In the isolated right atrium
In the isolated right atrium, activation of muscarinic receptors and adenosine receptors may block the generation of action potentials in the sinoatrial node, inducing cardiac arrest (Fig. 1, Fig. 2, Fig. 3, Fig. 4, Fig. 5) (Camara et al., 2015, Campbell et al., 1989). Physiologically, this condition can be observed after a massive release of Carboplatin australia or an exacerbated discharge of noradrenaline and ATP (Erlinge et al., 2005) which is converted to adenosine (Scamps and Vassort, 1990). In the clinic, cardiac arrest can be reversed by the use of a defibrillator. However, a specialized health professional is required to perform the procedure. In addition, electrical pulses of the defibrillator may injure cardiomyocytes (de Oliveira et al., 2008). Thus other interventional maneuvers, such as pharmacological treatment, are relevant therapeutic proposals to reverse cardiac arrest (Li et al., 2015, Ottani et al., 2014). To simulate in vitro what could occur in a situation of cardiac arrest, as described by some authors (Bohm et al., 1984, Bruckner et al., 1985, Jochem and Nawrath, 1983), we blocked spontaneous right atrial contractions by adding high concentrations of the endogenous agonists adenosine, ATP and acetylcholine to the organ bath. We observed that cardiac arrest induced by acetylcholine was spontaneously reversed, probably due to the action of cholinesterase in the isolated tissue (Blinks and Plummer, 1966). Therefore we decided to use carbachol as a drug to model the cardiac arrest induced by muscarinic receptors. In the rat right atrium, the negative chronotropic effect of adenosine and ATP is mediated by A1 adenosine receptors (Camara et al., 2015) while carbachol produces an inhibitory effect via muscarinic receptors (Dhein et al., 2001, Hammer and Giachetti, 1982). The A1 adenosine receptor competitive antagonist DPCPX reversed cardiac arrest induced by the three tested different adenosine receptor agonists (i.e. adenosine - Fig. 1; ATP – Supplementary Fig. 2; and CPA – Fig. 2). In addition, the muscarinic receptor antagonist atropine reversed cardiac arrest induced by carbachol (Fig. 4). Collectively, these data support the hypothesis that competing for the receptor binding site is an alternative to restore atrial contractions in receptor-mediated cardiac arrest. At equilibrium, the affinity of DPCPX for the A1 adenosine receptor is at the nanomolar range (Camara et al., 2015, Weyler et al., 2006); however, reversal of cardiac arrest by DPCPX was observed only at micromolar concentrations of the antagonist. This could be explained by the applied experimental setting, in which the antagonists were exposed at most for 10min, preventing equilibrium to be reached. In addition, the dependence on high concentrations of DPCPX to reverse cardiac arrest raised the possibility that secondary effects were contributing to the effect of the antagonist. To test this hypothesis we investigated whether DPCPX would be able to reverse the cardiac arrest induced by carbachol. DPCPX reversed the cardiac arrest induced by the muscarinic agonist (Fig. 3), demonstrating that this drug can restore spontaneous contractions independently of the canonical A1 adenosine receptor antagonism. Structurally, DPCPX share a xanthine core with some PDE inhibitors, for example IBMX (Beavo et al., 1970, Essayan, 2001, Lohse et al., 1987). Based on this structural similarity, we hypothesized that phosphodiesterase inhibition could well be the secondary mechanism mediating the reversal of carbachol-induced cardiac arrest by DPCPX. Therefore, we assessed whether phosphodiesterase inhibition by IBMX could reproduce the results obtained with DPCPX. We observed that IBMX also reversed the cardiac arrest induced by adenosine, CPA and carbachol. Besides that, enzymatic inhibition of PDE IV by DPCPX was observed in rat ventricular preparations (Ukena et al., 1993). Also, in addition to other xanthines, DPCPX inhibits PDE I to III in functional assays in the guinea pig left atrium at high concentrations (von der Leyen et al., 1989). These two works corroborate our hypothesis that inhibition of PDE is the secondary mechanism of DPCPX.