BETA(2)-ADRENOCEPTOR ACTIVATION BY ZINTEROL CAUSES PROTEIN-PHOSPHORYLATION, CONTRACTILE EFFECTS AND RELAXANT EFFECTS THROUGH A CAMP PATHWAYIN HUMAN ATRIUM

Evidence from ventricular preparations of cat, sheep, rat and dog sugg ests that both beta(1)-adrenoceptors (beta(1)AR) and beta(2)-adrenocep tors (beta(2)AR) mediate positive inotropic effects but that only beta (1)AR do it through activation of a cAMP pathway. On the other hand, o ur evidence has shown that both beta(1)AR and beta(2)AR hasten relaxat ion of isolated human myocardium consistent with a common cAMP pathway . We have now investigated in the isolated human right atrial appendag e, a tissue whose beta-AR comprise around 2/3 of beta(1)AR and 1/3 of beta(2)AR, whether or not beta(2)AR-mediated effects occur via activat ion of a cAMP pathway. We carried out experiments on atria obtained fr om patients without advanced heart failure undergoing open heart surge ry. To activate beta(2)AR, we used the beta(2)AR-selective ligand zint erol. Experiments were carried out on paced atrial strips (1 Hz) and t issue homogenates and membrane particles. Zinterol caused positive ino tropic and lusitropic (i.e. reduction of t(1/2) of relaxation) effects with EC(50) values of 3 and 2 nM, respectively. The zinterol-evoked e ffects were unaffected by the beta(1)AR-selective antagonist CGP 20712 A (300 nM) but blocked surmountably by the beta(2)AR-selective antagon ist ICI 118551 (50 nM) which reduced both EC(50) values to 1 mu M. Zin terol stimulated adenylyl cyclase activity with an EC(50) of 30 nM and intrinsic activity of 0.75 with respect to (-)-isoprenaline (600 mu M ); the effects were resistant to blockade by CGP 20712A (300 nM) but a ntagonised surmountably by ICI 118551 (50 nM). Zinterol bound to membr ane beta AR labelled with (-)-[I-125] cyanopindolol with higher affini ty for beta(2)AR than for beta(1)AR; the binding to beta(2)AR but not to beta(1)AR was reduced by GTP gamma S (10 mu M). In the presence of CGP 20712A (300 nM) (-)-isoprenaline (400 mu M) (to activate both beta (1)AR and beta(2)AR maximally) and zinterol (10 mu M) increased contra ctile force 3.4-fold and 2.5-fold respectively and reduced relaxation t(1/2) by 32% and 18% respectively. These effects of (-)-isoprenaline and zinterol were associated (5 min incubation) with phosphorylation ( pmol P/mg supernatant protein) of troponin I and C-protein to values o f 8.4 +/- 2.0 vs 12.4 +/- 2.3 and 10.1 +/- 2.5 vs 8.6 +/- 1.6 respecti vely. (-)-Isoprenaline and zinterol also caused phosphorylation of pho spholamban (1.8 +/- 0.3 vs 0.4 +/- 0.1 pmol P/mg respectively) specifi cally at serine residues. We conclude that in human atrial myocardium activation of both beta(1)AR and beta(2)AR leads to cAMP-dependent pho sphorylation of proteins involved in augmenting both contractility and relaxation.