Activation of multiple mechanisms including phospholipase D by endothelin-1 in rat aorta

1992 ◽  
Vol 262 (4) ◽  
pp. C941-C949 ◽  
Author(s):  
Y. Liu ◽  
B. Geisbuhler ◽  
A. W. Jones
Keyword(s):  
Phospholipase D ◽  
Channel Blocker ◽  
Platelet Activating Factor ◽  
Rat Aorta ◽  
Cellular Mechanisms ◽  
Endothelin 1 ◽  
Kinase C ◽  
Arachidonate Cascade ◽  
Alpha Response ◽  
Pkc Activation

This study investigated the cellular mechanisms underlying the endothelin-1 (ET-1)-induced contraction of rat aorta with focus on the involvement of phospholipase D (PLD). Preincubating rat aorta in Ca(2+)-free solution reduced the contraction by 80%, whereas diltiazem (10 microM), a voltage-operated Ca2+ channel blocker, caused only a small reduction (27%, P less than 0.05) of the contraction. In myo-[3H]inositol-labeled aorta, ET-1 stimulated the formation of [3H]inositol bisphosphate and [3H]inositol trisphosphate, indicating the activation of phospholipase C (PLC). In aorta labeled with 32PO4, [3H] myristic acid or [32P]lyso-platelet-activating factor followed by exposure to ethanol (0.5%), ET-1 stimulated phosphatidylethanol (PEt) production, suggesting that ET-1 activates PLD. The PEt response was not attenuated by staurosporine (ST, 0.1 microM), an inhibitor of protein kinase C (PKC) but was inhibited by removal of Ca2+. The ET-1-induced PEt response was at least additive to that induced by phorbol 12-myristate 13-acetate (1 microM). ET-1 also stimulated the release of 6-ketoprostaglandin F1 alpha (6-keto-PGF1 alpha) into the tissue medium. Unlike the PEt responses, the 6-keto-PGF1 alpha response could be inhibited by ST. Removal of Ca2+ abolished the response. These results suggest that 1) ET-1 activates multiple cellular mechanisms including PLC, PLD, and the arachidonate cascade; 2) PKC activation may not be essential for the ET-1 activation of PLD but may play an important role in the ET-1 stimulation of 6-keto-PGF1 alpha release; and 3) Ca2+ is an important factor in the ET-1-induced PLD activity and 6-keto-PGF1 alpha release.

scholarly journals Differential regulation of phospholipase D and phospholipase A2 by protein kinase C in P388D1 macrophages

1997 ◽  
Vol 321 (3) ◽  
pp. 805-810 ◽  
Author(s):  
Jesús BALSINDE ◽  
María A. BALBOA ◽  
Paul A. INSEL ◽  
Edward A. DENNIS
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase A2 ◽  
Phospholipase D ◽  
Phorbol Esters ◽  
Platelet Activating Factor ◽  
Lipid Mediator ◽  
Kinase C ◽  
Pkc Isoforms ◽  
Pkc Activation

Activation of P388D1 macrophages by phorbol myristate acetate (PMA) resulted in the translocation of the protein kinase C (PKC) isoforms α, Δ, and ε from the cytosol to membranes. Furthermore, PMA activated phospholipase D (PLD) in these cells, and potentiated the effect of the inflammatory lipid mediator platelet-activating factor (PAF) on PLD activation. PAF also activated phospholipase A2 (PLA2) and enhanced arachidonic acid (AA) release in P388D1 macrophages, and bacterial lipopolysaccharide (LPS) increased the responsiveness of these cells to PAF. In contrast with PLD, PLA2 activation in P388D1 macrophages was found to take place independently of PKC. This was supported by the following evidence: (i) PMA neither induced AA release nor enhanced the PAF response; (ii) inclusion of PMA along with LPS during priming did not have any effect on PAF-stimulated AA release; (iii) down-regulation of PMA-activatable PKC isoforms by chronic treatment with the phorbol ester had no effect on the PAF response; and (iv) the PKC inhibitor staurosporine did not alter the PAF-induced AA release. The present study provides an example of cells in which the direct activation of PKC by phorbol esters does not lead to a primed and/or enhanced AA release. As a unique example in which PKC activation is neither necessary nor sufficient for AA release to occur, this now allows study of the separate and distinct roles for PLD and PLA2 in signal-transduction processes. This has hitherto been difficult to achieve because of the lack of specific inhibitors of these two phospholipases.

scholarly journals Potassium channels modulate canine pulmonary vasoreactivity to protein kinase C activation

1999 ◽  
Vol 277 (3) ◽  
pp. L558-L565 ◽  
Author(s):  
Scott A. Barman
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Channel Blocker ◽  
Pressor Response ◽  
Vascular Occlusion ◽  
Membrane Depolarization ◽  
Delayed Rectifier ◽  
Vascular Compliance ◽  
Kinase C ◽  
Pkc Activation

The role of Ca2+-activated K+-channel, ATP-sensitive K+-channel, and delayed rectifier K+-channel modulation in the canine pulmonary vascular response to protein kinase C (PKC) activation was determined in the isolated blood-perfused dog lung. Pulmonary vascular resistances and compliances were measured with vascular occlusion techniques. The PKC activators phorbol 12-myristate 13-acetate (PMA; 10−7 M) and thymeleatoxin (THX; 10−7 M) significantly increased pulmonary arterial and pulmonary venous resistances and pulmonary capillary pressure and decreased total vascular compliance by decreasing both microvascular and large-vessel compliances. The Ca2+-activated K+-channel blocker tetraethylammonium ions (1 mM), the ATP-sensitive K+-channel inhibitor glibenclamide (10−5 M), and the delayed rectifier K+-channel blocker 4-aminopyridine (10−4 M) potentiated the pressor response to both PMA and THX on the arterial and venous segments and also further decreased pulmonary vascular compliance. In contrast, the ATP-sensitive K+-channel opener cromakalim (10−5 M) attenuated the vasoconstrictor effect of PMA and THX on both the arterial and venous vessels. In addition, membrane depolarization by 30 mM KCl elicited an increase in the pressor response to PMA. These results indicate that pharmacological activation of PKC elicits pulmonary vasoconstriction. Closure of the Ca2+-activated K+ channels, ATP-sensitive K+ channels, and delayed rectifier K+ channels as well as direct membrane depolarization by KCl potentiated the response to PMA and THX, indicating that K+ channels modulate the canine pulmonary vasoconstrictor response to PKC activation.

Mechanisms of action of endothelin 1 in maturing rabbit airway smooth muscle

1991 ◽  
Vol 260 (6) ◽  
pp. L434-L443 ◽  
Author(s):  
M. M. Grunstein ◽  
S. M. Rosenberg ◽  
C. M. Schramm ◽  
N. A. Pawlowski
Keyword(s):  
Smooth Muscle ◽  
Mechanisms Of Action ◽  
Maximal Response ◽  
Endothelin 1 ◽  
Kinase C ◽  
Adult Tissues ◽  
Age Dependent ◽  
Dose Dependent ◽  
Higher Doses ◽  
Pkc Activation

Maturational differences in the effects and mechanisms of action of endothelin 1 (ET-1) on airway contractility were investigated in tracheal smooth muscle (TSM) segments isolated from 2-wk-old and adult rabbits. In TSM under passive tension, ET-1 elicited dose-dependent contractions, with a potency of action that was significantly greater (P less than 0.001) in the 2-wk-old vs. adult tissues (i.e., mean +/- SE - log 50% of maximal response values: 8.59 +/- 0.17 vs. 7.79 +/- 0.15 - log M, respectively). In TSM half-maximally contracted with acetylcholine (ACh), however, ET-1 elicited dual and opposing dose-dependent effects. At lower doses (less than or equal to 10(-9) M), ET-1 induced TSM relaxation that was significantly greater in the adult vs. 2-wk-old TSM segments (i.e., approximately 100 vs. 26.5% decrease in active tension, respectively). The relaxant responses were associated with significantly enhanced (P less than 0.001) ET-1-induced release of prostaglandins E2 and I2 in the adult tissues. At higher doses (greater than 10(-9) M), ET-1 induced TSM contractions that were 1) attenuated to a relatively greater extent by the Ca2+ channel blocker, nifedipine (10(-5) M) in the 2-wk-old tissues and 2) associated with significantly (P less than 0.001) enhanced ET-1-stimulated accumulation of inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] in the immature TSM. Moreover, the TSM contractions were inhibited by the protein kinase C (PKC) antagonist, H-7, and the latter effect was more potent in the immature TSM. Collectively, these findings demonstrate that ET-1 exerts a potent duality of action in rabbit TSM which varies significantly with maturation, wherein 1) age-dependent differences in airway relaxation are associated with changes in the evoked release of bronchodilatory prostaglandins and 2) maturational differences in airway contraction are associated with changes in Ins(1,4,5)P3 accumulation and extracellular Ca2+ mobilization, coupled to differences in PKC activation.

scholarly journals Protein kinase C activity is not involved in N-formylmethionyl-leucyl-phenylalanine-induced phospholipase D activation in human neutrophils, but is essential for concomitant NADPH oxidase activation: studies with a staurosporine analogue with improved selectivity for protein kinase C

1993 ◽  
Vol 292 (3) ◽  
pp. 781-785 ◽  
Author(s):  
G C Kessels ◽  
K H Krause ◽  
A J Verhoeven
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Cyclic Amp ◽  
Nadph Oxidase ◽  
Phospholipase D ◽  
Respiratory Burst ◽  
Feedback Inhibition ◽  
Human Neutrophils ◽  
Kinase C ◽  
Pkc Activation

Stimulation of human neutrophils by the receptor agonist N-formylmethionyl-leucyl-phenylalanine (fMLP) results in a respiratory burst, catalysed by an NADPH oxidase. Concomitantly, phospholipase D (PLD) is activated. To investigate the role of protein kinase C (PKC) in these neutrophil responses, we have compared the effects of staurosporine and a structural analogue of staurosporine (cgp41251), that reflects a higher selectivity towards PKC [Meyer, Regenass, Fabbro, Alteri, Rösel, Müller, Caravatti and Matter (1989) Int. J. Cancer 43, 851-856]. Both staurosporine and cgp41251 dose-dependently inhibited the production of superoxide induced by phorbol 12-myristate 13-acetate (PMA). Both compounds also caused inhibition of the fMLP-induced respiratory burst, but with a lower efficacy during the initiation phase of this response. This latter observation cannot be taken as evidence against PKC involvement in the activation of the respiratory burst, because pretreatment of neutrophils with ionomycin before PMA stimulation also results in a lower efficacy of inhibition. Activation of PLD by fMLP was enhanced in the presence of staurosporine, but not in the presence of cgp41251. Enhancement of PLD activation was also observed in the presence of H-89, an inhibitor of cyclic-AMP-dependent protein kinase (PKA). Both staurosporine and H-89 reversed the dibutyryl-cyclic-AMP-induced inhibition of PLD activation, whereas cgp41251 was without effect. These results indicate that the potentiating effect of staurosporine on PLD activation induced by fMLP does not reflect a feedback inhibition by PKC activation, but instead a feedback inhibition by PKC activation. Taken together, our results indicate that in human neutrophils: (i) PKC activity is not essential for fMLP-induced activation of PLD; (ii) PKC activity does play an essential role in the activation of the respiratory burst by fMLP, other than mediating or modulating PLD activation; (iii) there exists a negative-feedback mechanism on fMLP-induced PLD activation by concomitant activation of PKA.

scholarly journals Participation of protein kinase C in endothelin-1-induced contraction in rat aorta: studies with a new tool, calphostin C

1992 ◽  
Vol 107 (2) ◽  
pp. 282-287 ◽  
Author(s):  
Hiroyuki Shimamoto ◽  
Yoriko Shimamoto ◽  
Chiu-Yin Kwan ◽  
Edwin E. Daniel
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Rat Aorta ◽  
Endothelin 1 ◽  
Calphostin C ◽  
Kinase C

scholarly journals Translocation-independent activation of protein kinase C by platelet-activating factor, thrombin and prostacyclin. Lack of correlation with polyphosphoinositide hydrolysis in rabbit platelets

1990 ◽  
Vol 267 (3) ◽  
pp. 689-696 ◽  
Author(s):  
H Salari ◽  
V Duronio ◽  
S Howard ◽  
M Demos ◽  
S L Pelech
Keyword(s):  
Protein Kinase C ◽  
Platelet Aggregation ◽  
Protein Kinase ◽  
Platelet Activating Factor ◽  
Kinase C ◽  
Camp Analogue ◽  
Rabbit Platelets ◽  
Rapid Hydrolysis ◽  
Hydrolysis Of ◽  
Pkc Activation

The relationship between polyphosphoinositide hydrolysis and protein kinase C (PKC) activation was explored in rabbit platelets treated with the agonists platelet-activating factor (PAF), thrombin and 12-O-tetradecanoylphorbol 13-acetate (TPA), and with the anti-aggregant prostacyclin (PGI2). Measurement of the hydrolysis of radiolabelled inositol-containing phospholipids relied upon the separation of the products [3H]inositol mono-, bis- and tris-phosphates by Dowex-1 chromatography. PKC activity, measured in platelet cytosolic and Nonidet-P40-solubilized particulate extracts that were fractionated by MonoQ chromatography, was based upon the ability of the enzyme to phosphorylate either histone H1 in the presence of the activators Ca2+, diacylglycerol and phosphatidylserine, or protamine in the absence of Ca2+ and lipid. Treatment of platelets for 1 min with PAF (2 nM) or thrombin (2 units/ml) led to the rapid hydrolysis of inositol-containing phospholipids, a 2-3-fold stimulation of both cytosolic and particulate-derived PKC activity, and platelet aggregation. Exposure to TPA (200 nM) for 5 min did not stimulate formation of phosphoinositides, but translocated more than 95% of cytosolic PKC into the particulate fraction, and induced a slower rate of aggregation. PGI2 (1 microgram/ml) did not enhance phosphoinositide production, and at higher concentrations (50 micrograms/ml) it antagonized the ability of PAF, but not that of thrombin, to induce inositol phospholipid turnover, even though platelet aggregation in response to both agonists was blocked by PGI2. On the other hand, PGI2 alone also appeared to activate (by 3-5-fold) cytosolic and particulate PKC by a translocation-independent mechanism. The activation of PKC by PGI2 was probably mediated via cyclic AMP (cAMP), as this effect was mimicked by the cAMP analogue 8-chlorophenylthio-cAMP. It is concluded that this novel mechanism of PKC regulation by platelet agonists may operate independently of polyphosphoinositide turnover, and that activation of cAMP-dependent protein kinase represents another route leading to PKC activation.

Voltage-independent calcium channels mediate lipopolysaccharide-induced hyporeactivity to endothelin-1 in the rat aorta

2009 ◽  
Vol 296 (5) ◽  
pp. H1408-H1415 ◽  
Author(s):  
Mohammed S. H. El-Awady ◽  
Sergey V. Smirnov ◽  
Malcolm L. Watson
Keyword(s):  
Rho Kinase ◽  
Treated Group ◽  
Rat Aorta ◽  
Receptor Expression ◽  
Initial Increase ◽  
Expression Levels ◽  
Endothelin 1 ◽  
Kinase C ◽  
Male Wistar Rats ◽  
Relaxation Responses

The roles of intracellular calcium concentration ([Ca2+]i) and Ca2+ sensitization in lipopolysaccharide (LPS)-induced vascular smooth muscle (VSM) hyporesponsiveness are incompletely understood. To investigate these roles, contraction responses to endothelin-1 (ET-1) and 80 mM KCl; relaxation responses to nifedipine; the expression levels of mRNAs of ET-1 and its receptors (ETA or ETB); the expression levels of protein kinase C (PKC) and phosphorylation of Rho kinase (ROKα), CPI-17, and myosin phosphatase target subunit-1 (MYPT1); and changes in aortic VSM cell [Ca2+]i were measured in LPS-treated aortic rings from male Wistar rats (250–300 g). LPS (10 μg/ml, 20 h) decreased contraction induced by ET-1 (0.3–100 nM) or 80 mM KCl. LPS-induced hypocontractility was not observed in the absence of external Ca2+, but LPS-treated aorta remained hypocontractile on subsequent stepwise restoration of extracellular Ca2+ (0.01–10 mM). Vascular relaxation to nifedipine; mRNA expression levels of ET-1, ETA, or ETB; protein expression levels of PKC; and phosphorylation levels of ROKα, CPI-17, and MYPT1 were not affected by LPS. In isolated aortic VSM cells, ET-1 caused a transient initial increase in [Ca2+]i, followed by a maintained tonic increase in [Ca2+]i, which was decreased by LPS pretreatment and was dependent on external Ca2+. Subsequent restoration of extracellular Ca2+ increased [Ca2+]i, but this increase was lower in the LPS-treated group. This difference in response to extracellular Ca2+ addition was not affected by diltiazem, but was abolished by SKF-96365. Therefore, LPS induces hyporeactivity to ET-1 in rat aorta that depends on external Ca2+ influx through non-voltage-operated Ca2+ channels, but not on ET-1 receptor expression or Ca2+ sensitization.

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