Muscarinic agonists acting through M2 acetylcholine receptors stimulate the migration of an NO-sensitive guanylyl cyclase to the plasma membrane of bovine tracheal smooth muscle

Abstract

Muscarinic agonists acting on bovine tracheal smooth muscle (BTSM) induce two separate cGMP signals, one at 20 sec associated with NO-sensitive-soluble-guanylyl-cyclase (NO-sGC) and another at 60 sec, linked to natriuretic-peptide-GC. The 20-sec-cGMP novel cascade starts with mAChRs, via unknown components, activates an NO-sGC. To unravel this cascade, in crude membranes isolated from intact BTSM strips exposed to muscarinic agonists, we detected GC activities increments at 20 sec and 60 sec. The 20-sec-GC is a NO-sensitive-GC, identified as α₁β₁-heterodimer. In reconstitution experiments with purified plasma membranes and cytosol, muscarinic agonists induced an NO-sGC migration in a dose-dependent manner, being inhibited by muscarinic antagonists displaying an M₂AChR profile and blocked by PTX, suggesting the involvement of G_o/G_i proteins. The NO-sGC related to migration was isolated and identified as an α₁β₁-heterodimer. This work shows that muscarinic agonists in BTSM induce a massive and selective α₁β₁-NO-sGC migration from cytoplasm to plasma membranes being responsible for the 20-sec-cGMP signal.

Keywords::

• Tracheal smooth muscle
• cGMP
• NO-sensitive sGC
• Muscarinic receptors
• G proteins.

Acknowledgments

This work was supported by grants from the Consejo de Desarrollo Científico y Humanístico, Universidad Central de Venezuela (CDCH-UCV) PG-09-7401-2008/1 and PI 09.006464.2006/2 (to RGA), PI-09-6030-2007 (to ILB),and a grant from the Fondo Nacional de Ciencia y Tecnología (FONACIT) of Venezuela # S1-2002000411 (to ILB). The authors also thank Dr. Coral Wynter for the English version of this manuscript and Dr. Marcelo A. Alfonzo-González for theFigures style. During the submission of this manuscript, some reviewers’ comments were incorporated into themanuscript and we are grateful.

Declaration of interest

This work was supported by grants from the Consejo de Desarrollo Científico y Humanístico, Universidad Central de Venezuela (CDCH-UCV) PG-09-7401-2008/1 and PI 09.006464.2006/2 (to RGA), PI-09-6030-2007 (to ILB),and a grant from the Fondo Nacional de Ciencia y Tecnología (FONACIT) of Venezuela # S1-2002000411 (to ILB). The authors also thank Dr. Coral Wynter for the English version of this manuscript and Dr. Marcelo A. Alfonzo-González for theFigures style. During the submission of this manuscript, some reviewers’ comments were incorporated into themanuscript and we are grateful.