Changes in extracellular pH and myocardial ischaemia alter the cardiac effects of diadenosine tetraphosphate and pentaphosphate

Abstract

1. The structural conformation of diadenosine tetraphosphate (Ap(4)A) and pentaphosphate (Ap(5)A) has been reported to alter as pH is reduced. As such, it is possible that the cardiac effects of Ap(4)A and Ap(5)A vary during acidosis and myocardial ischaemia due to changes in ligand structure, receptor proteins or intracellular signalling. 2. We investigated whether the cardiac electrophysiological and coronary vasomotor effects of Ap(4)A and Ap(5)A are preserved under conditions of extracellular acidosis (pH 6.5) and alkalosis (pH 8.5) and whether Ap(4)A has any electrophysiological or antiarrhythmic effects during ischaemia. 3. Transmembrane right ventricular action potentials, refractory periods and coronary perfusion pressure were recorded from isolated, Langendorff-perfused guinea-pig hearts under constant flow conditions. The effects of 1 nM and 1 microM Ap(4)A and Ap(5)A were studied at pH 7.4, 6.5 and 8.5. The effects of 1 microM Ap(4)A were studied during global low-flow ischaemia and reperfusion. 4. At pH 7.4, Ap(4)A and Ap(5)A increased action potential duration (APD(95)) and refractory period (RP) and reduced coronary perfusion pressure. The electrophysiological effects were absent at pH 6.5 while the reductions in perfusion pressure were attenuated. At pH 8.5, Ap(4)A increased RP but the effects of Ap(4)A and Ap(5)A on perfusion pressure were attenuated. During ischaemia, Ap(4)A had no antiarrhythmic or electrophysiological effects. 5. These data demonstrate the importance of extracellular pH in influencing the effects of Ap(4)A and Ap(5)A on the heart and indicate that any potentially cardioprotective effects of these compounds during normal perfusion at physiological pH are absent during ischaemia.

Figure 1

Effects of diadenosine tetraphosphate (Ap₄A) and pentaphosphate (Ap₅A) on coronary perfusion pressure at pH 7.4. (a) illustrates a typical recording (representative of five similar experiments which had similar results) from a heart receiving 1 nM Ap₄A followed by 1 μM. Exposure to 1 nM Ap₄A produced a transient reduction in perfusion pressure that recovered, while exposure to 1 μM Ap₄A produced a larger, maintained reduction in perfusion pressure. (b) illustrates the mean nadir responses of hearts to Ap₄A and Ap₅A at 1 nM and 1 μM. Data were obtained from six hearts for each compound and statistical significance determined by Bonferroni's Multiple Comparison test vs respective Ap_nA-free conditions (control).

Figure 2

Effect of diadenosine tetraphosphate (Ap₄A) and pentaphosphate (Ap₅A) on ventricular action potential duration and refractoriness at pH 7.4. (a) illustrates representative action potential recordings from hearts before and after 1 μM Ap₄A and Ap₅A (representative of five similar experiments with both compounds that had similar results). (b) and (c) illustrate the effects of Ap₄A and Ap₅A at concentrations of 1 nM and 1 μM on action potential duration measured at 95% repolarization (APD₉₅) and refractory period (RP), respectively. Data were obtained from six hearts for each compound and statistical significance determined by Dunnett's Multiple Comparison test vs respective Ap_nA-free conditions (control).

Figure 3

Effects of diadenosine tetraphosphate at 1 nM and 1 μM on action potential duration measured at 95% repolarization (APD₉₅, a), refractory period (RP, b), coronary perfusion pressure (c) and heart rate (in unpaced hearts, d) under alkalotic conditions (pH 8.5). Diadenosine tetraphosphate had no effect on APD₉₅ or RP at 1 nM under alkalotic conditions, but at 1 μM RP was increased. APD₉₅ tended to be increased by 1 μM diadenosine tetraphosphate, but this effect did not reach statistical significance (by Dunnett's Multiple Comparison test). Perfusion pressure and heart rate also tended to be reduced slightly by 1 μM diadenosine tetraphosphate but these changes did not reach statistical significance (by Bonferroni's and Dunnett's Multiple Comparison tests, respectively). Data were obtained from six hearts before (pH 8.5) and after admission of diadenosine tetraphosphate and the statistically significant difference indicated was made using Dunnett's test vs diadenosine tetraphosphate-free conditions (pH 8.5).

Figure 4

Effects of diadenosine pentaphosphate at 1 nM and 1 μM on action potential duration measured at 95% repolarization (APD₉₅, a), refractory period (RP, b), coronary perfusion pressure (c) and heart rate (in unpaced hearts, panel d) under alkalotic conditions (pH 8.5). Diadenosine pentaphosphate had no statistically significant effect on APD₉₅ or RP under alkalotic conditions (by Dunnett's Multiple Comparison test), but RP tended to be increased. Perfusion pressure and heart rate tended to be reduced slightly by 1 μM diadenosine pentaphosphate but these changes did not reach statistical significance (by Bonferroni's and Dunnett's Multiple Comparison tests, respectively). Data were obtained from six hearts before (pH 8.5) and after admission of diadenosine pentaphosphate.

Figure 5

Effects of diadenosine tetraphosphate (Ap₄A) at 1 μM during myocardial ischaemia and reperfusion. (a) Illustrates the effects of Ap₄A on action potential duration measured at 95% repolarization (APD₉₅) and (b) illustrates the effects on refractory period. No statistically significant differences (by Bonferroni's Multiple Comparison test) were detected between the groups. (Refractory periods were not measured between 15 and 30 min of ischaemia due to the risk of triggering ventricular arrhythmias). (c) Illustrates the percentage incidence of ventricular tachycardia (VT) and fibrillation (VF) during ischaemia in control hearts and hearts receiving Ap₄A, together with the incidence of spontaneous recovery from these arrhythmias during reperfusion. No statistically significant differences were observed (by Fisher's exact test). (d) Illustrates the onset time of ventricular tachycardia during ischaemia in the two groups of hearts. No statistically significant differences (by Student's t-test) were observed. Thus, Ap₄A had no antiarrhythmic or electrophysiological effects during ischaemia and reperfusion. Interestingly, the presence of Ap₄A was associated with a trend for an earlier onset and greater incidence of arrhythmias during ischaemia together with recovery in fewer hearts. Data were obtained from 10 control (drug-free) hearts and 10 hearts which received 1 μM Ap₄A.