title: Role of the NO-cGMP pathway in the muscarinic regulation of the L-type Ca2+ current in human atrial myocytes.
abstract: 1. The whole-cell patch-clamp technique was used to examine the participation of nitric oxide synthase (NOS) and soluble guanylyl cyclase in the muscarinic regulation of the L-type Ca2+ current (ICa) in freshly isolated human atrial myocytes. 2. Acetylcholine (ACh, 1 microM) decreased basal ICa by 39.1 +/- 5.5% (n = 8) under control conditions, and by 38.0 +/- 6.1% (n = 6) in the presence of 1H-[1,2,4]oxadiazolo[4,3-a]-quinoxaline-1-one (ODQ, 10 microM), a potent guanylyl cyclase inhibitor, and NG-monomethyl-L-arginine (L-NMMA, 1 mM), a competitive NOS inhibitor. L-NMMA alone had no effect on ICa, whilst ODQ increased ICa in 50% of the cells. 3. The accentuated antagonism of ACh on ICa, i.e. its ability to antagonize the stimulatory effect of beta-adrenergic agonists and, by extension, of other cAMP-elevating agents, was examined after the current was stimulated by either the beta-adrenergic agonist isoprenaline (Iso) or serotonin (5-HT). ACh (100 nM or 1 microM) completely blocked the stimulatory effects of 10 nM Iso or 10 nM 5-HT on ICa. 4. Extracellular application of Methylene Blue (MBlue, 10 microM), a guanylyl cyclase inhibitor, antagonized the inhibitory effect of 1 microM ACh on Iso- or 5-HT-stimulated ICa. However, this effect was overcome by a 100-fold higher ACh concentration and was not mimicked by an intracellular application of MBlue. 5. Inhibition of NOS and soluble guanylyl cyclase activities by addition of ODQ (10 microM) and L-NMMA (1 mM) to both extracellular and intracellular solutions, or by a 2 h pre-incubation of the cells with these inhibitors, modified neither the Iso (10 nM) response nor the inhibitory effect of ACh (100 nM or 1 microM) on Iso-stimulated ICa. 6. Extracellular application of the NO donor SNAP (S-nitroso-N-acetyl-D,L-penicillamine) at 100 nM produced a stimulatory effect on ICa in control conditions. This stimulatory effect was abolished by intracellular MBlue (20 microM) or by intracellular and extracellular application of ODQ (10 microM) in combination with L-NMMA (1 mM). 7. We conclude that the NO-cGMP pathway does not contribute significantly to the muscarinic regulation of ICa in human atrial myocytes.
authors.affiliation: Laboratoire de Cardiologie Cellulaire et Moléculaire, INSERM U-446, Université de Paris-Sud, Faculté de Pharmacie, Châtenay-Malabry, France.
authors.forename: G
authors.initials: G
authors.lastname: Vandecasteele
authors.forename: T
authors.initials: T
authors.lastname: Eschenhagen
authors.forename: R
authors.initials: R
authors.lastname: Fischmeister
mesh_terms: D000109:Acetylcholine / Q000037:antagonists & inhibitors / Q000494:pharmacology; D000293:Adolescent; D000328:Adult; D000368:Aged; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism*; D002648:Child; D004558:Electric Stimulation; D004594:Electrophysiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006321:Heart / Q000187:drug effects; D006801:Humans; D066298:In Vitro Techniques; D008564:Membrane Potentials / Q000502:physiology; D008875:Middle Aged; D009206:Myocardium / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D018408:Patch-Clamp Techniques; D010396:Penicillamine / Q000031:analogs & derivatives / Q000494:pharmacology; D011976:Receptors, Muscarinic / Q000187:drug effects / Q000378:metabolism*; D026423:S-Nitroso-N-Acetylpenicillamine; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors; D019323:omega-N-Methylarginine / Q000494:pharmacology
chemical_list: D015220:Calcium Channels; D004791:Enzyme Inhibitors; D011976:Receptors, Muscarinic; D019323:omega-N-Methylarginine; D009569:Nitric Oxide; D026423:S-Nitroso-N-Acetylpenicillamine; D019001:Nitric Oxide Synthase; D006162:Guanylate Cyclase; D000254:Sodium-Potassium-Exchanging ATPase; D010396:Penicillamine; D000109:Acetylcholine