Muscarinic inhibition of adenylyl cyclase regulates intracellular calcium in single airway smooth muscle cells
Madison, J.M.; Yamaguchi, H.
American Journal of Physiology 270(2 Pt 1): L208-L214
1996
ISSN/ISBN: 0002-9513 PMID: 8779989 Document Number: 465724
To determine whether muscarinic agonists attenuated isoproterenol-stimulated decreases in intracellular calcium concentration ((Ca-2+)-i), changes in (Ca-2+)-i were measured in single airway smooth muscle cells using ratiometric analysis of fura 2 fluorescence. Isoproterenol (10-5 M) and 8-bromoadenosine 3',5'-cyclic monophosphate (8-BrcAMP) decreased (Ca-2+)-i by 24 +- 3% (P lt 0.05, n = 28) and 17 +- 1% (P lt 0.05, n = 6), respectively. The decreased (Ca-2+)-i in response to isoproterenol was inhibited by propranolol (10-6 M) and Rp-adenosine 3',5'-cyclic monophosphothioate (Rp-cAMPS) (10-40 mu-M). In subsequent experiments assessing the effects of muscarinic agonists, isoproterenol did not decrease (Ca-2+)-i in the presence of carbachol (5 times 10-8 M) (6 +- 8% increase; NS, n = 8). To determine the mechanism underlying this inhibitory effect of carbachol, cells were loaded with 4,5-dimethoxy-2-nitrobenzyl adenosine-3',5'-cyclic monophosphate (caged cAMP). For cells loaded with 20 mu-M caged cAMP, photolysis of caged cAMP decreased basal (Ca-2+)-i by 28 +- 3% (P lt 0.05, n = 12). In the presence of carbachol (5 times 10-8 M), photolysis of caged cAMP still induced a 27 +- 4% decrease in (Ca-2+)-i (p lt 0.05, n = 12). We concluded that a low concentration of carbachol did attenuate isoproterenol-stimulated decreases in (Ca-2+)-i. Because low concentrations of carbachol attenuated the decreases in (Ca-2+)-i stimulated by isoproterenol but not the comparable decreases stimulated by cAMP directly, we concluded that the inhibition of adenylyl cyclase activity by muscarinic agonists contributed to the regulation of (Ca-2+)-i in airway smooth muscle cells. The findings suggested that physiological levels of cholinergic stimulation inhibit adenylyl cyclase, thereby attenuating the effects that beta-adrenergic agonists have on (Ca-2+)-i.