title: Cyclic GMP regulates M₃AChR activity at plasma membranes from airway smooth muscle.
abstract: Muscarinic acetylcholine receptors MAChRs from Bovine Tracheal Smooth Muscle (BTSM) plasma membranes are responsible for the cGMP rise and signal-amplitude peaks associated with smooth muscle contraction present in bronchial asthma. These MAChRs bind [(3)H]QNB and exhibit the classic G Protein Coupled-Receptor (GPCR) behavior towards muscarinic agonist and antagonists that is sensitive to sensitive to GTP analogs. Interestingly, the [(3)H]QNB binding activity was stimulated by cGMP and ATP, and was enhanced by IBMX and Zaprinast, inhibitors of cGMP-PDE. Cyclic GMP plus ATP affected the agonist-antagonist muscarinic binding activities. Thus, the high affinity agonist (Carbamylcholine) binding sites disappeared, whereas, 4-DAMP, a M3 selective antagonist displayed an additional high affinity-binding site. In contrast, non-selective (atropine) and M2-selective (methoctramine and gallamine) antagonists revealed one low binding site. Moreover, the 4-DAMP-mustard alkylation of the MAChRs blocked the cGMP effect indicating that the M3AChR is the main receptor target of cGMP. Interestingly, these cGMP effects were potentiated by an activator (Sp-8-pCPT-cGMPS), and diminished by an inhibitor (Rp-8-pCPT-CGMPS), of cGMP-dependent protein kinase (PKG-II), which was detected by Western blotting using specific PKG II antibodies. Finally, plasma membrane M3AChRs were phosphorylated in a cGMP-dependent manner and this novel post-translational reversible modification at M3AChRs may act as a feedback mechanism to terminate the cGMP dependent muscarinic signal transduction cascades at the sarcolema of BTSM.
authors.affiliation: Sección de Biomembranas, Instituto de Medicina Experimental (IME), Facultad de Medicina, Universidad Central de Venezuela (UCV) , Caracas , Venezuela.
authors.forename: Marcelo J
authors.initials: MJ
authors.lastname: Alfonzo
authors.forename: Ramona González
authors.initials: RG
authors.lastname: De Alfonzo
authors.forename: Marcelo A
authors.initials: MA
authors.lastname: Alfonzo-González
authors.forename: Itala Lippo
authors.initials: IL
authors.lastname: De Becemberg
mesh_terms: D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D006152:Cyclic GMP / Q000378:metabolism*; D062967:Cyclic GMP-Dependent Protein Kinase Type II / Q000276:immunology; D025461:Feedback, Physiological; D018721:Muscarinic Agonists / Q000378:metabolism; D018727:Muscarinic Antagonists / Q000378:metabolism; D009130:Muscle, Smooth / Q000378:metabolism*; D010880:Piperidines / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011813:Quinuclidinyl Benzilate / Q000378:metabolism; D011976:Receptors, Muscarinic / Q000378:metabolism*; D015398:Signal Transduction; D014132:Trachea / Q000378:metabolism*
chemical_list: D018721:Muscarinic Agonists; D018727:Muscarinic Antagonists; D010880:Piperidines; D011976:Receptors, Muscarinic; D011813:Quinuclidinyl Benzilate; C042375:4-diphenylacetoxy-1,1-dimethylpiperidinium; D062967:Cyclic GMP-Dependent Protein Kinase Type II; D006152:Cyclic GMP