Inhibition of nitric oxide synthesis increases adenosine production via an extracellular pathway through activation of protein kinase C

Background NO is known to deactivate protein kinase C (PKC). Because we have reported that the activation of PKC activates ecto-5'-nucleotidase, we examined whether the inhibition of NO synthesis increases ecto-5'-nucleotidase activity through the activation of PKC. Methods and Results The left anterior descending coronary artery (LAD) was cannulated and perfused with blood through a bypass tube from the left carotid artery in 65 open-chest dogs. The intracoronary administration of N-G-nitro-L-arginine methyl ester (L-NAME, 10 mu g.kg(-1).min(-1)), an NO synthase inhibitor, for 30 minutes increased (P<.05) adenosine levels in coronary venous blood (123+/-10 versus 21+/-3 pmol/mL) and ecto-5'-nucleotidase activity (64+/-6 versus 41+/-4 nmol.mg(-1).min(-1)) in the LAD-perfused myocardium, The intracoronary administration of alpha,beta-methyleneadenosine 5'-diphosphate, an inhibitor of ecto-5'-nucleotidase, or GF109203X or calphostin C, both of which are PKC Inhibitors, attenuated the L-NAME-induced increases in adenosine levels and ecto-5'-nucleotidase activity. Treatment of cultured human coronary arterial endothelial cells (HCAECs) with L-NAME: for 30 minutes increased ecto-5'-nucleotidase activity, which was inhibited by either GF109203X or calphostin C. NO releasers decreased both ecto-5'-nucleotidase and PKC activities in HCAECs. Treatment of HCAECs with zaprinast, a selective inhibitor of cGMP-specific phosphodiesterase, with or without atrial natriuretic peptide, increased intracellular cGMP concentrations but did not change ecto-5'-nucleotidase activity. Conclusions These results indicate that the inhibition of NO synthesis increases both adenosine production and ecto-5'-nucleotidase activity through the activation of PKC and that NO modulates ecto-5'-nucleotidase via cCMP-independent mechanisms.