Characterization of ERK1/2 signalling pathways induced by adenosine receptor subtypes in newborn rat cardiomyocytes

Abstract

1. Adenosine A 1, A 2A, and A 3 receptors (ARs) and extracellular signal-regulated kinase 1/2 (ERK1/2) play a major role in myocardium protection from ischaemic injury. In this study, we have characterized the adenosine receptor subtypes involved in ERK1/2 activation in newborn rat cardiomyocytes. 2. Adenosine (nonselective agonist), CPA (A 1), CGS 21680 (A 2A) or Cl-IB-MECA (A 3), all increased ERK1/2 phosphorylation in a time- and dose-dependent manner. The combined maximal response of the selective agonists was similar to adenosine alone. Theophylline (nonselective antagonist) inhibited completely adenosine-mediated ERK1/2 activation, whereas a partial inhibition was obtained with DPCPX (A 1), ZM 241385 (A 2A), and MRS 1220 (A 3). 3. PD 98059 (MEK1; ERK kinase inhibitor) abolished all agonist-mediated ERK1/2 phosphorylation. Pertussis toxin (PTX, G i/o blocker) inhibited completely CPA- and partially adenosine- and Cl-IB-MECA-induced ERK1/2 activation. Genistein (tyrosine kinase inhibitor) and Ro 318220 (protein kinase C, PKC inhibitor) partially reduced adenosine, CPA and Cl-IB-MECA responses, without any effect on CGS 21680-induced ERK1/2 phosphorylation. H89 (protein kinase A, PKA inhibitor) abolished completely CGS 21680 and partially adenosine and Cl-IB-MECA responses, without any effect on CPA response. 4. Cl-IB-MECA-mediated increases in cAMP accumulation suggest that A 3AR-induced ERK1/2 phosphorylation involves adenylyl cyclase activation via phospholipase C (PLC) and PKC stimulation. 5. In summary, we have shown that ERK1/2 activation by adenosine in cardiomyocytes results from an additive stimulation of A 1, A 2A, and A 3ARs, which involves G i/o proteins, PKC, and tyrosine kinase for A 1 and A 3ARs, and Gs and PKA for A 2AARs. Moreover, the A 3AR response also involves a cAMP/PKA pathway via PKC activation.