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)

Heparin is thought to inhibit InsP3 binding to receptors involved in the intracellular release of Ca2+. Injection of heparin into Limulus ventral photoreceptors to high intracellular concentrations reduces the amplitude and slows the rate of rise of voltage-clamp currents induced by brief flashes, tends to make the responses to long flashes more "square," and tends to block the light-induced rise in [Ca2+]i detected by arsenazo III. In these ways, intracellular heparin mimics the effects of high concentrations of intracellular BAPTA or EGTA. In addition, the effects of heparin are attenuated by prior injection of BAPTA to high intracellular concentrations. Neomycin and spermine are thought to inhibit phospholipase C activity. Injections of spermine or neomycin to low intracellular concentrations largely mimic the effects of intracellular heparin. These findings suggest that the predominant effect of polyamines is to inhibit light-induced production of InsP3 by phospholipase C activity and thereby reduce the light-induced increase in [Ca2+]i. Our findings suggest that excitation can proceed in the absence of InsP3-induced increases in [Ca2+]i, but (a) the gain and speed of transduction are reduced and (b) adaptation is largely blocked.
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PMID:Intracellular injection of heparin and polyamines. Effects on phototransduction in limulus ventral photoreceptors. 833 23

1. Whole cell recordings from dentate granule neurons in the hippocampal slice preparation reveal that (1 S, 3R)-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD), a selective agonist at metabotropic glutamate receptors (mGluRs), inhibits a calcium-activated potassium current (IAHP) responsible for the postspike after-hyperpolarization. Inclusion of 1 mM of the Ca2+ chelator ethylene glycol-bis (beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid in the patch pipette reduced the inhibitory action of ACPD on IAHP while having no effect on a similar action of serotonin (5-HT). Thus the known action of ACPD of mobilizing intracellular Ca2+ may be involved in this inhibitor action of ACPD. 2. Inhibition of IAHP is not secondary to effects on Ca2+ currents, because 10 microM ACPD, which inhibits IAHP by 95 +/- 5% (mean +/- SE), reduced the Ca2+ current by only 8 +/- 4%. 3. Activation of mGluRs accelerates the irreversible inhibition of IAHP that develops when 88 microM GTP-gamma-S is included in the pipette filling solution, whereas inclusion of 1 mM GDP-beta-S attenuated the inhibitory action of ACPD. These results indicate that the response to mGluR activation is G protein mediated. 4. Group I mGluRs, which includes mGluR1 and mGluR5, are G-protein-coupled receptors that are known to stimulate phospholipase C (PLC)-mediated hydrolysis of phosphoinositides to produce 1,4,5-triphosphate (IP3), which in turn is known to mobilize the release of intracellular Ca2+. The weak but selective mGluR1 agonist (S)-3-hydroxyphenylglycine (100 microM) completely inhibited IAHP, and the mGluR1 antagonist (S)-4-carboxyphenylglycine (500 microM) reduced IAHP inhibition produced by 5 microM ACPD from 73 +/- 6% to 22 +/- 4%. These results indicate that the mGluR responsible for IAHP inhibition has a similar pharmacological profile to that of those coupled to IP3 production. 5. The effects of agents known to interfere with IP3 production and action also support IP3 involvement in ACPD action. Neomycin (1 mM in pipette solution), which should reduce IP3 production through inhibition of PLC, reduced the ability of 10 microM ACPD to inhibit IAHP from almost 100% to 57 +/- 8% (n = 8). Heparin, an IP3 receptor antagonist that reduces Ca2+ mobilization, attenuated the inhibitory action 10 microM ACPD from almost 100% to 39 +/- 5% (n = 5). Heparin by itself increased the amplitude and duration of IAHP, suggesting that resting levels of IP3 are sufficient to suppress of IAHP partially. 6. In addition to the pool of intracellular Ca2+ that is mobilized by IP3, there is a distinct pool that is responsible for Ca(2+)-triggered Ca2+ release and is blocked by ryanodine or dantrolene. These drugs caused a small reduction of both IAHP and the inhibitory action of ACPD. Possibly the Ca2+ signal mobilized by IP3 is partially amplified by Ca2+ released from the ryanodine-sensitive stores. 7. Activation of PLC can also lead to the production of diacylglycerol and activation of protein kinase C (PKC). However, the inhibitory action of ACPD on IAHP was not affected by staurosporine at a concentration (1 microM) that inhibits both protein kinase A (PKA) and PKC and blocks the action of 5-HT to inhibit IAHP. 8. Activation of PKA by the adenylate cyclase activator forskolin led to inhibition of IAHP. Although activation of mGluR1 agonists can also stimulate adenylate cyclase and activate PKA, inhibition of PKA and the effect of forskolin on IAHP with the Walsh peptide did not affect ACPD inhibition of IAHP. 9. All of our results support the hypothesis that mGluR-mediated inhibition of IAHP is initiated by the production of IP3 and the mobilization of intracellular Ca2+.
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PMID:Metabotropic glutamate receptors coupled to IP3 production mediate inhibition of IAHP in rat dentate granule neurons. 889 38

Ammodytin L, purified from the venom of Vipera ammodytes, triggers a rapid and dramatic lytic process in myotubes in vitro, as well as in differentiated muscle cells in vivo, through a mechanism that is not well understood. Despite its great sequence similarity to phospholipase A2, it is devoid of any enzyme activity. Data on artificial membranes demonstrating a direct interaction between this toxin and the hydrophobic core of the lipid bilayer suggest that the toxin also acts on the lipid microenvironment in cell membranes. Recent experiments on living cells do not confirm this hypothesis, and a more intricate mechanism is proposed. In vitro, ammodytin L has necrotic effects only in well-differentiated myogenic cells, whereas other cell types such as platelets, red blood cells and lymphocytes show neither morphological nor functional alterations. In this work we demonstrate that rat 208F fibroblasts in culture after ammodytin L challenge increase [3H]thymidine incorporation, indicating that this toxin has a myogenic effect. Moreover, ammodytin L increases intracellular Ca2+ by acting on intracellular stores probably by activating a phosphatidylinositol-specific phospholipase C. Preincubation of the cells with ammodytin L did not prevent the massive Ca2+ release evoked by bradykinin, a phenomenon observed when fibroblasts were incubated with both thapsigargin and ionomycin. Heparin, an agent that inhibits the necrotic effect of the myotoxin in myotubes, also reduces the effect of ammodytin L on DNA synthesis. Heparin inhibits only the late sustained increase in intracellular Ca2+ induced by the toxin.
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PMID:Proliferative effect of ammodytin L from the venom of Vipera ammodytes on 208F rat fibroblasts in culture. 897 54

1. Spontaneous inhibitory postsynaptic currents (IPSCs) and evoked IPSCs were recorded by a whole-cell patch-recording technique from cultured Purkinje cells of the rat. The size of spontaneous IPSCs, after a train of depolarizing pulses was applied to the Purkinje cells, increased to 163 +/- 6% (mean +/- S.E.M., n = 7 cells) of the control levels measured before the stimulus train. 2. The GABAergic postsynaptic currents were recorded under voltage clamp from the synapse formed between two Purkinje cells. These IPSCs increased to 218 +/- 31% (n = 4) of control levels after depolarizing stimulation was applied to the postsynaptic Purkinje cells. Size-increased IPSCs were observed as long as recording continued and the phenomena will be called potentiation in this paper. 3. Intracellular application of Ruthenium Red (20 microM) did not block the potentiation of spontaneous IPSCs induced by the depolarizing stimulus (165 +/- 9%, n =6), but heparin (2 mg ml-1) partially blocked the potentiation (123 +/- 10%, n = 6). Heparin applied together with Ruthenium Red (20 microM) blocked potentiation completely (96 +/- 5%, n = 8) at concentrations higher than 1 mg ml-1. 4. Intracellular free calcium concentrations ([Ca2+]i) was monitored as the ratio of fura-2 fluorescences excited at 340 and 380 nm. In control cells, [Ca2+]i was increased by each depolarizing pulse. When Purkinje cells were dialysed with heparin or heparin with Ruthenium Red, the rise in [Ca2+]i was suppressed. 5. Bath application of thapsigargin (1 microM) blocked the potentiation (99 +/- 2%, n = 4) and suppressed the rise in [Ca2+]i. 6. When 30 mM BAPTA was applied intracellularly, a train of depolarizing pulses failed to induce potentiation of IPSCs and failed to raise [Ca2+]i. The results from points 3-6 suggest that the increase in [Ca2+]i, most probably coupled with the release from intracellular stores especially from the inositol trisphosphate (IP3)-sensitive stores, is crucial for the potentiation of IPSCs. 7. Bath application of a metabotropic glutamate receptor activator (t-ACPD, 200 microM) increased both the amplitude and frequency of spontaneous IPSCs and increased the [Ca2+]i slightly in dendrites. The inward current induced by the puff-applied GABA (2 microM) was increased, after t-ACPD application, to 186 +/- 36% of the control level (n = 3). Bath application of quisqualate (2 microM) caused a rapid increase in [Ca2+]i in dendrites and in the cell body and increased both the amplitude and frequency of spontaneous IPSCs. 8. The bath application of an inhibitor of phospholipase C (PLC), U73122 (1 microM), suppressed a rise in [Ca2+]i and blocked the potentiation (106 +/- 3%, n = 5). The inactive form, U73343 (1 microM), did not affect the potentiation (151 +/- 11%, n = 7) or the rise in [Ca2+]i. These observations suggest a possible involvement of the mechanism of Ca2+ activation of PLC and the IP3-induced Ca2+ release in the induction of IPSC potentiation in Purkinje cells.
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PMID:Release of Ca2+ is the crucial step for the potentiation of IPSCs in the cultured cerebellar Purkinje cells of the rat. 900 48

We have shown previously that the interaction between cytotoxic T lymphocytes (CTL) and ventricular myocytes, an in vitro model for heart transplant rejection, results in electrophysiological and morphological alterations indicative of overload of the intracellular [Ca2+] ([Ca2+]i). Since these deleterious effects cannot be accounted for by increased L-type Ca2+ current (ICa,L), we hypothesize that [Ca2+]i overload due to Ca2+ release from intracellular stores, e.g. sarcoplasmic reticulum (SR), is initiated by CTL-induced activation of the inositol trisphosphate (IP3) cascade. Patch-clamp and fura-2-fluorescence techniques were utilized to record transmembrane potentials and [Ca2+]i from ventricular myocytes bound to peritoneal exudate CTL (PEL). In ventricular myocyte-PEL conjugates (after 60 min), resting potential was reduced (compared with the nonconjugated state) from -80.9 +/- 0.7 to -59.9 +/- 2.5 mV, action potential amplitude from 139.5 +/- 1.4 to 80.6 +/- 1.7 mV and action potential duration to 50% repolarization (APD50) from 797 +/- 97 to 52 +/- 12 ms. The ratio of fluorescence at 340 and 380 nm (R340/380) increased from a control value (in nonconjugated myocytes) of 0.71 +/- 0.02 to 2.07 +/- 0.03, 30 min, after conjugate formation, and exceeded 4.0 at 60 min, before myocyte destruction. Heparin (50 micrograms/ml), an antagonist of IP3-induced Ca2+ release from SR channels, or U-73122 (2 microM), a phospholipase C (PLC) inhibitor (drugs were included in the pipette solution), prevented PEL-induced morphological and electrophysiological alterations. Accordingly, heparin attenuated the PEL-induced increase in [Ca2+]i; after 60 min of PEL-myocyte interaction, R340/380 was 1.15 +/- 0.09 (compared with approximately 4.0 in the absence of heparin). The results indicate that CTL-mediated damage to ventricular myocytes is, at least partially, mediated by PLC activation and IP3-induced Ca2+ release from intracellular stores. Pharmacological targeting of IP3 in heart transplant rejection is thus suggested.
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PMID:Involvement of the IP3 cascade in the damage to guinea-pig ventricular myocytes induced by cytotoxic T lymphocytes. 904 62

We have previously demonstrated that thrombin possesses an active yet cryptic Arg-Gly-Asp (RGD) site which upon exposure induces endothelial cell (EC) adhesion via alpha nu beta 3 integrin [Bar-Shavit et al. (1991): J Cell Biol 112:335]. This was achieved in the presence of cell surface-associated heparan sulfate proteoglycans (HSPG) and exceedingly low concentrations of plasmin [Bar-Shavit et al. (1993): J Cell Biol 123:1279]. A portion of the cell surface-associated HSPG (glypican) is anchored via a covalently linked glycosyl-phosphatidylinositol (PI) residue, which can be released by treatment with glycosyl-PI-specific phospholipase C (PI-PLC). We report here that exposure of either bovine aortic EC, smooth muscle cells (SMC), or wild-type CHO cells to PI-PLC released HSPG involved in the conversion of thrombin to an adhesive molecule. The adhesion-promoting activity of the released HSPG was abolished following treatment with heparinase but not chondroitinase ABC. Incubation of thrombin with heparan sulfate-deficient CHO cells or cells that were pretreated with PI-PLC failed to induce its conversion to an adhesive molecule, indicating that glypican was playing a major role in this conversion. Moreover, affinity-purified glypican, but not syndecan or fibroglycan, elicited efficient conversion of plasmin-treated thrombin into an adhesive molecule. Antibodies raised against the RGD site in thrombin failed to interact with native thrombin, prothrombin, or the RGD site in other adhesive proteins such as vitronectin, fibrinogen, or fibronectin. Anti-thrombin-RGD antibodies which blocked the adhesion-promoting activity of thrombin were also capable of recognizing thrombin that was first incubated with a suboptimal concentration of plasm in in the presence of PI-PLC-released HSPG. Heparin, heparan sulfate, and PI-PLC-released HSPG had no effect on other cellular properties of thrombin such as receptor binding and growth-promoting activity. Altogether we have demonstrated that the heparin binding domain in thrombin plays a specific role in promoting thrombin adhesive properties and that membrane-associated glypican is likely to be the major physiological inducer of this property.
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PMID:Specific involvement of glypican in thrombin adhesive properties. 917 91

Cytotoxic T lymphocytes (CTLs) that infiltrate the heart are important immune effectors implicated in heart transplant rejection, myocarditis, and other cardiomyopathies. To investigate the mechanism(s) underlying CTL damage to the myocardium through activation of the Fas receptor (Fas/CD95/Apo-1) by the Fas ligand, we explored the interaction between peritoneal exudate CTLs (PELs), derived from perforin gene-knockout (P-/-) mice, and murine ventricular myocytes. Fas expression on isolated ventricular myocytes was demonstrated immunohistochemically. Action potentials, [Ca2+]i transients, and contractions of myocytes conjugated to P-/- PELs or treated with the apoptosis-inducing anti-Fas monoclonal antibody Jo2 were recorded. Action potential characteristics of nonconjugated myocytes and myocytes conjugated with P-/- PELs were, respectively, as follows: Vm, -73.2+/-1.5 and -53.6+/-6.4 mV (mean+/-SEM); action potential amplitude, 117.9+/-3.9 and 74.3+/-21.2 mV; and action potential duration at 80% repolarization, 17+/-6 and 42+/-13 milliseconds (all P<.05). P-/- PELs also induced early and delayed afterdepolarizations as well as arrhythmogenic activity. Diastolic [Ca2+]i increased during the cytocidal interaction with P-/- PELs, from a fluorescence ratio of 0.82+/-0.05 (n=7) to 1.98+/-0.09 (n=13) (P<.05). All of the effects caused by P-/- PELs were reproduced by incubating the myocytes with Jo2. Heparin (50 microg/mL), an antagonist of inositol trisphosphate (IP3)-operated sarcoplasmic reticulum Ca2+ channels, or U-73122 (2 micromol/L), a phospholipase C inhibitor, but not the inactive agonist U-73343, prevented Fas-mediated myocyte dysfunction. Additionally, intracellular application (through the patch pipette) of the active IP3 analogue, inositol 1,4,5-trisphosphate, but not the inactive analogue, inositol 1,3,4-trisphosphate, caused electrophysiological changes resembling those resulting from P-/- PELs and Jo2, suggesting that CTL-induced Fas-based myocyte dysfunction is mediated by IP3. We conclude that a Fas-based perforin-independent mechanism of CTL action can account for the immunopathology seen in the allotransplanted heart, myocarditis, and dilated cardiomyopathy.
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PMID:Fas (CD95/Apo-1)-mediated damage to ventricular myocytes induced by cytotoxic T lymphocytes from perforin-deficient mice: a major role for inositol 1,4,5-trisphosphate. 950 4

In rat cerebellar slices, repetitive parallel fiber stimulation evokes an inward, postsynaptic current in Purkinje cells with a fast component mediated by alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainate receptors and a slower component mediated by metabotropic glutamate receptors (mGluR). The mGluR-mediated excitatory postsynaptic current (mGluR-EPSC) is evoked selectively by parallel fiber stimulation; climbing fiber stimulation is ineffective. The mGluR-EPSC is elicited most effectively with increasing frequencies of parallel fiber stimulation, from a threshold of 10 Hz to a maximum response at approximately 100 Hz. The amplitude of the mGluR-EPSC is a linear function of the number of stimulus pulses without any apparent saturation, even with >10 pulses. Thus mGluRs at the parallel fiber-Purkinje cell synapse can function as linear detectors of the number of spikes in a burst of activity in parallel fibers. The mGluR-EPSC is present from postnatal day 15 and persists into adulthood. It is inhibited by the generic mGluR antagonist (RS)-a-methyl-4-carboxyphenylglycine and by the group I mGluR antagonist (RS)-1-aminoindan-1,5-dicarboxylic acid at a concentration selective for mGluR1. Although the intracellular transduction pathway involves a G protein, the putative mediators of mGluR1 (phospholipase C and protein kinase C) are not directly involved, indicating that the mGluR-EPSC studied here is mediated by a different and still unidentified second-messenger pathway. Heparin, a nonselective antagonist of inositol-trisphosphate (IP3) receptors, has no significant effect on the mGluR-EPSC, suggesting that also IP3 might be not required for the response. Buffering intracellular Ca2+ with a high concentration of bis-(o-aminophenoxy)-N,N,N', N'-tetraacetic acid partially inhibits the mGluR-EPSC, indicating that Ca2+ is not directly responsible for the response but that resting Ca2+ levels exert a tonic potentiating effect on the mGluR-EPSC.
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PMID:Postsynaptic current mediated by metabotropic glutamate receptors in cerebellar Purkinje cells. 970 47

1. To examine the contributions of the putative Ca2+ releasers, inositol 1,4,5-trisphosphate (InsP3), cyclic ADP ribose (cADPR), and nicotinate adenine dinucleotide phosphate (NAADP), to carbachol (CCh)-induced contraction in airway smooth muscle, we measured force development of permeabilized rabbit tracheal smooth muscle, human bronchial smooth muscle and guinea-pig ileum longitudinal smooth muscle. 2. In the presence of 50 microM GTP, CCh and InsP3 contracted alpha-toxin-permeabilized tracheal smooth muscle dose dependently; the EC50 values for CCh and InsP3 were 1.84 microM and 363 microM, and the maximum responses (normalized to the 30 mM caffeine response) to 100 microM CCh and to 800 microM InsP3 were 206 +/- 13.4 % (mean +/- S.E.M.) and 84.4 +/- 5.3 %, respectively. 3. However, cADPR (10-300 microM), beta-NAD+ (2.5 mM), FK506 (30 microM) and NAADP (100 microM) neither contracted the strip by themselves nor affected the subsequent CCh (1 microM) response. alpha-Toxin-permeabilized bronchial smooth muscle and ileum smooth muscle also responded to caffeine, InsP3 and CCh but not to cADPR. 4. Both 100 microM 8-amino-cADPR, a selective cADPR antagonist, and 100 microM thionicotinamide-NADP, a selective NAADP antagonist, failed to inhibit the CCh response, although procaine abolished the caffeine, InsP3 and CCh responses in the permeabilized tracheal smooth muscle. 5. Although inhibition of the caffeine response by 30 microM ryanodine was nearly complete, approximately 30 % of the InsP3 (300 microM) plus GTP (50 microM) response was retained, and the resultant response disappeared after the caffeine response was evoked in the presence of ryanodine. 6. Heparin (300 microg ml-1) blocked InsP3 (300 microM) and CCh (3 microM) responses in beta-escin-permeabilized tracheal smooth muscle, while Ruthenium Red (100 microM) partially inhibited the CCh response. 7. Collectively, InsP3 but not cADPR or NAADP plays a key role in CCh-initiated contraction, and InsP3 utilizes a single compartment of the caffeine/ryanodine-sensitive stored Ca2+ in airway smooth muscle.
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PMID:InsP3, but not novel Ca2+ releasers, contributes to agonist-initiated contraction in rabbit airway smooth muscle. 971 70

The effects of heparin on ion channels formed by Staphylococcus aureus alpha-toxin (ST channel) in lipid bilayers were studied under voltage clamp conditions. Heparin concentrations as small as 100 pM induced a sharp dose-dependent increase in channel voltage sensitivity. This was only observed when heparin was added to the negative-potential side of lipid bilayers in the presence of divalent cations. Divalent cations differ in their efficiency: Zn2+>Ca2+>Mg2+. The apparent positive gating charge increased 2-3-fold with heparin addition as well as with acidification of the bathing solution. 'Free' carboxyl groups and carboxyl groups in ion pairs of the protein moiety are hypothesized to interact with sulfated groups of heparin through divalent cation bridges. The cis mouth of the channel (that protrudes beyond the membrane plane on the side of ST addition and to which voltage was applied) is less sensitive to heparin than the trans-mouth. It is suggested that charged residues which interact with heparin at the cis mouth of ST channels and which contribute to the effective gating charge at negative voltage may be physically different from those at the trans mouth and at positive voltage.
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PMID:Heparin influence on alpha-staphylotoxin formed channel. 1007 45


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