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)

Intracellular recordings were made from extraocular photoreceptor cells within isolated epistellar bodies of the lesser or northern octopus Eledone cirrhosa. The cells had resting potentials around -41+/-5 mV (mean +/- s.d., N=60) and showed light-flash-induced membrane depolarisation. The evoked response to a brief light flash consisted of a transient peak depolarisation, followed by a plateau component. The magnitude of the light-induced peak depolarisation response was decreased by bathing the epistellar body in artificial sea water (ASW) low in Na+, where choline+ replaced Na+, or by passing steady depolarising current. Replacement of external Na+ by Li+ had no effect on the light-stimulated response. The external application of the Na+ channel blocker tetrodotoxin (3 micromol l-1) increased the light-evoked response, but this was accompanied by a loss of action potential activity. The amplitude and duration of the response to a light flash was increased by bathing the epistellar body in ASW low in Ca2+, or in ASW containing 10 mmol l-1 Co2+, and after intracellular microinjection of the Ca2+ buffer EGTA. Intracellular microinjection of Ca2+ or inositol 1,4,5-trisphosphate, or external application of the phospholipase C inhibitor U-73122, had no apparent effect on the light-evoked response. These results are consistent with the interpretation that (1) the majority of the light-induced inward current is carried by Na+, probably via a non-selective cation channel, and (2) an increase in the intracellular free Ca2+ concentration, mediated by the phototransduction process, is involved in regulating the light-induced inward photocurrent and thus, in effect, determines the amplitude, time course and sensitivity of the receptor potential.
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PMID:Ionic mechanisms of phototransduction in photoreceptor cells from the epistellar body of the octopus eledone cirrhosa 1008 70

Exogenous electric fields induce cellular responses including redistribution of integral membrane proteins, reorganization of microfilament structures, and changes in intracellular calcium ion concentration ([Ca2+]i). Although increases in [Ca2+]i caused by application of direct current electric fields have been documented, quantitative measurements of the effects of alternating current (ac) electric fields on [Ca2+]i are lacking and the Ca2+ pathways that mediate such effects remain to be identified. Using epifluorescence microscopy, we have examined in a model cell type the [Ca2+]i response to ac electric fields. Application of a 1 or 10 Hz electric field to human hepatoma (Hep3B) cells induces a fourfold increase in [Ca2+]i (from 50 nM to 200 nM) within 30 min of continuous field exposure. Depletion of Ca2+ in the extracellular medium prevents the electric field-induced increase in [Ca2+]i, suggesting that Ca2+ influx across the plasma membrane is responsible for the [Ca2+]i increase. Incubation of cells with the phospholipase C inhibitor U73122 does not inhibit ac electric field-induced increases in [Ca2+]i, suggesting that receptor-regulated release of intracellular Ca2+ is not important for this effect. Treatment of cells with either the stretch-activated cation channel inhibitor GdCl3 or the nonspecific calcium channel blocker CoCl2 partially inhibits the [Ca2+]i increase induced by ac electric fields, and concomitant treatment with both GdCl3 and CoCl2 completely inhibits the field-induced [Ca2+]i increase. Since neither Gd3+ nor Co2+ is efficiently transported across the plasma membrane, these data suggest that the increase in [Ca2+]i induced by ac electric fields depends entirely on Ca2+ influx from the extracellular medium.
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PMID:Transmembrane calcium influx induced by ac electric fields. 1009 28

The phosphatidylcholine-preferring phospholipase C from Bacillus cereus (PC-PLC(Bc)) is a tri-Zn enzyme with two 'tight binding' and one 'loose binding' sites. The Zn2+ ions can be replaced with Co2+ and Cu2+ to afford metal-substituted derivatives. Two Cu2+-substituted derivatives are detected by means of 1H NMR spectroscopy, a 'transient' derivative and a 'stable' derivative. The detection of sharp hyperfine-shifted 1H NMR signals in the 'transient' derivative indicates the formation of a magnetically coupled di-Cu2+ center, which concludes that the Zn2+ ions in the dinuclear (Zn1 and Zn3) sites are more easily replaced by Cu2+ than that in the Zn2 site. This might possibly be the case for Co2+ binding. Complete replacement of the three Zn2+ ions can be achieved by extensive dialysis of the enzyme against excess Cu2+ to yield the final 'stable' derivative. This derivative has been determined to have five-coordinated His residues and an overall S'=1/2 spin state with NMR and EPR, consistent with the formation of a tri-Cu2+ center (i.e. a di-Cu2+/mono-Cu2+ center) in this enzyme. The binding of substrate to the inert tri-Cu2+ center to form an enzyme-substrate (ES) complex is clearly seen in the 1H NMR spectrum, which is not obtainable in the case of the native enzyme. The change in the spectral features indicates that the substrate binds directly to the trinuclear metal center. The studies reported here suggest that 1H NMR spectroscopy can be a valuable tool for the characterization of di- and multi-nuclear metalloproteins using the 'NMR friendly' magnetically coupled Cu2+ as a probe.
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PMID:Cobalt(II) and copper(II) binding of Bacillus cereus trinuclear phospholipase C: a novel 1H NMR spectrum of a 'Tri-Cu(II)' center in protein. 1173 Aug 96

Adenosine has a broad array of actions on neurons but astrocytes also possess adenosine receptors. We have previously shown that adenosine, by acting on astrocytes in the striatum, can modulate neuronal responses mediated by receptors coupled to phospholipase C through an astrocyto - neuronal interaction. In addition, adenosine was found to potentiate the alpha1-adrenergic production of inositol phosphates in astrocytes. The mechanism involved in this potentiation was further investigated by examining the effects of adenosine and alpha1-adrenergic receptor agonists on cytosolic Ca2+ in cultured striatal astrocytes from the embryonic mouse in primary culture. When used alone, methoxamine, a selective agonist of alpha-adrenergic receptors or 2-chloroadenosine, a stable analogue of adenosine, induced a transitory increase in cytosolic Ca2+, but their combined addition led to a sustained increase in cytosolic Ca2+, which seems to be due to a Ca2+ influx, because it was not observed in the absence of external Ca2+. Voltage independent Ca2+ channels contribute to this process and different blockers of voltage-operated calcium channels, such as dihydropyridines, phenylalkylamines, La3+ or Co2+ were ineffective in suppressing the sustained cytosolic Ca2+ elevation. Three observations suggest the implication of arachidonic acid in the observed potentiation: (i) arachidonic acid induced a sustained elevation of cytosolic Ca2+ similar to that evoked by the coapplication of methoxamine and 2-chloroadenosine; (ii) the addition of arachidonic acid during the calcic plateau produced by the combined application of the agonists did not increase further cytosolic Ca2+ levels; (iii) in the presence of methoxamine, 2-chloroadenosine induced a release of arachidonic acid. The stimulation of phospholipase C and the resulting activation of protein kinase C induced by methoxamine seem to be required for the potentiating effect of 2-chloroadenosine on cytosolic Ca2+. In fact, the direct activation of protein kinase C by an exogenous diacylglycerol analogue mimicked the effect of methoxamine because, in this condition, 2-chloroadenosine alone evoked a sustained elevation of cytosolic Ca2+. Therefore, methoxamine, through the successive activation of phospholipase C and protein kinase C, could allow a lipase, probably phospholipase A2, to be stimulated by 2-chloroadenosine. Arachidonic acid has already been shown to trigger the opening of K+ channels and the formation of inositol phosphates in other cell types. Therefore, in striatal astrocytes, 2-chloroadenosine, through an arachidonic acid-mediated hyperpolarization, could increase the Ca2+ driving force and thus improve Ca2+ influx through inositol phosphate-gated channels. This hypothesis is further supported by the suppressing effect of a 50 mM KCI-induced depolarization on the long lasting elevation of cytosolic Ca2+ seen in the combined presence of 2-chloroadenosine and methoxamine.
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PMID:Synergistic Regulation of Cytosolic Ca2+ Concentration by Adenosine and alpha1-Adrenergic Agonists in Mouse Striatal Astrocytes. 1210 86

Previous findings show that both the vanilloid receptor 1 and the insulin receptor are expressed on small primary sensory neurons. As insulin evokes activity in second messengers which could induce opening of the vanilloid receptor 1, we examined, by using the cobalt-uptake technique, whether or not insulin can activate cultured rat primary sensory neurons through activating the vanilloid receptor 1. Capsaicin (50, 100 and 500 nm) induced concentration-dependent labelling in primary sensory neurons. Preincubation of cells in insulin (10 micromoles) for 10 min followed by a 2-min wash did not produce significant change in the capsaicin-induced labelling. Coapplication of insulin (10 micromoles) with capsaicin, however, potentiated the 50 and 100 nm capsaicin-evoked staining. Insulin itself also produced cobalt labelling in a concentration-dependent manner. The size-frequency distributions of neurons showing capsaicin- or insulin-induced cobalt accumulation were similar. The insulin-induced cobalt labelling was significantly reduced by the tyrosine kinase inhibitor, tyrphostin AG1024, the vanilloid receptor 1 antagonists, ruthenium red and capsazepine, the protein kinase inhibitor, staurosporine and the phospholipase C inhibitor neomycin. Double immunostaining of cultured primary sensory neurons and sections from dorsal root ganglia revealed that about one-third of the cells coexpress the insulin receptor and vanilloid receptor 1. These findings suggest that insulin activates a subpopulation of primary sensory neurons, probably through phosphorylation- and/or phosphatidylinositol(4,5)biphosphate hydrolysis-evoked activation of the vanilloid receptor 1. Although the insulin-induced activation of vanilloid receptor 1 seems to be a short-lived effect in vitro, in vivo it might play a role in the development of burning pain sensation in hyperinsulinism.
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PMID:Insulin induces cobalt uptake in a subpopulation of rat cultured primary sensory neurons. 1462 48

Green tea has been receiving considerable attention as a possible preventive agent against cancer and cardiovascular disease. Epigallocatechin-3-gallate (EGCG) is a major polyphenol component of green tea. Using digital calcium imaging and an assay for [3H]-inositol phosphates, we determined whether EGCG increases intracellular [Ca2+] ([Ca2+]i) in non-excitable human astrocytoma U87 cells. EGCG induced concentration-dependent increases in [Ca2+]i. The EGCG-induced [Ca2+]i increases were reduced to 20.9% of control by removal of extracellular Ca2+. The increases were also inhibited markedly by treatment with the non-specific Ca2+ channel inhibitors cobalt (3 mM) for 3 min and lanthanum (1 mM) for 5 min. The increases were not significantly inhibited by treatment for 10 min with the L-type Ca2+ channel blocker nifedipine (100 nM). Treatment with the inhibitor of endoplasmic reticulum Ca2+-ATPase thapsigargin (1 micro M) also significantly inhibited the EGCG-induced [Ca2+]i increases. Treatment for 15 min with the phospholipase C (PLC) inhibitor neomycin (300 micro M) attenuated the increases significantly, while the tyrosine kinase inhibitor genistein (30 micro M) had no effect. EGCG increased [3H]-inositol phosphates formation via PLC activation. Treatment for 10 min with mefenamic acid (100 micro M) and flufenamic acid (100 micro M), derivatives of diphenylamine-2-carboxylate, blocked the EGCG-induced [Ca2+]i increase in non-treated and thapsigargin-treated cells but indomethacin (100 micro M) did not affect the increases. Collectively, these data suggest that EGCG increases [Ca2+]i in non-excitable U87 cells mainly by eliciting influx of extracellular Ca2+ and partly by mobilizing intracellular Ca2+ stores by PLC activation. The EGCG-induced [Ca2+]i influx is mediated mainly through channels sensitive to diphenylamine-2-carboxylate derivatives.
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PMID:Epigallocatechin-3-gallate increases intracellular [Ca2+] in U87 cells mainly by influx of extracellular Ca2+ and partly by release of intracellular stores. 1464 74

The aim of the present study was to determine whether cobalt poisoning induces haem oxidase isoenzyme-1 (HO-1) in coronary artery smooth muscle, or accounts for any changes in coronary smooth muscle cell (SMCs) membrane ionic currents that could result from this type of heavy metal poisoning. In SMCs isolated from cobalt-treated guinea-pig coronaries, K+ channel currents (IK) were much smaller than those in cells isolated from non-treated animals. Haemin (HO substrate) increased IK concentration dependently. This effect was mimicked by 1% CO and was abolished by pretreatment of cells with a competitive HO inhibitor, by inhibitors of guanylyl cyclase, protein kinase G or phospholipase C, as well as by blocking inositol trisphosphate-dependent Ca release, or sarcoplasmic reticulum Ca-ATPase, or by bathing cells in Ca-free external solution. Expression of the Na/Ca exchanger-1 (NCX-1) protein was reduced substantially in SMCs from coronary arteries of cobalt-treated animals. No expression of HO-1 was detected. It is concluded that acute cobalt poisoning in vivo depresses Ca-sensitive K currents via CO-dependent modulation of intracellular calcium availability, most probably by suppressing the expression of NCX-1 protein.
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PMID:Calcium-dependent changes in potassium currents in guinea-pig coronary artery smooth muscle cells after acute cobalt loading in vivo. 1534 Aug 49

A new tissue slice preparation of the cuttlefish eye is described that permits patch-clamp recordings to be acquired from intact photoreceptors during stimulation of the retina with controlled light flashes. Whole-cell recordings using this preparation, from the retinas of very young Sepia officinalis demonstrated that the magnitude, latency, and kinetics of the flash-induced photocurrent are closely dependent on the magnitude of the flash intensity. Depolarizing steps to voltages more positive than -40 mV, from a membrane holding potential of -60 mV, induced a transient inward current followed by a larger, more sustained outward current in these early-stage photoreceptors. The latter current resembled the delayed rectifier (I(K)) already identified in many other nerve cells, including photoreceptors. This current was activated at -30 mV from a holding potential of -60 mV, had a sustained time course, and was blocked in a dose-dependent manner by tetraethylammonium chloride (TEA). The smaller, transient, inward current appeared at potentials more positive than -50 mV, reached peak amplitude at -30 mV and decreased with further depolarization. This current was characterized as the sodium current (I(Na)) on the basis that it was inactivated at holding potentials above -40 mV, was blocked by tetrodotoxin (TTX) and was insensitive to cobalt. Intracellular perfusion of the photoreceptors, via the patch pipette, demonstrated that U-73122 and heparin blocked the evoked photocurrent in a dose-dependent manner, suggesting the involvement of the phospholipase C (PLC) and inositol 1,4,5-triphosphate (InsP(3)), respectively, in the phototransduction cascade. Perfusion with cyclic GMP increased significantly the evoked photocurrent, while the inclusion of phorbol-12,13-dibutyrate reduced significantly the evoked photocurrent, supporting the involvement of cGMP and the diacylglycerol (DAG) pathways, respectively, in the cuttlefish transduction process.
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PMID:Whole-cell recording of light-evoked photoreceptor responses in a slice preparation of the cuttlefish retina. 1607 10

The endogenous ligand, anandamide activates at least two receptors on nociceptors; the excitatory vanilloid type 1 transient receptor potential receptor, the activity of which is indispensable for the development and maintenance of inflammatory heat hyperalgesia, and the inhibitory cannabinoid 1 receptor, the activity of which reduces that pathological pain sensation. Recent data are equivocal on whether increasing anandamide levels at the peripheral terminals of nociceptors in pathological conditions increases or decreases inflammatory heat hyperalgesia. Here, by using the cobalt-uptake technique we examined whether vanilloid type 1 transient receptor potential receptor activity evoked by 10 nM-100 microM anandamide is increased or decreased in inflammatory conditions. An inflammatory milieu for cultured rat primary sensory neurons was established by incubating the cells in the presence of the inflammatory mediators, bradykinin and prostaglandin E2. Anandamide, similarly to the archetypical vanilloid type 1 transient receptor potential receptor agonist, capsaicin induced concentration-dependent cobalt-uptake in a proportion of neurons. However, the potency of anandamide was significantly lower than that of capsaicin. While pre-incubation of cultures with bradykinin and prostaglandin E2 alone did not evoke cobalt-entry, the inflammatory mediators potentiated the effect of both capsaicin and anandamide. Application of the competitive vanilloid type 1 transient receptor potential receptor antagonist, capsazepine, or inhibitors of protein kinase A, protein kinase C or phospholipase C inhibited the anandamide-evoked cobalt-uptake both in the presence and absence of bradykinin and prostaglandin E2. These findings show that inflammatory mediators significantly increase the excitatory potency and efficacy of anandamide on vanilloid type 1 transient receptor potential receptor, thus, increasing the anandamide concentration in, or around the peripheral terminals of nociceptors might rather evoke than decrease inflammatory heat hyperalgesia.
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PMID:Inflammatory mediators convert anandamide into a potent activator of the vanilloid type 1 transient receptor potential receptor in nociceptive primary sensory neurons. 1619 86

The capsaicin receptor, known as transient receptor potential channel vanilloid subtype 1 (TRPV1), is activated by a wide range of noxious stimulants and putative ligands such as capsaicin, heat, pH, anandamide, and phosphorylation by protein kinase C (PKC). However, the identity of endogenous activators for TRPV1 under physiological condition is still debated. Here, we report that diacylglycerol (DAG) directly activates TRPV1 channel in a membrane-delimited manner in rat dorsal root ganglion (DRG) neurons. 1-oleoyl-2-acetyl-sn-glycerol (OAG), a membrane-permeable DAG analog, elicited intracellular Ca2+ transients, cationic currents and cobalt uptake that were blocked by TRPV1-selective antagonists, but not by inhibitors of PKC and DAG lipase in rat DRG neurons or HEK 293 cells heterologously expressing TRPV1. OAG induced responses were about one fifth of capsaicin induced signals, suggesting that OAG displays partial agonism. We also found that endogenously produced DAG can activate rat TRPV1 channels. Mutagenesis of rat TRPV1 revealed that DAG-binding site is at Y511, the same site for capsaicin binding, and PtdIns(4,5)P2binding site may not be critical for the activation of rat TRPV1 by DAG in heterologous system. We propose that DAG serves as an endogenous ligand for rat TRPV1, acting as an integrator of Gq/11-coupled receptors and receptor tyrosine kinases that are linked to phospholipase C.
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PMID:Direct activation of transient receptor potential vanilloid 1(TRPV1) by diacylglycerol (DAG). 1882 53


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