Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.16.2 (PCP)
 3,761 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

[3H]ryanodine binding to and Ca2+ release from microsomal fractions derived from canine cerebrum (CBR) and cerebellum (CBL) were investigated. High-affinity ryanodine binding sites were detected in both cerebrum and cerebellum microsomes [CBR: maximal binding capacity (Bmax) = 446 fmol/mg protein, dissociation constant (Kd) = 9 nM, Hill coefficient (n) = 0.95; CBL: Bmax = 650, Kd = 12, n = 1.8]. Ryanodine binding in both fractions was increased by millimolar concentrations of ATP [or its nonhydrolyzable analogue beta, gamma-methyleneadenosine 5'-triphosphate (AMP-PCP)] and micromolar concentrations of Ca2+ but was decreased by micromolar concentrations of ruthenium red, similar to that found in sarcoplasmic reticulum (SR) of striated muscle. The addition of caffeine or the sudden elevation of extravesicular Ca2+ induced a rapid La(3+)-sensitive Ca2+ release from both CBR and CBL microsomal fractions with rate constants of approximately 100 s-1, as determined by stopped-flow photometry of the Ca2+ indicator arsenazo III. The release of Ca2+ was activated by either millimolar ATP or AMP-PCP, blocked by micromolar concentrations of La3+, and significantly inhibited by 50 microM ryanodine. Mg2+ and ruthenium red in millimolar and micromolar concentrations, respectively, caused only a slight inhibition of Ca2+ release. These results indicate that rapid Ca2+ release occurs from caffeine-, Ca2+- and ryanodine-sensitive Ca2+ stores in both CBR and CBL microsomal fractions.
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PMID:Caffeine- and ryanodine-sensitive Ca2+ stores of canine cerebrum and cerebellum neurons. 172 42

The locus coeruleus (LC) has been hypothesized to play an important role in opiate withdrawal. This hypothesis is supported, in part, by the finding that LC neurons greatly increase their activity during antagonist-precipitated morphine withdrawal and that this increased activity correlates temporally with withdrawal behavior. However, this withdrawal-induced increase in unit activity is not seen in vitro in brain slices taken from morphine-dependent animals, indicating that afferents to the LC play an important role in the withdrawal-induced activation of these neurons. This chapter reviews data indicating: (1) the morphine-withdrawal-induced activation of LC neurons is mediated predominantly by non-N-methyl-D-aspartate (NMDA) excitatory amino acid pathways in the brain; (2) the activation of the LC during morphine withdrawal may be mediated, at least in part, by an excitatory amino acid projection from the nucleus paragigantocellularis. The role of other excitatory amino acid pathways in the withdrawal-induced activation of the LC remains to be determined; (3) intrinsic changes in the G-protein/cyclic AMP system of LC cells may play an important role in mediating the effects of afferent inputs to the LC during morphine withdrawal; (4) NMDA antagonists (unlike the alpha 2 agonist clonidine) attenuate the behavioral signs of morphine withdrawal without blocking the withdrawal-induced increase of LC unit activity. In addition, non-competitive NMDA antagonists like MK801 may not be useful to alleviate opiate-withdrawal symptoms in man because of their PCP-like side effects. However, competitive NMDA antagonists like LY274614 could be of great benefit for alleviating opiate-withdrawal withdrawal symptoms in man.
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PMID:Afferent effects on locus coeruleus in opiate withdrawal. 183 35

ATP-induced increases of intracellular calcium concentration ([Ca2+]i) were measured as a function of flow rate in single cell recordings within a confluent endothelial cell monolayer. Although flow and its associated shear stress did not per se significantly alter basal [Ca2+]i, ATP-induced [Ca2+]i was exquisitely sensitive to flow. Step increases of flow in the presence of ATP triggered large [Ca2+]i transients that slowly (60-150 s) returned to basal values. ATP-releasable [Ca2+]i was mobilized from intracellular stores, as well as obtained from the extracellular medium. Since potent ectonucleotidases on the cell surface are expected to influence local ATP concentrations, experiments were repeated using the poorly hydrolyzable ATP analogue beta, gamma-methyleneadenosine 5'-triphosphate (AMP-PCP). Comparison between ATP and AMP-PCP responses suggested that flow regulates the mass transport of agonist to the endothelial cell surface by overcoming the local effects of degradative enzymes. An additional, quite different phenomenon of flow-mediated [Ca2+]i regulation in endothelial cells was observed when [Ca2+]i oscillations induced by AMP-PCP in the absence of flow were shown to be reversibly inhibited by step increases in flow. These results imply that the effectiveness of local or systemic agonists in stimulating endothelial transduction will vary with flow rates. Regional variations in hemodynamic shear stresses associated with altered flow patterns throughout the arterial system are predicted to result in large variations of vessel wall responsiveness to physiological and pathological agonists.
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PMID:Flow modulation of agonist (ATP)-response (Ca2+) coupling in vascular endothelial cells. 185 15

The motility of bile canaliculi was examined in hepatocyte couplets permeabilized with palmitoyl lysophosphatidyl choline in a dosage regimen that drastically affected secretory function, yet maintained relative integrity of the cellular cytoskeleton. The permeabilized cells showed no exclusion of trypan blue, notable cytoplasmic organelle and membrane damage, and no uptake or secretion of either fluorescein diacetate or sodium fluorescein. However, bile canalicular structure remained relatively intact and actin and myosin were localized immunocytochemically in the pericanalicular region. Coincident with the administration of 1 mM ATP, 2 mM Mg2+, and 1 microM Ca2+, the canaliculi contracted with partial or complete luminal closure. ADP, AMP, or AMP-PCP could not be substituted for ATP. A dose-dependent relationship was shown between ATP concentration and canalicular contraction rate. The permeabilization procedure also provided enhanced visualization of pericanalicular microfilaments, believed to be actin filaments, and their organization into two layers: an inner membrane-associated network, and an outer filament bundle that inserted into belt junctions (zonulae adherentes). The organization of the microfilament belt of contiguous hepatocytes was such that it formed a circumferential band of microfilaments around the canaliculus. It is analogous to contractile filament belts found in the apical terminal web region of other epithelia. It was also observed that with canalicular luminal closure, there was a change in the organization of the pericanalicular microfilaments. It is concluded that in hepatocyte couplets, differential sensitivity of cell components to permeabilization can be achieved with palmitoyl lysophosphatidyl choline. In addition, the results provide evidence that the bile canaliculus has the capacity to be a contractile structure even in the absence of secretion, that canalicular contraction is ATP-dependent, and hence is a dynamic process.
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PMID:Permeabilized hepatocyte couplets. Adenosine triphosphate-dependent bile canalicular contractions and a circumferential pericanalicular microfilament belt demonstrated. 188 Nov 22

Using lead citrate as a capture reagent and adenylate-(beta, gamma-methylene) diphosphate (AMP-PCP) as a substrate, we localized adenylate cyclase activity on the non-ruffled border plasma membrane of approximately half of the osteoclasts on trabecular bone surfaces in the tibial metaphyses of chickens fed a low (0.3%)-calcium diet. The enzyme was not detectable in osteoclasts when chickens were fed a normal calcium diet. Activity was observed on the entire plasma membrane of detached osteoclasts that were situated between osteoblasts on the bone surface and blood vessels in the marrow cavity. Detection of activity on detached osteoclasts required the presence of an activator, implying lower levels in these cells than in those with ruffled borders. Staining was greater on the lateral sides of osteoblasts and osteoclasts when they were in contact with each other. Reaction specificity was indicated by the demonstration of stimulation by forskolin, guanylate-(beta, gamma-methylene) diphosphate (GMP-PCP), dimethylsulfoxide, and NaF, inhibition by alloxan and 2',5'-dideoxyadenosine, and absence of activity when sections were incubated in substrate-free medium or when GMP-PCP replaced AMP-PCP as a substrate. The finding of adenylate cyclase in osteoclast plasma membrane provides structural evidence that the adenylate cyclase-cyclic AMP system has a role in regulation of osteoclast cell function. The low-calcium diet appears to have resulted in increased amounts of adenylate cyclase in osteoclasts.
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PMID:Ultrastructural localization of adenylate cyclase activity in chicken osteoclasts. 191 38

We have investigated the inhibition of Escherichia coli glutamine synthetase (GS) with alpha- and gamma-substituted analogues of phosphinothricin [L-2-amino-4-(hydroxymethylphosphinyl)butanoic acid (PPT)], a naturally occurring inhibitor of GS. These compounds display inhibition of bacterial GS that is competitive vs L-glutamate, with Ki values in the low micromolar range. At concentrations greater than Ki the phosphinothricins caused time-dependent loss of enzyme activity, while dilution after enzyme inactivation resulted in recovery of enzyme activity. ATP was required for inactivation; the nonhydrolyzable ATP analogue AMP-PCP failed to support inhibition of GS by the phosphinothricins. The binding of these inhibitors to the enzyme was also characterized by measurement of changes in protein fluorescence, which provided similar inactivation rate constants k1 and k2 for the entire series of compounds. Rate constants koff for recovery were also determined by fluorescence measurement and were comparable for both PPT and the gamma-hydroxylated analogue GHPPT and significantly greater for the alpha- and gamma-alkyl-substituted compounds. Electron paramagnetic resonance spectra provided information on the interaction of the phosphinothricins with the manganese form of the enzyme in the absence of ATP, and significant binding was observed for PPT and GHPPT. 31P NMR experiments confirmed that enzyme inactivation is accompanied by hydrolysis of ATP, although phosphorylated phosphinothricins could not be detected in solution. The kinetic behavior of these compounds is consistent with a mechanism involving inhibitor phosphorylation, followed by release from the active site and simultaneous hydrolysis to form Pi and free inhibitor.
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PMID:Inhibition of Escherichia coli glutamine synthetase by alpha- and gamma-substituted phosphinothricins. 196 48

1. The ontogeny of responses to purines and analogues of smooth muscle preparations was studied in rat duodenum and rat urinary bladder. 2. Responses to adenosine and to adenosine 5'-triphosphate (ATP) mediated by P1- and P2-purinoceptors respectively were present as early as postnatal day 2, the earliest day studied. 3. In rat bladder, adenosine was inhibitory and ATP and adenosine 5'-(beta, gamma-methylene) triphosphonate (AMP-PCP) were excitatory, acting on the P2X subtype of P2-purinoceptors. Adenosine was more potent in the neonate than in the adult, while the potency of the nucleotides initially increased with age but then declined, being highest between postnatal days 10 and 25. 4. In rat duodenum also, adenosine was inhibitory, its potency being less than the adult before day 15. 5. ATP at low concentrations was inhibitory in rat duodenum at every age studied and its potency increased with age, but higher concentrations of ATP (3 microM and above) were excitatory until day 15. Both relaxations and contractions were mediated by the P2Y subtype of P2-purinoceptors. These ATP-induced contractions were not inhibited by indomethacin (25 microM) or by tetrodotoxin (1 microM) and are therefore not due to prostaglandin synthesis or to ATP-induced release of transmitter substances from nerves. 6. These results show that responses to adenosine and to adenine nucleotides are present from birth and vary with age, and that the changes seen indicate a differential development for P1-, P2X- and P2Y-purinoceptors.
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PMID:The ontogeny of purinoceptors in rat urinary bladder and duodenum. 220 6

Uptake of the catecholamines (CA), dopamine (DA) and norepinephrine (NE) into synaptosomes prepared from rat and bovine brains was potentiated by ATP (from 0.1 to 5.0 mM) in a dose-dependent manner. Other nucleotides, particularly the nonhydrolyzable ATP analogs beta,gamma-imidoadenosine-5'-triphosphate (AMP-PNP) and beta,gamma-methyladenosine-5'-triphosphate (AMP-PCP) also potentiated [3H]DA and [3H]NE uptake. Several endogenous 5'-nucleotide triphosphates (e.g. GTP, UTP and CTP) potentiated [3H]CA uptake, but were less effective than ATP. Among the ATP metabolites, only ADP potentiated uptake whereas AMP and adenosine did not. [3H]Dopamine uptake measured in Krebs bicarbonate buffer had a Km of 2.1 microM and a Vmax of 163.9 pmol/mg prot./min. In presence of ATP, [3H]DA uptake had much higher affinity (Km = 0.56 microM) and larger capacity (Vmax = 333 pmol/mg prot./min) than uptake in absence of added ATP. Furthermore, [3H]DA uptake in presence of ATP had faster rate of uptake, and was independent of temperature while in absence of added ATP it was temperature-dependent. This ATP-dependent [3H]DA uptake was retained by synaptosomal ghosts that were obtained after lysing the striatal synaptosomes and removing their contents of synaptic vesicles and mitochondria. It is proposed that, in addition to the carrier-mediated (neuronal) uptake of CA, there is neuronal uptake that is regulated by ATP and inhibited by cocaine, which may be more relevant for terminating the synaptic action of CA because of its faster rate of uptake and larger capacity.
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PMID:ATP-regulated neuronal catecholamine uptake: a new mechanism. 240 89

Ionic gradients imposed by choline Cl replacement of K methanesulfonate (Mes) at constant [K][Cl] product stimulate 45Ca efflux from skinned muscle fibers; a small, sustained Ca2+-insensitive efflux component, observed in EGTA, appears to grade a much larger Ca2+-dependent component responsible for contractile activation and is likely to reflect intermediate steps in excitation-contraction coupling. The present studies examined ATP-related effects on the Ca2+-insensitive stimulation. 45Ca efflux was measured on segments of frog semitendinosus muscle skinned by microdissection, with isometric force monitored continuously. The Ca2+-insensitive component was potentiated by quercetin, a flavonoid thought to inhibit the sarcoplasmic reticulum (SR) Ca pump by stabilizing a phosphorylated intermediate. Quercetin increased the stimulated net 45Ca release in the absence of EGTA, as expected from inhibition of reaccumulation, but its effectiveness in EGTA indicated potentiation of unidirectional efflux as such. Quercetin also increased unstimulated (control) 45Ca efflux in EGTA, to a smaller extent; potentiation appeared to be a function of efflux, with stimulation above control loss increased approximately 2.6-fold. ATP removal before stimulation, which led to rigor force and increased stiffness, prevented all quercetin effects in EGTA. ATP removal by itself inhibited ionic stimulation of the Ca2+-insensitive component, with little residual increase above the parallel control loss. Addition of the nonhydrolyzable ATP analogue AMP-PCP ([adenylyl-beta,gamma-methylene]diphosphate) (0.8 mM) after ATP removal gave similar results to ATP-free solution, which suggests that adenine nucleotide binding alone does not support stimulation by choline Cl. These results imply a fundamental role for ATP in the excitation of skinned fibers by imposed diffusion potentials; they also suggest that ATP regulates the SR Ca efflux channel, in a manner that could provide the positive feedback in Ca2+-dependent Ca release.
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PMID:Excitation of skinned muscle fibers by imposed ion gradients. II. Influence of quercetin and ATP removal on the Ca2+-insensitive component of stimulated 45Ca efflux. 241 70

A subpopulation of canine cardiac sarcoplasmic reticulum vesicles has been found to contain a "Ca2+ release channel" which mediates the release of intravesicular Ca2+ stores with rates sufficiently rapid to contribute to excitation-contraction coupling in cardiac muscle. 45Ca2+ release behavior of passively and actively loaded vesicles was determined by Millipore filtration and with the use of a rapid quench apparatus using the two Ca2+ channel inhibitors, Mg2+ and ruthenium red. At pH 7.0 and 5-20 microM external Ca2+, cardiac vesicles released half of their 45Ca2+ stores within 20 ms. Ca2+-induced Ca2+ release was inhibited by raising and lowering external Ca2+ concentration, by the addition of Mg2+, and by decreasing the pH. Calmodulin reduced the Ca2+-induced Ca2+ release rate 3-6-fold in a reaction that did not appear to involve a calmodulin-dependent protein kinase. Under various experimental conditions, ATP or the nonhydrolyzable ATP analog, adenosine 5'-(beta, gamma-methylene)triphosphate (AMP-PCP), and caffeine stimulated 45Ca2+ release 2-500-fold. Maximal release rates (t1/2 = 10 ms) were observed in media containing 10 microM Ca2+ and 5 mM AMP-PCP or 10 mM caffeine. An increased external Ca2+ concentration (greater than or equal to 1 mM) was required to optimize the 45Ca2+ efflux rate in the presence of 8 mM Mg2+ and 5 mM AMP-PCP. These results suggest that cardiac sarcoplasmic reticulum contains a ligand-gated Ca2+ channel which is activated by Ca2+, adenine nucleotide, and caffeine, and inhibited by Mg2+, H+, and calmodulin.
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PMID:Rapid calcium release from cardiac sarcoplasmic reticulum vesicles is dependent on Ca2+ and is modulated by Mg2+, adenine nucleotide, and calmodulin. 243 95


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