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)

Uridine nucleotides, released into the extracellular environment, influence a variety of metabolic and other cellular activities in a wide range of target tissues. Here we have studied the effects of uridine nucleotides on gluconeogenesis in isolated rat proximal tubules. Gluconeogenesis, from a range of precursors, was stimulated following exposure of isolated proximal tubules to either UTP or UTPgammaS, but not when exposed to other uridine-containing nucleotides. UTP- and UTPgammaS-induced gluconeogenesis was diminished in the presence of purinoceptor antagonists (e.g. suramin, PPADS) indicative of a role for P2Y(2)-like purinoceptors in these effects. Likewise, agents that interfere with either phospholipase C activation or intracellular Ca2+ mobilization decreased UTP- and UTPgammaS-induced stimulation of gluconeogenesis consistent with a role for these secondary messenger systems in the mechanism of action of extracellular UTP and UTPgammaS on proximal tubule metabolism.
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PMID:Uridine nucleotide-induced stimulation of gluconeogenesis in isolated rat proximal tubules. 1212 2

The effects of P2 receptor agonists on cell size and intracellular calcium levels, [Ca(2+)](i), was investigated using cultured endothelial cells isolated from the caudal artery of male Wistar rats. Cell size and [Ca(2+)](i) were measured using a phase-contrast and fluorescent confocal microscopic image analyzer and a Calcium Green fluorescence probe. P2Y receptor agonists, 2-methylthio ATP (2meS-ATP), ADP, UTP and ATP decreased the cell size and increased [Ca(2+)](i) in endothelial cells from rat caudal artery. However, alpha,beta-methylene ATP, a P2X receptor agonist, did not induce these responses. The decrease in size and the increase in [Ca(2+)](i), by 2meS-ATP were blocked by PPADS (P2-antagonist), suramin (P2-antagonist), thapsigargin (Ca(2+) pump inhibitor) and U-73122 (phospholipase C inhibitor). The present results show that activation of P2Y receptors, not P2X receptors, induces a decrease in cell size and an increase in [Ca(2+)](i), and the pharmacological properties of these two responses are the same. We concluded that the size of endothelial cells is regulated by P2Y receptors via intracelluar Ca(2+) derived from Ca(2+) stores.
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PMID:P2Y-receptor regulates size of endothelial cells in an intracellular Ca2+ dependent manner. 1253 13

The effect of extracellular ATP (10-100 microM) on intracellular Ca2+ concentration ([Ca2+]i) and firing rate has been studied in single pacemaker cells isolated from the sinus venosus of cane toads. In spontaneously firing cells, ATP initially increased peak [Ca2+]i by 43 +/- 5 %, increased diastolic [Ca2+]i by 20 + 3 % and increased the firing rate by 58 +/- 8 %. These early effects were followed by a late phase in which both the peak [Ca2+]i and the firing rate declined. Adenosine, and UTP (respectively, P1- and P2Y2,4,6-selective agonists) caused no significant change in [Ca2+]i or firing rate, while alphabeta-methylene ATP (a P2X1,3 agonist) caused a small increase in firing rate but no changes in [Ca2+]i. In contrast the P2Y1-selective agonist 2-MesADP (1 microM) mimicked the biphasic effects of ATP and these effects were inhibited by the purinoceptor antagonists suramin and PPADS and by the P2Y1-selective antagonist MRS 2179. Immunohistochemistry established that P2Y1 purinoceptors were present on the cell surface. Western blotting analysis demonstrated that the P2Y1 antibody recognised a 57 kDa protein. After sarcoplasmic reticulum Ca2+ release was prevented with caffeine or ryanodine, ATP no longer had any effect on [Ca2+]i or firing rate. Furthermore, the SR Ca2+ store content was decreased during the late phase of 2-MesADP application. The effect of ATP was coupled to phospholipase C (PLC) activity because the PLC inhibitor U-73122 eliminated the effects of ATP. Our study shows that in toad pacemaker cells, the biphasic effects of ATP on pacemaker activity are mainly through P2Y1 purinoceptors, which are able to modulate Ca2+ release from the SR Ca2+ store.
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PMID:ATP modulates intracellular Ca2+ and firing rate through a P2Y1 purinoceptor in cane toad pacemaker cells. 1294 18

ATP receptors present on rat alveolar macrophages (NR8383 cells) were identified by recordings of membrane current, measurements of intracellular calcium, RT-PCR and immunocytochemistry. In whole-cell recordings with a sodium-based internal solution, ATP evoked an inward current at -60 mV. This reversed at 0 mV. The EC50 for ATP was 18 microM in normal external solution (calcium 2 mm, magnesium 1 mm). The currents evoked by 2',3-O-(4-benzoyl)benzoyl-ATP were about five-fold smaller than those observed with ATP. ADP, UTP and alphabeta-methylene-ATP (alphabetameATP) (up to 100 microM) had no effect. ATP-evoked currents were potentiated up to ten-fold by ivermectin and were unaffected by suramin (30-100 microM), pyridoxal-phosphate-6-azophenyl-(2,4-sulphonic acid) (30-100 microM), and brilliant blue G (1 microM). In whole-cell recordings with a potassium-based internal solution and low EGTA (0.01 mm), ATP evoked an inward current at -60 mV that was followed by larger outward current. ADP and UTP (1-100 microM) evoked only outward currents; these reversed polarity at the potassium equilibrium potential and were blocked by apamin (10 nm). Outward currents were also blocked by the phospholipase C inhibitor U73122 (1 microM), and they were not seen with higher intracellular EGTA (10 mm). Suramin (30 microM) blocked the outward currents evoked by ATP and UTP, but not that evoked by ADP. PPADS (10 microM) blocked the ADP-evoked outward current without altering the ATP or UTP currents. RT-PCR showed transcripts for P2X subunits 1, 4 and 7 (not 2, 3, 5, 6) and P2Y receptors 1, 2, 4 and 12 (not 6). Immunocytochemistry showed strong P2X4 receptor expression partly associated with the membrane, weak P2X7 staining that was not associated with the cell membrane, and no P2X1 receptor immunoreactivity. We conclude that rat alveolar macrophages express (probably homomeric) P2X4 receptors, but find no evidence for other functional P2X subtypes. The P2Y receptors are most likely P2Y1 and P2Y2 and these couple through phospholipase C to an increase in intracellular calcium and the opening of SK type potassium channels.
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PMID:P2X4, P2Y1 and P2Y2 receptors on rat alveolar macrophages. 1297 84

Kidney tubules are targets for the activation of locally released nucleotides through multiple P2 receptor types. Activation of these P2 receptors modulates cellular Ca(2+) signaling and downstream cellular function. The purpose of this study was to determine whether P2 receptors were present in mIMCD-3 cells, a mouse inner medullary collecting duct cell line, and if so, to examine their link with intracellular Ca(2+) homeostasis. To monitor intracellular Ca(2+) concentration ([Ca(2+)](i)), experiments were conducted using the fluorescent dye fura 2. ATP (0.1-100 microM) produced a dose-dependent increase in [Ca(2+)](i) in a physiological Ca(2+)-containing solution, with an EC(50) of 2.5 microM. The P2-receptor antagonist PPADS reduced the effect of ATP on [Ca(2+)](i), and the P1-receptor agonist adenosine caused only a small increase in [Ca(2+)](i). Preincubation of cells with the phospholipase C antagonist U-73122 blocked the ATP-induced increase in [Ca(2+)](i), indicating P2Y receptors were involved in this process. In a Ca(2+)-free bath solution, thapsigargin and ATP induced intracellular Ca(2+) release from an identical pool. Nucleotides caused an increase in [Ca(2+)](i) in the potency order of UTP = ATP > ATP gamma S > ADP > UDP that is best fitted with the P2Y(2) subtype profile. Although the P2Y agonist UTP induced a similar large transient increase in [Ca(2+)](i) as did ATP, a small but sustained increase in [Ca(2+)](i) occurred only in ATP-stimulated cells, suggesting the role of P2X receptors in Ca(2+) influx. The sustained increase in [Ca(2+)](i) could be blocked by either nonselective cation channel blockers Gd(3+) or P2X antagonists PPADS and PPNDS. Furthermore, when either Gd(3+) or PPNDS was applied to the bath solution before ATP application, the ATP-induced increase in [Ca(2+)](i) was significantly reduced. Both RT-PCR and Western blotting corroborated the presence of P2X(1) and P2Y(2) receptors. These studies demonstrate that mIMCD-3 cells have both P2X and P2Y subtype receptors and that the activation of both P2X and P2Y receptors by extracellular ATP appears to be required to regulate intracellular Ca(2+) signaling.
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PMID:Extracellular ATP-induced calcium signaling in mIMCD-3 cells requires both P2X and P2Y purinoceptors. 1506 72

We investigated the effects of P2-receptor agonists on cell size, intracellular calcium levels ([Ca(2+)](i)), and permeation of FITC-labeled dextran (FD-4) as well as the relationship between these effects in human umbilical vein endothelial cells (HUVEC). FD-4 concentration, cell size, and [Ca(2+)](i) were analyzed by HPLC with fluorescence, phase contrast microscopic imaging, and fluorescent confocal microscopic imaging, respectively. The P2Y(1)-receptor agonists 2-methylthio ATP (2meS-ATP) and ADP decreased cell size and increased [Ca(2+)](i) in HUVEC. The P2Y(2)-receptor agonist UTP increased [Ca(2+)](i), but did not influence cell size. The P2X-receptor agonist alpha,beta-methylene ATP did not induce either response. The decrease in size and increase in [Ca(2+)](i) by 2meS-ATP were blocked by pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS, P2Y(1)-antagonist), thapsigargin (Ca(2+)-pump inhibitor), and U73122 (phospholipase C inhibitor). Furthermore, 2meS-ATP (P2Y(1)-receptor agonist) enhanced permeation of FD-4 through the endothelial cell monolayer. The 2meS-ATP-induced enhancement of the permeation was also prevented by PPADS, thapsigargin, and U73122. These results indicate that activation of P2Y receptors induces a decrease in cell size, an increase in [Ca(2+)](i), and may participate in facilitating macromolecular permeability in HUVEC.
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PMID:P2Y receptor-mediated Ca(2+) signaling increases human vascular endothelial cell permeability. 1521 41

1. The object of the present study was to clarify the neurotransmitters controlling membrane responses to electrical field stimulation (EFS) in the longitudinal smooth muscle cells of the chicken anterior mesenteric artery. 2. EFS (5 pulses at 20 Hz) evoked a depolarization of amplitude 19.7+/-2.1 mV, total duration 29.6+/-3.1 s and latency 413.0+/-67.8 ms. This depolarization was tetrodotoxin (TTX)-sensitive and its amplitude was partially decreased by atropine (0.5 microM); however, its duration was shortened by further addition of prazosin (10 microM). 3. Atropine/prazosin-resistant component was blocked by the nonspecific purinergic antagonist, suramin, in a dose-dependent manner, indicating that this component is mediated by the neurotransmitter adenosine 5'-triphosphate (ATP). 4. Neither desensitization nor blocking of P2X receptor with its putative receptor agonist alpha,beta-methylene ATP (alpha,beta-MeATP, 1 microM) and its antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulfonic (PPADS, up to 50 microM), had significant effect on the purinergic depolarization. In contrast, either desensitization or blocking of P2Y receptor with its putative agonist 2-methylthioATP (2-MeSATP, 1 microM) and its antagonist Cibacron blue F3GA (CBF3GA, 10 microM) abolished the purinergic depolarization, indicating that this response is mediated through P2Y but not P2X receptor. 5. The purinergic depolarization was inhibited by pertussis toxin (PTX, 600 ng ml(-1)). Furthermore, it was significantly inhibited by a phospholipase C (PLC) inhibitor, U-73122 (10 microM), indicating that the receptors involved in mediating the purinergic depolarization are linked to a PTX-sensitive G-protein, which is involved in a PLC-mediated signaling pathway. 6. Data of the present study suggest that the EFS-induced excitatory membrane response occurring in the longitudinal smooth muscle of the chicken anterior mesenteric artery is mainly purinergic in nature and is mediated via P2Y purinoceptors.
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PMID:An electrophysiological study of excitatory purinergic neuromuscular transmission in longitudinal smooth muscle of chicken anterior mesenteric artery. 1568 11

The pancreatic hormone glucagon hyperpolarizes the liver cell membrane. In the present study, we investigated the cellular signalling pathway of glucagon-induced hyperpolarization of liver cells by using the conventional microelectrode method. The membrane potential was recorded in superficial liver cells of superfused mouse liver slices. In the presence of the K+ channel blockers tetraethylammonium (TEA, 1 mmol/l) and Ba2+ (BaCl2, 5 mmol/l) and the blocker of the Na+/K+ ATPase, ouabain (1 mmol/l), no glucagon-induced hyperpolarization was observed confirming previous findings. The hyperpolarizing effect of glucagon was abolished by the leukotriene B4 receptor antagonist CP 195543 (0.1 mmol/l) and the purinergic receptor antagonist PPADS (5 micromol/l). ATPgammaS (10 micromol/l), a non-hydrolyzable ATP analogue, induced a hyperpolarization of the liver cell membrane similar to glucagon. U 73122 (1 micromol/l), a blocker of phospholipase C, prevented both the glucagon- and ATPgammaS-induced hyperpolarization. These findings suggest that glucagon affects the hepatic membrane potential partly by inducing the formation and release of leukotrienes and release of ATP acting on purinergic receptors of the liver cell membrane.
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PMID:Leukotriene and purinergic receptors are involved in the hyperpolarizing effect of glucagon in liver cells. 1584 96

We have synthesized a series of derivatives of the known P2 receptor antagonist PPADS (pyridoxal-5'-phosphate-6-azo-phenyl-2,4-disulfonate) and examined their ability to inhibit functional activity of the recombinant human P2Y13 nucleotide receptor expressed in 1321N1 human astrocytoma cells co-expressing G(alpha)16 protein (AG32). Analogues of PPADS modified through substitution of the phenylazo ring, including halo and nitro substitution, and 5'-alkyl phosphonate analogues were synthesized and tested. A 6-benzyl-5'-methyl phosphonate analogue was prepared to examine the effect of stable replacement of the azo linkage. The highest antagonistic potency was observed for 6-(3-nitrophenylazo) derivatives of pyridoxal-5'-phosphate. The 2-chloro-5-nitro analogue (MRS 2211) and 4-chloro-3-nitro analogue (MRS 2603) inhibited ADP (100 nM)-induced inositol trisphosphate (IP3) formation with pIC50 values of 5.97 and 6.18, respectively, being 45- and 74-fold more potent than PPADS. The antagonism of MRS 2211 was competitive with a pA2 value of 6.3. MRS2211 and MRS2603 inhibited phospholipase C (PLC) responses to 30 nM 2-methylthio-ADP in human P2Y1 receptor-mediated 1321N1 astrocytoma cells with IC50 values of >10 and 0.245 microM, respectively. Both analogues were inactive (IC50>10 microM) as antagonists of human P2Y12 receptor-mediated PLC responses in 1321N1 astrocytoma cells. Thus, MRS2211 displayed >20-fold selectivity as antagonist of the P2Y13 receptor in comparison to P2Y1 and P2Y12 receptors, while MRS2603 antagonized both P2Y1 and P2Y13 receptors.
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PMID:Synthesis of pyridoxal phosphate derivatives with antagonist activity at the P2Y13 receptor. 1591 66

ATP is proposed to be a major inhibitory neurotransmitter in the gastrointestinal (GI) tract, causing hyperpolarization and smooth muscle relaxation. ATP activates small-conductance Ca(2+)-activated K(+) channels that are involved in setting the resting membrane potential and causing inhibitory junction potentials. No reports are available examining the effects of ATP on voltage-dependent inward currents in GI smooth muscle cells. We previously reported two types of voltage-dependent inward currents in murine proximal colonic myocytes: a low-threshold voltage-activated, nonselective cation current (I(VNSCC)) and a relatively high-threshold voltage-activated (L-type) Ca(2+) current (I(L)). Here we have investigated the effects of ATP on these currents. External application of ATP (1 mM) did not affect I(VNSCC) or I(L) in dialyzed cells. ATP (1 mM) increased I(VNSCC) and decreased I(L) in the perforated whole-cell configuration. UTP and UDP (1 mM) were more potent than ATP on I(VNSCC). ADP decreased I(L) but had no effect on I(VNSCC). The order of effectiveness was UTP = UDP > ATP > ADP. These effects were not blocked by pyridoxal phosphate-6-azo(benzene-2,4-disulfonic acid) (PPADS), but the phospholipase C inhibitor U-73122 reversed the effects of ATP on I(VNSCC). ATP stimulation of I(VNSCC) was also reversed by protein kinase C (PKC) inhibitors chelerythrine chloride or bisindolylmaleimide I. Phorbol 12,13-dibutyrate mimicked the effects of ATP. RT-PCR showed that P2Y(4) is expressed by murine colonic myocytes, and this receptor is relatively insensitive to PPADS. Our data suggest that ATP activates I(VNSCC) and depresses I(L) via binding of P2Y(4) receptors and stimulation of the phospholipase C/PKC pathway.
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PMID:Nucleotide regulation of the voltage-dependent nonselective cation conductance in murine colonic myocytes. 1672 14


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