Protective effect of M₃ receptor on H₂O₂-induced apoptosis of rat myocardial cells in vitro

AimTo observe the effect of activation of M₃ receptor on H₂O₂ induced apoptosis in cultured rat myocytes and to investigate its possible mechanisms. MethodsIsolated neonatal cardiomyocytes were cultured. Morphologic changes were observed by microscopy. The apoptosis in cardiomyocyte was detected by terminal deoxynucleotide transferase directed d-UTP nick and end labeling (TUNEL) assay. The expression of apoptosis-related protein in Bcl-2 and Fas was measured by immunohistochemistry assay. ［Ca²⁺］_i in single cardiomyocyte loaded with Fluo 3-AM was measured by confocal microscope. ResultsH₂O₂-mediated myocyte apoptosis was attenuated by M₃ receptor agonist choline (10 mmol·L^-1). Pretreatment of cardiac myocytes with choline also increased Bcl-2, decreased Fas expression, and inhibited the increase in FI value of ［Ca²⁺］_i in H₂O₂-stimulated cardiac myocytes. However, blockade of M₃ receptor by 4DAMP (10 nmol·L^-1) completely inhibited the effects of choline on H₂O₂-stimulated cardiac myocytes. ConclusionActivation of M₃ receptor showed protective effect on H₂O₂-induced apoptosis in cultured rat myocytes and this effect might be related to modulating the expression of some genes including Bcl-2 and Fas as well as the downregulation of ［Ca²⁺］_i.