scholarly journals Membrane depolarization is required for pressure-dependent pulmonary arterial tone but not enhanced vasoconstriction to endothelin-1 following chronic hypoxia

2020 ◽  
Vol 10 (4) ◽  
pp. 204589402097355
Author(s):  
Charles E Norton ◽  
Nikki L Jernigan ◽  
Benjimen R Walker ◽  
Thomas C Resta
Keyword(s):  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Membrane Depolarization ◽  
Src Kinases ◽  
Endothelin 1 ◽  
Receptor Inhibition ◽  
Pulmonary Arterial ◽  
Arterial Tone

Enhanced vasoconstriction is increasingly identified as an important contributor to the development of pulmonary hypertension. Chronic hypoxia results in enhanced Rho kinase mediated Ca2+ sensitization contributing to pressure-dependent pulmonary arterial tone as well as augmented vasoconstriction to endothelin-1 and depolarizing stimuli. We sought to investigate the interaction between these vasoconstrictor stimuli in isolated, pressurized, pulmonary arteries. We used the K+ ionophore, valinomycin, to clamp membrane potential (Vm) to investigate the role of membrane depolarization in endothelin-1 and pressure-dependent constriction, and endothelin-1 receptor inhibitors to determine whether membrane depolarization or stretch signal through endothelin-1 receptors. Clamping Vm prevented pressure-dependent tone, but not enhanced vasoconstriction to endothelin-1 following chronic hypoxia. Furthermore, endothelin-1 receptor inhibition had no effect on either pressure-dependent tone or vasoconstriction to KCl. As Src kinases contribute to both pressure-dependent tone and enhanced endothelin-1 vasoconstriction following chronic hypoxia, we further investigated their role in depolarization-induced vasoconstriction. Inhibition of Src kinases attenuated enhanced vasoconstriction to KCl. We conclude that membrane depolarization contributes to pressure-dependent tone but not enhanced vasoconstriction to ET-1, and that Src kinases serve as upstream mediators facilitating enhanced Rho kinase-dependent vasoconstriction following chronic hypoxia.

Sex Differences in Right Ventricular-Vascular Coupling and Pulmonary Artery Impedance in Response to Chronic Hypoxia and Recovery

2012 ◽  
Author(s):  
Aiping Liu ◽  
Lian Tian ◽  
Diana M. Tabima ◽  
Naomi C. Chesler
Keyword(s):  
Pulmonary Artery ◽  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
Pulmonary Artery Hypertension ◽  
Vascular Impedance ◽  
Female Predominance ◽  
Collagen Accumulation ◽  
Dominant Disease ◽  
Pulmonary Arterial

Pulmonary artery hypertension (PAH) is a female dominant disease (the female-to-male ratio is 4:1), characterized by small distal pulmonary arterial narrowing and large proximal arterial stiffening, which increase right ventricle (RV) afterload and ultimately lead to RV failure [1,2]. Our recent studies have shown that collagen accumulation induced by chronic hypoxia increases the stiffness of the large extralobar pulmonary arteries (PAs) [3], and affects pulmonary vascular impedance (PVZ) [4]. The role of collagen in the female predominance in developing PAH has not been explored to date.

scholarly journals Chronic hypoxia augments depolarization-induced Ca2+ sensitization in pulmonary vascular smooth muscle through superoxide-dependent stimulation of RhoA

2010 ◽  
Vol 298 (2) ◽  
pp. L232-L242 ◽  
Author(s):  
Brad R. S. Broughton ◽  
Nikki L. Jernigan ◽  
Charles E. Norton ◽  
Benjimen R. Walker ◽  
Thomas C. Resta
Keyword(s):  
Pulmonary Hypertension ◽  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Membrane Depolarization ◽  
Dependent Manner ◽  
Rock Inhibitor ◽  
Rhoa Activation

Rho kinase (ROCK)-dependent vasoconstriction has been implicated as a major factor in chronic hypoxia (CH)-induced pulmonary hypertension. This component of pulmonary hypertension is associated with arterial myogenicity and increased vasoreactivity to receptor-mediated agonists and depolarizing stimuli resulting from ROCK-dependent myofilament Ca2+ sensitization. On the basis of separate lines of evidence that CH increases pulmonary arterial superoxide (O2−) generation and that O2− stimulates RhoA/ROCK signaling in vascular smooth muscle (VSM), we hypothesized that depolarization-induced O2− generation mediates enhanced RhoA-dependent Ca2+ sensitization in pulmonary VSM following CH. To test this hypothesis, we determined effects of the ROCK inhibitor HA-1077 and the O2−-specific spin trap tiron on vasoconstrictor reactivity to depolarizing concentrations of KCl in isolated lungs and Ca2+-permeabilized, pressurized small pulmonary arteries from control and CH (4 wk at 0.5 atm) rats. Using the same vessel preparation, we examined effects of CH on KCl-dependent VSM membrane depolarization and O2− generation using sharp electrodes and the fluorescent indicator dihydroethidium, respectively. Finally, using a RhoA-GTP pull-down assay, we investigated the contribution of O2− to depolarization-induced RhoA activation. We found that CH augmented KCl-dependent vasoconstriction through a Ca2+ sensitization mechanism that was inhibited by HA-1077 and tiron. Furthermore, CH caused VSM membrane depolarization that persisted with increasing concentrations of KCl, enhanced KCl-induced O2− generation, and augmented depolarization-dependent RhoA activation in a O2−-dependent manner. These findings reveal a novel mechanistic link between VSM membrane depolarization, O2− generation, and RhoA activation that mediates enhanced myofilament Ca2+ sensitization and pulmonary vasoconstriction following CH.

scholarly journals Role of the RhoA/ROCK pathway in high-altitude associated neonatal pulmonary hypertension in lambs

2016 ◽  
Vol 310 (11) ◽  
pp. R1053-R1063 ◽  
Author(s):  
Nandy C. Lopez ◽  
German Ebensperger ◽  
Emilio A. Herrera ◽  
Roberto V. Reyes ◽  
Gloria Calaf ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
High Altitude ◽  
Acute Hypoxia ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Daily Administration ◽  
Rock Inhibitor ◽  
Pulmonary Arterial ◽  
Neonatal Lambs

Exposure to high-altitude chronic hypoxia during pregnancy may cause pulmonary hypertension in neonates, as a result of vasoconstriction and vascular remodeling. We hypothesized that susceptibility to pulmonary hypertension, due to an augmented expression and activity of the RhoA/Rho-kinase (ROCK) pathway in these neonates, can be reduced by daily administration of fasudil, a ROCK inhibitor. We studied 10 highland newborn lambs with conception, gestation, and birth at 3,600 m in Putre, Chile. Five highland controls (HLC) were compared with 5 highland lambs treated with fasudil (HL-FAS; 3 mg·kg−1·day−1 iv for 10 days). Ten lowland controls were studied in Lluta (50 m; LLC). During the 10 days of fasudil daily administration, the drug decreased pulmonary arterial pressure (PAP) and resistance (PVR), basally and during a superimposed episode of acute hypoxia. HL-FAS small pulmonary arteries showed diminished muscular area and a reduced contractile response to the thromboxane analog U46619 compared with HLC. Hypoxia, but not fasudil, changed the protein expression pattern of the RhoA/ROCKII pathway. Moreover, HL-FAS lungs expressed less pMYPT1T850 and pMYPT1T696 than HLC, with a potential increase of the myosin light chain phosphatase activity. Finally, hypoxia induced RhoA, ROCKII, and PKG mRNA expression in PASMCs of HLC, but fasudil reduced them (HL-FAS) similarly to LLC. We conclude that fasudil decreases the function of the RhoA/ROCK pathway, reducing the PAP and PVR in chronically hypoxic highland neonatal lambs. The inhibition of ROCKs by fasudil may offer a possible therapeutic tool for the pulmonary hypertension of the neonates.

scholarly journals Endothelin-1 contributes to increased NFATc3 activation by chronic hypoxia in pulmonary arteries

2011 ◽  
Vol 301 (2) ◽  
pp. C441-C450 ◽  
Author(s):  
Sergio de Frutos ◽  
Juan Manuel Ramiro Diaz ◽  
Carlos H. Nitta ◽  
Mingma L. Sherpa ◽  
Laura V. Gonzalez Bosc
Keyword(s):  
Nuclear Import ◽  
Actin Polymerization ◽  
Chronic Hypoxia ◽  
Ex Vivo ◽  
Pulmonary Arteries ◽  
Green Fluorescence Protein ◽  
Luciferase Reporter ◽  
Activated T Cells ◽  
Endothelin 1 ◽  
Pulmonary Arterial

Chronic hypoxia (CH) activates the Ca2+-dependent transcription factor nuclear factor of activated T cells isoform c3 (NFATc3) in mouse pulmonary arteries. However, the mechanism of this response has not been explored. Since we have demonstrated that NFATc3 is required for CH-induced pulmonary arterial remodeling, establishing how CH activates NFATc3 is physiologically significant. The goal of this study was to test the hypothesis that endothelin-1 (ET-1) contributes to CH-induced NFATc3 activation. We propose that this mechanism requires increased pulmonary arterial smooth muscle cell (PASMC) intracellular Ca2+ concentration ([Ca2+]i) and stimulation of RhoA/Rho kinase (ROK), leading to calcineurin activation and actin cytoskeleton polymerization, respectively. We found that: 1) CH increases pulmonary arterial pre-pro-ET-1 mRNA expression and lung RhoA activity; 2) inhibition of ET receptors, calcineurin, L-type Ca2+ channels, and ROK blunts CH-induced NFATc3 activation in isolated intrapulmonary arteries from NFAT-luciferase reporter mice; and 3) both ET-1-induced NFATc3 activation in isolated mouse pulmonary arteries ex vivo and ET-1-induced NFATc3-green fluorescence protein nuclear import in human PASMC depend on ROK and actin polymerization. This study suggests that CH increases ET-1 expression, thereby elevating PASMC [Ca2+]i and RhoA/ROK activity. As previously demonstrated, elevated [Ca2+]i is required to activate calcineurin, which dephosphorylates NFATc3, allowing its nuclear import. Here, we demonstrate that ROK increases actin polymerization, thus providing structural support for NFATc3 nuclear transport.

scholarly journals Contribution of Rho Kinase to Elevated Basal Pulmonary Arterial Tone Following Chronic Hypoxia

2006 ◽  
Vol 20 (4) ◽  
Author(s):  
Brad R.S. Broughton ◽  
Benjimen C. Walker ◽  
Thomas R. Resta
Keyword(s):  
Chronic Hypoxia ◽  
Rho Kinase ◽  
Pulmonary Arterial ◽  
Arterial Tone

scholarly journals Reduced membrane cholesterol limits pulmonary endothelial Ca2+ entry after chronic hypoxia

2017 ◽  
Vol 312 (6) ◽  
pp. H1176-H1184 ◽  
Author(s):  
Bojun Zhang ◽  
Jay S. Naik ◽  
Nikki L. Jernigan ◽  
Benjimen R. Walker ◽  
Thomas C. Resta
Keyword(s):  
Pulmonary Hypertension ◽  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
Barometric Pressure ◽  
Membrane Cholesterol ◽  
Potential Mechanism ◽  
Direct Role ◽  
Pulmonary Arterial ◽  
Arterial Endothelial Cells

Chronic hypoxia (CH)-induced pulmonary hypertension is associated with diminished production of endothelium-derived Ca2+-dependent vasodilators such as nitric oxide. Interestingly, ATP-induced endothelial Ca2+ entry as well as membrane cholesterol (Chol) are decreased in pulmonary arteries from CH rats (4 wk, barometric pressure = 380 Torr) compared with normoxic controls. Store-operated Ca2+ entry (SOCE) and depolarization-induced Ca2+ entry are major components of the response to ATP and are similarly decreased after CH. We hypothesized that membrane Chol facilitates both SOCE and depolarization-induced pulmonary endothelial Ca2+ entry and that CH attenuates these responses by decreasing membrane Chol. To test these hypotheses, we administered Chol or epicholesterol (Epichol) to acutely isolated pulmonary arterial endothelial cells (PAECs) from control and CH rats to either supplement or replace native Chol, respectively. The efficacy of membrane Chol manipulation was confirmed by filipin staining. Epichol greatly reduced ATP-induced Ca2+ influx in PAECs from control rats. Whereas Epichol similarly blunted endothelial SOCE in PAECs from both groups, Chol supplementation restored diminished SOCE in PAECs from CH rats while having no effect in controls. Similar effects of Chol manipulation on PAEC Ca2+ influx were observed in response to a depolarizing stimulus of KCl. Furthermore, KCl-induced Ca2+ entry was inhibited by the T-type Ca2+ channel antagonist mibefradil but not the L-type Ca2+ channel inhibitor diltiazem. We conclude that PAEC membrane Chol is required for ATP-induced Ca2+ entry and its two components, SOCE and depolarization-induced Ca2+ entry, and that reduced Ca2+ entry after CH may be due to loss of this key regulator. NEW & NOTEWORTHY This research is the first to examine the direct role of membrane cholesterol in regulating pulmonary endothelial agonist-induced Ca2+ entry and its components. The results provide a potential mechanism by which chronic hypoxia impairs pulmonary endothelial Ca2+ influx, which may contribute to pulmonary hypertension.

scholarly journals Chronic hypoxia upregulates pulmonary arterial ASIC1: a novel mechanism of enhanced store-operated Ca2+ entry and receptor-dependent vasoconstriction

2012 ◽  
Vol 302 (6) ◽  
pp. C931-C940 ◽  
Author(s):  
Nikki L. Jernigan ◽  
Lindsay M. Herbert ◽  
Benjimen R. Walker ◽  
Thomas C. Resta
Keyword(s):  
Pulmonary Hypertension ◽  
Smooth Muscle ◽  
Ion Channel ◽  
Vessel Wall ◽  
Wistar Rats ◽  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
Intracellular Ca ◽  
Pulmonary Arterial

Acid-sensing ion channel 1 (ASIC1) is a newly characterized contributor to store-operated Ca2+ entry (SOCE) in pulmonary vascular smooth muscle (VSM). Since SOCE is implicated in elevated basal VSM intracellular Ca2+ concentration ([Ca2+]i) and augmented vasoconstriction in chronic hypoxia (CH)-induced pulmonary hypertension, we hypothesized that ASIC1 contributes to these responses. To test this hypothesis, we examined effects of the specific pharmacologic ASIC1a inhibitor, psalmotoxin 1 (PcTX1), on vasoconstrictor and vessel wall [Ca2+]i responses to UTP and KCl (depolarizing stimulus) in fura-2-loaded, pressurized small pulmonary arteries from control and CH (4 wk at 0.5 atm) Wistar rats. PcTX1 had no effect on basal vessel wall [Ca2+]i, but attenuated vasoconstriction and increases in vessel wall [Ca2+]i to UTP in arteries from control and CH rats; normalizing responses between groups. In contrast, responses to the depolarizing stimulus, KCl, were unaffected by CH exposure or PcTX1. Upon examining potential Ca2+ influx mechanisms, we found that PcTX1 prevented augmented SOCE following CH. Exposure to CH resulted in a significant increase in pulmonary arterial ASIC1 protein. This study supports a novel role of ASIC1 in elevated receptor-stimulated vasoconstriction following CH which is likely mediated through increased ASIC1 expression and SOCE.

Hypoxic pulmonary hypertension: role of superoxide and NADPH oxidase (gp91phox)

2006 ◽  
Vol 290 (1) ◽  
pp. L2-L10 ◽  
Author(s):  
John Q. Liu ◽  
Igor N. Zelko ◽  
Efua M. Erbynn ◽  
James S. K. Sham ◽  
Rodney J. Folz
Keyword(s):  
Nadph Oxidase ◽  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
Superoxide Production ◽  
Wall Thickening ◽  
Wild Type ◽  
Enhanced Chemiluminescence ◽  
Pulmonary Arterial ◽  
Cuzn Superoxide Dismutase

Chronic exposure to low-O2tension induces pulmonary arterial hypertension (PAH), which is characterized by vascular remodeling and enhanced vasoreactivity. Recent evidence suggests that reactive oxygen species (ROS) may be involved in both processes. In this study, we critically examine the role superoxide and NADPH oxidase plays in the development of chronic hypoxic PAH. Chronic hypoxia (CH; 10% O2for 3 wk) caused a significant increase in superoxide production in intrapulmonary arteries (IPA) of wild-type (WT) mice as measured by lucigenin-enhanced chemiluminescence. The CH-induced increase in the generation of ROS was obliterated in NADPH oxidase (gp91phox) knockout (KO) mice, suggesting that NADPH oxidase was the major source of ROS. Importantly, pathological changes associated with CH-induced PAH (mean right ventricular pressure, medial wall thickening of small pulmonary arteries, and right heart hypertrophy) were completely abolished in NADPH oxidase (gp91phox) KO mice. CH potentiated vasoconstrictor responses of isolated IPAs to both 5-hydroxytryptamine (5-HT) and the thromboxane mimetic U-46619. Administration of CuZn superoxide dismutase to isolated IPA significantly reduced CH-enhanced superoxide levels and reduced the CH-enhanced vasoconstriction to 5-HT and U-46619. Additionally, CH-enhanced superoxide production and vasoconstrictor activity seen in WT IPAs were markedly reduced in IPAs isolated from NADPH oxidase (gp91phox) KO mice. These results demonstrate a pivotal role for gp91phox-dependent superoxide production in the pathogenesis of CH-induced PAH.

scholarly journals Actin polymerization contributes to enhanced pulmonary vasoconstrictor reactivity after chronic hypoxia

2018 ◽  
Vol 314 (5) ◽  
pp. H1011-H1021 ◽  
Author(s):  
Laura Weise-Cross ◽  
Michelle A. Sands ◽  
Joshua R. Sheak ◽  
Brad R. S. Broughton ◽  
Jessica B. Snow ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Actin Polymerization ◽  
Chronic Hypoxia ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Reactive Oxygen ◽  
Dependent Manner ◽  
Oxygen Species ◽  
Pulmonary Arterial ◽  
Arterial Constriction

Chronic hypoxia (CH) augments basal and endothelin-1 (ET-1)-induced pulmonary vasoconstrictor reactivity through reactive oxygen species (ROS) generation and RhoA/Rho kinase (ROCK)-dependent myofilament Ca2+ sensitization. Because ROCK promotes actin polymerization and the actin cytoskeleton regulates smooth muscle tension, we hypothesized that actin polymerization is required for enhanced basal and ET-1-dependent vasoconstriction after CH. To test this hypothesis, both end points were monitored in pressurized, endothelium-disrupted pulmonary arteries (fourth-fifth order) from control and CH (4 wk at 0.5 atm) rats. The actin polymerization inhibitors cytochalasin and latrunculin attenuated both basal and ET-1-induced vasoconstriction only in CH vessels. To test whether CH directly alters the arterial actin profile, we measured filamentous actin (F-actin)-to-globular actin (G-actin) ratios by fluorescent labeling of F-actin and G-actin in fixed pulmonary arteries and actin sedimentation assays using homogenized pulmonary artery lysates. We observed no difference in actin polymerization between groups under baseline conditions, but ET-1 enhanced actin polymerization in pulmonary arteries from CH rats. This response was blunted by the ROS scavenger tiron, the ROCK inhibitor fasudil, and the mDia (RhoA effector) inhibitor small-molecule inhibitor of formin homology domain 2. Immunoblot analysis revealed an effect of CH to increase both phosphorylated (inactive) and total levels of the actin disassembly factor cofilin but not phosphorylated cofilin-to-total cofilin ratios. We conclude that actin polymerization contributes to increased basal pulmonary arterial constriction and ET-1-induced vasoconstrictor reactivity after CH in a ROS- and ROCK-dependent manner. Our results further suggest that enhanced ET-1-mediated actin polymerization after CH is dependent on mDia but independent of changes in the phosphorylated cofilin-to-total cofilin ratio. NEW & NOTEWORTHY This research is the first to demonstrate a role for actin polymerization in chronic hypoxia-induced basal pulmonary arterial constriction and enhanced agonist-induced vasoconstrictor activity. These results suggest that a reactive oxygen species-Rho kinase-actin polymerization signaling pathway mediates this response and may provide a mechanistic basis for the vasoconstrictor component of pulmonary hypertension.

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