Naunyn-Schmiedeberg's Archives of Pharmacology

, Volume 328, Issue 3, pp 273–278 | Cite as

Modulation by ouabain and diphenylhydantoin of veratridine-induced 22Na influx and its relation to 45Ca influx and the secretion of catecholamines in cultured bovine adrenal medullary cells

  • Akihiko Wada
  • Futoshi Izumi
  • Nobuyuki Yanagihara
  • Hideyuki Kobayashi


The effects of ouabain and diphenylhydantoin on the secretion of catecholamines induced by veratridine were investigated in cultured bovine adrenal medullary cells with special reference to ion fluxes. Veratridine itself induced an influx of 22Na and 45Ca as well as secretion of catecholamines, which were antagonized by tetrodotoxin, a selective inhibitor of voltage dependent Na channels. The secretion of catecholamines caused by veratridine was not observed either in Na free or Ca free medium. Veratridineinduced influx of 45Ca did not occur in Na free medium, while veratridine-induced influx of 22Na occurred even in Ca free medium. Veratridine-induced influx of 22Na, 45Ca and secretion of catecholamines were all potentiated by ouabain, a potent inhibitor of Na, K-ATPase. Omission of K from the medium, a condition which suppresses the Na, K-ATPase activity, also augmented these cell responses caused by veratridine. On the contrary, diphenylhydantoin, which is known to decrease the intracellular concentration of Na, reduced the veratridine-induced influx of 22Na, 45Ca and secretion of catecholamines. The potentiating effects of ouabain on the veratridine-induced cell responses were all abolished by diphenylhydantoin. These findings imply that veratridine, ouabain and K removal as well as diphenylhydantoin modulate the intracellular accumulation of 22Na which is involved in the influx of 45Ca and the secretion of catecholamines.

Key words

Adrenal medulla Secretion 22Na influx 45Ca influx Veratridine 


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  1. Amy C, Kirshner N (1982) 22Na uptake and catecholamine secretion by primary cultures of adrenal medulla cells. J Neurochem 39:132–142Google Scholar
  2. 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: 31–40Google Scholar
  3. Ayala GF, Johnston D (1977) The influences of phenytoin on the fundamental electrical properties of simple neural systems. Epilepsia 18:299–307Google Scholar
  4. Baker PF (1972) Transport and metabolism of calcium in nerve. Prog Biophys Mol Biol 24:177–223Google Scholar
  5. Banks P (1967) The effect of ouabain on the secretion of catecholamines and on the intracellular concentration of potassium. J Physiol 193:631–637Google Scholar
  6. Banks P, Biggins R, Bishop R, Christian B, Currie N (1969) Sodium ions and the secretion of catecholamines. J Physiol 200: 797–805Google Scholar
  7. Biales B, Dichter M, Tischler A (1976) Electrical excitability of cultured adrenal chromaffin cells. J Physiol 262:743–753Google Scholar
  8. Blaustein MP (1974) The influence of sodium on calcium fluxes. Rev Physiol Biochem Pharmacol 70:33–82Google Scholar
  9. Brandt BL, Hagiwara S, Kidokoro Y, Miyazaki S (1976) Action potentials in the rat chromaffin cell and effects of acetylcholine. J Physiol 263:417–439Google Scholar
  10. Carafoli E, Tiozzo R, Lugli G, Crovetti F, Kratzing C (1974) The release of calcium from heart mitochondria by sodium. J Mol Cell Cardiol 6:361–371Google Scholar
  11. Donatsch P, Lowe DA, Richardson BP, Taylor P (1977) The functional significance of sodium channels in pancreatic beta-cell membranes. J Physiol 267:357–376Google Scholar
  12. Douglas WW (1975) Secretomotor control of adrenal medullary secretion: synaptic, membrane and ionic events in stimulussecretion coupling. In: Blaschko H, Sayers G, Smith AD (eds) Handbook of physiology, Section 7, vol. VI. American Physiological Society, Washington DC, pp 367–388Google Scholar
  13. Douglas WW, Poisner AM (1961) Stimulation of uptake of calcium-45 in the adrenal gland by acetylcholine. Nature 192:1299Google Scholar
  14. Douglas WW, Kanno T, Sampson SR (1967) Influence of the ionic environment on the membrane potential of adrenal chromaffin cells and on the depolarizing effect of acetycholine. J Physiol 191:107–121Google Scholar
  15. Feldberg W, Minz B, Tsudzimura H (1934) The mechanism of the nervous discharge of adrenaline. J Physiol 81:286–304Google Scholar
  16. Festoff BW, Appel SH (1968) Effect of diphenylhydantoin on synaptosome sodium-potassium-ATPase. J Clin Invest 47: 2752–2758Google Scholar
  17. Garcia AG, Hernandez M, Horga JF, Sanchez-Garcia P (1980) On the release of catecholamines and dopamine-β-hydroxylase evoked by ouabain in the perfused cat adrenal gland. Br J Pharmacol 68:571–583Google Scholar
  18. Garcia AG, Garcia-Lopez E, Montiel C, Nicolas GP, Sanchez-Garcia P (1981) Correlation between catecholamine release and sodium pump inhibition in the perfused adrenal gland of the cat. Br J Pharmacol 74:665–672Google Scholar
  19. Gutman Y, Boonyaviroj P (1977) Mechanism of inhibition of catecholamine release from adrenal medulla by diphenylhydantoin and low concentration of ouabain (10−10 M). Naunyn-Schmiedeberg's Arch Pharmacol 296:293–296Google Scholar
  20. Ito S, Nakazato Y, Ohga A (1978) Pharmacological evidence for the involvement of Na+ channels in the release of catecholamines from perfused adrenal glands. Br J Pharmacol 62:359–361Google Scholar
  21. Ito S, Nakazato Y, Ohga A (1979) The effect of veratridine on the release of catecholamines from the perfused adrenal gland. Br J Pharmacol 65:319–330Google Scholar
  22. Ito S, Nakazato Y, Ohga A (1980) Exocytotic release of catecholamine from perfused adrenal gland of guinea-pig induced by veratridine. Br J Pharmacol 70:527–535Google Scholar
  23. Kashimoto T, Izumi F, Wada A, Oka M (1980) Role of intracellular sodium in regulation of catecholamine release and calcium movement in perfused bovine adrenal gland. J Occup Environ Health 1:13–18Google Scholar
  24. Kidokoro Y, Ritchie AK (1980) Chromaffin cell action potentials and their possible role in adrenaline secretion from rat adrenal medulla. J Physiol 307:199–216Google Scholar
  25. Kilpatrick DL, Slepetis R, Kirshner N (1981) Ion channels and membrane potential in stimulus-secretion coupling in adrenal medulla cells. J Neurochem 36:1245–1255Google Scholar
  26. Kirpekar SM, Prat JC (1979) Release of catecholamines from perfused cat adrenal gland by veratridine. Proc Natl Acad Sci USA 76:2081–2083Google Scholar
  27. Knight DE, Baker PF (1983) Stimulus-secretion coupling in isolated bovine adrenal medullary cells. Q J Exp Physiol 68:123–143Google Scholar
  28. Knight DE, Whitaker MJ (1978) Veratridine-induced secretion in medullary cells isolated from the bovine adrenal gland. J Physiol 281:18p-19pGoogle Scholar
  29. Lewin E, Bleck V (1971) The effect of diphenylhydantoin administration on sodium-potassium activated ATPase in cortex. Neurology 21:647–651Google Scholar
  30. Lowe DA, Richardson BP, Taylor P, Donatsch P (1976) Increasing intracellular sodium triggers calcium release from bound pools. Nature 260:337–338Google Scholar
  31. Narahashi T (1974) Chemicals as tools in the study of excitable membranes. Physiol Rev 54:813–889Google Scholar
  32. Narahashi T, Moore JW, Scott W (1964) Tetrodotoxin blockage of sodium conductance increase in lobster giant axons. J Gen Physiol 47:965–974Google Scholar
  33. Ohta, M, Narahashi T, Keeler RF (1973) Effects of veratrum alkaloids on membrane potential and conductance of squid and crayfish giant axons. J Pharmacol Exp Ther 184:143–154Google Scholar
  34. Perry JG, McKinney L, Weer PD (1978) The cellular mode of action of the anti-epileptic drug 5,5-diphenylhydantoin. Nature 272:271–273Google Scholar
  35. Pocock G (1983a) Ionic and metabolic requirements for stimulation of secretion by ouabain in bovine adrenal medullary cells. Mol Pharmacol 23:671–680Google Scholar
  36. Pocock G (1983b) Ion movements in isolated bovine adrenal medullary cells treated with ouabain. Mol Pharmacol 23:681–697Google Scholar
  37. Reuter H (1974) Exchange of calcium ion in the mammalian myocardium. Mechanisms and physiological significance. Circ Res 34:599–605Google Scholar
  38. Sakurai S, Wada A, Izumi F, Kobayashi H, Yanagihara N (1983) Inhibition by α2-adrenoceptor agonists of the secretion of catecholamines from isolated adrenal medullary cells. Naunyn-Schmiedeberg's Arch Pharmacol 324:15–19Google Scholar
  39. Skou JC (1965) Enzymatic basis for active transport of sodium and potassium across cell membranes. Physiol Rev 45:596–617Google Scholar
  40. Wada A, Yanagihara N, Izumi F, Sakurai S, Kobayashi H (1983a) Trifluoperazine inhibits 45Ca2+ uptake and catecholamine secretion and synthesis in adrenal medullary cells. J Neurochem 40:481–486Google Scholar
  41. Wada A, Sakurai S, Kobayashi H, Yanagihara N, Izumi F (1983b) Suppression by phospholipase A2 inhibitors of secretion of catecholamines from isolated adrenal medullary cells by suppression of cellular calcium uptake. Biochem Pharmacol 32:1175–1178Google Scholar
  42. Wada A, Yashima N, Izumi F, Kobayashi H, Yanagihara N (1984) Involvement of Na influx in acetylcholine receptor mediated secretion of catecholamines from cultured bovine adrenal medulla cells. Neurosci Lett 47:75–80Google Scholar
  43. Weil-Malherbe H (1952) The chemical estimation of adrenaline like substances in blood. Biochem J 51:311–318Google Scholar
  44. Williams JA (1975) Na+ dependence of in vitro pancreatic amylase release. Am J Physiol 229:1023–1026Google Scholar

Copyright information

© Springer-Verlag 1985

Authors and Affiliations

  • Akihiko Wada
    • 1
  • Futoshi Izumi
    • 1
  • Nobuyuki Yanagihara
    • 1
  • Hideyuki Kobayashi
    • 1
  1. 1.Department of PharmacologyUniversity of Occupational and Environmental Health, School of MedicineFukuokaJapan

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