Although
protein kinase C (PKC) plays a key role in ischemic preconditioning (IPC), the actual mechanism of that protection is unknown. We recently found that protection from IPC requires activation of
adenosine receptors during early reperfusion. We, therefore, hypothesized that PKC might act to increase the heart's sensitivity to
adenosine. IPC limited
infarct size in isolated rabbit hearts subjected to 30-min regional
ischemia/2-h reperfusion and IPC's protection was blocked by the PKC inhibitor
chelerythrine given during early reperfusion revealing involvement of PKC at reperfusion. Similarly
chelerythrine infused in the early reperfusion period blocked the increased phosphorylation of the protective
kinases Akt and ERK1/2 observed after IPC. Infusing
phorbol 12-myristate 13-acetate (PMA), a PKC activator, during early reperfusion mimicked IPC's protection. As expected, the protection triggered by PMA at reperfusion was blocked by
chelerythrine, but surprisingly it was also blocked by
MRS1754, an
adenosine A(2b) receptor-selective antagonist, suggesting that PKC was somehow facilitating signaling from the A(2b) receptors.
NECA [5'-(N-ethylcarboxamido)
adenosine], a potent but not selective A(2b) receptor agonist, increased phosphorylation of Akt and ERK1/2 in a dose-dependent manner. Pretreating hearts with PMA or brief preconditioning
ischemia had no effect on phosphorylation of Akt or ERK1/2 per se but markedly lowered the threshold for
NECA to induce their phosphorylation.
BAY 60-6583, a highly selective A(2b) agonist, also caused phosphorylation of ERK1/2 and Akt.
MRS1754 prevented phosphorylation induced by
BAY 60-6583.
BAY 60-6583 limited
infarct size when given to ischemic hearts at reperfusion. These results suggest that activation of cardiac A(2b) receptors at reperfusion is protective, but because of the very low affinity of the receptors endogenous cardiac
adenosine is unable to trigger their signaling. We propose that the key protective event in IPC occurs when PKC increases the heart's sensitivity to
adenosine so that endogenous
adenosine can activate A(2b)-dependent signaling.