli V, Carbone E (2012) Calcium channel types contributing to chromaffin cell excitability, exocytosis and endocytosis. Cell Calcium 51:321–330. https://doi.org/10.1016/j.ceca.2012.01.005

    Article  CAS  PubMed  Google Scholar 

  24. Michelena P, Vega T, Montiel C, López MG, García-Perez LE, Gandía L, García AG (1995) Effects of tyramine and calcium on the kinetics of secretion in intact and electroporated chromaffin cells superfused at high speed. Pflugers Arch 431:283–296. https://doi.org/10.1007/BF00410202

    Article  CAS  PubMed  Google Scholar 

  25. Milla J, Montesinos MS, Machado JD, Borges R, Alonso E, Moreno-Ortega AJ, Cano-Abad MF, García AG, Ruiz-Nuño A (2011) Ouabain enhances exocytosis through the regulation of calcium handling by the endoplasmic reticulum of chromaffin cells. Cell Calcium 50:332–342. https://doi.org/10.1016/j.ceca.2011.06.002

    Article  CAS  PubMed  Google Scholar 

  26. Miranda-Ferreira R, de Pascual R, Smaili SS, Caricati-Neto A, Gandía L, García AG, Jurkiewicz A (2010) Greater cytosolic and mitochondrial calcium transients in adrenal medullary slices of hypertensive, compared with normotensive rats. Eur J Pharmacol 636:126–136. https://doi.org/10.1016/j.ejphar.2010.03.044

    Article  CAS  PubMed  Google Scholar 

  27. Mollard P, Seward EP, Nowycky MC (1995) Activation of nicotinic receptors triggers exocytosis from bovine chromaffin cells in the absence of membrane depolarization. Proc Natl Acad Sci U S A 92:3065–3069. https://doi.org/10.1073/pnas.92.7.3065

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Montero M, Alonso MT, Carnicero E, Cuchillo-Ibáñez I, Albillos A, García AG, García-Sancho J, Alvarez J (2000) Chromaffin-cell stimulation triggers fast millimolar mitochondrial Ca2+ transients that modulate secretion. Nat Cell Biol 2:57–61. https://doi.org/10.1038/35000001

    Article  CAS  PubMed  Google Scholar 

  29. Neher E (1998) Vesicle pools and Ca2+ microdomains: new tools for understanding their roles in neurotransmitter release. Neuron 20:389–399

    Article  CAS  Google Scholar 

  30. Novalbos J, Abad-Santos F, Zapater P, Alvarez J, Alonso MT, Montero M, García AG (1999) Novel antimigraineur dotarizine releases Ca2+ from caffeine-sensitive Ca2+ stores of chromaffin cells. Br J Pharmacol 128:621–626. https://doi.org/10.1038/sj.bjp.0702853

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Pan CY, Fox AP (2000) Rundown of secretion after depletion of intracellular calcium stores in bovine adrenal chromaffin cells. J Neurochem 75:1132–1139. https://doi.org/10.1046/j.1471-4159.2000.0751132.x

    Article  CAS  PubMed  Google Scholar 

  32. Poulsen JC, Caspersen C, Mathiasen D, East JM, Tunwell RE, Lai FA, Maeda N, Mikoshiba K, Treiman M (1995) Thapsigargin-sensitive Ca(2+)-ATPases account for Ca2+ uptake to inositol 1,4,5-trisphosphate-sensitive and caffeine-sensitive Ca2+ stores in adrenal chromaffin cells. Biochem J 307(Pt 3):749–758. https://doi.org/10.1042/bj3070749

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Rigual R, Montero M, Rico AJ, Prieto-Lloret J, Alonso MT, Alvarez J (2002) Modulation of secretion by the endoplasmic reticulum in mouse chromaffin cells. Eur J Neurosci 16:1690–1696. https://doi.org/10.1046/j.1460-9568.11-2.02244.x

    Article  PubMed  Google Scholar 

  34. von Rüden L, Neher E (1993) A Ca-dependent early step in the release of catecholamines from adrenal chromaffin cells. Science 262:1061–1065. https://doi.org/10.1126/science.8235626

    Article  Google Scholar 

  35. Segura F, Brioso MA, Gómez JF, Machado JD, Borges R (2000) Automatic analysis for amperometrical recordings of exocytosis. J Neurosci Methods 103:151–156. https://doi.org/10.1016/s0165-0270(00)00309-5

    Article  CAS  PubMed  Google Scholar 

  36. Voets T, Neher E, Moser T (1999) Mechanisms underlying phasic and sustained secretion in chromaffin cells from mouse adrenal slices. Neuron 23:607–615. https://doi.org/10.1016/s0896-6273(00)80812-0

    Article  CAS  PubMed  Google Scholar 

  37. Wightman RM, Jankowski JA, Kennedy RT, Kawagoe KT, Schroeder TJ, Leszczyszyn DJ, Near JA, Diliberto EJ, Viveros OH (1991) Temporally resolved catecholamine spikes correspond to single vesicle release from individual chromaffin cells. Proc Natl Acad Sci U S A 88:10754–10758. https://doi.org/10.1073/pnas.88.23.10754

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Wu P-C, Fann M-J, Kao L-S (2010) Characterization of Ca2+ signaling pathways in mouse adrenal medullary chromaffin cells. J Neurochem 112:1210–1222. https://doi.org/10.1111/j.1471-4159.2009.06533.x

    Article  CAS  PubMed  Google Scholar 

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Funding

We acknowledge the support received from the EU Horizon 2020 Research and Innovation Program under Maria Sklodowska-Curie (Grant Agreement No. 766124), the Ministerio de Economía y Competitividad (MINECO, Spain; Grant No. SAF2016-78892R), and the Fundación Teófilo Hernando.

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Authors and Affiliations

  1. Instituto Teófilo Hernando, Universidad Autónoma de Madrid, Madrid, Spain

    Carmen Martínez-Ramírez, Irene Gil-Gómez, Antonio M. G. de Diego & Antonio G. García

  2. Departamento de Farmacología, Universidad Autónoma de Madrid, Madrid, Spain

    Carmen Martínez-Ramírez, Irene Gil-Gómez, Antonio M. G. de Diego & Antonio G. García

  3. Fundación Teófilo Hernando, Parque científico de Madrid, Madrid, Spain

    Carmen Martínez-Ramírez, Irene Gil-Gómez, Antonio M. G. de Diego & Antonio G. García

  4. Instituto de Investigación Sanitaria del Hospital de La Princesa, Facultad de Medicina, Universidad Autónoma de Madrid, Madrid, Spain

    Antonio M. G. de Diego & Antonio G. García

  5. DNS Neuroscience, Instituto Teófilo Hernando, Department of Pharmacology, Universidad Autónoma de Madrid, Madrid, Spain

    Antonio M. G. de Diego & Antonio G. García

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  1. Carmen Martínez-Ramírez
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  2. Irene Gil-Gómez
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  4. Antonio G. García
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Correspondence to Antonio M. G. de Diego.

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All experiments were carried out in accordance with the recommendation of the Ethics Committee from Universidad Autónoma de Madrid, on the use of animals for laboratory experimentation.

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Martínez-Ramírez, C., Gil-Gómez, I., G. de Diego, A.M. et al. Acute reversible SERCA blockade facilitates or blocks exocytosis, respectively in mouse or bovine chromaffin cells. Pflugers Arch - Eur J Physiol 473, 273–286 (2021). https://doi.org/10.1007/s00424-020-02483-1

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