Abstract
Adrenal medullary chromaffin cells secrete catecholamines (CA) in response to cholinergic receptor activation by acetylcholine (ACh) released from splacnic nerve terminals. In cultured bovine chromaffin cells nicotinic receptors play a preponderant (> 90%) role in the control of CA release. By contrast, we found and report here that up to 40% of the ACh-evoked CA secretion from cultured porcine chromaffin cells can be associated with muscarinic receptor activation. The following results support our belief that in porcine adrenal medullary cells ACh (100 μM) evoked CA secretion is mediated by both nicotinic and muscarinic cholinergic receptors. 1) Hexamethonium (100 μM), a nicotinic receptor antagonist, inhibited ACh-induced CA secretion to ca. 40% of the control release and atropine (1 μM), a muscarinic receptor antagonist, inhibited to ca. 60% of the control value. 2) We also found that ACh (100 μM) evoked intracellular Ca2+ concentration ([Ca2+]i) rise was inhibited by these receptor antagonists to a different extent, and reversibly reduced by lowering the concentration of Ca2+ in the external medium ([Ca2+]o). This last maneuver ([Ca2+]o < 0.1 μM) per se caused a marked reduction in the peak phase of the [Ca2+]i rise evoked by ACh (40% of the control response). Switching the external medium back to physiologic [Ca2+]o in the continued presence of ACh caused a partial recovery of the elevated [Ca2+]i. This [Ca2+]o-dependent [Ca2+]i rise was blocked by hexamethonium (100 μM) but not by atropine (1 μM). Conversely, the ACh-evoked [Ca2+]i rise in low external [Ca2+]o was blocked by atropine but not by hexamethonium. From these data we conclude that in porcine adrenal medullary cells an important fraction (ca. 0.4) of both ACh-induced CA secretion and peak [Ca2+]i rise is due to muscarinic receptor activation.
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References
Douglas WW, Rubin RP: The role of calcium in the secretory response to the adrenal medulla to acetylcholine. J Physiol 159: 40–57, 1961
Wilson SP and Kirshner N: The acetylcholine receptor of the adrenal medulla. J Neurochem 28: 687–695, 1977
Holz RW, Senter RA, Frye RA: Relationship between Ca2+ uptake and catecholamine secretion in primary dissociated cultures of adrenal medulla. J Neurochem 39: 635–646, 1982
Kilpatrick DL, Slepetis RJ, Corcoran JJ and Kirshner N: Calcium up-take and catecholamine secretion by cultured bovine adrenal medulla cells. J Neurochem 38: 427–435, 1982
Kim KT, Westhead EW: Cellular responses to Ca2+ from extracellular and intracellular sources are different as shown by simultaneous measurements of cytosolic Ca2+ and secretion from bovine chromaffin cells. Proc Natl Acad Sci USA 86: 9881–9885, 1989
Knight DE and Baker PF: Observations on the muscarinic activation of catecholamine secretion in the chicken adrenal. Neuroscience 19: 357–366, 1986
Wakade AR and Wakade TD: Contribution of nicotinic and muscarinic receptors in the secretion of catecholamines evoked by endogenous and exogenous acetylcholine. Neuroscience 10: 973–978, 1983
Role LW, Perlman RL: Both nicotinic and muscarinic receptors mediated catecholamine secretion by isolated guinea-pig chromaffin cells. Neuroscience 10: 979–985, 1983
Poisner AM and Douglas WW: The need for calcium in adrenomedullary secretion evoked by biogenic amines, polypeptides, and muscarinic agents. Proc Soc Exp Biol Med 123: 62–64, 1966
Ohtsuki H, Morita K, Minami N, Suemitsu T, Tsujimoto A and Dohi T: Involvement of Ca2+ entry and inositol triphosphate-induced internal Ca2+ mobilization in muscarinic receptor-mediated catecholamine release in dog adrenal chromaffin cells. Neurochem Int 21: 259–267, 1992
Nassar-Gentina V, Pollard HB and Rojas E: Electrical and secretory response to cholinergic stimulation in mouse and human adrenal medullary chromaffin cells. ln: Hidalgo C, Bacigalupo J, Jaimovich E, Vergara J (eds). Transduction in biological systems. Plenum Press, New York, 1990, pp 143–162
Xu Y, Duarte EP and Forsberg EJ: Calcium dependency of muscarinic and nicotinic agonist-induced ATP and catecholamine secretion from porcine adrenal chromaffin cells. J Neurochem 56: 1889–1896, 1991
Kilpatrick DL, Slepetis R and Kirshner N: Ion channels and membrane potential in stimulus-secretion coupling in adrenal medulla cells. J Neurochem 36: 1245–1255, 1981
Knight DE, Kesteven NT: Evoked transient intracellular free Ca changes and secretion in isolated bovine adrenal medullary cells. Proc R Soc Ser B 218: 177–199, 1983
Abad F, Garrido B, Lopez MG, Garcia AG: The source of calcium for muscarinic-mediated catecholamine release from cat adrenals. J Physiol 445: 725–740, 1992
Yamagami K, Nishimura S, Sorimachi M: Internal Ca2+ mobilization by muscarinic stimulation increases secretion from adrenal chromaffin cells only in the presence of Ca2+ influx. J Neurochem 57: 1681–1689, 1991
Malhotra RK, Wakade TD, Wakade AR: Comparison of secretion of catecholamines from the rat adrenal medulla during continuous exposure to nicotine, muscarine or excess K. Neuroscience 26: 313–320, 1988
Misbahuddin M, Oka M: Muscarinic stimulation of guinea pig adrenal chromaffin cells stimulates catecholamine secretion without significant increase in Ca2+ uptake. Neurosci Lett 87: 266–270, 1988
Nakazato Y, Ohga A, Oleshansky M, Tomita U, Yamada Y: Voltage-independent catecholamine release mediated by the activation of muscarinic receptors in guinea-pig adrenal glands. Br J Pharmacol 93: 101–109, 1988
Pollard HB, Pazoles CJ, Creutz CE, Scott JH, Zinder O, Hotchkiss A: An osmotic mechanism for exocytosis from dissociated chromaffin cells. J Biol Chem 259: 1114–1121, 1984
Nassar-Gentina V, Rojas E and Luxoro M: Rise in cytoplasmic Ca2+ induced by monensin in bovine medullary chromaffin cells. Cell Calcium 16: 475–480, 1994
Kao JPY, Harootunian AT, Tsien RY: Photochemically generated cytosolic calcium pulses and their detection by fluo-3. J Biol Chem 264: 8171–8178, 1989
Anton AH, Sayre DF: A study of the factors affecting the aluminum oxide-trihydroxyindole procedure for the analysis of catecholamines. J Pharmacol Exp Ther 138: 360–375, 1962
Kao LS, Schneider AS: Muscarinic receptors on bovine adrenal chromaffin cells mediate a rise in cytosolic calcium that is independent of extracellular calcium. J Biol Chem 260: 2019–2022, 1985
Harish OE, Kao LS, Raffianello R, Wakade AR, Schneider AiS: Calcium dependence of muscarinic receptor-mediated catecholamine secretion from the perfused rat adrenal medulla. J Neurochem 48: 1730–1735, 1987
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Nassar-Gentina, V., Catalán, L. & Luxoro, M. Nicotonic and muscarinic components in acetylcholine stimulation of porcine adrenal medullary cells. Mol Cell Biochem 169, 107–113 (1997). https://doi.org/10.1023/A:1006867423715
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DOI: https://doi.org/10.1023/A:1006867423715