vasoactive-intestinal-peptide and 6-anilino-5-8-quinolinedione

1. We examined the role of the NO/cyclic GMP (cyclic GMP) pathway in nitric oxide (NO)- and vasoactive intestinal peptide (VIP)-induced relaxation of feline lower oesophageal sphincter (LES). Furthermore, it was studied whether methylene blue, LY83583 and ODQ, which are soluble guanylate cyclase (sGC) inhibitors, could inhibit NO-induced relaxation. 2. The nitric oxide synthase (NOS) inhibitor, N omega-nitro-L-arginine (L-NNA) had no effect in sodium nitropruside (SNP)-induced relaxation, but 3-morpholinosydnonimine-N-ethylcarbamide (SIN-1)-induced relaxation was decreased by the pretreatment of L-NNA, which showed that SIN-1, not SNP, could activate NOS to cause relaxation. Methylene blue and LY83583 did not inhibit the relaxation by SNP and SIN-1. However, the more specific sGC inhibitor ODQ blocked the relaxation induced by NO donors. 3. To identify the relationship of NOS, sGC and adenylate cyclase in VIP-induced relaxation, tissue were pretreated with L-NNA and ODQ and SQ22536. These inhibitors produced significant inhibition of this response to VIP. The adenylyl cyclase inhibitor SQ 22536 also inhibited relaxation by VIP. 4. In conclusion, our data showed that SNP- and SIN-1-induced relaxation was mediated by sGC. Of sGC inhibitors, methylene blue and LY83583 were not adequate for the examination of NO donor-induced feline LES smooth muscle relaxation. VIP also caused relaxation by the pathway involving NO and cGMP and cAMP.

Topics: Aminoquinolines; Animals; Anti-Inflammatory Agents, Non-Steroidal; Cats; Cyclic GMP; Dose-Response Relationship, Drug; Enzyme Inhibitors; Esophagogastric Junction; In Vitro Techniques; Indomethacin; Male; Methylene Blue; Molsidomine; Muscle Relaxation; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase; Nitroprusside; omega-N-Methylarginine; Oxadiazoles; Quinoxalines; Signal Transduction; Vasoactive Intestinal Peptide

Pituitary adenylate cyclase-activating peptide (PACAP) receptors and their signaling pathways were characterized in dispersed rabbit gastric muscle cells. 125I-PACAP-27 and 125I-vasoactive intestinal peptide (VIP) binding to muscle cells were inhibited equally by PACAP and VIP (mean inhibitory concentration 0.8 to 1.3 nM) and desensitized to the same extent (70-80%) by exposure to either peptide. PACAP, like VIP, increased cytosolic free Ca2+ and the formation of L-[3H]citrulline, NO-3/NO-2, guanosine 3',5'-cyclic monophosphate (cGMP), and adenosine 3'5'-cyclic monophosphate (cAMP) and induced relaxation (mean effective concentration 1.8 +/- 0.1 nM) that was partly inhibited by NG-nitro-L-arginine (L-NNA), VIP-(10-28), and PACAP 6-38. L-[3H]citrulline and cGMP formation were blocked by nifedipine, L-NNA, and pertussis toxin (PTx), implying activation of a G protein-coupled, Ca(2+)-calmodulin-dependent nitric oxide (NO) synthase. PACAP-induced relaxation was inhibited to the same extent (46-49%) by nifedipine, L-NNA, PTx, and the protein kinase G inhibitor KT-5823; the inhibition reflected the component of relaxation mediated by the NO-cGMP pathway. The residual relaxation was abolished by the protein kinase A inhibitor H-89. The pattern of inhibition of all responses was identical to that observed with VIP. Desensitization with VIP or PACAP abolished cAMP formation but had no effect on L-[3H]citrulline and cGMP formation induced by either peptide. Receptor protection with VIP or PACAP preserved fully all responses (L-[3H]citrulline, cGMP, and cAMP formation and relaxation) to either peptide. The complete cross-competition, cross-desensitization, cross-antagonism, and cross-protection of receptors by either VIP or PACAP are consistent with interaction of both peptides with the same receptors; the receptors consist of two classes, each coupled to a distinct signaling pathway.

Topics: Adenylate Cyclase Toxin; Alkaloids; Aminoquinolines; Animals; Calcium; Carbazoles; Cells, Cultured; Citrulline; Cyclic AMP; Cyclic GMP; Cytosol; Enzyme Inhibitors; GTP-Binding Proteins; Guanylate Cyclase; Indoles; Isoquinolines; Kinetics; Muscle Relaxation; Muscle, Smooth; Neuropeptides; Neurotransmitter Agents; Nifedipine; Nitric Oxide Synthase; Nitroarginine; Pertussis Toxin; Pituitary Adenylate Cyclase-Activating Polypeptide; Protein Kinase Inhibitors; Rabbits; Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; Receptors, Pituitary Hormone; Signal Transduction; Stomach; Sulfonamides; Vasoactive Intestinal Peptide; Virulence Factors, Bordetella

Vasoactive intestinal peptide release and L-[3H]citrulline production were examined in ganglia isolated from the myenteric plexus of guinea-pig intestine. The nicotinic agonist, 1,1-dimethyl-4-phenylpiperizinium stimulated vasoactive intestinal peptide release and L-[3H]citrulline production; the latter was considered an index of nitric oxide production. Both vasoactive intestinal peptide release and L-[3H]citrulline production were abolished by tetrodotoxin, hexamethonium, and the nitric oxide synthase inhibitor, NG-nitro-L-arginine. Inhibition of vasoactive intestinal peptide release by NG-nitro-L-arginine was reversed by L-arginine but not by D-arginine. Exogenous nitric oxide stimulated vasoactive intestinal peptide release whereas exogenous vasoactive intestinal peptide had no effect on L-[3H]citrulline production. The pattern of stimulation by nitric oxide and inhibition by NG-nitro-L-arginine implied that vasoactive intestinal peptide release is facilitated by and may be dependent on nitric oxide production. Consistent with this notion, vasoactive intestinal peptide release in response to either 1,1-dimethyl-4-phenylpiperizinium or nitric oxide was abolished by KT 5823, an inhibitor of cyclic GMP-dependent protein kinase activity and by LY83583, an inhibitor of soluble guanylate cyclase activity. The study provides the first direct evidence of nitric oxide production from enteric ganglia.

Topics: Alkaloids; Aminoquinolines; Animals; Arginine; Carbazoles; Citrulline; Dimethylphenylpiperazinium Iodide; Ganglia, Autonomic; Guinea Pigs; Hexamethonium; Hexamethonium Compounds; Ileum; In Vitro Techniques; Indoles; Kinetics; Muscle, Smooth; Myenteric Plexus; Nitric Oxide; Nitroarginine; Protein Kinase Inhibitors; SRS-A; Tetrodotoxin; Tritium; Vasoactive Intestinal Peptide

The mechanism of action of vasoactive intestinal peptide (VIP) was examined in isolated gastric and taenia coli muscle cells and compared with that of nitric oxide (NO), sodium nitroprusside (SNP), and isoproterenol. In gastric muscle cells, VIP stimulated NO production, increased adenosine 3',5'-cyclic monophosphate (cAMP) and guanosine 3',5'-cyclic monophosphate (cGMP) levels, and induced relaxation in a concentration-dependent fashion. The NO synthase inhibitor NG-nitro-L-arginine abolished NO and cGMP production and partly inhibited relaxation. The soluble guanylate cyclase inhibitor LY 83583 abolished cGMP production and partly inhibited relaxation. (R)-p-adenosine 3',5'-cyclic phosphorothioate [(R)-p-cAMPS], a preferential inhibitor of cAMP-dependent protein kinase (cAK), and KT5823, a preferential inhibitor of cGMP-dependent protein kinase (cGK), partly inhibited relaxation separately and abolished relaxation in combination. The pattern implied that VIP induced relaxation by activation of cAK and by NO-mediated stimulation of cGMP and activation of cGK. In taenia coli muscle cells, VIP did not increase NO production or cGMP levels: relaxation was accompanied by an increase in cAMP and was partly inhibited by (R)-p-cAMPS and KT5823 and abolished by a combination of both inhibitors. Isoproterenol increased only cAMP levels in both cell types, which induced relaxation by activating cAK at low concentrations of agonist and both cAK and cGK at high concentrations in a pattern identical to that observed with VIP in taenia coli muscle cells. SNP and NO increased only cGMP levels in both cell types, which induced relaxation by activating cGK only. We conclude that cAK and cGK can be activated separately and mediate relaxation independently.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Aminoquinolines; Animals; Arginine; Cell Separation; Colon; Cyclic AMP; Cyclic GMP; Digestive System; Digestive System Physiological Phenomena; Enzyme Activation; Gastric Fundus; Guinea Pigs; Isoproterenol; Muscle Relaxation; Muscles; Nitrates; Nitric Acid; Nitroarginine; Nitroprusside; Protein Kinase C; SRS-A; Vasoactive Intestinal Peptide