Involvement of K+ channels in adenosine A2A and A2B receptor-mediated hyperpolarization of porcine coronary artery endothelial cells

This study investigated the effects of the following adenosine agonists: 5'-ethylcarboxamidoadenosine (NECA), N-6-cyclopentyadenosine (CPA) 2- [p- (2-carboxyethyl)]phenylamino-5'N-ethylcarboxamidoadenosine (CGS-21680), and 2-chloroadenosine (CAD) and its antagonist, 4-(2-[7-amino-2-{2-furyl}]1,2,4-triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM-241385), a selective A(2A) adenosine receptor antagonist, and the involvement of the K-ATP(+), and K-Ca channels on the resting membrane potential (RMP) of confluent monolayers of cultured porcine coronary artery endothelial cells (PCAECs). Adenosine agonists and K-ATP(+) channel openers (pinacidil, cromakalim) hyperpolarized cultured PCAECs. The average RMP was -32.31 +/- 1.2 mV. Adenosine agonists at 10(-5) M caused a significant increase in RMP to -65.0 +/- 1.5 mV for CAD (a nonselective adenosine receptor agonist) to -75.9 +/- 1.6 mV for CGS-21680 (a selective A(2A) receptor agonist) and to -87.0 +/- 3.5 mV for NECA (a nonselective A(1)/A(2A)/A(2B) receptor agonist). Pinacidil and cromakalim at 10 muM increased the membrane potential to -76.2 +/- 1.2 mV and -75.22 +/- 0.12 mV, respectively. The hyperpolarization induced by adenosine receptor agonists and K-ATP openers was inhibited by an application of the K-ATP(+) channel blocker glibenclamide (10 muM), indicating the involvement of the K-ATP(+) channel in the adenosine-mediated hyperpolarization of PCAECs. Moreover, 1-EB10, a selective opener of the maxi-K-Ca channel, hyperpolarized PCAECs, and the effect of 1-EB10 was completely blocked by a selective, irreversible blocker of the high conductance K-Ca (maxi-K) channels (penitrem A), but it only partially blocked the effect of NECA. ZM-241385 has no effect on hyperpolarization elicited by K-ATP(+) and K-Ca channel openers. However, ZM-241385 significantly blocked the hyperpolarization effect of CAD and CGS-21680. ZM-241385 partially blocked the hyperpolarizing effect of NECA, and a combination of ZM-241385 and penitrem A further blocked the hyperpolarizing effect of NECA. These results further support the involvement of K+ channels in adenosine A(2A) and A(2B) receptor-mediated hyperpolarization of PCAECs.