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. The relationship between muscarinic receptor-mediated phosphatidylinositol 4,5-bisphosphate (PIP2) breakdown and the increase of intracellular Ca2+ ([Ca2+])i has been examined in canine cultured tracheal smooth muscle cells (TSMCs). 2. Addition of acetylcholine (ACh) and carbachol led to a 2-3 fold increase in [Ca2+]i over the resting level as determined by fura-2, with half-maximal stimulation (EC50) obtained at concentrations of 97 and 340 nM, respectively. Addition of the partial agonist, bethanechol, showed a smaller increase in PIP2 turnover and [Ca2+]i than did ACh or carbachol. 3. Addition of ACh or carbachol to TSMCs that had been prelabelled with [3H]-inositol led to the rapid (5-15 s) release of inositol mono, bis and trisphosphates IP1, IP2 and IP3. The time course of IP3 accumulation is correlated with the time course of the peak rise in [Ca2+]i. 4. Inclusion of EGTA lowered the resting [Ca2+]i and markedly reduced the extent of the agonist-induced rise in [Ca2+]i. When assayed under conditions similar to those used for the [Ca2+]i measurements, EGTA reduced the muscarinic agonist-stimulated inositol phosphates (IPs) accumulation. Conversely, ionomycin could stimulate IPs accumulation and elevate [Ca2+]i. The addition of Ca2+ (2.7-617 nM) to digitonin-permeabilized TSMCs directly stimulated IPs accumulation. 5. Both Ca2+ and guanosine-5'-O-(3-thiotriphosphate) (GTP gamma S) stimulated the formation of IPs in digitonin-permeabilized TSMCs prelabelled with [3H]-inositol. A further calcium-dependent increase in IPs accumulation was obtained by inclusion of either GTP gamma S or carbachol. The combined presence of carbachol and GTP gamma S elicited a synergistic effect on IPs accumulation, with half-maximal stimulation observed at approximately 8 nM free Ca2+.6. These results indicate that (i) the magnitude of the initial rise in [Ca2+], is directly related to the production of IPs and (ii) the phospholipase C-mediated PIP2 breakdown in TSMCs is sensitive to regulation by physiologically relevant concentrations of free Ca2+ ([Ca2+]f).
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PMID:Muscarinic regulation of cytosolic free calcium in canine tracheal smooth muscle cells: Ca2+ requirement for phospholipase C activation. 829 14

The cellular content of phosphatidic acid can increase in response to several agonists either by phosphorylation of diacylglycerol after phospholipase C-catalyzed hydrolysis of phospholipids or directly through activation of phospholipase D. Although previous findings indicated that the generation of phosphatidic acid was exclusively a means of regulation of the cellular concentration of diacylglycerol, more recent studies have indicated that phosphatidic acid may also directly regulate several cellular functions. Accordingly, the present study was performed to assess whether phosphatidic acid could stimulate cardiac phospholipase C in intact adult rabbit ventricular myocytes. The mass of inositol 1,4,5-trisphosphate [Ins (1,4,5)P3] was determined by a specific and sensitive binding protein assay and by direct mass measurement using anion exchange chromatography for separation of selected inositol phosphates and gas chromatography and mass spectrometry for quantification of inositol monophosphate (IP1), inositol bisphosphate (IP2), inositol trisphosphate (IP3), and inositol tetrakisphosphate (IP4). Phosphatidic acid (10(-9)-10(-6) M) elicited a rapid concentration-dependent increase in Ins (1,4,5)P3 accumulation, with the peak fourfold to fivefold increase at 30 seconds of stimulation; the concentration required for 50% of maximal stimulation was 4.4 x 10(-8) M. The time course of individual inositol phosphates indicated a successive increase in the mass of IP3, IP4, IP2, and IP1 in response to stimulation with phosphatidic acid. The production of Ins (1,4,5)P3 in response to phosphatidic acid was not altered in the absence of extracellular calcium or in the presence of extracellular EGTA (10(-3) M). Thus, these findings indicate that phosphatidic acid is a potent activator of inositol phosphate production in adult ventricular myocytes.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Phosphatidic acid stimulates inositol 1,4,5-trisphosphate production in adult cardiac myocytes. 843 93

Both gastrin and cholecystokinin (CCK) can stimulate pepsinogen release from chief cells, but controversy exists about the receptors or intracellular mediators involved. In the present study, we prepared isolated chief cells from guinea pig stomach (> 90% pure) to investigate the ability of gastrin and CCK to alter cell function. The COOH-terminal octapeptide of CCK (CCK-8) caused an eightfold increase in pepsinogen release (EC50, 54 nM). Both CCK-8 and gastrin increased inositol phosphates, with CCK-8 (1 microM) and gastrin (3 microM) causing a 40- and 14-fold increase in [3H]IP1, 10- and 6-fold for [3H]IP2, and 8- and 4-fold for [3H]IP3. CCK-8 caused a half-maximal increase in [3H]IP3 at 2 nM, and the dose-response curve was monophasic, whereas with gastrin the curve was biphasic, with an EC50 of the initial component (20% maximal) at 38 nM and the second component at 10 microM. L-364,718 (0.1 microM) inhibited the secondary increase seen with gastrin concentrations > 10 nM. The CCK-A-selective agonist A-71378 was 85-90% as efficacious as CCK-8 and was equally potent. With 0.1 microM L-364,718, A-71378 caused no increase in [3H]inositol phosphates until > 10 nM, whereas CCK-8 caused 15% of maximal increase at concentrations > 0.3 nM. Similar results were obtained with cytosolic calcium measured using fura-2 or on CCK-8- or gastrin-stimulated pepsinogen release. These results demonstrate that gastrin and CCK-8 can alter chief cell function by interacting with either a CCK-A or CCK-B/gastrin receptor. Both receptors are coupled to phospholipase C and cause changes in inositol phosphates, cytosolic calcium, and pepsinogen release; however, the intracellular amplification differs between the two receptor subtypes. Activation by CCK-related peptides of the CCK-A receptor subtype accounts for 85-90% of the maximal changes in cellular function, and activation of the CCK-B/gastrin receptor accounts for 10-20% of maximal changes.
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PMID:Gastrin and CCK activate phospholipase C and stimulate pepsinogen release by interacting with two distinct receptors. 847 56

The signalling mechanisms whereby high-density lipoproteins (HDL) and low-density lipoproteins (LDL) affect a number of cellular functions in fibroblasts are unclear. This study has analyzed the influence of HDL3 and LDL on the phosphatidylinositol specific phospholipase C pathway in human skin fibroblasts. Exposure of myo-[2-3H]-inositol prelabelled fibroblasts to HDL3 or LDL elicited major increases in IP1 and minor increases in IP2 and IP3 within 30 s. In fura-2 loaded suspended fibroblasts, HDL3 and LDL increased intracellular Ca2+ concentrations ([Ca2+]i) with comparable rapid, transient kinetics. The dose-profiles for HDL3- and LDL-induced increases in [Ca2+]i were also comparable, with half-maximally and maximally effective concentrations being approximately 15 micrograms/mL and approximately 50 micrograms/mL, respectively. HDL3- and LDL-induced increases in [Ca2+]i were diminished by approximately 60% (vs. control fibroblasts) in thapsigargin-pretreated fibroblasts, indicating that release of Ca2+ from intracellular pools is the major contributor toward lipoprotein-induced increases in [Ca2+]i. Pertussis toxin-pretreatment of cells completely abolished lipoprotein induced Ca(2+)-transient, indicating the involvement of a guanine nucleotide-binding protein in the signalling process. In [3H]-palmitic acid-prelabelled fibroblasts, both HDL3 and LDL were observed to stimulate production of DAG. Activation of protein kinase C (PKC) was analysed by determining the cytosol-to-membrane translocation of both enzymatic activity and immunoreactivity of specific PKC isoforms (alpha, delta, epsilon, and zeta). Stimulation with HDL3 and LDL evoked a rapid (within 2.5 min) translocation of PKC activity, with PKC alpha and PKC epsilon being the isoforms translocated. It is concluded that HDL3 and LDL acutely stimulate a phosphoinositide-specific phospholipase C pathway in human skin fibroblasts. However, the specific cell membrane events mediating this signal transduction remain to be further elucidated.
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PMID:High-density lipoprotein and low-density lipoprotein-mediated signal transduction in cultured human skin fibroblasts. 851 99

The beneficial effects of n-3 polyunsaturated fatty acids of fish oil in the prevention of fatal arrhythmias in myocardial ischemia were suggested to be at least in part mediated by a modulation of dihydropyridine-sensitive L-type calcium channels. As cardiac alpha 1-adrenoceptor stimulation has been suggested to have no significant effect on L-type calcium channels, the aim of this study using cultured neonatal rat cardiomyocytes was to investigate whether chronic n-3 polyunsaturated fatty acid exposure may have an influence on alpha 1-adrenoceptor-induced positive inotropic effects and induction of arrhythmias. Pretreatment of the rat cardiomyocytes for 3 days in the presence of the n-3 polyunsaturated fish oil-derived fatty acid docosahexaenoic acid (60 mumol/l) markedly decreased alpha 1-adrenoceptor-stimulated increase in contraction velocity and induction of arrhythmias. The increase in contraction velocity of the cardiomyocytes induced by the beta-adrenoceptor agonist isoprenaline was also markedly reduced by the n-3 fatty acid pretreatment. Basal contractile amplitude and spontaneous beating frequency of the cardiomyocytes were not significantly altered by the docosahexaenoic acid exposure. The pretreatment of the rat cardiomyocytes for 3 days in the presence of docosahexaenoic acid (60 mumol/l) decreased alpha 1-adrenoceptor-stimulated formation of the calcium-mobilizing second messenger IP3 and its metabolites IP2 and IP1 by 55%. The depression of IP3 formation by docosahexaenoic acid treatment was not mediated by a decreased uptake of myo-inositol into the cardiomyocytes nor by a decreased synthesis of phosphatidylinositol bisphosphate (PIP2), the substrate of phospholipase C. The level of glycerol-3-phosphate, an important substrate of the phosphoinositide cycle, was unaltered by the docosahexaenoic acid pretreatment. Receptor binding studies revealed that the dissociation constant and maximal binding capacity of the alpha 1-adrenoceptor antagonist (3H)prazosin was unchanged by the n-3 polyunsaturated fatty acid exposure. Beta-Adrenoceptor-and forskolin-stimulated adenylyl cyclase activities were not diminished by the docosahexaenoic acid pretreatment. Chronic exposure of the cardiomyocytes to the n-6 polyunsaturated fatty acid arachidonic acid (60 mumol/l) did neither significantly alter alpha 1-adrenoceptor-induced inositol phosphate formation nor alpha 1-adrenoceptor-stimulated increase in contraction velocity. The results presented show that chronic n-3 polyunsaturated fatty acid pretreatment of rat cardiomyocytes leads to a marked impairment of alpha 1-adrenoceptor-induced positive inotropic effects and induction of arrhythmias concomitant with a n-3 fatty acid-induced decrease in IP3 formation. This derangement of the phosphoinositide pathway by chronic n-3 fatty acid exposure may, thus, contribute to the beneficial effects of fish oil-derived fatty acids in the prevention of fatal arrhythmias in myocardial ischemia.
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PMID:Exposure to the n-3 polyunsaturated fatty acid docosahexaenoic acid impairs alpha 1-adrenoceptor-mediated contractile responses and inositol phosphate formation in rat cardiomyocytes. 885 87

The pungent sesquiterpenoid unsaturated dialdehydes polygodial and isovelleral, have previously been shown to increase the intracellular free calcium concentration [Ca2+]i in human neuroblastoma SH-SY5Y cells, partly by a release from intracellular Ca2+ stores, whereas the non-pungent compound epipolygodial, had no effect on the [Ca2+]i. In this study, we investigated the effect of isovelleral, polygodial and epipolygodial on inositol phosphate (IP) formation on the assumption that there might be a correlation between the release of intracellular Ca2+ and pungency of the compounds. It was found that polygodial induced IP mobilization in a concentration dependent way, whereas isovelleral had no effect on the IP formation in the SH-SY5Y cells. Phosphoinositide (PPI) turnover was activated by epipolygodial, but only at concentrations 40-fold higher than for polygodial, which emphasizes the importance of the correct stereometry in the dialdehyde configuration for the biological activity of polygodial. The polygodial-induced formation of IP1 was reduced by 71% under extracellular calcium-free conditions, which suggests feedback interactions between the IP formation and the increase in [Ca2+]i to account for a periodic activation of phospholipase C(PLC).
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PMID:Polygodial induces inositol phosphate turnover in human neuroblastoma SH-SY5Y cells. 890 37

The aim of this study was to assess the contribution of alpha 1-adrenoceptor subtypes to noradrenaline (NA)-induced inositol phosphate formation in rat striatum. In cross-chopped slices and in the presence of 10 mM LiCl, NA stimulated the accumulation of [3H]inositol phosphates. After 60-min incubation with 100 microM NA, [3H]IP1 was the major product detected (82 +/- 3% of total [3H]inositol phosphates). Best-fit values for the concentration-response curve for NA-induced [3H]IP1 accumulation yielded an EC50 of 9.4 +/- 1.1 microM, maximum effect of 210 +/- 3% of basal, and Hill coefficient (nH) of 1.1 +/- 0.1. Pre-treatment of the slices for 30 min with the alkylating agent chloroethylclonidine (100 microM) failed to decrease significantly the response to 100 microM NA. Inhibition curves for four alpha 1-antagonists, (+)-niguldipine, prazosin, WB-4101 and 5-methylurapidil (5-MU), best-fit to a single-site model with pKi values of 9.4 ((+)-niguldipine), 9.2 (prazosin and WB-4101) and 8.8 (5-MU). The putative alpha 1 D-selective antagonist BMY 7378 reduced the response to NA only partially (30 +/- 3% inhibition at 1 microM: pKi 7.24). NA-induced [3H]IP1 accumulation was significantly reduced (to 20 +/- 9% of controls) by Ca2+ removal and increased as the extracellular Ca2+ concentration was raised from nominally zero (no added Ca2+) to 1 mM Ca2+. NA-induced [3H]IP1 accumulation was reduced by both the non-selective Ca2+ channel blocker Ni2+ (58 +/- 3% inhibition at 2 mM) and nimodipine, an antagonist of L-type voltage-operated Ca2+ channels (77 +/- 4% inhibition at 3 microM). Taken together these results indicate that NA-induced inositol phosphate formation in striatal slices is mediated by activation of alpha 1A-adrenoceptors coupled to Ca2+ entry and Ca2+ activation of phospholipase C.
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PMID:Noradrenaline-induced inositol phosphate formation in rat striatum is mediated by alpha 1A-adrenoceptors. 902 8

The molecular mechanisms for the stimulation of inositol 1-phosphate (IP1) formation by vinconate were investigated using preparations of rat brain. Vinconate (10(-8)-10(-3) M) dose-dependently inhibited the binding of [3H]quinuclidinyl benzilate ([3H]QNB) to muscarinic acetylcholine receptors and its IC50 value for [3H]QNB binding was 1.7 x 10(-5) M. The rightward shift of carbachol displacement curve of [3H]QNB binding by GTP (10(-4) M) was completely abolished by vinconate (10(-5) M). Carbachol (10(-8)-10(-2) M) increased [3H]IP1 formation in a dose-dependent manner and the carbachol-induced [3H]IP1 formation was significantly accentuated by vinconate (10(-5) M). The enhancement of [3H]IP1 accumulation by vinconate was inhibited by approximately 50% in the presence of atropine (10(-5) M), although phentolamine and ketanserin had no effects on the stimulatory effect of vinconate on [3H]IP1 formation. Vinconate showed no alteration in the binding of [3H]guanosine 5'-(beta, gamma-imino) triphosphate ([3H]Gpp(NH)p) to the crude synaptic membranes. The enhancement of phosphatidylinositol 4,5-biphosphate (PIP2)-specific phospholipase C (PLC) activity by GTP was unaffected in the presence of 10(-3) M vinconate, whereas vinconate alone dose-dependently enhanced the activities of both PIP2-specific and cytosolic PLC. These results suggest that vinconate may induce the facilitation of phosphatidylinositide (PI) turnover via the stimulation of muscarinic acetylcholine receptors, the enhancement of coupling between muscarinic acetylcholine receptors and GTP-binding protein, and the direct activations of PIP2-specific and cytosolic PLC.
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PMID:Vinconate, a cognitive enhancer, and PI turnover-phospholipase C systems in the brain. 906 64

The rat urinary bladder possesses NK1, NK2 (but not NK3) and 'septide-sensitive' tachykinin receptors coupled to a phospholipase C. The present study performed with SR48968 (10(-6) M) to avoid any interaction of the tested peptides with NK2 receptors, indicates that substance P(6-11) (with a high potency), neurokinin A, neurokinin B and to a lesser extent neuropeptide K (with a lower potency) stimulate [3H]-inositol monophosphate ([3H]-IP1) formation in this tissue by acting on the 'septide-sensitive' tachykinin receptors. Substance P(6-11) had little affinity for NK1 binding sites and stimulated [3H]-IP1 formation with an EC50 value and a maximal amplitude similar to those of septide. As previously observed with septide, this maximal response of substance P(6-11) (insensitive to 10(-6) M SR48968) which was about three-fold that of substance P, was blocked by the NK1 receptor antagonist RP67580 and prevented by [Pro9]substance P (NK1 receptor agonist). Similarly, substance P and several substance P C-terminal fragments prevented the substance P(6-11)-evoked response. In addition, neurokinin A, neuropeptide K and neurokinin B induced SR48968-resistant responses which exhibited a maximal amplitude similar to that of substance P (6-11) and were blocked by RP67580 and totally or partially (neuropeptide K) prevented by [Pro9]substance P.
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PMID:Substance P(6-11) and natural tachykinins interact with septide-sensitive tachykinin receptors coupled to a phospholipase C in the rat urinary bladder. 924 21

Bombesin (Bn) receptor subtype 3 (BRS-3) is an orphan receptor that is a predicted member of the heptahelical G-protein receptor family and so named because it shares a 50% amino acid homology with receptors for the mammalian bombesin-like peptides neuromedin B (NMB) and gastrin-releasing peptide. In a recent targeted disruption study, in which BRS-3-deficient mice were generated, the mice developed obesity, diabetes, and hypertension. To date, BRS-3's natural ligand remains unknown, its pharmacology unclear, and cellular basis of action undetermined. Furthermore, there are few tissues or cell lines found that express sufficient levels of BRS-3 protein for study. To define the intracellular signaling properties of BRS-3, we examined the ability of [D-Phe6,beta-Ala11,Phe13, Nle14]Bn-(6-14), a newly discovered peptide with high affinity for BRS-3, and various Bn receptor agonists and antagonists to alter cellular function in hBRS-3-transfected BALB 3T3 cells and hBRS-3-transfected NCI-H1299 non-small cell lung cancer cells, which natively express very low levels of hBRS-3. This ligand stimulated a 4-9-fold increase in [3H]inositol phosphate formation in both cell lines under conditions where it caused no stimulation in untransfected cells and also stimulated an increase in [3H]IP1, [3H]IP2, and 3H]IP3. The elevation of [3H]IP was concentration-dependent, with an EC50 of 20-35 nM in both cell lines. [D-Phe6,beta-Ala11,Phe13,Nle14]Bn-(6-14) stimulated a 2-3-fold increase in [Ca2+]i, a 3-fold increase in tyrosine phosphorylation of p125(FAK) with an EC50 of 0.2-0.7 nM, but failed to either stimulate increases in cyclic AMP or inhibit forskolin-stimulated increases. None of nine naturally occurring Bn peptides or three synthetic Bn analogues reported to activate hBRS-3 did so with high affinity. No high affinity Bn receptor antagonists had high affinity for the hBRS-3 receptor, although two low affinity antagonists for gastrin-releasing peptide and NMB receptors, [D-Arg1,D-Trp7,9, Leu11]substance P and [D-Pro4,D-Trp7,9,10]substance P-(4-11), inhibited hBRS-3 receptor activation. The NMB receptor-specific antagonist D-Nal,Cys,Tyr,D-Trp,Lys,Val, Cys,Nal-NH2 inhibited hBRS-3 receptor activation in a competitive fashion (Ki = 0.5 microM). Stimulation of p125(FAK) tyrosine phosphorylation by hBRS-3 activation was not inhibited by the protein kinase C inhibitor, GF109203X, or thapsigargin, alone or in combination. These results show that hBRS-3 receptor activation increases phospholipase C activity, which causes generation of inositol phosphates and changes in [Ca2+]i and is also coupled to tyrosine kinase activation, but is not coupled to adenylate cyclase activation or inhibition. hBRS-3 receptor activation results in tyrosine phosphorylation of p125(FAK), and it is not dependent on activation of either limb of the phospholipase C cascade. Although the natural ligand is not a known bombesin-related peptide, the availability of [D-Phe6,beta-Ala11, Phe13,Nle14]Bn-(6-14), which functions as a high affinity agonist in conjunction with hBRS-3-transfected cell lines and the recognition of three classes of receptor antagonists including one with affinity of 0.5 microM, should provide important tools to assist in the identification of its natural ligand, the development of more potent selective receptor antagonists and agonists, and further exploration of the signaling properties of the hBRS-3 receptor.
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PMID:Ability of various bombesin receptor agonists and antagonists to alter intracellular signaling of the human orphan receptor BRS-3. 959 99


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