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Increasing resting tension enhances cGMP production in normotensive and hypertensive rat aortas
Author(s) -
Silva Bruno R,
Grando Marcella Daruge,
Bendhack Lusiane Maria
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.1017.15
Subject(s) - enos , phosphorylation , medicine , soluble guanylyl cyclase , phenylephrine , endocrinology , extracellular , chemistry , western blot , aorta , calmodulin , guanylate cyclase , calcium , nitric oxide synthase , nitric oxide , blood pressure , biochemistry , gene
NO‐Synthase (NOS) is a Ca 2+ ‐Calmodulin‐dependent enzyme that is activated by phosphorylation of Ser 1177 (P‐Ser) producing NO and inactivated by phosphorylation of its site Thr 495 (P‐Thr). The main target in NO signaling is soluble guanylyl‐cyclase (sGC) and the production of cGMP. Shear stress‐mediated phosphorylation of NOS regulates NOS activity. However, the signaling in renal hypertensive (2K‐1C) rat aorta submitted to increasing resting tension (RT) is not known. Our hypothesis is that RT can activate the endothelial NOS (eNOS) increasing NO production and consequently cGMP in normotensive (2K) and 2K‐1C rat aortas. This study aimed to evaluate the effects of RT of 1.5 g (control) and 3.0 g on the eNOS phosphorylation in phenylephrine (PE)‐induced contractile response in isolated aorta from 2K‐1C and 2K rats. Concentration‐effect curves for PE and for extracellular CaCl 2 were constructed in aortas under resting tension of 1.5 g and 3.0 g in the absence or presence of NOS inhibitor (L‐NAME, 100 μM). The expression of P‐Ser and P‐Thr were evaluated by Western Blot. The cGMP was measured by ELISA‐Immunoassay. This study was performed in accordance with the Ethics Committee CEUA‐USP (15.1.477.60.1). PE‐induced maximum contractile effect (ME) on RT of 1.5 g was lower in 2K‐1C (1.2 ± 0.2 g, n=7; p<0.001) than in normotensive 2K (2.2 ± 0.1g; n=5), which effects were normalized by NOS inhibition with L‐NAME. PE‐induced contractile response was similar between 2K‐1C and 2K under RT of 3.0 g. Similar responses were obtained for CaCl 2 . However, NOS inhibition increased the contractile response induced by CaCl 2 in RT‐dependent way only in 2K‐1C (1.5 g: 0.72 ± 0.09 g; 3.0 g: 1.03 ± 0.10 g; n=4; p<0.05). P‐Ser 1177 expression stimulated by PE was higher in 2K‐1C than in 2K aortas. Increasing of RT from 1.5 g to 3.0 g decreased P‐Ser and increased P‐Thr in 2K aorta, but it was not altered in 2K‐1C. Increasing RT from 1.5 g to 3.0 g increased cGMP production in 2K (1.5 g: 4.4 ± 0.7 pmol/ml; 3.0 g: 7.1 ± 1.0 pmol/ml; n=5; p<0.05) and 2K‐1C (1.5 g: 2.5 ± 0.5 pmol/ml; 3.0 g: 4.9 ± 0.5 pmol/ml; n=5; p<0.05), which was higher in 2K (p<0.05) than in 2K‐1C. Taken together, our results demonstrate that the anti‐contractile effect induced by PE in 2K‐1C rat aortas under 1.5 g tension is due to NOS activation by calcium transient and higher P‐Ser 1177 . The anti‐contractile effect induced by PE in 2K‐1C aorta is abolished by increasing RT to 3.0 g. The increased RT in 2K‐1C aortas to 3.0 g also increases cGMP production despite of this production to stay lower than in 2K rat aortas. Support or Funding Information Supported by FAPESP, CNPq and CAPES