Induction of apoptosis in rat cardiocytes by A(3) adenosine receptor activation and its suppression by isoproterenol

The purpose of the present study was to investigate the mechanisms involved in the induction of apoptosis in newborn cultured cardiomyocytes by activa tion of adenosine (ADO) A(3) receptors and to examine the protective effect s of beta-adrenoceptors, The selective agonist for A(3) ADO receptors Cl-IB -MECA (2-chloro-N-6-iodobenzyl-5-N-methylcarboxamidoadenosine) and the anta gonist MRS1523 (5-propyl-2-ethyl-4-propyl-3-(ethylsulfanylcarbonyl)-6-pheny lpyridin-5-carboxylate) were used. High concentrations of the Cl-IBMECA (gr eater than or equal to 10 mu M) agonist induced morphological modifications of myogenic cells, such as rounding and retraction of cell body and dissol ution of contractile filaments, followed by apoptotic death. In addition, C l-IB-MECA caused a sustained and reversible increase in [Ca2+](i), which wa s prevented by the selective antagonist MRS1523. Furthermore, MRS1523 prote cted the cardiocytes if briefly exposed to CI-IBMECA and partially protecte d from prolonged (48 h) agonist exposure. Apoptosis induced by Cl-IB-MECA w as not redox-dependent, since the mitochondrial membrane potential remained constant until the terminal stage of cell death. Cl-IB-MECA activated casp ase-3 protease in a concentration-dependent manner after 7 h of treatment a nd more effectively after 18 h of exposure. Bcl-2 protein was readily detec ted in control cells, and its expression was significantly decreased after 24 and 48 h of treatment with Cl-IB-MECA. beta-Adrenergic stimulation antag onized the pro-apoptotic effects of Cl-IB-MECA, probably through a cAMP/pro tein kinase A-independent mechanism, since addition of dibutyryl-cAMP did n ot abolish the apoptosis induced by Cl-IB-MECA. Incubation of cultured myoc ytes with isoproterenol (5 mu M) for 3 or 24 h almost completely abolished the increase in [Ca2+]i. Prolonged incubation of cardiomyocytes with isopro terenol and Cl-IB-MECA did not induce apoptosis, Our data suggest that the apoptosis-inducing signal from activation of adenosine A(3) receptors (or c ounteracting beta-adrenergic signal) leads to the activation of the G-prote in-coupled enzymes and downstream pathways to a self-amplifying cascade. Ex pression of different genes within this cascade is responsible for orchestr ating either cardiomyocyte apoptosis or its protection. (C) 2000 Academic P ress.