OBJECTIVES: The present study was designed to investigate the characteristic effects of anabasine on secretion of catecholamines (CA) from the isolated perfused rat adrenal gland and to establish its mechanism of adrenomedullary secretion. METHODS: The adrenal gland was isolated by a modification of the Wakade method, and perfused with normal Krebs-bicarbonate solution. The content of CA was measured using fluorometry. RESULTS: The perfusion of anabasine(30-300 microM) into an adrenal vein for 60 min resulted in great increases in CA secretions in a dose-dependent fashion. Upon repeated injection of anabasine (100 microM) at 120 min-intervals, CA secretion was rapidly decreased after the third injection of anabasine. However, there was no statistical difference between the CA secretory responses of both 1st and 2nd treated groups by the successive administration of anabasine at 120 min-intervals. Tachyphylaxis to the releasing effects of CA evoked by anabasine was observed by repeated administration. Therefore, in all subsequent experiments, anabasine was not administered successively more than twice at only 120 min-intervals. The CA-releasing effects of anabasine were depressed by pretreatment with chlorisondamine (selective neuronal nicotinic receptor antagonist, 1 microM), atropine (muscarinic receptor antagonist, 2 microM), nicardipine (L-type dihydropyridine Ca2+ channel blocker, 1 microM), TMB-8 (anti-releaser of intracellular Ca2 +, 30 microM), and perfusion of EGTA (Ca2+ chelator, 5 mM) plus Ca2+ -free medium. In the presence of anabasine (100 microM), the CA secretory responses induced by acetylcholine (5.32 mM), high K+ (direct membrane-depolarizer, 56 mM), DMPP(selective neuronal nicotinic receptor agonist, 10(-4) M), and McN-A-343 (selective muscarinic M1 receptor agonist, 10(-4) M) were maximally enhanced in the first 4 min. However, as time elapsed, these responses became more inhibited at later periods. Furthermore, the perfusion of nicotine (30 microM) into an adrenal vein for 60 min also caused a great increase in CA secretion, leading to peak response in the first 0-5 min period. In the presence of nicotine (30 microM), the CA secretory responses induced by acetylcholine, high K+, DMPP and McN-A-343 were also enhanced for the first 4min, but later reduced to less than the control release. CONCLUSIONS: Taken together, these experimental results indicate that anabasine affects rat adrenomedullary CA secretion in a calcium-dependent fashion. This facilitatory effect of anabasine may be mediated by activation of both cholinergic nicotinic and muscarinic receptors, which is relevant to both stimulation of Ca2+ influx into adrenomedullary chromaffin cells and Ca2+ release from cytoplasmic Ca2+ Anabasine may be less potent than nicotine in rat adrenomedullary CA secretion. Anabasine, in addition to nicotine, alkaloids present in tobacco smoke may be a risk factor in causing cardiovascular diseases.
OBJECTIVES: The present study was designed to investigate the characteristic effects of anabasine on secretion of catecholamines (CA) from the isolated perfused rat adrenal gland and to establish its mechanism of adrenomedullary secretion. METHODS: The adrenal gland was isolated by a modification of the Wakade method, and perfused with normal Krebs-bicarbonate solution. The content of CA was measured using fluorometry. RESULTS: The perfusion of anabasine(30-300 microM) into an adrenal vein for 60 min resulted in great increases in CA secretions in a dose-dependent fashion. Upon repeated injection of anabasine (100 microM) at 120 min-intervals, CA secretion was rapidly decreased after the third injection of anabasine. However, there was no statistical difference between the CA secretory responses of both 1st and 2nd treated groups by the successive administration of anabasine at 120 min-intervals. Tachyphylaxis to the releasing effects of CA evoked by anabasine was observed by repeated administration. Therefore, in all subsequent experiments, anabasine was not administered successively more than twice at only 120 min-intervals. The CA-releasing effects of anabasine were depressed by pretreatment with chlorisondamine (selective neuronal nicotinic receptor antagonist, 1 microM), atropine (muscarinic receptor antagonist, 2 microM), nicardipine (L-type dihydropyridine Ca2+ channel blocker, 1 microM), TMB-8 (anti-releaser of intracellular Ca2 +, 30 microM), and perfusion of EGTA (Ca2+ chelator, 5 mM) plus Ca2+ -free medium. In the presence of anabasine (100 microM), the CA secretory responses induced by acetylcholine (5.32 mM), high K+ (direct membrane-depolarizer, 56 mM), DMPP(selective neuronal nicotinic receptor agonist, 10(-4) M), and McN-A-343 (selective muscarinic M1 receptor agonist, 10(-4) M) were maximally enhanced in the first 4 min. However, as time elapsed, these responses became more inhibited at later periods. Furthermore, the perfusion of nicotine (30 microM) into an adrenal vein for 60 min also caused a great increase in CA secretion, leading to peak response in the first 0-5 min period. In the presence of nicotine (30 microM), the CA secretory responses induced by acetylcholine, high K+, DMPP and McN-A-343 were also enhanced for the first 4min, but later reduced to less than the control release. CONCLUSIONS: Taken together, these experimental results indicate that anabasine affects rat adrenomedullary CA secretion in a calcium-dependent fashion. This facilitatory effect of anabasine may be mediated by activation of both cholinergic nicotinic and muscarinic receptors, which is relevant to both stimulation of Ca2+ influx into adrenomedullary chromaffin cells and Ca2+ release from cytoplasmic Ca2+ Anabasine may be less potent than nicotine in rat adrenomedullary CA secretion. Anabasine, in addition to nicotine, alkaloids present in tobacco smoke may be a risk factor in causing cardiovascular diseases.
Authors: Paula L Vieira-Brock; Eleanor I Miller; Shannon M Nielsen; Annette E Fleckenstein; Diana G Wilkins Journal: J Chromatogr B Analyt Technol Biomed Life Sci Date: 2011-09-18 Impact factor: 3.205