Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

1. Single smooth muscle cells obtained by enzymic dispersion of the longitudinal muscle layer of rabbit jejunum were held under voltage clamp using patch pipettes and membrane currents measured. The effects of carbachol or caffeine applied externally were examined in cells dialysed with normal pipette solutions or with a solution containing heparin (which blocks receptors for D-myo-inositol 1,4,5-trisphosphate, InsP3), guanosine 5-O-(gamma-thio)triphosphate (GTP gamma S) or guanosine 5-O-(beta-thio)diphosphate (GDP beta S). 2. Outward current in response to application of carbachol or caffeine was considered to represent the opening of calcium-activated potassium channels in response to a localized rise in the free ionized calcium concentration occasioned by the rapid discharge of stored calcium (Ca) by these agents. 3. Heparin included in the pipette solution blocked outward current to muscarinic receptor activation by carbachol but not that to caffeine, suggesting that receptor-evoked discharge of stored cellular Ca is caused by InsP3 action. However, heparin did not affect muscarinic-receptor inward current. 4. After dialysis with 0.1-0.5 mM-GTP gamma S, carbachol inward current was evoked in two out of three of the cells; after dialysis with 0.1-0.2 mM-GTP gamma S for an average of 7.7 min it was 80% of the normal response; after dialysis for an average of 8.6 min with 0.5 mM-GTP gamma S it was 31% of the normal response. In contrast, 0.1 mM-GTP gamma S reduced caffeine outward current by 93% after an average 4.5 min dialysis and spontaneous transient outward currents (STOCs) were abolished in 2.9 min on average. 5. Carbachol inward current (at -40 or -50 mV) and carbachol outward current (at 0 mV) in responding cells were reduced only by half after 8-10 min dialysis with 1 mM-GDP beta S which has been shown in portal vein cells to antagonize the depletion of Ca stores by intracellular GTP gamma S (Komori & Bolton, 1989). After 8-10 min dialysis with 5 mM-GDP beta S outward current was 27% of normal. However, if GDP beta S was present, outward current generally could not be evoked by a second application of carbachol. 6. The discharge of Ca stores by dialysis with 0.1 mM-GTP gamma S was prevented completely by heparin included in the pipette solution, suggesting that activation of a G-protein associated with phospholipase C (PLC) enzyme accelerates PLC activity. InsP3 production and depletion of Ca stores.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Role of G-proteins in muscarinic receptor inward and outward currents in rabbit jejunal smooth muscle. 212 Apr 27

Receptor-mediated arachidonic acid release and its relationship to phospholipase A2 and phospholipase C activation were investigated in Chinese hamster ovary cells transfected with and expressing the m5 muscarinic receptor. Carbachol, a muscarinic receptor agonist, stimulated the release of arachidonic acid and inositol phosphates with similar potencies. In addition, carbachol and the phorbol ester, phorbol-12-myristate, 13-acetate (PMA), stimulated protein kinase C (PKC) activity. PMA potentiated the carbachol-stimulated release of arachidonic acid, but had no effect on release of inositol phosphates. Long-term preincubation with PMA or carbachol inhibited PKC activity and prevented carbachol-stimulated release of arachidonic acid, but not inositol phosphates, suggesting that release of arachidonic acid, but not release of inositol phosphates, required activation of PKC. Carbachol stimulated the release of [3H]lysophosphatidylcholine from [3H]choline prelabeled cells, suggesting that phospholipase A2 was involved in the release of arachidonic acid. The role of calcium in carbachol-stimulated release of arachidonic acid was also investigated. Carbachol stimulated a transient followed by a sustained increase in intracellular calcium. In the absence of extracellular calcium, the transient rise in intracellular calcium was maintained but the sustained increase in intracellular calcium and the release of arachidonic acid were abolished. Carbachol stimulated a sustained influx of 45Ca++. We conclude that the combined effect of PKC activation and sustained elevation of intracellular calcium, from an extracellular source, is essential for m5 muscarinic receptor activation of phospholipase A2.
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PMID:A transfected m5 muscarinic acetylcholine receptor stimulates phospholipase A2 by inducing both calcium influx and activation of protein kinase C. 212 20

Brain cortex membranes labeled with [14C]arachidonic acid were used as the source of substrate and enzyme for the assay of arachidonic acid (AA) liberation. A significant amount of AA was released Ca2(+)-independently, mainly from phosphatidic acid, polyphosphoinositides and phosphatidylserine. Quinacrine, inhibitor of phospholipase A2 (PLA2), suppressed AA release by 60% and neomycin, inhibitor of phospholipase C (PLC) by about 30%. Both inhibitors applied together have an additive effect. Physiological calcium level elevated AA liberation by 50%, whereas 2 mM calcium enhanced this process by a further 30%. Carbachol, exclusively in the presence of calcium, activated AA release selectively from phosphatidylinositol and diglycerides. We suggest that Ca2(+)-independent PLA2 and PLC play an important role in AA liberation, and that physiological increments of calcium may have serious implications.
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PMID:Regulation of arachidonic acid release by enzyme(s) of rat brain cortex. 212 31

The muscarinic acetylcholine receptor-mediated inhibition of adenylate cyclase was studied in slices of guinea-pig cerebral cortex under normal and depolarizing conditions. Carbachol (1 mM) inhibited basal and isoproterenol (50 microM)-stimulated cyclic AMP (cAMP) accumulation by 20% and 25%, respectively, in normal Krebs-Ringer bicarbonate buffer solution (KRB). High-K+ medium (42 mM K+) increased cAMP accumulation to 330% of the basal level and abolished the inhibitory effect of carbachol. It also abolished the effect of morphine, an agonist of opioid receptors. Low-Ca2+ KRB or the presence of the Ca2+ channel blocker nifedipine, counteracted the effect of high K+ and restored the inhibitory effect of carbachol on the cAMP level. Pretreatment of slices with W-7 or trifluoperazine, two calmodulin antagonists, had the same effect as low Ca2+ or nifedipine on high-K(+)-stimulated cAMP accumulation and caused reappearance of the inhibitory effects of carbachol and morphine. On the contrary, H-7, an inhibitor of protein kinase C, and neomycin, an inhibitor of phospholipase C, had no significant effect on high-K(+)-induced phenomena and did not restore the effect of carbachol. These data suggest that the Ca2(+)-calmodulin system activated by membrane depolarization regulates the cAMP level directly and also by affecting the receptor-mediated process in nerve cells.
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PMID:Disappearance in high-K+ medium of receptor-mediated inhibition of adenylate cyclase in guinea-pig cortical slices. 217 2

In alpha-toxin-permeabilized guinea-pig ileum smooth muscle, a step increase in Ca2+ caused a rapid rise in force and myosin light chain (LC20) phosphorylation, followed by their spontaneous decline to a low steady level even though Ca2+ remained constant. Carbachol resensitized the muscles to Ca2+, causing an increase in both the steady state force and LC20 phosphorylation at constant Ca2+. In beta-escin permeabilized preparations, calmodulin and okadaic acid converted the phasic responses to Ca2+ to more tonic ones. We conclude that Ca2(+)-sensitivity of force is modulated through changes in LC20 kinase/phosphatase activity ratio by Ca2+ itself (desensitization) and by agonists (sensitization).
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PMID:Desensitization and muscarinic re-sensitization of force and myosin light chain phosphorylation to cytoplasmic Ca2+ in smooth muscle. 224 12

Carbachol, histamine and bradykinin activate phospholipase C in pertussis toxin-insensitive manner in human astrocytoma cells. Pretreatments of the cells with these agonists resulted in the reduction of GTP gamma S-induced accumulation of inositol phosphates in membrane preparations. Treatment of cells with carbachol mobilized GTP gamma S binding activities as well as muscarinic receptors from heavy membrane fraction to light fraction, reflecting from an agonist-induced desensitization. The treatment of the cells with agonists reduced a 32 kDa GTP binding protein in heavy membrane fraction, determined by a photoaffinity labeling with [35S]GTP gamma S. The data suggest that the 32 kDa GTP binding protein is involved in desensitization by agonists which activate phospholipase C in human astrocytoma cells.
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PMID:GTP gamma S binding activities were reduced in heavy membrane fraction during desensitization by Ca-mobilizing agonists in human astrocytoma cells. 250 Jun 83

Activation of rat uterine myometrial muscarinic receptors with a variety of agonists results in increased phosphatidylinositol metabolism. Activation with carbachol is concentration- and time-dependent and is most apparent by following the accumulation of inositol monophosphate although there are small but significant increases of inositol bisphosphate and inositol trisphosphate. Carbachol stimulation of phospholipid turnover is greatest in the upper third of the uterus. The carbachol-induced increase of inositol monophosphate is antagonized by atropine and by the selective M-3 muscarinic receptor antagonist 4-diphenylacetoxy-N-methylpiperidine methobromide. Pirenzepine, a selective M-1 receptor antagonist is less active, whereas gallamine and 11-2[[(diethylamino)methyl]-1-piperidinyl]acetyl]-5, 11-dihydro-6H-pyrido[2,3-b][1,4]benzodiazepine-6-one, selective M-2 receptor antagonists, are minimally effective suggesting that muscarinic M-3 receptors modulate phospholipid turnover in the rat myometrium. Displacement of tritium-quinuclidinyl benzilate binding by muscarinic antagonists also supports the presence of M-3 receptors in the uterus. Incubation with phorbol 12, 13-dibutyrate significantly reduced the accumulation of inositol monophosphate induced by carbachol implying that protein kinase C might modulate the responsiveness of the M-3 receptors in the rat uterus. Our results suggest that the intracellular concentration of calcium required for the contraction of the rat myometrium may be modulated, in part, through M-3 muscarinic receptors coupled to phospholipase C-activated turnover of phosphoinositides.
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PMID:Pharmacological characterization of the muscarinic receptors mediating phosphoinositide hydrolysis in rat myometrium. 254 Mar 9

We examined the relationship between phosphatidylcholine (PC) hydrolysis, phosphoinositide hydrolysis, and diacylglycerol (DAG) formation in response to muscarinic acetylcholine receptor (mAChR) stimulation in 1321N1 astrocytoma cells. Carbachol increases the release of [3H]choline and [3H]phosphorylcholine ([3H]Pchol) from cells containing [3H]choline-labeled PC. The production of Pchol is rapid and transient, while choline production continues for at least 30 min. mAChR-stimulated release of Pchol is reduced in cells that have been depleted of intracellular Ca2+ stores by ionomycin pretreatment, whereas choline release is unaffected by this pretreatment. Phorbol 12-myristate 13-acetate (PMA) increases the release of choline, but not Pchol, from 1321N1 cells, and down-regulation of protein kinase C blocks the ability of carbachol to stimulate choline production. Taken together, these results suggest that Ca2+ mobilization is involved in mAChR-mediated hydrolysis of PC by a phospholipase C, whereas protein kinase C activation is required for mAChR-stimulated hydrolysis of PC by a phospholipase D. Both carbachol and PMA rapidly increase the formation of [3H]phosphatidic acid ([3H]PA) in cells containing [3H]myristate-labeled PC. [3H]Diacylglycerol ([3H]DAG) levels increase more slowly, suggesting that the predominant pathway for PC hydrolysis is via phospholipase D. When cells are labeled with [3H]myristate and [14C]arachidonate such that there is a much greater 3H/14C ratio in PC compared with the phosphoinositides, the 3H/14C ratio in DAG and PA increases with PMA treatment but decreases in response to carbachol. By analyzing the increase in 3H versus 14C in DAG, we estimate that the DAG that is formed in response to PMA arises largely from PC. Muscarinic receptor activation also causes formation of DAG from PC, but approximately 20% of carbachol-stimulated DAG appears to arise from hydrolysis of the phosphoinositides.
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PMID:Muscarinic receptor activation of phosphatidylcholine hydrolysis. Relationship to phosphoinositide hydrolysis and diacylglycerol metabolism. 254 33

Guanine nucleotides have been shown to stimulate phosphoinositide breakdown in brain membranes, but no potentiation of such an effect by agonist was demonstrated. We have studied the effect of carbachol and histamine on guanosine 5'-[gamma-thio]triphosphate (GTP[S]) stimulation of inositol phosphates formation in [3H]inositol-labelled rat brain cortical membranes. In this preparation, GTP[S] enhancement of phosphoinositide hydrolysis required the presence of MgATP and low Ca2+ concentration (100 nM). Carbachol potentiation of the GTP[S] effect was only observed when 1 mM-deoxycholate was also added. Under these conditions, stimulated production of [3H]inositol phosphates was linear for at least 15 min, and [3H]inositol bisphosphate [( 3H]IP2) accounted for approx. 80%, whereas the amount of [3H]inositol trisphosphate [( 3H]IP3) was very low. Stimulation by GTP[S] was concentration-dependent (half-maximal effect at 0.86 microM), and its maximal effect (815% over basal) was increased by 1 mM-carbachol (1.9-fold) and -histamine (1.7-fold). Both agonists decreased the slope index of the GTP[S] concentration/effect curve to values lower than unity, suggesting the appearance of some heterogeneity in the population of guanine-nucleotide-binding proteins (G-proteins) involved. The carbachol and histamine effects were also concentration-dependent, and were inhibited by atropine and mepyramine respectively. Fluoroaluminate stimulated phosphoinositide hydrolysis to a higher extent than GTP[S] plus carbachol, and these stimulations were not additive, indicating that the same polyphosphoinositide phospholipase C-coupled G-protein mediates both effects.
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PMID:Carbachol and histamine stimulation of guanine-nucleotide-dependent phosphoinositide hydrolysis in rat brain cortical membranes. 254 64

Exogenously added phosphatidylinositol 4,5-bisphosphate (PtdInsP2) is rapidly associated with cerebral-cortical membranes. Substrate association with membranes was promoted by Mg2+, but inhibited by bivalent chelators. Once associated with the membrane, the PtdInsP2 was resistant to displacement by EDTA. The apparent phospholipase C activity was dependent on the degree of association of substrate with membranes. After preincubation of membranes with substrate, PtdInsP2 hydrolysis was independent of the incubation volume, indicating that substrate and membrane-associated phospholipase C were not independently diluted. Hydrolysis of the membrane-associated substrate was stimulated by Ca2+, guanosine 5'-[beta gamma-imido]triphosphate (p[NH]ppG), guanosine 5'[gamma-thio]triphosphate and carbachol in the presence of p[NH]ppG. Carbachol in the absence of guanine nucleotides, GDP, GTP, ATP and pyrophosphate was ineffective. These results demonstrate that exogenously added PtdInsP2 substrate is rapidly associated with membranes and hydrolysed by a phospholipase C whose activity is regulated by guanine nucleotides and agonist in the presence of guanine nucleotides. Use of exogenously added substrate for studies on the regulation of membrane phospholipase C requires consideration as to possible effects of incubation conditions on the partitioning of substrate into membranes.
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PMID:Interaction of cerebral-cortical membranes with exogenously added phosphatidylinositol 4,5-bisphosphate. Effects on measured phospholipase C activity. 254 69


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