Abstract:
Differential regulation of muscarinic M2 and M3
receptor signaling in gastrointestinal smooth muscle by caveolin-1. Am J
Physiol Cell Physiol 305: C334?C347, 2013. First published June 19,
2013; doi:10.1152/ajpcell.00334.2012.?Caveolae act as scaffolding proteins
for several G protein-coupled receptor signaling molecules to
regulate their activity. Caveolin-1, the predominant isoform in smooth
muscle, drives the formation of caveolae. The precise role of caveolin-1
and caveolae as scaffolds for G protein-coupled receptor signaling and
contraction in gastrointestinal muscle is unclear. Thus the aim of this
study was to examine the role of caveolin-1 in the regulation of Gq- and
Gi-coupled receptor signaling. RT-PCR, Western blot, and radioligandbinding
studies demonstrated the selective expression of M2 and M3
receptors in gastric smooth muscle cells. Carbachol (CCh) stimulated phosphatidylinositol
(PI) hydrolysis, Rho kinase and zipper-interacting protein
(ZIP) kinase activity, induced myosin phosphatase 1 (MYPT1) phosphorylation
(at Thr696) and 20-kDa myosin light chain (MLC20) phosphorylation
(at Ser19) and muscle contraction, and inhibited cAMP formation.
Stimulation of PI hydrolysis, Rho kinase, and ZIP kinase activity, phosphorylation
of MYPT1 and MLC20, and muscle contraction in response
to CCh were attenuated by methyl -cyclodextrin (MCD) or caveolin-1
small interfering RNA (siRNA). Similar inhibition of PI hydrolysis, Rho
kinase, and ZIP kinase activity and muscle contraction in response to
CCh and gastric emptying in vivo was obtained in caveolin-1-knockout
mice compared with wild-type mice. Agonist-induced internalization of
M2, but not M3, receptors was blocked by MCD or caveolin-1 siRNA.
Stimulation of PI hydrolysis, Rho kinase, and ZIP kinase activities in
response to other Gq-coupled receptor agonists such as histamine and
substance P was also attenuated by MCD or caveolin-1 siRNA. Taken
together, these results suggest t