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)

Isolated islets from adult rats or obese hyperglycemic (ob/ob) mice were incubated with human recombinant interleukin 1 beta in order to study whether the acute effects of the cytokine on islet insulin release are associated with changes in islet phospholipase C activity, Ca2+ handling or protein phosphorylation. The cytokine stimulated insulin release both at low and high glucose concentrations during one hour incubations. In short-term incubations (less than 1 min) interleukin 1 beta did not affect the production of inositoltrisphosphate. Addition of interleukin 1 beta affected neither the cytoplasmic free Ca2+ concentration at rest nor that observed subsequent to stimulation with a high concentration of glucose. Furthermore, the endogenous protein kinase C activity, as visualized by immunoprecipitation of a 32P-labelled substrate for this enzyme, was not altered by interleukin 1 beta. Separation of 32P-labelled proteins by means of 2-dimensional gel electrophoresis failed to reveal any specific effects of the cytokine on the total protein phosphorylation activity. These results suggest that the stimulatory effects on insulin release exerted by interleukin 1 beta are not caused by acute activation of phospholipase C and protein kinase C or by an alteration of islet Ca2+ handling of the B-cells.
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PMID:Human interleukin 1 beta stimulates islet insulin release by a mechanism not dependent on changes in phospholipase C and protein kinase C activities or Ca2+ handling. 268 30

Glomerular inositol content and the turnover of polyphosphoinositides was reduced by 58% in 1-2 wk streptozotocin diabetic rats. Addition of inositol to the incubation medium increased polyphosphoinositide turnover in glomeruli from diabetic rats to control values. Despite the reduction in inositol content and polyphosphoinositide turnover, protein kinase C was activated in glomeruli from diabetic rats, as assessed by an increase in the percentage of enzyme activity associated with the particulate cell fraction. Total protein kinase C activity was not different between glomeruli from control and diabetic rats. Treatment of diabetic rats with insulin to achieve near euglycemia prevented the increase in particulate protein kinase C. Moreover, incubation of glomeruli from control rats with glucose (100-1,000 mg/dl) resulted in a progressive increase in labeled diacylglycerol production and in the percentage of protein kinase C activity which was associated with the particulate fraction. These results support a role for hyperglycemia per se in the enhanced state of activation of protein kinase C seen in glomeruli from diabetic rats. Glucose did not appear to increase diacylglycerol by stimulating inositol phospholipid hydrolysis in glomeruli. Other pathways for diacylglycerol production, including de novo synthesis and phospholipase C mediated hydrolysis of phosphatidylcholine or phosphatidyl-inositol-glycan are not excluded.
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PMID:Protein kinase C is activated in glomeruli from streptozotocin diabetic rats. Possible mediation by glucose. 270 28

Brush-border membrane vesicles were prepared from rat kidney cortex by Mg precipitation. Using quin2 (free acid), intravesicular [Ca2+] was found to be 44 microM and less than 300 nM when vesicles were incubated in 0.2 mM CaCl2 or Ca-free buffer, respectively. In Ca-loaded vesicles, the initial D-glucose uptake, measured in the presence of 150 mM Na+ and 0.1 mM D-glucose inward gradients, was reduced to 30% of the control uptake. This reduction persisted when the extra-vesicular Ca2+ was chelated by ethylene glycol-bis(beta-amino-ethyl ether)-N,N,N',N'-tetraacetic acid but was abolished in the presence of saturating concentrations of D-glucose. Whereas KG0.5 (g) for D-glucose at constant [Na] in the Ca-loaded membranes increased by approximately 50% of the control value (0.5 +/- 0.1 mM), no significant change in Jmax was observed. In contrast, both Jmax and KG0.5 (Na) for glucose, measured as a function of [Na] in the extravesicular fluid, were found to be significantly reduced. Na uptake, determined in the presence of 0.5 mM amiloride, was found to increase by approximately 30% of the value for control membrane. This increase was abolished when vesicles were preincubated with 0.5 mM neomycin or 0.5 mM phlorizin. The results suggest that the effect of Ca2+ on Na entry may be mediated in part by activation of phospholipase C and are consistent with a model of cotransport in which Ca2+ increases the mobility of the binary Na-sugar-translocator complex, thus leading to uncoupling of Na transport from glucose uptake ("slipping") and in part with Ca-induced Na entry by nonmediated leakage.
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PMID:Effect of Ca on Na-D-glucose cotransport across isolated renal brush-border membranes. 275 Sep 17

The stimulation of adenylate cyclase by glucagon and isoproterenol in cell lysates of hepatocytes isolated from fetal and adult female rats were measured after pretreatment with 12-O-tetradecanoyl-phorbol-13-acetate (TPA). Glucagon stimulation of adenylate cyclase activation was found to decrease in the hepatocyte lysate of adult female rats. Application of guanyl-5'-yl-imidodiphosphate (GppNHp) and forskolin showed this effect to be localized on the receptor level. However, glucagon stimulation of glucose liberation from phorbol ester-treated hepatocytes from adult female rats was not influenced. Maximum effects of glucose liberation were observed at glucagon concentrations which did not stimulate adenylate cyclase. The results are in agreement with the proposed existence of a low and high affinity glucagon receptor coupled to two different transducing systems. It is concluded that TPA uncouples in the liver of adult female rats--most likely by phosphorylation--the low affinity receptor from the adenylate cyclase system, whereas the high affinity receptor and phospholipase C/inositol 1,4,5-trisphosphate (IP3)/diacylglycerol (DAG) signalling systems do not seem to be affected. Such TPA effects could not be found in the liver of fetal rats.
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PMID:Age-dependent effects of phorbol ester on adenylate cyclase stimulation by glucagon in liver of female rats. 275 35

To ascertain whether mannose 6-phosphate affects insulin-like growth factor (IGF) II stimulation of phospholipase C activity in the basolateral membrane of the renal proximal tubular cell, we determined the effect of mannose 6-phosphate on IGF II-stimulated production of inositol trisphosphate (Ins-P3) in isolated basolateral membranes. Production of Ins-P3 measured in the presence of 10(-10), 10(-9), or 10(-8) M rat IGF II was potentiated approximately 2-fold by inclusion of 5 mM mannose 6-phosphate in incubations. Mannose 6-phosphate had no effect on Ins-P3 production in the absence of IGF II. Neither mannose 1-phosphate, mannose, glucose 6-phosphate, nor fructose 1-phosphate exerted similar potentiation. Enhancement of IGF II-stimulated Ins-P3 production required concentrations on the order of several millimolar mannose 6-phosphate. Total and specific binding of 10(-10) M 125I-IGF II to basolateral membranes was significantly increased by 5 mM mannose 6-phosphate. However, there was no significant effect on total or specific binding of 10(-9) or 10(-8) M 125I-IGF II. Our findings suggest that mannose 6-phosphate potentiates stimulation of phospholipase C by IGF II in the basolateral membrane of the renal proximal tubular cell and that potentiation is mediated via a mechanism in addition to enhanced binding of IGF II. Such potentiation could reflect a role for the mannose 6-phosphate moiety as a modulator of IGF II "signal" transmission in vivo.
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PMID:Mannose 6-phosphate potentiates insulin-like growth factor II-stimulated inositol trisphosphate production in proximal tubular basolateral membranes. 278 38

Incubation of human platelets with myo-[3H]inositol in a low-glucose Tyrode's solution containing MnCl2 enhanced the labelling of phosphoinositides about sevenfold and greatly facilitated the measurement of [3H]inositol phosphates formed by the activation of phospholipase C. Labelled platelets were permeabilized by high-voltage electric discharges and equilibrated at 0 degree C with ATP, Ca2+ buffers and guanine nucleotides, before incubation in the absence or presence of thrombin. Incubation of these platelets with ATP in the presence or absence of Ca2+ ions led to the conversion of [3H]phosphatidylinositol to [3H]phosphatidylinositol 4-phosphate and [3H]phosphatidylinositol 4,5-bisphosphate ([3H]PtdInsP2). At a pCa of 6, addition of 100 microM GTP[gamma S] both prevented this accumulation of [3H]PtdInsP2 and stimulated its breakdown; the formation of [3H]inositol phosphates was increased ninefold. After 5 min these comprised 70% [3H]inositol monophosphate ([3H]InsP), 28% [3H]inositol bisphosphate ([3H]InsP2) and 2% [3H]inositol trisphosphate ([3H]InsP3). In shorter incubations higher percentages of [3H]InsP2 and [3H]InsP3 were found. In the absence of added Ca2+, the formation of [3H]inositol phosphates was decreased by over 90%. Incubation of permeabilized platelets with GTP[gamma S] in the presence of 10 mM Li+ decreased the accumulation of [3H]InsP and increased that of [3H]InsP2, without affecting [3H]InsP3 levels. Addition of unlabelled InsP3 decreased the intracellular hydrolysis of exogenous [32P]InsP3 but did not trap additional [3H]InsP3. These results and the time course of [3H]inositol phosphate formation suggest that GTP[gamma S] stimulated the action of phospholipase C on a pool of [3H]phosphatidylinositol 4-phosphate that was otherwise converted to [3H]PtdInsP2 and that much less hydrolysis of [3H]phosphatidylinositol to [3H]InsP or of [3H]PtdInsP2 to [3H]InsP3 occurred. At a pCa of 6, addition of thrombin (2 units/ml) to permeabilized platelets caused small increases in the formation of [3H]InsP and [3H]InsP2. This action of thrombin was enhanced twofold by 10-100 microM GTP and much more potently by 4-40 microM GTP[gamma S]. In the presence of the latter, thrombin also increased [3H]InsP3. The total formation of [3H]inositol phosphates by permeabilized platelets incubated with thrombin and GTP[gamma S] was comparable with that observed on addition of thrombin alone to intact platelets. However, HPLC of the [3H]inositol phosphates formed indicated that about 75% of the [3H]InsP accumulating in permeabilized platelets was the 4-phosphate, whereas in intact platelets stimulated by thrombin, up to 80% was the 1-phosphate.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Effects of guanosine 5'-[gamma-thio]triphosphate and thrombin on the phosphoinositide metabolism of electropermeabilized human platelets. 283 Oct 52

The metabolism of D-glucose is believed to initiate and regulate insulin secretion by islet beta-cells, although the identity of the metabolite which couples glucose metabolism to the cellular events involved in insulin secretion is unknown. An alternative hypothesis involves the presence of a glucoreceptor for which there has been no biochemical evidence. We have investigated whether glucose recognition by the beta-cell is coupled to phospholipase C. We have used digitonin-permeabilized, [3H]inositol-prelabeled islets to study glucose and carbachol activation of phospholipase C. In this model, carbachol recognition by its muscarinic receptor was coupled to phospholipase C activation. D-Glucose (but not L-glucose) also stimulated phospholipase C activity in these permeabilized islets. This effect was not due to glucose metabolism since glucose 6-phosphate did not affect phospholipase C activity and since phosphorylation of [3H]glucose was not detectable in digitonin-permeabilized islets. Glucose had no effect on the myo-inositol-1,4,5-trisphosphate-5-phosphatase or 3-kinase activities. In the absence of agonist, free Ca2+ concentrations between 0.1 and 1 microM (as determined with a Ca2+-specific electrode) did not influence phospholipase C activity. Stimulation of phospholipase C activity by either carbachol or glucose required Ca2+ in the submicromolar range and was optimal at 0.5 microM free Ca2+.myo-Inositol-1,3,4,5-tetrakisphosphate production from permeabilized islets was synergistically augmented by Ca2+ (0.5-10 microM) and glucose. Phospholipase C activity in islets is therefore not directly activated by free Ca2+ concentrations in the submicromolar range. Furthermore, glucose per se activates phospholipase C activity independently of glucose metabolism. A working hypothesis based on these findings is that glucose is recognized by a site which is coupled to phospholipase C in islets.
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PMID:Studies of the Ca2+ requirements for glucose- and carbachol-induced augmentation of inositol trisphosphate and inositol tetrakisphosphate accumulation in digitonin-permeabilized islets. Evidence for a glucose recognition site in insulin secretion. 283 Nov 91

Diacylglycerols (DAG) modulate secretory responses by the activation of protein kinase C. Early changes in DAG formation induced by the muscarinic receptor agonist carbachol were compared to those caused by the nutrient secretagogue glucose in pancreatic islets. Turnover rates of DAG were investigated in radiolabeling experiments, whereas changes in total mass and fatty acid composition of DAG were assessed by gas-liquid chromatography. When islet lipids were labeled to steady state in tissue culture with [3H]glycerol, carbachol induced a rapid (10 s) and sustained increase of [3H]DAG generation. In contrast, glucose stimulation failed to increase [3H]glycerol containing DAG, and this was probably due to the isotopic dilution of the label secondary to enhanced glycolysis. This was substantiated by following the transfer of 14C from glucose into DAG. Within 1 min of acute exposure of islets to D-[U-14C]-glucose at stimulatory concentrations, DAG labeling increased fivefold representing up to 2% of total glucose usage. Similar stimulation of 14C incorporation into other neutral lipids and inositol phospholipids was observed, suggesting the enhanced de novo synthesis of phosphatidic acid, the common precursor for DAG, and inositol phospholipids from glycolytic intermediates. Transfer of 14C from glucose was not stimulated by agents such as carbachol and exogenous phospholipase C that act primarily on inositol phospholipid breakdown. The total mass of islet DAG was increased by 60% after both carbachol and glucose stimulation. However, analysis of the fatty acid composition of carbachol-generated DAG revealed at the early time point (10 s) a prevalent stearoyl-arachidonoyl configuration similar to that reported for inositol phospholipids. This pattern shifted to a DAG enriched in palmitic acid at a later time point. Glucose-stimulated islets displayed a predominance of palmitic acid containing DAG, indicating increased de novo synthesis of the putative second messenger rather than its formation by inositol phospholipid hydrolysis. Indeed, steady-state labeling of these phospholipids with [3H]inositol confirmed this idea since only carbachol caused detectable inositol phospholipid hydrolysis. Thus, although protein kinase C may be activated by both carbachol and glucose, the two secretagogues generate diacylglycerols through different mechanisms.
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PMID:Glucose and carbachol generate 1,2-diacylglycerols by different mechanisms in pancreatic islets. 283 45

In isolated islets, the hydrolysis of membrane phosphoinositides (PI) participates in the transduction of both extracellular and intracellular signals into an effective insulin secretory response. A wide variety of potential second-messenger molecules are generated during the phospholipase C-mediated cleavage of these strategically situated membrane phospholipids. Several distinct but interrelated issues are addressed in this perspective. These include 1) methodological approaches utilized to assess PI turnover, 2) the synergistic relationship between PI-derived second messengers and cAMP, 3) the contribution of changing PI turnover rates to the biphasic pattern of insulin output induced by 20 mM glucose, and 4) the role played by PI turnover in the phenomenon of "memory" displayed by islets after prior stimulation with various agonists. The concept that events unique to PI turnover contribute to beta-cell activation is well founded. Because of uncertainty regarding the exact nature of all PI-derived messengers, however, it is not yet possible to mold the available information into a comprehensive theory of beta-cell activation. Future studies will have to address various important unresolved issues.
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PMID:Modulation of insulin secretion from beta-cells by phosphoinositide-derived second-messenger molecules. 283 85

We have investigated the effects of the specific platelet-activating factor (PAF; 1-alkyl-2-acetyl-glycerophosphocholine) antagonist BN52021 on free fatty acid (FFA) and diacylglycerol (DG) accumulation and on the loss of fatty acids from phosphatidylinositol-4,5-bisphosphate (PIP2) in mouse brain. Mice were pretreated with BN52021 (10 mg/kg, i.p.) 30 min before electroconvulsive shock (ECS) or postdecapitation ischemia. These procedures cause rapid breakdown of PIP2 and accumulation of FFA and DG. Lipid extracts were prepared from microwave-fixed cerebrum and fractionated by TLC, and the fatty acid methyl esters were prepared by methanolysis and quantified by capillary GLC. In saline or vehicle (dimethyl sulfoxide)-treated mice, ECS caused marked accumulation of FFA and DG and loss of mainly stearic (18:0) and arachidonic (20:4) acids from PIP2. BN52021 pretreatment of ECS-treated mice decreased the accumulation of free palmitic (16:0), 18:0, 20:4, and docosahexaenoic (22:6) acids with no effect on the fatty acids in DG or the loss of PIP2. BN52021 had no effect on basal levels of FFA, DG, or PIP2. One minute of postdecapitation ischemia induced PIP2 loss and accumulation of FFA and DG. BN52021 attenuated the accumulation of free 20:4 and 22:6 acids, decreased the content of oleic (18:1), 20:4, and 22:6 acids in DG, but had no effect on PIP2 loss. These data indicate that BN52021 reduces the injury-induced activation of phospholipase A2 and lysophospholipase, which mediate the accumulation of FFA in brain, while having a negligible effect on phospholipase C-mediated degradation of PIP2.
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PMID:Platelet-activating factor antagonist BN52021 decreases accumulation of free polyunsaturated fatty acid in mouse brain during ischemia and electroconvulsive shock. 284 88


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