Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.4.3 (phospholipase C)
 18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The stimulation of phospholipase D (PLD) activity by endothelin-1 (ET1) was investigated in rabbit iris sphincter prelabelled with [3H]myristic acid. In the presence of 0.5% ethanol, ET1 caused a time- and dose-dependent increase in the production of [3H]phosphatidylethanol ([3H]PEt). Within 30 s the peptide increased PEt formation by 30% and after 5 min increased it by 140%. The EC50 value for ET1-stimulated PEt formation was found to be 30 nM. This value is appreciably lower than the EC50 we previously obtained for ET1-induced inositol trisphosphate production (45 nM), but considerably higher than that for arachidonic acid release (1 nM). PEt formation was significantly stimulated by prostaglandin F20, phorbol 12,13-dibutyrate (PDBu), chloroform, A23187 and A1F4-, but it was not affected by carbachol or the platelet-activating factor. PDBu-stimulated PEt formation was blocked by staurosporine and it was not potentiated by A23187. Staurosporine had no effect on ET1-stimulated PEt formation. Our data indicate that ET1 stimulation of PLD occurs independently of protein kinase C activation, phospholipase C activation and intracellular Ca2+ mobilization, and phospholipase A2 activation. In this tissue the ET1 receptor is probably coupled to the three phospholipases through several G-proteins, and this appears to be species and receptor type specific.
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PMID:Activation of phospholipase D by endothelin-1 and other pharmacological agents in rabbit iris sphincter smooth muscle. 148 66

We have previously reported that endothelin-1 stimulates phospholipase C-induced hydrolysis of phosphatidylinositol-4,5-bisphosphate. Other signal transduction pathways that hydrolyze alternative phospholipids through phospholipase D may also mediate endothelin-stimulated cellular responses. We initially evaluated endothelin-dependent generation of 32P-phosphatidic acid as an indirect indication of phospholipase D activity in rat mesangial cells. Endothelin (10(-7) M) induced an elevation of phosphatidic acid that was maximal at 15 min and persisted upward of 60 min. Pretreatment with the diacylglycerol-kinase inhibitor, R59022, did not reduce formation of endothelin-stimulated 32P-phosphatidic acid, demonstrating that the sequential actions of phospholipase C/diacylglycerol kinase do not contribute to endothelin-stimulated phosphatidic acid formation. We next conclusively identified a role for phospholipase D in the generation of phosphatidic acid by assessing the formation of 3H-phosphatidylethanol from 3H-alkyl lyso glycerophosphocholine and exogenous ethanol. Endothelin stimulated 3H-alkyl phosphatidylethanol formation in the presence but not the absence of 0.5% ethanol. Also, endothelin induced a concomitant elevation of 3H-alkyl-phosphatidic acid that was significantly reduced when the cells were exposed to exogenous ethanol, reflecting the formation of phosphatidylethanol. In addition, endothelin stimulated the release of 3H-choline and 3H-ethanolamine, demonstrating that additional phospholipids may serve as substrates for phospholipase D. Phorbol esters and synthetic diglycerides mimicked the effects of endothelin to stimulate phospholipase D and inhibitors of protein kinase C significantly reduced endothelin-stimulated phospholipase D. In addition, endothelin did not stimulate phosphatidylethanol formation in protein kinase C down-regulated cells. The calcium ionophore, ionomycin, did not stimulate phospholipase D and mesangial cells pretreated with BAPTA to chelate cytosolic calcium did not show a diminished endothelin-stimulated phospholipase D. Thus these data demonstrate that mesangial cells possess a protein kinase C-regulated phospholipase D activity that can be stimulated with endothelin.
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PMID:Endothelin stimulates phosphatidic acid formation in cultured rat mesangial cells: role of a protein kinase C-regulated phospholipase D. 153 86

The effects of endothelin, a novel vasoconstrictive peptide, on the delayed rectifier K+ current (IK) were examined in single dialyzed cells from guinea pig ventricles. Either big endothelin or endothelin-1 enhanced IK at a dissociation constant of 2 nM with L-type Ca2+ current being unaffected. Under intracellular perfusion with pCa 7.6 solution, 3 nM big endothelin increased IK by 55 +/- 38.5%. Either pretreatment with 10 microM 1-(5-isoquinolinylsulfonyl)-2-methyl-piperazine (H 7) or a low Ca2+ [10 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) and minus CaCl2] internal solution diminished the enhancement. Preceding stimulation of protein kinase C (PKC) by 10-20 nM 12-O-tetradecanoylphorbol-13-acetate also reduced the degree of enhancement. When Na+ was eliminated from the solutions, endothelin increased IK distinctively in cells internally dialyzed with a low Ca2+ solution. This enhancement was not abolished by either pretreatment with H 7 or by removal of Ca2+ from the external perfusate but by increasing the internal EGTA concentration to 40 mM. Preincubation with ryanodine or internal perfusion with heparin also reduced the IK enhancement under Na(+)-free conditions. Intracellular application of 200 microM guanosine 5'-O-(3-thiotriphosphate) effectively attenuated the effect of endothelin. It is concluded that endothelin enhances IK via phospholipase C-mediated PKC activation and intracellular Ca2+ mobilization. GTP-binding protein is involved in these reactions.
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PMID:Endothelin enhances delayed potassium current via phospholipase C in guinea pig ventricular myocytes. 153 93

The mechanisms by which endothelin-1 (ET-1) and endothelin-3 (ET-3) stimulate Ca2+ mobilization were investigated in rat aortic smooth muscle cells. Both ET-1 and ET-3 potently stimulated mobilization of Ca2+ from intracellular stores, however only ET-1-stimulated Ca2+ mobilization appeared to occur as a consequence of an elevation in cellular inositol trisphosphate (IP3) concentration. Neomycin, an inhibitor of phospholipase C, inhibited both the increase in [3H]IP3 formation and the mobilization of Ca2+ induced by ET-1, however it did not affect Ca2+ mobilization induced by ET-3. Together these findings indicate that ET-1 stimulates Ca2+ mobilization via an increase in IP3, whereas the effect of ET-3 appears to be mediated by a separate, IP3-independent signalling pathway.
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PMID:Endothelin-1 and endothelin-3 stimulate calcium mobilization by different mechanisms in vascular smooth muscle. 155 May 75

The effects of the expression of the protein tyrosine kinase pp60v-src on endothelin- and thrombin-stimulated inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) production and calcium responses were investigated in Rat-1 fibroblasts. The ability of endothelin-1 to induce the accumulation of these second messengers was dramatically amplified by v-src transformation, with 6- and 3-fold enhancements of the peak Ins(1,4,5)P3 and peak calcium responses, respectively. In contrast, thrombin-dependent responses were slightly reduced following v-src transformation, demonstrating that the augmentation of endothelin-stimulated signal transduction is a selective effect. The magnitude of the stimulated accumulation of Ins(1,4,5)P3 presumably depends upon both the functional activation of phospholipase C to produce Ins(1,4,5)P3, and the activity of the enzymes that metabolize Ins(1,4,5)P3. Although the metabolism of Ins(1,4,5)P3 was strikingly altered by expression of pp60v-src, with a bias towards the production of higher inositol polyphosphates that is consistent with an activated Ins(1,4,5)P3 3-kinase, this change could not account for the marked increase in endothelin-stimulated signaling induced by v-src transformation. This suggests that an effect of pp60v-src is expressed at the level of the plasma membrane, through an interaction with one or more components in the receptor/guanine nucleotide binding protein (G protein)/phospholipase C system that transduces the endothelin signal into Ins(1,4,5)P3 production. Preparation of membranes from normal and v-src-transformed cells showed that, while there was no change in the number of high-affinity endothelin binding sites, the release of Ins(1,4,5)P3 in response to guanine nucleotides and endothelin-1 was significantly increased following v-src transformation. In contrast, the Ins(1,4,5)P3 responses to thrombin and high Ca2+ concentrations were unaffected by transformation. Thus the selective interactions within the G protein system that couples the endothelin receptor to phospholipase C are potential sites at which the v-src transformation process may act to amplify endothelin-dependent Ins(1,4,5)P3 production.
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PMID:Selective amplification of endothelin-stimulated inositol 1,4,5-trisphosphate and calcium signaling by v-src transformation of rat-1 fibroblasts. 155 85

We tested the hypothesis that increased systemic vascular resistance in spontaneously hypertensive rats may be secondary to enhanced phospholipase C activity in response to vasoconstrictor stimuli. Activation of phospholipase C by angiotensin II (Ang II), thromboxane A2, arginine vasopressin, and endothelin-1 was compared in cultured glomerular mesangial cells and mesenteric vascular smooth muscle cells taken from 13- to 14-week-old hypertensive and normotensive Wistar-Kyoto rats (blood pressure, 185 +/- 1 versus 135 +/- 2 mm Hg). Phospholipase C was assessed by measuring cytosolic free calcium and by the accumulation of radiolabeled inositol phosphates. Basal cytosolic calcium did not differ between mesangial cells taken from both strains but was greater in smooth muscle cells from hypertensive rats (210.1 +/- 8.2 versus 149.2 +/- 4.7 nM). The responsiveness of cytosolic calcium and inositol phosphate accumulation to Ang II was significantly enhanced in mesangial cells from hypertensive rats (10(-7) M Ang II: peak increase of calcium, 1,266 +/- 181 versus 603 +/- 93 nM; percent increment of inositol phosphates at 1 minute, 266 +/- 26 versus 98 +/- 10%). Vascular smooth muscle cells from hypertensive rats, when compared with normotensive rats, showed a similar augmentation of Ang II-stimulated intracellular calcium and inositol phosphates. Thromboxane A2-induced enhancement of intracellular calcium and inositol phosphate accumulation in vascular smooth muscle cells was also greater in hypertensive animals. However, the responses to vasopressin and endothelin in mesangial or vascular smooth muscle cells did not differ between the normotensive and hypertensive animals. There was no significant difference in Ang II receptor number and affinity between hypertensive- and normotensive-derived mesangial cells. We conclude that genetically increased blood pressure in rats may be secondary to enhanced post-receptor signaling in glomerular mesangial cells activated by Ang II and to enhanced signaling in vascular smooth muscle cells stimulated by either Ang II or thromboxane A2.
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PMID:Phospholipase C responses in cells from spontaneously hypertensive rats. 156 63

In isolated rat aorta, 72.7 mM KCI, 10 microM prostaglandin F2 alpha, 30 nM endothelin-1 and 1 microM norepinephrine increased muscle tension, cytosolic Ca++ concentration ([Ca++]i) and 20 kDa myosin light chain (MLC) phosphorylation. The levels of contractile tension and MLC phosphorylation at a given [Ca++]i were greatest in the presence of endothelin-1 followed by prostaglandin F2 alpha greater than norepinephrine greater than high K+. Verapamil inhibited the high K(+)-induced increments to their respective resting levels. Verapamil also almost completely inhibited the receptor agonist-induced increments in [Ca++]i and MLC phosphorylation, although a part of the contraction was not inhibited. Ethylene glycol bis(beta-aminoethyl ether)-N,N'-tetraacetic acid further decreased [Ca++]i and muscle tension, suggesting that a part of the contraction is regulated by [Ca++]i below a resting level. Receptor agonists induced sustained contraction in the absence of external Ca++ which was not followed by the increase in [Ca++]i or MLC phosphorylation. This contraction was followed by the increments in shortening velocity and stiffness. In the rabbit mesenteric artery permeabilized with Staphylococcus aureus, alpha-toxin, norepinephrine and endothelin-1 shifted the Ca(++)-tension curve to the left in the presence of GTP. From these results, it is suggested that high K(+)-induced sustained contraction of vascular smooth muscle is attributable to an increase in [Ca++]i followed by an increase in MLC phosphorylation. In addition to this fundamental mechanism, receptor agonists increase Ca+ sensitivity of MLC phosphorylation when [Ca++]i is higher than resting level resulting in a greater contraction than that induced by high K+ for a given increase in [Ca++]i.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Receptor agonists induce myosin phosphorylation-dependent and phosphorylation-independent contractions in vascular smooth muscle. 157 67

The effects of endothelin-1 and noradrenaline on phospholipase C activity in the rabbit isolated corpus cavernosum were investigated by measuring the accumulation of inositol phosphates. Both endothelin-1 and noradrenaline caused a time- and concentration-dependent increase in the accumulation of 3H-inositol phosphates in preparations prelabelled with 3H-myo-inositol. The reaction was slow in onset with no significant accumulation of 3H-inositol phosphates, including inositol trisphosphate, demonstrable during the first 15 minutes. At 60 minutes, the mean increases in 3H-inositol inositol phosphates induced by 3 x 10(-7) M endothelin-1 and 10(-3) M noradrenaline amounted to 341 and 530% of time-matched controls, respectively. However, when given at concentrations having the same contractile amplitude on rabbit corpus cavernosum, there was no difference in the amounts of 3H-inositol phosphates generated by endothelin-1 and noradrenaline. Prazosin (10(-6) M) significantly inhibited the stimulatory effect of noradrenaline on phosphoinositide hydrolysis. Pretreatment with 10(-6) M nimodipine did not reduce the increases in 3H-inositol phosphates induced by 3 x 10(-7) M endothelin-1 and 10(-3)M noradrenaline. Also in Ca(2+)-free medium, both agonists had significant stimulatory effects on phosphoinositide turnover, although under this condition, the responses were greatly reduced. The results suggest that exogenous endothelin-1 and noradrenaline activate phospholipase C in corpus cavernosum, and that this mechanism is partly independent of extracellular Ca2+. Considering the slow onset of action, phospholipase C activation is probably not directly involved in rapid contractile events, but may be of importance in the long-term regulation of penile smooth muscle tone.
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PMID:Phospholipase C activation by endothelin-1 and noradrenaline in isolated penile erectile tissue from rabbit. 159 12

The authors investigated the effects of endothelin-1 (ET1) on inositol trisphosphate (IP3) production, 1, 2-diacylglycerol (DAG) formation, measured as phosphatidic acid (PA), cAMP formation, and contraction in iris sphincter of different mammalian species. They found that ET1 is a potent agonist for IP3 production, DAG formation, and contraction in rabbit, dog, cat, and pig iris sphincters, and for cAMP formation in all species that were investigated--rabbit, dog, cat, pig, bovine, monkey, and human sphincters. In the bovine model, ET1 induced cAMP formation in a dose-dependent manner, with an EC50 of 28 nM. This is the first report that showed an effect of the peptide on the adenylate cyclase system. In rabbit sphincter, ET1 induced a significant increase in IP3 production by 30 sec and reached a 6-fold level more than control within 1 and 5 min. ET1-stimulated IP3 production is dose dependent with an EC50 of 45 nM, this value is about 100- and 56-fold lower than those we reported for substance P and carbachol, respectively. ET1 also increased 32P labeling of PA more than 6-fold; and in rabbit sphincter, ET1 is a more potent agonist in contracting the sphincter than in contracting the dilator (the EC50 values for sphincter and dilator were 46 and 120 nM, respectively). L-type Ca2+ channels are not involved in IP3- and contraction responses because several blockers of these channels did not affect the ET1-induced responses, implying that in the iris sphincter, ET1 elicits the physiologic response through the G protein activation of phospholipase C and/or adenylate cyclase and not through the activation of voltage-dependent Ca2+ channels.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Species differences in the effects of endothelin-1 on myo-inositol trisphosphate accumulation, cyclic AMP formation and contraction of isolated iris sphincter of rabbit and other species. 164 47

Endothelins are a novel group of potent vasoconstrictor peptides originally isolated from cultured porcine endothelial cells. We and others have previously reported the presence of endothelin receptors in the central nervous system, and this study was designed to further characterize endothelin receptors and their transduction mechanism in cultured neurohybrid NG108-15 cells. Specific binding of [125I]endothelin-1 to NG108-15 cells reached saturation within 60 min at 22 degrees C and was only partially reversible. Scatchard analysis of the saturation binding revealed the presence of one class of high-affinity binding sites with an apparent dissociation constant of 160 pM and a maximal binding capacity of 3.3 x 10(4) sites/cell. Unlabeled endothelin analogues competitively inhibited [125I]endothelin-1 binding to NG108-15 cells and the apparent dissociation constant values obtained from the competition curves correlated well with the EC50 values obtained for inducing elevation of intracellular free Ca2+ level. Endothelin stimulated phosphoinositide metabolism in a dose-dependent manner with an EC50 value of 5.4 nM for inositol trisphosphate formation. The protein kinase C-activator phorbol ester dose-dependently inhibited endothelin-induced phosphoinositide turnover and intracellular free Ca2+ increase, suggesting the involvement of protein kinase C in the regulation of endothelin-induced responses. Neither endothelin-induced phosphoinositide hydrolysis nor endothelin-induced increase in intracellular free Ca2+ were affected by pertussis toxin. These data indicate that endothelin receptors are present on NG108-15 cells and the G protein coupled to endothelin receptor for inducing activation of phospholipase C and increase of free intracellular Ca2+ is insensitive to pertussis toxin.
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PMID:Endothelin receptor binding and cellular signal transduction in neurohybrid NG108-15 cells. 166 18


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