Immunohistochemical and functional studies for M3 muscarinic receptors and cyclo-oxygenase-2 expressed in the mouse atrium
- In mouse atrium, M2 and M3 muscarinic receptors (M2R and M3R) are involved in biphasic (negative and positive) inotropic actions of muscarinic agonists, and the positive inotropic action is reduced by indomethacin. The aim of our study was to determine the localization of M2R, M3R and cyclo-oxygenase (COX) in mouse atrium and to characterize muscarinic receptor-mediated positive inotropy.
- M2R immunoreactivity was found only on atrial myocardium, but M3R immunoreactivity was localized on both the myocardium and endocardial endothelium. COX-1 and COX-2 immunoreactivities were identified in both myocardial and endocardial endothelium.
- In electrically stimulated left atria, carbachol caused M2R-mediated negative inotropy followed by M3R-mediated positive inotropy. Removal of atrial endothelium reduced the positive inotropy without affecting the negative inotropy, suggesting that stimulation of endothelial M3R mediates the positive inotropy.
- N-[2-(cyclohexyloxy)-4-nitrophenyl]-methanesulfonamide (NS398, COX-2 inhibitor) decreased the carbachol-induced positive inotropy; however, 5-(4-chlorophenyl)-1-(4-methoxyphenyl)-3-trifluoromethylpyrazole (SC560, COX-1 inhibitor), 1-[[4,5-bis(4-methoxyphenyl)-2-thiazolyl]carbonyl]-4-methylpiperazine (FR122047, COX-1 inhibitor) and l-nitroarginine methylester did not affect the inotropic response.
- M3R activation caused positive chronotropy in spontaneously beating right atria when M2R-mediated negative chronotropy was suppressed and rate of contraction was low, <350 beats min−1.
- Our results indicate that although M3Rs are located on both myocardial cells and endocardial endothelial cells, only endothelial M3Rs mediate positive inotropy in response to muscarinic agonists via activation of COX-2 in the mouse atrium. M3R-mediated positive chronotropy counteracting M2R-mediated negative chronotropy was also demonstrated.