The PKC activator PMA preconditions rabbit heart in the presence of adenosine receptor blockade:: Is 5′-nucleotidase important?

While there is good evidence that both protein kinase C (PKC) and adenosine are involved in ischemic preconditioning, their sequence in the intracellular signaling cascade is in dispute. One hypothesis proposes that PKC activation causes release of adenosine which then protects the heart, while the other proposes that adenosine stimulates PKC which in turn causes protection. Accordingly, we studied the effects of specified sequences of pharmacologic triggers and blockers on the infarct-sparing effect of a preconditioning protocol. The combination of the adenosine receptor agonist R(-)N-6-(2-phenylisopropyl) adenosine (PIA) and the PKC blocker chelerythrine would be protective only if the first hypothesis were correct. On the other hand, the combination of the adenosine receptor blocker 8-(p-sulfophenyl) theophylline (SPT) and a PKC activator would be protective only if the second hypothesis were correct. Isolated, Krebs-perfused rabbit hearts experienced 30 min of regional ischemia and 2 h of reperfusion. Infarct size was quantitated by triphenyltetrazolium chloride staining. In untreated control hearts, 30.0+/-2.7% of the risk zone infarcted, Fifty nmol/l PIA for 20 min starting 10 min prior to ischemia resulted in only 8.4+/-1.9% infarction (P<0.01), while the combination of PIA and 5 mu mol/l chelerythrine resulted in large infarcts of 27.8 +/- 3.2%. This attenuation of the protective effect continued to be observed even when the PIA infusion was continued to the end of the reperfusion period. Conversely 0.2 nmol of the PKC activator phorbol 12-myristate 13-acetate (PMA) infused during the 10-min interval prior to ischemia protected the hearts (6.5 +/- 1.3% infarction, P<0.01 v control). And protection persisted when PMA-treated hearts were also exposed to 100 mu M SPT for 35 min starting 5 min prior to ischemia (9.5 +/- 1.9% infarction, P<0.01 v control). When PKC activation by the PKC-coupled agonist phenylephrine was continued to the end of ischemia and adenosine blockade was extended throughout the reperfusion period by prolonged infusion of SPT, protection was unaffected. The administration of either SPT or chelerythrine alone did not confer any protection (32.5 +/- 3.3 and 34.0 +/- 3.2% infarction, respectively). Thus, because the combination of PKC activation and adenosine receptor blockade was protective while that of adenosine receptor agonist and PKC blockade was not, adenosine receptors must be upstream of PKC in preconditioning. (C) 1998 Academic Press.