Journal
BRITISH JOURNAL OF PHARMACOLOGY
Volume 143, Issue 3, Pages 397-403Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/sj.bjp.0705933
Keywords
Akt; beta(2)-adrenoceptors; endothelium; nitric oxide; phosphatidylinositol 3-kinase; protein kinase A; rat aorta
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1 Vasorelaxation to beta(2)-adrenoceptor stimulation occurs through both endothelium-dependent and endothelium-independent mechanisms, and the former is mediated through Ca2+-independent activation of endothelial-type nitric oxide synthase (NOS-3). Since Ca2+-independent NOS-3 activation may occur through its serine phosphorylation via protein kinase A (PKA) or Akt, we determined the PKA and Akt dependency of beta(2)-adrenergic relaxation of rat aorta. 2 Rat aortic rings were pre-incubated with the PKA inhibitor H-89 (10(-7) M), the phosphatidylinositol 3-kinase (PI3K) inhibitor wortmannin (5 x 10(-7) M), Akt inhibitor (10(-5) M), or vehicle, in the absence or presence of the NOS inhibitor N-G-nitro-L-arginine methyl ester (L-NAME, 10(-4) M). Rings were then contracted with phenylephrine (10(-7) M), and concentration - relaxation responses determined to the beta(2)-adrenoceptor agonist albuterol. 3 Rings exhibited a concentration-dependent relaxation to albuterol: pEC(50) 6.9 +/- 0.2, E-max 88.2 +/- 4.0%. L-NAME attenuated E-max to 60.2 +/- 3.5% (P<0.001). 4 In the presence of L-NAME, wortmannin or Akt inhibitor did not influence albuterol responses, whereas H-89 reduced E-max further, to 27.5 +/- 2.2% (P<0.001). 5 In the absence of L-NAME, E-max to albuterol was reduced by H-89, wortmannin or Akt inhibitor, to 56.2 +/- 2.2, 56.0 +/- 1.6 and 55.4 +/- 1.8%, respectively (P<0.001 for each); the combinations H-89 plus wortmannin or H-89 plus Akt inhibitor reduced Emax further still. 6 Western blotting of NOS-3 immunoprecipitates from rat aortas confirmed that albuterol increased serine phosphorylation of NOS-3, and this increase was attenuated by H-89 or Akt inhibitor. 7 Our results indicate that beta(2)-adrenoceptor stimulation relaxes rat aorta through both NO-dependent and independent mechanisms. The latter is predominantly PKA-mediated, whereas the former occurs through both PKA and PI3K/Akt activation.
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