Abstract 15211: Calcium/Calmodulin-Dependent Protein Kinase Kinase β Plays a Key Role for the Diverse Functions of Endothelial Nitric Oxide Synthases System between Conduit and Resistance Arteries in Mice
Background: We have previously demonstrated in animals and humans that endothelium-derived hydrogen peroxide (H2O2) is an endothelium-derived hyperpolarizing factor (EDHF) and that endothelial NO synthase (eNOS)-derived superoxide anions is a major precursor of EDHF/H2O2. We also have recently demonstrated in mice deficient of the whole NOSs system that endothelial NOSs system plays diverse roles depending on vessel size, mainly serving as a superoxide generating system to cause EDHF/H2O2 responses in resistance arteries (e.g. mesenteric arteries, MA) while serving as a NO generating system in conduit arteries (e.g. the aorta, Ao). In this study, we aimed to elucidate the mechanisms involved in the diverse roles of endothelial NOSs system in mice.
Methods and Results: We used male wild-type mice (10–16 weeks-old) (n=4∼6 in all protocols). Isometric tensions were recorded in organ chamber experiments. Since we previously reported that pathological eNOS uncoupling is not the mechanism for the divergent roles of eNOS between Ao and MA, we further examined this point. The extent of eNOS phosphorylation at Ser1177 (stimulatory site) was significantly greater in Ao than in MA, whereas that at Thr495 (inhibitory site) was significantly greater in MA than in Ao. STO-609 (5 μM for 1 hour), an inhibitor of calcium/calmodulin-dependent protein kinase kinase β (CaMKKβ) that lies upstream of eNOS Ser1177 phosphorylation pathway, significantly decreased EDHF-mediated relaxations and hyperpolarizations to acetylcoline (ACh) in MA but not in Ao. STO-609 had no effects on contractions or endothelium-independent relaxations. In response to ACh, eNOS Ser1177 phosphorylation was enhanced in both MA and Ao, which was inhibited by STO-609 in MA more than in Ao. The extent of eNOS Thr495 phosphorylation was unchanged by STO-609 in MA or Ao. The AMP-activated protein kinase (AMPK) inhibitor compound C and the phosphoinositide 3-kinase (PI3K) inhibitor wortmannin had no effects on EDHF-mediated relaxations or hyperpolarizations to ACh in MA and Ao.
Conclusions: These results suggest that eNOS activity is significantly attenuated in MA and that CaMKKβ plays a key role for the diverse functions of endothelial NOSs system between conduit and resistance arteries.
- © 2010 by American Heart Association, Inc.