Abstract 2305: Endothelin-1 Alters Nitric Oxide-Dependent Cerebrovascular Responses by Modulating The Phosphorylation State of eNOS via Rho Kinase

Stroke ◽  
2012 ◽  
Vol 43 (suppl_1) ◽  
Author(s):  
Giuseppe Faraco ◽  
Joseph Anrather ◽  
Costantino Iadecola
Keyword(s):  
Nitric Oxide ◽  
Rho Kinase ◽  
Smooth Muscle Relaxation ◽  
No Production ◽  
Rock Inhibitor ◽  
No Donor ◽  
Doppler Flowmetry ◽  
Phosphorylation State ◽  
Endothelin 1 ◽  
Cranial Window

Hypertension (HTN) alters vital homeostatic mechanisms regulating cerebral blood flow (CBF) and increases the risk of stroke and dementia. HTN exerts some of its damaging effects by counteracting the beneficial vascular actions of nitric oxide (NO). The potent vasoconstrictor endothelin-1 (ET1) has been implicated in the pathogenesis of HTN, but its role in the cerebrovascular effects of HTN is unknown. We examined whether ET1 disrupts CBF regulation. CBF (laser-Doppler flowmetry) was assessed in the somatosensory cortex in anesthetized male C57Bl/6 mice (n=5/group) equipped with a cranial window. ET1 (35 pmol/kg/min; i.v. for 45 min) increased mean arterial pressure from 72±4 to 99±6 mmHg (p<0.05), without reducing resting CBF (p>0.05). However, ET1 attenuated the CBF increase produced by neocortical application of the endothelium-dependent vasodilator acetylcholine (ACh; -37±1%; p<0.05) and by whisker stimulation (-31±1%; p<0.05), responses dependent on NO. The CBF response to adenosine was intact (p>0.05) indicating that ET1 did not act by compromising smooth muscle relaxation. The effects of ET1 were prevented by the ET type A (ET A ) receptor antagonist BQ123 (1µM; p<0.05), by the Rho kinase (ROCK) inhibitor Y27632 (1 µM; p<0.05), but not by the ET B antagonist BQ788 (100nM; p>0.05). ET-1 did not affect the CBF increase produced by the NO donor SNAP and did not increase free radicals, suggesting that ET-1 did not act by reducing NO vasoactivity or bioavailability. However, in brain endothelial cell cultures ET1 (10-100nM) attenuated the NO production induced by ACh (-49±4% at 50nM; p<0.05), an effect blocked by BQ123 and Y27632. ET1 increased eNOS phosphorylation at Thr495, which inhibits eNOS, and reduced phosphorylation at Ser1177, which activates eNOS, effects blocked by Y27632. These findings, collectively, suggest that ET1 alters key regulatory mechanisms of the cerebral circulation by modulating the phosphorylation state of eNOS via ROCK. The resulting downregulation of eNOS activity is responsible for the neurovascular dysregulation induced by ET1. ET A receptors may be a valuable target to counteract the deleterious cerebrovascular actions of HTN.

Acute vasodilator effects of Rho-kinase inhibitors in neonatal rats with pulmonary hypertension unresponsive to nitric oxide

2008 ◽  
Vol 294 (2) ◽  
pp. L205-L213 ◽  
Author(s):  
Patrick J. McNamara ◽  
Prashanth Murthy ◽  
Crystal Kantores ◽  
Lilian Teixeira ◽  
Doreen Engelberts ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Pulmonary Hypertension ◽  
Kinase Inhibitors ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Neonatal Rat ◽  
Rock Inhibitor ◽  
No Donor ◽  
Rat Models ◽  
Pulmonary Hemodynamic

Pulmonary hypertension (PHT) in neonates is often refractory to the current best therapy, inhaled nitric oxide (NO). The utility of a new class of pulmonary vasodilators, Rho-kinase (ROCK) inhibitors, has not been examined in neonatal animals. Our objective was to examine the activity and expression of RhoA/ROCK in normal and injured pulmonary arteries and to determine the short-term pulmonary hemodynamic (assessed by pulse wave Doppler) effects of ROCK inhibitors (15 mg/kg ip Y-27632 or 30 mg/kg ip fasudil) in two neonatal rat models of chronic PHT with pulmonary vascular remodeling (chronic hypoxia, 0.13 FiO2, or 1 mg·kg−1·day−1 ip chronic bleomycin for 14 days from birth). Activity of the RhoA/ROCK pathway and ROCK expression were increased in hypoxia- and bleomycin-induced PHT. In both models, severe PHT [characterized by raised pulmonary vascular resistance (PVR) and impaired right ventricular (RV) performance] did not respond acutely to inhaled NO (20 ppm for 15 min) or to a single bolus of a NO donor, 3-morpholinosydnonimine hydrochloride (SIN-1; 2 μg/kg ip). In contrast, a single intraperitoneal bolus of either ROCK inhibitor (Y-27632 or fasudil) completely normalized PVR but had no acute effect on RV performance. ROCK-mediated vasoconstriction appears to play a key role in chronic PHT in our two neonatal rat models. Inhibitors of ROCK have potential as a testable therapy in neonates with PHT that is refractory to NO.

Endothelin-1-induced vasoconstriction is inhibited during erection in rats

2001 ◽  
Vol 281 (2) ◽  
pp. R476-R483 ◽  
Author(s):  
T. M. Mills ◽  
D. M. Pollock ◽  
R. W. Lewis ◽  
H. S. Branam ◽  
C. J. Wingard
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Mean Arterial Pressure ◽  
Arterial Pressure ◽  
Muscle Relaxation ◽  
Corpus Cavernosum ◽  
Smooth Muscle Relaxation ◽  
No Donor ◽  
Intracavernosal Injection ◽  
Endothelin 1

Recent evidence indicates that endothelin-1 (ET-1) might be a principal vasoconstrictor in the penis. We report that ET-1 injection into the cavernous sinuses before erection sharply reduced the magnitude of subsequent erections. Corpus cavernosum pressure-to-mean arterial pressure ratios (CCP/MAP), with maximal ganglionic stimulation, were 0.62 ± 0.05 before ET-1 injection and 0.31 ± 0.05 after, indicating that ET-1 acted as a vasoconstrictor. When ET-1 was injected during a maximal neurally induced erection, the ability of ET-1 to attenuate subsequent erections was diminished (CCP/MAP 0.75 ± 0.02 before ET-1, 0.61 ± 0.03 after). At submaximal stimulation voltages, injection of ET-1 during erection also attenuated its vasoconstrictive effect. Similarly, when ET-1 was injected during erection induced by intracavernosal injection of the nitric oxide (NO) donor NOR-1, subsequent erections were not significantly suppressed (CCP/MAP 0.53 ± 0.04 before ET-1, 0.45 ± 0.04 after). These findings that ET-1-induced vasoconstriction is attenuated during erection are consistent with the hypothesis that NO mediates erection both by initiating pathways that cause smooth muscle relaxation and by inhibiting the vasoconstrictive actions of ET-1.

scholarly journals Blockade of Nitric Oxide Synthesis in Rats Strongly Attenuates the CBF Response to Extracellular Acidosis

1993 ◽  
Vol 13 (3) ◽  
pp. 535-539 ◽  
Author(s):  
Kiyoshi Niwa ◽  
Ute Lindauer ◽  
Arno Villringer ◽  
Ulrich Dirnagl
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthesis ◽  
Cerebral Vasculature ◽  
Regional Cerebral Blood ◽  
Extracellular Acidification ◽  
No Donor ◽  
Doppler Flowmetry ◽  
Cranial Window ◽  
Extracellular Acidosis ◽  
First Time

We tested the hypothesis that the CBF response to extracellular acidosis is mediated by nitric oxide (NO). A closed cranial window, superfused with artificial CSF (aCSF), was implanted over the parietal cortex in anesthetized and ventilated Wistar rats. Regional cerebral blood flow (rCBF) was measured continuously with laser-Doppler flowmetry (LDF). The reaction of rCBF to hypercapnia (Paco2 from 30.5 ± 1.8 to 61.3 ± 5.8 mm Hg by adding CO2 to the inspiratory gas) was 2.9 ± 1.4%/mm Hg, and the reaction of rCBF to H+ (superfusion of acidic aCSF, pH 7.07 ± 0.05) was 101.7 ± 24.7%/pH unit. The regional NO synthase (NOS) activity was blocked by superfusing aCSF containing 10−3 M Nω-nitro-L-arginine (L-NA, n = 10). After 30 min of L-NA superfusion, rCBF was reduced to 80.1 ± 6.5% of baseline, and the rCBF responses to hypercapnia (Paco2 from 30.9 ± 2.9 to 58.8 ± 7.7 mm Hg) and extracellular acidosis (aCSF pH 7.08 ± 0.06) were reduced to 0.8 ± 1.1%/ mm Hg and 10.1 ± 23.0%/pH unit, respectively (both p < 0.001). This effect was stereospecific since aCSF containing 10−3 M Nω-nitro-D-arginine affected neither baseline rCBF nor the response to H+ ( n = 5). The NOS blockade did not affect the vasodilatation by the NO donor sodium nitroprusside ( n = 5, 114.3 ± 25.1% before vs. 130.2 ± 24.7% after NOS blockade). The results confirm the involvement of NO in the CBF reaction to hypercapnia and demonstrate for the first time that NOS blockade also strongly attenuates the H+ response of the cerebral vasculature. We speculate that extracellular acidification triggers the production of NO.

scholarly journals Inverse Relationship between Nitric Oxide Synthases and Endothelin-1 Synthesis in Bovine Corpus Luteum: Interactions at the Level of Luteal Endothelial Cell

Endocrinology ◽  
2006 ◽  
Vol 147 (11) ◽  
pp. 5228-5235 ◽  
Author(s):  
Maya Rosiansky-Sultan ◽  
Eyal Klipper ◽  
Katharina Spanel-Borowski ◽  
Rina Meidan
Keyword(s):  
Nitric Oxide ◽  
Corpus Luteum ◽  
Inverse Relationship ◽  
Prostaglandin F2α ◽  
Inos Mrna ◽  
No Production ◽  
Dependent Manner ◽  
No Donor ◽  
Main Site ◽  
Endothelin 1

Endothelin-1 (ET-1) and nitric oxide (NO) play pivotal roles in corpus luteum (CL) function. The present study examined the interplay between NO and ET-1 synthesis in the bovine CL. We found similar inducible and endothelial NO synthase (iNOS and eNOS, respectively) activities in the young CL (d 1–5) expressing the highest levels of both eNOS and iNOS mRNA. These values later declined at mid-cycle (d 8–15) and remained low at later stages (d 16–18). Luteolysis, initiated by prostaglandin F2α analog administration, further reduced NOS mRNA and by 24 h, NOS values dropped to approximately 15% of those at mid-cycle. eNOS protein levels followed a similar pattern to its mRNA. Because endothelial cells (ECs) are the main site for ET-1 and NO production in the CL, we examined the direct effects of the NO donor, NONOate on luteal ECs (LECs). Elevated NO levels markedly decreased ET-1 mRNA, and peptide concentrations in cultured and freshly isolated LECs in a dose-dependent manner. In agreement, NOS inhibitor, NG-nitro-l-arginine methyl ester, stimulated ET-1 mRNA expression in these cells. Interestingly, NO also up-regulated prostaglandin F2α receptors in LECs. These data show that there is an inverse relationship between NOS and ET-1 throughout the CL life span, and imply that this pattern may be the result of their interaction within the resident LECs. NOS are expressed in a physiologically relevant manner: elevated NO at an early luteal stage is likely to play an important role in angiogenesis, whereas reduced levels of NO during luteal regression may facilitate the sustained up-regulation of ET-1 levels during luteolysis.

scholarly journals Nitric Oxide Modulates the CBF Response to Increased Extracellular Potassium

1995 ◽  
Vol 15 (6) ◽  
pp. 914-919 ◽  
Author(s):  
J. P. Dreier ◽  
K. Körner ◽  
A. Görner ◽  
U. Lindauer ◽  
M. Weih ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Extracellular Potassium ◽  
Regional Cerebral Blood ◽  
High Potassium ◽  
Subsequent Increase ◽  
No Donor ◽  
Doppler Flowmetry ◽  
Cranial Window ◽  
Artificial Cerebrospinal Fluid ◽  
Oxide Synthase

The response of the regional cerebral blood flow (rCBF) to brain topical superfusion of 20 m M K+ was characterized in a closed cranial window preparation in barbiturate anesthetized and ventilated rats; Increasing K+ in the artificial cerebrospinal fluid (ACSF) induced a rCBF elevation (measured by laser–Doppler flowmetry) of +85 ± 37% above baseline (n = 19). This elevation was stable for >3 h with continuous superfusion of increased K+ (n = 5) and partially reversible to a level of + 18 ± 19% above baseline when returning to a physiological K+ concentration. Nitric oxide synthase (NOS) inhibition by brain topical superfusion with Nω-nitro-L-arginine (L-NA) revealed (a) Addition of L-NA to high-potassium ACSF reduced the rCBF increase from + 94 ± 36% to + 21 ± 18% (p ≤ 0.01, n = 7). (b) When L-NA was superfused for 60 min before increasing K+, rCBF decreased to – 17 ± 7% below baseline. Subsequent coapplication of L-NA and increased K+ induced only an elevation of +7 ± 4% above baseline (n = 4). (c) When the NO donor S-nitroso- N-acetylpenicillamine (SNAP) was added during NOS inhibition to restore basal tissue NO levels, the resultant level of rCBF was +28 ± 54% above baseline. Subsequent increase of K+ in the presence of NOS inhibition and SNAP elevated rCBF to + 137 ± 89% above baseline (n = 4). Statistical analysis comparing K+-induced elevation of rCBF (a) without any added drugs, (b) in the presence of NOS inhibition with L-NA, and (c) in the presence of both NOS inhibition and SNAP revealed that K+-induced elevation in the presence of NOS inhibition was significantly reduced (p ≤ 0.05) whereas no statistical difference was found between K+-induced elevation of rCBF without drugs compared with the K+-induced elevation of rCBF in the presence of L-NA and SNAP. We conclude that NO is a modulator of the rCBF elevation to increased extracellular K+ concentration.

Effect of nitric oxide and potassium channel agonists and inhibitors on basilar artery diameter

1997 ◽  
Vol 272 (1) ◽  
pp. H256-H262 ◽  
Author(s):  
C. G. Sobey ◽  
F. M. Faraci
Keyword(s):  
Nitric Oxide ◽  
Sodium Nitroprusside ◽  
Basilar Artery ◽  
Selective Inhibitor ◽  
No Production ◽  
No Donor ◽  
K Channels ◽  
Cranial Window ◽  
Endogenous Production ◽  
Direct Activator

The first goal of this study was to examine the hypothesis that dilatation of the basilar artery in response to activation of ATP-sensitive K+ channels is mediated by nitric oxide (NO). Diameter of the basilar artery (209 +/- 5 microns, mean +/- SE) was measured using a cranial window in anesthetized rats. Aprikalim (a direct activator of ATP-sensitive K+ channels) dilated the basilar artery under control conditions. Inhibition of endogenous NO production with NG-nitro-L-arginine (L-NNA, 10(-4) M) did not alter responses to aprikalim. The second goal was to determine whether vasodilatation in response to NO is dependent on activation of calcium-activated K+ channels. Tetraethylammonium (TEA, 10(-3) M), an inhibitor of calcium-activated K+ channels, did not affect dilator responses to sodium nitroprusside (an NO donor) under control conditions. Responses to nitroprusside (10(-8) and 10(-7) M) were augmented more than twofold during application of L-NNA. In the presence of L-NNA, the augmented portion of the response to nitroprusside was inhibited by TEA and iberiotoxin (5 x 10(-8) M, a highly selective inhibitor of calcium-activated K+ channels), but it was not inhibited by glibenclamide (10(-6) M), an inhibitor of ATP-sensitive K+ channels. These findings suggest that dilator responses of the basilar artery to an activator of ATP-sensitive potassium channels are not mediated by NO. Calcium-activated K+ channels may not normally contribute to dilator responses of the basilar artery to nitroprusside. The effects of TEA and iberiotoxin suggest that when endogenous production of NO is inhibited, sodium nitroprusside causes the opening of calcium-activated K+ channels, contributing to an augmented vasodilator response.

Inducible nitric oxide synthase augments injury elicited by oxidative stress in rat cardiac myocytes

1998 ◽  
Vol 274 (1) ◽  
pp. C245-C252 ◽  
Author(s):  
Junsuke Igarashi ◽  
Masashi Nishida ◽  
Shiro Hoshida ◽  
Nobushige Yamashita ◽  
Hiroaki Kosaka ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Nitric Oxide ◽  
Cardiac Myocytes ◽  
Myocardial Injury ◽  
No Production ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
No Donor ◽  
Oxide Synthase ◽  
Gpx Activity

The effects of nitric oxide (NO) produced by cardiac inducible NO synthase (iNOS) on myocardial injury after oxidative stress were examined. Interleukin-1β induced cultured rat neonatal cardiac myocytes to express iNOS. After induction of iNOS,l-arginine enhanced NO production in a concentration-dependent manner. Glutathione peroxidase (GPX) activity in myocytes was attenuated by elevated iNOS activity and by an NO donor, S-nitroso- N-acetyl-penicillamine (SNAP). Although NO production by iNOS did not induce myocardial injury, NO augmented release of lactate dehydrogenase from myocyte cultures after addition of H2O2(0.1 mM, 1 h). Inhibition of iNOS with Nω-nitro-l-arginine methyl ester ameliorated the effects of NO-enhancing treatments on myocardial injury and GPX activity. SNAP augmented the myocardial injury induced by H2O2. Inhibition of GPX activity with antisense oligodeoxyribonucleotide for GPX mRNA increased myocardial injury by H2O2. Results suggest that the induction of cardiac iNOS promotes myocardial injury due to oxidative stress via inactivation of the intrinsic antioxidant enzyme, GPX.

scholarly journals Hydrogen Sulfide Increases Nitric Oxide Production and Subsequent S-Nitrosylation in Endothelial Cells

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Ping-Ho Chen ◽  
Yaw-Syan Fu ◽  
Yun-Ming Wang ◽  
Kun-Han Yang ◽  
Danny Ling Wang ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Hydrogen Sulfide ◽  
Protein Kinase ◽  
Fluorescent Probe ◽  
Acid Methyl Ester ◽  
High Specificity ◽  
Protein S ◽  
No Production ◽  
No Donor

Hydrogen sulfide (H2S) and nitric oxide (NO), two endogenous gaseous molecules in endothelial cells, got increased attention with respect to their protective roles in the cardiovascular system. However, the details of the signaling pathways between H2S and NO in endothelia cells remain unclear. In this study, a treatment with NaHS profoundly increased the expression and the activity of endothelial nitric oxide synthase. Elevated gaseous NO levels were observed by a novel and specific fluorescent probe, 5-amino-2-(6-hydroxy-3-oxo-3H-xanthen-9-yl)benzoic acid methyl ester (FA-OMe), and quantified by flow cytometry. Further study indicated an increase of upstream regulator for eNOS activation, AMP-activated protein kinase (AMPK), and protein kinase B (Akt). By using a biotin switch, the level of NO-mediated protein S-nitrosylation was also enhanced. However, with the addition of the NO donor, NOC-18, the expressions of cystathionine-γ-lyase, cystathionine-β-synthase, and 3-mercaptopyruvate sulfurtransferase were not changed. The level of H2S was also monitored by a new designed fluorescent probe, 4-nitro-7-thiocyanatobenz-2-oxa-1,3-diazole (NBD-SCN) with high specificity. Therefore, NO did not reciprocally increase the expression of H2S-generating enzymes and the H2S level. The present study provides an integrated insight of cellular responses to H2S and NO from protein expression to gaseous molecule generation, which indicates the upstream role of H2S in modulating NO production and protein S-nitrosylation.

Maturational changes in the regulation of pulmonary vascular tone by nitric oxide in neonatal rats

2007 ◽  
Vol 293 (5) ◽  
pp. L1261-L1270 ◽  
Author(s):  
Louis G. Chicoine ◽  
Michael L. Paffett ◽  
Mark R. Girton ◽  
Matthew J. Metropoulus ◽  
Mandar S. Joshi ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Guanylyl Cyclase ◽  
Vascular Tone ◽  
Age Groups ◽  
Soluble Guanylyl Cyclase ◽  
No Production ◽  
Sprague Dawley ◽  
No Donor ◽  
Early Postnatal Development ◽  
Old Rats

Nitric oxide (NO) is an important regulator of vasomotor tone in the pulmonary circulation. We tested the hypothesis that the role NO plays in regulating vascular tone changes during early postnatal development. Isolated, perfused lungs from 7- and 14-day-old Sprague-Dawley rats were studied. Baseline total pulmonary vascular resistance (PVR) was not different between age groups. The addition of KCl to the perfusate caused a concentration-dependent increase in PVR that did not differ between age groups. However, the nitric oxide synthase (NOS) inhibitor Nω-nitro-l-arginine augmented the K+-induced increase in PVR in both groups, and the effect was greater in lungs from 14-day-old rats vs. 7-day-old rats. Lung levels of total endothelial, inducible, and neuronal NOS proteins were not different between groups; however, the production rate of exhaled NO was greater in lungs from 14-day-old rats compared with those of 7-day-old rats. Vasodilation to 0.1 μM of the NO donor spermine NONOate was greater in 14-day lungs than in 7-day lungs, and lung levels of both soluble guanylyl cyclase and cGMP were greater at 14 days than at 7 days. Vasodilation to 100 μM of the cGMP analog 8-(4-chlorophenylthio)guanosine-3′,5′-cyclic monophosphate was greater in 7-day lungs than in 14-day lungs. Our results demonstrate that the pulmonary vascular bed depends more on NO production to modulate vascular tone at 14 days than at 7 days of age. The observed differences in NO sensitivity may be due to maturational increases in soluble guanylyl cyclase protein levels.

Nitric oxide decreases endothelin-1 secretion through the activation of soluble guanylate cyclase

2004 ◽  
Vol 286 (5) ◽  
pp. L984-L991 ◽  
Author(s):  
Lisa K. Kelly ◽  
Stephen Wedgwood ◽  
Robin H. Steinhorn ◽  
Stephen M. Black
Keyword(s):  
Nitric Oxide ◽  
Guanylate Cyclase ◽  
Soluble Guanylate Cyclase ◽  
Primary Cultures ◽  
No Donor ◽  
Endothelin 1 ◽  
Cgmp Analog ◽  
Pulmonary Arterial ◽  
Long Acting ◽  
Arterial Endothelial Cells

The use of exogenous nitric oxide (NO) has been shown to alter the regulation of other endothelially derived mediators of vascular tone, such as endothelin-1 (ET-1). However, the interaction between NO and ET-1 appears to be complex and remains incompletely understood. One of the major actions of NO is the activation of soluble guanylate cyclase (sGC) with the subsequent generation of cGMP. Therefore, we undertook this study to test the hypothesis that NO regulates ET-1 production via the activation of the sGC/cGMP pathway. The results obtained indicated that the exposure of primary cultures of 4-wk-old ovine pulmonary arterial endothelial cells (4-wk PAECs) to the long-acting NO donor DETA NONOate induced both a dose- and time-dependent decrease in secreted ET-1. This decrease in ET-1 secretion occurred in the absence of changes in endothelin-converting enzyme-1 or sGC expression but in conjunction with a decrease in prepro-ET-1 mRNA. The changes in ET-1 release were inversely proportional to the cellular cGMP content. Furthermore, the NO-independent activator of sGC, YC-1, or treatment with a cGMP analog also produced significant decreases in ET-1 secretion. Conversely, pretreatment with the sGC inhibitor ODQ blocked the NO-induced decrease in ET-1. Therefore, we conclude that exposure of 4-wk PAECs to exogenous NO decreases secreted ET-1 resulting from the activation of sGC and increased cGMP generation.

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