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:3.6.1.3 (ATPase)
65,361 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

It has recently been shown that two novel tachykinins, ranakinin and [Leu3, Ile7]neurokinin A, are present in fibers innervating the frog adrenal gland, and it has been demonstrated that tachykinins stimulate corticosteroid secretion in vitro through activation of chromaffin cells. The purpose of the present study was to investigate the effect of ranakinin on cytosolic free calcium concentrations ([Ca2+]i) and to determine the source of calcium involved. Cultured adrenal cells were loaded with the fluorescent calcium indicator indo-1, and changes in [Ca2+]i were studied using dual emission wavelength microfluorimetry. Administration of a brief pulse of ranakinin (1 microM; 1 sec) in the vicinity of chromaffin cells caused an immediate and transient increase in [Ca2+]i. Repeated pulses of ranakinin resulted in a gradual decline in the [Ca2+]i response, suggesting the occurrence of a desensitization phenomenon. Preincubation of the cells with the calcium channel blockers nifedipine (10 microM) and omega-conotoxin (1 microM) did not alter the response of chromaffin cells to ranakinin. Chelation of extracellular calcium by EGTA (10 mM) caused a marked decrease in the basal [Ca2+]i, but did not suppress the ranakinin-induced [Ca2+]i increase. Conversely, incubation of the cells with thapsigargin (10 microM), an inhibitor of calcium adenosine triphosphatase activity, abolished the stimulatory effect of ranakinin, indicating that the increase in [Ca2+]i can be ascribed to mobilization of calcium from intracellular stores. Preincubation of adrenal cells with the phospholipase C antagonist U-73122 (1 microM; 18 min) or with pertussis toxin (10 microM; 18 h) totally blocked the ranakinin-induced [Ca2+]i rise. Taken together, these data indicate that in frog adrenochromaffin cells, ranakinin causes mobilization of calcium from intracellular stores. The effect of ranakinin is mediated through activation of a phospholipase C via a pertussis toxin-sensitive G protein.
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PMID:Effect of ranakinin, a novel tachykinin, on cytosolic free calcium in frog adrenochromaffin cells. 766 74

Agents that deplete cells of K+ without grossly disrupting the plasma membrane were found to stimulate the cleavage of pro-interleukin (IL)-1 beta to mature IL-1 beta. Agents examined in this study included staphylococcal alpha-toxin and gramicidin, both of which selectively permeabilize plasma membranes for monovalent ions, the ionophores nigericin and valinomycin, and the Na+/K+ ATPase inhibitor ouabain. K+ depletion by brief hypotonic shock also triggered processing of pro-IL-1 beta. The central role of K+ depletion for inducing IL-1 beta maturation was demonstrated in cells permeabilized with alpha-toxin: processing of pro-IL-1 beta was totally blocked when cells were suspended in medium that contained high K+, but could be induced by replacing extracellular K+ with Na+, choline+ or sucrose. To test whether K+ flux might also be important in physiological situations, monocytes were stimulated with lipopolysaccharide (LPS) for 1-2 h to trigger pro-IL-1 beta synthesis, and transferred to K(+)-rich medium. This maneuver totally suppressed IL-1 beta maturation. Even after 16 h, however, removal of K+ from the medium resulted in rapid processing and export of IL-1 beta. Ongoing export of mature IL-1 beta from cells stimulated with LPS for 2-6 h could also be arrested by transfer to K(+)-rich medium. Moreover, a combination of two K+ channel blockers inhibited processing of IL-1 beta in LPS-stimulated monocytes. We hypothesize that K+ movement and local K+ concentrations directly or indirectly influence the action of interleukin-1 beta-converting enzyme (ICE) and, possibly, of related intracellular proteases.
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PMID:Potassium-inhibited processing of IL-1 beta in human monocytes. 773 13

Smooth muscle was made permeable with alpha-toxin and beta-escin. ATPase activity was measured using a phosphoenolpyruvate-pyruvate kinase regenerating system for ATP that was monitored by NADH fluorescence changes, and Ca2+ was measured using fura 2 fluorescence. alpha-Toxin-and beta-escin-treated bundles of cells had a high ATPase activity, which was reduced 80% when exposed to 1% Triton X-100. This Triton-sensitive ATPase activity was increased by approximately 20% when GTP or GTP gamma S was added to the solutions and was of much greater magnitude than the Ca(2+)-activated ATPase associated with contraction. This high membrane ATPase activity will cause a gradient of ATP into and ADP out of the bundle of cells. Thus modulation of this ATPase by G-protein-receptor mechanisms could alter the force at a constant Ca2+ concentration by changing the ADP/ATP ratio within the cells. Measurements of the fura 2 fluorescence ratio (340/380) in alpha-toxin-treated bundles of cells following sudden changes in extracellular Ca2+ showed that the cells were not freely permeable to Ca EGTA. Similar experiments in beta-escin-treated cells showed the cells to be much more permeable to Ca EGTA. These experiments indicate that great care must be taken in alpha-toxin- and beta-escin-treated fibers to make sure that the intracellular ATP, ADP, and Ca2+ are held constant.
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PMID:Relationship between ATPase activity, Ca2+, and force in alpha-toxin- and beta-escin-treated smooth muscle. 776 79

The Drosophila proteins, Trp and Trpl, are suggested to be cation channels responsible for depolarization of the receptor potential associated with stimulation of insect photoreceptor cells by light. Consistent with this hypothesis, we recently showed that recombinant Trpl forms Ca(2+)- and Ba(2+)-permeable non-selective cation channels when expressed in Sf9 cells using the baculovirus expression vector. As Trpl may be activated in the photoreceptor cell after stimulation of phospholipase C, we hypothesized that a similar regulation of recombinant Trpl may be observed in the Sf9 cell after activation of heterologous membrane receptors linked to Ca(2+)-signal-transduction pathways. To test this hypothesis, Ca2+ signalling was examined in Fura-2-loaded Sf9 cells infected with baculovirus containing cDNA for the M5 muscarinic receptor alone (M5 cells) or in cells co-infected with both M5 and Trpl-containing baculoviruses (M5-Trpl cells). Addition of carbachol (100 microM) to M5 cells produced an increase in cytosolic free Ca2+ concentration ([Ca2+]i) (mean +/- S.D.; n = 17) from 101 +/- 20 to 762 +/- 178 nM which declined to a sustained elevated level of 384 +/- 102 nM after 3 min. The sustained component was eliminated by removal of extracellular Ca2+ or by addition of La3+ or Gd3+ (10 microM). In M5-Trpl cells, basal [Ca2+]i increased as a function of time after infection. To evaluate the contribution of Ca2+ influx to the overall profile observed, Ba2+, a Ca2+ surrogate that is not a substrate for the Ca2+ pump, was used. The increase in basal [Ca2+]i seen in M5-Trpl cells was associated with an increase in basal Ba2+ influx. Addition of carbachol to M5-Trpl cells at 30-36 h after infection produced a large increase in [Ca2+]i to a sustained value of 677 +/- 143 nM. This change in [Ca2+]i was (1) blocked by atropine, (2) attenuated in the absence of extracellular Ca2+, and (3) relatively insensitive to La3+, but blocked by Gd3+ in the 0.1-1 mM range. In the presence of 10 microM Gd3+ to block the endogenous-receptor-mediated Ca(2+)-influx in M5-Trpl cells. In sharp contrast increase in Ba2+ influx in M5-Trpl cells. In sharp contrast, neither Ca2+ nor Ba2+ influx through Trpl was affected by thapsigargin, a selective inhibitor of the endoplasmic reticulum Ca(2+)-ATPase pump.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Receptor-mediated activation of recombinant Trpl expressed in Sf9 insect cells. 783 80

ATP and UTP cause mobilization of Ca2+ from the intracellular stores with similar potency in several cell types including both undifferentiated and differentiated HL60 cells. We show here that, in HL60 cells with Ca2+ stores that had been fully and irreversibly emptied using the endomembrane Ca(2+)-ATPase inhibitor thapsigargin, both nucleotides produced a biphasic effect on Ca2+ entry, first rapid inhibition and then delayed (about 15 s) activation. ATP was more effective at producing the initial inhibition of Ca2+ entry, whereas UTP was more effective at activating the delayed Ca2+ entry. Previous incubation with UTP desensitized the Ca2+ mobilization and the delayed activation of Ca2+ entry induced by ATP but not the inhibition of Ca2+ entry. The ATP analogue 2-methylthioATP (2-MeSATP) barely mobilized stored Ca2+ but inhibited Ca2+ entry. These results could be explained by the presence of two receptors: (i) a P2u receptor sensitive to ATP and UTP, responsible for activation of phospholipase C and Ca2+ mobilization, early inhibition of Ca2+ entry and delayed activation of Ca2+ entry and (ii) a P2y-like receptor sensitive to ATP and 2-MeSATP which produces only inhibition of Ca2+ entry. The inhibition of Ca2+ entry by nucleotides increased greatly during differentiation. Given that Ca2+ mobilization by nucleotides is not modified by differentiation, this suggests that a component of the mechanism of inhibition of Ca2+ entry is gradually expressed during differentiation of HL60 cells.
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PMID:Biphasic and differential modulation of Ca2+ entry by ATP and UTP in promyelocytic leukaemia HL60 cells. 784 89

The mechanisms by which guanosine 3',5'-cyclic monophosphate (cGMP) modulates the contraction induced by ATP were investigated in small mesenteric resistance arteries of the rat. The nitric oxide donors 3-morpholinosydnonimine (SIN-1, 10 microM) and sodium nitroprusside (SNP, 10 microM) increased cGMP but not adenosine 3',5'-cyclic monophosphate (cAMP) content of the tissue. SIN-1, SNP, and 8-bromoguanosine 3',5'-cyclic monophosphate (8-BrcGMP, 100 microM) inhibited the myosin light chain phosphorylation and the contractile response to ATP. Both effects were completely reversed by the selective inhibitor of cGMP protein kinase, Rp-8-bromoguanosine 3',5'-cyclic monophosphorothioate (30 microM). The sensitivity to Ca2+ of arteries permeabilized with Staphylococcus aureus alpha-toxin (4,000 hemolytic units/ml) was not affected by 8-BrcGMP. The two nitric oxide donors and 8-BrcGMP decreased the rise in intracellular Ca2+ induced by ATP. The vasodilator agents abolished the contractile response to the exogenous calcium in vessels that were exposed to 3 mM ATP after depletion of intracellular Ca2+ stores. Thapsigargin (1 microM), an inhibitor of the sarcoplasmic reticulum Ca(2+)-adenosinetriphosphatase, reversed the inhibitory effect of the vasodilator agents when the contraction induced by ATP was elicited in the presence of the Ca2+ entry blocker nitrendipine (1 microM) or in Ca(2+)-free medium. These results show that cGMP inhibits ATP-induced contraction by decreasing intracellular Ca2+ concentration in small resistance arteries. They indicate that this effect results from decreased Ca2+ influx and enhanced Ca2+ sequestration through a thapsigargin-sensitive pump via activation of a cGMP protein kinase.
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PMID:Effects of cGMP on calcium handling in ATP-stimulated rat resistance arteries. 790 Aug 76

The relationship between the phospholipid composition of sarcoplasmic reticulum and the activity of the Ca2+, Mg2+-stimulated ATPase was analyzed by digestion of membrane phospholipids with phospholipase C and A2 enzymes of diverse specificity and by detergent extraction. Phospholipase C of Clostridium perfringens and Clostridium welchii, that hydrolyze preferentially phosphatidylcholine (PC), inhibited the Ca2+-ATPase activity parallel with the depletion of phosphatidylcholine from the membrane. Phospholipase C of Bacillus cereus hydrolyzed in addition to PC, phosphatidylethanolamine (PE) and phosphatidylserine (PS), causing complete inhibition of Ca2+-stimulated ATPase activity. Digestion of sarcoplasmic reticulum with the phospholipase A2 of snake or bee venom produced similar effects. The phosphatidylinositol (PI)-specific phospholipases of B. cereus and Bacillus thuringiensis caused less than 10% inhibition of the Ca2+-ATPase, accompanied by the hydrolysis of more than 70% of the phosphatidylinositol content of the membrane, without significant change in PC, PE and PS content. The inhibition of ATPase activity by the C type phospholipases was nearly completely reversed by octaethyleneglycol dodecyl ether (C12E8). These experiments suggest that the full phospholipid content of native sarcoplasmic reticulum (congruent to 100 mol phospholipid per mol Ca2+-ATPase), is required for ATPase activity and there is no indication that PE, PS, and PI play a specific role in ATP hydrolysis. Extraction of sarcoplasmic reticulum phospholipids by detergents such as deoxycholate, cholate and C12E8 also caused proportional inhibition of ATPase activity with the decrease in phospholipid content; the parallel extraction of PC, PE and PI left the phospholipid composition largely unchanged during delipidation. These observations do not support the requirement for a 'lipid annulus' of congruent to 30 phospholipid molecules/Ca2+-ATPase as proposed by Hesketh et al. ((1976) Biochemistry 15, 4145-4151) or the specific interaction of phosphatidylethanolamine with the ATPase molecule proposed by Bick et al. ((1991) Arch. Biochem. Biophys. 286, 346-352).
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PMID:The relationship between phospholipid content and Ca2+-ATPase activity in the sarcoplasmic reticulum. 798 4

1. Small strips from third-order branches of rabbit mesenteric artery (approximately 150-200 microM wide) contracted in response to noradrenaline (10 microM) or 5-hydroxytryptamine (5-HT; 10 microM) in oxygenated Krebs solution containing 2.5 mM Ca2+. In a Ca(2+)-free mock intracellular solution (0 Ca2+ plus 0.2 mM EGTA), noradrenaline (10 microM) and caffeine (10 mM) induced only a single, transient contraction in artery strips, while 5-HT (10 microM) failed to induce any response. 2. In strips of mesenteric artery which had been permeabilized with Staphylococcus alpha-toxin and bathed in Ca(2+)-free mock intracellular solution, noradrenaline (10 microM), caffeine (10 mM) and D-myo-inositol (1,4,5)-trisphosphate (IP3, 100 microM), but not 5-HT (10 or 100 microM) induced a transient contraction. In contrast to the non-permeabilized strips, contractions to noradrenaline, caffeine and IP3 were restored by prior incubation (10 min) in solution containing 0.08 microM Ca2+. The contractions to noradrenaline and IP3 in permeabilized muscle strips required the presence of 100 microM guanosine 5'-triphosphate (GTP), although in the absence of Ca2+. GTP alone did not induce contraction. 3. Exposure of permeabilized mesenteric artery strips to IP3 significantly reduced the subsequent contractile responses to caffeine. Contractile responses to caffeine and IP3 were abolished by the Ca(2+)-ATPase inhibitor, thapsigargin (1 microM). 4. Ca2+ (0.1-10 microM) induced concentration-dependent contraction in permeabilized artery strips. In strips which were submaximally contracted with 0.5 microM Ca2+/100 microM GTP, the subsequent addition of 5-HT (10 microM) stimulated further contraction. The protein kinase C inhibitor, H-7 (1 microM) abolished the 5-HT/GTP-induced contraction, but did not alter the contraction to Ca2+. 5. In non-permeabilized, endothelium-denuded segments of rabbit mesenteric artery bathed in Ca2+-replete Krebs solution, noradrenaline (10 microM) stimulated a rapid, transient accumulation of IP3. 5-HT(100 microM) failed to stimulate IP3 accumulation during exposure periods of up to 5 min. 5-HT (100 microM)did stimulate IP3 accumulation if the external K+ concentration was raised (to around 25 mM). This concentration of K+ alone did not stimulate IP3 production and the 5-HT-stimulated IP3 accumulation in the presence of elevated extracellular [K+] was abolished by the alpha l-adrenoceptor antagonist, prazosin(O.1 microM).6. These results suggest that intracellular Ca2+ release does not play an important role in 5-HT-induced smooth muscle contraction in the rabbit mesenteric artery. This is despite the fact that a significant intracellular Ca2+ pool is present in these cells, which can be discharged by either noradrenaline or IP3.However, 5-HT did stimulate smooth muscle contraction in the presence of raised intracellular calcium,suggesting that a component of the contraction to 5-HT will reflect an increase in myofilament Ca2+sensitivity, possibly due to the activation of protein kinase C.
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PMID:Importance of inositol (1,4,5)-trisphosphate, intracellular Ca2+ release and myofilament Ca2+ sensitization in 5-hydroxytryptamine-evoked contraction of rabbit mesenteric artery. 800 97

Rabbit portal veins were permeabilized using Staphylococcus aureus alpha-toxin, and adenosinetriphosphatase (ATPase) was measured as the formation of [3H]ADP, [3H]AMP, and [3H]adenosine from [3H]ATP in the solution bathing the muscle. The resting ATPase (1.96 +/- 0.15 mM/min, n = 13) is approximately 5-10 times higher than that measured in Triton X-100-permeabilized muscles (0.28 +/- 0.01 mM/min, n = 4), with nucleotide accumulating as ADP, AMP, and adenosine. The ATPase activity is also seen when the intact muscle is incubated in a Krebs solution containing 1 mM MgATP (2.76 +/- 0.10 mM/min, n = 73). This suggests that it is due primarily to an ecto-ATPase. The ectoenzyme is capable of hydrolyzing both ATP and ADP, and in both cases there is a higher rate at 3 than at 1 mM nucleotide. The high resting ATPase compromises the control of nucleotide concentrations within the permeabilized tissue even in the presence of an ATP-regenerating system consisting of phosphocreatine (PCr, 35mM) and creatine kinase (1 mg/ml). Treatment of the intact muscle with the ectonucleotidase inhibitor 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) followed by alpha-toxin permeabilization and inclusion of sodium azide in subsequent solutions reduces the ecto-ATPase by approximately 70%. Addition of PCr and creatine kinase then results in the maintenance of high [ATP] and low [ADP] in the muscle, and importantly, there are no significant changes in [ATP], [ADP], [adenosine/AMP], or the ADP-to-ATP ratio upon activation of the muscle in pCa 4.5. In general, the force output in high Ca2+ increased as the metabolic profile of the muscle improved. When ATPase was measured as the appearance of [32P]Pi from [32P]PCr and [gamma-32P]ATP, the alpha-toxin-permeabilized muscle subjected to the above treatment showed only approximately 30% higher total ATPase under activated conditions compared with the freeze-glycerinated Triton-treated portal vein. The suprabasal ATPase is similar in both preparations. We conclude that the reduction of the basal ATPase by the DIDS-azide treatment permits both rigorous control of nucleotide contents and accurate measurement of ATPase activity in alpha-toxin-permeabilized smooth muscle.
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PMID:Metabolic characteristics of alpha-toxin-permeabilized smooth muscle. 802 97

The effect of neuropeptide Y on the number and affinity of catecholamine receptors in the ventricular myocardium was investigated. Receptor binding studies showed that incubation of cardiac membrane in the presence of neuropeptide Y (NPY, 10(-7) M) decreased the number of alpha/beta-adrenoceptor binding sites (Bmax) without affecting the affinity (KD) of these receptors. Although not able to modulate the contractility by itself, NPY was able to decrease the positive inotropic effects of phenylephrine and isoproterenol in the isolated, perfused myocardium. Ca2+/Mg(2+)-ATPase activity, measured from the sarcolemma, sarcoplasmic reticulum and myofibrils, was unaltered whereas the activity of sarcolemmal Na+/K(+)-ATPase was decreased when NPY was included in the media. On the other hand, NPY was shown to increase the phosphoinositide-phospholipase C associated with the sarcolemma. These findings support the hypothesis that NPY modulates postsynaptic adrenergic receptors in the myocardium and can affect the adrenergic-induced, inotropic response.
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PMID:Adrenoreceptor-mediated effect of neuropeptide Y decreases cardiac inotropic responses. 803 15


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