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Sample records for ca2 mg2 atpase

  1. [Kinetics of inhibitory effect of calix[4]arene C-90 on activity of transporting plasma membrane Ca2+, Mg2+-ATPase of smooth muscle cells].

    Science.gov (United States)

    Veklich, T O; Shkrabak, O A; Mazur, Iu Iu; Rodik, R V; Kal'chenko, V I; Kosterin, S O

    2014-01-01

    In experiments on the suspension of myometrium cell plasma membrane, processed by 0.1% digitonin, the inhibitory action of calix[4]arene C-90 (5,11,17,23-tetra(threeftor)methyl(phenilsulphonilimino)-methylamino-25,26,27,28-tetrapropoxy-calix[4]arene) on the activity of Ca2+,Mg2+-ATPase was investigated. The authors also examined the influence of calix[4]arene in different concentration on affinity of enzyme (Ca2,Mg2+-ATPase) for the ATP and ions of Mg and Ca, and its influence on cooperative effect and maximum velocity of ATP hydrolysis. It is shown that calix[4]arene does not influence the affinity of Ca2+,Mg2+-ATPase for the ATP, which means that these two compounds have different binding centers. Also calix[4]arene has no influence on affinity and cooperative effect of Ca ions, if it is used in concentration lower than 50 μM. Calix[4]arene slightly increases coefficient of Ca2+,Mg2+-ATPase activation by magnesium chloride. In all three cases, where ATP, Mg and Ca ions are used to test the impact of calix[4]arene, maximum velocity of ATP hydrolysis significantly decreases. All these results clarify that calix[4]arene implements its inhibitory action through mechanism of uncompetitive inhibition of Ca2+,Mg2+-ATPase activity.

  2. Increased oxidative stress and decreased activities of Ca2+/Mg2+-ATPase and Na+/K+-ATPase in the red blood cells of the hibernating black bear

    Science.gov (United States)

    Chauhan, V.P.S.; Tsiouris, J.A.; Chauhan, A.; Sheikh, A.M.; Brown, W. Ted; Vaughan, M.

    2002-01-01

    During hibernation, animals undergo metabolic changes that result in reduced utilization of glucose and oxygen. Fat is known to be the preferential source of energy for hibernating animals. Malonyldialdehyde (MDA) is an end product of fatty acid oxidation, and is generally used as an index of lipid peroxidation. We report here that peroxidation of lipids is increased in the plasma and in the membranes of red blood cells in black bears during hibernation. The plasma MDA content was about four fold higher during hibernation as compared to that during the active, non-hibernating state (P oxidative stress, and have reduced activities of membrane-bound enzymes such as Ca2+/Mg2+-ATPase and Na+/K+-ATPase. These changes can be considered part of the adaptive for survival process of metabolic depression. ?? 2002 Elsevier Science Inc. All rights reserved.

  3. Ca2+- and Mg2+-ATPase activities in winter wheat root plasma membranes as affected by NaCl stress during growth

    NARCIS (Netherlands)

    Mansour, MMF; van Hasselt, PR; Kuiper, PJC

    1998-01-01

    Winter wheat seedlings were grown in Hoagland nutrient solution with or without 100 mmol/L NaCl added. Plasma membranes from root cells were prepared by aqueous polymer two phase partitioning and the stimulation of plasma membrane ATPase activity by Mg2+ and Ca2+ was investigated. The enzyme was act

  4. MODULATION OF Na + /K + , Mg 2 + and Ca 2+ ATPase ACTIVITY IN DIFFERENT REGIONS OF RAT BRAIN DURING ROTENONE INDUCED PARKINSON'S DISEASE AND PROTECTIVE ROLE OF BACOPA MONNIERI

    Directory of Open Access Journals (Sweden)

    Gunduluru Swathi

    2013-02-01

    Full Text Available Bacopa monnieri(BM; Family: Scrophulariaceae, also referred as Brahmi or Jalbrahmi has been used for centuries in Ayurvedic system of medicine as a brain tonic, memory enhancer, revitaliser of sensory organs, anti-anxiety, cardio-tonic, diuretic, antidepressant and anticonvulsant agent, and the pharmacological actions are mainly attributed to the saponin compounds present in the alcoholic extract of the plant. The present study was carried out with a specific aim to examine the neuroprotective effect of Bacopa monnieriduring Rotenone (RT induced Parkinson’s disease (PD with particular reference to Na+/K+, Mg2+and Ca2+-ATPase activities in different regions of rat brain. In the experiment conducted rats were divided into four groups of six in each group, group 1 received Salinewater (1 ml/kg, group 2 received RT (2.5 mg/kg through i.p. route administration for 60 days to induce PD. The third group received BM extract (180 mg/kg/day for 20 days orally before induction of PD and group 4 received Levodopa (LD (10 mg/kg/day orally which is referred as drug control. The levels of Na+/K+, Mg2+and Ca2+-ATPase activities were measured. Na+/K+, Mg2+and Ca2+-ATPase activities were significantly depleted in different brain regions of rat during RT induced PD when compared to control rats. Treatment with BM and LD caused significant elevation in the activity levels of Na+/K+, Mg2+and Ca2+-ATPase in different brain regions of rats when compared to induced PD rats. Our results suggest the ability of BM extract to modulate Na+/K+, Mg2+and Ca2+- ATPase activities in different brain regions of RT induced rodent model of PD and thus offers effective management in the treatment of PD.

  5. Effect of organic solvents on nervous cell membrane as measured by changes in the (Ca2+/Mg2+) ATPase activity and fluidity of synaptosomal membrane.

    Science.gov (United States)

    Edelfors, S; Ravn-Jonsen, A

    1992-03-01

    The effect of various solvents on the central nervous system was studied by using rat brain synaptosomal membranes as an in vitro model. The activity of (Ca2+/Mg2+) ATPase and the membrane fluidity was determined. The alteration of the ATPase activity depended on the physio-chemical characteristics of the solvent in question. Incubation with aliphatic alkanes caused a stimulation of the ATPase activity whereas mixed hydrocarbons as kerosene, white spirit and gasoline inhibited the enzyme. Incubation with chlorinated hydrocarbons caused a biphasic response dependent on the concentration. Oxygen-containing hydrocarbons exhibited various effects as found after incubation with hydrocarbons. The different effects of the solvents on the ATPase activity suggest that the lipophilicity of the solvents is one of more parameters affecting the membrane. Furthermore, the biphasic response following the incubation with chlorinated hydrocarbons indicates that more mechanisms are involved in the enzyme effect. The membrane fluidity is increased with higher concentrations of the solvents. From the results it is concluded that the ATPase activity depends not only on the membrane fluidity and volume, but also on the hydrophilic vicinity of the enzyme molecule. PMID:1533717

  6. Effects of aqueous extract of Hibiscus sabdariffa on renal Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities in Wistar rats%玫瑰茄提取物对Wistar大鼠肾Na+-K+-ATP酶及Ca2+-Mg2+-ATP酶活性的影响

    Institute of Scientific and Technical Information of China (English)

    Lawrence A.Olatunji; Taofeek O.Usman; Joseph O.Adebayo; Victoria A.Olatunji

    2012-01-01

    OBJECTIVE:To investigate the effects of oral administration of aqueous extract of Hibiscus sabdariffa on renal Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities in rats.METHODS:The 25 and 50 mg/(kg · d) of aqueous extracts of H.sabdariffa were respectively given to rats in the experimental groups for 28 d,and rats in the control group received an appropriate volume of distilled water as vehicle.Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities in the kidney were assayed by spectrophotometric method. RESULTS:Administrations of 25 and 50 mg/(kg · d) of aqueous extract of H.sabdariffa significantly decreased the Ca2+-Mg2+-ATPase activity in the kidney of rats (P <0.05).However,the renal Na+-K+-ATpase activity of the experimental rats was not affected by either dose of the extract.And the plasma Na+,K+ and Ca2+ levels of the experimental rats had no significant changes.Administration of either dose of the extract did not result in any significant changes in body and kidney weights,the concentrations of plasma albumin and total protein,and alkaline phosphatase,aspartate aminotransferase and alanine aminotransferase activities.However,concentrations of creatinine and urea were significantly reduced by 50 mg/kg of the extract (P<0.05). CONCLUSION:The present study indicates that oral administration of aqueous extract of H.sabdariffa may preserve the renal function despite a decreased renal Ca2+-Mg2+-ATPase activity.%目的:研究口服玫瑰茄水提取物对大鼠肾Na+-K+-ATP酶和Ca2+- Mg2+-ATP酶活性的影响.方法:连续28 d分别给予实验大鼠口服25和50 mg/kg的玫瑰茄水提物,同时给予对照组大鼠灌胃适当剂量的蒸馏水.用光谱测定法分析大鼠肾脏中Na+-K+-ATP酶和Ca2+ -Mg2+-ATP酶的活性.结果:口服25和50 mg/kg的玫瑰茄提取物后,实验组大鼠肾Ca2+-Mg2+-ATP酶活性显著降低(P<0.05),然而肾Na+ -K+ -ATP酶的活性却未受到任何影响.实验组大鼠体质量、肾脏质量,血浆白蛋白和总蛋白浓

  7. 电离辐射对大鼠咬肌钙泵活性和表达的短期影响%Effect of radiation on the activity and expression of Ca2+-Mg2+-ATPase in rat masseter muscle

    Institute of Scientific and Technical Information of China (English)

    李志民; 马绪臣; 曲兴民; 徐寿平; 马林

    2009-01-01

    -ATP酶本身功能异常造成的.%ase of ATPase activity played an important role in the cause of radiation-induced skeletal muscle injury, while there was no significant reduction in the expression of Ca2+-Mg2+ -ATPase protein in irradiated rat masseter muscle.

  8. Effects of percutaneous midband pulse current stimulation in hepatic region on the activity of hepatic mitochondrial Na+-K+-ATPase and Ca2+-Mg2+-ATPase in exercise-induced fatigued rats

    OpenAIRE

    Yi-zong ZHAI; Chang-lin HUANG; Chang, Qi; Wang, Jiu-Qing; Zhang, Jia; Guo, Yan-Ling

    2015-01-01

    Objective To explore the effects of percutaneous impulsive current stimulation in hepatic region on the activity of hepatic mitochondrial Na+-K+-ATPase and Ca2+-Mg2+-ATPase in exercise-induced fatigued rats, in order to investigate the effect of exercise-induced fatigue. Methods Seventy-two 8-week old male Wistar rats were randomly divided into 4 groups (18 each): control group (group A), fatigue group (group B), stimulation before fatigue group (group C) and stimulation after fatigue group (...

  9. Effects of anthropogenic sound on digging behavior, metabolism, Ca2+/Mg2+ ATPase activity, and metabolism-related gene expression of the bivalve Sinonovacula constricta

    Science.gov (United States)

    Peng, Chao; Zhao, Xinguo; Liu, Saixi; Shi, Wei; Han, Yu; Guo, Cheng; Jiang, Jingang; Wan, Haibo; Shen, Tiedong; Liu, Guangxu

    2016-04-01

    Anthropogenic sound has increased significantly in the past decade. However, only a few studies to date have investigated its effects on marine bivalves, with little known about the underlying physiological and molecular mechanisms. In the present study, the effects of different types, frequencies, and intensities of anthropogenic sounds on the digging behavior of razor clams (Sinonovacula constricta) were investigated. The results showed that variations in sound intensity induced deeper digging. Furthermore, anthropogenic sound exposure led to an alteration in the O:N ratios and the expression of ten metabolism-related genes from the glycolysis, fatty acid biosynthesis, tryptophan metabolism, and Tricarboxylic Acid Cycle (TCA cycle) pathways. Expression of all genes under investigation was induced upon exposure to anthropogenic sound at ~80 dB re 1 μPa and repressed at ~100 dB re 1 μPa sound. In addition, the activity of Ca2+/Mg2+-ATPase in the feet tissues, which is directly related to muscular contraction and subsequently to digging behavior, was also found to be affected by anthropogenic sound intensity. The findings suggest that sound may be perceived by bivalves as changes in the water particle motion and lead to the subsequent reactions detected in razor clams.

  10. Effects of percutaneous midband pulse current stimulation in hepatic region on the activity of hepatic mitochondrial Na+-K+-ATPase and Ca2+-Mg2+-ATPase in exercise-induced fatigued rats

    Directory of Open Access Journals (Sweden)

    Yi-zong ZHAI

    2015-06-01

    Full Text Available Objective To explore the effects of percutaneous impulsive current stimulation in hepatic region on the activity of hepatic mitochondrial Na+-K+-ATPase and Ca2+-Mg2+-ATPase in exercise-induced fatigued rats, in order to investigate the effect of exercise-induced fatigue. Methods Seventy-two 8-week old male Wistar rats were randomly divided into 4 groups (18 each: control group (group A, fatigue group (group B, stimulation before fatigue group (group C and stimulation after fatigue group (group D. Exhaustion of animals in B, C and D groups were reproduced by prolonged swimming. Current stimulation (1024Hz, 10mA, current cycle 1sec for 20 minutes was given to the rats of group C before swimming, and to those in group D after exhaustion. At the weekend of 1st, 3rd and 5th week after modeling, the rats were sacrificed in batches from each group (6 each. The activities of hepatic mitochondrial Na+-K+-ATPase and Ca2+-Mg2+-ATPase were determined by spectrophotometry, and Bradfood protein quantification was employed to quantitate the protein in rats' hepatic mitochondria. Results No significant difference was found in swimming-exhaustion time among 3 groups at the first weekend (P>0.05, while the swimming-exhaustion time was significantly prolonged at the 3rd and 5th weekends in group D than in group B and C (P0.05, while the enzyme activities were obviously lower at the 3rd and 5th weekend in group B than that in groups A, C and D (P<0.05, and they were also lower in group C than that in group D (P<0.05. Conclusions Exercise-induced fatigue can lower the activity of hepatic mitochondrial Na+-K+-ATPase and Ca2+-Mg2+-ATPase. Percutaneous pulsive current stimulating hepatic region of exercise-induced fatigued rats may improve the enzyme activity, reduce the concentration of free calcium and calcium overload in mitochondria, stimulate the oxidative phosphorylation, accelerate the rate of respiratory chain, promote exercise endurance and score, and

  11. Hereditary tubular transport disorders: implications for renal handling of Ca2+ and Mg2+

    DEFF Research Database (Denmark)

    Dimke, Henrik; Hoenderop, Joost G; Bindels, René J;

    2010-01-01

    change tubular transport of Ca2+ and Mg2+. In the distal convolutions, several proteins involved in Mg2+ transport have been identified [TRPM6 (transient receptor potential melastatin 6), proEGF (pro-epidermal growth factor) and FXYD2 (Na+/K+-ATPase gamma-subunit)]. In addition, conditions...

  12. 甘草黄酮对大强度耐力运动大鼠肾脏组织Ca2+、Mg2+-ATPase及Bax,Bc1-2蛋白表达的影响%Effect of High-Intensity Endurance Exercise on Ca2+,Mg2+-ATPase and Bax, Bcl-2 Protein Expression With Glycyrrhiza Flavonoids in rat Nephridial Tissue

    Institute of Scientific and Technical Information of China (English)

    王东旭; 陈艳艳

    2013-01-01

    Objective To explore Glycyrrhiza Elavonoids on the rat nephridial tissue of Ca2+, Mg2+-ATPase and Bax, Bcl-2 protein expression with high-intensity endurance exercise. Methods The twenty-four healthy male rats were randomly divided into quiet groups, high-intensity exercise group and exercise plus Glycyrrhiza Elavonoids group, After 6 weeks of treadmill training, Using the box of reagent and immunity histochemistry examined the changing of Ca2+, Mg2+-ATPase and Bax, Bcl-2 protein expression on each groups . Results Compared with the quiet groups, the activity of Ca2+, Mg2+-ATPase both had significant droped (P<0.01), and the groups of plus drog had very difference increased than high-intendity exerxise groups (P<0.01); High-intensity endurance exercise group and exercise dosing rats AI apoptosis index increased in varying degrees;high-intensity exercise group (MOD) were very significant difference(P<0.01), exercise plus drug group Bac protein expression (MOD)were very significant difference (P<0.01); Exercise plus drug group Bcl-2 protein expression(MOD) with the high-intersity exercise group had significant difference(P<0.01), High-intensity exercise group and exercise plus drug group Bax/Bcl-2 ratio of distribution is significantly difference degrees of difference(P<0.05,P<0.01).%目的:探讨甘草黄酮对大强度耐力运动大鼠肾脏组织Ca2+、Mg2+-ATPase及Bax、Bcl-2表达的影响。方法:选取SD雄性健康大鼠24只,随机分为安静组、大强度运动组和运动加药组;采用跑台训练6周后取材,应用试剂盒和免疫组织化学法测检测各组大鼠肾脏组织Ca2+、Mg2+-TPase活性及Bax和Bcl-2表达的变化。结果:与安静对照组相比,大强度运动组和运动加药组肾脏组织Ca2+、Mg2+-TPase活性均呈非常显著性下降(P<0.01);其中运动加药组Ca2+、Mg2+-TPase活性均较大强度运动组具有非常显著差异性提高(P<0.01);大强度耐力运动组和运动加

  13. Crystal Structure of the Vanadate-Inhibited Ca(2+)-ATPase.

    Science.gov (United States)

    Clausen, Johannes D; Bublitz, Maike; Arnou, Bertrand; Olesen, Claus; Andersen, Jens Peter; Møller, Jesper Vuust; Nissen, Poul

    2016-04-01

    Vanadate is the hallmark inhibitor of the P-type ATPase family; however, structural details of its inhibitory mechanism have remained unresolved. We have determined the crystal structure of sarcoplasmic reticulum Ca(2+)-ATPase with bound vanadate in the absence of Ca(2+). Vanadate is bound at the catalytic site as a planar VO3(-) in complex with water and Mg(2+) in a dephosphorylation transition-state-like conformation. Validating bound VO3(-) by anomalous difference Fourier maps using long-wavelength data we also identify a hitherto undescribed Cl(-) site near the dephosphorylation site. Crystallization was facilitated by trinitrophenyl (TNP)-derivatized nucleotides that bind with the TNP moiety occupying the binding pocket that normally accommodates the adenine of ATP, rationalizing their remarkably high affinity for E2P-like conformations of the Ca(2+)-ATPase. A comparison of the configurations of bound nucleotide analogs in the E2·VO3(-) structure with that in E2·BeF3(-) (E2P ground state analog) reveals multiple binding modes to the Ca(2+)-ATPase.

  14. Relations between erythrocyte Ca2+ , Mg2+ levels and cell membrane ATPase activities in patients with hypertension in obese children%肥胖儿童红细胞钙镁水平及ATP酶活性与高血压关系的探讨

    Institute of Scientific and Technical Information of China (English)

    朱树森; 符云峰; 卢振敏; 王素敏; 孙爱丽

    2001-01-01

    目的探讨细胞离子代谢紊乱在肥胖儿童高血压发病中的作用。方法测定125例(正常对照37例,正常血压肥胖34例,正常体重高血压21例,高血压肥胖33例)12~16岁中学生的红细胞膜ATP酶活性、血浆和红细胞胞浆Ca2+、Mg2+水平。结果正常血压肥胖组(ONT)儿童红细胞膜Na+-K+-ATP酶和Ca2+-ATP酶活性较正常对照组(NT)显著降低,正常体重高血压组(NOHT)和高血压肥胖组(OHT)两酶活性又显著低于ONT组。ONT组、NOHT组和OHT组红细胞胞浆Ca2+水平三组之间无显著差异,但均显著高于NT组。红细胞胞浆Mg2+,血浆Ca2+、Mg2+在NT组和ONT组之间无显著差异,在NOHT组和OHT组均显著降低。结论红细胞膜Na+-K+-ATP酶和Ca2+-ATP酶活性降低可能在肥胖儿童高血压的发病机制中有重要作用。%Objective To explore the roles of the metabolic disorders of cellular ions in pathogenesis of hypertension in obese children. Methods Erythrocyte membrane ATPase activities,plasma and cellular Ca2 + , Mg2 + levels were measured in 125(37 non-obese normotensives,34 obese normotensives,21 hypertensives and 33 obese hypertensives)middle school students aged 12~16 years. Results Erythrocyte membrane Na+ -K+-ATPase and Ca2+ -ATPase activities were significantly lower in obese children with normotension than those in non-obese normotensive children,and they were also significantly lower in two groups of hypertensive children than those in obese children with normotension. Erythrocyte Ca2+ in normotensive obese and two group of hypertensive children was significantly higher than that in non-obese normotensive children, but there were no significant differences between the three groups. There were no significant differences found in erythrocyte Mg2+ , plasma Ca2+ and Mg2+ between normotensive obese and normotensive non-obese children, but significant decreases were found in both groups of hypertensive children. Conclusion Decreased Na +-K+-ATPase

  15. The Influences of Mg2+ , Ca2+ and Mg2+/Ca2+ Ratio in Mixed Seawater on the Emergence Rate of Penaeus japonicus Postlarva

    Institute of Scientific and Technical Information of China (English)

    臧维玲; 戴习林; 江敏; 姚庆祯; 蔡云龙; 罗春芳; 徐桂荣; 丁福江

    2003-01-01

    This paper reports the approprite ranges of Mg2+ , Ca2 + and their ratio Mg2 +/Ca2 + inmixed seawater for rearing of Penaeus japonicus larvae. The ranges for the above three indices are1150- 1450 mg/L, 360- 440 mg/L and 2.8 - 3.4, respectively. The proper sahnity range ofmixed seawater is 22.1 - 33.8 obtained by mixing estuarine water and concentrated seawater.

  16. Transmembrane Ca2+ gradient-mediated phosphatidylcholine modulating sarcoplasmic reticulum Ca2+-ATPase

    Institute of Scientific and Technical Information of China (English)

    屠亚平; 徐红; 杨福愉

    1995-01-01

    The sarcoplasmic reticulum (SR) Ca2+-ATPase was purified and reconstituted into the sealed phospholipids vesicles with or without transmembrane Ca2+ gradient. The role ofphospholipids, especially phosphatidylcholine(PC), in the modulation of Ca2+-ATPase by transmembrane Ca2+ gradient was investigated. The results are as follows, (i) Incubated with phospholiplds, the enzyme activity of the delipidated Ca2+-ATPase is inhibited by Ca2+ and the highest inhibition is observed in the presence of PC. (ii) When there exists a transmembrane Ca2+ gradient (higher Ca2+ concentration inside vesicles, 1 000μmol/L:50μmol/L, similar to the physiological condition), the inhibition of Ca2+-ATPase by transmembrane Ca2+ gradient can be only observed in the vesicles containing PC:PE, but not in those containing PS:PE or PG:PE. The highest inhibition is obtained at a 50.50 molar ratio of PC:PE. (iii) By comparing the effects of PC differing in acyl chains, higher inhibition of Ca2+-ATPase is observed in vesicles containin

  17. Acid-base status determines the renal expression of Ca2+ and Mg2+ transport proteins.

    NARCIS (Netherlands)

    Nijenhuis, T.; Renkema, K.Y.R.; Hoenderop, J.G.J.; Bindels, R.J.M.

    2006-01-01

    Chronic metabolic acidosis results in renal Ca2+ and Mg2+ wasting, whereas chronic metabolic alkalosis is known to exert the reverse effects. It was hypothesized that these adaptations are mediated at least in part by the renal Ca2+ and Mg2+ transport proteins. The aim of this study, therefore, was

  18. Effect of Ca2+ and Mg2+ on CO2 Corrosion Behavior of Tube Steel

    Institute of Scientific and Technical Information of China (English)

    ZHAO Guo-xian; LI Jian-ping; HAO Shi-ming; L(U) Xiang-hong; LI He-lin

    2005-01-01

    Effects of Ca2+ and Mg2+ on the CO2 corrosion behaviors of tube steel were studied in simulated oil-fieldenvironment. The influence of Ca2+ and Mg2+ on the corrosion rate and morphologies of corrosion product layerwas determined by scanning electron microscope and measuring mass loss. Potentiodynamic polarization and im-pedance spectroscopy were used to investigate the change of electrochemical characteristic parameters of corrosionproduct layer and corrosion dynamic process. The results show that with Ca2+ and Mg2+ in electrolyte, the mor-phologies and microstructures of corrosion product layer changed obviously, thus affecting the corrosion process.

  19. Ganglioside GM3 modulates conformation of reconstituted Ca2+ -ATPase

    Institute of Scientific and Technical Information of China (English)

    王丽华; 杨小毅; 屠亚平; 催肇春; 杨福愉

    1997-01-01

    Using steady-state fluorescence and nanosecond time-resolved fluorescence techniques, the Ca 2+-ATPase conformational changes induced by ganglioside GM3 were studied with different quenchers. The results showed that GM3 could significantly increase the lifetime of intrinsic fluorescence of Ca2 + -ATPase reconstituted into proteoliposomes, and could also weaken the intrinsic fluorescence quenching by KI or hypocrellin B, HB. Further-more, by using quenching kinetic analysis of the time-resolved fluorescence, in the presence of GM3, the quenching constant (Ksv) and quenching efficiency were significantly lowered. The obtained results suggest that the oligosaccha-ride chain and the ceramide moieties of the GM3 molecule could interact with its counterparts of the Ca2+ -ATPase re-spectively, thus change the conformation of the hydrophobic domain of the enzyme, making the tryptophan residues in different regions shift towards the hydrophilic-hydrophobic interface, and hence shorten the distance between the hy

  20. Interaction of phosphatidic acid and phosphatidylserine with the Ca2+-ATPase of sarcoplasmic reticulum and the mechanism of inhibition.

    Science.gov (United States)

    Dalton, K A; East, J M; Mall, S; Oliver, S; Starling, A P; Lee, A G

    1998-02-01

    The sarcoplasmic reticulum of skeletal muscle contains anionic phospholipids as well as the zwitterionic phosphatidylcholine and phosphatidylethanolamine. Here we study the effects of anionic phospholipids on the activity of the Ca2+-ATPase purified from the membrane. Reconstitution of the Ca2+-ATPase into dioleoylphosphatidylserine [di(C18:1)PS] or dioleoylphosphatidic acid [di(C18:1)PA] leads to a decrease in ATPase activity. Measurements of the quenching of the tryptophan fluorescence of the ATPase by brominated phospholipids give a relative binding constant for the anionic lipids compared with dioleoylphosphatidylcholine close to 1 and suggest that phosphatidic acid only binds to the ATPase at the bulk lipid sites around the ATPase. Addition of di(C18:1)PS or di(C18:1)PA to the ATPase in the short-chain dimyristoleoylphosphatidylcholine [di(C14:1)PC] reverse the effects of the short-chain lipid on ATPase activity and on Ca2+ binding, as revealed by the response of tryptophan fluorescence intensity to Ca2+ binding. It is concluded that the lipid headgroup and lipid fatty acyl chains have separate effects on the function of the ATPase. The anionic phospholipids have no significant effect on Ca2+ binding to the ATPase; the level of Ca2+ binding to the ATPase, the affinity of binding and the rate of dissociation of Ca2+ are unchanged by reconstitution into di(C18:1)PA. The major effect of the anionic lipids is a reduction in the maximal level of binding of MgATP. This is attributed to the formation of oligomers of the Ca2+-ATPase, in which only one molecule of the ATPase can bind MgATP dimers in di(C18:1)PS and trimers or tetramers in di(C18:1)PA. The rates of phosphorylation and dephosphorylation for the proportion of the ATPase still able to bind ATP are unaffected by reconstitution. Larger changes were observed in the level of phosphorylation of the ATPase by Pi, which became very low in the anionic phospholipids. The fluorescence response to Mg2+ for the ATPase

  1. Ca2+ and Mg2+ binding induce conformational stability of Calfumirin-1 from Dictyostelium discoideum

    Indian Academy of Sciences (India)

    Bairagi C Mallick; Sa-Ouk Kang; Suman Jha

    2014-05-01

    The apo-Calfumirin-1 (CAF-1) binds to Ca2+ with high affinity and also to Mg2+ with high positive cooperativity. The thermal unfolding curves of wtCAF-1 monitored at neutral pH by CD spectroscopy are reversible and show different thermal stabilities in the absence or presence of Ca2+ and Mg2+ ions. Metalfree wtCAF-1 shows greater thermal stability than EF-IV mutant protein. We observed that GdnHCl-induced unfolding of apo-wtCAF-1 monitored by CD and fluorescence spectroscopies increases co-operative folding with approximately same C values. Binding of Ca2+ and Mg2+ ions to CAF-1 dramatically altered the fluorescence and CD spectra, indicating metal ion-induced conformational changes both in the wild-type and mutant proteins. The hydrophobic probe, ANS is used to observe alteration in surface hydrophobicity of the protein in different ligation states. In apo-wtCAF-1, the exposed hydrophobic surfaces are able to bind ANS which is in contrast to the unfolded or the metal ions ligated conformations. Isothermal titration calorimetry (ITC) resultsshow two possible independent binding sites of comparable affinity for the metal ions. However, their binding to the EF-IV E helix-loop-F helix mutant apo-protein happens with different affinities. The present study demonstrates that Ca2+ or Mg2+ binding plays a possible role in the conformational stability of the protein.

  2. Chelate titrations of Ca(2+) and Mg(2+) using microfluidic paper-based analytical devices.

    Science.gov (United States)

    Karita, Shingo; Kaneta, Takashi

    2016-06-14

    We developed microfluidic paper-based analytical devices (μPADs) for the chelate titrations of Ca(2+) and Mg(2+) in natural water. The μPAD consisted of ten reaction zones and ten detection zones connected through narrow channels to a sample zone located at the center. Buffer solutions with a pH of 10 or 13 were applied to all surfaces of the channels and zones. Different amounts of ethylenediaminetetraacetic acid (EDTA) were added to the reaction zones and a consistent amount of a metal indicator (Eriochrome Black T or Calcon) was added to the detection zones. The total concentrations of Ca(2+) and Mg(2+) (total hardness) in the water were measured using a μPAD containing a buffer solution with a pH of 10, whereas only Ca(2+) was titrated using a μPAD prepared with a potassium hydroxide solution with a pH of 13. The μPADs permitted the determination of Ca(2+) and Mg(2+) in mineral water, river water, and seawater samples within only a few minutes using only the naked eye-no need of instruments.

  3. Chelate titrations of Ca(2+) and Mg(2+) using microfluidic paper-based analytical devices.

    Science.gov (United States)

    Karita, Shingo; Kaneta, Takashi

    2016-06-14

    We developed microfluidic paper-based analytical devices (μPADs) for the chelate titrations of Ca(2+) and Mg(2+) in natural water. The μPAD consisted of ten reaction zones and ten detection zones connected through narrow channels to a sample zone located at the center. Buffer solutions with a pH of 10 or 13 were applied to all surfaces of the channels and zones. Different amounts of ethylenediaminetetraacetic acid (EDTA) were added to the reaction zones and a consistent amount of a metal indicator (Eriochrome Black T or Calcon) was added to the detection zones. The total concentrations of Ca(2+) and Mg(2+) (total hardness) in the water were measured using a μPAD containing a buffer solution with a pH of 10, whereas only Ca(2+) was titrated using a μPAD prepared with a potassium hydroxide solution with a pH of 13. The μPADs permitted the determination of Ca(2+) and Mg(2+) in mineral water, river water, and seawater samples within only a few minutes using only the naked eye-no need of instruments. PMID:27181645

  4. Comparison of Ca2+ and Mg2+ enhancing aerobic granulation in SBR

    International Nuclear Information System (INIS)

    Two sequencing batch reactors (SBRs) were operated to investigate the effect of Ca2+ and Mg2+ augmentation on aerobic granulation. Reactor R1 was augmented with Ca2+ at 40 mg/L, while Mg2+ was added to the reactor R2 with 40 mg/L. Results showed that the reactor R1 had a faster granulation process compared with R2, and the mature granules in R1 showed better physical characteristics. However, the mature granules in R2 had the higher production yield of polysaccharides and proteins, and aerobic granules in R2 experienced a faster substrate biodegradation. Microbial and genetic characteristics in mature granules were analyzed using polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE) techniques. The results revealed that Mg2+ addition led to higher microbial diversity in mature granules. In addition, an uncultured bacterium (AB447697) was major specie in R1, and β-proteobacterium was dominant in R2. It can be concluded that Ca2+ had an important effect on physical properties of aerobic granules, while Mg2+ played a key role on biological properties during the sludge granulation.

  5. Molecular basis of epithelial Ca2+ and Mg2+ transport: insights from the TRP channel family

    DEFF Research Database (Denmark)

    Dimke, Henrik Anthony; Hoenderop, Joost G J; Bindels, René J M

    2011-01-01

    active transcellular movement of divalent cations from the lumen into the enterocyte. Furthermore, in bone, TRPV channels play important roles by influencing the osteoclastic resorption process, thereby contributing importantly to overall bone mineral content. The divalent cation-permeable TRPV5 and TRPM......Maintenance of plasma Ca(2+) and Mg(2+) levels is of vital importance for many physiological functions. This is achieved via a coordinated interplay between the intestine, bone and kidney by amending the rate of absorption, storage and excretion, respectively. Discovery of the transient receptor...... potential (TRP) family identified several new ion channels acting as gatekeepers of Ca(2+) and Mg(2+) transport in these epithelia, greatly increasing our understanding of the molecular processes that facilitate the movement of these minerals. In the intestine, TRP channels contribute to the saturable...

  6. Roles of transmembrane segment M1 of Na(+),K (+)-ATPase and Ca (2+)-ATPase, the gatekeeper and the pivot

    DEFF Research Database (Denmark)

    Einholm, Anja P.; Andersen, Jens Peter; Vilsen, Bente

    2007-01-01

    In this review we summarize mutagenesis work on the structure-function relationship of transmembrane segment M1 in the Na(+),K(+)-ATPase and the sarco(endo)plasmic reticulum Ca(2+)-ATPase. The original hypothesis that charged residues in the N-terminal part of M1 interact with the transported...... cations can be rejected. On the other hand hydrophobic residues in the middle part of M1 turned out to play crucial roles in Ca(2+) interaction/occlusion in Ca(2+)-ATPase and K(+) interaction/occlusion in Na(+),K(+)-ATPase. Leu(65) of the Ca(2+)-ATPase and Leu(99) of the Na(+),K(+)-ATPase, located...... of the extracytoplasmic gate in both the Ca(2+)-ATPase and the Na(+),K(+)-ATPase. Udgivelsesdato: 2007-Dec...

  7. Structural studies of Ca2+-ATPase ligand and regulatory complexes

    DEFF Research Database (Denmark)

    Drachmann, Nikolaj Düring

    2015-01-01

    against their concentration gradient upon ATP hydrolysis. The ion gradients are used to drive several key cellular processes, like the action potential in nerve tissue, acidification of the gastric juice, cell signalling and muscle contraction. The Ca2+-ATPase is an important part of mammalian cells...... choline lipids with different aliphatic chain length and saturation show three specific lipid binding sites. The four different lipids analysed bind to the same binding sites with varying degrees of disorder. The study contributes to understanding the complex interplay between the surrounding membrane......-of-concept Ca2+ bound crystal form, indicated that the information content of SFX data is higher than synchrotron data, and ligands and ions can be detected with low redundant data. The data of the E2 stabilised form was processed to 5 Å resolution, and it was possible to extract useful anomalous data showing...

  8. Interactions of Na+, K+, Mg2+, and Ca 2+ with benzene self-assembled monolayers

    DEFF Research Database (Denmark)

    Pedersen, Morten Rimmen; Matthiesen, Jesper; Bovet, Nicolas Emile;

    2014-01-01

    that are most common in the natural world, namely, Na+, K+, Mg 2+, and Ca2+. Specifically, we investigated how these ions affect the interactions between surfaces covered by self-Assembled monolayers (SAMs) terminated with benzene molecules. We used a flat oxidized silicon substrate and an atomic force...... from X-ray photoelectron spectroscopy (XPS) allowed us to conclude that K+ binds in the benzene layers, creating a positive surface charge on the benzene-covered surfaces, thus leading to lower adhesion in KCl solutions than in pure water. Evidence suggested that Ca2+ does not bind to the surfaces...... measurements. The results of our studies clearly show that even a nonpolar, hydrophobic molecule, such as benzene, has a role to play in the behavior of aqueous solutions and that it interacts differently depending on which ions are present. Even ions from the same column in the periodic table behave...

  9. Mg(2+) differentially regulates two modes of mitochondrial Ca(2+) uptake in isolated cardiac mitochondria: implications for mitochondrial Ca(2+) sequestration.

    Science.gov (United States)

    Blomeyer, Christoph A; Bazil, Jason N; Stowe, David F; Dash, Ranjan K; Camara, Amadou K S

    2016-06-01

    The manner in which mitochondria take up and store Ca(2+) remains highly debated. Recent experimental and computational evidence has suggested the presence of at least two modes of Ca(2+) uptake and a complex Ca(2+) sequestration mechanism in mitochondria. But how Mg(2+) regulates these different modes of Ca(2+) uptake as well as mitochondrial Ca(2+) sequestration is not known. In this study, we investigated two different ways by which mitochondria take up and sequester Ca(2+) by using two different protocols. Isolated guinea pig cardiac mitochondria were exposed to varying concentrations of CaCl2 in the presence or absence of MgCl2. In the first protocol, A, CaCl2 was added to the respiration buffer containing isolated mitochondria, whereas in the second protocol, B, mitochondria were added to the respiration buffer with CaCl2 already present. Protocol A resulted first in a fast transitory uptake followed by a slow gradual uptake. In contrast, protocol B only revealed a slow and gradual Ca(2+) uptake, which was approximately 40 % of the slow uptake rate observed in protocol A. These two types of Ca(2+) uptake modes were differentially modulated by extra-matrix Mg(2+). That is, Mg(2+) markedly inhibited the slow mode of Ca(2+) uptake in both protocols in a concentration-dependent manner, but not the fast mode of uptake exhibited in protocol A. Mg(2+) also inhibited Na(+)-dependent Ca(2+) extrusion. The general Ca(2+) binding properties of the mitochondrial Ca(2+) sequestration system were reaffirmed and shown to be independent of the mode of Ca(2+) uptake, i.e. through the fast or slow mode of uptake. In addition, extra-matrix Mg(2+) hindered Ca(2+) sequestration. Our results indicate that mitochondria exhibit different modes of Ca(2+) uptake depending on the nature of exposure to extra-matrix Ca(2+), which are differentially sensitive to Mg(2+). The implications of these findings in cardiomyocytes are discussed.

  10. Depressing effect of phenoxyl acetic acids on flotation of minerals containing Ca2+/Mg2+ gangues

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Phenoxyl acetic acids were applied to determine their depressing effect on minerals containing Ca2+/Mg2+ gangues. Calcite,mixture of calcite and fluorite, and nickel ore were used in the flotation. And the depression mechanism was studied by the determination of contact angle, zeta potential, adsorptive capacity of collector, and IR analysis as well. It is found that 0.1 mmol/L of phenoxyl acetic acid derived from pyrogallol or gallic acid exhibits strong depressing ability on calcite in almost zero yields at pH value of 9.8, and calcite can be depressed in the flotation of calcite/fluorite mixture for approximate 87% yield of fluorite. The flotation result of practical nickel ore containing serpentine indicates that these two depressants may also show better depression performance to serpentine than traditional depressants such as sodium fluosilicate and carboxylmethyl cellulose. Analysis for the depression mechanism reveals that there exists strong chemical interaction between the depressants and minerals.

  11. Isotopic fractionation of Mg 2+(aq), Ca 2+(aq), and Fe 2+(aq) with carbonate minerals

    Science.gov (United States)

    Rustad, James R.; Casey, William H.; Yin, Qing-Zhu; Bylaska, Eric J.; Felmy, Andrew R.; Bogatko, Stuart A.; Jackson, Virgil E.; Dixon, David A.

    2010-11-01

    Density-functional electronic structure calculations are used to compute the equilibrium constants for 26Mg/ 24Mg and 44Ca/ 40Ca isotope exchange between carbonate minerals and uncomplexed divalent aquo ions. The most reliable calculations at the B3LYP/6-311++G(2d,2p) level predict equilibrium constants K, reported as 10 3ln ( K) at 25 °C, of -5.3, -1.1, and +1.2 for 26Mg/ 24Mg exchange between calcite (CaCO 3), magnesite (MgCO 3), and dolomite (Ca 0.5Mg 0.5CO 3), respectively, and Mg 2+(aq), with positive values indicating enrichment of the heavy isotope in the mineral phase. For 44Ca/ 40Ca exchange between calcite and Ca 2+(aq) at 25 °C, the calculations predict values of +1.5 for Ca 2+(aq) in 6-fold coordination and +4.1 for Ca 2+(aq) in 7-fold coordination. We find that the reduced partition function ratios can be reliably computed from systems as small as M(CO)610- and M(HO)62+ embedded in a set of fixed atoms representing the second-shell (and greater) coordination environment. We find that the aqueous cluster representing the aquo ion is much more sensitive to improvements in the basis set than the calculations on the mineral systems, and that fractionation factors should be computed using the best possible basis set for the aquo complex, even if the reduced partition function ratio calculated with the same basis set is not available for the mineral system. The new calculations show that the previous discrepancies between theory and experiment for Fe 3+-hematite and Fe 2+-siderite fractionations arise from an insufficiently accurate reduced partition function ratio for the Fe 3+(aq) and Fe 2+(aq) species.

  12. CO2和O3体积分数升高对银杏希尔反应活力和叶绿体ATP酶活性的影响%Effects of elevated atmospheric CO2 and O3 on Hill activity and Ca2+/Mg2+-ATPase activity of Ginkgo biloba L

    Institute of Scientific and Technical Information of China (English)

    郭丹; 赵天宏; 张兆伟; 王美玉; 付士磊; 何兴元

    2007-01-01

    近年来,随着温室气体体积分数不断上升,研究CO2和O3体积分数升高对植物的影响已取得一定进展,但二者对植物的复合作用及生理研究不够深入.文章利用开顶式气室研究了大气CO2和O3体积分数升高对银杏(Ginkgo biloba L.)光合特性的影响.结果表明,在整个生长季内,与对照相比,在大气CO2体积分数为700×10-6条件下,银杏叶片净光合速率显著增加(P《0.05),希尔反应活力增大,Ca2+/Mg2+-ATPase活性增强,光合产物可溶性糖和淀粉含量增多;而在O3体积分数为80×10-9的情况下,银杏叶片净光合速率下降,希尔反应活力减小,Ca2+/Mg2+-ATPase活性减弱,光合产物可溶性糖和淀粉含量减少;在CO2和O3 复合作用(700×10-6+80×10-9)条件下,银杏叶片净光合速率、希尔反应活力、可溶性糖和淀粉均有所增加,且淀粉含量增加极显著(P《0.01),而Ca2+-ATPase活性先增强后减弱,Mg2+-ATPase活性先减弱后增强.说明CO2可缓解O3对银杏的负效应,而O3亦对CO2的正效应有削弱作用.

  13. Effects of Ca2+ and Mg2+ on the Enzymatic Properties of Cardiac Muscle Myosin%Ca2+、Mg2+对心肌肌球蛋白ATP酶活性的影响

    Institute of Scientific and Technical Information of China (English)

    朱彬; 万朝敏; 刘荣韬; 孙爱民; 黄勋; 王正荣

    2002-01-01

    目的研究Ca2+、Mg2+对心肌肌球蛋白ATP酶活性的影响.方法采用一种简便、快速的方法从心肌组织中提取肌球蛋白,根据酶反应ATP分解释放的无机磷含量检测Ca2+、Mg2+激活肌球蛋白ATP酶的Km值及最大反应速度Vmax,以及不同浓度的Ca2+、Mg2+、pH值对肌球蛋白ATP酶活性的影响.结果 Ca2+、Mg2+-肌球蛋白ATP酶Km值为5.27±2.10 mmol、7.04±2.06 mmol,Vmax分别为:1.10±0.13 μmol·mg-1·min-1、0.617±0.09 μmol·mg-1·min-1;Ca2+较Mg2+具有较大的酶激活能力(P<0.01),但Mg2+的浓度影响着肌球蛋白ATP酶对Ca2+的敏感性,当Mg2+浓度大于6 mmol/L时,酶对Ca2+的存在无反应;不同的pH值,对Mg2+激活的酶反应影响较小.结论 Ca2+、Mg2+对肌球蛋白ATP酶的作用机理不同,Mg2+是维系肌球蛋白具有酶活性构象所必需,而Ca2+在肌肉收缩过程中可能主要充当的是信号传导功能及调控功能.

  14. Temperature and Ca2+-dependence of the sarcoplasmic reticulum Ca2(+)-ATPase in haddock, salmon, rainbow trout and zebra cichlid

    DEFF Research Database (Denmark)

    Godiksen, Helene; Jessen, Flemming

    2002-01-01

    Temperature dependence of Ca2+-ATPase from the sarcoplasmic reticulum (SR) in rabbit muscle has been widely studied, and it is generally accepted that a break point in Arrhenius plot exist at approximately 20 degreesC. Whether the break point arises as a result of temperature dependent changes...... in the enzyme or its membrane lipid environment is still a matter of discussion. In this study we compared the temperature dependence and Ca2+-dependence of SR Ca2+-ATPase in haddock (Melanogrammus aeglefinus), salmon (Salmo, salar), rainbow trout (Oncorhynchus mykiss) and zebra cichlid (Cichlasoma...... nigrofasciatum). The Arrhenius plot of zebra cichlid showed a break point at 20 degreesC, and the haddock Arrhenius plot was non-linear with pronounced changes in slope in the. temperature area, 6-14 degreesC. In Arrhenius plot from both salmon and rainbow trout a plateau exists with an almost constant SR Ca2...

  15. The interplay between plasma membrane and endoplasmic reticulum Ca(2+)ATPases in agonist-induced temporal Ca(2+) dynamics.

    Science.gov (United States)

    Cicek, Figen Amber; Ozgur, Ekin Ozge; Ozgur, Erol; Ugur, Mehmet

    2014-12-01

    A change in the intracellular free Ca(2+) concentration ([Ca(2+)]i) functions as a transmitter for signal transduction and shows a broad temporal pattern. Even genetically homogeneous cell types show different Ca(2+) response patterns under permanent agonist stimulation. In Ca(2+) signaling, the dynamics of the Ca(2+) release from the Ca(2+) channels during continuous agonist stimulation and the simultaneous effect of the pumps are unclear. In this study, the dynamic interaction of the Ca(2+) ATPases in the plasma membrane (PMCA) and the endoplasmic reticulum membrane (SERCA) during continuous ACh stimulation is monitored using Fluo-3 and Fura-2 loaded HEK 293 cells. We characterize Ca(2+) release patterns at the sub-maximal and maximal stimulation doses in the absence of extracellular Ca(2+). We analyze the responses regarding their types, oscillation frequency and response times. La(3+) (PMCA blocker) do not change the frequency and time courses in sub-maximal ACh treatment, while with the maximal stimulation oscillation frequency increase as oscillations superimpose on robust release, and response time of [Ca(2+)]i is elongated. A similar effect of La(3+) is observed in quantal Ca(2+) release phenomenon. In the presence of CPA, a SERCA blocker, oscillations are completely abolished, but response time does not change. We also observe that during continuous receptor stimulation, Ca(2+) release do not cease. These data may suggest that Ca(2+) release continues during agonist stimulation, but SERCA and PMCA form a new steady state and return [Ca(2+)]i to its physiological concentration. PMID:25331516

  16. Detailed characterization of the cooperative mechanism of Ca(2+) binding and catalytic activation in the Ca(2+) transport (SERCA) ATPase.

    Science.gov (United States)

    Zhang, Z; Lewis, D; Strock, C; Inesi, G; Nakasako, M; Nomura, H; Toyoshima, C

    2000-08-01

    Expression of heterologous SERCA1a ATPase in Cos-1 cells was optimized to yield levels that account for 10-15% of the microsomal protein, as revealed by protein staining on electrophoretic gels. This high level of expression significantly improved our characterization of mutants, including direct measurements of Ca(2+) binding by the ATPase in the absence of ATP, and measurements of various enzyme functions in the presence of ATP or P(i). Mutational analysis distinguished two groups of amino acids within the transmembrane domain: The first group includes Glu771 (M5), Thr799 (M6), Asp800 (M6), and Glu908 (M8), whose individual mutations totally inhibit binding of the two Ca(2+) required for activation of one ATPase molecule. The second group includes Glu309 (M4) and Asn796 (M6), whose individual or combined mutations inhibit binding of only one and the same Ca(2+). The effects of mutations of these amino acids were interpreted in the light of recent information on the ATPase high-resolution structure, explaining the mechanism of Ca(2+) binding and catalytic activation in terms of two cooperative sites. The Glu771, Thr799, and Asp800 side chains contribute prominently to site 1, together with less prominent contributions by Asn768 and Glu908. The Glu309, Asn796, and Asp800 side chains, as well as the Ala305 (and possibly Val304 and Ile307) carbonyl oxygen, contribute to site 2. Sequential binding begins with Ca(2+) occupancy of site 1, followed by transition to a conformation (E') sensitive to Ca(2+) inhibition of enzyme phosphorylation by P(i), but still unable to utilize ATP. The E' conformation accepts the second Ca(2+) on site 2, producing then a conformation (E' ') which is able to utilize ATP. Mutations of residues (Asp813 and Asp818) in the M6/M7 loop reduce Ca(2+) affinity and catalytic turnover, suggesting a strong influence of this loop on the correct positioning of the M6 helix. Mutation of Asp351 (at the catalytic site within the cytosolic domain

  17. Action of erythropoietin in vitro on rabbit reticulocyte membrane Ca2+-ATPase activity.

    OpenAIRE

    Lawrence, W D; Davis, P J; Blas, S D

    1987-01-01

    The mechanism of action of erythropoietin is thought to require specific interaction with the target cell surface and involve alteration of cellular calcium metabolism. Using the rabbit reticulocyte membrane as a model of the immature red cell membrane, we investigated the effects of human recombinant erythropoietin on membrane Ca2+-ATPase (calcium pump) activity in vitro. Erythropoietin in a concentration range of 0.025 to 3.0 U/ml progressively decreased membrane Ca2+-ATPase activity by up ...

  18. Mg(2+)/Ca(2+) promotes the adhesion of marine bacteria and algae and enhances following biofilm formation in artificial seawater.

    Science.gov (United States)

    He, Xiaoyan; Wang, Jinpeng; Abdoli, Leila; Li, Hua

    2016-10-01

    Adhesion of microorganisms in the marine environment is essential for initiation and following development of biofouling. A variety of factors play roles in regulating the adhesion. Here we report the influence of Ca(2+) and Mg(2+) in artificial seawater on attachment and colonization of Bacillus sp., Chlorella and Phaeodactylum tricornutum on silicon wafer. Extra addition of the typical divalent cations in culturing solution gives rise to significantly enhanced adhesion of the microorganisms. Mg(2+) and Ca(2+) affect the adhesion of Bacillus sp. presumably by regulating aggregation and formation of extracellular polymeric substances (EPS). The ions alter quantity and types of the proteins in EPS, in turn affecting subsequent adhesion. However, it is noted that Mg(2+) promotes adhesion of Chlorella likely by regulating EPS formation and polysaccharide synthesis. Ca(2+) plays an important role in protein expression to enhance the adhesion of Chlorella. For Phaeodactylum tricornutum, Ca(2+) expedites protein synthesis for enhanced adhesion. The results shed some light on effective ways of utilizing divalent cations to mediate formation of biofilms on the marine structures for desired performances.

  19. Mg(2+)/Ca(2+) promotes the adhesion of marine bacteria and algae and enhances following biofilm formation in artificial seawater.

    Science.gov (United States)

    He, Xiaoyan; Wang, Jinpeng; Abdoli, Leila; Li, Hua

    2016-10-01

    Adhesion of microorganisms in the marine environment is essential for initiation and following development of biofouling. A variety of factors play roles in regulating the adhesion. Here we report the influence of Ca(2+) and Mg(2+) in artificial seawater on attachment and colonization of Bacillus sp., Chlorella and Phaeodactylum tricornutum on silicon wafer. Extra addition of the typical divalent cations in culturing solution gives rise to significantly enhanced adhesion of the microorganisms. Mg(2+) and Ca(2+) affect the adhesion of Bacillus sp. presumably by regulating aggregation and formation of extracellular polymeric substances (EPS). The ions alter quantity and types of the proteins in EPS, in turn affecting subsequent adhesion. However, it is noted that Mg(2+) promotes adhesion of Chlorella likely by regulating EPS formation and polysaccharide synthesis. Ca(2+) plays an important role in protein expression to enhance the adhesion of Chlorella. For Phaeodactylum tricornutum, Ca(2+) expedites protein synthesis for enhanced adhesion. The results shed some light on effective ways of utilizing divalent cations to mediate formation of biofilms on the marine structures for desired performances. PMID:27362920

  20. Divergent effects of the malignant hyperthermia-susceptible Arg(615)-->Cys mutation on the Ca(2+) and Mg(2+) dependence of the RyR1.

    OpenAIRE

    Balog, E M; Fruen, B R; Shomer, N H; Louis, C F

    2001-01-01

    The sarcoplasmic reticulum (SR) Ca(2+) release channel (RyR1) from malignant hyperthermia-susceptible (MHS) porcine skeletal muscle has a decreased sensitivity to inhibition by Mg(2+). This diminished Mg(2+) inhibition has been attributed to a lower Mg(2+) affinity of the inhibition (I) site. To determine whether alterations in the Ca(2+) and Mg(2+) affinity of the activation (A) site contribute to the altered Mg(2+) inhibition, we estimated the Ca(2+) and Mg(2+) affinities of the A- and I-si...

  1. Effects of type 1 diabetes, sprint training and sex on skeletal muscle sarcoplasmic reticulum Ca2+ uptake and Ca2+-ATPase activity.

    Science.gov (United States)

    Harmer, A R; Ruell, P A; Hunter, S K; McKenna, M J; Thom, J M; Chisholm, D J; Flack, J R

    2014-02-01

    Calcium cycling is integral to muscle performance during the rapid muscle contraction and relaxation of high-intensity exercise. Ca(2+) handling is altered by diabetes mellitus, but has not previously been investigated in human skeletal muscle. We investigated effects of high-intensity exercise and sprint training on skeletal muscle Ca(2+) regulation among men and women with type 1 diabetes (T1D, n = 8, 3F, 5M) and matched non-diabetic controls (CON, n = 8, 3F, 5M). Secondarily, we examined sex differences in Ca(2+) regulation. Subjects undertook 7 weeks of three times-weekly cycle sprint training. Before and after training, performance was measured, and blood and muscle were sampled at rest and after high-intensity exercise. In T1D, higher Ca(2+)-ATPase activity (+28%) and Ca(2+) uptake (+21%) than in CON were evident across both times and days (P women across both times and days. Intense exercise did not alter Ca(2+)-ATPase activity in T1D or CON. However, sex differences were evident: Ca(2+)-ATPase was reduced with exercise among men but increased among women across both days (time × sex interaction, P Sprint training reduced Ca(2+)-ATPase (-8%, P Sprint training reduced Ca(2+)-ATPase in T1D and CON. Sex differences in Ca(2+)-ATPase activity were evident and may be linked with fibre type proportion differences.

  2. Ultrastructural and immunohistochemical localization of plasma membrane Ca2+-ATPase 4 in Ca2+-transporting epithelia

    DEFF Research Database (Denmark)

    Alexander, R Todd; Beggs, Megan R; Zamani, Reza;

    2015-01-01

    role in transcellular Ca(2+) flux and investigated the localization and regulation of Pmca4 in Ca(2+)-transporting epithelia. Using antibodies directed specifically against Pmca4, we found it expressed only in the smooth muscle layer of mouse and human intestine, while pan-specific Pmca antibodies...... the cortical thick ascending limbs, macula densa, and early distal tubules as well as smooth muscle layers surrounding renal vessels. In human kidney, a similar pattern of distribution was observed, with highest PMCA4 expression in NCC positive tubules. Electron microscopy demonstrated Pmca4 localization...... in distal nephron cells at both the basolateral membrane and intracellular perinuclear compartments, but not submembranous vesicles, suggesting rapid trafficking to the plasma membrane is unlikely to occur in vivo. Pmca4 expression was not altered by perturbations in Ca(2+) balance, pointing...

  3. Photoproducts of tetracycline and oxytetracycline involving self-sensitized oxidation in aqueous solutions: Effects of Ca2+ and Mg2+

    Institute of Scientific and Technical Information of China (English)

    Yong Chen; Hua Li; Zongping Wang; Tao Tao; Chun Hu

    2011-01-01

    Tetracyclines constitute one of the most important antibiotic families and represent a classic example of phototoxicity.The photoproducts of tetracyclines and their parent compounds have potentially adverse effects on natural ecosystem.In this study,the self-sensitized oxidation products of tetracycline (TC) and oxytetracycline (OTC) were determined and the effects of Ca2+ and Mg2+on self-sensitized degradation were investigated.The Ca2+ and Mg2+ in the natural water sample accounted for enhancement (pH 7.3)and inhibition (pH 9.0) of photodegradation of TC and OTC due to the formation of metal-ions complexes.The formation of Mg2+ complexes was unfavorable for the photodegradation of the tetracyclines at both pH values.In contrast,the Ca2+ complexes facilitated the attack of singlet oxygen (1O2) arising from self-sensitization at pH 7.3 and enhanced TC photodegradation.For the first time,selfsensitized oxidation products of TC and OTC were verified by quenching experiments and detected by LC/ESI-DAD-MS.The products had a nominal mass 14 Da higher than the parent drugs (designated M+14),which resulted from the 1O2 attack of the dimethylamino group on the C-4 atom of the tetracyclines.The presence of Ca2+ and Mg2+ also affected the generation of M+14 due to the formation of metal-ions complexes with TC and OTC.The findings suggest that the metal-ion complexation has significant impact on the selfsensitized oxidation processes and the photoproducts of tetracyclines.

  4. Photophysical study of a polyoxo ethylene linked naphthalene-based fluorescent chemosensor for Mg2+ and Ca2+ detection.

    Science.gov (United States)

    Xiang, Xiaoyan; Wang, Dan; Guo, Yali; Liu, Weisheng; Qin, Wenwu

    2013-07-01

    A naphthalene-based bichromophoric fluorescent sensor 2,2'-[oxy-bis(2-oxatetramethyleneoxy)]-bis[N-(2-naphthyl)-benzamide)] (1) was synthesized and characterized. Fluorescence decay for 1 in alcoholic solvents in the region of 415-460 nm revealed bi-exponential behavior. The faster component of the decay can be attributed to the formation of dimers. Above 480 nm, besides the dimer, there is also a little excimer formation and this excimer emits at longer wavelengths than the dimer. The observation of the change of the fluorescence emission spectra upon addition of water in EtOH-water mixtures is in line with the formation of water-bridged complexes preventing excimer formation. The sensor shows an increase in fluorescence intensity upon increasing Mg(2+) or Ca(2+) concentration in EtOH because the formation of the excimer can be hindered upon complexation with Mg(2+) or Ca(2+) ions. Because of the competition between hydrated metal ions and the water-bridged complex, spectral changes by complexation with Mg(2+) or Ca(2+) in EtOH-H2O (9 : 1 v/v) are quite different from those in neat ethanol. The ground-state dissociation constant K(d) estimated for the complex with Mg(2+) or Ca(2+) was found to be around 2.0 mM in EtOH-H2O (9 : 1 v/v), which makes it suitable for the measurement of the concentrations of these ions in physiologically relevant concentration ranges.

  5. Strong Dependence of Hydration State of F-Actin on the Bound Mg(2+)/Ca(2+) Ions.

    Science.gov (United States)

    Suzuki, Makoto; Imao, Asato; Mogami, George; Chishima, Ryotaro; Watanabe, Takahiro; Yamaguchi, Takaya; Morimoto, Nobuyuki; Wazawa, Tetsuichi

    2016-07-21

    Understanding of the hydration state is an important issue in the chemomechanical energetics of versatile biological functions of polymerized actin (F-actin). In this study, hydration-state differences of F-actin by the bound divalent cations are revealed through precision microwave dielectric relaxation (DR) spectroscopy. G- and F-actin in Ca- and Mg-containing buffer solutions exhibit dual hydration components comprising restrained water with DR frequency f2 (fw). The hydration state of F-actin is strongly dependent on the ionic composition. In every buffer tested, the HMW signal Dhyme (≡ (f1 - fw)δ1/(fwδw)) of F-actin is stronger than that of G-actin, where δw is DR-amplitude of bulk solvent and δ1 is that of HMW in a fixed-volume ellipsoid containing an F-actin and surrounding water in solution. Dhyme value of F-actin in Ca2.0-buffer (containing 2 mM Ca(2+)) is markedly higher than in Mg2.0-buffer (containing 2 mM Mg(2+)). Moreover, in the presence of 2 mM Mg(2+), the hydration state of F-actin is changed by adding a small fraction of Ca(2+) (∼0.1 mM) and becomes closer to that of the Ca-bound form in Ca2.0-buffer. This is consistent with the results of the partial specific volume and the Cotton effect around 290 nm in the CD spectra, indicating a change in the tertiary structure and less apparent change in the secondary structure of actin. The number of restrained water molecules per actin (N2) is estimated to be 1600-2100 for Ca2.0- and F-buffer and ∼2500 for Mg2.0-buffer at 10-15 °C. These numbers are comparable to those estimated from the available F-actin atomic structures as in the first water layer. The number of HMW molecules is roughly explained by the volume between the equipotential surface of -kT/2e and the first water layer of the actin surface by solving the Poisson-Boltzmann equation using UCSF Chimera. PMID:27332748

  6. Regulation of plant plasma membrane H+- and Ca2+-ATPases by terminal domains

    DEFF Research Database (Denmark)

    Bækgaard, Lone; Fuglsang, Anja Thoe; Palmgren, Michael Gjedde

    2005-01-01

    In the last few years, major progress has been made to elucidate the structure, function, and regulation of P-type plasma membrane H(+)-and Ca(2+)-ATPases. Even though a number of regulatory proteins have been identified, many pieces are still lacking in order to understand the complete regulatory...... mechanisms of these pumps. In plant plasma membrane H(+)- and Ca(2+)-ATPases, autoinhibitory domains are situated in the C- and N-terminal domains, respectively. A model for a common mechanism of autoinhibition is discussed....

  7. All Ca(2+)-binding loops of light-sensitive ctenophore photoprotein berovin bind magnesium ions: The spatial structure of Mg(2+)-loaded apo-berovin.

    Science.gov (United States)

    Burakova, Ludmila P; Natashin, Pavel V; Malikova, Natalia P; Niu, Fengfeng; Pu, Mengchen; Vysotski, Eugene S; Liu, Zhi-Jie

    2016-01-01

    Light-sensitive photoprotein berovin accounts for a bright bioluminescence of ctenophore Beroe abyssicola. Berovin is functionally identical to the well-studied Ca(2+)-regulated photoproteins of jellyfish, however in contrast to those it is extremely sensitive to the visible light. Berovin contains three EF-hand Ca(2+)-binding sites and consequently belongs to a large family of the EF-hand Ca(2+)-binding proteins. Here we report the spatial structure of apo-berovin with bound Mg(2+) determined at 1.75Å. The magnesium ion is found in each functional EF-hand loop of a photoprotein and coordinated by oxygen atoms donated by the side-chain groups of aspartate, carbonyl groups of the peptide backbone, or hydroxyl group of serine with characteristic oxygen-Mg(2+) distances. As oxygen supplied by the side-chain of the twelfth residue of all Ca(2+)-binding loops participates in the magnesium ion coordination, it was suggested that Ca(2+)-binding loops of berovin belong to the mixed Ca(2+)/Mg(2+) rather than Ca(2+)-specific type. In addition, we report an effect of physiological concentration of Mg(2+) on bioluminescence of berovin (sensitivity to Ca(2+), rapid-mixed kinetics, light-sensitivity, thermostability, and apo-berovin conversion into active protein). The different impact of physiological concentration of Mg(2+) on berovin bioluminescence as compared to hydromedusan photoproteins was attributed to different affinities of the Ca(2+)-binding sites of these photoproteins to Mg(2+). PMID:26690016

  8. All Ca(2+)-binding loops of light-sensitive ctenophore photoprotein berovin bind magnesium ions: The spatial structure of Mg(2+)-loaded apo-berovin.

    Science.gov (United States)

    Burakova, Ludmila P; Natashin, Pavel V; Malikova, Natalia P; Niu, Fengfeng; Pu, Mengchen; Vysotski, Eugene S; Liu, Zhi-Jie

    2016-01-01

    Light-sensitive photoprotein berovin accounts for a bright bioluminescence of ctenophore Beroe abyssicola. Berovin is functionally identical to the well-studied Ca(2+)-regulated photoproteins of jellyfish, however in contrast to those it is extremely sensitive to the visible light. Berovin contains three EF-hand Ca(2+)-binding sites and consequently belongs to a large family of the EF-hand Ca(2+)-binding proteins. Here we report the spatial structure of apo-berovin with bound Mg(2+) determined at 1.75Å. The magnesium ion is found in each functional EF-hand loop of a photoprotein and coordinated by oxygen atoms donated by the side-chain groups of aspartate, carbonyl groups of the peptide backbone, or hydroxyl group of serine with characteristic oxygen-Mg(2+) distances. As oxygen supplied by the side-chain of the twelfth residue of all Ca(2+)-binding loops participates in the magnesium ion coordination, it was suggested that Ca(2+)-binding loops of berovin belong to the mixed Ca(2+)/Mg(2+) rather than Ca(2+)-specific type. In addition, we report an effect of physiological concentration of Mg(2+) on bioluminescence of berovin (sensitivity to Ca(2+), rapid-mixed kinetics, light-sensitivity, thermostability, and apo-berovin conversion into active protein). The different impact of physiological concentration of Mg(2+) on berovin bioluminescence as compared to hydromedusan photoproteins was attributed to different affinities of the Ca(2+)-binding sites of these photoproteins to Mg(2+).

  9. An improvement to the ligand optimisation method (LOM) for measuring the apparent dissociation constant and ligand purity in Ca2+ and Mg2+ buffer solutions.

    Science.gov (United States)

    McGuigan, John A S; Kay, James W; Elder, Hugh Y

    2014-01-01

    In Ca(2+)/Mg(2+) buffers the calculated ionised concentrations ([X(2+)]) can vary by up to a factor of seven. Since there are no defined standards it is impossible to check calculated [X(2+)], making measurement essential. The ligand optimisation method (LOM) is an accurate method to measure [X(2+)] in Ca(2+)/Mg(2+) buffers; independent estimation of ligand purity extends the method to pK(/) buffers, to calculate electrode and buffer characteristics as a function of Σ. Ca(2+)-electrodes have a Σ buffers. These results demonstrated that it is pK(/) that is normally distributed. Until defined standards are available, [X(2+)] in Ca(2+)/Mg(2+) buffers have to be measured. The most appropriate method is to use Ca(2+)/Mg(2) electrodes combined with the Excel programs SALE or AEC.

  10. The Ca2+/H+ antiporter TMEM165 expression, localization in the developing, lactating and involuting mammary gland parallels the secretory pathway Ca2+ATPase (SPCA1)

    Science.gov (United States)

    Plasma membrane Ca2+-ATPase 2 (PMCA2) knockout mice showed that ~ 60 % of calcium in milk is transported across the mammary cells apical membrane by PMCA2. The remaining milk calcium is thought to arrive via the secretory pathway through the actions of secretory pathway Ca2+-ATPase’s 1 and/or 2 (SP...

  11. A high affinity Ca2(+)-ATPase on the surface membrane of Leishmania donovani promastigote

    Energy Technology Data Exchange (ETDEWEB)

    Ghosh, J.; Ray, M.; Sarkar, S.; Bhaduri, A. (Indian Institute of Chemical Biology, Calcutta (India))

    1990-07-05

    A Ca2(+)-dependent ATP-hydrolytic activity was detected in the crude membrane ghost of the promastigote or vector form of the protozoal parasite Leishmania donovani, the pathogen responsible for kala azar. The Ca2(+)-ATPase was purified to apparent homogeneity after solubilization with deoxycholate. The enzyme consists of two subunits of Mr = 51,000 and 57,000 and has an apparent molecular weight of 215,000 +/- 12,000. The enzyme activity is exclusively dependent on Ca2+, and the pure enzyme can hydrolyze 1.6 mumol of ATP/min/mg of protein. The apparent Km for Ca2+ is 35 nM, which is further reduced to 12 nM in the presence of heterologous calmodulin. The enzyme is sensitive to vanadate, but is insensitive to oligomycin and ouabain. The enzyme is strongly associated with the plasma membrane and has its catalytic site oriented toward the cytoplasmic face. The enzyme spans across the plasma membrane as surface labeling with radioiodine shows considerable radioactivity in the completely purified enzyme. The localization and orientation of this high affinity, calmodulin-sensitive Ca2(+)-ATPase suggest some role of this enzyme in Ca2+ movement in the life cycle of this protozoal parasite.

  12. Direct demonstration of Ca2+ binding defects in sarco-endoplasmic reticulum Ca2+ ATPase mutants overexpressed in COS-1 cells transfected with adenovirus vectors.

    Science.gov (United States)

    Strock, C; Cavagna, M; Peiffer, W E; Sumbilla, C; Lewis, D; Inesi, G

    1998-06-12

    Single mutations of specific amino acids within the membrane-bound region of the sarco-endoplasmic reticulum Ca2+ (SERCA)-1 ATPase interfere with Ca2+ inhibition of ATPase phosphorylation by Pi (1), suggesting that these residues may be involved in complexation of two Ca2+ that are known to bind to the enzyme. However, direct measurements of Ca2+ binding in the absence of ATP have been limited by the low quantities of available mutant protein. We have improved the transfection efficiency by means of recombinant adenovirus vectors, yielding sufficient expression of wild type and mutant SERCA-1 ATPase for measurements of Ca2+ binding to the microsomal fraction of the transfected cells. We find that in the presence of 20 microM Ca2+ and in the absence of ATP, the Glu771 --> Gln, Thr799 --> Ala, Asp800 --> Asn, and Glu908 --> Ala mutants exhibit negligible binding, indicating that the oxygen functions of Glu771, Thr799, Asp800, and Glu908 are involved in interactions whose single disruption causes major changes in the highly cooperative "duplex" binding. Total loss of Ca2+ binding is accompanied by loss of Ca2+ inhibition of the Pi reaction. We also find that, at pH 7.0, the Glu309 --> Gln and the Asn796 --> Ala mutants bind approximately half as much Ca2+ as the wild type ATPase and do not interfere with Ca2+ inhibition of the Pi reaction. At pH 6.2, the Glu309 --> Gln mutant does not bind any Ca2+, and its phosphorylation by Pi is not inhibited by Ca2+. On the contrary, the Asn796 --> Ala mutant retains the behavior displayed at pH 7.0. This suggests that in the Glu309 --> Gln mutant, ionization of acidic functions in other amino acids (e.g. Glu771 and Asp800) occurs as the pH is shifted, thereby rendering Ca2+ binding possible. In the Asn796 --> Ala mutant, on the other hand, the Glu309 carboxylic function allows binding of inhibitory Ca2+ even at pH 6.2. In all cases mutational interference with the inhibition of the Pi reaction by Ca2+ can be overcome by raising

  13. Two-Dimensional Crystallization of the Ca(2+)-ATPase for Electron Crystallography.

    Science.gov (United States)

    Glaves, John Paul; Primeau, Joseph O; Young, Howard S

    2016-01-01

    Electron crystallography of two-dimensional crystalline arrays is a powerful alternative for the structure determination of membrane proteins. The advantages offered by this technique include a native membrane environment and the ability to closely correlate function and dynamics with crystalline preparations and structural data. Herein, we provide a detailed protocol for the reconstitution and two-dimensional crystallization of the sarcoplasmic reticulum calcium pump (also known as Ca(2+)-ATPase or SERCA) and its regulatory subunits phospholamban and sarcolipin.

  14. Biochemical Evidences for Scopoletin lnhibits Ca2+-ATPase Activity in the Carmine Spider Mite, Tetranychus cinnabarinus (Boisduval)

    Institute of Scientific and Technical Information of China (English)

    Qiuli HOU; Dan WANG; Bingchuan ZHANG; Wei DlNG; Yongqiang ZHANG

    2015-01-01

    Objective] This study almed to investigate the acaricidal effect of scopo-Ietin, and provide the biochemical evidences of scopoIetin infIuences Ca2+-ATPase activity and gene expressions in the Carmine Spider Mite, Tetranychus cinnabarinus. [Method] The acaricidal effects of scopoIetin were investigated by sIip-dip method. Exposeed to different concentrations of scopoIetin (0.16-2.5 mg/mI), Ca2+-ATPase ac-tivity in vivo and protein contents were investigated. For assessing the in vitro ef-fect, Ca2+-ATPase enzyme (200 μI) prepared from normal mites were incubated with different concentrations of scopoIetin reagents. [Result] ScopoIetin exhibited signifi-cant inhibitory effect on Ca2+-ATPase activity both in vivo and in vitro, and resuIted in increased protein contents; kinetic analysis showed that the catalytic capabiIity of Ca2+-ATPase was significantIy reduced by scopoIetin. [Conclusion] ScopoIetin exhibits a significant inhibitory effect on Ca2+-ATPase , and its acaricidal effect agalnst T . cinnabarinus might be due to the direct inhibition of Ca2+-ATPase.

  15. Sarco/Endoplasmic reticulum Ca2+-ATPases (SERCA contribute to GPCR-mediated taste perception.

    Directory of Open Access Journals (Sweden)

    Naoko Iguchi

    Full Text Available The sense of taste is important for providing animals with valuable information about the qualities of food, such as nutritional or harmful nature. Mammals, including humans, can recognize at least five primary taste qualities: sweet, umami (savory, bitter, sour, and salty. Recent studies have identified molecules and mechanisms underlying the initial steps of tastant-triggered molecular events in taste bud cells, particularly the requirement of increased cytosolic free Ca(2+ concentration ([Ca(2+](c for normal taste signal transduction and transmission. Little, however, is known about the mechanisms controlling the removal of elevated [Ca(2+](c from the cytosol of taste receptor cells (TRCs and how the disruption of these mechanisms affects taste perception. To investigate the molecular mechanism of Ca(2+ clearance in TRCs, we sought the molecules involved in [Ca(2+](c regulation using a single-taste-cell transcriptome approach. We found that Serca3, a member of the sarco/endoplasmic reticulum Ca(2+-ATPase (SERCA family that sequesters cytosolic Ca(2+ into endoplasmic reticulum, is exclusively expressed in sweet/umami/bitter TRCs, which rely on intracellular Ca(2+ release for signaling. Serca3-knockout (KO mice displayed significantly increased aversive behavioral responses and greater gustatory nerve responses to bitter taste substances but not to sweet or umami taste substances. Further studies showed that Serca2 was mainly expressed in the T1R3-expressing sweet and umami TRCs, suggesting that the loss of function of Serca3 was possibly compensated by Serca2 in these TRCs in the mutant mice. Our data demonstrate that the SERCA family members play an important role in the Ca(2+ clearance in TRCs and that mutation of these proteins may alter bitter and perhaps sweet and umami taste perception.

  16. Pharmacological evidence that potentiation of plasmalemmal Ca(2+)-extrusion is functionally coupled to inhibition of SR Ca(2+)-ATPases in vascular smooth muscle cells.

    Science.gov (United States)

    Zhang, Wen-Bo; Kwan, Chiu-Yin

    2016-04-01

    Cyclopiazonic acid (CPA), a specific inhibitor of sarcoplasmic reticulum (SR) Ca(2+)-ATPases, causes slowly developing and subsequently diminishing characteristic contractions in vascular smooth muscle, and the second application of CPA has incompletely repeatable effects, depending on the vessel type. The objective of the present study was to examine the mechanisms underlying the significant decrease of CPA-induced contractions upon the second application. A pharmacological intervention of Ca(2+) extrusion process as a strategy was performed to modulate vasoconstrictor effects of CPA in rat aortic ring preparations. CPA-induced contractions, expressed as percentages of the contractions induced by KCl (80 mM), were significantly decreased from 44.1 ± 5.7 to 7.6 ± 1.8 % (P CPA-induced contractions were sustained and completely repeatable in Na(+)-free and low Na(+) medium. Furthermore, we found that the contractions were completely repeatable in the presence of 2',4'-dichlorobenzamil, an inhibitor of the forward mode of Na(+)/Ca(2+) exchangers, but not of KBR7943, an inhibitor of the reverse mode of Na(+)/Ca(2+) exchangers. Our findings indicate that CPA by inducing a transient rise in cytosolic Ca(2+) level causes a long-lasting upregulation of plasma membrane (PM) Ca(2+) extruders and thus leads to a diminished contraction upon its second application in blood vessels. This suggests that there is a functional coupling between PM Ca(2+) extruders and SR Ca(2+)-ATPases in rat aortic smooth muscle cells. PMID:26842648

  17. Propofol regulates Ca2+, Mg2+, Cu2+ and Zn2+ balance in the spinal cord after ischemia/reperfusion injury

    Institute of Scientific and Technical Information of China (English)

    Shuzhou Yin; Qijing Yu; Ji Hu; Jie Yang; Juan Chen

    2011-01-01

    This study assessed concentrations of Ca2+, Mg2+, Cu2+ and Zn2+ in blood serum and spinal cord tissues, as well as the possible mechanisms by which propofol may protect spinal cord tissues during ischemia/reperfusion injury. With prolonged duration of ischemia/reperfusion injury, serum Ca2+ and Cu2+ concentrations gradually increased, but Mg 2+ and Zn2+ concentrations gradually decreased. Seven days after spinal cord injury, changes in Ca2+, Mg2+, Cu2+ and Zn2+ concentrations were significant. After 7 days of reperfusion, changes in the concentrations of Ca2+, Mg2+, Cu2+ and Zn2+ in spinal cord homogenates were consistent with those in the serum. After propofol treatment, no significant changes in Ca2+, Mg2+, Cu2+ and Zn2+ concentrations in serum and spinal cord homogenates were noted during ischemia/reperfusion injury. These findings suggest that propofol exerts protective effects against spinal cord injury by stabilizing or recovering metal ion balance in ischemic regions.

  18. Epigallocatechin-3-Gallate Protects Erythrocyte Ca2+-ATPase and Na+/K+-ATPase Against Oxidative Induced Damage During Aging in Humans

    Directory of Open Access Journals (Sweden)

    Prabhanshu Kumar

    2014-10-01

    Full Text Available Purpose: The main purpose of this study was to investigate the protective role of epigallocatechin-3-gallate on tertiary butyl hydroperoxide induced oxidative damage in erythrocyte during aging in humans. Methods: Human erythrocyte membrane bound Ca2+-ATPase and Na+/K+-ATPase activities were determined as a function of human age. Protective role of epigallocatechin-3-gallate was evaluated by in vitro experiments by adding epigallocatechin-3-gallate in concentration dependent manner (final concentration range 10-7M to 10-4M to the enzyme assay medium. Oxidative stress was induced in vitro by incubating washed erythrocyte ghosts with tertiary butyl hydroperoxide (10-5 M final concentration. Results: We have reported concentration dependent effect of epigallocatechin-3-gallate on tertiary butyl hydroperoxide induced damage on activities of Ca2+-ATPase and Na+/K+-ATPase during aging in humans. We have detected a significant (p < 0.001 decreased activity of Ca2+-ATPase and Na+/K+ -ATPase as a function of human age. Epigallocatechin-3-gallate protected ATPases against tertiary butyl hydroperoxide induced damage in concentration dependent manner during aging in humans. Conclusion: Epigallocatechin-3-gallate is a powerful antioxidant that is capable of protecting erythrocyte Ca2+-ATPase and Na+/K+ -ATPase against oxidative stress during aging in humans. We may propose hypothesis that a high intake of catechin rich diet may provide some protection against development of aging and age related diseases.

  19. Effect of ultrasonic exposure on Ca2+-ATPase activity in plasma membrane from Aloe arborescens callus cells.

    Science.gov (United States)

    Liu, Yiyao; Yang, Hong; Takatsuki, Hideyo; Sakanishi, Akio

    2006-04-01

    We investigated the effect of ultrasound on plasma membrane (PM) Ca2+-ATPase activity of Aloe arborescens callus cells in solid culture. The calluses were exposed by a 20 kHz digital sonifier at the powers of 2 and 10 W from the effective exposure times of 2-10 s. PM Ca2+-ATPase activity was almost significantly higher at 2 W both in continuous wave and 10% duty cycle than that of the control (no ultrasound) at effective exposure times of 5 and 10 s. However, its activity decreased at 10 W in continuous wave exposure. It is possible that the PM Ca2+-ATPase configuration or structure may be partly damaged by high-energy ultrasound at 10 W. Our results showed that low-energy ultrasound exposure was a useful physical field to stimulate A. arborescens callus cells to adapt environmental stress through PM Ca2+-ATPase activity increase. PMID:15936236

  20. Plasma membrane Ca2+-ATPases in the nervous system during development and ageing

    Institute of Scientific and Technical Information of China (English)

    Ana; M; Mata; M; Rosario; Sepulveda

    2010-01-01

    Calcium signaling is used by neurons to control a variety of functions,including cellular differentiation,synaptic maturation,neurotransmitter release,intracellular signaling and cell death.This review focuses on one of the most important Ca2+regulators in the cell,the plasma membrane Ca2+-ATPase(PMCA),which has a high affinity for Ca2+and is widely expressed in brain.The ontogeny of PMCA isoforms,linked to specific requirements of Ca2+ during development of different brain areas,is addressed, as well as their function in the adult tissue.This is based on the high diversity of variants in the PMCA family in brain,which show particular kinetic differences possibly related to specific localizations and functions of the cell. Conversely,alterations in the activity of PMCAs could lead to changes in Ca2+homeostasis and,consequently,to neural dysfunction.The involvement of PMCA isoforms in certain neuropathologies and in brain ageing is also discussed.

  1. Photoluminescence of Vanadate Garnet Ca2NaMg(2-x)V3O12:xEu3+ Phosphors Synthesized by Solution Combustion Method.

    Science.gov (United States)

    Kim, H; Kim, J; Lim, S; Park, K

    2016-02-01

    In this study, a series of nano-sized Ca2NaMg(2-x)V3O12:xEu3+ (0.06 phosphors is synthesized by solution combustion method. The microstructure and photoluminescence properties of the Ca2NaMg(2-x)V3O12:xEu3+ phosphors are studied in accordance with the Eu3+ content. Annealed Ca2NaMg(2-x)V3O12:xEu3+ phosphors form a single phase with the cubic garnet structure and Ia3d space group. The emission from (VO4)3- in the Ca2NaMg(2-x)V3O12:xEu3+ phosphors is almost completely quenched due to the efficient energy transfer from the (VO4)3- to the Eu3+ ions. The emission intensity increases sharply with the increased Eu3+ content, reaching a maximum value at x = 0.15, and then decreases with further Eu3+ content. Ca2NaMg1.85V3O12:0.15Eu3+ shows great potential as a red phosphor for white light-emitting diodes. PMID:27433680

  2. Coordinated regulation of cardiac Na(+)/Ca (2+) exchanger and Na (+)-K (+)-ATPase by phospholemman (FXYD1).

    Science.gov (United States)

    Cheung, Joseph Y; Zhang, Xue-Qian; Song, Jianliang; Gao, Erhe; Chan, Tung O; Rabinowitz, Joseph E; Koch, Walter J; Feldman, Arthur M; Wang, JuFang

    2013-01-01

    Phospholemman (PLM) is the founding member of the FXYD family of regulators of ion transport. PLM is a 72-amino acid protein consisting of the signature PFXYD motif in the extracellular N terminus, a single transmembrane (TM) domain, and a C-terminal cytoplasmic tail containing three phosphorylation sites. In the heart, PLM co-localizes and co-immunoprecipitates with Na(+)-K(+)-ATPase, Na(+)/Ca(2+) exchanger, and L-type Ca(2+) channel. The TM domain of PLM interacts with TM9 of the α-subunit of Na(+)-K(+)-ATPase, while its cytoplasmic tail interacts with two small regions (spanning residues 248-252 and 300-304) of the proximal intracellular loop of Na(+)/Ca(2+) exchanger. Under stress, catecholamine stimulation phosphorylates PLM at serine(68), resulting in relief of inhibition of Na(+)-K(+)-ATPase by decreasing K(m) for Na(+) and increasing V(max), and simultaneous inhibition of Na(+)/Ca(2+) exchanger. Enhanced Na(+)-K(+)-ATPase activity lowers intracellular Na(+), thereby minimizing Ca(2+) overload and risks of arrhythmias. Inhibition of Na(+)/Ca(2+) exchanger reduces Ca(2+) efflux, thereby preserving contractility. Thus, the coordinated actions of PLM during stress serve to minimize arrhythmogenesis and maintain inotropy. In acute cardiac ischemia and chronic heart failure, either expression or phosphorylation of PLM or both are altered. PLM regulates important ion transporters in the heart and offers a tempting target for development of drugs to treat heart failure.

  3. Plasma membrane Ca2+-ATPases:Targets of oxidative stress in brain aging and neurodegeneration

    Institute of Scientific and Technical Information of China (English)

    Asma; Zaidi

    2010-01-01

    The plasma membrane Ca2+-ATPase(PMCA)pumps play an important role in the maintenance of precise levels of intracellular Ca2+[Ca2+]i,essential to the functioning of neurons.In this article,we review evidence showing age-related changes of the PMCAs in synaptic plasma membranes(SPMs).PMCA activity and protein levels in SPMs diminish progressively with increasing age. The PMCAs are very sensitive to oxidative stress and undergo functional and structural changes when exposed to oxidants of physiological relevance.The major signatures of oxidative modification in the PMCAs are rapid inactivation,conformational changes,aggregation, internalization from the plasma membrane and proteolytic degradation.PMCA proteolysis appears to be mediated by both calpains and caspases.The predominance of one proteolytic pathway vs the other,the ensuing pattern of PMCA degradation and its consequence on pump activity depends largely on the type of insult,its intensity and duration.Experimental reduction of PMCA expression not only alters the dynamics of cellular Ca2+ handling but also has a myriad of downstream conse-quences on various aspects of cell function,indicating a broad role of these pumps.Age-and oxidation-related down-regulation of the PMCAs may play an important role in compromised neuronal function in the aging brain and its several-fold increased susceptibility to neurodegenerative disorders such as Alzheimer’s disease, Parkinson’s disease,and stroke.Therapeutic approaches that protect the PMCAs and stabilize[Ca2+]i homeostasis may be capable of slowing and/or preventing neuronal degeneration.The PMCAs are therefore emerging as a new class of drug targets for therapeutic interventions in various chronic degenerative disorders.

  4. 海水对兔眼晶状体的Na,K-ATP酶及Ca2,Mg2-ATP酶功能的影响%Experimental Study on the Effects of Seawater on Na+,K+-ATP ase and Ca2+,Mg2+-ATP ase in Lens of Rabbits

    Institute of Scientific and Technical Information of China (English)

    徐绍娟; 彭秀军; 魏翠荣; 竹颖

    2013-01-01

    Objective:To investigate the changes of Na+,K+-ATP ase and Ca2+,Mg2+-ATP ase in lens of rabbits with seawater injected into anterior chambers.Methods:After 0.2 ml seawater were injected into anterior chambers,the rabbit’s eye balls were enucleated respectively in deferent time to examine the activity of Na+,K+-ATP ase and Ca2+,Mg2+-ATP ase in lens by biochemistry method.Results:The activity of Na+,K+-ATP ase and Ca2+,Mg2+-ATP ase in lens of seawater group decreased significantly at first,then increased and recovered partly or completely at last.No significant difference of the activity of two kinds of enzyme in lens was found between control group and normal group.Conclusion:The activity of Na+,K+-ATP ase and Ca2+,Mg2+-ATP ase in rabbit lens are damaged in deferent degree with seawater injected into anterior chamber.%  目的:观察海水对兔眼晶状体Na+,K+-ATP酶及Ca2+,Mg2+-ATP酶功能的影响.方法:在兔眼前房内注入0.2 ml海水后不同时间摘除眼球,分别测定其Na+,K+-ATP酶及Ca2+,Mg2+-ATP酶活性.结果:海水组兔眼晶状体两酶活性均先降低,再增高,最终完全或部分恢复正常;而对照组两酶的活性与正常组比较没有显著差别.结论:海水可使晶状体的Na+,K+-ATP酶及Ca2+,Mg2+-ATP酶的活性不同程度受损.

  5. Changes of mitochondrial structure, ATPase and Ca2+ concentration in spermatogenic cells of mouse testes induced by low dose radiation

    International Nuclear Information System (INIS)

    Objective: To observe the ultrastructure, ATPase activity and Ca2+ concentration ([Ca2+]i) of mitochondria in the sperematogenic cells of mouse testes 3-24 h after low dose radiation with 0.025-0.200 Gy X-rays, and illuminate the effects of mitochondrion structure and relative biological function on apoptosis. Methods: The ultrastructure changes of mitochondria in the spermatogenic cells were observed with transmission electron microscope; the ATPase activity was measured with protein enzymic method; [Ca2+]i was measured indirectly by flow cytometry with Fluo-3 probes. Results: The mitochondria swelled and vacuolizated, and their cristae were broken in the spermatogonia and spermatocytes 12 h after irradiation, and their nuclei were karyopyknosis, the acrosomal vesicle structure was ambiguity, the membrane structure was unclear, and the mitochondria in spermatids were vacuolization. The activities of Na+-K+-ATPase in mouse testis tissue 12 h after irradiated with 0.025-0.200 Gy decreased compared with those with 0 Gy, the Na+-K+-ATPase activities of the cells irradiated with 0.05-0.200 Gy decreased significantly compared with those with 0 Gy (P2+-ATPase of the cells irradiated with 0.025-0.200 Gy decreased significantly compared with those with 0 Gy (P2+]i in mouse testis spermatogenic cells had similar dose-response relationship, [Ca2+]i after irradiated with 0.075 Gy decreased compared with those with 0 Gy (P+-K+-ATPase in mouse testis tissues decreased obviously compared with those at 0 h (P2+-ATPase in mouse testis tissues increased slightly at 3 h, then decreased at 6-24 h compared with those at 0 h (P2+]i in mouse testis spermatogenic cells had similar time course-response relationship, [Ca2+]i at 12 h decreased significantly compared with at 0 h (P2+]i induced by low dose radiation. (authors)

  6. Contribution of plasma membrane Ca2+ ATPase to cerebellar synapse function

    Institute of Scientific and Technical Information of China (English)

    Helena; Huang; Raghavendra; Y; Nagaraja; Molly; L; Garside; Walther; Akemann; Thomas; Knpfel; Ruth; M; Empson

    2010-01-01

    The cerebellum expresses one of the highest levels of the plasma membrane Ca2+ATPase,isoform 2 in the mammalian brain.This highly efficient plasma membrane calcium transporter protein is enriched within the main output neurons of the cerebellar cortex;i.e. the Purkinje neurons(PNs) .Here we review recent evidence,including electrophysiological and calcium imaging approaches using the plasma membrane calcium ATPase 2(PMCA2) knockout mouse,to show that PMCA2 is critical for the physiological control of calcium at cerebellar synapses and cerebellar dependent behaviour.These studies have also revealed that deletionof PMCA2 throughout cerebellar development in the PMCA2 knockout mouse leads to permanent signalling and morphological alterations in the PN dendrites. Whilst these findings highlight the importance of PMCA2 during cerebellar synapse function and development,they also reveal some limitations in the use of the PMCA2 knockout mouse and the need for additional experimental approaches including cell-specific and reversible manipulation of PMCAs.

  7. A sulphonated carbon dot-chitosan hybrid hydrogel nanocomposite as an efficient ion-exchange film for Ca2+ and Mg2+ removal

    Science.gov (United States)

    Baruah, Upama; Konwar, Achyut; Chowdhury, Devasish

    2016-04-01

    We have developed a hybrid hydrogel nanocomposite film via conjugation of oxidised carbon dots synthesized from 11-mercaptoundecanoic acid with chitosan. The potential applicability of the film was then successfully tested for the removal of Ca2+ and Mg2+ ions from solution.We have developed a hybrid hydrogel nanocomposite film via conjugation of oxidised carbon dots synthesized from 11-mercaptoundecanoic acid with chitosan. The potential applicability of the film was then successfully tested for the removal of Ca2+ and Mg2+ ions from solution. Electronic supplementary information (ESI) available: The ESI includes the detailed synthesis and characterization of carbon dots both before and after oxidation and of the carbon dot-chitosan nanocomposite films viz. DLS, SEM, UV-visible, FTIR, PL spectroscopy and TGA. See DOI: 10.1039/c6nr01129b

  8. Thapsigargin, a tumor promoter, discharges intracellular Ca2+ stores by specific inhibition of the endoplasmic reticulum Ca2(+)-ATPase

    DEFF Research Database (Denmark)

    Thastrup, Ole; Cullen, P J; Drøbak, B K;

    1990-01-01

    Thapsigargin, a tumor-promoting sesquiterpene lactone, discharges intracellular Ca2+ in rat hepatocytes, as it does in many vertebrate cell types. It appears to act intracellularly, as incubation of isolated rat liver microsomes with thapsigargin induces a rapid, dose-dependent release of stored Ca...

  9. Molecular dynamics simulation exploration of cooperative migration mechanism of calcium ions in sarcoplasmic reticulum Ca2+-ATPase.

    Science.gov (United States)

    Huang, Yongqi; Li, Huifang; Bu, Yuxiang

    2009-10-01

    Calcium ATPase is a member of the P-type ATPase, and it pumps calcium ions from the cytoplasm into the reticulum against a concentration gradient. Several X-ray structures of different conformations have been solved in recent years, providing basis for elucidating the active transport mechanism of Ca2+ ions. In this work, molecular dynamics (MD) simulations were performed at atomic level to investigate the dynamical process of calcium ions moving from the outer mouth of the protein to their binding sites. Five initial locations of Ca2+ ions were considered, and the simulations lasted for 2 or 6 ns, respectively. Specific pathways leading to the binding sites and large structural rearrangements around binding sites caused by uptake of calcium ions were identified. A cooperative binding mechanism was observed from our simulation. Firstly, the first Ca2+ ion binds to site I, and then, the second Ca2+ ion approaches. The interactions between the second Ca2+ and the residues around site I disturb the binding state of site I and weaken its binding ability for the first bound Ca2+. Because of the electrostatic repulsion of the second Ca2+ and the electrostatic attraction of site II, the first bound Ca2+ shifts from site I to site II. Concertedly, the second Ca2+ binds to site I, forming a binding state with two Ca2+ ions, one at site I and the other at site II. Both of Glu908 and Asp800 coordinate with the two Ca2+ ions simultaneously during the concerted binding process, which is believed to be the hinge to achieve the concerted binding. In our simulations, four amino acid residues that serve as the channel to link the outer mouth and the binding sites during the binding process were recognized, namely Tyr837, Tyr763, Asn911, and Ser767. The analyses regarding the activity of the proteins via mutations of some key residues also supported our cooperative mechanism. PMID:19242958

  10. Cyclopiazonic Acid Is Complexed to a Divalent Metal Ion When Bound to the Sarcoplasmic Reticulum Ca2+-ATPase

    DEFF Research Database (Denmark)

    Laursen, Mette; Bublitz, Maike; Moncoq, Karine;

    2009-01-01

    is expected to be part of the cytoplasmic cation access pathway. Our model is consistent with the biochemical data on CPA function and provides new measures in structure-based drug design targeting Ca2+-ATPases, e. g. from pathogens. We also present an extended structural basis of ATP modulation pinpointing...

  11. The Relationship Between Senescence and Ca2+-ATPase Activity of Microsomal Membrane and Lipid Peroxidation in Harvested Peach Fruit

    Institute of Scientific and Technical Information of China (English)

    GUAN Jun-feng; FAN Xiu-cai; DOU Shi-juan; ZHANG Ji-shu; LI Guang-min

    2006-01-01

    Peach fruit easily soften and have a short storage time at normal temperature. In this study, peach fruit (Prunus persica sieb et Zucc cv. Yingqing) were picked and stored at 25 and 4℃ to investigate the senescence in correlation with Ca2+- ATPase activity of microsomal membrane and lipid peroxidation during ripening and senescence. In comparison with that stored at 25℃, the fruit stored at 4℃ exhibited a higher flesh firmness, lower respiration rate, and generated the late bigger peak value of Ca2+-ATPase activity as well as maintained the higher activity of the enzyme. Meanwhile, the lower levels of super oxygen radical (O2-) production and content of malondialdehyde (MDA), a product of membrane lipid peroxidation were observed. Sodium orthovanadate (SO) and erythrosin B (EB), as Ca2+-ATPase inhibitors, could stimulate the respiration rate. The results suggested that the slower senescence rate of peach fruit was closely related to the higher peak value and longer duration of Ca2+-ATPase activity in microsomal membrane, with the slighter membrane lipid peroxidation and lower O2(-) production rate.

  12. Role of platelet plasma membrane Ca2+-ATPase in health and disease

    Institute of Scientific and Technical Information of China (English)

    William; L; Dean

    2010-01-01

    Platelets have essential roles in both health and disease. Normal platelet function is required for hemostasis.Inhibition of platelet function in disease or by pharmacological treatment results in bleeding disorders.On the other hand,hyperactive platelets lead to heart attack and stroke.Calcium is a major second messenger in platelet activation,and elevated intracellular calcium leads to hyperactive platelets.Elevated platelet calcium has been documented in hypertension and diabetes;both conditions increase the likelihood of heart attack and stroke. Thus,proper regulation of calcium metabolism in the platelet is extremely important.Plasma membrane Ca2+-ATPase(PMCA)is a major player in platelet calcium metabolism since it provides the only significant route for calcium efflux.In keeping with the important role of calcium in platelet function,PMCA is a highly regulated transporter.In human platelets,PMCA is activated by Ca2+/calmodulin,by cAMP-dependent phosphorylation and by calpain-dependent removal of the inhibitory peptide.It is inhibited by tyrosine phosphorylation and calpain-dependent proteolysis.In addition,the cellular location of PMCA is regulated by a PDZ-domain-dependent interaction with the cytoskeleton during platelet activation.Rapid regulation by phosphorylation results in changes in the rate of platelet activation,whereas calpain-dependent proteolysis and interaction with the cytoskeleton appears to regulate later events such as clot retraction.In hypertension and diabetes,PMCA expression is upregulated while activity is decreased, presumably due to tyrosine phosphorylation.Clearly,a more complete understanding of PMCA function in human platelets could result in the identification of new ways to control platelet function in disease states.

  13. Cellular Pathophysiology of an Adrenal Adenoma-Associated Mutant of the Plasma Membrane Ca(2+)-ATPase ATP2B3.

    Science.gov (United States)

    Tauber, Philipp; Aichinger, B; Christ, C; Stindl, J; Rhayem, Y; Beuschlein, F; Warth, R; Bandulik, S

    2016-06-01

    Adrenal aldosterone-producing adenomas (APAs) are a main cause for primary aldosteronism leading to arterial hypertension. Physiologically, aldosterone production in the adrenal gland is stimulated by angiotensin II and high extracellular potassium. These stimuli lead to a depolarization of the plasma membrane and, as a consequence, an increase of intracellular Ca(2+). Mutations of the plasma membrane Ca(2+)-ATPase ATP2B3 have been found in APAs with a prevalence of 0.6%-3.1%. Here, we investigated the effects of the APA-associated ATP2B3(Leu425_Val426del) mutation in adrenocortical NCI-H295R and human embryonic kidney (HEK-293) cells. Ca(2+) measurements revealed a higher basal Ca(2+) level in cells expressing the mutant ATP2B3. This rise in intracellular Ca(2+) was even more pronounced under conditions with high extracellular Ca(2+) pointing to an increased Ca(2+) influx associated with the mutated protein. Furthermore, cells with the mutant ATP2B3 appeared to have a reduced capacity to export Ca(2+) suggesting a loss of the physiological pump function. Surprisingly, expression of the mutant ATP2B3 caused a Na(+)-dependent inward current that strongly depolarized the plasma membrane and compromised the cytosolic cation composition. In parallel to these findings, mRNA expression of the cytochrome P450, family 11, subfamily B, polypeptide 2 (aldosterone synthase) was substantially increased and aldosterone production was enhanced in cells overexpressing mutant ATP2B3. In summary, the APA-associated ATP2B3(Leu425_Val426del) mutant promotes aldosterone production by at least 2 different mechanisms: 1) a reduced Ca(2+) export due to the loss of the physiological pump function; and 2) an increased Ca(2+) influx due to opening of depolarization-activated Ca(2+) channels as well as a possible Ca(2+) leak through the mutated pump. PMID:27035656

  14. Response of Ca2+-ATPase to clinorotaion of pea seedlings. O. M. Nedukha and E. L. Kordyum

    Science.gov (United States)

    Nedukha, Olena

    2016-07-01

    The present study was aimed to reveal of response of Ca2+-ATPase activity of cortex cells in distal elongation zone of Pisum sativum root to slow clinorotation. Pea seedlings were grown on a horizontal clinostat (2 rpm) and in the stationary control for 6 days. The electron-cytochemical method was used to examine the effects of imitated microgravity on the distribution of Ca2+-ATPase in outer layers of root cortex. The quantitative analysis of the density of cytochemical reaction products was measured using the Image J program. Electron microscopy showed the presence of electron-dense lead phosphate precipitated grains, the enzymatic activity reaction products on the plasma membrane, membranes of vesicular structures, endoplasmic reticulum (ER) and on organelles envelope in both of samples of the stationary control and clinorotated seedlings. We revealed the sensitivity of Ca2+-ATPase to clinorotation. The quantitative analysis of the area and density of enzymatic activity reaction products revealed that clinorotation led to the decrease of 3.4 times the density of reaction products on the plasma membrane and the increase of reaction products density on endomembranes and organelles membranes, in particular: in 2.2 times on mitochondria membranes; in 1.3 times - on membranes of ER; in 2.5 times - on tonoplast; by an order of magnitude greater - on contacting membranes of organelles with plasma membrane in comparison with such in cells of control samples. The data analysis can indicate an intensification of calcium pump on endomembranes, on envelopes of cytoplasmic organelles and nucleus. The obtained data suggest that the redistribution of Ca2+-ATPase activity in cells can be mediated by the activation of certain isoforms of enzyme or/and by an activation of Ca2+/H+ antiporter in plasma membrane that helps to maintain optimal calcium balance in plant cells under imitated microgravity.

  15. Polyamines cause plasma membrane depolarization, activate Ca2+-, and modulate H+-ATPase pump activity in pea roots.

    Science.gov (United States)

    Pottosin, Igor; Velarde-Buendía, Ana María; Bose, Jayakumar; Fuglsang, Anja T; Shabala, Sergey

    2014-06-01

    Polyamines regulate a variety of cation and K(+) channels, but their potential effects on cation-transporting ATPases are underexplored. In this work, noninvasive microelectrode ion flux estimation and conventional microelectrode techniques were applied to study the effects of polyamines on Ca(2+) and H(+) transport and membrane potential in pea roots. Externally applied spermine or putrescine (1mM) equally activated eosin yellow (EY)-sensitive Ca(2+) pumping across the root epidermis and caused net H(+) influx or efflux. Proton influx induced by spermine was suppressed by EY, supporting the mechanism in which Ca(2+) pump imports 2 H(+) per each exported Ca(2+). Suppression of the Ca(2+) pump by EY diminished putrescine-induced net H(+) efflux instead of increasing it. Thus, activities of Ca(2+) and H(+) pumps were coupled, likely due to the H(+)-pump inhibition by intracellular Ca(2+). Additionally, spermine but not putrescine caused a direct inhibition of H(+) pumping in isolated plasma membrane vesicles. Spermine, spermidine, and putrescine (1mM) induced membrane depolarization by 70, 50, and 35 mV, respectively. Spermine-induced depolarization was abolished by cation transport blocker Gd(3+), was insensitive to anion channels' blocker niflumate, and was dependent on external Ca(2+). Further analysis showed that uptake of polyamines but not polyamine-induced cationic (K(+)+Ca(2+)+H(+)) fluxes were a main cause of membrane depolarization. Polyamine increase is a common component of plant stress responses. Activation of Ca(2+) efflux by polyamines and contrasting effects of polyamines on net H(+) fluxes and membrane potential can contribute to Ca(2+) signalling and modulate a variety of transport processes across the plasma membrane under stress. PMID:24723394

  16. Reproductive organ and vascular specific promoter of the rice plasma membrane Ca2+ATPase mediates environmental stress responses in plants.

    Directory of Open Access Journals (Sweden)

    Kazi Md Kamrul Huda

    Full Text Available BACKGROUND: Plasma membrane Ca(2+ATPase is a transport protein in the plasma membrane of cells and helps in removal of calcium (Ca(2+ from the cell, hence regulating Ca(2+ level within cells. Though plant Ca(2+ATPases have been shown to be involved in plant stress responses but their promoter regions have not been well studied. RESULTS: The 1478 bp promoter sequence of rice plasma membrane Ca(2+ATPase contains cis-acting elements responsive to stresses and plant hormones. To identify the functional region, serial deletions of the promoter were fused with the GUS sequence and four constructs were obtained. These were differentially activated under NaCl, PEG cold, methyl viologen, abscisic acid and methyl jasmonate treatments. We demonstrated that the rice plasma membrane Ca(2+ATPase promoter is responsible for vascular-specific and multiple stress-inducible gene expression. Only full-length promoter showed specific GUS expression under stress conditions in floral parts. High GUS activity was observed in roots with all the promoter constructs. The -1478 to -886 bp flanking region responded well upon treatment with salt and drought. Only the full-length promoter presented cold-induced GUS expression in leaves, while in shoots slight expression was observed for -1210 and -886 bp flanking region. The -1210 bp deletion significantly responded to exogenous methyl viologen and abscisic acid induction. The -1210 and -886 bp flanking region resulted in increased GUS activity in leaves under methyl jasmonate treatments, whereas in shoots the -886 bp and -519 bp deletion gave higher expression. Salicylic acid failed to induce GUS activities in leaves for all the constructs. CONCLUSIONS: The rice plasma membrane Ca(2+ATPase promoter is a reproductive organ-specific as well as vascular-specific. This promoter contains drought, salt, cold, methyl viologen, abscisic acid and methyl jasmonate related cis-elements, which regulated gene expression. Overall, the

  17. 中医不同治法对骨质疏松症大鼠骨密度及骨骼肌Ca2+-Mg2+-ATP酶影响的比较研究%Effect of varied traditional Chinese therapies on bone mineral density and Ca2+-Mg2+-ATP enzyme in the skeletal muscle of glucocorticoid-induced osteoporosis in rats

    Institute of Scientific and Technical Information of China (English)

    杨芳; 郑洪新; 王剑; 朱辉; 张国哲

    2011-01-01

    Objective To observe the effect of the different treating methods with traditional Chinese medicine (TCM) on bone mineral density (BMD) and Ca2+ -Mg2+-ATP enzyme in the skeletal muscle in rats with glucocorticoid-induced osteoporosis and to discuss the mechanism of TCM on prevention and treatment of osteoporosis.Methods One hundred and twenty rats with half of each sex were randomly divided into 6 groups, including normal control group, model control group, reinforcing the kidney herb group, strengthening the spleen herb group, promoting blood circulation to remove stasis herb group, and GUSUKANG herb group.Intramuscular injection of dexamethasone was use to establish the model.At the end of the experiment, the rats were killed by withdrawing blood from the abdominal aorta.Ca2+ -Mg2+ -ATP enzyme was measured using enzyme-linked immunosorbent assay and BMD of the upper 1/3 of rat femur was detected using dual energy X-ray absorptiometry.Results ①Compared to normal control group, BMD of upper 1/3 of the femur in model control group decreased significantly (P < 0.01 ).Compared to model control group, BMD of upper 1/3 of the femur increased in all treatment groups at different degrade, with the most significant increase in reinforcing the kidney herb group ( P < 0.01 ).The BMD difference between model control group and the other treatment groups had no statistical significance.②Compared to normalcontrol group, Ca2+ -Mg2+ -ATP enzyme in the skeletal muscle in all other groups decreased significantly (P < 0.01 ).Compared to model control group, Ca2+-Mg2+ -ATP enzyme in the skeletal muscle of all treatment groups increased significantly ( P < 0.01 ).Ca2+ -Mg2+ -ATP enzyme in the skeletal muscle in reinforcing the kidney herb group increased the most significantly, compared to GUSUKANG group, promoting blood circulation group, and strengthening the spleen group with statistical significance (P < 0.01 ).The increase of Ca2+-Mg2+-ATP enzyme in the skeletal

  18. Engineering a prototypic P-type ATPase Listeria Monocytogenes Ca(2+)-ATPase 1 for single-molecule FRET studies

    DEFF Research Database (Denmark)

    Dyla, Mateusz; Andersen, Jacob; Kjaergaard, Magnus;

    2016-01-01

    Approximately 30% of the ATP generated in the living cell is utilized by P-type ATPase primary active transporters to generate and maintain electrochemical gradients across biological membranes. P-type ATPases undergo large conformational changes during their functional cycle to couple ATP hydrol...

  19. Adsorption of acetanilide herbicides on soil and its components II. Adsorption and catalytic hydrolysis of diethatyl-ethyl on saturated Na+-, K+-, Ca2+-, and Mg2+-montmorillonite

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Adsorption and catalytic hydrolysis of the herbicidediethatyl-ethyl [N-chloroacetyl-N-(2,6-diethylphenyl)glycine ethyl ester] on homoionic Na+-, K+-, Ca2+-, and Mg2+-montmorillonite clays were investigated in water solution. The Freundlich adsorption coefficient, Kf, got from isotherms on clay followed the order of Na+ K+ > Mg2+ Ca2+. Analysis of FT-IR spectra of diethatyl-ethyl adsorbed on clay suggests probable bonding at the carboxyl and amide carbonyl groups of the herbicide. The rate of herbicide hydrolysis in homoionic clay suspensions followed the same order as that for adsorption, indicating that adsorption may have preceded and thus caused hydrolysis. Preliminary product identification showed that hydrolysis occurred via nucleophilic substitution at the carboxyl carbon, causing the cleavage of the ester bond and formation of diethatyl and its dechlorinated derivative, and at the amide carbon, yielding an ethyl ester derivative and its acid.These pathways also suggest that hydrolysis of diethatyl-ethyl was catalyzed by adsorption on the clay surface.

  20. Mechanism of fluorescence and conformational changes of the sarcoplasmic calcium binding protein of the sand worm Nereis diversicolor upon Ca2+ or Mg2+ binding.

    Science.gov (United States)

    Sillen, Alain; Verheyden, Stefan; Delfosse, Lotte; Braem, Tania; Robben, Johan; Volckaert, Guido; Engelborghs, Yves

    2003-09-01

    The calcium-binding protein isolated from the sarcoplasm of the muscles of the sand worm Nereis diversicolor has four EF-hands and three active binding sites for Ca(2+) or Mg(2+). Nereis diversicolor sarcoplasmic calcium-binding protein contains three tryptophan residues at positions 4, 57, and 170, respectively. The Wt protein shows a very limited fluorescence increase upon binding of Ca(2+) or Mg(2+). Single-tryptophan-containing mutants were produced and purified. The fluorescence titrations of these mutants show a limited decrease of the affinity for calcium, but no alterations of the cooperativity. Upon adding calcium, Trp170 shows a strong fluorescence increase, Trp57 an extensive fluorescence decrease, and Trp4 shows no fluorescence change. Therefore mutant W4F/W170F is ideally suited to analyze the fluorescence titrations and to study the binding mechanism. Mutations of the calcium ligands at the z-position in the three binding sites show no effect at site I and a total loss of cooperativity at sites III and IV. The quenching of Trp57 upon calcium binding is dependent on the presence of arginine R25, but this residue is not just a simple dynamic quencher. The role of the salt bridge R25-D58 is also investigated. PMID:12944301

  1. Effect of ionizing radiation on catalytic properties of Ca2+-ATP-ase from sarcoplasmic reticulum of skeletal muscle

    International Nuclear Information System (INIS)

    It was studied kinetic and thermodynamic characteristics of Ca2+-ATP-ase of rat skeletal muscle (membranes of sarcoplasmic reticulum) after irradiation in doses 0,5, 4,0 and 8,0 Gy. It was shown that external gamma-irradiation at different doses changed kinetic and thermodynamic characteristics of the enzyme of sarcoplasmic reticulum membranes of skeletal muscle. These alterations probably correlate with disbalance of hormonal regulation of intracellular calcium metabolism and changes in membrane structure and functions

  2. Effect of ionizing radiation on catalytic properties of Ca2+-ATPase from sarcoplasmic reticulum of skeletal muscle

    International Nuclear Information System (INIS)

    It was studied kinetic and thermodynamic characteristics of Ca2+-ATPase of rat skeletal muscle (membranes of sarcoplasmic reticulum) after irradiation in doses 0,5, 4,0 and 8,0 Gy. It was shown that external gamma-irradiation at different doses changed kinetic and thermodynamic characteristics of the enzyme of sarcoplasmic reticulum membranes of skeletal muscle. These alterations probably correlate with dis balance of hormonal regulation of intracellular calcium metabolism and changes in membrane structure and functions

  3. Affinity of Smectite and Divalent Metal Ions (Mg(2+), Ca(2+), Cu(2+)) with L-leucine: An Experimental and Theoretical Approach Relevant to Astrobiology.

    Science.gov (United States)

    Pandey, Pramod; Pant, Chandra Kala; Gururani, Kavita; Arora, Priyanka; Pandey, Neetu; Bhatt, Preeti; Sharma, Yogesh; Negi, Jagmohan Singh; Mehata, Mohan Singh

    2015-12-01

    Earth is the only known planet bestowed with life. Several attempts have been made to explore the pathways of the origin of life on planet Earth. The search for the chemistry which gave rise to life has given answers related to the formation of biomonomers, and their adsorption on solid surfaces has gained much attention for the catalysis and stabilization processes related to the abiotic chemical evolution of the complex molecules of life. In this communication, surface interactions of L-leucine (Leu) on smectite (SMT) group of clay (viz. bentonite and montmorillonite) and their divalent metal ion (Mg(2+), Ca(2+) and Cu(2+)) incorporated on SMT has been studied to find the optimal conditions of time, pH, and concentration at ambient temperature (298 K). The progress of adsorption was followed spectrophotometrically and further characterized by FTIR, SEM/EDS and XRD. Leu, a neutral/non polar amino acid, was found to have more affinity in its zwitterionic form towards Cu(2+)- exchanged SMT and minimal affinity for Mg(2+)- exchanged SMT. The vibrational frequency shifts of -NH3 (+) and -COO(-) favor Van der Waal's forces during the course of surface interaction. Quantum calculations using density functional theory (DFT) have been applied to investigate the absolute value of metal ion affinities of Leu (Leu-M(2+) complex, M = Mg(2+), Ca(2+), Cu(2+)) with the help of their physico-chemical parameters. The hydration effect on the relative stability and geometry of the individual species of Leu-M(2+) × (H2O)n, (n =2 and 4) has also been evaluated within the supermolecule approach. Evidence gathered from investigations of surface interactions, divalent metal ions affinities and hydration effects with biomolecules may be important for better understanding of chemical evolution, the stabilization of biomolecules on solid surfaces and biomolecular-metal interactions. These results may have implications for understanding the origin of life and the preservation of

  4. Affinity of Smectite and Divalent Metal Ions (Mg2+, Ca2+, Cu2+) with L-leucine: An Experimental and Theoretical Approach Relevant to Astrobiology

    Science.gov (United States)

    Pandey, Pramod; Pant, Chandra Kala; Gururani, Kavita; Arora, Priyanka; Pandey, Neetu; Bhatt, Preeti; Sharma, Yogesh; Negi, Jagmohan Singh; Mehata, Mohan Singh

    2015-12-01

    Earth is the only known planet bestowed with life. Several attempts have been made to explore the pathways of the origin of life on planet Earth. The search for the chemistry which gave rise to life has given answers related to the formation of biomonomers, and their adsorption on solid surfaces has gained much attention for the catalysis and stabilization processes related to the abiotic chemical evolution of the complex molecules of life. In this communication, surface interactions of L-leucine (Leu) on smectite (SMT) group of clay (viz. bentonite and montmorillonite) and their divalent metal ion (Mg2+, Ca2+ and Cu2+) incorporated on SMT has been studied to find the optimal conditions of time, pH, and concentration at ambient temperature (298 K). The progress of adsorption was followed spectrophotometrically and further characterized by FTIR, SEM/EDS and XRD. Leu, a neutral/non polar amino acid, was found to have more affinity in its zwitterionic form towards Cu2+- exchanged SMT and minimal affinity for Mg2+- exchanged SMT. The vibrational frequency shifts of —NH3 + and —COO- favor Van der Waal's forces during the course of surface interaction. Quantum calculations using density functional theory (DFT) have been applied to investigate the absolute value of metal ion affinities of Leu (Leu—M2+ complex, M = Mg2+, Ca2+, Cu2+) with the help of their physico-chemical parameters. The hydration effect on the relative stability and geometry of the individual species of Leu—M2+ × (H2O)n, ( n =2 and 4) has also been evaluated within the supermolecule approach. Evidence gathered from investigations of surface interactions, divalent metal ions affinities and hydration effects with biomolecules may be important for better understanding of chemical evolution, the stabilization of biomolecules on solid surfaces and biomolecular-metal interactions. These results may have implications for understanding the origin of life and the preservation of biomarkers.

  5. P160L mutation in the Ca(2+) ATPase 2A domain in a patient with severe Darier disease.

    Science.gov (United States)

    Godic, Aleksandar; Glavac, Damjan; Korosec, Branka; Miljković, Jovan; Potocnik, Marko; Kansky, Aleksej

    2004-01-01

    Darier disease (DD) is caused by mutations of the ATP2A2 gene, which encodes the sarco/endoplasmic reticulum Ca(2+)-ATPase isoform 2 (SERCA2). The mutations affect protein expression, degradation and activity. We report a patient with severe sporadic DD, who did not respond adequately to repeated courses of orally administered acitretin and isotretinoin. He was found to harbor the missense P160L mutation of the ATP2A2 gene in a heterozygous state in the A domain of SERCA2 and polymorphism in intron 18 (2741 + 54 G --> A). The A domain plays a key role in translocation of Ca(2+) from cytoplasm to endoplasmic reticulum lumen, thus establishing a low intracellular Ca(2+) concentration.

  6. Effects of dissolved Ca2+, Mg2+, and Na+ ions on the supramolecular aggregation of natural organic matter in aqueous solutions

    Science.gov (United States)

    Ahn, W.; Kalinichev, A. G.; Clark, M. M.

    2008-12-01

    The complexation of natural organic matter (NOM) with metal ions, minerals and organic species in soil and water allows NOM to form water-soluble and water-insoluble aggregates of widely differing chemical and biological stabilities. Metal-NOM interaction induces strong correlations between the concentration of natural organic matter and the speciation, solubility and toxicity of many metals in the environment. In water purification and desalination, NOM is also implicated in fouling of nanofiltration and reverse osmosis membranes, either as the primary foulant or as a conditioning layer for microbial attachment ("biofouling"). In this work we investigated the effects of various metal ions on NOM aggregation in aqueous solutions, by a combination of dynamic light scattering (DLS), small angle neutron scattering (SANS) and large-scale molecular dynamics (MD) computer simulations. This allows a detailed molecular-scale statistical analysis of the size and the structural topology of metal-NOM aggregates. The DLS measurements show that Ca2+ ions present in a Suwannee River NOM (SRNOM) solution lead to the formation of a wide range of supramolecular structures with sizes between 100 and 1,000 nm. In contrast, Mg2+ and Na+ do not affect the aggregation of SRNOM as strongly. SANS data are inconclusive but indicate the presence of quite large (>50 nm) fractal particles formed presumably through a cluster-cluster aggregation. MD simulations confirm these observations and show that NOM can aggregate in aqueous solutions by two different mechanisms. On the one hand, NOM molecules can spontaneously aggregate by hydrogen bonding between their functional groups when only Na+ and Mg2+ are present as background cations. This promotes the formation of uniformly shaped NOM clusters. On the other hand, if Ca2+ ions are present in solution, they can more strongly bind two different NOM molecules by co-complexing the carboxylate groups, thus promoting the formation of longer linear and

  7. Aluminum resistance in wheat involves maintenance of leaf Ca(2+) and Mg(2+) content, decreased lipid peroxidation and Al accumulation, and low photosystem II excitation pressure.

    Science.gov (United States)

    Moustaka, Julietta; Ouzounidou, Georgia; Bayçu, Gülriz; Moustakas, Michael

    2016-08-01

    The phytotoxic aluminum species (Al(3+)) is considered as the primary factor limiting crop productivity in over 40 % of world's arable land that is acidic. We evaluated the responses of two wheat cultivars (Triticum aestivum L.) with differential Al resistance, cv. Yecora E (Al-resistant) and cv. Dio (Al-sensitive), exposed to 0, 37, 74 and 148 μM Al for 14 days in hydroponic culture at pH 4.5. With increasing Al concentration, leaf Ca(2+) and Mg(2+) content decreased, as well as the effective quantum yield of photosystem II (PSII) photochemistry (Φ PSII ), while a gradual increase in leaf membrane lipid peroxidation, Al accumulation, photoinhibition (estimated as F v /F m ), and PSII excitation pressure (1 - q p ) occurred. However, the Al-resistant cultivar with lower Al accumulation, retained larger concentrations of Ca(2+) and Mg(2+) in the leaves and kept a larger fraction of the PSII reaction centres (RCs) in an open configuration, i.e. a higher ratio of oxidized to reduced quinone A (QA), than plants of the Al-sensitive cultivar. Four times higher Al concentration in the nutrient solution was required for Al-resistant plants (148 μM Al) than for Al-sensitive (37 μM Al), in order to establish the same closed RCs. Yet, the decline in photosynthetic efficiency in the cultivar Dio was not only due to closure of PSII RCs but also to a decrease in the quantum yield of the open RCs. We suggest that Al(3+) toxicity may be mediated by nutrient deficiency and oxidative stress, and that Al-resistance of the wheat cultivar Yecora E, may be due at least partially, from the decreased Al accumulation that resulted to decreased reactive oxygen species (ROS) formation. However, under equal internal Al accumulation (exposure Al concentration: Dio 74 μM, Yecora E 148 μM) that resulted to the same oxidative stress, the reduced PSII excitation pressure and the better PSII functioning of the Al-resistant cultivar was probably due to the larger concentrations of Ca

  8. Density functional investigation of metal encapsulated X-C12Si8 heterofullerene (X=Li+, Na+, K+, Be2+, Mg2+, Ca2+, Al3+, Ga3+)

    International Nuclear Information System (INIS)

    The stability and the possible application of our recently reported SiC heterofullerenes inspire the investigation of their further stabilization through ion encapsulation. The endohedral complexes X-C12Si8, where X=Li+, Na+, K+, Be2+, Mg2+, Ca2+, Al3+, and Ga3+, are probed at the MPWB1K/6-311G* and B3LYP/6-311G* levels of theory. The optimized geometries show the expanding or contracting capability of C12Si8 in order to accommodate metal ion guests. The inclusion energies indicate the stability of the complexes compared to the components. Meanwhile, the calculated binding energies show the stabilization of C12Si8 through the inclusion of Be2+, Mg2+, Al3+, and Ga3+. The host-guest interaction that is probed through NBO atomic charges supports the obtained results. This study refers to 'metal ion encapsulation' as a strategy for stabilization of SiC heterofullerenes.

  9. Affinity of Smectite and Divalent Metal Ions (Mg(2+), Ca(2+), Cu(2+)) with L-leucine: An Experimental and Theoretical Approach Relevant to Astrobiology.

    Science.gov (United States)

    Pandey, Pramod; Pant, Chandra Kala; Gururani, Kavita; Arora, Priyanka; Pandey, Neetu; Bhatt, Preeti; Sharma, Yogesh; Negi, Jagmohan Singh; Mehata, Mohan Singh

    2015-12-01

    Earth is the only known planet bestowed with life. Several attempts have been made to explore the pathways of the origin of life on planet Earth. The search for the chemistry which gave rise to life has given answers related to the formation of biomonomers, and their adsorption on solid surfaces has gained much attention for the catalysis and stabilization processes related to the abiotic chemical evolution of the complex molecules of life. In this communication, surface interactions of L-leucine (Leu) on smectite (SMT) group of clay (viz. bentonite and montmorillonite) and their divalent metal ion (Mg(2+), Ca(2+) and Cu(2+)) incorporated on SMT has been studied to find the optimal conditions of time, pH, and concentration at ambient temperature (298 K). The progress of adsorption was followed spectrophotometrically and further characterized by FTIR, SEM/EDS and XRD. Leu, a neutral/non polar amino acid, was found to have more affinity in its zwitterionic form towards Cu(2+)- exchanged SMT and minimal affinity for Mg(2+)- exchanged SMT. The vibrational frequency shifts of -NH3 (+) and -COO(-) favor Van der Waal's forces during the course of surface interaction. Quantum calculations using density functional theory (DFT) have been applied to investigate the absolute value of metal ion affinities of Leu (Leu-M(2+) complex, M = Mg(2+), Ca(2+), Cu(2+)) with the help of their physico-chemical parameters. The hydration effect on the relative stability and geometry of the individual species of Leu-M(2+) × (H2O)n, (n =2 and 4) has also been evaluated within the supermolecule approach. Evidence gathered from investigations of surface interactions, divalent metal ions affinities and hydration effects with biomolecules may be important for better understanding of chemical evolution, the stabilization of biomolecules on solid surfaces and biomolecular-metal interactions. These results may have implications for understanding the origin of life and the preservation of

  10. Dynamic Changes of Ca~(2+) and Ca~(2+)-ATPase in Sieve Elements in the Developing Caryopsis of Triticum aestivum L.%Ca~(2+)和Ca~(2+)-ATPase在小麦颖果筛分子分化中的动态变化

    Institute of Scientific and Technical Information of China (English)

    李继伟; 邓祥宜; 周竹青; 王利凯; 阳超男; 樊海燕

    2009-01-01

    [Objective] Previous study revealed that sieve elements (SEs) in the developing caryopsis of Triticum aestivum L.underwent a unique type of programmed cell death (PCD).In this paper,the dynamic changes and the roles of Ca~(2+) and Ca~(2+)-ATPase in SEs during the PCD were studied.[Method] The ultrastructural aspects of phloem cells in wheat caryopsis were examined by transmission electron microscopy (TEM).Using specific fluorescence staining and potassium pyroantimonate precipitation method,Ca~(2+) was localized at histological and sub-cellular levels in SEs in the developing wheat caryopsis.TEM and lead nitrate were used to locate Ca~(2+)-ATPase in SEs.[Result] TEM studies showed that the cell walls of SEs thickened at the beginning of differentiation,and then became thinner and smoother.Fluorescence staining showed that the fluorescence due to Ca~(2+) appeared in cell walls of SEs from 6 to 10 d after flowering.The fluorescence due to Ca~(2+) in cell walls of SEs was most notable on 9 d after flowering and disappeared on 14 d after flowering.Sub-celluar localization of Ca~(2+) showed that Ca~(2+) was localized on plasma membrane and in nuclei from 1 to 2 d after flowering.On 4 d after flowering,Ca2+ was localized in cytoplasm and mitochondria of SEs.From 5 to 8 d after flowering,Ca~(2+) was transported to the cell walls of SEs and no Ca~(2+) precipitates were observed in mitochondria.From 10 to 18 d after flowering,Ca~(2+) was transported into the cytoplasm again from cell walls and no Ca~(2+) precipitates were observed on 20 d after flowering.In intermediary cells (ICs),Ca~(2+) precipitates were observed from 1 to 18 d after flowering,and Ca~(2+) mainly distributed on intine and tonoplast.The activity of Ca~(2+)-ATPase changed obviously during the SEs differentiation.There was lowest activity of Ca~(2+)-ATPase on 3 d after flowering in SEs.High levels of Ca~(2+)-ATPase activity were found from 4 to 14 d after flowering in SEs,and the enzyme was mainly localized

  11. Effects of Ca2+, Mg2+ of Foliar Application and Combined with SA on Resistance to Botrytis cinerea in Tomato Seedlings%叶面喷施Ca2+、Mg2+、SA及其组合对番茄抗灰霉病的影响

    Institute of Scientific and Technical Information of China (English)

    李琳琳; 潘晓爱; 郭秋城; 易知利

    2016-01-01

    矿质元素不仅可使植物旺盛、健壮生长,而且多数元素可作为病原物营养需要或毒害作用而影响病原物的侵染扩散和繁殖,增强植物抗病能力. 选用番茄灰霉病敏感型番茄 L402 为试材,叶片施用Ca2+、Mg2+、SA及其组合处理,对番茄五叶幼苗期抗灰霉病的效果进行调查. 结果表明:叶面增施Mg2+不显著改变番茄灰霉病的病情指数,病情指数为78. 25;Mg2+与SA不同顺序复合施用的病情性指数虽低于对照,但均显著高于SA单一处理;Mg2+不仅没有提高番茄抗灰霉病的作用,也没有提高SA诱导番茄抗灰霉病的作用;Ca2+与SA不同顺序配施对番茄幼苗抗灰霉病的影响效果不同. 明确了先施Ca2+再施用SA处理抗病效果最好. 即Ca+SA处理的病情指数比SA降低17. 93 %,比SA+Ca降低13. 34 %,比单纯施Ca2+降低45. 22 %;而SA+Ca处理的病情指数与SA处理无显著差异. 说明先施Ca2+再施SA, Ca2+具有显著增强SA诱导番茄抗灰霉病的作用. 实验结果将有助于了解和提高SA诱导抗性机制,为提高番茄产量和品种提供理论基础和现实依据.%Mineral elements as importance nutrition substance can make the plants grow strong and vigoroust, meanwhile, most elements can be used as nutritional or toxic substance of patho-gens to affect its spread, infection and reproduction, in order to enhance plant disease resist-ance. The cultivated tomato 'L402', which was sensitive to Botrytis cinerea, was employed in this experiment. In the experimet, the treatments of with Ca2+, Mg2+, SA and combinations was performed. The treatments effect on resistance on Botrytis cinerea inoculation in tomato five-leaves-stage seedlings were surveied. It was clear that the disease index was 78. 25, which was not significantly changed by foliar applying Mg2+ compared with CK. The disease index of Mg2+ application combined SA were lower than control, but significantly higher than SA single treatment. The results

  12. Pycnogenol® and Ginkgo biloba extract: effect on peroxynitrite-oxidized sarcoplasmic reticulum Ca2+-ATPase

    OpenAIRE

    Žižková, Petronela; Viskupičová, Jana; Horáková, L'ubica

    2010-01-01

    The effect of two natural standardized plant extracts, Pycnogenol® and EGb 761, on sarcoplasmic reticulum Ca2+-ATPase (SERCA) activity and posttranslational modifications induced by peroxynitrite was investigated to assess their possible protective role. EGb 761 was found to have a protective effect on SERCA activity in the concentration range of 5–40 µg/ml. On the other hand, Pycnogenol® caused a decrease of SERCA activity at concentrations of 25 µg/ml. EGb 761 did not prevent protein carbon...

  13. 女贞子提取物对大鼠不同组织Ca2+-ATPase活性的影响%Effects of Ligustrum lucidum Extracts on Ca2+-ATPase Activity in Different Tissues of Rat

    Institute of Scientific and Technical Information of China (English)

    马云慧; 熊正英

    2012-01-01

    研究了女贞子提取物对大强度耐力训练大鼠不同组织Ca2+ -ATPase活性的影响,探讨了女贞子提取物对大鼠运动能力的作用机制.结果表明:运动组和运动+女贞子组大鼠各组织Ca2+-ATPasee活性均显著低于安静组,运动+女贞子组大鼠不同组织Ca2+-ATPase活性显著高于运动组;运动+女贞子组大鼠力竭运动时间比运动组延长23.09%.女贞子提取物可以调节大鼠不同组织Ca2+ -ATPase活性,延长运动至疲劳的时间.%To study the mechanism of Ligustrum lucidum extract on the exercise performance of rat through examining the effects of Ligustrum lucidum extracts on Ca2+ -ATPase activity in different tissue of rats in endurance training. The results showed that the Ca2+ -ATPase activity in the rat tissue of exercise control group and exercise+ extract feeding group was significantly lower than that of the sedentary control group (/> increase Ca2+-ATPase activity, and extend the time from exercise to fatigue.

  14. Expression, purification, crystallization and preliminary X-ray analysis of calmodulin in complex with the regulatory domain of the plasma-membrane Ca2+-ATPase ACA8

    DEFF Research Database (Denmark)

    Tidow, Henning; Hein, Kim Langmach; Bækgaard, Lone;

    2010-01-01

    of calcium-bound calmodulin (Ca(2+)-CaM) to this tail and a conformational change that displaces the autoinhibitory tail from the catalytic domain. The complex between calmodulin and the regulatory domain of the plasma-membrane Ca(2+)-ATPase ACA8 from Arabidopsis thaliana has been crystallized. The......Plasma-membrane Ca(2+)-ATPases (PMCAs) are calcium pumps that expel Ca(2+) from eukaryotic cells to maintain overall Ca(2+) homoeostasis and to provide local control of intracellular Ca(2+) signalling. They are of major physiological importance, with different isoforms being essential, for example...... crystals belonged to space group C2, with unit-cell parameters a = 176.8, b = 70.0, c = 69.8 A, beta = 113.2 degrees. A complete data set was collected to 3.0 A resolution and structure determination is in progress in order to elucidate the mechanism of PMCA activation by calmodulin....

  15. Effect of Turmeric, Turmerin and Curcumin on Ca2+, Na/K+ Atpases in Concanavalin A-Stimulated Human Blood Mononuclear Cells

    Directory of Open Access Journals (Sweden)

    Suman K. Das

    2003-01-01

    Full Text Available Abstract: Ion transport enzymes may play an important role in T cell activation. This study investigates the role of turmeric and its individual components, turmerin-and curcumin-on Ca2+ and Na/K+ adenosine triphosphatases (ATPase in the course of T cell activation. Concanavalin A (Con A stimulated human blood mononuclear T cell proliferation paradigm was investigated for 3, 5 and 7 day periods with different concentrations of turmeric, curcumin and turmerin. Con A-stimulated cells treated with turmeric (250, 50, 5 μg/ml for 3 and 5 days inhibited ATPase levels when compared to base levels obtained by cells in media alone. At day 7, there was a 3-fold increase for Ca2+ATPase levels and a 2-fold increase for Na/K+ATPase. Curcumin (250, 50, 5 μg/ml showed the same pattern for ATPase activity as turmeric at 3 and 5 days with a 2-fold increase at day 7. Turmerin (2500, 1250, 250, 25 ng/ml for Na/K+ ATPase activity showed an increase at day 3, a decrease on day 5, and a 2-fold increase on day 7. Ca2+ ATPase activity in the presence of turmerin showed an increase in ATPase levels at day 3 (except at 2500ng/ml where it decreased and a decrease in day 5 (except at 25 ng/ml where it increased. Turmeric and curcumin generally inhibited Ca2+ATPase and Na/K+ATPases in early (day 3 and intermediate (day 5 stages of mitogen stimulation. However, the effect after 7 days incubation for turmeric, curcumin and turmerin showed a marked increase up to three fold.

  16. Mg2+-dependent ATPase activity in cardiac myofibrils from the insulin-resistant JCR:LA-cp rat.

    Science.gov (United States)

    Misra, T; Russell, J C; Clark, T A; Pierce, G N

    2001-01-01

    There is a great deal of information presently available documenting a cardiomyopathic condition in insulin-deficient models of diabetes. Less information is available documenting a similar status in non insulin-dependent models of diabetes. We have studied the functional integrity of the myofibrils isolated from hearts of JCR:LA rats. The JCR:LA rat is hyperinsulinemic, hyperlipidemic, glucose intolerant and obese. As such, it carries many of the characteristics found in humans with non insulin-dependent diabetes mellitus. These animals also have many indications of heart disease. However, it is not clear if the hearts suffer from vascular complications or are cardiomyopathic in nature. We examined Mg2+-dependent myofibrillar ATPase in hearts of JCR:LA-cp/cp rats and their corresponding control animals (+/?) and found no significant differences (P> 0.05). This is in striking contrast to the depression in this activity exhibited by cardiac myofibrils isolated from insulin-deficient models of diabetes. Our data demonstrate that myofibrillar functional integrity is normal in JCR:LA-cp rats and suggest that these hearts are not in a cardiomyopathic state. Insulin status may be critical in generating a cardiomyopathic condition in diabetes.

  17. Synthesis of a ruthenium(II) bipyridyl complex coordinated by a functionalized Schiff base ligand: Characterization, spectroscopic and isothermal titration calorimetry measurements of M 2+ binding and sensing (M 2+ = Ca 2+, Mg 2+)

    Science.gov (United States)

    Dixit, Namrata; Mishra, Lallan; Mustafi, Sourajit M.; Chary, Kandala V. R.; Houjou, Hirohiko

    2009-07-01

    Bis-[methylsalicylidine-4'benzoic acid]-ethylene (LH 2) complexed with cis-Ru(bpy) 2Cl 2·2H 2O provides a complex of composition [Ru(bpy) 2L]·2NH 4PF 6 ( 1), which has been characterized spectroscopically. Its binding behaviour towards Mg 2+ and Ca 2+ ions is monitored using 1H NMR titration, isothermal titration calorimetry (ITC) and luminescence microscopy. The luminescent ruthenium complex binds Ca 2+ in a more selective manner as compared to Mg 2+.

  18. Critical Roles of Hydrophobicity and Orientation of Side Chains for Inactivation of Sarcoplasmic Reticulum Ca2+-ATPase with Thapsigargin and Thapsigargin Analogs*

    OpenAIRE

    Winther, Anne-Marie L.; Liu, Huizhen; Sonntag, Yonathan; Olesen, Claus; le Maire, Marc; Soehoel, Helmer; Olsen, Carl-Erik; Christensen, S. Brøgger; Nissen, Poul; Møller, Jesper V.

    2010-01-01

    Thapsigargin (Tg), a specific inhibitor of sarco/endoplasmic Ca2+-ATPases (SERCA), binds with high affinity to the E2 conformation of these ATPases. SERCA inhibition leads to elevated calcium levels in the cytoplasm, which in turn induces apoptosis. We present x-ray crystallographic and intrinsic fluorescence data to show how Tg and chemical analogs of the compound with modified or removed side chains bind to isolated SERCA 1a membranes. This occurs by uptake via the membrane lipid followed b...

  19. 不同冷敏感型甘蔗茎尖Ca2+和Ca2+-ATP酶活性对低温的响应%Response of Ca2+ and Ca2+-ATPase activity in the stem tip of sugarcane to low temperature stress

    Institute of Scientific and Technical Information of China (English)

    李素丽; 杨丽涛; 李志刚; 李杨瑞; 韩春旺; 梁兆宙

    2011-01-01

    为了探明不同冷敏感型甘蔗品种茎尖Ca2+和Ca2+-ATP酶活性对低温胁迫的响应机制,本研究利用植物组织化学技术结合电子显微镜观察了2个不同冷敏感型甘蔗品种茎尖在低温胁迫前后Ca2+和Ca2+-ATP酶的变化规律.供试品种为桂糖28号(抗冷型)和园林6号(冷敏感型),在0℃条件下,分别处理0、2、4和6 d后切取茎尖进行组织化学定位,获得如下结果:1)低温胁迫前后桂糖28号形态和细胞结构变化不明显,但园林6号发生质壁分离现象,线粒体空泡化,细胞崩溃,组织水溃状明显;2)低温胁迫开始,2个甘蔗品种细胞质和细胞核Ca2+沉淀颗粒均增多,但随着低温胁迫时间的延长桂糖28号细胞质和细胞核Ca2+沉淀颗粒减少,并维持在一个低稳态水平,而园林6号细胞质和细胞核一直维持在高Ca2+浓度水平;3)低温对桂糖28号Ca2+-ATP酶活性与分布影响不大,其活性一直维持在较高水平,而园林6号Ca2+-ATP酶活性随着低温胁迫时间的延长而变弱.结果说明,低温条件下细胞质和细胞核Ca2+浓度增高是导致甘蔗茎尖细胞受伤害的重要原因,维持较高Ca2+-ATP酶活性有助于避免Ca2+-中毒.%Low temperature in winter is an adverse abjotic stress to sugarcane industry,and caused significant lose. In order to investigate the response of Ca2+ and Ca2+-ATPase activity in the stem tip of different cold sensitivity sugarcane cultivars, Electron microscope (EMS) and phytohistochemistry technology were employed to detect the changes of Ca2+ level and Ca2+ -ATPase activity in the stem tips of two sugarcane varieties YL6 (cold sensitive) and GT28 (cold resistant) before and after low temperature treatment. The plants were treated at 0 ℃ in fridge and samples were taken after treatment at 0,2,4 and 6 d, respectively. The results showed that there was no significant morphological difference and cellular structures in the stem tip of GT28 before and after low

  20. [Changes of sarcolemma Na+/K+ ATPase and sarcoplasmic reticulum membrane Ca2+ ATPase activity after stem cell transplantation in chronic heart failure].

    Science.gov (United States)

    Fan, Zhongcai; Chen, Mao; Deng, Juelin; Liu, Xiaojing; Zhang, Li; Rao, Li; Yang, Qing; Huang, Dejia

    2007-02-01

    To assess the changes of sarcolemma Na+/K+ ATPase (CMNKA) and sarcoplasmic reticulum membrane Ca2+ ATPase (SERCA) activities after stem cells transplantation in heart failure. Rabbit was used as heart failure model by intravenously injecting adriamycin. Autologous bone marrow mononuclear cells (BMCs), bone marrow mesenchymal stem cells (MSCs) or skeletal myoblasts (SMs) were introduced into coronary arteies through the root of aorta when two balloons occluding just above sinus of Valsalva. After 4 weeks, left ventricular ejection fraction (LVEF)was evaluated by echocardiography, and the activities of CMNKA and SERCA were measured by colorimeter. In BMCs (n=8)and MSCs (n=8) group, LVEF were significantly improved (P SMs group (n=6) compared to sham group (n=8). The CMNKA activity in all stem cells groups was significantly increased compared to sham group (P < 0.05). Meanwhile, in comparison with sham group, the incremental tendencies of SERCA activity were seen in stem cells groups. In conclusion, stem cells transplantation could increase the activities of CMNKA and SERCA in heart failure, a possible mechanism to improve heart function. PMID:17333908

  1. Modulation of sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase activity and oxidative modification during the development of adjuvant arthritis.

    Science.gov (United States)

    Strosova, Miriam K; Karlovska, Janka; Zizkova, Petronela; Kwolek-Mirek, Magdalena; Ponist, Silvester; Spickett, Corinne M; Horakova, Lubica

    2011-07-01

    Adjuvant arthritis (AA) was induced by intradermal administration of Mycobacterium butyricum to the tail of Lewis rats. In sarcoplasmic reticulum (SR) of skeletal muscles, we investigated the development of AA. SR Ca(2+)-ATPase (SERCA) activity decreased on day 21, suggesting possible conformational changes in the transmembrane part of the enzyme, especially at the site of the calcium binding transmembrane part. These events were associated with an increased level of protein carbonyls, a decrease in cysteine SH groups, and alterations in SR membrane fluidity. There was no alteration in the nucleotide binding site at any time point of AA, as detected by a FITC fluorescence marker. Some changes observed on day 21 appeared to be reversible, as indicated by SERCA activity, cysteine SH groups, SR membrane fluidity, protein carbonyl content and fluorescence of an NCD-4 marker specific for the calcium binding site. The reversibility may represent adaptive mechanisms of AA, induced by higher relative expression of SERCA, oxidation of cysteine, nitration of tyrosine and presence of acidic phospholipids such as phosphatidic acid. Nitric oxide may regulate cytoplasmic Ca(2+) level through conformational alterations of SERCA, and decreasing levels of calsequestrin in SR may also play regulatory role in SERCA activity and expression. PMID:21531199

  2. Modulation of sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase activity and oxidative modification during the development of adjuvant arthritis.

    Science.gov (United States)

    Strosova, Miriam K; Karlovska, Janka; Zizkova, Petronela; Kwolek-Mirek, Magdalena; Ponist, Silvester; Spickett, Corinne M; Horakova, Lubica

    2011-07-01

    Adjuvant arthritis (AA) was induced by intradermal administration of Mycobacterium butyricum to the tail of Lewis rats. In sarcoplasmic reticulum (SR) of skeletal muscles, we investigated the development of AA. SR Ca(2+)-ATPase (SERCA) activity decreased on day 21, suggesting possible conformational changes in the transmembrane part of the enzyme, especially at the site of the calcium binding transmembrane part. These events were associated with an increased level of protein carbonyls, a decrease in cysteine SH groups, and alterations in SR membrane fluidity. There was no alteration in the nucleotide binding site at any time point of AA, as detected by a FITC fluorescence marker. Some changes observed on day 21 appeared to be reversible, as indicated by SERCA activity, cysteine SH groups, SR membrane fluidity, protein carbonyl content and fluorescence of an NCD-4 marker specific for the calcium binding site. The reversibility may represent adaptive mechanisms of AA, induced by higher relative expression of SERCA, oxidation of cysteine, nitration of tyrosine and presence of acidic phospholipids such as phosphatidic acid. Nitric oxide may regulate cytoplasmic Ca(2+) level through conformational alterations of SERCA, and decreasing levels of calsequestrin in SR may also play regulatory role in SERCA activity and expression.

  3. The changes of cardioelectrical activity of rat with myocardial infarction receiving sarcoplasmic reticulum Ca2+-ATPase gene modified bone marrow stem cell transplantation by microelectrode array technology

    Institute of Scientific and Technical Information of China (English)

    范平

    2012-01-01

    Objective Therapy effects and cardiac electrical activity comparison of bone marrow stem cells (BMSCs) transplantation and sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) gene modified BMSCs transplantation after acute myocardial infarction(AMI) in rats.Methods Rats with AMI were divided

  4. Effects of fructose-1,6-diphosphate on concentration of calcium and activities of sarcoplosnic Ca2+-ATPase in cardiomyocytes of Adriamycin-treated rats

    Institute of Scientific and Technical Information of China (English)

    CAI Wei; CHEN Jun-zhu; RUAN Li-ming; WANG Yi-na

    2005-01-01

    Objective: To observe the effects of fructose-1,6-diphosphate (FDP) on serum levels of cardiac troponin I (cTnI) and creatine kinase-MB (CK-MB), as well as the concentration of calcium in cardiomyocytes (Myo[Ca2+]) and activity of sarcoplosnic Ca2+-ATPase (SRCa2+-ATPase) in Adriamycin (ADR)-treated rats. Methods: Rats were intraperitoneally injected with ADR (2.5mg/kg every other day for 6 times) and then with different dosages of FDP (every other day for twenty-one times). Bi-antibodies sandwich Enzyme linked immune absorption assay (ELISA) was performed to detect serum level of cTnI. CK-MB was detected by monoclonal antibody, Myo[Ca2+] was detected by fluorescent spectrophotometry and the activity of SRCa2+-ATPase was detected by inorganic phosphate method. Results: FDP (300, 600, 1200 mg/kg) significantly reduced the serum levels of cTnI and CK-MB, while at the same time decreased calcium concentration and increased SRCa2+-ATPase activity in cardiomyocytes of ADR-treated rats (P<0.01). Conclusions: FDP might alleviate the cardiotoxic effects induced by ADR through decreasing calcium level as well as increasing SRCa2+-ATPase activity in cardiomyocytes.

  5. Effects of Glycinin on the Activities of Ca2+-ATPase and Superoxide Dismutase in Small Intestine%大豆球蛋白对小肠Ca2+吸收相关酶Ca2+-ATP酶和超氧化物歧化酶活性的影响

    Institute of Scientific and Technical Information of China (English)

    李兴起; 汪秀志; 陈晶; 秦学功

    2013-01-01

    To evaluate the effects and mechanism of glycinin on Ca2+absorption in small intestine,the Ca2+concentrations of the inner liquid in the everted gut sacs were determined by automatic biochemistry analyzer. Then the homogenized small intestine was applied to evaluate the activities of Ca2+-ATPase and Superoxide Dismutase (SOD). The results showed that single usage of glycinin (0.4~1 mg·mL-1)to everted gut sacs did not significantly change the Ca2+ absorption,whereas with dose-dependent inhibitions of Ca2+-ATPase activities and enhancement of superoxide dismutase activities. But 1 mg·mL -1 glycinin could enhance the Ca2+absorptions with calcium gluconate addition to culture medium. Our work indicated that glycin inhibited Ca2 + absorption and transportation by inhibiting the activity of Ca2+-ATPase,and with feedback mode,the activity of SOD was enhanced to resist the damage of oxygen free radicals to intestinal mucosa.%为探讨大豆球蛋白对小肠内微量元素Ca2+吸收的影响及相关机制。采用自动生化分析仪测定小肠囊内液中Ca2+浓度,并测定小肠囊中段组织中与Ca2+吸收相关的Ca2+-ATP酶和超氧化物歧化酶活性。结果显示,单独应用大豆球蛋白(0.4~1 mg·mL-1)对小肠囊Ca2+吸收无明显影响,但可剂量依赖性抑制小肠粘膜内Ca2+-ATP酶活性,同时剂量依赖性的反馈性诱导增强小肠粘膜内超氧化物歧化酶活性。1 mg·mL-1大豆球蛋白通过抑制Ca2+-ATP酶活性抑制葡萄糖酸钙Ca2+吸收。综上表明,大豆球蛋白通过抑制Ca2+-ATP酶活性阻碍Ca2+的吸收及转运,并可能通过反馈性的增强SOD的活性以减少氧化自由基对粘膜的进一步损伤。

  6. Effects of combination of irbesartan and perindopril on calcineurin expression and sarcoplasmic reticulum Ca2+-ATPase activity in rat cardiac pressure-overload hypertrophy

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Aim: To observe effects of angiotensin (Ang) Ⅱ receptor antagonist (AT1) irbesartan and angiotensin-converting enzyme (ACE) inhibitor perindopril on rat myocardium calcineurin expression and sarcoplasmic reticulum Ca2+-ATPase activity in the model of pressure-overload cardiac hypertrophy. Methods: Forty male adult Sprague Dawley rats were divided into 5 groups.One group was treated by sham operation; four groups were myocardium hypertrophy cases caused by banding aortic above renal artery. Drugs were given one week after operation. Group 1: sham group, rats (n=8) were gavaged with normal saline 2 ml/(kg·d)(ig); Group 2: control group, rats (n=8) were treated with normal saline 2 ml/(kg·d) (ig); Group 3: rats (n=8) were given perindopril 2 mg/(kg·d) (ig); Group 4: rats (n=8) were treated with irbesartan 20 mg/(kg·d) (ig); Group 5: rats (n=8) were given irbesartan 20 mg/(kg·d) plus perindopril 2 mg/(kg·d) (ig). Morphometric determination, calcineurin expression and sarcoplasmic reticulum Ca2+-ATPase activity were done at the end of 6 week of drug intervention. Expression of calcineurin in myocardium was detected by immunohistochemistry. Results: Left ventricular mass index (LVMI), transverse diameter of myocardial cell (TDM), calcineurin activity were remarkably decreased after drug intervention and this decrease was most remarkable in the combination drug therapy group. Sarcoplasmic reticulum Ca2+-ATPase activity was increased after drug intervention, especially in the combined drug therapy group. Calcineurin expression in myocardium was remarkably decreased after drug intervention. LVMI was positively correlated with TDM and calcineurin, negatively correlated with sarcoplasmic reticulum Ca2+-ATPase. Conclusion:These data suggest that irbesartan and perindopril inhibit cardiac hypertrophy through the increased activity of sarcoplasmic reticulum Ca2+-ATPase and decreased expression of calcineurin. Their combination had better effects on regressing of

  7. Synergistic toxic effect of calcium, magnesium and copper on marine biofouling organisms%Ca2+、Mg2+、Cu2+对海洋污损生物的协同毒性效应

    Institute of Scientific and Technical Information of China (English)

    刘晓军; 刘贵昌; 宋树军; 铁镝

    2010-01-01

    以东方小藤壶(Chthamalus challengengerl Hoek)Ⅱ期无节幼虫作为试验对象,研究了Ca2+、Mg2+、Cu2+三种金属离子对藤壶Ⅱ期无节幼虫的单独毒性效应及协同毒性效应.研究发现Mg2+对藤壶Ⅱ期无节幼虫存在毒性效应,ca2'基本没有毒性效应,但当Ca2+、Mg2+共同作用时对藤壶Ⅱ期幼虫的毒性效应远远大于其单独作用.Cu2+对藤壶Ⅱ期幼虫具有明显的毒性效应,并且Ca2+、Mg2+浓度的升高显著增加了铜离子的毒性效应.

  8. The N Terminus of Sarcolipin Plays an Important Role in Uncoupling Sarco-endoplasmic Reticulum Ca2+-ATPase (SERCA) ATP Hydrolysis from Ca2+ Transport

    DEFF Research Database (Denmark)

    Sahoo, Sanjaya K; Shaikh, Sana A; Sopariwala, Danesh H;

    2015-01-01

    to bind SERCA throughout its kinetic cycle and promotes uncoupling of Ca(2+) transport from ATP hydrolysis. To determine the structural regions of SLN that mediate uncoupling of SERCA, we employed mutagenesis and generated chimeras of PLB and SLN. In this study we demonstrate that deletion of SLN N...

  9. The investigation for the relationship among serum leptin, erythrocyte membrane Ca2+-ATPase activity and hypertensive disorder complicating pregnancy

    Institute of Scientific and Technical Information of China (English)

    Chunfang Li; Wenli Gou; Xuelian Chen; Shuping Zhang

    2007-01-01

    Objective: To study the significance of Leptin and the activity of erythrocyte membrane Ca2+-ATPase (EMCA) in the development of hypertensive disorder complicating pregnancy. Methods: Radioimmunoassay was used to test the level of serum Leptin,and the activity of EMCA was determined chemically in 38 pregnant women with hypertensive disorder complicating pregnancy and 36 normotensive pregnant women. Results: The level of serum Leptin in hypertensive disorder complicating pregnancy(gestational hypertension: 13.76 ± 3.46 ng/ml; preeclampsia:15.76 ± 5.47 ng/ml; eclampsia: 18.32 ± 6.38 ng/ml)was significantly higher than that in normotensive pregnant women (11.33 ± 2.93 ng/ml), respectively. The average EMCA activity of patients with hypertensive disorder complicating pregnancy (gestational hypertension: 1.65 ± 0.24 μmol· pi/mg·h; preeclampsia: 1.37 ± 0.19 μmol·pi/mg·h; eclampsia:1.12 ± 0.14 μ mol·pi/mg·h) was significantly lower than that of normotensive pregnant women(1.83 ±0.38 μ mol·pi/mg·h),respectively. There was a negative correlation between the level of serum Leptin and the activity of RMCA in hypertensive disorder complicating pregnancy (r = -0.63). Conclusion: Inhibition of EMCA activity of erythrocyte in hypertensive disorder complicating pregnancy may increase cytoplasmic free calcium, which contributes to the development of hypertensive disorder complicating pregnancy. The negative correlation between the level of serum Leptin and the activity of EMCA, also suggested that serum Leptin and the activity of EMCA may play a role in the development of hypertensive disorder complicating pregnancy.

  10. Secretory pathway Ca2+/Mn2+-ATPase isoform 2 and lactation: specific localization of plasmalemmal and secretory pathway Ca2+ pump isoforms in the mammary gland

    Energy Technology Data Exchange (ETDEWEB)

    Faddy, Helen M.; Smart, Chanel E.; Xu, Ren; Lee, Genee Y.; Kenny, Paraic A.; Feng, Mingye; Rao, Rajini; Brown, Melissa A.; Bissell, Mina J.; Roberts-Thomson, Sarah J.; Monteith, Gregory R.

    2008-04-09

    The supply of calcium to the developing neonate via milk is an important physiological process. Until recently the mechanism for the enrichment of milk with calcium was thought to be almost entirely mediated via the secretory pathway. However, recent studies suggest that a specific isoform of the plasma membrane calcium ATPase, PMCA2, is the primary mechanism for calcium transport into milk, highlighting a major role for apical calcium transport. We compared the expression of the recently identified secretory calcium ATPase, SPCA2, and SPCA1, in the mouse mammary gland during different stages of development. SPCA2 levels increased over 35 fold during lactation, while SPCA1 increased only a modest two fold. The potential importance of SPCA2 in lactation was also highlighted by its localization to luminal secretory cells of the mammary gland during lactation, while SPCA1 was expressed throughout the cells of the mammary gland. We also observed major differences in the localization of PMCA2 and PMCA1 during lactation. Using the SCp2 mouse mammary epithelial cell 3D culture model, differences in the sub-cellular distribution of PMCA2 and PMCA1 were clear. These studies highlight the likely specific roles of PMCA2 and SPCA2 in lactation, and link the recently characterized SPCA2 calcium pump to the supply of calcium into milk and the regulation of Golgi resident enzymes important in lactation. They also indicate that calcium transport into milk is a complex interplay between apical and secretory pathways.

  11. Cytokines downregulate the sarcoendoplasmic reticulum pump Ca2+ ATPase 2b and deplete endoplasmic reticulum Ca2+, leading to induction of endoplasmic reticulum stress in pancreatic beta-cells

    DEFF Research Database (Denmark)

    Cardozo, Alessandra K; Ortis, Fernanda; Storling, Joachim;

    2005-01-01

    , beta-cells showed marked sensitivity to apoptosis induced by SERCA blockers, as compared with fibroblasts. Cytokine-induced ER Ca(2+) depletion was paralleled by an NO-dependent induction of CHOP protein and activation of diverse components of the ER stress response, including activation of inositol......Cytokines and free radicals are mediators of beta-cell death in type 1 diabetes. Under in vitro conditions, interleukin-1beta (IL-1beta) + gamma-interferon (IFN-gamma) induce nitric oxide (NO) production and apoptosis in rodent and human pancreatic beta-cells. We have previously shown......, by microarray analysis of primary beta-cells, that IL-1beta + IFN-gamma decrease expression of the mRNA encoding for the sarcoendoplasmic reticulum pump Ca(2+) ATPase 2b (SERCA2b) while inducing expression of the endoplasmic reticulum stress-related and proapoptotic gene CHOP (C/EBP [CCAAT/enhancer binding...

  12. 丛枝菌根真菌(AMF)对盐胁迫下芦笋幼苗生长及体内Na+、K+、Ca2+、Mg2+含量和分布的影响%Effects of arbuscular mycorrhizal fungi (AMF) on seedling growth and Na+, K+, Ca2+,Mg2+ contents and distribution in asparagus under salt stress

    Institute of Scientific and Technical Information of China (English)

    曹岩坡; 代鹏; 戴素英; 贺超兴

    2015-01-01

    以芦笋盐敏感品种‘NJ978’为材料,采用盆栽试验,研究了接种丛枝菌根真菌(AMF)对NaCl胁迫下芦笋幼苗生长及体内Na+、K+、Ca2+、Mg2+吸收和分布的影响.结果表明:在NaCl胁迫下,幼苗株高、鲜重、干重均显著降低,接种AMF可以有效缓解盐胁迫对芦笋幼苗生长的抑制;NaCl处理的芦笋幼苗根系和地上部Na+含量显著高于对照,K+、Ca2+、Mg2+的含量则显著减少;AMF+NaCl处理的芦笋幼苗根系K+、Ca2+、Mg2+含量与NaCl处理相比,分别增加了76.9%、23.1%和22.5%,而Na+含量则减少了27.4%;AMF+NaCl处理的芦笋幼苗地上部K+、Ca2+、Mg2+含量与NaCl处理相比,分别增加了58.4%、50.4%和76.0%,而Na+含量则减少了42.3%.与NaCl处理相比,接种AMF可以降低盐胁迫下芦笋幼苗根系和地上部Na+/K+、Na+/Ca2+、Na+/Mg2+,提高根系选择吸收性ASK.Na、ASCa,Na、ASMg,Na和根系向地上部的选择运输性TSK,Na、TSca.Na、TSMg,Na.由此表明,盐胁迫下接种AMF可以通过调节芦笋体内的离子平衡,从而缓解盐胁迫对植株的伤害.

  13. Sarco(endoplasmic Reticulum Ca2+-ATPase-2 Gene: Structure and Transcriptional Regulation of the Human Gene

    Directory of Open Access Journals (Sweden)

    Angel Zarain-Herzberg

    2002-01-01

    Full Text Available The sarco(endoplasmic reticulum Ca2+-ATPases (SERCAs belong to a family of active calcium transport enzymes encoded by the SERCA1, 2, and 3 genes. In this study, we describe the complete structure of the human SERCA2 gene and its 5’ -regulatory region. The hSERCA2 gene is located in chromosome 12 position q24.1 in Contig NT_009770.8, spans 70 kb, and is organized in 21 exons intervened by 20 introns. The last two exons of the pre-mRNA produce by alternatively splicing the cardiac/slow-twitch muscle-specific SERCA2a isoform and the ubiquitous SERCA2b isoform. The sequence of the proximal 225-bp regulatory region of the SERCA2 genes is 80% G+C-rich and is conserved among human, rabbit, rat, and mouse species. It contains a TATA-like-box, an E-box/USF sequence, a CAAT-box, four Sp1 binding sites, and a thyroid hormone responsive element (TRE. There are two other conserved regulatory regions located between positions -410 to -661 bp and from -919 to -1410 bp. Among the DNA cis-elements present in these two regulatory regions there are potential binding sites for: GATA-4, -5, -6, Nkx-2.5/Csx, OTF-1, USF, MEF-2, SRF, PPAR/RXR, AP-2, and TREs. Upstream from position -1.5 kb, there is no significant homology among the SERCA2 genes cloned. In addition, the human gene has several repeated sequences mainly of the Alu and L2 type located upstream from position -1.7 kb, spanning in a continuous fashion for more than 40 kb. In this study, we report the cloning of 2.4 kb of 5’-regulatory region and demonstrate that the proximal promoter region is sufficient for expression in cardiac myocytes, and the region from -225 to -1232 bp contains regulatory DNA elements which down-regulate the expression of the SERCA2 gene in neonatal cardiomyocytes.

  14. Structure of Na+,K+-ATPase at 11-A resolution: comparison with Ca2+-ATPase in E1 and E2 states.

    OpenAIRE

    Rice, W J; Young, H S; Martin, D W; Sachs, J R; Stokes, D.L.

    2001-01-01

    Na+,K+-ATPase is a heterodimer of alpha and beta subunits and a member of the P-type ATPase family of ion pumps. Here we present an 11-A structure of the heterodimer determined from electron micrographs of unstained frozen-hydrated tubular crystals. For this reconstruction, the enzyme was isolated from supraorbital glands of salt-adapted ducks and was crystallized within the native membranes. Crystallization conditions fixed Na+,K+-ATPase in the vanadate-inhibited E2 conformation, and the cry...

  15. Activity of the Na,K-ATPase alpha4 isoform is important for membrane potential, intracellular Ca2+, and pH to maintain motility in rat spermatozoa.

    Science.gov (United States)

    Jimenez, Tamara; Sánchez, Gladis; Wertheimer, Eva; Blanco, Gustavo

    2010-05-01

    While the function of the ubiquitous Na,K-ATPase alpha1 subunit has been well documented, the role of the sperm-specific alpha4 isoform of this ion transporter is less known. We have explored the importance of alpha4 in rat sperm physiology by taking advantage of the high sensitivity of this isoform for the inhibitor ouabain. Using concentrations that selectively block alpha4 activity, we found ouabain to reduce not only sperm total motility, but also multiple parameters of sperm movement, including progressive motility, straight line, curvilinear, and average path velocities, lateral head displacement, beat cross frequency, and linearity. According to a direct role of alpha4 in Na(+) transport, ouabain inhibition of alpha4 increased [Na(+)](i) in the male gametes. In addition, interference of alpha4 activity with ouabain produced cell membrane depolarization, diminished pH, and increased [Ca(2)(+)](i) in spermatozoa. Inhibition of alpha4 was sufficient to cause all these effects and additional blockage of alpha1, the other Na,K-ATPase alpha isoform expressed in sperm, and higher doses of ouabain did not result in further changes in the cell parameters studied. These results show that alpha4 is the Na,K-ATPase isoform primarily involved in controlling the transmembrane Na(+) gradient in sperm, and that alpha4 activity is necessary for maintaining membrane potential, [Ca(2)(+)](i), and [H(+)](i) in the cells. The high dependence of sperm motility on membrane excitability, [Ca(2)(+)](i), and acid-base balance suggests that their regulation is the mechanism by which alpha4 maintains motility of the male gametes.

  16. Different Na+/K+-ATPase signal pathways was involved in the increase of [Ca2+]i induced by strophanthidin in normal and failing isolated guinea pig ventricular myocytes

    Institute of Scientific and Technical Information of China (English)

    Ya-juan QI; Su-wen SU; Jun-xia LI; Ji-he LI; Fang GUO; Yong-li WANG

    2008-01-01

    Aim: To determine whether different Na+/K+-ATPase signal transduction pathways have positive inotropic effects on normal ventricular myocytes (NC) and failing ventricular myocytes (FC), and are involved in an increase of [Ca2+]i induced by strophanthidin (Str). Methods: A guinea pig model of congestive heart failure was made by constricting descending aorta. The left ventricular myocytes were enzymatically isolated. The effects of 25 μmol/L Str with different signal-transducing inhibitors on contractility and the calcium transient of NC or FC from guinea pigs were simultaneously assessed and compared with those in the 25 μmol/L Str-only group by a video-based, motion-edge detection system. Results: Str at 1, 10, and 25 μmol/L in NC and Str at 0.1, 1, 10, and 25 μmol/L) in FC elevated the calcium transient amplitude and increased the positive inotropic effects in a concentration-dependent manner, respectively. At the same concentration, the effects of Str were more potent in FC than in NC. In FC, both the mitogen-activated protein kinase (MAPK) and reactive oxygen species (ROS) signal transduction pathway of Na+/K+-ATPase were involved in the increase of the calcium transient induced by Str, but only activation of the MAPK pathway increased the calcium transient in NC. However, only the ROS pathway was involved in positive inotropic effects both in NC and FC. Conclusion: The present study suggests that Na+/K+-ATPase signaling pathways involved in the inotropic effects of Str in NC and FC are consistent, and Na+/K+-ATPase signaling pathways involved in the increase of [Ca2+]i by Str in NC and FC are different.

  17. Endomembrane Ca2+-AtPases play a significant role in virus-induced adaptation to oxidative stress

    DEFF Research Database (Denmark)

    Shabala, Sergey; Bækgaard, Lone; Shabala, Lana;

    2011-01-01

    in adaptive responses to oxidative stress by removing excessive Ca2+ from the cytosol, and that their functional expression is significantly altered in PVX-inoculated plants. These findings highlight the crucial role of Ca2+ efflux systems in acquired tolerance to oxidative stress and open up prospects...... for practical applications in agriculture, after in-depth comprehension of the fundamental mechanisms involved in common responses to environmental factors at the genomic, cellular and organismal levels....

  18. Calcium- and CaMKII-dependent chloride secretion induced by the microsomal Ca(2+)-ATPase inhibitor 2,5-di-(tert-butyl)-1,4-hydroquinone in cystic fibrosis pancreatic epithelial cells.

    OpenAIRE

    Chao, A C; Kouyama, K; Heist, E K; Dong, Y. J.; Gardner, P

    1995-01-01

    Microsomal Ca(2+)-ATPase inhibitors such as thapsigargin (THG), cyclopiazonic acid (CPA) and 2,5-di-(tert-butyl)-1,4-hydroquinone (DBHQ) have been shown to inhibit Ca2+ reuptake by the intracellular stores and increase cytosolic free Ca2+ ([Ca2+]i). DBHQ is a commercially available non-toxic synthetic compound chemically unrelated to THG and CPA. In this study, we tested the feasibility of utilizing DBHQ to improve Cl- secretion via the Ca(2+)-dependent pathway, in the cystic fibrosis (CF)-de...

  19. Some commonly used brominated flame retardants cause Ca2+-ATPase inhibition, beta-amyloid peptide release and apoptosis in SH-SY5Y neuronal cells.

    Directory of Open Access Journals (Sweden)

    Fawaz Al-Mousa

    Full Text Available Brominated flame retardants (BFRs are chemicals commonly used to reduce the flammability of consumer products and are considered pollutants since they have become widely dispersed throughout the environment and have also been shown to bio-accumulate within animals and man. This study investigated the cytotoxicity of some of the most commonly used groups of BFRs on SH-SY5Y human neuroblastoma cells. The results showed that of the BFRs tested, hexabromocyclododecane (HBCD, tetrabromobisphenol-A (TBBPA and decabromodiphenyl ether (DBPE, all are cytotoxic at low micromolar concentrations (LC(50 being 2.7 ± 0.7 µM, 15 ± 4 µM and 28 ± 7 µM, respectively. They induced cell death, at least in part, by apoptosis through activation of caspases. They also increased intracellular [Ca(2+] levels and reactive-oxygen-species within these neuronal cells. Furthermore, these BFRs also caused rapid depolarization of the mitochondria and cytochrome c release in these neuronal cells. Elevated intracellular [Ca(2+] levels appear to occur through a mechanism involving microsomal Ca(2+-ATPase inhibition and this maybe responsible for Ca(2+-induced mitochondrial dysfunction. In addition, µM levels of these BFRs caused β-amyloid peptide (Aβ-42 processing and release from these cells with a few hours of exposure. These results therefore shows that these pollutants are both neurotoxic and amyloidogenic in-vitro.

  20. X-ray effects on the activity of a Mg2+-dependent, Na+- and K+-activable microsomal membrane ATP-ase system

    International Nuclear Information System (INIS)

    The bahviour of a Mg2+-dependent, Na+- and K+-activable ATP-ase sytem on irradiation was investigated using a microsome fraction of guinea pig myocardial cells prepared by fractionated centrifugation. The Na+- and K+-activable component, transport-ATPase, was particularly radiation-sensitive. Three stages of development were observed for a 1,500 R radiation damage until 24 h p.r.. In the first stage, until 30 minutes p.r., the activity of transport-ATP-ase was inhibited. This was followed by repair processes which had reached a peak value clearly higher than the control values at 4 hours p.r.. In the third stage, the activity was reduced again; 15 and 24 hours after termination of exposure, values again were nearly the same as after 30 minutes where a maximum was observed for this radiation dose. Radiation-induced electrolyte displacements, active transport, and radiation-induced inhibition of transport-ATP-ase were correlated and discussed; the assumption was that changes in, the electrolyte conditions in the membranes on irradiation are at least partly due to the described inhibition of transport-ATP-ase. (orig./AJ)

  1. Influence of a protein hydrolysate from green algae on the activity of some ATPase systems in frog skeletal muscle.

    Science.gov (United States)

    Ivanov, R; Georgieva, B; Naumova, P; Mileva, K; Radicheva, N

    1999-06-01

    The present study investigated the effect of a protein hydrolysate from green algae cultured in the Bulgarian region of Rupy, on the enzyme activity of frog skeletal muscle. The activity of pure Mg(2+)-ATPase, Mg2+,Ca(2+)-ATPase, NaHCO3-stimulated Mg(2+)-ATPase and the latter in the presence of the inhibitors NaSCN and NaN3 in mitochondrial (B-3) and membrane (B-12) fractions were determined before and after treatment with the protein hydrolysate from green algae (30 and 300 micrograms/ml). The differences between ATPase activity of mitochondrial and membrane fractions were described and it was established that in the B-3 fraction, the activity of the NaHCO3-stimulated Mg(2+)-ATPase and Ca(2+)-dependent Mg(2+)-ATPase were accelerated by increasing concentrations of the algae protein hydrolysate. Irrespective of the different (equal or inverse) dose-dependent effects, the protein hydrolysate stimulated Mg(2+)-ATPase and that inhibited by NaSCN an NaN3 bicarbonate-stimulated Mg(2+)-ATPase activity. In most of the probes, the protein hydrolysate produced some increase in enzyme activity of NaHCO3-stimulated Mg(2+)-ATPase and Ca(2+)-dependent Mg(2+)-ATPase in B-12 fractions. The observed properties of the algae protein hydrolysate suggest that it is capable of stimulating enzyme processes in addition to having some antitoxic effect in skeletal muscle. PMID:10420389

  2. 硫酸锌溶液脱除钙镁试验及生产应用%Experiment and production application of removal of Ca2+ and Mg2+ from zinc sulphate solution

    Institute of Scientific and Technical Information of China (English)

    李天杰

    2012-01-01

    Calcium and magnesium contents of one zinc sulphate plant's zinc sulphate product were on the high side and its product's main content was also very low, because zinc oxide ore ,the raw material had high contents of calcium and magnesium, resulting in high content of calcium and magnesium in zinc sulphate solution during the sulfuric acid leaching process. Therefore, the product quality was affected.The removal of Ca2+ and Mg2+ from aqueus zinc sulphate solution with hydrofluoric acid precipitation process was experimentally studied and the production application was also made.Using hydrofluoric acid as precipitant at low temperature and high pH conditions ,Ca2+ and Mg2+ could be removed effectively from the solution.Moreover, the accumulation of fluorine ions in the solution could also be controlled.Thus the product's quality could be improved.%某硫酸锌生产厂,由于原料氧化锌矿中钙镁含量较高,用硫酸浸出后的硫酸锌溶液中钙镁含量也较高,导致生产的硫酸锌产品钙镁含量偏高,主含量偏低,影响了硫酸锌产品质量.对氢氟酸沉淀法脱除硫酸锌溶液中的钙镁离子进行了试验研究及生产应用.用氢氟酸作为沉淀剂,在较低温度和较高pH条件下可有效脱除硫酸锌溶液中的钙镁离子,并能控制氟在溶液中的累积,使制得的硫酸锌产品质量得以提高.

  3. Effects of Sodium Sulfate and Sodium Chloride on Ca2+ , Mg2+ Removal from Glauber Type Brine by Sodium Hydroxide and Sodium Carbonate%芒硝型卤水中盐硝组分对碱法脱除钙镁的影响

    Institute of Scientific and Technical Information of China (English)

    董泽亮; 张琦; 王俐聪; 蔡荣华; 马来波; 黄西平

    2013-01-01

    Ca2+ 、Mg2+ removal from Glauber type brine by sodium hydroxide and sodium carbonate has been studied.The effects of sodium sulfate and sodium chloride on removal efficiency of Ca2+,Mg2+ at room temperature were investigated in detail when addition amount of sodium hydroxide and sodium carbonate was theoretical value,reaction time was 30 min,aging time was 60 min.The experimental results show that the presence of sodium sulfate has large effect on removal efficiency of Ca2+.The removal rate of Ca2+ is more than 90% when the concentration of sodium sulfate is below 30 g/L.The presence of sodium sulfate has good but little effect on removal efficiency of Mg2+.The presence of sodium chloride has small effect on removal efficiency of Ca2+ and Mg2+.When the concentration of sodium chloride increases,the removal rate of Ca2+ increases significantly,but the removal rate of Mg2+ declines slightly.%采用“烧碱-纯碱”法,对芒硝型卤水中Ca2+和Mg2+的脱除进行了研究,在常温、两碱用量为理论用量、反应时间为30 min和陈化时间为60 min的条件下详细考察了卤水中氯化钠和硫酸钠含量的变化对Ca2+和Mg2脱除效果的影响.结果表明,硫酸钠组分对Ca2+的脱除效果影响较大,当硫酸钠含量在30 g/L以下时,Ca2+脱除率在90%以上,硫酸钠浓度增加有利于Mg2+的脱除,但影响不大.氯化钠组分对Ca2Mg2+脱除效果的影响相对较小,氯化钠含量增加,Ca2+脱除率明显增加,而Mg2+脱除率略有下降.

  4. Inhibition of the Formation of the Spf1p Phosphoenzyme by Ca2.

    Science.gov (United States)

    Corradi, Gerardo R; Czysezon, Nicolas A; Mazzitelli, Luciana R; Sarbia, Nicolas; Adamo, Hugo P

    2016-04-01

    P5-ATPases are important for processes associated with the endosomal-lysosomal system of eukaryotic cells. In humans, the loss of function of P5-ATPases causes neurodegeneration. In the yeastSaccharomyces cerevisiae, deletion of P5-ATPase Spf1p gives rise to endoplasmic reticulum stress. The reaction cycle of P5-ATPases is poorly characterized. Here, we showed that the formation of the Spf1p catalytic phosphoenzyme was fast in a reaction medium containing ATP, Mg(2+), and EGTA. Low concentrations of Ca(2+)in the phosphorylation medium decreased the rate of phosphorylation and the maximal level of phosphoenzyme. Neither Mn(2+)nor Mg(2+)had an inhibitory effect on the formation of the phosphoenzyme similar to that of Ca(2+) TheKmfor ATP in the phosphorylation reaction was ∼1 μmand did not significantly change in the presence of Ca(2+) Half-maximal phosphorylation was attained at 8 μmMg(2+), but higher concentrations partially protected from Ca(2+)inhibition. In conditions similar to those used for phosphorylation, Ca(2+)had a small effect accelerating dephosphorylation and minimally affected ATPase activity, suggesting that the formation of the phosphoenzyme was not the limiting step of the ATP hydrolytic cycle. PMID:26858246

  5. Geographic structuring of the Plasmodium falciparum sarco(endoplasmic reticulum Ca2+ ATPase (PfSERCA gene diversity.

    Directory of Open Access Journals (Sweden)

    Ronan Jambou

    Full Text Available Artemisinin, a thapsigargin-like sesquiterpene has been shown to inhibit the Plasmodium falciparum sarco/endoplasmic reticulum calcium-ATPase PfSERCA. To collect baseline pfserca sequence information before field deployment of Artemisinin-based Combination therapies that may select mutant parasites, we conducted a sequence analysis of 100 isolates from multiple sites in Africa, Asia and South America. Coding sequence diversity was large, with 29 mutated codons, including 32 SNPs (average of one SNP/115 bp, of which 19 were novel mutations. Most SNP detected in this study were clustered within a region in the cytosolic head of the protein. The PfSERCA functional domains were very well conserved, with non synonymous mutations located outside the functional domains, except for the S769N mutation associated in French Guiana with elevated IC(50 for artemether. The S769N mutation is located close to the hinge of the headpiece, which in other species modulates calcium affinity and in consequence efficacy of inhibitors, possibly linking calcium homeostasis to drug resistance. Genetic diversity was highest in Senegal, Brazil and French Guiana, and few mutations were identified in Asia. Population genetic analysis was conducted for a partial fragment of the gene encompassing nucleotide coordinates 87-2862 (unambiguous sequence available for 96 isolates. This supported a geographic clustering, with a separation between Old and New World samples and one dominant ancestral haplotype. Genetic drift alone cannot explain the observed polymorphism, suggesting that other evolutionary mechanisms are operating. One possible contributor could be the frequency of haemoglobinopathies that are associated with calcium dysregulation in the erythrocyte.

  6. Cardiac function improved by sarcoplasmic reticulum Ca2+-ATPase overexpression in a heart failure model induced by chronic myocardial ischemia

    Directory of Open Access Journals (Sweden)

    Wei XIN

    2011-04-01

    Full Text Available Objective Chronic myocardial ischemia(CMI has become an important cause of heart failure(HF.The aim of present study was to examine the effects of Sarco-endoplasmic reticulum calcium ATPase(SERCA2a gene transfer in HF model in large animal induced by CMI.Methods HF was reproduced in minipigs by ligating the initial segment of proximal left anterior descending(LAD coronary artery with an ameroid constrictor to produce progressive vessel occlusion and ischemia.After confirmation of myocardial perfusion defect and cardiac function impairment by SPECT and echocardiography in the model,animals were divided into 4 groups: HF group;HF+enhanced green fluorescent protein(EGFP group;HF+SERCA2a group;and sham operation group as control.rAAV1-EGFP and rAAV1-SERCA2a(1×1012 vg for each animal were directly and intramyocardially injected to the animals of HF+EGFP and HF+SERCA2a groups.Sixty days after the gene transfer,the expression of SERCA2a at the protein level was examined by Western blotting and immunohistochemistry,the changes in cardiac function were determined by echocardiographic and hemodynamic analysis,and the changes in serum inflammatory and neuro-hormonal factors(including BNP,TNF-a,IL-6,ET-1 and Ang II were determined by radioimmunoassay.Results Sixty days after gene transfer,LVEF,Ev/Av and ±dp/dtmax increased significantly(P < 0.05,along with an increase of SERCA2a protein expression in the ischemic myocardium(PP < 0.05,accompanied by a significant decrease of inflammatory and neural-hormonal factors(PP < 0.05 in HF+SERCA2a group as compared with HF/HF+EGFP group.Conclusions Overexpression of SERCA2a may significantly improve the cardiac function of the ischemic myocardium of HF model induced by CMI and reverse the activation of neural-hormonal factors,implying that it has a potential therapeutic significance in CMI related heart failure.

  7. Improvement in cardiac function after sarcoplasmic reticulum Ca2+-ATPase gene transfer in a beagle heart failure model

    Institute of Scientific and Technical Information of China (English)

    MI Ya-fei; LI Xiao-ying; TANG Li-jiang; LU Xiao-chun; FU Zhi-qing; YE Wei-hua

    2009-01-01

    Background Heart failure (HF) is a major cause of morbidity and mortality worldwide, but current treatment modalities cannot reverse the underlying pathological state of the heart. Gene-based therapies are emerging as promising therapeutic modalities in HF patients. Our previous studies have shown that recombinant adeno-associated viral (rAAV) gene transfer of Sarco-endoplasmic reticulum calcium ATPase (SERCA2a) can be effective in treating rats with chronic heart failure (CHF). The aim of this study was to examine the effects of SERCA2a gene transfer in a large HF animal model.Methods HF was induced in beagles by rapid right ventricular pacing (230 beats/min) for 30 days. A reduced rate ventricular pacing (180 beats/min) was continued for another 30 days. The beagles were assigned to four groups: (a) control group (n=4); (b) HF group (n=4); (c) enhanced green fluorescent protein group (n=4); and (d) SERCA2.a group (n=4). rAAVl-EGFP (lx1012 μg) and rAAVl-SERCA2a (lx1012 μg) were delivered intramyocardially. SERCA2.a expression was assessed by Western blotting and immunohistochemistry.Results Following 30 days of SERCA2a gene transfer in HF beagles its protein expression was significantly higher than in the HF group than in the control group (P <0.05). Heart function improved along with the increase in SERCA2a expression. Left ventricular systolic function significantly improved, including the ejection fraction, left ventricular systolic pressure, maximal rate of rise of left ventricular pressure (+dp/dtmax), and the maximal rate of decline of left ventricular pressure (-dp/dtmax) (P <0.05). Left ventricular end-diastole pressure significantly decreased (P <0.05). The expression of SERCA2a in the myocardial tissue was higher in the SERCA2a group than in the HF group (P<0.05). Conclusions Intramyocardial injection of rAAVl-SERCA2a can improve the cardiac function in beagles induced with HE We expect further studies on SERCA2a's long-term safety, efficacy, dosage

  8. Chelating agents related to ethylenediamine bis(2-hydroxyphenyl)acetic acid (EDDHA): synthesis, characterization, and equilibrium studies of the free ligands and their Mg2+, Ca2+, Cu2+, and Fe3+ chelates.

    Science.gov (United States)

    Yunta, Felipe; García-Marco, Sonia; Lucena, Juan J; Gómez-Gallego, Mar; Alcázar, Roberto; Sierra, Miguel A

    2003-08-25

    Iron chelates such as ethylenediamine-N,N'-bis(2-hydroxyphenyl)acetic acid (EDDHA) and their analogues are the most efficient soil fertilizers to treat iron chlorosis in plants growing in calcareous soils. EDDHA, EDDH4MA (ethylenediamine-N,N'-bis(2-hydroxy-4-methylphenyl)acetic acid), and EDDCHA (ethylenediamine-N,N'-bis(2-hydroxy-5-carboxyphenyl)acetic acid) are allowed by the European directive, but also EDDHSA (ethylenediamine-N,N'-bis(2-hydroxy-5-sulfonylphenyl)acetic acid) and EDDH5MA (ethylenediamine-N,N'-bis(2-hydroxy-5-methylphenyl)acetic acid) are present in several commercial iron chelates. In this study, these chelating agents as well as p,p-EDDHA (ethylenediamine-N,N'-bis(4-hydroxyphenyl)acetic acid) and EDDMtxA (ethylenediamine-N,N'-bis(2-metoxyphenyl)acetic acid) have been obtained following a new synthetic pathway. Their chemical behavior has been studied to predict the effect of the substituents in the benzene ring on their efficacy as iron fertilizers for soils above pH 7. The purity of the chelating agents has been determined using a novel methodology through spectrophotometric titration at 480 nm with Fe(3+) as titrant to evaluate the inorganic impurities. The protonation constants were determined by both spectrophotometric and potentiometric methods, and Ca(2+) and Mg(2+) stability constants were determined from potentiometric titrations. To establish the Fe(3+) and Cu(2+) stability constants, a new spectrophotometric method has been developed, and the results were compared with those reported in the literature for EDDHA and EDDHMA and their meso- and rac-isomers. pM values have been also determined to provide a comparable basis to establish the relative chelating ability of these ligands. The purity obtained for the ligands is higher than 87% in all cases and is comparable with that obtained by (1)H NMR. No significant differences have been found among ligands when their protonation and stability constants were compared. As expected, no Fe(3

  9. Protective effects of riboflavin and selenium on brain microsomal Ca2+-ATPase and oxidative damage caused by glyceryl trinitrate in a rat headache model.

    Science.gov (United States)

    Nazıroğlu, Mustafa; Çelik, Ömer; Uğuz, Abdulhadi Cihangir; Bütün, Ayşe

    2015-03-01

    Migraine headaches are considered to be associated with increased mitochondrial energy metabolism. Mitochondrial oxidative stress is also important in migraine headache pathophysiology although riboflavin and selenium (Se) induced a modulator role on mitochondrial oxidative stress in the brain. The current study aimed to determine the effects of Se with/without riboflavin on the microsomal membrane Ca(2+)-ATPase (MMCA), lipid peroxidation, antioxidant, and electroencephalography (EEG) values in glyceryl trinitrate (GTN)-induced brain injury rats. Thirty-two rats were randomly divided into four groups. The first group was used as the control, and the second group was the GTN group. Se and Se plus oral riboflavin were administered to rats constituting the third and fourth groups for 10 days prior to GTN administration. The second, third, and fourth groups received GTN to induce headache. Ten hours after the administration of GTN, the EEG records and brain cortex samples were obtained for all groups. Brain cortex microsomes were obtained from the brain samples. The brain and microsomal lipid peroxidation levels were higher in the GTN group compared to the control group, whereas they were decreased by selenium and selenium + riboflavin treatments. Vitamin A, vitamin C, vitamin E, and reduced glutathione (GSH) concentrations of the brain and MMCA, GSH and glutathione peroxidase values of microsomes were decreased by the GTN administration, although the values and β-carotene concentrations were increased by Se and Se + riboflavin treatments. There was no significant change in EEG records of the four groups. In conclusion, Se with/without riboflavin administration protected against GTN-induced brain oxidative toxicity by inhibiting free radicals and the modulation of MMCA activity and supporting the antioxidant redox system.

  10. Sub-chronic effect of neem based pesticide (Vepacide) on acetylcholinesterase and ATPases in rat.

    Science.gov (United States)

    Rahman, M F; Siddiqui, M K; Jamil, K

    1999-09-01

    Acetylcholinesterases (AChE), Na(+)-K+, Mg2+ and Ca(2+)-ATPases were monitored in rat brain when treated orally with 80, 160 and 320 mg/kg of Vepacide, an active ingredient from neem seed oil, daily for 90 days. Brain AChE, Na(+)-K+ and Ca(2+)-ATPases were inhibited whereas Mg(2+)-ATPase levels were enhanced in both the sexes after 45 and 90 days of treatment. The relative sensitivities of these ATPases to Vepacide indicated that Ca(2+)-ATPase being more sensitive than Na(+)-K(+)-ATPase in both the sexes. The magnitude of Ca(2+)-ATPase inhibited by this compound was higher than that of brain AChE. It appears to be sexual dimorphism in the alterations of brain AChE, Na(+)-K+ and Mg(2+)-ATPases by Vepacide with females being significant when compared with males. After 28 days of post treatment the alterations observed were approached to those of controls both in male and female rats showing reversal of the toxicity. These results indicated that the ATPases were potently inhibited by Vepacide and seemed to be its precise target among the enzyme studied. This can be used as biochemical marker of exposure to this neem derived product. PMID:10466107

  11. Ultracytochemical localization of Ca2+ ATPase activity in the inner ear of the guinea pig%豚鼠耳蜗Ca2+-ATP酶活性的超微细胞化学定位

    Institute of Scientific and Technical Information of China (English)

    孙建和; 张德添

    2006-01-01

    @@ In contrast to the perilymph,cochlear endolymph shows the special ionic components of a high K+ and low Na+.Bosher et al reported that the cochlear endolymph of laboratory rats contains very low Ca2+.

  12. Hydrolysis and Synthesis of ATP by Membrane-Bound ATPase from a Motile Streptococcus

    NARCIS (Netherlands)

    Drift, C. van der; Janssen, D.B.; Wezenbeek, P.M.G.F. van

    1978-01-01

    ATPase was detected in the membranes of a motile Streptococcus. Maximal enzymic activity was observed at pH 8 and ATP/Mg2+ ratio of 2. Mn2+ and Ca2+ could replace Mg2+ to some extent. Besides ATP, GTP and ITP were substrates. The enzyme was inhibited by N,N'-dicyclohexylcarbodiimide but not by sodiu

  13. 女贞子提取物对大强度耐力训练大鼠不同组织Mg2+-ATPase活性的影响%Effect of Fructus Ligustri Lucidi Extracts on Mg2+-ATPase Activity in Different Tissues of Rats of High-Intensity Endurance Training and Exercise Capacity

    Institute of Scientific and Technical Information of China (English)

    马云慧; 熊正英

    2013-01-01

    To analyze the change of Mg2+-ATPase activity in different tissue of rats like heart,liver,brain,kidney and quadriceps after the rats were fed with Fructus Ligustri Lucidi extracts (FLLE) and trained with high-intensity endurance training.To study the effect of FLLE as the exercise nutrition tonifying formula on the exercise capacity in rats.SD rats were randomly divided Sedentary control group,exercise control group and exercise + FLLE group,n =8.Exercise control group received high-intensity treadmill training for 6 weeks,exercise+FLLE group was fed with 2mL 400 mg/kg FLLE extract besides high-intensity treadmill training daily.Sedentary control group and exercise control group was given with 2 mL 0.5 % Tween-80 solution.After 6 weeks,sedentary control group was in rest state,and after exercise control and exercise+FLLE group were given an exhaustive exercise,determination of the different tissue Mg2+-ATPase activity of each group.The results show the exercise control group and exercise+ FLLE group of the different tissue Mg2+-ATPase activity was significantly lower than the sedentary control group,exercise+FLLE group of the different tissue Mg2+-ATPase activity was higher than the exercise control group (P<0.01 or P<0.05); with the sedentary control group,exercise control rat heart,liver,brain,kidney and quadriceps Mg2+-ATPase activity decreased by 21.01%,10.06 %,11.15 %,19.89 % and 12.36 %; with the exercise control group,exercise+FLLE group rat heart,liver,brain,kidney and quadriceps Mg2+-ATPase activity were increased by 21.62%,9.21%,8.24%,21.94% and 7.05%; compared with that of rats in the control group,the time of exhaustive exercise of rats in the exercise+FLLE group was prolonged by 23.09 %.That indicates,complementing FLLE can increase the concentration of antioxidants in the body,preventing oxidative damage to cell membranes,maintaining the steady-state concentrations of intracellular ion,and ensuring the mitochondria

  14. Total soil electrical conductivity and critical soil K+ to Ca2+ and Mg2+ ratio for potato crops Condutividade elétrica e níveis críticos da relação entre K+ e Ca+ + Mg+ no solo para cultura da batata

    Directory of Open Access Journals (Sweden)

    Roberto Anjos Reis Jr.

    1999-10-01

    Full Text Available Soil K+ to Ca2+ and Mg2+ ratio as well as the total salinity were evaluated in response to potassium fertilizer application onto potato. Potassium was applied at six different rates (0, 60, 120, 240, 480 and 960 kg ha-1 of K2O, as K2SO4, and was placed during planting time in the furrow. Soil from the 0-200 mm layer was collected in the furrow, 20 and 48 days after plant emergence (DAE to evaluate soil pH, K+, Ca2+ and Mg2+ contents and the total electrical conductivity (EC. A factorial design (6x2, with six K rates and two sampling times was set up in a randomized block design with four replications. The application of K fertilizer increased exchangeable K, did not affect pH and exchangeable Ca and Mg contents, but caused a linear increase of the soil K+/(Ca2++Mg2+1/2 ratio as well as EC. At 20 DAE, the critical soil K+/(Ca2++Mg2+1/2ratio and the EC associated with maximum tuber yield (30.5 Mg.ha-1, with 353.4 kg ha-1 of K2O were 1.79 and 1.6 dS m-1, respectively. The highest soil K+/(Ca2++Mg2+1/2 ratio and EC were obtained with the highest application of K fertilizer, which led to a reduction in the potato tuber yield.Com o objetivo de avaliar a relação entre K e Ca + Mg e a salinidade no solo em resposta à adubação potássica no cultivo da batateira (cultivar Baraka, foi instalado experimento fatorial a nível de campo com seis doses de potássio (0, 60, 120, 240, 480 e 960 kg ha-1 de K2O e duas épocas de amostragem, 20 e 48 dias após a emergência das plantas, (DAE delineado em blocos casualizados com quatro repetições. O potássio foi aplicado como K2SO4 no sulco de plantio. O solo foi amostrado (0-200 mm de profundidade para avaliar o pH, a condutividade elétrica e os teores de K, Ca e Mg. A adubação potássica aumentou o K trocável, não afetou o pH e os teores de Ca e Mg trocáveis no solo, e elevou linearmente a condutividade elétrica e a relação K+/(Ca2++ Mg2+1/2. Aos 20 DAE, a máxima produção de tubérculos foi

  15. [ATPase and phosphatase activity of drone brood].

    Science.gov (United States)

    Bodnarchuk, L I; Stakhman, O S

    2004-01-01

    Most researches on insect enzymes concern carbohydrate and nitrogenous exchange. Data on ATPase activity for larval material of drone brood are absent in the available literature. The drone brood is one of the least investigated apiproducts. Allowing for the important role of ATPase in the vital functions of the insect cells our work was aimed at the study of ATPase of the drone blood activity and that of alkaline and acid phosphatases. When studying liophylised preparations of the drone brood homogenate we have found out high activity of Mg2+, Na+, K+-, Ca2+- and Mg2+-ATPase and of alkaline and acid phosphatase, that is the possible explanation of the high-intensity power and plastic processes proceeding during growth and development of larvae. PMID:16350755

  16. Hailey-Hailey disease and tight junctions: Claudins 1 and 4 are regulated by ATP2C1 gene encoding Ca2+/Mn2+ ATPase SPCA1 in cultured keratinocytes

    OpenAIRE

    Raiko, Laura; Siljamäki, Elina; Mahoney, Mỹ G.; Putaala, Heli; Suominen, Erkki; Peltonen, Juha; Peltonen, Sirkku

    2012-01-01

    Mutations in the ATP2C1 gene encoding Ca2+/Mn2+ ATPase SPCA1 cause Hailey-Hailey disease (HHD, OMIM 16960). HHD is characterized by epidermal acantholysis. We attempted to model HHD using normal keratinocytes in which the SPCA1 mRNA was down-regulated with the small inhibitory RNA (siRNA) method. SiRNA inhibition significantly down-regulated the SPCA1 mRNA, as demonstrated by qPCR, and decreased the SPCA1 protein beyond detectable level, as shown by western analysis. The expression of selecte...

  17. 铁·镁·钙元素对白腐真菌生长及降解石油的影响%Effect of Fe2+ , Mg2+ and Ca2+ on Oil Degradation and the Growth of White Rot Fungi

    Institute of Scientific and Technical Information of China (English)

    谭丽泉; 黄敏; 余梅; 梁俊豪

    2012-01-01

    [目的]探讨黄孢原毛平革菌的生长及对石油的降解能力的优化条件.[方法]选用白腐真菌的典型菌种——黄孢原毛平革菌作为降解菌,研究其生长及对石油的降解性能,以不同浓度的Fe、Mg、Ca元素作为研究对象,在筛选单因素最佳水平的基础上,设计L9(33)正变表进行正变试验,对各因素的重要性及其最优水平进行分析.[结果]单因素试验表明,体系中添加Fe、Mg、Ca元素对白腐真菌的生长及其对石油的降解能力有较大的影响.正交试验结果表明,3种元素对P.C.菌降解石油的影响大小依次为Fe2+> Ca2+ >Mg2+;溶液中铁、镁、钙元素的最佳组合为0.008 g/L的Fe2、0.8g/L的Mg2+、1.5g/L的Ca2+,该条件下10d后白腐真菌对石油的降解率为70.9%.[结论]该研究为探索白腐真菌的生长及其对石油降解能力的优化提供了科学依据.%[Objective] The study aimed to discuss the growth and degradation of petroleum by white rot fungi. [Method] Taking the typical white rot fungi P. chrysosporium as the degrading fungus,its growth and degradation on the petroleum were studied. The effect of cations including Fe2+ 、Mg2+ and Ca2+ were selected as the effecting factors,on the basis of screening the best level of single factor, the orthogonal experiment with the L L9(33 )orthogonal design was conducted to analyze the importance of the factors and their optimal level. [ Result]The single test showed that Fe2+ ,Mg2+ and Ca2+ had a greater influence on the growth of white rot fungi, and biodegradation of petroleum. The results of orthogonal experiment showed that Fe2+ 、Mg2+ and Ga2 + had significant influence on the degradation of petroleum by white rot fungus, the effect was Fe2+ > Ca2+ >Mg2+ , the Degradation can reached up to 70.9% under the optimum conditions: [Fe2+ ] = 0.008 g/L,[ Mg2+ ] =0.8 g/L,[ Ca2+ ] =1.5 g/L. [Gonclusion] The study provided the scientific basis for exploring the effective method of

  18. Effects of Four Interior-warming Drugs on the Tension of Ileum Smooth Muscle and Ca2+-ATPase in Rabbits%4种温里药对兔回肠平滑肌张力及Ca2+-ATP 酶的影响

    Institute of Scientific and Technical Information of China (English)

    黄庆芳; 陈艳芬; 杨全; 杨超燕; 唐春萍; 明露; 黎洁玲; 陶曙红

    2016-01-01

    Objective To investigate the effects of four interior-warming drugs( galangal,cinnamon,evodia rutaecarpa,and dried ginger)on the tension of ileum smooth muscle and Ca2+-ATPase on the cell membrane in rabbits. Methods The effects of galangal,cinnamon,evodia rutaecarpa,and dried ginger were examined on normal ileum smooth muscle,in vitro intestinal muscle contraction caused by acetylcholine(ACh),barium chloride(BaCl2 )and histamine(His), and ACh-induced calcium release by using BL-420E+ biological signal collection and processing system.The average tension was measured within 1 min before delivery and within 3 minutes after the treatment,and the inhibition rate was calculated according to the average tension value.The effects of sera containing galangal,cinnamon,evodia rutaecarpa,and dried ginger on Ca2+-ATPase activity on the cell membrane of the intestinal smooth muscle were examined by phosphorus method. Results Galangal,cinnamon,evodia rutaecarpa,and dried ginger at high concentrations could restrain in vitro intestinal contraction in normal circumstances(P<0.05 or P<0.01).Significant inhibitory effects on intestinal contraction caused by ACh,His and BaCl2 were found in low,medium and high concentration groups(P<0.01).There was a dose-effectiveness relationship between the inhibition rate and final drug concentrations.The ACh-induced intracellular and extracellular calcium dependent contraction were significantly inhibited by the four interior-warming drugs( P < 0. 05 or P < 0. 01). The Ca2+-ATPase activities were( 0. 384 ± 0.070),(0.302±0.016),(0.307±0.016),(0.296±0.016),(0.313±0.003)U·mg-1 ,respectively,in intestinal smooth muscle in normal control group and high concentration groups of galangal,cinnamon,evodia rutaecarpa,and dried ginger(P<0.01). Conclusion Interior-warming drugs may relax intestinal smooth muscle by reducing the intracellular calcium release and the extracellular calcium inflow via receptor-controlled calcium channels,and inhibiting the

  19. 颈椎椎后肌肉组织Ca2+-ATP酶与颈椎病及颈部软组织病理变化的相关性%Correlation of Ca2+-ATPase activity with pathological changes in cervical vertebra muscular tissue and cervical syndrome

    Institute of Scientific and Technical Information of China (English)

    罗才贵; 温元强; 朱德良; 罗建

    2007-01-01

    目的:通过分析Ca2+与骨骼肌的关系,进一步阐述颈椎椎后肌肉组织内Ca2+-ATP酶与颈椎病的关系,及颈部软组织的病理变化.方法:应用计算机检索PubMed1985-01/2006-12相关骨骼肌损伤与肌组织Ca2+-ATP酶关系方面的文献,检索词"Ca2+pump,Ca2+-ATPase,skeletal muscle".限定文献语言种类为English.同时计算机检索CNKI与万方数据库1990-01/2006-12相关钙ATP酶与骨骼肌损伤的关系,及颈椎病发病病因的文献.检索词"钙ATP酶(Ca2+-ATP酶),骨骼肌,颈椎病病因",限定文献语言种类为中文.对资料进行初审,选取包括Ca2+-ATP酶与肌组织损伤相关的文献,开始查找全文.纳入标准:Ca2+-ATP酶活性变化与骨骼肌损伤密切相关的文献研究.排除标准:重复研究,Meta分析类文章.共检索到4 050篇关于Ca2+-ATP酶活性变化,骨骼肌损伤及颈椎病发病原因等方面的文献,最终纳入24篇符合标准的文献.结果:目前对酶活性变化与骨骼肌损伤的关系研究已较为广泛.而骨骼肌损伤作为颈椎病发病的一个因素,以酶活性变化作为指标来观察颈椎病发病时颈部软组织病理变化情况,及治疗后软组织病理变化情况的研究则较为少见.众多研究表明Ca2+是参与骨骼肌收缩的重要离子之一,与骨骼肌的生理病理有密切关系,而肌细胞内质网膜上的Ca2+-ATP酶是调节细胞内Ca2+浓度的重要蛋白质之一.骨骼肌慢性劳损易导致ATP酶活性下降,而酶活性的下降加重骨骼肌损伤,而引发系列疾病.强迫屈颈体位作为颈椎病发病的危险因素之一,可使颈椎椎后肌肉Ca2+-ATP酶活性降低,酶活性降低致使肌细胞损伤,并最终导致骨骼肌损伤而发病.结论:Ca2+-ATP酶活性变化是颈部软组织病理变化的一个重要指标之一.同时,该酶的活性变化也与颈椎病发病有着密切的关系.

  20. Antioxidation and ATPase activity in the gill of mud crab Scylla serrata under cold stress

    Institute of Scientific and Technical Information of China (English)

    KONG Xianghui; WANG Guizhong; LI Shaojing

    2007-01-01

    Mud crab (Scylla serrata) is an important commercial crustacean in China. An experiment was designed to study the effect of cold stress on S. serrata. After a one-week adaptation at 28 ℃, the temperature is suddenly reduced to 4 ℃. The crabs were sampled every 2 h for 10 h and dissected immediately to measure the enzyme activity. The crabs at room temperature (28 ℃) were used as the control group. The activity of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX), the content of malondialdehyde (MDA) and the activity of 4 ATPases (Na+, K+-ATPase;Mg2+-ATPase; Ca2+-ATPase; Ca2+, Mg2+-ATPase) were measured biochemically. In contrast to the control group, the SOD activity increased significantly from 2 to 6 h after the cold stress, and then decreased. The CAT and GPX activities increased in 2 h, and then decreased gradually. The content of MDA increased gradually in 4 h. The activity ofNa+, K+-ATPase decreased in 2 h, increased up to the top value at Hour 6,then decreased again. The activities of Mg2+-ATPase, Ca2+-ATPase and Ca2+, Mg2+-ATPase increased significantly in 6 h, insignificantly in any other hours. Under cold stress, the activity of antioxidative enzymes in S. serrata was reduced at first then stabilized, ROS-scavenging weakened, and MDA accumulated gradually in the gill after 6 h. The activity of the 4 ATPases in the crab decreased after 6 h,suggesting that the ability to regulate ion concentration has been paralyzed. Therefore, the maximum period to sustain healthy meat in the crab under cold stress is 6 hours.

  1. Changes of Ca2+ in mitochondria and Ca2+-ATPase activity in sarcoplasmic reticulum(SR) of different skeletal muscle fiber types in rat after exhaustive exercise%力竭运动后大鼠骨骼肌不同肌纤维线粒体钙含量和肌浆网Ca2+-ATP酶活性变化

    Institute of Scientific and Technical Information of China (English)

    王翔; 魏源

    2002-01-01

    为探讨急性力竭运动后大鼠骨骼肌不同肌纤维内Ca2+转运功能变化,测定了股四头肌红肌和白肌线粒体钙含量和肌浆网Ca2+-ATP酶活性,结果红肌和白肌线粒体Ca2+含量增加,肌浆网Ca2+-ATP酶活性降低,说明急性力竭运动后肌细胞内Ca2+转动功能发生了改变,影响了肌肉收缩特性,从而导致了运动性疲劳的产生.

  2. 脱硫废弃物对碱胁迫下水稻叶片钙分布、Ca2+-ATPase活性及抗氧化特征的影响%Effects of desulfurization waste on calcium distribution, Ca2+-ATPase activity, and antioxidant characteristics of rice leaf under alkali stress

    Institute of Scientific and Technical Information of China (English)

    毛桂莲; 许兴; 曾瑾; 岳自慧; 杨淑娟

    2012-01-01

    为了探讨脱硫废弃物提高水稻抗盐碱的作用机制,采用盆栽法,研究脱硫废弃物对碱胁迫下水稻幼苗叶片总钙含量、Ca2+分布、细胞膜Ca2+-ATPase活性及活性氧含量等的变化.结果表明:对照处理的细胞中钙颗粒零星分布于细胞壁和叶绿体中,添加脱硫废弃物和CaSO4处理的细胞质膜、细胞间隙、细胞壁和液泡中有大量的钙颗粒分布;随着脱硫废弃物和CaSO4添加量的增加,叶片总钙含量增加,质膜和液泡膜Ca2+-ATPase活性呈上升趋势,质膜透性、MDA含量和活性氧O2-产生速率呈下降趋势,SOD、POD等保护酶活性升高.添加脱硫废弃物在一定程度上能够减缓碱胁迫对水稻造成的细胞伤害,起主要作用的物质可能是其主要成分CaSO4.%To approach the action mechanisms of desulfurization waste on alleviating alkali stress-induced injury of rice, a pot experiment was conducted to study the variations of leaf total calcium content, calcium distribution, plasma membrane Ca2+-ATPase activity, and reactive oxygen content of rice seedlings under alkali stress after the application of desulfurization waste. In the control, a few calcium particulates scattered in the cell wall and chloroplasts, while applying desulfurization waste or CaS04 increased the calcium particulates in the plasma membrane, intercellular space, cell wall, and vacuole significantly. With the increasing application rate of desulfurization waste or CaSO4, the leaf total calcium content increased, Ca2+ -ATPase activity in plasma membrane and tonoplast presented an increasing trend, plasma membrane relative permeability, MDA content, and O2' production rate decreased, and SOD and POD activities increased. The desulfurization waste could relieve the alkali stress to rice in some extent, and the main reactive compound in the waste could be CaS04.

  3. Expression and secretion of plasma membrane Ca2+-ATPase 4a (PMCA4a) during murine estrus: association with oviductal exosomes and uptake in sperm.

    Science.gov (United States)

    Al-Dossary, Amal A; Strehler, Emanuel E; Martin-Deleon, Patricia A

    2013-01-01

    PMCA4, a membrane protein, is the major Ca(2+) efflux pump in murine sperm where its deletion leads to a severe loss of hyperactivated motility and to male infertility. We have previously shown that the PMCA4b splice variant interacts with CASK (Ca(2+/)CaM-dependent serine kinase) in regulating sperm Ca(2+). More recently we detected that PMCA4a isoform, in addition to its presence in testis, is secreted in the epididymal luminal fluid and transferred to sperm. Here we show that Pmca4 mRNA is expressed in both the 4a and 4b variants in the vagina, uterus, and oviduct. Immunofluorescence reveals that PMCA4a is similarly expressed and is elevated during estrus, appearing in the glandular and luminal epithelia. Western analysis detected PMCA4a in all tissues and in the luminal fluids (LF) of the vagina (VLF), uterus (ULF), and the oviduct (OLF) collected during estrus. It was ~9- and 4-fold higher in OLF than in VLF and ULF, and only marginally present in LF collected at metestrus/diestrus. Fractionation of the LF collected at estrus, via ultracentrifugation, revealed that 100% of the PMCA4a resides in the vesicular fraction of the ULF and OLF. Transmission electron microscopy (TEM) revealed that OLF vesicles have an exosomal orientation (with the cytoplasmic-side inward), a size range of 25-100 nm, with the characteristic CD9 biomarker. Thus, we dubbed these vesicles "oviductosomes", to which PMCA4a was immunolocalized. Incubation of caudal sperm in the combined LF or exosomes resulted in up to a ~3-fold increase of sperm PMCA4a, as detected by flow cytometry, indicating in vitro uptake. Our results are consistent with the increased requirement of Ca(2+) efflux in the oviduct. They show for the first time the presence of oviductal exosomes and highlight their role, along with uterosomes and vaginal exosomes, in post-testicular sperm acquisition of PMCA4a which is essential for hyperactivated motility and fertility. PMID:24244642

  4. Expression and secretion of plasma membrane Ca2+-ATPase 4a (PMCA4a during murine estrus: association with oviductal exosomes and uptake in sperm.

    Directory of Open Access Journals (Sweden)

    Amal A Al-Dossary

    Full Text Available PMCA4, a membrane protein, is the major Ca(2+ efflux pump in murine sperm where its deletion leads to a severe loss of hyperactivated motility and to male infertility. We have previously shown that the PMCA4b splice variant interacts with CASK (Ca(2+/CaM-dependent serine kinase in regulating sperm Ca(2+. More recently we detected that PMCA4a isoform, in addition to its presence in testis, is secreted in the epididymal luminal fluid and transferred to sperm. Here we show that Pmca4 mRNA is expressed in both the 4a and 4b variants in the vagina, uterus, and oviduct. Immunofluorescence reveals that PMCA4a is similarly expressed and is elevated during estrus, appearing in the glandular and luminal epithelia. Western analysis detected PMCA4a in all tissues and in the luminal fluids (LF of the vagina (VLF, uterus (ULF, and the oviduct (OLF collected during estrus. It was ~9- and 4-fold higher in OLF than in VLF and ULF, and only marginally present in LF collected at metestrus/diestrus. Fractionation of the LF collected at estrus, via ultracentrifugation, revealed that 100% of the PMCA4a resides in the vesicular fraction of the ULF and OLF. Transmission electron microscopy (TEM revealed that OLF vesicles have an exosomal orientation (with the cytoplasmic-side inward, a size range of 25-100 nm, with the characteristic CD9 biomarker. Thus, we dubbed these vesicles "oviductosomes", to which PMCA4a was immunolocalized. Incubation of caudal sperm in the combined LF or exosomes resulted in up to a ~3-fold increase of sperm PMCA4a, as detected by flow cytometry, indicating in vitro uptake. Our results are consistent with the increased requirement of Ca(2+ efflux in the oviduct. They show for the first time the presence of oviductal exosomes and highlight their role, along with uterosomes and vaginal exosomes, in post-testicular sperm acquisition of PMCA4a which is essential for hyperactivated motility and fertility.

  5. Structure-based drug design studies of the interactions of ent-kaurane diterpenes derived from Wedelia paludosa with the Plasmodium falciparum sarco/endoplasmic reticulum Ca2+-ATPase PfATP6

    Directory of Open Access Journals (Sweden)

    Daniel Silqueira Martins Guimarães

    2015-04-01

    Full Text Available Malaria is responsible for more deaths around the world than any other parasitic disease. Due to the emergence of strains that are resistant to the current chemotherapeutic antimalarial arsenal, the search for new antimalarial drugs remains urgent though hampered by a lack of knowledge regarding the molecular mechanisms of artemisinin resistance. Semisynthetic compounds derived from diterpenes from the medicinal plant Wedelia paludosa were tested in silico against the Plasmodium falciparum Ca2+-ATPase, PfATP6. This protein was constructed by comparative modelling using the three-dimensional structure of a homologous protein, 1IWO, as a scaffold. Compound 21 showed the best docking scores, indicating a better interaction with PfATP6 than that of thapsigargin, the natural inhibitor. Inhibition of PfATP6 by diterpene compounds could promote a change in calcium homeostasis, leading to parasite death. These data suggest PfATP6 as a potential target for the antimalarial ent-kaurane diterpenes.

  6. Green fluorescent protein-tagged sarco(endo)plasmic reticulum Ca2+-ATPase overexpression in Paramecium cells: isoforms, subcellular localization, biogenesis of cortical calcium stores and functional aspects.

    Science.gov (United States)

    Hauser, K; Pavlovic, N; Klauke, N; Geissinger, D; Plattner, H

    2000-08-01

    We have followed the time-dependent transfection of Paramecium cells with a vector containing the gene of green fluorescent protein (GFP) attached to the C-terminus of the PtSERCA1 gene. The outlines of alveolar sacs (ASs) are labelled, as is the endoplasmic reticulum (ER) throughout the cell. When GFP fluorescence is compared with previous anti-PtSERCA1 antibody labelling, the much wider distribution of GFP (ER+ASs) indicates that only a small amount of SERCA molecules is normally retained in the ER. A second isoform, PtSERCA2, also occurs and its C-terminal GFP-tagging results in the same distribution pattern. However, when GFP is inserted in the major cytoplasmic loop, PtSERCA1 and two fusion proteins are mostly retained in the ER, probably because of the presence of the overt C-terminal KKXX ER-retention signal and/or masking of a signal for transfer into ASs. On the overall cell surface, new SERCA molecules seem to be permanently delivered from the ER to ASs by vesicle transport, whereas in the fission zone of dividing cells ASs may form anew. In cells overexpressing PtSERCA1 (with C-terminal GFP) in ASs, [Ca2+]i regulation during exocytosis is not significantly different from controls, probably because their Ca2+ pump has to mediate only slow reuptake.

  7. Effect of endurance swimming on rat cardiac myofibrillar ATPase with experimental diabetes.

    Science.gov (United States)

    Belcastro, A N; Maybank, P; Rossiter, M; Secord, D

    1985-09-01

    Diabetes is characterized by depressed cardiac functional properties attributed to Ca2+-activated ATPase activity. In contrast, endurance swimming enhances the cardiac functional properties and Ca2+-activated myofibril ATPase. Thus, the purpose of this study was to observe if the changes associated with experimental diabetes can be ameliorated with training. Diabetes was induced with a single i.v. injection of streptozotocin (60 mg/kg). Blood and urine glucose concentrations were 802 +/- 44 and 6965 +/- 617 mg/dL, respectively. The training control and training diabetic animals were made to swim (+/- 2% body weight) 4 days/week for 8 weeks. Cardiac myofibril, at 10 microM free Ca2+ concentration was reduced by 54% in the sedentary diabetics compared with sedentary control animals (p less than 0.05). Swim training enhanced the Ca2+-activated myofibril ATPase activities for the normal animals. The diabetic animals, which swam for 8 weeks, had further reduced their Ca2+-activated myofibril ATPase activity when compared with sedentary diabetics (p less than 0.05). Similarly, the Mg2+-stimulated myofibril ATPase activity was depressed by 31% in diabetics following endurance swimming. It is concluded that the depressed Ca2+-activated myofibril ATPase activity of diabetic hearts is not reversible with endurance swimming. PMID:2932207

  8. Materiales de Al2O3 - MgAl2O4 - CaAl12O19 - Ca2Mg2Al28O46 obtenidos mediante un proceso de sinterización reactiva entre Al2O3 y CaMg(CO3)2

    OpenAIRE

    Aza Moya, Antonio H. de; Peña, P.; Moset, M.

    2002-01-01

    [ES] Utilizado la información suministrada por el diagrama de equilibrio de fases Al2O3-MgO-CaO se ha diseñado y obtenido un material de Al2O3-MgAl2O4-CaAl12O19-Ca2Mg2Al28O46 mediante sinterización reactiva de una mezcla de Al2O3 y CaMg(CO3)2. Las reacciones que tienen lugar en la mezcla durante el proceso se han estudiado usando técnicas de análisis térmico diferencial, termogravimetrico y dilatometría. Muestras reaccionadas a temperaturas seleccionadas se han estudiado por difracci...

  9. 离子色谱法测定奶粉中氯化胆碱、钠、钾、镁、钙的含量%Determination of Choline Chloride,Na^+,K^+, Mg^2+ and Ca^2+ in Milk Powder by Ion Chromatography

    Institute of Scientific and Technical Information of China (English)

    曹文军; 崔晗; 沈葆真; 苏海滨; 贺舒文; 黄大亮; 李莉

    2012-01-01

    A method was established to determine choline chloride,Na+,K+,Mg2+ and Ca2+ in milk powder by using ion exchange chromatography with conductivity detector.The samples were separated on a IonPac CS-12A(250mm×4 mm) cation exchange column using gradient elution of MSA,and the velocity was 1.0 mL/min.The detection limit of the method was 0.5~10 mg/L.The relative standard deviation of this method is 2.3%-4.7%(n=6)and the recovery is the range of 74.7%~93.5%.The method is accurate and simple,and is suitable for rapid detection choline chloride,Na+,K+,Mg2+ and Ca2+in milk powder.%建立离子色谱定量测定奶粉中钠、钾、氯化胆碱、镁、钙的方法,应用IonPac CS-12A(250 mm×4 mm)阳离子交换柱,淋洗液为20 mmol/L MSA,等浓度淋洗,流速为1.0 mL/min,电导检测器检测,方法检出限为0.5~10 mg/kg。6次测定平行样,相对标准偏差2.3%~4.7%,5种阳离子加标回收率74.7%~93.5%,该方法具有准确、操作简便等特点,可用于奶粉中钠、钾、氯化胆碱、镁、钙的检测。

  10. Comparison of endogenous and exogenous sources of ATP in fueling Ca2+ uptake in smooth muscle plasma membrane vesicles

    OpenAIRE

    1992-01-01

    A smooth muscle plasma membrane vesicular fraction (PMV) purified for the (Ca2+/Mg2+)-ATPase has endogenous glycolytic enzyme activity. In the presence of glycolytic substrate (fructose 1,6-diphosphate) and cofactors, PMV produced ATP and lactate and supported calcium uptake. The endogenous glycolytic cascade supports calcium uptake independent of bath [ATP]. A 10-fold dilution of PMV, with the resultant 10-fold dilution of glycolytically produced bath [ATP] did not change glycolytically fuel...

  11. Effects of incremental load training on the activity of Na+,K+-ATPase and Ca2+-ATPase in rats'skeletal muscle sarcoplasmic reticulum%递增负荷训练对大鼠骨骼肌肌浆网Na+,K+-ATP酶和Ca2+-ATP酶活性的影响

    Institute of Scientific and Technical Information of China (English)

    张敏; 陈立军; 周蔚

    2010-01-01

    [目的]研究递增负荷训练对大鼠骨骼肌肌浆网Na+,K+-ATP酶和Ca2+-ATP酶活性的影响.[方法]参照BEDFORD TG 标准,采用跑台运动方式,建立大鼠递增负荷训练模型.将24只大鼠随机分为3组:正常对照组(n=8)、递增负荷运动4周组(n=8)和递增负荷运动6周组(n=8).超速离心法提取大鼠骨骼肌肌浆网,测定肌浆网Na+,K+-ATP酶和Ca2+-ATP酶的活性.[结果]递增负荷运动组Na+,K+-ATP酶和Ca2+-ATP酶的活性较正常对照组显著升高(P<0.05).[结论]实验结果表明,一定时间的递增负荷训练可提升骨骼肌肌浆网Na+,K+-ATP酶和Ca2+-ATP酶的活性.

  12. Secretory pathway Ca(2+)-ATPase isoform 1 knockdown promotes Golgi apparatus stress injury in a mouse model of focal cerebral ischemia-reperfusion: In vivo and in vitro study.

    Science.gov (United States)

    Fan, Yongmei; Zhang, Changjie; Peng, Wenna; Li, Ting; Yin, Jing; Kong, Ying; Lan, Chunna; Li, Xiaofang; Wang, Rumi; Hu, Zhiping

    2016-07-01

    The present study was designed to investigate the potential role of secretory pathway Ca(2+)-ATPase isoform 1(SPCA1) in experimental focal cerebral ischemia-reperfusion injury. Cerebral ischemia-reperfusion was induced by transient middle cerebral artery occlusion (MCAO) for 2h s in Sprague-Dawley rats, and then the expression levels of SPAC1 mRNA and protein were determined. Results showed that SPCA1 level was transiently increased 1 day after reperfusion in peri-infarction area, while markedly increased in infarction core on 3day and 7 day after reperfusion. Then a SPCA1 lentivirus was used to achieve knockdown of SPCA1 gene: Ca(2+) transporting type 2C, member 1 (ATP2C1) gene. It has been observed that SPCA1 knockdown by lentivirus markedly increased cerebral infarction volume in vivo. Meanwhile, SPCA1 knockdown also facilitated per-oxidative production, including nitric oxide (NO) and 3-nitrotyrosine (3-NT) and decreased the expression of total superoxide dismutase (SOD) and manganese superoxide dismutase (MnSOD). Moreover, in vitro study showed that SPCA1 knockdown increased hydrogen peroxide (H2O2)-induced lactate dehydrogenase (LDH) leakage dose-dependently, and elevated caspase3 level in neuro-2a (N2a) cells. In addition, SPCA1 knockdown increased H2O2-induced production of nitric oxide and 3-NT dose-dependently, and reversed the increased activity of total SOD and MnSOD in neuro-2a cells. In conclusion, the present study indicated that SPCA1 could suppress over active Golgi apparatus (GA) stress thus attenuate cerebral ischemia-reperfusion injury. PMID:27038757

  13. 急性运动对大鼠骨骼肌线粒体Ca2+-ATP酶和H+-ATP酶活性的影响%Effects of Acute Exercise on Activities of Mitochondrial Ca2+-ATPase and H+-ATPase in Skeletal Muscles of Rats

    Institute of Scientific and Technical Information of China (English)

    周锦琳; 田野

    2001-01-01

    采用不同强度的跑台运动,观察大鼠运动后即刻骨骼肌线粒体Ca2+-ATP酶和H+-ATP酶活性的变化.结果发现:与对照组相比,股四头肌和腓肠肌线粒体Ca2+-ATP酶活性大强度运动后略有变化;中等强度运动后显著下降,分别下降了60.60%和57.53%(P<0.01).股四头肌和腓肠肌线粒体H+-ATP酶活性运动后非常明显地增加,大强度分别增加了39.83%和48.08%(P<0.01);中等强度运动后,股四头肌H+-ATP酶活性增加了35.97%(P<0.01).结果表明线粒体Ca2+-ATP酶活性下降可能是造成中等强度长时间运动后骨骼肌疲劳的原因之一.运动后即刻骨骼肌线粒体H+-ATP酶活性非常明显地增加,但这并不能肯定运动后H+-ATP酶合成ATP的能力一定增强.

  14. Crystal Structure of the Vanadate-Inhibited Ca2+-ATPase

    DEFF Research Database (Denmark)

    Clausen, Johannes D.; Bublitz, Maike; Arnou, Bertrand Jean-Paul;

    2016-01-01

    was facilitated by trinitrophenyl (TNP)-derivatized nucleotides that bind with the TNP moiety occupying the binding pocket that normally accommodates the adenine of ATP, rationalizing their remarkably high affinity for E2P-like conformations of the Ca2+-ATPase. A comparison of the configurations of bound......Vanadate is the hallmark inhibitor of the P-type ATPase family; however, structural details of its inhibitory mechanism have remained unresolved. We have determined the crystal structure of sarcoplasmic reticulum Ca2+-ATPase with bound vanadate in the absence of Ca2+. Vanadate is bound...... at the catalytic site as a planar VO3− in complex with water and Mg2+ in a dephosphorylation transition-state-like conformation. Validating bound VO3− by anomalous difference Fourier maps using long-wavelength data we also identify a hitherto undescribed Cl− site near the dephosphorylation site. Crystallization...

  15. A many-body model to study proteins. I. Applications to MLnm+ complexes, Mm+=Li+, Na+, K+, Mg2+, Ca2+, and Zn2+, L=H2O, CH3OH, HCONH2, n=1-6, and to small hydrogen bonded systems

    Science.gov (United States)

    Masella, Michel; Cuniasse, Philippe

    2003-07-01

    A new model to study proteinic systems including a many-body polarization and a hydrogen bond energy contribution is presented. This model represents an extension of an earlier water many-body model [M. Masella and J.-P. Flament, J. Chem. Phys. 107 9105 (1997)]. As in this earlier model, the new model is developed to reproduce quantum computations on small molecular aggregates, and, in this first paper, we focus our efforts in developing an accurate potential to describe interactions among all nonbonded atoms occurring in proteins, and among those atoms and six cations of biological interest: Li+, Na+, K+, Mg2+, Ca2+, and Zn2+. Intramolecular degrees of freedom are described as in classical two-body force fields. In the present paper, the new model is applied to investigate the properties of small ion-neutral [M,Ln]m+ complexes and of small hydrogen-bonded systems. The results showed that this model is able to reproduce most of the theoretical quantum predictions and experimental data published until now regarding those systems.

  16. Motion of the Ca2+-pump captured.

    Science.gov (United States)

    Yokokawa, Masatoshi; Takeyasu, Kunio

    2011-09-01

    Studies of ion pumps, such as ATP synthetase and Ca(2+)-ATPase, have a long history. The crystal structures of several kinds of ion pump have been resolved, and provide static pictures of mechanisms of ion transport. In this study, using fast-scanning atomic force microscopy, we have visualized conformational changes in the sarcoplasmic reticulum Ca(2+)-ATPase (SERCA) in real time at the single-molecule level. The analyses of individual SERCA molecules in the presence of both ATP and free Ca(2+) revealed up-down structural changes corresponding to the Albers-Post scheme. This fluctuation was strongly affected by the ATP and Ca(2+) concentrations, and was prevented by an inhibitor, thapsigargin. Interestingly, at a physiological ATP concentrations, the up-down motion disappeared completely. These results indicate that SERCA does not transit through the shortest structure, and has a catalytic pathway different from the ordinary Albers-Post scheme under physiological conditions. PMID:21707923

  17. Effect of bacoside A on membrane-bound ATPases in the brain of rats exposed to cigarette smoke.

    Science.gov (United States)

    Anbarasi, K; Vani, G; Balakrishna, K; Devi, C S Shyamala

    2005-01-01

    Membrane-bound enzymes play a vital role in neuronal function through maintenance of membrane potential and impulse propagation. We have evaluated the harmful effects of chronic cigarette smoking on membrane-bound ATPases and the protective effect of Bacoside A in rat brain. Adult male albino rats were exposed to cigarette smoke for a period of 12 weeks and simultaneously administered with Bacoside A (the active principle isolated from Bacopa monniera) at a dosage of 10 mg/kg b.w/day, p.o. The levels of lipid peroxides as marker for evaluating the extent of membrane damage, the activities of Na+/K+-ATPase, Ca2+-ATPase and Mg2+-ATPase, and associated cations sodium (Na+), potassium (K+), calcium (Ca2+), and magnesium (Mg2+) were investigated in the brain. Neuronal membrane damage was evident from the elevated levels of lipid peroxides and decreased activities of membrane-bound enzymes. Disturbances in the electrolyte balance with accumulation of Na+ and Ca2+ and depletion of K+ and Mg2+ were also observed. Administration of Bacoside A inhibited lipid peroxidation, improved the activities of ATPases, and maintained the ionic equilibrium. The results of our study indicate that Bacoside A protects the brain from cigarette smoking induced membrane damage.

  18. Oxidative damage increases intracellular free calcium [Ca2+]i concentration in human erythrocytes incubated with lead.

    Science.gov (United States)

    Quintanar-Escorza, M A; González-Martínez, M T; del Pilar, Intriago-Ortega Ma; Calderón-Salinas, J V

    2010-08-01

    One important effect of lead toxicity in erythrocytes consists of increasing [Ca(2+)](i) which in turn may cause alterations in cell shape and volume and it is associated with cellular rigidity, hemolysis, senescence and apoptosis. In this work, we proposed the use of erythrocytes incubated with Pb(2+) to assess association of the mechanisms of lead erythrocyte oxidative damage and calcium homeostasis. Lead incubation produced an increase in [Ca(2+)](i) dose- and time-dependent, which mainly involved Ca(2+) entry mechanism. Additionally, in this in vitro model alterations similar to erythrocytes of lead-exposed workers were produced: Increase in Ca(2+) influx, decrease in (Ca(2+)-Mg(2+))-ATPase activity and GSH/GSGG ratio; increase in lipoperoxidation, protein carbonylation and osmotic fragility accompanied of dramatic morphological changes. Co-incubation with trolox, a soluble vitamin-E analog is able to prevent these alterations indicating that lead damage mechanism is strongly associated with oxidative damage with an intermediate toxic effect via [Ca(2+)](i) increase. Furthermore, erythrocytes oxidation induced with a free radical generator (APPH) showed effects in [Ca(2+)](i) and oxidative damage similar to those found in erythrocytes incubated with lead. Co-incubation with trolox prevents the oxidative effects induced by AAPH in erythrocytes. These results suggest that increase of [Ca(2+)](i) depends on the oxidative status of the erythrocytes incubated with lead. We consider that this model contributes in the understanding of the relation between oxidative damage induced by lead exposure and Ca(2+) homeostasis, the consequences related to these phenomena and the molecular basis of lead toxicity in no excitable cells. PMID:20460147

  19. Differential effects of insecticides on mitochondrial membrane lfuidity and ATPase activity between the wolf spider and the rice stem borer

    Institute of Scientific and Technical Information of China (English)

    LI Hai-ping; CHANG Jing; FENG Tao; GAO Xi-wu

    2015-01-01

    Differential effects of methamidophos and three pyrethroids on ATPase activity and membrane lfuidity of mitochondria were investigated between the wolf spider (Pirata subpiraticus(Boes. et Str.)) and the rice stem borer (Chilo suppressalis (Walker)). Based on a comparison of LD50values, the toxicities of the tested insecticides were higher to the wolf spider than to the rice stem borer. Cyhalothrin at 1×10–4 mmol L–1 caused inhibition of the mitochondrial Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities, and it’s inhibitions on Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities were signiifcantly higher in the wolf spider (44 and 28%) than in the rice stem borer (19 and 11%). Methamidophos at 1×10–4 mmol L–1 decreased Ca2+-Mg2+-ATPase activity by 16 and 27% in the wolf spider and the rice stem borer, respectively, but no signiifcant effect on the speciifc activity of Na+-K+-ATPase was observed. The DPH (1,6-diphenyl-1,3,5-hexatriene) lfuorescence polarization values of mitochondrial membranes were not signiifcantly affected by methamidophos in either species. However, cyhalothrin and alpha-cyperme-thrin induced the values of DPH polarization of mitochondrial membrane increasing with the concentration of cyhalothrin and alpha-cypermethrin from 20 to 100 µmol L–1 in the rice stem borer and the wolf spider. Effect of ethofenprox on lfuidity of the wolf spider and the rice stem borer was contrary. These results suggest that both inhibition of membrane ATPase and changes of membrane lfuidity could be appended to the action mechanisms of pyrethroid insecticides.

  20. Effects of exogenous creatine phosphate on glutamic acid and Ca2+-ATPase activity in brain of mice after exhaustive exercise%外源性磷酸肌酸对游泳力竭小鼠大脑中谷氨酸和钙-ATP酶活力的影响

    Institute of Scientific and Technical Information of China (English)

    马集; 卢畅; 姜茜; 殷林波; 刘彦娜; 刘克敏

    2013-01-01

    Objective:To observe the effects of exogenous creatine phosphate on glutamic acid level and Ca2+-ATPase activity in brain of mice after exhaustive exercise and to further reveal the mechanism of exogenous creatine phosphate in allaying tiredness.Methods:All 36 mice,6-week-age,were divided into 4 groups:exhaustive swimming control group (group A); exhaustive swimming with medication group (group B); 8-min swimming control group (group C);and 8-min swimming with medication group (group D).The method of mice weight-loading swimming was used to sets up the model of exhaustive exercise,and each mouse loaded weight with 6% of the mass of itself.Thirty min before the experiment,mice in groups B and D were given the intraperitoneal injection with creatine phosphate sodium by the standard of 1000 mg/kg,and the mice in groups A and C were given the same proportionate normal saline as placebo.The exhaustive swimming time was recorded,and glutamic level and Ca2+-ATPase activity were measured by using biochemical kits.Results:After testing,the exhaustion time in group B was longer than that in group A (P<0.05).The Glu contents in groups B and D were significantly lower than in groups A and C (P<0.05).Ca2+-ATPase activity in groups B and group D was significantly higher than that in groups A and C (P<0.05).Conclusion:The mechanism of exogenous creatine phosphate in allaying tiredness may be closely related with increased Ca2+-ATPase activity and reduced glutamic level.%目的:研究外源性磷酸肌酸(PCr)对游泳力竭小鼠大脑中谷氨酸(Glu)和钙-ATP酶(Ca2+-ATPase)活力的影响,以进一步揭示PCr的抗疲劳机制.方法:将44只6周龄小鼠分为力竭对照组12只(A组)、力竭给药组12只(B组)、游泳8min对照组10只(C组)、游泳8min给药组10只(D组),采取小鼠负重游泳的力竭运动模型,每只小鼠负重量为自身体质量的6%.于游泳前30min,B、D组小鼠经腹腔注射磷酸肌酸钠溶液1000mg/kg;A、C组小鼠注

  1. Differential expression of P-type ATPases in intestinal epithelial cells: Identification of putative new atp1a1 splice-variant

    International Nuclear Information System (INIS)

    P-type ATPases are membrane proteins that couple ATP hydrolysis with cation transport across the membrane. Ten different subtypes have been described. In mammalia, 15 genes of P-type ATPases from subtypes II-A, II-B and II-C, that transport low-atomic-weight cations (Ca2+, Na+, K+ and H+), have been reported. They include reticulum and plasma-membrane Ca2+-ATPases, Na+/K+-ATPase and H+/K+-ATPases. Enterocytes and colonocytes show functional differences, which seem to be partially due to the differential expression of P-type ATPases. These enzymes have 9 structural motifs, being the phosphorylation (E) and the Mg2+ATP-binding (H) motifs the most preserved. These structural characteristics permitted developing a Multiplex-Nested-PCR (MN-PCR) for the simultaneous identification of different P-type ATPases. Thus, using MN-PCR, seven different cDNAs were cloned from enterocytes and colonocytes, including SERCA3, SERCA2, Na+/K+-ATPase α1-isoform, H+/K+-ATPase α2-isoform, PMCA1, PMCA4 and a cDNA-fragment that seems to be a new cassette-type splice-variant of the atp1a1 gen. PMCA4 in enterocytes and H+/K+-ATPase α2-isoform in colonocytes were differentially expressed. This cell-specific expression pattern is related with the distinctive enterocyte and colonocyte functions.

  2. Characterization of ATPase Activity of Recombinant Human Pif1

    Institute of Scientific and Technical Information of China (English)

    Yu HUANG; Deng-Hong ZHANG; Jin-Qiu ZHOU

    2006-01-01

    Saccharomyces cerevisiae Pif1p helicase is the founding member of the Pif1 subfamily that is conserved from yeast to human. The potential human homolog of the yeast PIF1 gene has been cloned from the cDNA library of the Hek293 cell line. Here, we described a purification procedure of glutathione Stransferase (GST)-fused N terminal truncated human Pif1 protein (hPif1△N) from yeast and characterized the enzymatic kinetics of its ATP hydrolysis activity. The ATPase activity of human Pif1 is dependent on divalent cation, such as Mg2+, Ca2+ and single-stranded DNA. Km for ATP for the ATPase activity is approximately 200 μM. As the ATPase activity is essential for hPif1's helicase activity, these results will facilitate the further investigation on hPif1.

  3. ATP-ases of synaptic plasma membranes in striatum: enzymatic systems for synapses functionality by in vivo administration of L-acetylcarnitine in relation to Parkinson's Disease.

    Science.gov (United States)

    Villa, R F; Ferrari, F; Gorini, A

    2013-09-17

    The maximum rate (Vmax) of some enzymatic activities related to energy consumption was evaluated in synaptic plasma membranes from rat brain striatum, the synaptic energy state being a crucial factor in neurodegenerative diseases etiopathogenesis. Two types of synaptic plasma membranes were isolated from rats subjected to in vivo treatment with L-acetylcarnitine at two different doses (30 and 60 mg × kg(-1) i.p., 28 days, 5 days/week). The following enzyme activities were evaluated: acetylcholinesterase (AChE); Na(+), K(+), Mg(2+)-ATP-ase; ouabain insensitive Mg(2+)-ATP-ase; Na(+), K(+)-ATP-ase; direct Mg(2+)-ATP-ase; Ca(2+), Mg(2+)-ATP-ase; and low- and high-affinity Ca(2+)-ATP-ase. In control (vehicle-treated) animals, enzymatic activities are differently expressed in synaptic plasma membranes type I (SPM1) with respect to synaptic plasma membranes type II (SPM2), the evaluated enzymatic activities being higher in SPM2. Subchronic treatment with L-acetylcarnitine decreased AChE on SPM1 and SPM2 at the dose of 30 mg × kg(-1). Pharmacological treatment decreased ouabain insensitive Mg(2+)-ATP-ase activity and high affinity Ca(2+)-ATP-ase activity at the doses of 30 and 60 mg × kg(-1) respectively on SPM1, while it decreased Na(+), K(+)-ATP-ase, direct Mg(2+)-ATP-ase and Ca(2+), Mg(2+)-ATP-ase activities at the dose of 30 mg × kg(-1) on SPM2. These results suggest that the sensitivity to drug treatment is different between these two populations of synaptic plasma membranes from the striatum, confirming the micro-heterogeneity of these subfractions, possessing different metabolic machinery with respect to energy consumption and utilization and the regional selective effect of L-acetylcarnitine on cerebral tissue, depending on the considered area. The drug potential effect at the synaptic level in Parkinson's Disease neuroprotection is also discussed with respect to acetylcholine and energy metabolism.

  4. 质膜Ca2+-ATP酶异构体2基因多态性与突发性耳聋的关系%Association between poly-morphism of Ca2 + -ATPase isomer 2 gene in plasma membrane and sudden deafness

    Institute of Scientific and Technical Information of China (English)

    邓嘉虹; 金磊

    2016-01-01

    目的:探讨质膜 Ca2+-ATP 酶异构体2(PMCA2)基因的多态性与突发性耳聋的关系。方法采用分组研究的方法,对164名受检者进行调查和听力测试,按听力学评价的结果将其分为感音神经性听力损失的突发性耳聋组(n=82)和听力正常组(n=82);用 PCR 和等位基因特意扩增法检测 PMCA2基因上 rs2289274和 rs6790640两个单核苷酸位点的多态性。结果在突发性耳聋组中,rs2289274位点基因型频率分别为 AA 55.8%,AG 17.4%,GG 26.8%,等位基因频率 A 64.5%和 G 35.5%。在听力正常组中,rs2289274位点基因型频率分别为 AA 26.8%,AG 28.0%,GG 45.2%,等位基因频率 A 41.1%和 G 58.9%。在突发性耳聋组中,rs6790640位点基因型频率分别为 CC 18.3%,CT 35.4%,TT 46.3%,等位基因频率 C 36.3%和 T 63.7%。在听力正常患者组中,rs6790640位点基因型频率分别为 CC 2.4%,CT 63.4%,TT 34.1%,等位基因频率 C 34.1%和 G 65.9%。两位点的基因型分布及其等位基因频率在突发性耳聋组和听力正常患者组之间部分差异有统计学意义(P<0.05)。结论 PMCA2基因 rs2289274和 rs6790640两个单核苷酸位点的多态性可能是突发性耳聋的遗传易感性因素。%Objective To investigate the association between polymorphisms of Ca2 + -ATPase isomer 2 gene (PMCA2) in plasma membrane and the development of sudden deafness .Methods Totally ,164 patients were investigated and hearing tests were conducted .According to the results of audiometry ,they were divided into two groups ,sensorineural hearing loss group(n= 82) and normal hearing group(n= 82) .Polymorphisms of two single nucleotide loci rs2289274 and rs6790640 in the PMCA2 gene were de-termined by polymerase chain reaction followed by allele specific amplication analysis .Results In the sudden deafness group ,fre-quencies of

  5. The Ca2+ Over Loading Effect of Oxidized Low Density Lipoprotein on Macrophages

    Institute of Scientific and Technical Information of China (English)

    谭健苗; 杨向东; 姜志胜; 李亮

    2007-01-01

    Purpose The Ca2+ over loading effect of oxidized low density lipoprotein (OLDL) on macrophages and its mechanisms were explored in these studies. Methods C57BL/6J mouse peritoneal macrophages were incubated in 10mg/L oxidized low density lipoprotein (OLDL), the intracellular Ca2+ level was determined with the technique of Ca2+ fluorescent indicator, and the membranous Ca2+-ATPase activity was determined with the assay of NADH-oxidizing coupling spectrum-alteration. Results The intracellular Ca2+ level was 2.7 times higher than the control, and the membranous Ca2+-ATPase activity was 24.0% of the control. Conclusion OLDL exerted a Ca2+ over loading effect on macrophages, and this was probably related to the opening of the membranous Ca2+ channels and the inactivating of the membranous Ca2+-ATPase.

  6. Stabilization of membrane bound ATPases and lipid peroxidation by carotenoids from Chlorococcum humicola in Benzo(a)pyrene induced toxicity

    Institute of Scientific and Technical Information of China (English)

    Bhagavathy S; Sumathi P

    2012-01-01

    Objective: To identify the alteration of the membrane potential and the effect of carotenoid extracts from Chlorococcum humicola (C. humicola) on membrane bound ATPases and lipid peroxidation. Methods: The total carotenoids were extracted from C. humicola. Four groups of Swiss albino mice were treated as control, Benzo(a)pyrene [B(a)P], total carotenoids, B(a)P +total carotenoids respectively for a period of 60 days. Membrane lipid peroxidation and ATPases (Total ATPases, Ca2+- ATPases, Mg2+ - ATPases, Na+K+ - ATPase) were determined in lung, liver and erythrocyte samples. Results: The activity of total ATPase was found to be significantly increased in the B(a)P treated liver and lung tissue. Erythrocyte membrane also showed higher ATPase activity which was significantly reverted on total carotenoid treatment. Conclusions:It can be concluded that the changes in membrane potential favour the functional deterioration of physiological system. The overall findings demonstrates that the animals post treated with carotenoid extract from C. humicola may maintains the alterations in membrane bound ATPase and lipid peroxidation in tissues against the carcinogenic chemical and hence aid in establishing the membrane potential action. Therefore C. humicola can be further extended to exploits its possible application for various health benefits as neutraceuticals and food additives.

  7. Effects of exercise training at different work intensities on the activity of Na+,K+-ATPase and Ca2+-ATP in rat skeletal muscle sarcoplasmic reticulum%骨骼肌肌浆网Na+,K+-ATP酶和Ca2+-ATP酶活性在不同训练条件下的变化

    Institute of Scientific and Technical Information of China (English)

    张敏; 陈立军; 史娜; 周蔚

    2010-01-01

    背景:Na+,K+-ATP酶和Ca2+-ATP酶存物质运送、能量转换以及信息传递方面具有重要作用.肌浆网在肌肉兴奋-收缩耦联过程中起关键作用,与运动性骨骼肌疲劳的发生密切关.目的:通过建立SD大鼠有氧和无氧训练模型,观察不同训练负荷条件对大鼠骨骼肌肌浆网Na+,K+-ATP酶和Ca2+-ATP酶活性的影响.方法:参照Bedford TG标准,建立有氧和无氧运动大鼠跑台训练模型,有氧运动组采用递增负荷训练,无氧运动组采用高速间歇训练,正常对照组大鼠正常笼内生活,不运动.各组动物训练结束后用超速离心法提取大鼠骨骼肌肌浆网,紫外分光光度汁检测大鼠骨骼肌肌浆网Na+,K+-ATP酶和Ca2+-ATP酶的活性.结果与结论:训练4周后,两个运动组大鼠骨骼肌肌浆网Na+,K+-ATP酶和Ca2+-ATP酶的活性逐渐升高(P<0.05);训练6周,仅有氧运动组升高(P<0.05),无氧运动组则活性降低(P<0.05).结果提示有氧训练更有利十保护大鼠骨骼肌肌浆网Na+,K+-ATP酶和Ca2+-ATP酶的活性,但需要一定的时间累积.

  8. Influence of valsartan on sarcoplasmic reticulum Ca2+-ATPase and phospholamban in rats with dilated cardiomyopathy%缬沙坦对扩张型心肌病心衰大鼠肌浆网Ca2+-ATP酶及其调控蛋白的影响

    Institute of Scientific and Technical Information of China (English)

    尹春阳; 李卫东; 王成华; 俞捷; 汪建飞; 张国辉

    2010-01-01

    目的 探讨血管紧张素Ⅱ1型受体阻滞剂(ARB)缬沙坦对扩张型心肌病(DCM)心衰大鼠心肌肌浆网Ca2+-ATP酶(SERCA2a)及其调控蛋白(PLB)的影响及意义.方法 腹腔注射多柔比星建立SD大鼠DCM模型.随机分为DCM组(M组,n=11)、缬沙坦组(V组,n=10)、另设正常对照组(C组,n=10),V组予以缬沙坦30 mg·kg-1·d-1,C组和M组予以生理盐水灌胃,8周后行血流动力学检测,采用RT-PCR和Western-blot法分别检测心肌组织SERCA2a、PLB的mRNA和蛋白含量.结果 与正常对照组相比,DCM组左心室压峰值(LVSP)、左室最大压力上升及下降速度(±dp/dtmax)显著下降(均P0.05).结论 缬沙坦改善DCM心功能可以与其部分纠正SERCA2a、PLB的异常有关.

  9. Substantial depletion of the intracellular Ca2+ stores is required for macroscopic activation of the Ca2+ release-activated Ca2+ current in rat basophilic leukaemia cells.

    Science.gov (United States)

    Fierro, L; Parekh, A B

    2000-01-15

    1. Tight-seal whole-cell patch clamp experiments were performed to examine the ability of different intracellular Ca2+ mobilising agents to activate the Ca2+ release-activated Ca2+ current (ICRAC) in rat basophilic leukaemia (RBL-1) cells under conditions of weak cytoplasmic Ca2+ buffering. 2. Dialysis with a maximal concentration of inositol 1,4,5-trisphosphate (IP3) routinely failed to activate macroscopic ICRAC in low buffer (0.mM EGTA, BAPTA or dimethyl BAPTA), whereas it activated the current to its maximal extent in high buffer (10 mM EGTA). Dialysis with a poorly metabolisable analogue of IP3, with ionomycin, or with IP3 and ionomycin all failed to generate macroscopic ICRAC in low Ca2+ buffering conditions. 3. Dialysis with the sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) pump blocker thapsigargin was able to activate ICRAC even in the presence of low cytoplasmic Ca2+ buffering, albeit at a slow rate. Exposure to IP3 together with the SERCA blockers thapsigargin, thapsigargicin or cyclopiazonic acid rapidly activated ICRAC in low buffer. 4. Following activation of ICRAC by intracellular dialysis with IP3 and thapsigargin in low buffer, the current was very selective for Ca2+ (apparent KD of 1 mM) Sr2+ and Ba2+ were less effective charge carriers and Na+ was not conducted to any appreciable extent. The ionic selectivity of ICRAC was very similar in low or high intracellular Ca2+ buffer. 5. Fast Ca2+-dependent inactivation of ICRAC occurred at a similar rate and to a similar extent in low or high Ca2+ buffer. Ca2+-dependent inactivation is not the reason why macroscopic ICRAC cannot be seen under conditions of low cytoplasmic Ca2+ buffering. 6. ICRAC could be activated by combining IP3 with thapsigargin, even in the presence of 100 microM Ca2+ and the absence of any exogenous Ca2+ chelator, where ATP and glutamate represented the only Ca2+ buffers in the pipette solution. 7. Our results suggest that a threshold exists within the IP3-sensitive Ca2+ store

  10. Plasmalemmal Na+/Ca2+ exchanger modulates Ca2+-dependent exocytotic release of glutamate from rat cortical astrocytes

    Directory of Open Access Journals (Sweden)

    Reno C Reyes

    2012-01-01

    Full Text Available Astroglial excitability operates through increases in Ca2+cyt (cytosolic Ca2+, which can lead to glutamatergic gliotransmission. In parallel fluctuations in astrocytic Na+cyt (cytosolic Na+ control metabolic neuronal-glial signalling, most notably through stimulation of lactate production, which on release from astrocytes can be taken up and utilized by nearby neurons, a process referred to as lactate shuttle. Both gliotransmission and lactate shuttle play a role in modulation of synaptic transmission and plasticity. Consequently, we studied the role of the PMCA (plasma membrane Ca2+-ATPase, NCX (plasma membrane Na+/Ca2+ exchanger and NKA (Na+/K+-ATPase in complex and coordinated regulation of Ca2+cyt and Na+cyt in astrocytes at rest and upon mechanical stimulation. Our data support the notion that NKA and PMCA are the major Na+ and Ca2+ extruders in resting astrocytes. Surprisingly, the blockade of NKA or PMCA appeared less important during times of Ca2+ and Na+ cytosolic loads caused by mechanical stimulation. Unexpectedly, NCX in reverse mode appeared as a major contributor to overall Ca2+ and Na+ homoeostasis in astrocytes both at rest and when these glial cells were mechanically stimulated. In addition, NCX facilitated mechanically induced Ca2+-dependent exocytotic release of glutamate from astrocytes. These findings help better understanding of astrocyte-neuron bidirectional signalling at the tripartite synapse and/or microvasculature. We propose that NCX operating in reverse mode could be involved in fast and spatially localized Ca2+-dependent gliotransmission, that would operate in parallel to a slower and more widely distributed gliotransmission pathway that requires metabotropically controlled Ca2+ release from the ER (endoplasmic reticulum.

  11. Kinetics of Ca2+ carrier in rat liver mitochondria.

    Science.gov (United States)

    Bragadin, M; Pozzan, T; Azzone, G F

    1979-12-25

    The rate of aerobic Ca2+ transport is limited by the rate of the H+ pump rather than by the Ca2+ carrier. The kinetics of the Ca2+ carrier has therefore been studied by using the K+ diffusion potential as the driving force. The apparent Vmax of the Ca2+ carrier is, at 20 degrees C, about 900 nmol (mg of protein)-1 min-1, more than twice the rate of the H+ pump. The apparent Vmax is depressed by Mg2+ and Li+. This supports the view that the electrolytes act as noncompetitive inhibitors of the Ca2+ carrier. The degree of sigmoidicity of the kinetics of Ca2+ transport increases with the lowering of the temperature and proportionally with the concentration of impermeant electrolytes such as Mg2+ and Li+ but not choline. The effects of temperature and of electrolyte do not support the view that the sigmoidicity is due to modifications of the surface potential. Rather, they suggest that Ca2+ transport occurs through a multisubunit carrier, where cooperative phenomena are the result of ligand-induced conformational changes due to the interaction of several allosteric effectors with the carrier subunits. In contrast with La3+ which acts as a competitive inhibitor, Ruthenium Red affects the kinetics by inducing phenomena both of positive and of negative cooperativity. The Ruthenium Red induced kinetics has been reproduced through curve-fitting procedures by applying the Koshland sequential interaction hypothesis to a four-subunit Ca2+ carrier model. PMID:42437

  12. Dietary calcium attenuates platelet aggregation and intracellular Ca2+ mobilization in spontaneously hypertensive rats

    Science.gov (United States)

    Otsuka, K.; Watanabe, M.; Yue, Q.; McCarron, D. A.; Hatton, D.

    1997-01-01

    Spontaneously hypertensive rats (SHR) are known to be blood pressure sensitive to dietary calcium. The effects of dietary calcium on platelet aggregation and intracellular Ca2+ mobilization were assessed by turbidimetric methods and fura-2 methods, respectively, in washed platelets of SHR. Ca2+ ATPase activity was examined in aortic membrane fractions. Six weeks of dietary calcium supplementation attenuated the increase of systolic blood pressure (SBP 199 +/- 16 v 170 +/- 9 mm Hg, P ionomycin-induced intracellular calcium ([Ca2+]i) peak in the absence of external Ca2+, which reflects [Ca2+]i storage size, and thrombin-evoked [Ca2+]i release from [Ca2+]i storage were decreased by 2.0% Ca diet (472 +/- 55 v 370 +/- 23 nmol/L, P ionomycin-induced [Ca2+]i (r = 0.591, P = .0415), respectively. However, there was no significant effect of dietary calcium on Ca2+-ATPase activity in aortic membranes. These results suggest that dietary calcium supplementation had a beneficial effect on platelets of SHR by attenuating [Ca2+]i mobilization from [Ca2+]i storage. The hypotensive effect of dietary calcium might be associated with attenuated [Ca2+]i mobilization in SHR.

  13. Study of the effect of Chinese herbal drugs upon the activities of Na + -K + -ATPase, Ca2 + -ATPase and ECG in cardiac muscle of rats damaged by nickel sulfate%中药防治硫酸镍对大鼠两种心肌酶活性及心电图影响的实验研究 .

    Institute of Scientific and Technical Information of China (English)

    赵健雄; 朱玉真; 孙应彪; 王学习; 黄渊

    2001-01-01

    目的探讨扶正解毒汤(FJD)对硫酸镍(NiSO4)致心脏损伤的防治作用及其机理.方法腹腔注射NiSO4染毒,FJD灌胃防治,检测心肌Na+-K+-ATP酶、Ca2+-ATP酶活性,心电图和心肌病理,进行评价.结果NiSO4组与NS组比较,两种酶活性均明显抑制(P<0.001),同时出现心肌病理和心电图异常.用FJD防治后,两种酶活性明显改善(P<0.01),心肌病理损伤和心电图恢复正常.结论FJD可有效防治NiSO4对心肌的损伤,其机理与恢复心肌Na+-K+-ATP酶、Ca2+-ATP酶活性有关.

  14. Effects of C-terminal truncations on trafficking of the yeast plasma membrane H+-ATPase.

    Science.gov (United States)

    Mason, A Brett; Allen, Kenneth E; Slayman, Carolyn W

    2006-08-18

    Within the large family of P-type cation-transporting ATPases, members differ in the number of C-terminal transmembrane helices, ranging from two in Cu2+-ATPases to six in H+-, Na+,K+-, Mg2+-, and Ca2+-ATPases. In this study, yeast Pma1 H+-ATPase has served as a model to examine the role of the C-terminal membrane domain in ATPase stability and targeting to the plasma membrane. Successive truncations were constructed from the middle of the major cytoplasmic loop to the middle of the extended cytoplasmic tail, adding back the C-terminal membrane-spanning helices one at a time. When the resulting constructs were expressed transiently in yeast, there was a steady increase in half-life from 70 min in Pma1 delta452 to 348 min in Pma1 delta901, but even the longest construct was considerably less stable than wild-type ATPase (t(1/2) = 11 h). Confocal immunofluorescence microscopy showed that 11 of 12 constructs were arrested in the endoplasmic reticulum and degraded in the proteasome. The only truncated ATPase that escaped the ER, Pma1 delta901, traveled slowly to the plasma membrane, where it hydrolyzed ATP and supported growth. Limited trypsinolysis showed Pma1 delta901 to be misfolded, however, resulting in premature delivery to the vacuole for degradation. As model substrates, this series of truncations affirms the importance of the entire C-terminal domain to yeast H+-ATPase biogenesis and defines a sequence element of 20 amino acids in the carboxyl tail that is critical to ER escape and trafficking to the plasma membrane.

  15. Role of inositol (1,4,5)trisphosphate in epidermal growth factor-induced Ca2+ signaling in A431 cells

    DEFF Research Database (Denmark)

    Hughes, A R; Bird, G S; Obie, J F;

    1991-01-01

    + as well as the ability to increase Ca2+ entry from the extracellular space. The Ca(2+)-ATPase inhibitor thapsigargin also activated Ca2+ entry, and neither epidermal growth factor nor the guanine nucleotide-dependent protein-linked receptor agonist bradykinin activated additional Ca2+ entry over that due...... activates intracellular Ca2+ release as well as Ca2+ entry, the latter most likely resulting from an indirect effect due to the depletion of intracellular Ca2+ pools, (ii) that the actions of epidermal growth factor on Ca2+ homeostasis can be fully accounted for by inositol 1,4,5-trisphosphate formation...

  16. ATP hydrolysis is critically required for function of CaV1.3 channels in cochlear inner hair cells via fueling Ca2+ clearance.

    Science.gov (United States)

    Weiler, Simon; Krinner, Stefanie; Wong, Aaron B; Moser, Tobias; Pangršič, Tina

    2014-05-14

    Sound encoding is mediated by Ca(2+) influx-evoked release of glutamate at the ribbon synapse of inner hair cells. Here we studied the role of ATP in this process focusing on Ca(2+) current through CaV1.3 channels and Ca(2+) homeostasis in mouse inner hair cells. Patch-clamp recordings and Ca(2+) imaging demonstrate that hydrolyzable ATP is essential to maintain synaptic Ca(2+) influx in inner hair cells via fueling Ca(2+)-ATPases to avoid an increase in cytosolic [Ca(2+)] and subsequent Ca(2+)/calmodulin-dependent inactivation of CaV1.3 channels.

  17. Cellular mechanisms of reduced sarcoplasmic reticulum Ca2+ content in L-thyroxin-induced rat ventricular hypertrophy

    Institute of Scientific and Technical Information of China (English)

    Lai-jing SONG; Guan-lei WANG; Jie LIU; Qin-ying QIU; Jing-hua OU; Yong-yuan GUAN

    2008-01-01

    Aim:To examine how the sarcoplasmic reticulum (SR) Ca2+ content changes and the underlying mechanism in L-thyroxin-induced cardiac hypertrophy. Methods:Echocardiography was used to confirm the establishment of the cardiac hypertro-phy model. The confocal microscopy and fluorescent indicator Fluo-3 was ap-plied to examine the intracellular Ca2+ concentration ([Ca2+]I), the Ca2+ sparks, and the caffeine-induced Ca2+ transient in freshly isolated cardiac ventricular myocytes. The activity of sarcolemmal and SR Ca2+-ATPase 2a (SERCA2a) in the ventricular tissue was also measured, respectively. Results:L-thyroxin (1 mg/kg injection for 10 d) induces left ventricular cardiac hypertrophy with normal myocardial function. The decreased caffeine-induced Ca2+ transient in the Ca2+-free solution was detected. The spontaneous Ca2+ sparks in hypertrophied myocytes occurred more frequently than in normal cells, with similar duration and spatial spread, but smaller amplitude. Then the basal [Ca2+]I increase was observed in quiescent left ventricular myocytes from hyperthyroidism rats. The activity of sarcolemmal and SR Ca2+-ATPase was decreased in the hypertrophied ventricle tissue. Conclusion:The results suggested that the reduced SR Ca2+ content may be associated with an increased Ca2+ leak and reduced SERCA2a activity, contributing to abnormal intracellular Ca2+ handling during hypertrophy in hyperthyroidism rats.

  18. State-dependent firing determines intrinsic dendritic Ca2+ signaling in thalamocortical neurons.

    Science.gov (United States)

    Errington, Adam C; Renger, John J; Uebele, Victor N; Crunelli, Vincenzo

    2010-11-01

    Activity-dependent dendritic Ca(2+) signals play a critical role in multiple forms of nonlinear cellular output and plasticity. In thalamocortical neurons, despite the well established spatial separation of sensory and cortical inputs onto proximal and distal dendrites, respectively, little is known about the spatiotemporal dynamics of intrinsic dendritic Ca(2+) signaling during the different state-dependent firing patterns that are characteristic of these neurons. Here we demonstrate that T-type Ca(2+) channels are expressed throughout the entire dendritic tree of rat thalamocortical neurons and that they mediate regenerative propagation of low threshold spikes, typical of, but not exclusive to, sleep states, resulting in global dendritic Ca(2+) influx. In contrast, actively backpropagating action potentials, typical of wakefulness, result in smaller Ca(2+) influxes that can temporally summate to produce dendritic Ca(2+) accumulations that are linearly related to firing frequency but spatially confined to proximal dendritic regions. Furthermore, dendritic Ca(2+) transients evoked by both action potentials and low-threshold spikes are shaped by Ca(2+) uptake by sarcoplasmic/endoplasmic reticulum Ca(2+) ATPases but do not rely on Ca(2+)-induced Ca(2+) release. Our data demonstrate that thalamocortical neurons are endowed with intrinsic dendritic Ca(2+) signaling properties that are spatially and temporally modified in a behavioral state-dependent manner and suggest that backpropagating action potentials faithfully inform proximal sensory but not distal corticothalamic synapses of neuronal output, whereas corticothalamic synapses only "detect" Ca(2+) signals associated with low-threshold spikes.

  19. State-dependent firing determines intrinsic dendritic Ca2+ signaling in thalamocortical neurons.

    Science.gov (United States)

    Errington, Adam C; Renger, John J; Uebele, Victor N; Crunelli, Vincenzo

    2010-11-01

    Activity-dependent dendritic Ca(2+) signals play a critical role in multiple forms of nonlinear cellular output and plasticity. In thalamocortical neurons, despite the well established spatial separation of sensory and cortical inputs onto proximal and distal dendrites, respectively, little is known about the spatiotemporal dynamics of intrinsic dendritic Ca(2+) signaling during the different state-dependent firing patterns that are characteristic of these neurons. Here we demonstrate that T-type Ca(2+) channels are expressed throughout the entire dendritic tree of rat thalamocortical neurons and that they mediate regenerative propagation of low threshold spikes, typical of, but not exclusive to, sleep states, resulting in global dendritic Ca(2+) influx. In contrast, actively backpropagating action potentials, typical of wakefulness, result in smaller Ca(2+) influxes that can temporally summate to produce dendritic Ca(2+) accumulations that are linearly related to firing frequency but spatially confined to proximal dendritic regions. Furthermore, dendritic Ca(2+) transients evoked by both action potentials and low-threshold spikes are shaped by Ca(2+) uptake by sarcoplasmic/endoplasmic reticulum Ca(2+) ATPases but do not rely on Ca(2+)-induced Ca(2+) release. Our data demonstrate that thalamocortical neurons are endowed with intrinsic dendritic Ca(2+) signaling properties that are spatially and temporally modified in a behavioral state-dependent manner and suggest that backpropagating action potentials faithfully inform proximal sensory but not distal corticothalamic synapses of neuronal output, whereas corticothalamic synapses only "detect" Ca(2+) signals associated with low-threshold spikes. PMID:21048143

  20. Effects of mibefradil on intracellular Ca2+ release in cultured rat cardiac fibroblasts and human platelets.

    Science.gov (United States)

    Eberhard, M; Miyagawa, K; Hermsmeyer, K; Erne, P

    1995-12-01

    The Ca2+ antagonist mibefradil at supratherapeutic concentrations induced a sustained increase of cytosolic Ca2+ in cultured rat cardiac fibroblasts and human platelets which lack sensitivity to K+ depolarization and Ca2+ channel block by verapamil or other Ca2+ antagonists. At concentrations above 10 microM, mibefradil elevated substantially cytosolic [Ca2+] without affecting the peak level of agonist-induced Ca2+ transients. These Ca2+-mobilizing actions of 10 or 100 microM mibefradil stand in contrast to the Ca2+ antagonism and relaxation of vascular muscle at 1 microM concentrations. Since a substantial part of mibefradil-induced increase in cytosolic Ca2+ was independent of extracellular Ca2+, and in order to define better the mechanism of Ca2+ increase, we exposed permeabilized cultured rat cardiac fibroblasts and human platelets to mibefradil at concentrations sufficiently high to identify covert effects. In permeabilized fibroblasts or platelets mibefradil at concentrations above 10 microM activated dose-dependent Ca2+ release from intracellular Ca2+ stores. Verapamil had no effect at concentrations of up to 100 microM. Mibefradil-induced Ca2+ release was not affected by ryanodine, thapsigargin, removal of ATP or dithioerythreitol, indicating that neither Ca2+ - nor disulfide reagent-induced Ca2+ release were involved and that mibefradil did not release Ca2+ by inhibition of the Ca2+-ATPase pump of endoplasmic reticulum. The rate, but not the amplitude, of mibefradil-induced Ca2+ release is increased up to fourfold in the presence of pentosan polysulphate or heparin, two potent inhibitors of inositol 1,4,5-trisphosphate-induced Ca2+ release. Depletion of Ca2+ stores of permeabilized cells inositol 1,4,5-trisphosphate in the presence of thapsigargin completely blocked mibefradil-induced Ca2+ release, and depletion of Ca2+ stores by mibefradil prevented further Ca2+ release by inositol 1,4,5-trisphosphate. Mibefradil at supratherapeutic concentrations (> or

  1. Diversity and regulation of plant Ca2+ pumps: insights from expression in yeast

    Science.gov (United States)

    Sze, H.; Liang, F.; Hwang, I.; Curran, A. C.; Harper, J. F.; Evans, M. L. (Principal Investigator)

    2000-01-01

    The spatial and temporal regulation of calcium concentration in plant cells depends on the coordinate activities of channels and active transporters located on different organelles and membranes. Several Ca2+ pumps have been identified and characterized by functional expression of plant genes in a yeast mutant (K616). This expression system has opened the way to a genetic and biochemical characterization of the regulatory and catalytic features of diverse Ca2+ pumps. Plant Ca(2+)-ATPases fall into two major types: AtECA1 represents one of four or more members of the type IIA (ER-type) Ca(2+)-ATPases in Arabidopsis, and AtACA2 is one of seven or more members of the type IIB (PM-type) Ca(2+)-ATPases that are regulated by a novel amino terminal domain. Type IIB pumps are widely distributed on membranes, including the PM (plasma membrane), vacuole, and ER (endoplasmic reticulum). The regulatory domain serves multiple functions, including autoinhibition, calmodulin binding, and sites for modification by phosphorylation. This domain, however, is considerably diverse among several type IIB ATPases, suggesting that the pumps are differentially regulated. Understanding of Ca2+ transporters at the molecular level is providing insights into their roles in signaling networks and in regulating fundamental processes of cell biology.

  2. Effect of human proinsulin C-peptide on erythrocyte Na+-K+-ATPase activity and Ca2+concentration in patients with type H diabetes mellitus in vitro%胰岛素原C肽对2型糖尿病患者离体红细胞膜Na+-K+-ATP酶活性及红细胞内Ca2+浓度的影响

    Institute of Scientific and Technical Information of China (English)

    李素霞; 袁勤生

    2006-01-01

    目的研究外源性胰岛素原C肽对2型糖尿病患者离体红细胞膜Na+-K+-ATP酶活性及红细胞内Ca2+浓度的影响.方法取2型糖尿病患者空腹血,肝素抗凝,分离红细胞,生理盐水重新悬浮,加外源性C肽,分别测定细胞膜上的Na+-K+-ATP酶活性和细胞内Ca2+浓度.结果外源性C肽的加入可升高2型糖尿病患者红细胞膜的Na+-K+-ATP酶活性,并降低红细胞内的Ca2+浓度.结论胰岛素原C肽可以改善2型糖尿病患者降低的红细胞膜Na+-K+-ATP酶活性.

  3. Store-operated Ca(2+) entry in rhabdomyosarcoma cells.

    Science.gov (United States)

    Schmid, Evi; Stagno, Matias Julian; Yan, Jing; Stournaras, Christos; Lang, Florian; Fuchs, Jörg; Seitz, Guido

    2016-08-12

    Rhabdomyosarcoma (RMS), the most common pediatric soft tissue sarcoma, has an intrinsic or early-acquisition of resistance to chemo- and radiation therapy. Molecular determinants pivotal for RMS migration, metastatic invasion, cell proliferation, and survival are incompletely identified. Migration and cell proliferation were shown to correlate with cytosolic Ca(2+) activity ([Ca(2+)]i). Store-operated Ca(2+)-entry (SOCE) that increases intracellular [Ca(2+)] is accomplished by Orai1, a pore-forming ion channel unit, the expression of which is stimulated by the transcription factor NFκB. The present study explored the expression of Orai1 and its regulators STIM1 and NFκB in human rhabdomyosarcoma cell lines and analyzed their impact on cell proliferation and migration. For the study human rhabdomyosarcoma cell lines RD (embryonal) and RH30 (alveolar) were analyzed for Orai1, STIM1, and NFκB transcription by RT-PCR and their corresponding proteins in Western blot. [Ca(2+)]i was detected via Fura-2 fluorescence and SOCE - resulting from [Ca(2+)]i increase following store depletion with extracellular Ca(2+) removal and inhibition of the sarcoendoplasmatic reticular Ca(2+) ATPase - detected with thapsigargin. Cell migration was analyzed in transwell and mitotic cell death with the clonogenic assay. In summary, Orai1, STIM1, and NFκB are expressed in embryonal (RD) and alveolar (RH30) rhabdomyosarcoma. SOCE inhibitor BTP2, Orai1 inhibitor 2-APB, or NFκB inhibitor wogonin virtually abrogated (BTP2, 2-APB) or significantly reduced (wogonin) SOCE. Moreover, SOCE inhibitors 2-APB and BTP2 and wogonin significantly inhibited migration and proliferation of both, RD and RH30 cells. These results suggest that Orai1 signaling is involved in SOCE into rhabdomyosarcoma cells thus contributing to migration, invasion and proliferation. PMID:27291153

  4. 肌浆网/内质网钙ATP酶在大鼠肝再生过程中的表达和活性分析%Expression Patterns and Activity Analysis of the Sarco (endo) Plasmic Reticulum Ca2+-ATPase in the Liver Regeneration of Rats

    Institute of Scientific and Technical Information of China (English)

    吴灿; 侯国俊; 支樑健; 王颖; 叶波平

    2012-01-01

    肌浆网/内质网钙ATP酶(SERCA)主要位于内质网和肌浆网膜上,是生物体内重要的钙离子转运酶.为进一步探讨肝再生过程中SERCA的表达变化,该研究构建了大鼠2/3肝脏切除模型,用实时荧光定量RT-PCR和测定酶活性的方法研究该酶的两种亚型(SERCA2b和SERCA3)在肝切除后O,1,12,24,48,72,120,168,216 h共9个时间点的表达变化,结果发现肝再生早期两种亚型表达量均下调,但在一定时间内显著应激性上升,分别在12h和48h达到峰值,之后表达量下调至原始水平.同时,酶活性测定结果与基因表达变化相类似,在12~48 h酶活性达到峰值,以上结果表明SERCA参与了肝脏再生过程,可能的途径是调控内质网钙离子,进而影响细胞质的钙震荡和细胞周期.%Sarco ( endo) plasmic reticulum Ca2+-ATPase, SERCA, is thought to play an important role in Ca2+-transporting. It is mainly located in the membranes of sarco ( endo) plasmic reticulum. To characterize its role in liver regeneration, the expression patterns of its two isoforms( SERCA2b,SERCA3) at different time points of 0,1 ,12,24,48,72,120,168,216 h were studied after 2/3 partial hepatectomy(PH) by Real-time quantiatative PCR. And the total enzyme activity was also determined. The results showed that the mRNA level of the two isoforms was down-regulated at the early stage of liver regeneration. However they increased dramtically responding to the PH. SERCA2b peaked at 12 h and the SERCA3 peaked at 48 h. After reaching the peak,the level decreased to the normal. The change of total enzyme activity was consistent with the mRNA level and kept the highest level between the 12~48 h. All these implied that SERCA involved in the liver regeneration. It may play an important role in charging the calcium oscillatory in the cytoplasmic and cell cycle by regulateing the calcium in the ER.

  5. Subcellular localization and kinetic characterization of a gill (Na+, K+)-ATPase from the giant freshwater prawn Macrobrachium rosenbergii.

    Science.gov (United States)

    França, Juliana L; Pinto, Marcelo R; Lucena, Malson N; Garçon, Daniela P; Valenti, Wagner C; McNamara, John C; Leone, Francisco A

    2013-07-01

    The stimulation by Mg(2+), Na(+), K(+), NH4 (+), and ATP of (Na(+), K(+))-ATPase activity in a gill microsomal fraction from the freshwater prawn Macrobrachium rosenbergii was examined. Immunofluorescence labeling revealed that the (Na(+), K(+))-ATPase α-subunit is distributed predominantly within the intralamellar septum, while Western blotting revealed a single α-subunit isoform of about 108 kDa M r. Under saturating Mg(2+), Na(+), and K(+) concentrations, the enzyme hydrolyzed ATP, obeying cooperative kinetics with V(M) = 115.0 ± 2.3 U mg(-1), K(0.5) = 0.10 ± 0.01 mmol L(-1). Stimulation by Na(+) (V(M) = 110.0 ± 3.3 U mg(-1), K(0.5) = 1.30 ± 0.03 mmol L(-1)), Mg(2+) (V(M) = 115.0 ± 4.6 U mg(-1), K(0.5) = 0.96 ± 0.03 mmol L(-1)), NH4 (+) (V(M) = 141.0 ± 5.6 U mg(-1), K(0.5) = 1.90 ± 0.04 mmol L(-1)), and K(+) (V(M) = 120.0 ± 2.4 U mg(-1), K(M) = 2.74 ± 0.08 mmol L(-1)) followed single saturation curves and, except for K(+), exhibited site-site interaction kinetics. Ouabain inhibited ATPase activity by around 73% with K(I) = 12.4 ± 1.3 mol L(-1). Complementary inhibition studies suggest the presence of F0F1-, Na(+)-, or K(+)-ATPases, but not V(H(+))- or Ca(2+)-ATPases, in the gill microsomal preparation. K(+) and NH4(+) synergistically stimulated enzyme activity (≈25%), suggesting that these ions bind to different sites on the molecule. We propose a mechanism for the stimulation by both NH4(+), and K(+) of the gill enzyme.

  6. Biochemical and ultrastructural aspects of Ca2+ transport by mitochondria of the hepatopancreas of the blue crab Callinectes sapidus.

    Science.gov (United States)

    Chen, C H; Greenawalt, J W; Lehninger, A L

    1974-05-01

    Mitochondria isolated from the hepatopancreas of the blue crab Callinectes sapidus show up to 12-fold stimulation of respiration on addition of Ca(2+), which is accompanied by Ca(2+) accumulation (Ca(2+):site = 1.9) and H(+) ejection (H(+):Ca(2+) = 0.85). Sr(2+) and Mn(2+) are also accumulated; Mg(2+) is not. A strongly hypertonic medium (383 mosM), Mg(2+), and phosphate are required for maximal Ca(2+) uptake. Ca(2+) uptake takes precedence over oxidative phosphorylation of ADP for respiratory energy. Once Ca(2+) is accumulated by the crab mitochondria, it is stable and only very slowly released, even by uncoupling agents. ATP hydrolysis also supports Ca(2+) uptake. Respiration-inhibited crab hepatopancreas mitochondria show both high-affinity and low-affinity Ca(2+)-binding sites, which are inactive in the presence of uncoupling agents. Crab hepatopancreas mitochondria have an enormous capacity for accumulation of Ca(2+), up to 5,500 ng-atoms Ca(2+) per mg protein, with an equivalent amount of phosphate. Freshly isolated mitochondria contain very large amounts of Ca(2+), Mg(2+), phosphate, K(+), and Na(+); their high Ca(2+) content is a reflection of the vary large amount of extra-mitochondrial Ca(2+) in the whole tissue. Electron microscopy of crab mitochondria loaded with Ca(2+) and phosphate showed large electron-dense deposits, presumably of precipitated calcium phosphate. They consisted of bundles of needle-like crystals, whereas Ca(2+)-loaded rat liver mitochondria show only amorphous deposits of calcium phosphate under similar conditions. The very pronounced capacity of crab hepatopancreas mitochondria for transport of Ca(2+) appears to be adapted to a role in the storage and release of Ca(2+) during the molting cycle of this crustacean.

  7. Routes of Ca2+ Shuttling during Ca2+ Oscillations: FOCUS ON THE ROLE OF MITOCHONDRIAL Ca2+ HANDLING AND CYTOSOLIC Ca2+ BUFFERS.

    Science.gov (United States)

    Pecze, László; Blum, Walter; Schwaller, Beat

    2015-11-20

    In some cell types, Ca(2+) oscillations are strictly dependent on Ca(2+) influx across the plasma membrane, whereas in others, oscillations also persist in the absence of Ca(2+) influx. We observed that, in primary mesothelial cells, the plasmalemmal Ca(2+) influx played a pivotal role. However, when the Ca(2+) transport across the plasma membrane by the "lanthanum insulation method" was blocked prior to the induction of the serum-induced Ca(2+) oscillations, mitochondrial Ca(2+) transport was found to be able to substitute for the plasmalemmal Ca(2+) exchange function, thus rendering the oscillations independent of extracellular Ca(2+). However, in a physiological situation, the Ca(2+)-buffering capacity of mitochondria was found not to be essential for Ca(2+) oscillations. Moreover, brief spontaneous Ca(2+) changes were observed in the mitochondrial Ca(2+) concentration without apparent changes in the cytosolic Ca(2+) concentration, indicating the presence of a mitochondrial autonomous Ca(2+) signaling mechanism. In the presence of calretinin, a Ca(2+)-buffering protein, the amplitude of cytosolic spikes during oscillations was decreased, and the amount of Ca(2+) ions taken up by mitochondria was reduced. Thus, the increased calretinin expression observed in mesothelioma cells and in certain colon cancer might be correlated to the increased resistance of these tumor cells to proapoptotic/pronecrotic signals. We identified and characterized (experimentally and by modeling) three Ca(2+) shuttling pathways in primary mesothelial cells during Ca(2+) oscillations: Ca(2+) shuttled between (i) the endoplasmic reticulum (ER) and mitochondria, (ii) the ER and the extracellular space, and (iii) the ER and cytoplasmic Ca(2+) buffers.

  8. Dense core secretory vesicles revealed as a dynamic Ca2+ store in neuroendocrine cells with a vesicle-associated membrane protein aequorin chimaera

    Science.gov (United States)

    Mitchell, Kathryn J.; Pinton, Paolo; Varadi, Aniko; Tacchetti, Carlo; Ainscow, Edward K.; Pozzan, Tullio; Rizzuto, Rosario; Rutter, Guy A.

    2001-01-01

    The role of dense core secretory vesicles in the control of cytosolic-free Ca2+ concentrations ([Ca2+]c) in neuronal and neuroendocrine cells is enigmatic. By constructing a vesicle-associated membrane protein 2–synaptobrevin.aequorin chimera, we show that in clonal pancreatic islet β-cells: (a) increases in [Ca2+]c cause a prompt increase in intravesicular-free Ca2+ concentration ([Ca2+]SV), which is mediated by a P-type Ca2+-ATPase distinct from the sarco(endo) plasmic reticulum Ca2+-ATPase, but which may be related to the PMR1/ATP2C1 family of Ca2+ pumps; (b) steady state Ca2+ concentrations are 3–5-fold lower in secretory vesicles than in the endoplasmic reticulum (ER) or Golgi apparatus, suggesting the existence of tightly bound and more rapidly exchanging pools of Ca2+; (c) inositol (1,4,5) trisphosphate has no impact on [Ca2+]SV in intact or permeabilized cells; and (d) ryanodine receptor (RyR) activation with caffeine or 4-chloro-3-ethylphenol in intact cells, or cyclic ADPribose in permeabilized cells, causes a dramatic fall in [Ca2+]SV. Thus, secretory vesicles represent a dynamic Ca2+ store in neuroendocrine cells, whose characteristics are in part distinct from the ER/Golgi apparatus. The presence of RyRs on secretory vesicles suggests that local Ca2+-induced Ca2+ release from vesicles docked at the plasma membrane could participate in triggering exocytosis. PMID:11571310

  9. Dense core secretory vesicles revealed as a dynamic Ca(2+) store in neuroendocrine cells with a vesicle-associated membrane protein aequorin chimaera.

    Science.gov (United States)

    Mitchell, K J; Pinton, P; Varadi, A; Tacchetti, C; Ainscow, E K; Pozzan, T; Rizzuto, R; Rutter, G A

    2001-10-01

    The role of dense core secretory vesicles in the control of cytosolic-free Ca(2+) concentrations ([Ca(2+)](c)) in neuronal and neuroendocrine cells is enigmatic. By constructing a vesicle-associated membrane protein 2-synaptobrevin.aequorin chimera, we show that in clonal pancreatic islet beta-cells: (a) increases in [Ca(2+)](c) cause a prompt increase in intravesicular-free Ca(2+) concentration ([Ca(2+)]SV), which is mediated by a P-type Ca(2+)-ATPase distinct from the sarco(endo) plasmic reticulum Ca(2+)-ATPase, but which may be related to the PMR1/ATP2C1 family of Ca(2+) pumps; (b) steady state Ca(2+) concentrations are 3-5-fold lower in secretory vesicles than in the endoplasmic reticulum (ER) or Golgi apparatus, suggesting the existence of tightly bound and more rapidly exchanging pools of Ca(2+); (c) inositol (1,4,5) trisphosphate has no impact on [Ca(2+)](SV) in intact or permeabilized cells; and (d) ryanodine receptor (RyR) activation with caffeine or 4-chloro-3-ethylphenol in intact cells, or cyclic ADPribose in permeabilized cells, causes a dramatic fall in [Ca(2+)](SV). Thus, secretory vesicles represent a dynamic Ca(2+) store in neuroendocrine cells, whose characteristics are in part distinct from the ER/Golgi apparatus. The presence of RyRs on secretory vesicles suggests that local Ca(2+)-induced Ca(2+) release from vesicles docked at the plasma membrane could participate in triggering exocytosis.

  10. Ca2+ transport by mitochondria from L1210 mouse ascites tumor cells.

    Science.gov (United States)

    Reynafarje, B; Lehninger, A L

    1973-06-01

    Mitochondria isolated from the ascites form of L1210 mouse leukemia cells readily accumulate Ca(2+) from the suspending medium and eject H(+) during oxidation of succinate in the presence of phosphate and Mg(2+), with normal stoichiometry between Ca(2+) uptake and electron transport. Ca(2+) loads up to 1600 ng-atoms per mg of protein are attained. As is the case in mitochondria from normal tissues, Ca(2+) uptake takes precedence over oxidative phosphorylation. However, Ca(2+) transport by the L-1210 mitochondria is unusual in other respects, which may possibly have general significance in tumor cells. The apparent affinity of the L1210 mitochondria for Ca(2+) in stimulation of oxygen uptake is about 3-fold greater than in normal liver mitochondria; moreover, the maximal rate of Ca(2+) transport is also considerably higher. Furthermore, when Ca(2+) pulses are added to L1210 mitochondria in the absence of phosphate or other permeant anions, much larger amounts of Ca(2+) are bound and H(+) ejected per atom of oxygen consumed than in the presence of phosphate; up to 7 Ca(2+) ions are bound per pair of electrons passing each energy-conserving site of the electron-transport chain. Such "superstoichiometry" of Ca(2+) uptake can be accounted for by two distinct types of respiration-dependent interaction of Ca(2+) with the L1210 mitochondria. One is the stimulation of oxygen consumption, which is achieved by relatively low concentrations of Ca(2+) (K(m) congruent with 8 muM) and is accompanied by binding of Ca(2+) up to 40 ng-atoms per mg of protein. The second process, also dependent on electron transport, is the binding of further Ca(2+) from the medium in exchange with previously stored membrane-bound protons, in which the affinity for Ca(2+) is much lower (K(m) congruent with 120 muM).

  11. Human keratinocyte ATP2C1 localizes to the Golgi and controls Golgi Ca2+ stores

    OpenAIRE

    Behne, M J; Tu, Chia-Ling L; Aronchik, I; Epstein, E; Bench, G.; Bikle, D D; Pozzan, T; Mauro, T.M.

    2003-01-01

    Hailey-Hailey disease (MIM16960) is a blistering skin disease caused by mutations in the Ca2+ ATPase ATP2C1. We found that the abnormal Ca2+ signaling seen in Hailey-Hailey disease keratinocytes correlates with decreased protein levels of ATP2C1. Human ATP2C1 protein approximated 115 kDa in size. The ATP2C1 is localized to the Golgi apparatus in human keratinocytes, similar to its localization in yeast and Caenorhabditis elegans. To test whether the ATP2C1 controls Golgi Ca2+ stores, we measu...

  12. Kinetic and mesoscopic non-equilibrium description of the Ca2+ pump: a comparison

    NARCIS (Netherlands)

    Lervik, A.; Bedeaux, D.; Kjelstrup, S.H.

    2012-01-01

    We analyse the operation of the Ca2?-ATPase ion pump using a kinetic cycle diagram. Using the methodology of Hill, we obtain the cycle fluxes, entropy production and efficiency of the pump. We compare these results with a mesoscopic non-equilibrium description of the pump and show that the kinetic a

  13. Superstoichiometric Ca2+ uptake supported by hydrolysis of endogenous ATP in rat liver mitochondria.

    Science.gov (United States)

    Brand, M D; Lehninger, A L

    1975-10-10

    The nature of the energy store causing rapid superstoichiometric leads to H+/2e minus ejection and leads to Ca2+/2e minus uptake ratios in rat liver mitochondria pulsed with Ca2+ has been investigated. The extent and the rate of the initial fast superstoichiometric phase of H plus ejection were greatly reduced by oligomycin and other ATPase inhibitors; the subsequent shoichiometric phase was unaffected. No such inhibition was seen with atractyloside. Similarly, the initial fast phase of Ca2+ uptake was reduced in extent by oligomycin, whereas the slower stoichiometric phase was unaffected. Moreover, the ATP content of mitochondria previously incubated with succinate decreased by about 80% within 5 s after pulsing with Ca2+. The energy store for superstoichiometric Ca2+ uptake and H plus injection is thus identified as endogenous ATP.

  14. Superstoichiometric Ca2+ uptake supported by hydrolysis of endogenous ATP in rat liver mitochondria.

    Science.gov (United States)

    Brand, M D; Lehninger, A L

    1975-10-10

    The nature of the energy store causing rapid superstoichiometric leads to H+/2e minus ejection and leads to Ca2+/2e minus uptake ratios in rat liver mitochondria pulsed with Ca2+ has been investigated. The extent and the rate of the initial fast superstoichiometric phase of H plus ejection were greatly reduced by oligomycin and other ATPase inhibitors; the subsequent shoichiometric phase was unaffected. No such inhibition was seen with atractyloside. Similarly, the initial fast phase of Ca2+ uptake was reduced in extent by oligomycin, whereas the slower stoichiometric phase was unaffected. Moreover, the ATP content of mitochondria previously incubated with succinate decreased by about 80% within 5 s after pulsing with Ca2+. The energy store for superstoichiometric Ca2+ uptake and H plus injection is thus identified as endogenous ATP. PMID:1176454

  15. EMRE Is a Matrix Ca(2+) Sensor that Governs Gatekeeping of the Mitochondrial Ca(2+) Uniporter.

    Science.gov (United States)

    Vais, Horia; Mallilankaraman, Karthik; Mak, Don-On Daniel; Hoff, Henry; Payne, Riley; Tanis, Jessica E; Foskett, J Kevin

    2016-01-26

    The mitochondrial uniporter (MCU) is an ion channel that mediates Ca(2+) uptake into the matrix to regulate metabolism, cell death, and cytoplasmic Ca(2+) signaling. Matrix Ca(2+) concentration is similar to that in cytoplasm, despite an enormous driving force for entry, but the mechanisms that prevent mitochondrial Ca(2+) overload are unclear. Here, we show that MCU channel activity is governed by matrix Ca(2+) concentration through EMRE. Deletion or charge neutralization of its matrix-localized acidic C terminus abolishes matrix Ca(2+) inhibition of MCU Ca(2+) currents, resulting in MCU channel activation, enhanced mitochondrial Ca(2+) uptake, and constitutively elevated matrix Ca(2+) concentration. EMRE-dependent regulation of MCU channel activity requires intermembrane space-localized MICU1, MICU2, and cytoplasmic Ca(2+). Thus, mitochondria are protected from Ca(2+) depletion and Ca(2+) overload by a unique molecular complex that involves Ca(2+) sensors on both sides of the inner mitochondrial membrane, coupled through EMRE.

  16. Regulation of calpain activity in rat brain with altered Ca2+ homeostasis.

    Science.gov (United States)

    Averna, Monica; Stifanese, Roberto; De Tullio, Roberta; Passalacqua, Mario; Defranchi, Enrico; Salamino, Franca; Melloni, Edon; Pontremoli, Sandro

    2007-01-26

    Activation of calpain occurs as an early event in correlation with an increase in [Ca2+]i induced in rat brain upon treatment with a high salt diet for a prolonged period of time. The resulting sequential events have been monitored in the brain of normal and hypertensive rats of the Milan strain, diverging for a constitutive alteration in the level of [Ca2+]i found to be present in nerve cells of hypertensive animals. After 2 weeks of treatment, the levels of the plasma membrane Ca2+-ATPase and of native calpastatin are profoundly decreased. These degradative processes, more pronounced in the brain of hypertensive rats, are progressively and efficiently compensated in the brain of both rat strains by different incoming mechanisms. Along with calpastatin degradation, 15-kDa still-active inhibitory fragments are accumulated, capable of efficiently replacing the loss of native inhibitor molecules. A partial return to a more efficient control of Ca2+ homeostasis occurs in parallel, assured by an early increase in the expression of Ca2+-ATPase and of calpastatin, both producing, after 12 weeks of a high salt (sodium) diet, the restoration of almost original levels of the Ca2+ pump and of significant amounts of native inhibitor molecules. Thus, conservative calpastatin fragmentation, associated with an increased expression of Ca2+-ATPase and of the calpain natural inhibitor, has been demonstrated to occur in vivo in rat brain. This represents a sequential adaptive response capable of overcoming the effects of calpain activation induced by a moderate long term elevation of [Ca2+]i.

  17. STIM1 enhances SR Ca2+ content through binding phospholamban in rat ventricular myocytes.

    Science.gov (United States)

    Zhao, Guiling; Li, Tianyu; Brochet, Didier X P; Rosenberg, Paul B; Lederer, W J

    2015-08-25

    In ventricular myocytes, the physiological function of stromal interaction molecule 1 (STIM1), an endo/sarcoplasmic reticulum (ER/SR) Ca(2+) sensor, is unclear with respect to its cellular localization, its Ca(2+)-dependent mobilization, and its action on Ca(2+) signaling. Confocal microscopy was used to measure Ca(2+) signaling and to track the cellular movement of STIM1 with mCherry and immunofluorescence in freshly isolated adult rat ventricular myocytes and those in short-term primary culture. We found that endogenous STIM1 was expressed at low but measureable levels along the Z-disk, in a pattern of puncta and linear segments consistent with the STIM1 localizing to the junctional SR (jSR). Depleting SR Ca(2+) using thapsigargin (2-10 µM) changed neither the STIM1 distribution pattern nor its mobilization rate, evaluated by diffusion coefficient measurements using fluorescence recovery after photobleaching. Two-dimensional blue native polyacrylamide gel electrophoresis and coimmunoprecipitation showed that STIM1 in the heart exists mainly as a large protein complex, possibly a multimer, which is not altered by SR Ca(2+) depletion. Additionally, we found no store-operated Ca(2+) entry in control or STIM1 overexpressing ventricular myocytes. Nevertheless, STIM1 overexpressing cells show increased SR Ca(2+) content and increased SR Ca(2+) leak. These changes in Ca(2+) signaling in the SR appear to be due to STIM1 binding to phospholamban and thereby indirectly activating SERCA2a (Sarco/endoplasmic reticulum Ca(2+) ATPase). We conclude that STIM1 binding to phospholamban contributes to the regulation of SERCA2a activity in the steady state and rate of SR Ca(2+) leak and that these actions are independent of store-operated Ca(2+) entry, a process that is absent in normal heart cells. PMID:26261328

  18. Fine tuning of cytosolic Ca 2+ oscillations

    Science.gov (United States)

    Dupont, Geneviève; Combettes, Laurent

    2016-01-01

    Ca 2+ oscillations, a widespread mode of cell signaling, were reported in non-excitable cells for the first time more than 25 years ago. Their fundamental mechanism, based on the periodic Ca 2+ exchange between the endoplasmic reticulum and the cytoplasm, has been well characterized. However, how the kinetics of cytosolic Ca 2+ changes are related to the extent of a physiological response remains poorly understood. Here, we review data suggesting that the downstream targets of Ca 2+ are controlled not only by the frequency of Ca 2+ oscillations but also by the detailed characteristics of the oscillations, such as their duration, shape, or baseline level. Involvement of non-endoplasmic reticulum Ca 2+ stores, mainly mitochondria and the extracellular medium, participates in this fine tuning of Ca 2+ oscillations. The main characteristics of the Ca 2+ exchange fluxes with these compartments are also reviewed. PMID:27630768

  19. 云芝多糖对运动训练大鼠脑组织抗氧化能力和ATPase活性的影响%Effect of Krestin Polysaccharide on Antioxidant Capability and ATPase Activity in Brain Tissues of Rats with High Intensity Exercise

    Institute of Scientific and Technical Information of China (English)

    习雪峰; 王单一; 熊正英; 张林

    2012-01-01

    目的:探讨云芝多糖对力竭运动大鼠脑组织的部分抗氧化酶活性和Na^2+,K^+-ATP酶(Na^+,K^+.ATPasc),Ca^2+,Mg^2+-ATP酶(Ca^2+,Mg^2+.ATPase)活性影响。方法:选取成年雄性SD大鼠24只。将大鼠随机分为3组:安静对照组8只,运动对照组8只,运动加药组8只,进行为期8周的大强度耐力跑台训练。测定大鼠脑组织部分抗氧化酶和Na^+,K^+-ATPase,Ca^2+,Mg^2+.ATPase活性的变化。结果:与安静对照组相比,运动对照组脑组织中超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-Px)、总抗氧化能力(T-AOC)、Na^+,K^+-ATPase、Can,Mg^2+-ATPase活性和血糖含量显著下降(P〈0.01或P〈0.05),MDA生成显著增多(P〈0.05);云芝多糖提高了力竭运动大鼠脑组织SOD、CAT、GSH.Px、T-AOC、Na^+,K^+-ATPase、Ca^2+,Mg^2+.ATPase活性和血糖含量(P〈0.01或P〈0.05),减少MDA生成(P〈0.05)。结论:云芝多糖可以提高力竭运动大鼠脑组织中抗氧化酶活性和Na^+,D.ATPase和Ca^2+,Mg^2+-ATPase的活性,这对于维持运动中能量的供应和抗氧化能力,从而提高大鼠的运动能力具有重要意义。%Objective: To explore the effect of Krestin polysaechatide (PSK) on the activities of antioxidant enzymes and Na^+- K^+-ATPase as well as Ca^2 +-Mg^2+-ATPase in brain tissues of rats after exhaustive exercise. Methods: 24 SD rats were divided into three groups including the control group without exercise, exercise group and exercise plus PSK group. Each group included 8 rats. The rats from the exercise and exercise plus PSK groups were subjected to high-intensity endurance running for 8 consecutive weeks. The changes in the activities of antioxidant enzymes, Na^+-K^+-ATPase and Ca^2+-Mg^2+-ATPase were measured. Results: The activities of SOD

  20. Ca2+ entry in gonadotrophs and alpha T3-1 cells: does store-dependent Ca2+ influx mediate gonadotrophin-releasing hormone action?

    Science.gov (United States)

    McArdle, C A; Forrest-Owen, W; Davidson, J S; Fowkes, R; Bunting, R; Mason, W T; Poch, A; Kratzmeier, M

    1996-04-01

    In pituitary gonadotrophs GnRH causes biphasic (spike and plateau) increases in cytosolic Ca2+ ([Ca2+]i) and gonadotrophin release. The spike phases reflect mobilization of stored Ca2+ and the plateau responses are attributed, in part, to Ca2+ influx via voltage-sensitive Ca2+ channels. In recent years, store-dependent Ca2+ influx (SDCI), in which depletion of the intracellular inositol 1,4,5-trisphosphate-mobilizable pool stimulates Ca2+ influx, has emerged as a major form of Ca2+ entry activated by phosphoinositidase C-coupled receptors in non-excitable cells. More recent evidence also indicates a role for SDCI in excitable cells. We have used dynamic video imaging of [Ca2+]i in alpha T3-1 cells (a gonadotroph-derived cell line) and manipulation of the filling state of the GnRH-mobilizable Ca2+ pool to test the possible role of SDCI in GnRH action. In Ca(2+)-containing medium, GnRH caused a biphasic increase in [Ca2+]i whereas in Ca(2+)-free medium only a transient increase occurred. The response to a second stimulation with GnRH in Ca(2+)-free medium was reduced by > 95% (demonstrating that Ca2+ pool depletion had occurred) and was recovered after brief exposure to Ca(2+)-containing medium (which enables refilling of the pool). Ionomycin (a Ca2+ ionophore) and thapsigargin (which inhibits the Ca(2+)-sequestering ATPase of the endoplasmic reticulum) also transiently increased [Ca2+]i in Ca(2+)-free medium and depleted the GnRH-mobilizable pool as indicated by greatly reduced subsequent responses to GnRH. Pool depletion also occurs on stimulation with GnRH in Ca(2+)-containing medium because addition of ionomycin and Ca(2+)-free medium during the plateau phase of the GnRH response caused only a reduction in [Ca2+]i rather than the transient increase seen without GnRH. To deplete intracellular Ca2+ pools, cells were pretreated in Ca(2+)-free medium with thapsigargin or GnRH and then, after extensive washing, returned to Ca(2+)-containing medium. Pretreatment with

  1. Regulation by L channels of Ca(2+)-evoked secretory responses in ouabain-treated chromaffin cells.

    Science.gov (United States)

    De Pascual, Ricardo; Colmena, Inés; Ruiz-Pascual, Lucía; Baraibar, Andrés Mateo; Egea, Javier; Gandía, Luis; García, Antonio G

    2016-10-01

    It is known that the sustained depolarisation of adrenal medullary bovine chromaffin cells (BCCs) with high K(+) concentrations produces an initial sharp catecholamine release that subsequently fades off in spite depolarisation persists. Here, we have recreated a sustained depolarisation condition of BCCs by treating them with the Na(+)/K(+) ATPase blocker ouabain; in doing so, we searched experimental conditions that permitted the development of a sustained long-term catecholamine release response that could be relevant during prolonged stress. BCCs were perifused with nominal 0Ca(2+) solution, and secretion responses were elicited by intermittent application of short 2Ca(2+) pulses (Krebs-HEPES containing 2 mM Ca(2+)). These pulses elicited a biphasic secretory pattern with an initial 30-min period with secretory responses of increasing amplitude and a second 30-min period with steady-state, non-inactivating responses. The initial phase was not due to gradual depolarisation neither to gradual increases of the cytosolic calcium transients ([Ca(2+)]c) elicited by 2Ca(2+) pulses in BBCs exposed to ouabain; both parameters increased soon after ouabain addition. Νifedipine blocked these responses, and FPL64176 potentiated them, suggesting that they were triggered by Ca(2+) entry through non-inactivating L-type calcium channels. This was corroborated by nifedipine-evoked blockade of the L-type Ca(2+) channel current and the [Ca(2+)]c transients elicited by 2Ca(2+) pulses. Furthermore, the plasmalemmal Na(+)/Ca(2+) exchanger (NCX) blocker SEA0400 caused a mild inhibition followed by a large rebound increase of the steady-state secretory responses. We conclude that these two phases of secretion are mostly contributed by Ca(2+) entry through L calcium channels, with a minor contribution of Ca(2+) entry through the reverse mode of the NCX.

  2. Rat vas deferens SERCA2 is modulated by Ca2+/calmodulin protein kinase II-mediated phosphorylation

    Directory of Open Access Journals (Sweden)

    J.B.R. Rodriguez

    2013-03-01

    Full Text Available Ca2+ pumps are important players in smooth muscle contraction. Nevertheless, little information is available about these pumps in the vas deferens. We have determined which subtype of sarco(endoplasmic reticulum Ca2+-ATPase isoform (SERCA is expressed in rat vas deferens (RVD and its modulation by calmodulin (CaM-dependent mechanisms. The thapsigargin-sensitive Ca2+-ATPase from a membrane fraction containing the highest SERCA levels in the RVD homogenate has the same molecular mass (∼115 kDa as that of SERCA2 from the rat cerebellum. It has a very high affinity for Ca2+ (Ca0.5 = 780 nM and a low sensitivity to vanadate (IC50 = 41 µM. These facts indicate that SERCA2 is present in the RVD. Immunoblotting for CaM and Ca2+/calmodulin-dependent protein kinase II (CaMKII showed the expression of these two regulatory proteins. Ca2+ and CaM increased serine-phosphorylated residues of the 115-kDa protein, indicating the involvement of CaMKII in the regulatory phosphorylation of SERCA2. Phosphorylation is accompanied by an 8-fold increase of thapsigargin-sensitive Ca2+ accumulation in the lumen of vesicles derived from these membranes. These data establish that SERCA2 in the RVD is modulated by Ca2+ and CaM, possibly via CaMKII, in a process that results in stimulation of Ca2+ pumping activity.

  3. Multiple Ca2+ sensors in secretion

    DEFF Research Database (Denmark)

    Walter, Alexander M; Groffen, Alexander J; Sørensen, Jakob Balslev;

    2011-01-01

    Regulated neurotransmitter secretion depends on Ca(2+) sensors, C2 domain proteins that associate with phospholipids and soluble N-ethylmaleimide-sensitive fusion attachment protein receptor (SNARE) complexes to trigger release upon Ca(2+) binding. Ca(2+) sensors are thought to prevent spontaneous...... fusion at rest (clamping) and to promote fusion upon Ca(2+) activation. At least eight, often coexpressed, Ca(2+) sensors have been identified in mammals. Accumulating evidence suggests that multiple Ca(2+) sensors interact, rather than work autonomously, to produce the complex secretory response...... observed in neurons and secretory cells. In this review, we present several working models to describe how different sensors might be arranged to mediate synchronous, asynchronous and spontaneous neurotransmitter release. We discuss the scenario that different Ca(2+) sensors typically act on one shared...

  4. Features of Mg2Si Layer Growth in Si/Mg2Si Multilayers

    Directory of Open Access Journals (Sweden)

    L.E. Konotopskyi

    2016-06-01

    Full Text Available Features of magnesium siliced layer growth in Si/Mg2Si multilayers in initial state and after thermal annealing were studied by methods of transmission electron microscopy and X-Ray scattering. As-deposited magnesium silicide layers are amorphous with nanocrystal inclusions of metastable h-Mg2Si. Formation of Mg2Si in hexagonal modification occurs under the influence of stress produced by silicon layers. At T = 723 К Mg2Si layers finished crystallizes in hexagonal modification, with some coarsening of grains. That is accompanied with 7.3 % reduction in period of the Si/Mg2Si multilayer.

  5. Rotary ATPases

    Science.gov (United States)

    Stewart, Alastair G.; Sobti, Meghna; Harvey, Richard P.; Stock, Daniela

    2013-01-01

    Rotary ATPases are molecular rotary motors involved in biological energy conversion. They either synthesize or hydrolyze the universal biological energy carrier adenosine triphosphate. Recent work has elucidated the general architecture and subunit compositions of all three sub-types of rotary ATPases. Composite models of the intact F-, V- and A-type ATPases have been constructed by fitting high-resolution X-ray structures of individual subunits or sub-complexes into low-resolution electron densities of the intact enzymes derived from electron cryo-microscopy. Electron cryo-tomography has provided new insights into the supra-molecular arrangement of eukaryotic ATP synthases within mitochondria and mass-spectrometry has started to identify specifically bound lipids presumed to be essential for function. Taken together these molecular snapshots show that nano-scale rotary engines have much in common with basic design principles of man made machines from the function of individual “machine elements” to the requirement of the right “fuel” and “oil” for different types of motors. PMID:23369889

  6. Retinoic acid inhibits calmodulin binding to human erythrocyte membranes and reduces membrane Ca2(+)-adenosine triphosphatase activity.

    OpenAIRE

    Davis, F B; Smith, T. J.; Deziel, M R; Davis, P J; Blas, S D

    1990-01-01

    Ca2(+)-ATPase activity in human red cell membranes is dependent on the presence of calmodulin. All trans-retinoic acid inhibited human red cell membrane Ca2(+)-ATPase activity in vitro in a concentration-dependent manner (10(-8) to 10(-4) M). In contrast, retinol, retinal, 13-cis-retinoic acid and the benzene ring analogue of retinoic acid did not alter enzyme activity. Purified calmodulin (up to 500 ng/ml, 3 X 10(-8) M) added to red cell membranes, in the presence of inhibitory concentration...

  7. When Isolated at Full Receptivity, in Vitro Fertilized Wheat (Triticum aestivum, L. Egg Cells Reveal [Ca2+]cyt Oscillation of Intracellular Origin

    Directory of Open Access Journals (Sweden)

    Zsolt Pónya

    2014-12-01

    Full Text Available During in vitro fertilization of wheat (Triticum aestivum, L. in egg cells isolated at various developmental stages, changes in cytosolic free calcium ([Ca2+]cyt were observed. The dynamics of [Ca2+]cyt elevation varied, reflecting the difference in the developmental stage of the eggs used. [Ca2+]cyt oscillation was exclusively observed in fertile, mature egg cells fused with the sperm cell. To determine how [Ca2+]cyt oscillation in mature egg cells is generated, egg cells were incubated in thapsigargin, which proved to be a specific inhibitor of the endoplasmic reticulum (ER Ca2+-ATPase in wheat egg cells. In unfertilized egg cells, the addition of thapsigargin caused an abrupt transient increase in [Ca2+]cyt in the absence of extracellular Ca2+, suggesting that an influx pathway for Ca2+ is activated by thapsigargin. The [Ca2+]cyt oscillation seemed to require the filling of an intracellular calcium store for the onset of which, calcium influx through the plasma membrane appeared essential. This was demonstrated by omitting extracellular calcium from (or adding GdCl3 to the fusion medium, which prevented [Ca2+]cyt oscillation in mature egg cells fused with the sperm. Combined, these data permit the hypothesis that the first sperm-induced transient increase in [Ca2+]cyt depletes an intracellular Ca2+ store, triggering an increase in plasma membrane Ca2+ permeability, and this enhanced Ca2+ influx results in [Ca2+]cyt oscillation.

  8. Cytoplasmic free Ca2+ is essential for multiple steps in malaria parasite egress from infected erythrocytes

    Directory of Open Access Journals (Sweden)

    Glushakova Svetlana

    2013-01-01

    Full Text Available Abstract Background Egress of Plasmodium falciparum, from erythrocytes at the end of its asexual cycle and subsequent parasite invasion into new host cells, is responsible for parasite dissemination in the human body. The egress pathway is emerging as a coordinated multistep programme that extends in time for tens of minutes, ending with rapid parasite extrusion from erythrocytes. While the Ca2+ regulation of the invasion of P. falciparum in erythrocytes is well established, the role of Ca2+ in parasite egress is poorly understood. This study analysed the involvement of cytoplasmic free Ca2+ in infected erythrocytes during the multistep egress programme of malaria parasites. Methods Live-cell fluorescence microscopy was used to image parasite egress from infected erythrocytes, assessing the effect of drugs modulating Ca2+ homeostasis on the egress programme. Results A steady increase in cytoplasmic free Ca2+ is found to precede parasite egress. This increase is independent of extracellular Ca2+ for at least the last two hours of the cycle, but is dependent upon Ca2+ release from internal stores. Intracellular BAPTA chelation of Ca2+ within the last 45 minutes of the cycle inhibits egress prior to parasitophorous vacuole swelling and erythrocyte membrane poration, two characteristic morphological transformations preceding parasite egress. Inhibitors of the parasite endoplasmic reticulum (ER Ca2+-ATPase accelerate parasite egress, indicating that Ca2+ stores within the ER are sufficient in supporting egress. Markedly accelerated egress of apparently viable parasites was achieved in mature schizonts using Ca2+ ionophore A23187. Ionophore treatment overcomes the BAPTA-induced block of parasite egress, confirming that free Ca2+ is essential in egress initiation. Ionophore treatment of immature schizonts had an adverse effect inducing parasitophorous vacuole swelling and killing the parasites within the host cell. Conclusions The parasite egress

  9. Preparation of pure and intact Plasmodium falciparum plasma membrane vesicles and partial characterisation of the plasma membrane ATPase

    Directory of Open Access Journals (Sweden)

    Smith Pete J

    2002-04-01

    Full Text Available Abstract Background In host erythrocytes, the malaria parasite must contend with ion and drug transport across three membranes; its own plasma membrane, the parasitophorous membrane and the host plasma membrane. Isolation of pure and intact Plasmodium falciparum plasma membrane would provide a suitable model to elucidate the possible role played by the parasite plasma membrane in ion balance and drug transport. Results This study describes a procedure for isolating parasite plasma membrane from P. falciparum-infected erythrocytes. With this method, the trophozoites released by saponin treatment were cleansed of erythrocyte membranes using anti-erythrocyte antibodies fixed to polystyrene beads. These trophozoites were then biotinylated and the parasite plasma membrane was disrupted by nitrogen cavitation. This process allows the membranes to reform into vesicles. The magnetic streptavidin beads bind specifically to the biotinylated parasite plasma membrane vesicles facilitating their recovery with a magnet. These vesicles can then be easily released from the magnetic beads by treatment with dithiotreithol. The parasite plasma membrane showed optimal ATPase activity at 2 mM ATP and 2 mM Mg2+. It was also found that Ca2+ could not substitute for Mg2+ ATPase activity in parasite plasma membranes whereas activity was completely preserved when Mn2+ was used instead of Mg2+. Other nucleoside triphosphates tested were hydrolysed as efficiently as ATP, while the nucleoside monophosphate AMP was not. Conclusions We have described the successful isolation of intact P. falciparum plasma membrane vesicles free of contaminating organelles and determined the experimental conditions for optimum ATPase activity.

  10. Calcium-ATPases: Gene disorders and dysregulation in cancer.

    Science.gov (United States)

    Dang, Donna; Rao, Rajini

    2016-06-01

    Ca(2+)-ATPases belonging to the superfamily of P-type pumps play an important role in maintaining low, nanomolar cytoplasmic Ca(2+) levels at rest and priming organellar stores, including the endoplasmic reticulum, Golgi, and secretory vesicles with high levels of Ca(2+) for a wide range of signaling functions. In this review, we introduce the distinct subtypes of Ca(2+)-ATPases and their isoforms and splice variants and provide an overview of their specific cellular roles as they relate to genetic disorders and cancer, with a particular emphasis on recent findings on the secretory pathway Ca(2+)-ATPases (SPCA). Mutations in human ATP2A2, ATP2C1 genes, encoding housekeeping isoforms of the endoplasmic reticulum (SERCA2) and secretory pathway (SPCA1) pumps, respectively, confer autosomal dominant disorders of the skin, whereas mutations in other isoforms underlie various muscular, neurological, or developmental disorders. Emerging evidence points to an important function of dysregulated Ca(2+)-ATPase expression in cancers of the colon, lung, and breast where they may serve as markers of differentiation or novel targets for therapeutic intervention. We review the mechanisms underlying the link between calcium homeostasis and cancer and discuss the potential clinical relevance of these observations. This article is part of a Special Issue entitled: Calcium and Cell Fate. Guest Editors: Jacques Haiech, Claus Heizmann, Joachim Krebs, Thierry Capiod and Olivier Mignen. PMID:26608610

  11. SERCA and PMCA pumps contribute to the deregulation of Ca2+ homeostasis in human CF epithelial cells.

    Science.gov (United States)

    Philippe, Réginald; Antigny, Fabrice; Buscaglia, Paul; Norez, Caroline; Becq, Frédéric; Frieden, Maud; Mignen, Olivier

    2015-05-01

    Cystic Fibrosis (CF) disease is caused by mutations in the CFTR gene (CF transmembrane conductance regulator). F508 deletion is the most represented mutation, and F508del-CFTR is absent of plasma membrane and accumulates into the endoplasmic reticulum (ER) compartment. Using specific Ca2+ genetics cameleon probes, we showed in the human bronchial CF epithelial cell line CFBE that ER Ca2+ concentration was strongly increased compared to non-CF (16HBE) cells, and normalized by the F508del-CFTR corrector agent, VX-809. We also showed that ER F508del-CFTR retention increases SERCA (Sarcoplasmic/Reticulum Ca2+ ATPase) pump activity whereas PMCA (Plasma Membrane Ca2+ ATPase) activities were reduced in these CF cells compared to corrected CF cells (VX-809) and non-CF cells. We are showing for the first time CFTR/SERCA and CFTR/PMCA interactions that are modulated in CF cells and could explain part of Ca2+ homeostasis deregulation due to mislocalization of F508del-CFTR. Using ER or mitochondria genetics Ca2+ probes, we are showing that ER Ca2+ content, mitochondrial Ca2+ uptake, SERCA and PMCA pump, activities are strongly affected by the localization of F508del-CFTR protein. PMID:25661196

  12. Novel nuclear targeting coiled-coil protein of Helicobacter pylori showing Ca(2+)-independent, Mg(2+)-dependent DNase I activity.

    Science.gov (United States)

    Kwon, Young Chul; Kim, Sinil; Lee, Yong Seok; Lee, Je Chul; Cho, Myung-Je; Lee, Woo-Kon; Kang, Hyung-Lyun; Song, Jae-Young; Baik, Seung Chul; Ro, Hyeon Su

    2016-05-01

    HP0059, an uncharacterized gene of Helicobacter pylori, encodes a 284-aa-long protein containing a nuclear localization sequence (NLS) and multiple leucine-rich heptad repeats. Effects of HP0059 proteins in human stomach cells were assessed by incubation of recombinant HP0059 proteins with the AGS human gastric carcinoma cell line. Wild-type HP0059 proteins showed cytotoxicity in AGS cells in a concentration-dependent manner, whereas NLS mutant protein showed no effect, suggesting that the cytotoxicity is attributed to host nuclear localization. AGS cells transfected with pEGFP-HP0059 plasmid showed strong GFP signal merged to the chromosomal DNA region. The chromosome was fragmented into multiple distinct dots merged with the GFP signal after 12 h of incubation. The chromosome fragmentation was further explored by incubation of AGS chromosomal DNA with recombinant HP0059 proteins, which leaded to complete degradation of the chromosomal DNA. HP0059 protein also degraded circular plasmid DNA without consensus, being an indication of DNase I activity. The DNase was activated by MgCl2, but not by CaCl2. The activity was completely blocked by EDTA. The optimal pH and temperature for DNase activity were 7.0-8.0 and 55°C, respectively. These results indicate that HP0059 possesses a novel DNase I activity along with a role in the genomic instability of human gastric cells, which may result in the transformation of gastric cells. PMID:27095458

  13. Downregulation of Ca(2+) and Mg(2+) transport proteins in the kidney explains tacrolimus (FK506)-induced hypercalciuria and hypomagnesemia.

    NARCIS (Netherlands)

    Nijenhuis, T.; Hoenderop, J.G.J.; Bindels, R.J.M.

    2004-01-01

    FK506 (tacrolimus) and dexamethasone are potent immunosuppressants known to induce significant side effects on mineral homeostasis, including hypercalciuria and hypomagnesemia. However, the underlying molecular mechanisms remain unknown. The present study investigated the effects of FK506 and dexame

  14. Dissection of local Ca(2+) signals inside cytosol by ER-targeted Ca(2+) indicator.

    Science.gov (United States)

    Niwa, Fumihiro; Sakuragi, Shigeo; Kobayashi, Ayana; Takagi, Shin; Oda, Yoichi; Bannai, Hiroko; Mikoshiba, Katsuhiko

    2016-10-01

    Calcium (Ca(2+)) is a versatile intracellular second messenger that operates in various signaling pathways leading to multiple biological outputs. The diversity of spatiotemporal patterns of Ca(2+) signals, generated by the coordination of Ca(2+) influx from the extracellular space and Ca(2+) release from the intracellular Ca(2+) store the endoplasmic reticulum (ER), is considered to underlie the diversity of biological outputs caused by a single signaling molecule. However, such Ca(2+) signaling diversity has not been well described because of technical limitations. Here, we describe a new method to report Ca(2+) signals at subcellular resolution. We report that OER-GCaMP6f, a genetically encoded Ca(2+) indicator (GECI) targeted to the outer ER membrane, can monitor Ca(2+) release from the ER at higher spatiotemporal resolution than conventional GCaMP6f. OER-GCaMP6f was used for in vivo Ca(2+) imaging of C. elegans. We also found that the spontaneous Ca(2+) elevation in cultured astrocytes reported by OER-GCaMP6f showed a distinct spatiotemporal pattern from that monitored by plasma membrane-targeted GCaMP6f (Lck-GCaMP6f); less frequent Ca(2+) signal was detected by OER-GCaMP6f, in spite of the fact that Ca(2+) release from the ER plays important roles in astrocytes. These findings suggest that targeting of GECIs to the ER outer membrane enables sensitive detection of Ca(2+) release from the ER at subcellular resolution, avoiding the diffusion of GECI and Ca(2+). Our results indicate that Ca(2+) imaging with OER-GCaMP6f in combination with Lck-GCaMP6f can contribute to describing the diversity of Ca(2+) signals, by enabling dissection of Ca(2+) signals at subcellular resolution.

  15. A kinetic study of the gill (Na+, K+)-ATPase, and its role in ammonia excretion in the intertidal hermit crab, Clibanarius vittatus.

    Science.gov (United States)

    Gonçalves, Rúbia R; Masui, Douglas C; McNamara, John C; Mantelatto, Fernando L M; Garçon, Daniela P; Furriel, Rosa P M; Leone, Francisco A

    2006-11-01

    To better comprehend the role of gill ion regulatory mechanisms, the modulation by Na(+), K(+), NH(4)(+) and ATP of (Na(+), K(+))-ATPase activity was examined in a posterior gill microsomal fraction from the hermit crab, Clibanarius vittatus. Under saturating Mg(2+), Na(+) and K(+) concentrations, two well-defined ATP hydrolyzing sites were revealed. ATP was hydrolyzed at the high-affinity sites at a maximum rate of V=19.1+/-0.8 U mg(-1) and K(0.5)=63.8+/-2.9 nmol L(-1), obeying cooperative kinetics (n(H)=1.9); at the low-affinity sites, hydrolysis obeyed Michaelis-Menten kinetics with K(M)=44.1+/-2.6 mumol L(-1) and V=123.5+/-6.1 U mg(-1). Stimulation by Na(+) (V=149.0+/-7.4 U mg(-1); K(M)=7.4+/-0.4 mmol L(-1)), Mg(2+) (V=132.0+/-5.3 U mg(-1); K(0.5)=0.36+/-0.02 mmol L(-1)), NH(4)(+) (V=245.6+/-9.8 U mg(-1); K(M)=4.5+/-0.2 mmol L(-1)) and K(+) (V=140.0+/-4.9 U mg(-1); K(M)=1.5+/-0.1 mmol L(-1)) followed a single saturation curve and, except for Mg(2+), obeyed Michaelis-Menten kinetics. Under optimal ionic conditions, but in the absence of NH(4)(+), ouabain (K(I)=117.3+/-3.5 mumol L(-1)) and orthovanadate inhibited up to 67% of the ATPase activity. The inhibition studies performed suggest the presence of F(0)F(1), V- and P-ATPases, but not Na(+)-, K(+)- or Ca(2+)-ATPases as contaminants in the gill microsomal preparation. (Na(+), K(+))-ATPase activity was synergistically modulated by NH(4)(+) and K(+). At 20 mmol L(-1) K(+), a maximum rate of V=290.8+/-14.5 U mg(-1) was seen as NH(4)(+) concentration was increased up to 50 mmol L(-1). However, at fixed NH(4)(+) concentrations, no additional stimulation was found for increasing K(+) concentrations (V=135.2+/-4.1 U mg(-1) and V=236.6+/-9.5 U mg(-1) and for 10 and 30 mmol L(-1) NH(4)(+), respectively). This is the first report to detail ionic modulation of gill (Na(+), K(+))-ATPase in C. vittatus, revealing an asymmetrical, synergistic stimulation of the enzyme by K(+) and NH(4)(+), as yet undescribed for other (Na

  16. EMRE Is a Matrix Ca2+ Sensor that Governs Gatekeeping of the Mitochondrial Ca2+ Uniporter

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    Horia Vais

    2016-01-01

    Full Text Available The mitochondrial uniporter (MCU is an ion channel that mediates Ca2+ uptake into the matrix to regulate metabolism, cell death, and cytoplasmic Ca2+ signaling. Matrix Ca2+ concentration is similar to that in cytoplasm, despite an enormous driving force for entry, but the mechanisms that prevent mitochondrial Ca2+ overload are unclear. Here, we show that MCU channel activity is governed by matrix Ca2+ concentration through EMRE. Deletion or charge neutralization of its matrix-localized acidic C terminus abolishes matrix Ca2+ inhibition of MCU Ca2+ currents, resulting in MCU channel activation, enhanced mitochondrial Ca2+ uptake, and constitutively elevated matrix Ca2+ concentration. EMRE-dependent regulation of MCU channel activity requires intermembrane space-localized MICU1, MICU2, and cytoplasmic Ca2+. Thus, mitochondria are protected from Ca2+ depletion and Ca2+ overload by a unique molecular complex that involves Ca2+ sensors on both sides of the inner mitochondrial membrane, coupled through EMRE.

  17. Regulation of free Ca2+ by liver mitochondria and endoplasmic reticulum.

    Science.gov (United States)

    Becker, G L; Fiskum, G; Lehninger, A L

    1980-10-10

    Electrode measurements were made of the free Ca2+ concentration maintained by suspensions of isolated rat liver mitochondria and microsomes, as well as by hepatocytes whose plasma membranes had been made permeable by treatment with digitonin. When the KCl, ATP, Mg2+, and phosphate concentrations were made similar to that of cytosol, the steady state free Ca2+ concentration in the presence of respiring mitochondria alone was about 0.5 microM. The additional presence of rat liver microsomes resulted in a steady state level of close to 0.2 microM, which was maintaied for greater than 1 h at 25 degrees C. This concentration of Ca2+ was also maintained by suspensions of hepatocytes permeabilized by digitonin and thus may approximate the actual cytosolic free Ca2+ concentration in vivo. The "set point" for free Ca2+ homeostasis in these systems is determined by mitochondrial Ca2+ influx-efflux cycling, which is dependent on the level of intramitochondrial Ca2+ and can be adjusted by sequestration of Ca2+ in microsomes. PMID:7410406

  18. Na(+) and Ca(2+) pumps in the gills, epipodites and branchiostegites of the european lobster Homarus gammarus: effects of dilute sea water.

    Science.gov (United States)

    Flik, G; Haond, C

    2000-01-01

    Crude homogenates and plasma-membrane-enriched fractions were prepared from the epithelium of the gills, epipodites and branchiostegites of intermoult European lobsters Homarus gammarus, and Na(+)/K(+)-ATPase, Ca(2+)-ATPase and Na(+)/Ca(2+) exchange activities were quantified in these tissues. Lobsters were kept in sea water (salinity 35 ) or were adapted to dilute sea water (22.1 ). The lobster hyperregulates haemolymph osmolarity and Ca(2+) levels in both media. Homogenates of the podobranchs, arthrobranchs and pleurobranchs had comparable Na(+)/K(+)-ATPase specific activities, and mean activities increased significantly for all three types of gills when the animals were kept in dilute sea water. In the epipodites and branchiostegites, Na(+)/K(+)-ATPase specific activities exceeded those in the gills, and exposure to dilute sea water greatly enhanced these activities. In sea water, 80 % of the total Na(+)/K(+)-ATPase activity is associated with the gills and epipodites (each tissue containing 40 %) and 20 % with the branchiostegites; in dilute sea water, the gills contained approximately 25 %, the epipodites 40 % and the branchiostegites approximately 35 % of the total activity, indicating the relative importance of the epipodites and branchiostegites for ionic hyperregulation in dilute media. In plasma membrane vesicles isolated from the gills, epipodites and branchiostegites, Ca(2+) transport driven by ATP and by a Na(+ )gradient was demonstrated. Exposure to dilute sea water enhanced Na(+)/Ca(2+ )exchange and Ca(2+)-ATPase activities in the epipodites and branchiostegites; in the gills, however, Ca(2+) transport activities decreased. The role of these tissues and enzymes in Na(+) and Ca(2+) handling by the lobster is discussed. PMID:10607531

  19. Hemolymph ion regulation and kinetic characteristics of the gill (Na⁺, K⁺)-ATPase in the hermit crab Clibanarius vittatus (Decapoda, Anomura) acclimated to high salinity.

    Science.gov (United States)

    Lucena, Malson N; Garçon, Daniela P; Mantelatto, Fernando L M; Pinto, Marcelo R; McNamara, John C; Leone, Francisco A

    2012-04-01

    We examine hemolymph ion regulation and the kinetic properties of a gill microsomal (Na(+), K(+))-ATPase from the intertidal hermit crab, Clibanarius vittatus, acclimated to 45‰ salinity for 10 days. Hemolymph osmolality is hypo-regulated (1102.5 ± 22.1 mOsm kg(-1) H(2)O) at 45‰ but elevated compared to fresh-caught crabs (801.0 ± 40.1 mOsm kg(-1) H(2)O). Hemolymph [Na(+)] (323.0 ± 2.5 mmol L(-1)) and [Mg(2+)] (34.6 ± 1.0 mmol L(-1)) are hypo-regulated while [Ca(2+)] (22.5 ± 0.7 mmol L(-1)) is hyper-regulated; [K(+)] is hyper-regulated in fresh-caught crabs (17.4 ± 0.5 mmol L(-1)) but hypo-regulated (6.2 ± 0.7 mmol L(-1)) at 45‰. Protein expression patterns are altered in the 45‰-acclimated crabs, although Western blot analyses reveal just a single immunoreactive band, suggesting a single (Na(+), K(+))-ATPase α-subunit isoform, distributed in different density membrane fractions. A high-affinity (Vm=46.5 ± 3.5 Umg(-1); K(0.5)=7.07 ± 0.01 μmol L(-1)) and a low-affinity ATP binding site (Vm=108.1 ± 2.5 U mg(-1); K(0.5)=0.11 ± 0.3 mmol L(-1)), both obeying cooperative kinetics, were disclosed. Modulation of (Na(+), K(+))-ATPase activity by Mg(2+), K(+) and NH(4)(+) also exhibits site-site interactions, but modulation by Na(+) shows Michaelis-Menten kinetics. (Na(+), K(+))-ATPase activity is synergistically stimulated up to 45% by NH(4)(+) plus K(+). Enzyme catalytic efficiency for variable [K(+)] and fixed [NH(4)(+)] is 10-fold greater than for variable [NH(4)(+)] and fixed [K(+)]. Ouabain inhibited ≈80% of total ATPase activity (K(I)=464.7 ± 23.2 μmol L(-1)), suggesting that ATPases other than (Na(+), K(+))-ATPase are present. While (Na(+), K(+))-ATPase activities are similar in fresh-caught (around 142 nmol Pi min(-1)mg(-1)) and 45‰-acclimated crabs (around 154 nmol Pi min(-1)mg(-1)), ATP affinity decreases 110-fold and Na(+) and K(+) affinities increase 2-3-fold in 45‰-acclimated crabs. PMID:22260788

  20. Identification of calcium-transporting ATPases of Entamoeba histolytica and cellular localization of the putative SERCA.

    Science.gov (United States)

    Martinez-Higuera, Aarón; Salas-Casas, Andrés; Calixto-Gálvez, Mercedes; Chávez-Munguía, Bibiana; Pérez-Ishiwara, D Guillermo; Ximénez, Cecilia; Rodríguez, Mario A

    2013-09-01

    Calcium has an important role on signaling of different cellular processes in the protozoa parasite Entamoeba histolytica, including development and pathogenesis. However, the systems that control calcium responses in this parasite are incompletely understood. Calcium-ATPases (Ca(2+)-ATPases) are proteins that play an important role in calcium homeostasis by catalyzing the active efflux of this ion from cytoplasm and are essential to the correct functioning of the cell machinery. Here, we reported the identification of five E. histolytica genes encoding putative Ca(2+)-ATPases, three related to PMCA, and two related to organellar ATPases. RT-PCR assays showed that all those genes are expressed in trophozoites and specific antibodies against the SERCA-like member located this protein in a continuous cytoplasmic network, supporting the hypothesis that it corresponds to the Ca(2+)-ATPase responsible to sequester calcium in the endoplasmic reticulum of this parasite.

  1. By Regulating Mitochondrial Ca2+-Uptake UCP2 Modulates Intracellular Ca2+.

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    Lukas Jaroslaw Motloch

    Full Text Available The possible role of UCP2 in modulating mitochondrial Ca2+-uptake (mCa2+-uptake via the mitochondrial calcium uniporter (MCU is highly controversial.Thus, we analyzed mCa2+-uptake in isolated cardiac mitochondria, MCU single-channel activity in cardiac mitoplasts, dual Ca2+-transients from mitochondrial ((Ca2+m and intracellular compartment ((Ca2+c in the whole-cell configuration in cardiomyocytes of wild-type (WT and UCP2-/- mice.Isolated mitochondria showed a Ru360 sensitive mCa2+-uptake, which was significantly decreased in UCP2-/- (229.4±30.8 FU vs. 146.3±23.4 FU, P0.05 and transsarcolemmal Ca2+-influx was inhibited suggesting a possible compensatory mechanism. Additionally, we observed an inhibitory effect of ATP on mCa2+-uptake in WT mitoplasts and (Ca2+m of cardiomyocytes leading to an increase of (Ca2+c while no ATP dependent effect was observed in UCP2-/-.Our results indicate regulatory effects of UCP2 on mCa2+-uptake. Furthermore, we propose, that previously described inhibitory effects on MCU by ATP may be mediated via UCP2 resulting in changes of excitation contraction coupling.

  2. 亚低温治疗对脑创伤后三磷酸腺苷酶的影响%Effects of hypothermia on ATPase following brain trauma in rats

    Institute of Scientific and Technical Information of China (English)

    黄慧玲; 只达石; 张琳瑛; 王颖; 张玲

    2001-01-01

    目的观察亚低温治疗对大鼠脑外伤后脑组织Na+-K+-ATP酶、Mg2+-ATP酶以及Ca2+-ATP酶的影响。方法 75只大鼠随机分为常温对照组(33只)、常温受伤组(22只)和亚低温治疗组(20只),后两组用自由落体方法致大鼠左侧脑外伤,亚低温治疗组受伤后用冰袋全身降温至脑温30 ℃后维持1 h,然后加热复温至37℃。每组大鼠在伤后3 h、1,3, 5 和7 d取大脑组织,测定组织匀浆液中ATP酶的活性。结果 (1)Na+-K+-ATP酶:常温受伤组和亚低温治疗组在3 h明显高于对照组,而后明显下降。亚低温治疗组在第3天明显高于常温受伤组;(2)Mg2+-ATP酶:常温受伤组和亚低温治疗组在1 d后才开始明显下降,但亚低温治疗组在1 d和3 d中较常温组下降速度明显变慢;(3)Ca2+-ATP酶:常温受伤组第1天就较常温对照组明显下降,而亚低温治疗组3 h和第1天保持正常,第3天才明显下降,但仍然显著高于常温受伤组。结论 (1)脑外伤大鼠脑细胞Na+-K+-ATP酶早期对脑外伤有应激反应,亚低温治疗对细胞钠通道的作用不明显;(2)亚低温对钙泵有明显的调节作用,较常温受伤组显著提高脑细胞Ca2+-Mg2+-ATP酶的活性;(3)亚低温治疗能延缓脑细胞钙通道的损伤时间,且在7 d内脑细胞的钠通道和钙通道在低水平保持相对稳定,从而减少Ca2+的内流,减轻脑水肿。%Objective To investigate the effects of hypothermia on the contents of Na+-K+-ATPase, Mg2+-ATPase and Ca2+-ATPase in traumatic rats.  Methods  Seventy five Wistar rats were divided randomly into three groups : the non-traumatic control group (n=33), the normothermia traumatic group (n=22) and the hypothermia-treated group (n=20). Brain trauma was induced on the left cerebrum by free falling objects. The whole body of the animals in the hypothermia-treated group was cooled to 30℃ for 1 hour, then heated to 37℃. The cerebrum

  3. Myofibril ATPase activity of cardiac and skeletal muscle of exhaustively exercised rats.

    Science.gov (United States)

    Belcastro, A N; Turcotte, R; Rossiter, M; Secord, D; Maybank, P E

    1984-01-01

    The activation characteristics of Mg-ATP and Ca2+ on cardiac and skeletal muscle myofibril ATPase activity were studied in rats following a run to exhaustion. In addition, the effect of varying ionic strength was determined on skeletal muscle from exhausted animals. The exhausted group (E) ran at a speed of 25 m min-1 with an 8% incline. Myofibril ATPase activities for control (C) and E were determined with 1, 3 and 5 mM Mg-ATP and 1 and 10 microM Ca2+ at pH 7.0 and 30 degrees C. For control skeletal muscle, at 1 and 10 microM Ca2+, there was an increase in ATPase activity from 1 to 5 mM Mg-ATP (P less than 0.05). For E animals the myofibril ATPase activities at 10 microM Ca2+ and all Mg-ATP concentrations were similar to C (P greater than 0.05). At 1.0 microM Ca2+ and all Mg-ATP concentrations were similar to C (P greater than 0.05). At 1.0 microM Ca2+ the activities at 3 and 5 mM Mg-ATP were greater for the E animals (P less than 0.05). Increasing KCl concentrations resulted in greater inhibition for E animals. With cardiac muscle, the myofibril ATPase activities at 1.0 microM free Ca2+ were lower for E at all Mg-ATP levels (P less than 0.05). In contrast, at 10 microM Ca2+, the E group exhibited an elevated myofibril ATPase activity. The results indicate that Mg-ATP and Ca2+ activation of cardiac and skeletal muscle myofibril ATPase is altered with exhaustive exercise. PMID:6230276

  4. Vacuolar ATPase regulates surfactant secretion in rat alveolar type II cells by modulating lamellar body calcium.

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    Narendranath Reddy Chintagari

    Full Text Available Lung surfactant reduces surface tension and maintains the stability of alveoli. How surfactant is released from alveolar epithelial type II cells is not fully understood. Vacuolar ATPase (V-ATPase is the enzyme responsible for pumping H(+ into lamellar bodies and is required for the processing of surfactant proteins and the packaging of surfactant lipids. However, its role in lung surfactant secretion is unknown. Proteomic analysis revealed that vacuolar ATPase (V-ATPase dominated the alveolar type II cell lipid raft proteome. Western blotting confirmed the association of V-ATPase a1 and B1/2 subunits with lipid rafts and their enrichment in lamellar bodies. The dissipation of lamellar body pH gradient by Bafilomycin A1 (Baf A1, an inhibitor of V-ATPase, increased surfactant secretion. Baf A1-stimulated secretion was blocked by the intracellular Ca(2+ chelator, BAPTA-AM, the protein kinase C (PKC inhibitor, staurosporine, and the Ca(2+/calmodulin-dependent protein kinase II (CaMKII, KN-62. Baf A1 induced Ca(2+ release from isolated lamellar bodies. Thapsigargin reduced the Baf A1-induced secretion, indicating cross-talk between lamellar body and endoplasmic reticulum Ca(2+ pools. Stimulation of type II cells with surfactant secretagogues dissipated the pH gradient across lamellar bodies and disassembled the V-ATPase complex, indicating the physiological relevance of the V-ATPase-mediated surfactant secretion. Finally, silencing of V-ATPase a1 and B2 subunits decreased stimulated surfactant secretion, indicating that these subunits were crucial for surfactant secretion. We conclude that V-ATPase regulates surfactant secretion via an increased Ca(2+ mobilization from lamellar bodies and endoplasmic reticulum, and the activation of PKC and CaMKII. Our finding revealed a previously unrealized role of V-ATPase in surfactant secretion.

  5. Vacuolar ATPase regulates surfactant secretion in rat alveolar type II cells by modulating lamellar body calcium.

    Science.gov (United States)

    Chintagari, Narendranath Reddy; Mishra, Amarjit; Su, Lijing; Wang, Yang; Ayalew, Sahlu; Hartson, Steven D; Liu, Lin

    2010-01-01

    Lung surfactant reduces surface tension and maintains the stability of alveoli. How surfactant is released from alveolar epithelial type II cells is not fully understood. Vacuolar ATPase (V-ATPase) is the enzyme responsible for pumping H(+) into lamellar bodies and is required for the processing of surfactant proteins and the packaging of surfactant lipids. However, its role in lung surfactant secretion is unknown. Proteomic analysis revealed that vacuolar ATPase (V-ATPase) dominated the alveolar type II cell lipid raft proteome. Western blotting confirmed the association of V-ATPase a1 and B1/2 subunits with lipid rafts and their enrichment in lamellar bodies. The dissipation of lamellar body pH gradient by Bafilomycin A1 (Baf A1), an inhibitor of V-ATPase, increased surfactant secretion. Baf A1-stimulated secretion was blocked by the intracellular Ca(2+) chelator, BAPTA-AM, the protein kinase C (PKC) inhibitor, staurosporine, and the Ca(2+)/calmodulin-dependent protein kinase II (CaMKII), KN-62. Baf A1 induced Ca(2+) release from isolated lamellar bodies. Thapsigargin reduced the Baf A1-induced secretion, indicating cross-talk between lamellar body and endoplasmic reticulum Ca(2+) pools. Stimulation of type II cells with surfactant secretagogues dissipated the pH gradient across lamellar bodies and disassembled the V-ATPase complex, indicating the physiological relevance of the V-ATPase-mediated surfactant secretion. Finally, silencing of V-ATPase a1 and B2 subunits decreased stimulated surfactant secretion, indicating that these subunits were crucial for surfactant secretion. We conclude that V-ATPase regulates surfactant secretion via an increased Ca(2+) mobilization from lamellar bodies and endoplasmic reticulum, and the activation of PKC and CaMKII. Our finding revealed a previously unrealized role of V-ATPase in surfactant secretion. PMID:20169059

  6. Involvement of Ca(2+)-Dependent Hyperpolarization in Sleep Duration in Mammals.

    Science.gov (United States)

    Tatsuki, Fumiya; Sunagawa, Genshiro A; Shi, Shoi; Susaki, Etsuo A; Yukinaga, Hiroko; Perrin, Dimitri; Sumiyama, Kenta; Ukai-Tadenuma, Maki; Fujishima, Hiroshi; Ohno, Rei-ichiro; Tone, Daisuke; Ode, Koji L; Matsumoto, Katsuhiko; Ueda, Hiroki R

    2016-04-01

    The detailed molecular mechanisms underlying the regulation of sleep duration in mammals are still elusive. To address this challenge, we constructed a simple computational model, which recapitulates the electrophysiological characteristics of the slow-wave sleep and awake states. Comprehensive bifurcation analysis predicted that a Ca(2+)-dependent hyperpolarization pathway may play a role in slow-wave sleep and hence in the regulation of sleep duration. To experimentally validate the prediction, we generate and analyze 21 KO mice. Here we found that impaired Ca(2+)-dependent K(+) channels (Kcnn2 and Kcnn3), voltage-gated Ca(2+) channels (Cacna1g and Cacna1h), or Ca(2+)/calmodulin-dependent kinases (Camk2a and Camk2b) decrease sleep duration, while impaired plasma membrane Ca(2+) ATPase (Atp2b3) increases sleep duration. Pharmacological intervention and whole-brain imaging validated that impaired NMDA receptors reduce sleep duration and directly increase the excitability of cells. Based on these results, we propose a hypothesis that a Ca(2+)-dependent hyperpolarization pathway underlies the regulation of sleep duration in mammals. PMID:26996081

  7. Ca2+ sparks and Ca2+ glows in superior cervical ganglion neurons

    Institute of Scientific and Technical Information of China (English)

    Li-jun YAO; Cai-hong WU; Jie LIU; Zhuan ZHOU; He-ping CHENG; Gang WANG; Kun-fu OU-YANG; Chao-liang WEI; Xian-hua WANG; Shi-rong WANG; Wei YAO; Hong-ping HUANG; Jian-hong LUO

    2006-01-01

    Aim: Ca2+ release from the endoplasmic reticulum (ER) is an integral component of neuronal Ca2+ signaling. The present study is to investigate properties of local Ca2+ release events in superior cervical ganglion (SCO) neurons. Methods: Primary cultured SCO neurons were prepared from neonatal rats (P3-P7). Low concentration of caffeine was used to induce Ca2+ release from the ER Ca2+ store, and intracellular Ca2+ was recorded by high-resolution line scan confocal imaging and the Ca2+ indicator Fluo-4. Results: Two populations of local Ca2+ release events with distinct temporal characteristics were evoked by 1.5 mmol/L caffeine near the surface membrane in the soma and the neurites of SCG neurons. Brief events similar to classic Ca2+ sparks lasted a few hundreds of milliseconds, whereas long-lasting events displayed duration up to tens of seconds. Typical somatic and neurite sparks were of 0.3- and 0.52-fold increase in local Fluo-4 fluorescence, respectively. Typical Ca2+ glows were brighter (△F/F0 approximately 0.6), but were highly confined in space. The half maximum of full duration of neurite sparks was much longer than those in the soma (685 vs 381 ms). Conclusion: Co-existence of Ca2+ sparks and Ca2+ glows in SCG neurons indicates distinctive local regulation of Ca2+ release kinetics. The local Ca2+ signals of variable, site-specific temporal length may bear important implications in encoding a "memory" of the trigger signal.

  8. Interleukin-1β activates an Src family kinase to stimulate the plasma membrane Ca2+ pump in hippocampal neurons.

    Science.gov (United States)

    Ghosh, Biswarup; Green, Matthew V; Krogh, Kelly A; Thayer, Stanley A

    2016-04-01

    The plasma membrane Ca(2+) ATPase (PMCA) plays a major role in clearing Ca(2+) from the neuronal cytoplasm. The cytoplasmic Ca(2+) clearance rate affects neuronal excitability, synaptic plasticity, and neurotransmission. Here, we examined the modulation of PMCA activity by PTKs in hippocampal neurons. PMCA-mediated Ca(2+) clearance slowed in the presence of pyrazolopyrimidine 2, an inhibitor of Src family kinases (SFKs), and accelerated in the presence of C2-ceramide, an activator of PTKs. Ca(2+) clearance kinetics were attenuated in cells expressing a dominant-negative Src mutant, suggesting that the pump is tonically stimulated by a PTK. Tonic stimulation was reduced in hippocampal neurons expressing short hairpin (sh)RNA directed to mRNA for Yes. shRNA-mediated knockdown of PMCA isoform 1 (PMCA1) removed tonic stimulation of Ca(2+) clearance, indicating that the kinase stimulates PMCA1. IL-1β accelerated Ca(2+) clearance in a manner blocked by an IL-1β receptor antagonist or by an inhibitor of neutral sphingomyelinase, the enzyme that produces ceramide. Thus IL-1β activates an SFK to stimulate the plasma membrane Ca(2+) pump, decreasing the duration of Ca(2+) transients in hippocampal neurons. PMID:26843596

  9. Inhibition of oxidative phosphorylation in ascites tumor mitochondria and cells by intramitochondrial Ca2+.

    Science.gov (United States)

    Villalobo, A; Lehninger, A L

    1980-03-25

    Accumulation of Ca2+ (+ phosphate) by respiring mitochondria from Ehrlich ascites or AS30-D hepatoma tumor cells inhibits subsequent phosphorylating respiration in response to ADP. The respiratory chain is still functional since a proton-conducting uncoupler produces a normal stimulation of electron transport. The inhibition of phosphorylating respiration is caused by intramitochondrial Ca2+ (+ phosphate). ATP + Mg2+ together, but not singly, prevents the inhibitory action of Ca2+. Neither AMP, GTP, GDP, nor any other nucleoside 5'-triphosphate or 5'-diphosphate could replace ATP in this effect. Phosphorylating respiration on NAD(NADP)-linked substrates was much more susceptible to the inhibitory effect of intramitochondrial Ca2+ than succinate-linked respiration. Significant inhibition of oxidative phosphorylation is given by the endogenous Ca2+ present in freshly isolated tumor mitochondria. The phosphorylating respiration of permeabilized Ehrlich ascites tumor cells is also inhibited by Ca2+ accumulated by the mitochondria in situ. Possible causes of the Ca2+-induced inhibition of oxidative phosphorylation are considered.

  10. The relative contribution of NMDARs to excitatory postsynaptic currents is controlled by Ca2+-induced inactivation.

    Directory of Open Access Journals (Sweden)

    Fliza eValiullina

    2016-01-01

    Full Text Available NMDA receptors (NMDARs are important mediators of excitatory synaptic transmission and plasticity. A hallmark of these channels is their high permeability to Ca2+. At the same time, they are themselves inhibited by the elevation of intracellular Ca2+ concentration. It is unclear however, whether the Ca2+ entry associated with single NMDAR mediated synaptic events is sufficient to self-inhibit their activation. Such auto-regulation would have important effects on the dynamics of synaptic excitation in several central networks. Therefore, we studied NMDAR-mediated synaptic currents in mouse hippocampal CA1 pyramidal neurons. Postsynaptic responses to subthreshold Schaffer collateral stimulation depended strongly on the absence or presence of intracellular Ca2+ buffers. Loading of pyramidal cells with exogenous Ca2+ buffers increased the amplitude and decay time of NMDAR mediated EPSCs (EPSP and prolonged the time window for action potential generation.Our data indicate that the Ca2+ influx mediated by unitary synaptic events is sufficient to produce detectable self-inhibition of NMDARs even at a physiological Mg2+ concentration. Therefore, the contribution of NMDARs to synaptic excitation is strongly controlled by both previous synaptic activity as well as by the Ca2+ buffer capacity of postsynaptic neurons.

  11. Inhibition of oxidative phosphorylation in ascites tumor mitochondria and cells by intramitochondrial Ca2+.

    Science.gov (United States)

    Villalobo, A; Lehninger, A L

    1980-03-25

    Accumulation of Ca2+ (+ phosphate) by respiring mitochondria from Ehrlich ascites or AS30-D hepatoma tumor cells inhibits subsequent phosphorylating respiration in response to ADP. The respiratory chain is still functional since a proton-conducting uncoupler produces a normal stimulation of electron transport. The inhibition of phosphorylating respiration is caused by intramitochondrial Ca2+ (+ phosphate). ATP + Mg2+ together, but not singly, prevents the inhibitory action of Ca2+. Neither AMP, GTP, GDP, nor any other nucleoside 5'-triphosphate or 5'-diphosphate could replace ATP in this effect. Phosphorylating respiration on NAD(NADP)-linked substrates was much more susceptible to the inhibitory effect of intramitochondrial Ca2+ than succinate-linked respiration. Significant inhibition of oxidative phosphorylation is given by the endogenous Ca2+ present in freshly isolated tumor mitochondria. The phosphorylating respiration of permeabilized Ehrlich ascites tumor cells is also inhibited by Ca2+ accumulated by the mitochondria in situ. Possible causes of the Ca2+-induced inhibition of oxidative phosphorylation are considered. PMID:6766937

  12. Plasma membrane Ca2+-ATPase isoforms composition regulates cellular pH homeostasis in differentiating PC12 cells in a manner dependent on cytosolic Ca2+ elevations

    DEFF Research Database (Denmark)

    Boczek, Tomasz; Lisek, Malwina; Ferenc, Bozena;

    2014-01-01

    Plasma membrane Ca2+-ATPase (PMCA) by extruding Ca2+ outside the cell, actively participates in the regulation of intracellular Ca2+ concentration. Acting as Ca2+/H+ counter-transporter, PMCA transports large quantities of protons which may affect organellar pH homeostasis. PMCA exists in four is...

  13. Regulated release of Ca2+ from respiring mitochondria by Ca2+/2H+ antiport.

    Science.gov (United States)

    Fiskum, G; Lehninger, A L

    1979-07-25

    Simultaneous measurements of oxygen consumption and transmembrane transport of Ca2+, H+, and phosphate show that the efflux of Ca2+ from respiring tightly coupled rat liver mitochondria takes place by an electroneutral Ca2+/2H+ antiport process that is ruthenium red-insensitive and that is regulated by the oxidation-reduction state of the mitochondrial pyridine nucleotides. When mitochondrial pyridine nucleotides are kept in a reduced steady state, the efflux of Ca2+ is inhibited; when they are in an oxidized state, Ca2+ efflux is activated. These processes were demonstrated by allowing phosphate-depleted mitochondria respiring on succinate in the presence of rotenone to take up Ca2+ from the medium. Upon subsequent addition of ruthenium red to block Ca2+ transport via the electrophoretic influx pathway, and acetoacetate, to bring mitochondrial pyridine nucleotides into the oxidized state, Ca2+ efflux and H+ influx ensued. The observed H+ influx/Ca2+ efflux ratio was close to the value 2.0 predicted for the operation of an electrically neutral Ca2+/2H+ antiport process.

  14. Reduced endogenous Ca2+ buffering speeds active zone Ca2+ signaling.

    Science.gov (United States)

    Delvendahl, Igor; Jablonski, Lukasz; Baade, Carolin; Matveev, Victor; Neher, Erwin; Hallermann, Stefan

    2015-06-01

    Fast synchronous neurotransmitter release at the presynaptic active zone is triggered by local Ca(2+) signals, which are confined in their spatiotemporal extent by endogenous Ca(2+) buffers. However, it remains elusive how rapid and reliable Ca(2+) signaling can be sustained during repetitive release. Here, we established quantitative two-photon Ca(2+) imaging in cerebellar mossy fiber boutons, which fire at exceptionally high rates. We show that endogenous fixed buffers have a surprisingly low Ca(2+)-binding ratio (∼ 15) and low affinity, whereas mobile buffers have high affinity. Experimentally constrained modeling revealed that the low endogenous buffering promotes fast clearance of Ca(2+) from the active zone during repetitive firing. Measuring Ca(2+) signals at different distances from active zones with ultra-high-resolution confirmed our model predictions. Our results lead to the concept that reduced Ca(2+) buffering enables fast active zone Ca(2+) signaling, suggesting that the strength of endogenous Ca(2+) buffering limits the rate of synchronous synaptic transmission. PMID:26015575

  15. Regulated release of Ca2+ from respiring mitochondria by Ca2+/2H+ antiport.

    Science.gov (United States)

    Fiskum, G; Lehninger, A L

    1979-07-25

    Simultaneous measurements of oxygen consumption and transmembrane transport of Ca2+, H+, and phosphate show that the efflux of Ca2+ from respiring tightly coupled rat liver mitochondria takes place by an electroneutral Ca2+/2H+ antiport process that is ruthenium red-insensitive and that is regulated by the oxidation-reduction state of the mitochondrial pyridine nucleotides. When mitochondrial pyridine nucleotides are kept in a reduced steady state, the efflux of Ca2+ is inhibited; when they are in an oxidized state, Ca2+ efflux is activated. These processes were demonstrated by allowing phosphate-depleted mitochondria respiring on succinate in the presence of rotenone to take up Ca2+ from the medium. Upon subsequent addition of ruthenium red to block Ca2+ transport via the electrophoretic influx pathway, and acetoacetate, to bring mitochondrial pyridine nucleotides into the oxidized state, Ca2+ efflux and H+ influx ensued. The observed H+ influx/Ca2+ efflux ratio was close to the value 2.0 predicted for the operation of an electrically neutral Ca2+/2H+ antiport process. PMID:36390

  16. Enhancing the potency of lithospermate B for inhibiting Na+/K+-ATPase activity by forming transition metal ion complexes

    Institute of Scientific and Technical Information of China (English)

    Nan-Hei LIN; Tse-Yu CHUNG; Feng-Yin LI; Hsin-An CHEN; Jason TC TZEN

    2013-01-01

    Aim:To determine whether replacing Mg2+ in magnesium lithospermate B (Mg-LSB) isolated from danshen (Salvia miltiorrhiza) with other metal ions could affect its potency in inhibition of Na+/K+-ATPase activity.Methods:Eight metal ions (Na+,K+,Mg2+,Cr3+,Mn2+,Co2+,Ni2+,and Zn2+) were used to form complexes with LSB.The activity of Na+/K+-ATPase was determined by measuring the amount of inorganic phosphate (Pi) liberated from ATP.Human adrenergic neuroblastoma cell line SH-SY5Y was used to assess the intracellular Ca2+ level fluctuation and cell viability.The metal binding site on LSB and the binding mode of the metal-LSB complexes were detected by NMR and visible spectroscopy,respectively.Results:The potencies of LSB complexed with Cr3+,Mn2+,Co2+,or Ni2+ increased by approximately 5 times compared to the naturally occurring LSB and Mg-LSB.The IC50 values of Cr-LSB,Mn-LSB,Co-LSB,Ni-LSB,LSB,and Mg-LSB in inhibition of Na+/K+-ATPase activity were 23,17,26,25,101,and 128 μmoVL,respectively.After treatment of SH-SY5Y cells with the transition metal-LSB complexes (25 μmol/L),the intracellular Ca2+ level was substantially elevated,and the cells were viable for one day.The transition metals,as exemplified by Co2+,appeared to be coordinated by two carboxylate groups and one carbonyl group of LSB.Titration of LSB against Co2+ demonstrated that the Co-LSB complex was formed with a C02+:LSB molar ratio of 1:2 or 1:1,when [Co2+] was less than half of the [LSB] or higher than the [LSB],respectively.Conclusion:LSB complexed with Cr3+,Mn2+,Co2+,or Ni2+ are stable,non-toxic and more potent in inhibition of Na+/K+-ATPase.The transition metal-LSB complexes have the potential to be superior substitutes for cardiac glycosides in the treatment of congestive heart failure.

  17. Obesity induces upregulation of genes involved in myocardial Ca2+ handling

    Directory of Open Access Journals (Sweden)

    A.P. Lima-Leopoldo

    2008-07-01

    Full Text Available Obesity is a complex multifactorial disorder that is often associated with cardiovascular diseases. Research on experimental models has suggested that cardiac dysfunction in obesity might be related to alterations in myocardial intracellular calcium (Ca2+ handling. However, information about the expression of Ca2+-related genes that lead to this abnormality is scarce. We evaluated the effects of obesity induced by a high-fat diet in the expression of Ca2+-related genes, focusing the L-type Ca2+ channel (Cacna1c, sarcolemmal Na+/Ca2+ exchanger (NCX, sarcoplasmic reticulum Ca2+ ATPase (SERCA2a, ryanodine receptor (RyR2, and phospholamban (PLB mRNA in rat myocardium. Male 30-day-old Wistar rats were fed a standard (control or high-fat diet (obese for 15 weeks. Obesity was defined as increased percent of body fat in carcass. The mRNA expression of Ca2+-related genes in the left ventricle was measured by RT-PCR. Compared with control rats, the obese rats had increased percent of body fat, area under the curve for glucose, and leptin and insulin plasma concentrations. Obesity also caused an increase in the levels of SERCA2a, RyR2 and PLB mRNA (P < 0.05 but did not modify the mRNA levels of Cacna1c and NCX. These findings show that obesity induced by high-fat diet causes cardiac upregulation of Ca2+ transport_related genes in the sarcoplasmic reticulum.

  18. Heat shock protein 70 protects PC12 cells against ischemia-hypoxia/reoxygenation by maintaining intracellular Ca2+ homeostasis

    Directory of Open Access Journals (Sweden)

    Yuan Liu

    2016-01-01

    Full Text Available Heat shock protein 70 (HSP70 maintains Ca2+ homeostasis in PC12 cells, which may protect against apoptosis; however, the mechanisms of neuroprotection are unclear. Therefore, in this study, we examined Ca2+ levels in PC12 cells transfected with an exogenous lentiviral HSP70 gene expression construct, and we subsequently subjected the cells to ischemia-hypoxia/reoxygenation injury. HSP70 overexpression increased neuronal viability and ATPase activity, and it decreased cellular reactive oxygen species levels and intracellular Ca2+ concentration after hypoxia/reoxygenation. HSP70 overexpression enhanced the protein and mRNA expression levels of sarcoplasmic/endoplasmic reticulum Ca2+-ATPase (SERCA, but it decreased the protein and mRNA levels of inositol 1,4,5-trisphosphate receptor (IP3R, thereby leading to decreased intracellular Ca2+ concentration after ischemia-hypoxia/reoxygenation. These results suggest that exogenous HSP70 protects against ischemia-hypoxia/reoxygenation injury, at least in part, by maintaining cellular Ca2+ homeostasis, by upregulating SERCA expression and by downregulating IP3R expression.

  19. Release of Ca2+ from the Endoplasmic Reticulum Contributes to Ca2+ Signaling in Dictyostelium discoideum

    OpenAIRE

    Wilczynska, Zofia; Happle, Kathrin; Müller-Taubenberger, Annette; Schlatterer, Christina; Malchow, Dieter; Fisher, Paul R.

    2005-01-01

    Ca2+ responses to two chemoattractants, folate and cyclic AMP (cAMP), were assayed in Dictyostelium D. discoideum mutants deficient in one or both of two abundant Ca2+-binding proteins of the endoplasmic reticulum (ER), calreticulin and calnexin. Mutants deficient in either or both proteins exhibited enhanced cytosolic Ca2+ responses to both attractants. Not only were the mutant responses greater in amplitude, but they also exhibited earlier onsets, faster rise rates, earlier peaks, and faste...

  20. Localized Ca2+ uncaging induces Ca2+ release through IP3R in smooth muscle

    Institute of Scientific and Technical Information of China (English)

    Min WANG; Zheng CHEN; Yan XING; Xu ZHANG; Xian-zhi DONG; Guang-ju JI

    2006-01-01

    Aim: Our previous study indicated that there are two types of Ca2+ release events seen in intact mouse bladder tissue. In this study our aim is to investigate the mechanism that underlies the phenomena of Ca2+ release in smooth muscle. Methods: Single cells were isolated and tissue segments were prepared by cutting the detrusor into 0.1 cm × O.5 cm strips running along the axis from the neck to the fundus. Single cells and intact tissue strips were co-loaded with the Ca2+ indicator and caged Ca2+ by incubation with 10 μmol/L Fluo-4 AM and DMNP-EDTA-AM. Fluo-4 AM fluorescence was detected by laser scanning confocal microscopy, and local uncaging of DMNP-EGTA was achieved by brief exposure to the output of a diode-pumped, Ti:sapphire laser tuned to 730 nm. Results: Local uncaging of caged Ca2+ was able to trigger Ca2+ release events in both single cells and tissue strips from mouse bladder. The Ca2+ release events could not be blocked by ryanodine alone, but the property of the Ca2+ release was markedly altered. Surprisingly, in the presence of ryanodine, Xestospongin C completely inhibited the Ca2+ release events both in single cell and tissue experiments. Conclusion: (1) Two photon flash photolysis (TPFP) triggers Ca2+ induced Ca2+ release. This process involves release through type 2 ryanodine receptor channels; (2) TPFP results in the release of Ca2+ through inositol 1,4,5-trisphosphate receptors in the absence of phospholipase C activation.

  1. Characterization of the Ca2+ current in freshly dissociated crustacean peptidergic neuronal somata.

    Science.gov (United States)

    Richmond, J E; Sher, E; Cooke, I M

    1995-06-01

    suggest that hyperpolarizing pulses are more effective in removing voltage-dependent inactivation, but also allow some recovery from Ca(2+)-dependent inactivation. 6. In the crab saline, which contained 24 mM Mg2+, the amplitudes of currents carried by 52 mM Ca2+, Sr2+ and Ba2+ were similar. Removing the Mg2+ from the saline augmented both the Ba2+ and Sr2+ currents relative to the Ca2+ current.(ABSTRACT TRUNCATED AT 400 WORDS)

  2. Endothelin-1 induces intracellular [Ca2+] increase via Ca2+ influx through the L-type Ca2+ channel, Ca2+-induced Ca2+ release and a pathway involving ETA receptors, PKC, PKA and AT1 receptors in cardiomyocytes

    Institute of Scientific and Technical Information of China (English)

    ZENG QingHua; LI XingTing; ZHONG GuoGan; ZHANG WenJie; SUN ChengWen

    2009-01-01

    Using fura-2-acetoxymethyl eater (AM) fluorescence imaging and patch clamp techniques, we found that endothelin-1 (ET-1) significantly elevated the intracellular calcium level ([Ca2+]1) in a dose-dependent manner and activated the L-type Ca2+ channel in cardiomyocytes isolated from rats.The effect of ET-1 on [Ca2+]1 elevation was abolished in the presence of the ETA receptor blocker BQ123,but was not affected by the ETa receptor blocker BQ788. ET-1-induced an increase in [Ca2+]1, which was inhibited 46.7% by pretreatment with a high concentration of ryanodine (10 μmol/L), a blocker of the ryanodine receptor. The ET-1-induced [Ca2+]i increase was also inhibited by the inhibltors of protein kinase A (PKA), protein kinase C (PKC) and angiotensin type 1 receptor (AT1 receptor). We found that ET-1 induced an enhancement of the amplitude of the whole cell L-type Ca2+ channel current and an Increase of open-state probability (NPo) of an L-type single Ca2+ channel. BQ123 completely blocked the ET-1-induced increase in calcium channel open-state probability. In this study we demonstrated that ET-1 regulates calcium overload through a series of mechanisms that include L-type Ca2+ channel activation and Ca2+-induced Ca2+ release (CICR). ETa receptors, PKC, PKA and AT1 receptors may also contribute to this pathway.

  3. Endothelin-1 induces intracellular [Ca2+] increase via Ca2+ influx through the L-type Ca2+ channel, Ca2+-induced Ca2+ release and a pathway involving ETA receptors, PKC, PKA and AT1 receptors in cardiomyocytes

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Using fura-2-acetoxymethyl ester (AM) fluorescence imaging and patch clamp techniques, we found that endothelin-1 (ET-1) significantly elevated the intracellular calcium level ([Ca2+]i) in a dose-dependent manner and activated the L-type Ca2+ channel in cardiomyocytes isolated from rats. The effect of ET-1 on [Ca2+]i elevation was abolished in the presence of the ETA receptor blocker BQ123, but was not affected by the ETB receptor blocker BQ788. ET-1-induced an increase in [Ca2+]i, which was inhibited 46.7% by pretreatment with a high concentration of ryanodine (10 μmol/L), a blocker of the ryanodine receptor. The ET-1-induced [Ca2+]i increase was also inhibited by the inhibitors of protein kinase A (PKA), protein kinase C (PKC) and angiotensin type 1 receptor (AT1 receptor). We found that ET-1 induced an enhancement of the amplitude of the whole cell L-type Ca2+ channel current and an increase of open-state probability (NPo) of an L-type single Ca2+ channel. BQ123 completely blocked the ET-1-induced increase in calcium channel open-state probability. In this study we demonstrated that ET-1 regulates calcium overload through a series of mechanisms that include L-type Ca2+ channel activation and Ca2+-induced Ca2+ release (CICR). ETA receptors, PKC, PKA and AT1 receptors may also contribute to this pathway.

  4. [Cationic specificity of a Ca2+-accumulating system in smooth muscle cell mitochondria].

    Science.gov (United States)

    Veklich, T O; Kosterin, S O; Shynlova, O P

    2002-01-01

    In the experiments conducted with application of an isotopic technique (45Ca2+) on the myometrium cells suspension treated by digitonin solution (0.1 mg/ml) some properties of Ca ions accumulation system in the mitochondria--cationic and substrate specificity as well as effects of Mg2+ and some other bivalent metals ions on the Ca2+ accumulation velocity have been estimated. Ca ions accumulation from the incubation medium containing 3 mM sodium succinate Na, 2 mM Pi (as potassium K(+)-phosphate buffer, pH 7.4 at 37 degrees C), 0.01 mM (40CaCl2 + 45CaCl2) and 100 nM thapsigargin--selective inhibiting agent of endoplasmatic reticulum calcium pump were demonstrated as detected just only in presence of Mg, while not Ni, Co or Cu ions. The increase of Mg2+ concentration from 1 x 10(-6) to 10(-3) M induced the ATP dependent transport activation in the myometrium mitochondria. Under [Mg2+] increase till 40 mM this cation essentially decreased Ca2+ accumulation (by 65% from the maximal value). The optimum for Ca2+ transport in the myometrium cells suspension is Mg2+ 10 mM concentration. Ka activation apparent constant along Mg2+ value (in presence 3 mM ATP and 3 mM sodium succinate) is 4.27 mM. The above listed bivalent metals decreased Mg2+, ATP-dependent accumulation of calcium, values of inhibition apparent constants for ions Co2+, Ni2+ and Cu2+ were--2.9 x 10(-4) M, 5.1 x 10(-5) M and 4.2 x 10(-6) M respectively. For Mg2+, ATP-dependent Ca2+ transport in the uterus myocytes mitocondria a high substrate specificity is a characteristic phenomenon in elation to ATP: GTP, CTP and UTP practically fail to provide for Ca accumulation process. PMID:12199098

  5. Heat and hyposmotic stimulation increase in [Ca2+]i by Ca2+ influx in rat synoviocytes

    Institute of Scientific and Technical Information of China (English)

    SUN WenWu; HU Fen; YANG WenXiu

    2008-01-01

    Rheumatoid arthritis (RA), which is marked by inflammatory synovitis, is a common, chronic autoimmune-disease, whose pathogenesis is complex and still unclear. In order to explore the effects of heat and hyposmotic stimuli on synoviocytes in rheumatoid arthritis, the changes of [Ca2+]i induced by heat, hyposmotic and 4α-PDD stimuli were observed in synoviocytes. [Ca2+]i elevation induced by heat ≥ 28℃, hyposmotic and 4α-PDD stimuli is found to be positively relative to increasing temperature, decreasing osmolality and rising concentration of 4α-PDD. Results show that there is reciprocity among these stimuli and desensitization, and that [Ca2+]i elevation depends on Ca2+ influx, but not necessarily links to Ca2+ release from intracellular stores and voltage-dependent Ca2+ channel in synoviocytes. The above characteristics of Ca2+ influx are similar to those of TRPV4. A probable mechanism has been suggested that heat and hyposmotic stimulation might increase the level of [Ca2+]i by activating the TRPV4-like channel and Ca2+ influx in the synoviocytes.

  6. Inhibition mechanism of the intracellular transporter Ca2+-pump from sarco-endoplasmic reticulum by the antitumor agent dimethyl-celecoxib.

    Directory of Open Access Journals (Sweden)

    Ramón Coca

    Full Text Available Dimethyl-celecoxib is a celecoxib analog that lacks the capacity as cyclo-oxygenase-2 inhibitor and therefore the life-threatening effects but retains the antineoplastic properties. The action mechanism at the molecular level is unclear. Our in vitro assays using a sarcoplasmic reticulum preparation from rabbit skeletal muscle demonstrate that dimethyl-celecoxib inhibits Ca2+-ATPase activity and ATP-dependent Ca2+ transport in a concentration-dependent manner. Celecoxib was a more potent inhibitor of Ca2+-ATPase activity than dimethyl-celecoxib, as deduced from the half-maximum effect but dimethyl-celecoxib exhibited higher inhibition potency when Ca2+ transport was evaluated. Since Ca2+ transport was more sensitive to inhibition than Ca2+-ATPase activity the drugs under study caused Ca2+/Pi uncoupling. Dimethyl-celecoxib provoked greater uncoupling and the effect was dependent on drug concentration but independent of Ca2+-pump functioning. Dimethyl-celecoxib prevented Ca2+ binding by stabilizing the inactive Ca2+-free conformation of the pump. The effect on the kinetics of phosphoenzyme accumulation and the dependence of the phosphoenzyme level on dimethyl-celecoxib concentration were independent of whether or not the Ca2+-pump was exposed to the drug in the presence of Ca2+ before phosphorylation. This provided evidence of non-preferential interaction with the Ca2+-free conformation. Likewise, the decreased phosphoenzyme level in the presence of dimethyl-celecoxib that was partially relieved by increasing Ca2+ was consistent with the mentioned effect on Ca2+ binding. The kinetics of phosphoenzyme decomposition under turnover conditions was not altered by dimethyl-celecoxib. The dual effect of the drug involves Ca2+-pump inhibition and membrane permeabilization activity. The reported data can explain the cytotoxic and anti-proliferative effects that have been attributed to the celecoxib analog. Ligand docking simulation predicts interaction of

  7. Swelling-activated Ca2+ channels trigger Ca2+ signals in Merkel cells.

    Directory of Open Access Journals (Sweden)

    Henry Haeberle

    Full Text Available Merkel cell-neurite complexes are highly sensitive touch receptors comprising epidermal Merkel cells and sensory afferents. Based on morphological and molecular studies, Merkel cells are proposed to be mechanosensory cells that signal afferents via neurotransmission; however, functional studies testing this hypothesis in intact skin have produced conflicting results. To test this model in a simplified system, we asked whether purified Merkel cells are directly activated by mechanical stimulation. Cell shape was manipulated with anisotonic solution changes and responses were monitored by Ca2+ imaging with fura-2. We found that hypotonic-induced cell swelling, but not hypertonic solutions, triggered cytoplasmic Ca2+ transients. Several lines of evidence indicate that these signals arise from swelling-activated Ca2+-permeable ion channels. First, transients were reversibly abolished by chelating extracellular Ca2+, demonstrating a requirement for Ca2+ influx across the plasma membrane. Second, Ca2+ transients were initially observed near the plasma membrane in cytoplasmic processes. Third, voltage-activated Ca2+ channel (VACC antagonists reduced transients by half, suggesting that swelling-activated channels depolarize plasma membranes to activate VACCs. Finally, emptying internal Ca2+ stores attenuated transients by 80%, suggesting Ca2+ release from stores augments swelling-activated Ca2+ signals. To identify candidate mechanotransduction channels, we used RT-PCR to amplify ion-channel transcripts whose pharmacological profiles matched those of hypotonic-evoked Ca2+ signals in Merkel cells. We found 11 amplicons, including PKD1, PKD2, and TRPC1, channels previously implicated in mechanotransduction in other cells. Collectively, these results directly demonstrate that Merkel cells are activated by hypotonic-evoked swelling, identify cellular signaling mechanisms that mediate these responses, and support the hypothesis that Merkel cells contribute

  8. Calmodulin transduces Ca2+ oscillations into differential regulation of its target proteins.

    Science.gov (United States)

    Slavov, Nikolai; Carey, Jannette; Linse, Sara

    2013-04-17

    Diverse physiological processes are regulated differentially by Ca(2+) oscillations through the common regulatory hub calmodulin. The capacity of calmodulin to combine specificity with promiscuity remains to be resolved. Here we propose a mechanism based on the molecular properties of calmodulin, its two domains with separate Ca(2+) binding affinities, and target exchange rates that depend on both target identity and Ca(2+) occupancy. The binding dynamics among Ca(2+), Mg(2+), calmodulin, and its targets were modeled with mass-action differential equations based on experimentally determined protein concentrations and rate constants. The model predicts that the activation of calcineurin and nitric oxide synthase depends nonmonotonically on Ca(2+)-oscillation frequency. Preferential activation reaches a maximum at a target-specific frequency. Differential activation arises from the accumulation of inactive calmodulin-target intermediate complexes between Ca(2+) transients. Their accumulation provides the system with hysteresis and favors activation of some targets at the expense of others. The generality of this result was tested by simulating 60 000 networks with two, four, or eight targets with concentrations and rate constants from experimentally determined ranges. Most networks exhibit differential activation that increases in magnitude with the number of targets. Moreover, differential activation increases with decreasing calmodulin concentration due to competition among targets. The results rationalize calmodulin signaling in terms of the network topology and the molecular properties of calmodulin.

  9. Mitochondrial Ca2+ uptake in skeletal muscle health and disease

    CERN Document Server

    Zhou, Jingsong; Yi, Jianxun

    2016-01-01

    Muscle uses Ca2+ as a messenger to control contraction and relies on ATP to maintain the intracellular Ca2+ homeostasis. Mitochondria are the major sub-cellular organelle of ATP production. With a negative inner membrane potential, mitochondria take up Ca2+ from their surroundings, a process called mitochondrial Ca2+ uptake. Under physiological conditions, Ca2+ uptake into mitochondria promotes ATP production. Excessive uptake causes mitochondrial Ca2+ overload, which activates downstream adverse responses leading to cell dysfunction. Moreover, mitochondrial Ca2+ uptake could shape spatio-temporal patterns of intracellular Ca2+ signaling. Malfunction of mitochondrial Ca2+ uptake is implicated in muscle degeneration. Unlike non-excitable cells, mitochondria in muscle cells experience dramatic changes of intracellular Ca2+ levels. Besides the sudden elevation of Ca2+ level induced by action potentials, Ca2+ transients in muscle cells can be as short as a few milliseconds during a single twitch or as long as min...

  10. Mitochondrial calcium uniporter MCU supports cytoplasmic Ca2+ oscillations, store-operated Ca2+ entry and Ca2+-dependent gene expression in response to receptor stimulation.

    Science.gov (United States)

    Samanta, Krishna; Douglas, Sophie; Parekh, Anant B

    2014-01-01

    Ca2+ flux into mitochondria is an important regulator of cytoplasmic Ca2+ signals, energy production and cell death pathways. Ca2+ uptake can occur through the recently discovered mitochondrial uniporter channel (MCU) but whether the MCU is involved in shaping Ca2+ signals and downstream responses to physiological levels of receptor stimulation is unknown. Here, we show that modest stimulation of leukotriene receptors with the pro-inflammatory signal LTC4 evokes a series of cytoplasmic Ca2+ oscillations that are rapidly and faithfully propagated into mitochondrial matrix. Knockdown of MCU or mitochondrial depolarisation, to reduce the driving force for Ca2+ entry into the matrix, prevents the mitochondrial Ca2+ rise and accelerates run down of the oscillations. The loss of cytoplasmic Ca2+ oscillations appeared to be a consequence of enhanced Ca2+-dependent inactivation of InsP3 receptors, which arose from the loss of mitochondrial Ca2+ buffering. Ca2+ dependent gene expression in response to leukotriene receptor activation was suppressed following knockdown of the MCU. In addition to buffering Ca2+ release, mitochondria also sequestrated Ca2+ entry through store-operated Ca2+ channels and this too was prevented following loss of MCU. MCU is therefore an important regulator of physiological pulses of cytoplasmic Ca2+.

  11. Mitochondrial calcium uniporter MCU supports cytoplasmic Ca2+ oscillations, store-operated Ca2+ entry and Ca2+-dependent gene expression in response to receptor stimulation.

    Directory of Open Access Journals (Sweden)

    Krishna Samanta

    Full Text Available Ca2+ flux into mitochondria is an important regulator of cytoplasmic Ca2+ signals, energy production and cell death pathways. Ca2+ uptake can occur through the recently discovered mitochondrial uniporter channel (MCU but whether the MCU is involved in shaping Ca2+ signals and downstream responses to physiological levels of receptor stimulation is unknown. Here, we show that modest stimulation of leukotriene receptors with the pro-inflammatory signal LTC4 evokes a series of cytoplasmic Ca2+ oscillations that are rapidly and faithfully propagated into mitochondrial matrix. Knockdown of MCU or mitochondrial depolarisation, to reduce the driving force for Ca2+ entry into the matrix, prevents the mitochondrial Ca2+ rise and accelerates run down of the oscillations. The loss of cytoplasmic Ca2+ oscillations appeared to be a consequence of enhanced Ca2+-dependent inactivation of InsP3 receptors, which arose from the loss of mitochondrial Ca2+ buffering. Ca2+ dependent gene expression in response to leukotriene receptor activation was suppressed following knockdown of the MCU. In addition to buffering Ca2+ release, mitochondria also sequestrated Ca2+ entry through store-operated Ca2+ channels and this too was prevented following loss of MCU. MCU is therefore an important regulator of physiological pulses of cytoplasmic Ca2+.

  12. Demethoxycurcumin is a potent inhibitor of P-type ATPases from diverse kingdoms of life

    DEFF Research Database (Denmark)

    Dao, Trong Tuan; Sehgal, Pankaj; Thanh Tung, Truong;

    2016-01-01

    the curcuminoids, demethoxycurcumin was the most potent inhibitor of all tested P-type ATPases from fungal (Pma1p; H+-ATPase), plant (AHA2; H+-ATPase) and animal (SERCA; Ca2+-ATPase) cells. All three curcuminoids acted as non-competitive antagonist to ATP and hence may bind to a highly conserved allosteric site......P-type ATPases catalyze the active transport of cations and phospholipids across biological membranes. Members of this large family are involved in a range of fundamental cellular processes. To date, a substantial number of P-type ATPase inhibitors have been characterized, some of which are used...... as drugs. In this work a library of natural compounds was screened and we first identified curcuminoids as plasma membrane H+-ATPases inhibitors in plant and fungal cells. We also found that some of the commercial curcumins contain several curcuminoids. Three of these were purified and, among...

  13. The impact of extracellular and intracellular Ca2+ on ethanol-induced smooth muscle contraction

    Institute of Scientific and Technical Information of China (English)

    Naciye YAKTUBAY DONDAS; Mahir KAPLAN; Derya KAYA; Ergin SiNGiRiK

    2009-01-01

    Aim:To evaluate the impact of extracellular and intracellular Ca~(2+) on contractions induced by ethanol in smooth muscle.Methods: Longitudinal smooth muscle strips were prepared from the gastric fundi of mice. The contractions of smooth muscle strips were recorded with an isometric force displacement transducer.Results: Ethanol (164 mmol/L) produced reproducible contractions in isolated gastric fundal strips of mice. Although lidocaine (50 and 100 μmol/L), a local anesthetic agent, and hexamethonium (100 and 500 μmol/L), a ganglionic blocking agent, failed to affect these contractions, verapamil (1-50 μmol/L) and nifedipine (1-50 μmol/L), selective blockers of L-type Ca~(2+) channels, significantly inhibited the contractile responses of ethanol. Using a Ca~(2+)-free medium nearly eliminated these contractions in the same tissue. Ryanodine (1-50 μmol/L) and ruthenium red (10-100 μmol/L), selective blockers of intracellular Ca~(2+) channels/ryanodine receptors; cyclopiazonic acid (CPA; 1-10 μmol/L), a selective inhibitor of sarcoplasmic reticulum (SR) Ca~(2+)-ATPase; and caffeine (0.5-5 mmol/L), a depleting agent of intracellular Ca~(2+) stores, significantly inhibited the contractile responses induced by ethanol. In addition, the com-bination of caffeine (5 mmol/L) plus CPA (10 μmol/L), and ryanodine (10 μmol/L) plus CPA (10 μmol/L), caused further inhibition of contractions in response to ethanol. This inhibition was significantly different from those associated with caffeine, ryanodine or CPA. Furthermore the combination of caffeine (5 mmol/L), ryanodine (10 μmol/L) and CPA(10 μmol/L) eliminated the contractions induced by ethanol in isolated gastric fundal strips of mice.Conclusion: Both extracellular and intracellular Ca~(2+) may have important roles in regulating contractions induced by ethanol in the mouse gastric fundus.

  14. Effects of benzo(a)pyrene exposure on oxidative stress and ATPase in the hippocampus of rats%苯并[a]芘对大鼠海马组织氧化应激及ATP酶的影响

    Institute of Scientific and Technical Information of China (English)

    段利; 汤艳; 陈承志; 彭斌; 邱崇莹; 戚友宾; 涂白杰

    2013-01-01

    目的 通过研究苯并[a]芘(B[a]P)对大鼠行为学、海马氧化应激及ATP酶的影响,探讨B[a]P的神经行为毒性分子机制.方法 将120只21d龄雄性SD大鼠,随机分为空白对照组、植物油组(溶剂对照组),2.5、5.0、10.0 mg/kg B[a]P染毒组,每组24只.腹腔注射给药,每天1次,连续4周.染毒结束后,用Morris水迷宫和穿梭箱检测学习记忆能力;用化学比色法测定海马超氧化物歧化酶(SOD)、Na+-K+-ATP酶和Ca2+-Mg2+-ATP酶的活力及丙二醛(MDA)含量;用荧光标记方法测定海马Ca2+浓度.结果 各染毒组大鼠的水迷宫逃避潜伏期、穿梭箱主动回避反应潜伏期(AARL)和被动回避反应潜伏期(RARL)均明显高于空白对照组和溶剂对照组,水迷宫末次跨平台次数和穿梭箱主动回避反应次数(AARF)均明显低于空白对照组和溶剂对照组,差异均有统计学意义(P<0.05);且呈剂量-效应关系.与空白对照组和溶剂对照组比较,染毒组大鼠海马组织SOD活力、Na+-K+-ATP酶和ca2+-Mg2+-ATP酶活力明显下降,且呈剂量-效应关系,差异均有统计学意义(P<0.05).染毒组大鼠海马组织MDA含量、Ca2+浓度均明显高于空白对照组和溶剂对照组,且呈剂量-效应关系,差异均有统计学意义(P<0.05).结论 B[a]P所致神经行为毒性,可能与染毒后大鼠海马组织氧化应激受损,Na+-K+-ATP酶和Ca2+-Mg2+-ATP酶活力下降有关.%Objective To investigate the effects of benzo[a]pyrene (B[a]P) exposure on the behaviors and hippocampal oxidative stress and ATPase in rats and the molecular mechanism of neurobehavioral toxicity of B[a]P.Methods A total of 120 male SD rats (21 days old) were randomly and equally assigned to five groups:blank control group,vegetable oil (solvent control) group,and 2.5,5,and 10 mg/kg B[a]P exposure groups.The rats in B [a]P exposure groups were injected intraperitoneally with B[a]P once a day for 4 consecutive weeks.Then,Morris water maze and

  15. Modeling the contributions of Ca2+ flows to spontaneous Ca2+ oscillations and cortical spreading depression-triggered Ca2+ waves in astrocyte networks.

    Directory of Open Access Journals (Sweden)

    Bing Li

    Full Text Available Astrocytes participate in brain functions through Ca(2+ signals, including Ca(2+ waves and Ca(2+ oscillations. Currently the mechanisms of Ca(2+ signals in astrocytes are not fully clear. Here, we present a computational model to specify the relative contributions of different Ca(2+ flows between the extracellular space, the cytoplasm and the endoplasmic reticulum of astrocytes to the generation of spontaneous Ca(2+ oscillations (CASs and cortical spreading depression (CSD-triggered Ca(2+ waves (CSDCWs in a one-dimensional astrocyte network. This model shows that CASs depend primarily on Ca(2+ released from internal stores of astrocytes, and CSDCWs depend mainly on voltage-gated Ca(2+ influx. It predicts that voltage-gated Ca(2+ influx is able to generate Ca(2+ waves during the process of CSD even after depleting internal Ca(2+ stores. Furthermore, the model investigates the interactions between CASs and CSDCWs and shows that the pass of CSDCWs suppresses CASs, whereas CASs do not prevent the generation of CSDCWs. This work quantitatively analyzes the generation of astrocytic Ca(2+ signals and indicates different mechanisms underlying CSDCWs and non-CSDCWs. Research on the different types of Ca(2+ signals might help to understand the ways by which astrocytes participate in information processing in brain functions.

  16. Ca2+ Alternans in a Cardiac Myocyte Model that Uses Moment Equations to Represent Heterogeneous Junctional SR Ca2+

    OpenAIRE

    Huertas, Marco A; Smith, Gregory D.; Györke, Sándor

    2010-01-01

    Multiscale whole-cell models that accurately represent local control of Ca2+-induced Ca2+ release in cardiac myocytes can reproduce high-gain Ca2+ release that is graded with changes in membrane potential. Using a recently introduced formalism that represents heterogeneous local Ca2+ using moment equations, we present a model of cardiac myocyte Ca2+ cycling that exhibits alternating sarcoplasmic reticulum (SR) Ca2+ release when periodically stimulated by depolarizing voltage pulses. The model...

  17. Thermodynamic properties of Mg2Si and Mg2Ge investigated by first principles method

    International Nuclear Information System (INIS)

    The lattice dynamics and thermodynamic properties of Mg2Si and Mg2Ge are studied based on the first principles calculations. We obtain the phonon dispersion curves and phonon density of states spectra using the density functional perturbation theory with local density approximations. By employing the quasi-harmonic approximation, we calculate the temperature dependent Helmholtz free energy, bulk modulus, thermal expansion coefficient, specific heat, Debye temperature and overall Grueneisen coefficient. The results are in good agreement with available experimental data and previous theoretical studies. The thermal conductivities of both compounds are then estimated with the Slack's equation. By carefully choosing input parameters, especially the acoustic Debye temperature, we find that the calculated thermal conductivities agree fairly well with the experimental values above 80 K for both compounds. This demonstrates that the lattice thermal conductivity of simple cubic semiconductors may be estimated with satisfactory accuracy by combining the Slack's equation with the necessary thermodynamics parameters derived completely from the first principles calculations.

  18. Curcumin modulation of Na,K-ATPase: phosphoenzyme accumulation, decreased K+ occlusion, and inhibition of hydrolytic activity

    OpenAIRE

    Mahmmoud, Yasser A.

    2005-01-01

    Curcumin, the major constitute of tumeric, is an important nutraceutical that has been shown to be useful in the treatment of many diseases. As an inhibitor of the sarcoplasmic reticulum Ca2+-ATPase, curcumin was shown to correct cystic fibrosis (CF) defects in some model systems, whereas others have reported no or little effects on CF after curcumin treatment, suggesting that curcumin effect is not due to simple inhibition of the Ca2+-ATPase.We tested the hypothesis that curcumin may modulat...

  19. Neuronal Ca(2+) dyshomeostasis in Huntington disease.

    Science.gov (United States)

    Giacomello, Marta; Oliveros, Juan C; Naranjo, Jose R; Carafoli, Ernesto

    2013-01-01

    The expansion of the N-terminal poly-glutamine tract of the huntingtin (Htt) protein is responsible for Huntington disease (HD). A large number of studies have explored the neuronal phenotype of HD, but the molecular aethiology of the disease is still very poorly understood. This has hampered the development of an appropriate therapeutical strategy to at least alleviate its symptoms. In this short review, we have focused our attention on the alteration of a specific cellular mechanism common to all HD models, either genetic or induced by treatment with 3-NPA, i.e. the cellular dyshomeostasis of Ca(2+). We have highlighted the direct and indirect (i.e. transcriptionally mediated) effects of mutated Htt on the maintenance of the intracellular Ca(2+) balance, the correct modulation of which is fundamental to cell survival and the disturbance of which plays a key role in the death of the cell.

  20. Mitochondrial Ca(2+) uptake in skeletal muscle health and disease.

    Science.gov (United States)

    Zhou, Jingsong; Dhakal, Kamal; Yi, Jianxun

    2016-08-01

    Muscle uses Ca(2+) as a messenger to control contraction and relies on ATP to maintain the intracellular Ca(2+) homeostasis. Mitochondria are the major sub-cellular organelle of ATP production. With a negative inner membrane potential, mitochondria take up Ca(2+) from their surroundings, a process called mitochondrial Ca(2+) uptake. Under physiological conditions, Ca(2+) uptake into mitochondria promotes ATP production. Excessive uptake causes mitochondrial Ca(2+) overload, which activates downstream adverse responses leading to cell dysfunction. Moreover, mitochondrial Ca(2+) uptake could shape spatio-temporal patterns of intracellular Ca(2+) signaling. Malfunction of mitochondrial Ca(2+) uptake is implicated in muscle degeneration. Unlike non-excitable cells, mitochondria in muscle cells experience dramatic changes of intracellular Ca(2+) levels. Besides the sudden elevation of Ca(2+) level induced by action potentials, Ca(2+) transients in muscle cells can be as short as a few milliseconds during a single twitch or as long as minutes during tetanic contraction, which raises the question whether mitochondrial Ca(2+) uptake is fast and big enough to shape intracellular Ca(2+) signaling during excitation-contraction coupling and creates technical challenges for quantification of the dynamic changes of Ca(2+) inside mitochondria. This review focuses on characterization of mitochondrial Ca(2+) uptake in skeletal muscle and its role in muscle physiology and diseases. PMID:27430885

  1. Anoxia-induced elevation of cytosolic Ca2+ concentration depends on different Ca2+ sources in rice and wheat protoplasts.

    Science.gov (United States)

    Yemelyanov, Vladislav V; Shishova, Maria F; Chirkova, Tamara V; Lindberg, Sylvia M

    2011-08-01

    The anoxia-dependent elevation of cytosolic Ca(2+) concentration, [Ca(2+)](cyt), was investigated in plants differing in tolerance to hypoxia. The [Ca(2+)](cyt) was measured by fluorescence microscopy in single protoplasts loaded with the calcium-fluoroprobe Fura 2-AM. Imposition of anoxia led to a fast (within 3 min) significant elevation of [Ca(2+)](cyt) in rice leaf protoplasts. A tenfold drop in the external Ca(2+) concentration (to 0.1 mM) resulted in considerable decrease of the [Ca(2+)](cyt) shift. Rice root protoplasts reacted upon anoxia with higher amplitude. Addition of plasma membrane (verapamil, La(3+) and EGTA) and intracellular membrane Ca(2+)-channel antagonists (Li(+), ruthenium red and cyclosporine A) reduced the anoxic Ca(2+)-accumulation in rice. Wheat protoplasts responded to anoxia by smaller changes of [Ca(2+)](cyt). In wheat leaf protoplasts, the amplitude of the Ca(2+)-shift little depended on the external level of Ca(2+). Wheat root protoplasts were characterized by a small shift of [Ca(2+)](cyt) under anoxia. Plasmalemma Ca(2+)-channel blockers had little effect on the elevation of cytosolic Ca(2+) in wheat protoplasts. Intact rice seedlings absorbed Ca(2+) from the external medium under anoxic treatment. On the contrary, wheat seedlings were characterized by leakage of Ca(2+). Verapamil abolished the Ca(2+) influx in rice roots and Ca(2+) efflux from wheat roots. Anoxia-induced [Ca(2+)](cyt) elevation was high particularly in rice, a hypoxia-tolerant species. In conclusion, both external and internal Ca(2+) stores are important for anoxic [Ca(2+)](cyt) elevation in rice, whereas the hypoxia-intolerant wheat does not require external sources for [Ca(2+)](cyt) rise. Leaf and root protoplasts similarly responded to anoxia, independent of their organ origin.

  2. Metal ion determinants of conantokin dimerization as revealed in the X-ray crystallographic structure of the Cd(2+)/Mg (2+)-con-T[K7gamma] complex.

    Science.gov (United States)

    Cnudde, Sara E; Prorok, Mary; Castellino, Francis J; Geiger, James H

    2010-06-01

    Predatory sea snails from the Conus family produce a variety of venomous small helical peptides called conantokins that are rich in gamma-carboxyglutamic acid (Gla) residues. As potent and selective antagonists of the N-methyl-D: -aspartate receptor, these peptides are potential therapeutic agents for a variety of neurological conditions. The two most studied members of this family of peptides are con-G and con-T. Con-G has Gla residues at sequence positions 3, 4, 7, 10, and 14, and requires divalent cation binding to adopt a helical conformation. Although both Ca(2+) and Mg(2+) can fulfill this role, Ca(2+) induces dimerization of con-G, whereas the Mg(2+)-complexed peptide remains monomeric. A variant of con-T, con-T[K7gamma] (gamma is Gla), contains Gla residues at the same five positions as in con-G and behaves very similarly with respect to metal ion binding and dimerization; each peptide binds two Ca(2+) ions and two Mg(2+) ions per helix. To understand the difference in metal ion selectivity, affinity, and the dependence on Ca(2+) for dimer formation, we report here the structure of the monomeric Cd(2+)/Mg(2+)-con-T[K7gamma] complex, and, by comparison with the previously published con-T[K7gamma]/Ca(2+) dimer structure, we suggest explanations for both metal ion binding site specificity and metal-ion-dependent dimerization.

  3. Aerobic interval training partly reverse contractile dysfunction and impaired Ca2+ handling in atrial myocytes from rats with post infarction heart failure.

    Directory of Open Access Journals (Sweden)

    Anne Berit Johnsen

    Full Text Available BACKGROUND: There is limited knowledge about atrial myocyte Ca(2+ handling in the failing hearts. The aim of this study was to examine atrial myocyte contractile function and Ca(2+ handling in rats with post-infarction heart failure (HF and to examine whether aerobic interval training could reverse a potential dysfunction. METHODS AND RESULTS: Post-infarction HF was induced in Sprague Dawley rats by ligation of the left descending coronary artery. Atrial myocyte shortening was depressed (p<0.01 and time to relaxation was prolonged (p<0.01 in sedentary HF-rats compared to healthy controls. This was associated with decreased Ca(2+ amplitude, decreased SR Ca(2+ content, and slower Ca(2+ transient decay. Atrial myocytes from HF-rats had reduced sarcoplasmic reticulum Ca(2+ ATPase activity, increased Na(+/Ca(2+-exchanger activity and increased diastolic Ca(2+ leak through ryanodine receptors. High intensity aerobic interval training in HF-rats restored atrial myocyte contractile function and reversed changes in atrial Ca(2+ handling in HF. CONCLUSION: Post infarction HF in rats causes profound impairment in atrial myocyte contractile function and Ca(2+ handling. The observed dysfunction in atrial myocytes was partly reversed after aerobic interval training.

  4. Leishmania amazonensis: PKC-like protein kinase modulates the (Na++K+)ATPase activity.

    Science.gov (United States)

    Almeida-Amaral, Elmo Eduardo de; Caruso-Neves, Celso; Lara, Lucienne Silva; Pinheiro, Carla Mônica; Meyer-Fernandes, José Roberto

    2007-08-01

    The present study aimed to identify the presence of protein kinase C-like (PKC-like) in Leishmania amazonensis and to elucidate its possible role in the modulation of the (Na(+)+K(+))ATPase activity. Immunoblotting experiments using antibody against a consensus sequence (Ac 543-549) of rabbit protein kinase C (PKC) revealed the presence of a protein kinase of 80 kDa in L. amazonensis. Measurements of protein kinase activity showed the presence of both (Ca(2+)-dependent) and (Ca(2+)-independent) protein kinase activity in plasma membrane and cytosol. Phorbol ester (PMA) activation of the Ca(2+)-dependent protein kinase stimulated the (Na(+)+K(+))ATPase activity, while activation of the Ca(2+)-independent protein kinase was inhibitory. Both effects of protein kinase on the (Na(+)+K(+))ATPase of the plasma membrane were lower than that observed in intact cells. PMA induced the translocation of protein kinase from cytosol to plasma membrane, indicating that the maximal effect of protein kinase on the (Na(+)+K(+))ATPase activity depends on the synergistic action of protein kinases from both plasma membrane and cytosol. This is the first demonstration of a protein kinase activated by PMA in L. amazonensis and the first evidence for a possible role in the regulation of the (Na(+)+K(+))ATPase activity in this trypanosomatid. Modulation of the (Na(+)+K(+))ATPase by protein kinase in a trypanosomatid opens up new possibilities to understand the regulation of ion homeostasis in this parasite. PMID:17475255

  5. Palmitoylation of the Cysteine Residue in the DHHC Motif of a Palmitoyl Transferase Mediates Ca2+ Homeostasis in Aspergillus.

    Directory of Open Access Journals (Sweden)

    Yuanwei Zhang

    2016-04-01

    Full Text Available Finely tuned changes in cytosolic free calcium ([Ca2+]c mediate numerous intracellular functions resulting in the activation or inactivation of a series of target proteins. Palmitoylation is a reversible post-translational modification involved in membrane protein trafficking between membranes and in their functional modulation. However, studies on the relationship between palmitoylation and calcium signaling have been limited. Here, we demonstrate that the yeast palmitoyl transferase ScAkr1p homolog, AkrA in Aspergillus nidulans, regulates [Ca2+]c homeostasis. Deletion of akrA showed marked defects in hyphal growth and conidiation under low calcium conditions which were similar to the effects of deleting components of the high-affinity calcium uptake system (HACS. The [Ca2+]c dynamics in living cells expressing the calcium reporter aequorin in different akrA mutant backgrounds were defective in their [Ca2+]c responses to high extracellular Ca2+ stress or drugs that cause ER or plasma membrane stress. All of these effects on the [Ca2+]c responses mediated by AkrA were closely associated with the cysteine residue of the AkrA DHHC motif, which is required for palmitoylation by AkrA. Using the acyl-biotin exchange chemistry assay combined with proteomic mass spectrometry, we identified protein substrates palmitoylated by AkrA including two new putative P-type ATPases (Pmc1 and Spf1 homologs, a putative proton V-type proton ATPase (Vma5 homolog and three putative proteins in A. nidulans, the transcripts of which have previously been shown to be induced by extracellular calcium stress in a CrzA-dependent manner. Thus, our findings provide strong evidence that the AkrA protein regulates [Ca2+]c homeostasis by palmitoylating these protein candidates and give new insights the role of palmitoylation in the regulation of calcium-mediated responses to extracellular, ER or plasma membrane stress.

  6. Ca2+-Clock-Dependent Pacemaking in the Sinus Node Is Impaired in Mice with a Cardiac Specific Reduction in SERCA2 Abundance

    Science.gov (United States)

    Logantha, Sunil Jit R. J.; Stokke, Mathis K.; Atkinson, Andrew J.; Kharche, Sanjay R.; Parveen, Sajida; Saeed, Yawer; Sjaastad, Ivar; Sejersted, Ole M.; Dobrzynski, Halina

    2016-01-01

    Background: The sarcoplasmic reticulum Ca2+-ATPase (SERCA2) pump is an important component of the Ca2+-clock pacemaker mechanism that provides robustness and flexibility to sinus node pacemaking. We have developed transgenic mice with reduced cardiac SERCA2 abundance (Serca2 KO) as a model for investigating SERCA2's role in sinus node pacemaking. Methods and Results: In Serca2 KO mice, ventricular SERCA2a protein content measured by Western blotting was 75% (P 70% reduction in SERCA2 activity. Conclusions: Serca2 KO mice show a disrupted Ca2+-clock-dependent pacemaker mechanism contributing to impaired sinus node and atrioventricular node function. PMID:27313537

  7. Reduced expression of Ca2+-regulating proteins in the upper gastrointestinal tract of patients with achalasia

    Institute of Scientific and Technical Information of China (English)

    Harald Fischer; Judith Fischer; Peter Boknik; Ulrich Gergs; Wilhelm Schmitz; Wolfram Domschke; Jan W Konturek; Joachim Neumann

    2006-01-01

    AIM: To compare expression of Ca2+-regulating proteins in upper gastrointestinal (GI) tract of achalasia patients and healthy volunteers and to elucidate their role in achalasia.METHODS: Sarcoplasmic reticulum Ca2+ ATPase (SERCA)isoforms 2a and 2b, phospholamban (PLB), calsequestrin (CSQ), and calreticulin (CRT) were assessed by quantitative Western blotting in esophagus and heart of rats, rabbits, and humans. Furthermore, expression profiles of these proteins in biopsies of lower esophageal sphincter and esophagus from patients with achalasia and healthy volunteers were analyzed.RESULTS: SERCA 2a protein expression was much higher in human heart (cardiac ventricle) compared to esophagus. However, SERCA 2b was expressed predominantly in the esophagus. The highest CRT expression was noted in the human esophagus, while PLB, although highly expressed in the heart, was below our detection limit in upper GI tissue. Compared to healthy controls, CSQ and CRT expression in lower esophageal sphincter and distal esophageal body were significantly reduced in patients with achalasia (P < 0.05).CONCLUSION: PLB in the human esophagus might be of lesser importance for regulation of SERCA than in heart. Lower expression of Ca2+ storage proteins (CSQ and CRT) might contribute to increased lower esophageal sphincter pressure in achalasia, possibly by increasing free intracellular Ca2+.

  8. The role of Na(+), K(+)-ATPase in the hypoxic vasoconstriction in isolated rat basilar artery.

    Science.gov (United States)

    Shen, Haitao; Liang, Peng; Qiu, Suhua; Zhang, Bo; Wang, Yongli; Lv, Ping

    2016-06-01

    Hypoxia-induced cerebrovascular dysfunction is a key factor in the occurrence and the development of cerebral ischemia. Na(+), K(+)-ATPase affects the regulation of intracellular Ca(2+) concentration and plays an important role in vascular smooth muscle function. However, the potential role of Na(+), K(+)-ATPase in hypoxia-induced cerebrovascular dysfunction is unknown. In this study, we found that the KCl-induced contraction under hypoxia in rat endothelium-intact basilar arteries is similar to that of denuded arteries, suggesting that hypoxia may cause smooth muscle cell (SMC)-dependent vasoconstriction in the basilar artery. The Na(+), K(+)-ATPase activity of the isolated basilar artery with or without endothelium significantly reduced with prolonged hypoxia. Blocking the Na(+)-Ca(2+) exchanger with Ni(2+) (10(-3)M) or the L-type Ca(2+) channel with nimodipine (10(-8)M) dramatically attenuated KCl-induced contraction under hypoxia. Furthermore, prolonged hypoxia significantly reduced Na(+), K(+)-ATPase activity and increased [Ca(2+)]i in cultured rat basilar artery SMCs. Hypoxia reduced the protein and mRNA expression of the α2 isoform of Na(+), K(+)-ATPase in SMCs in vitro. We used a low concentration of the Na(+), K(+)-ATPase inhibitor ouabain, which possesses a high affinity for the α2 isoform. The contractile response in the rat basilar artery under hypoxia was partly inhibited by ouabain pretreatment. The decreased Na(+), K(+)-ATPase activity in isolated basilar artery and the increased [Ca(2+)]i in SMCs induced by hypoxia were partly inhibited by pretreatment with a low concentration of ouabain. These results suggest that hypoxia may educe Na(+), K(+)-ATPase activity in SMCs through the α2 isoform contributing to vasoconstriction in the rat basilar artery.

  9. Nitric oxide inhibits capacitative Ca2+ entry by suppression of mitochondrial Ca2+ handling

    Science.gov (United States)

    Thyagarajan, Baskaran; Malli, Roland; Schmidt, Kurt; Graier, Wolfgang F; Groschner, Klaus

    2002-01-01

    Nitric oxide (NO) is a key modulator of cellular Ca2+ signalling and a determinant of mitochondrial function. Here, we demonstrate that NO governs capacitative Ca2+ entry (CCE) into HEK293 cells by impairment of mitochondrial Ca2+ handling. Authentic NO as well as the NO donors 1-[2-(carboxylato)pyrrolidin-1-yl]diazem-1-ium-1,2-diolate (ProliNO) and 2-(N,N-diethylamino)-diazenolate-2-oxide (DEANO) suppressed CCE activated by thapsigargin (TG)-induced store depletion. Threshold concentrations for inhibition of CCE by ProliNO and DEANO were 0.3 and 1 μM, respectively. NO-induced inhibition of CCE was not mimicked by peroxynitrite (100 μM), the peroxynitrite donor 3-morpholino-sydnonimine (SIN-1, 100 μM) or 8-bromoguanosine 3′,5′-cyclic monophosphate (8-BrcGMP, 1 mM). In addition, the guanylyl cyclase inhibitor 1H-[1,2,4] oxadiazole[4,3-a] quinoxalin-1-one (ODQ, 30 μM) failed to antagonize the inhibitory action of NO on CCE. DEANO (1–10 μM) suppressed mitochondrial respiration as evident from inhibition of cellular oxygen consumption. Experiments using fluorescent dyes to monitor mitochondrial membrane potential and mitochondrial Ca2+ levels, respectively, indicated that DEANO (10 μM) depolarized mitochondria and suppressed mitochondrial Ca2+ sequestration. The inhibitory effect of DEANO on Ca2+ uptake into mitochondria was confirmed by recording mitochondrial Ca2+ during agonist stimulation in HEK293 cells expressing ratiometric-pericam in mitochondria. DEANO (10 μM) failed to inhibit Ba2+ entry into TG-stimulated cells when extracellular Ca2+ was buffered below 1 μM, while clear inhibition of Ba2+ entry into store depleted cells was observed when extracellular Ca2+ levels were above 10 μM. Moreover, buffering of intracellular Ca2+ by use of N,N′-[1,2-ethanediylbis(oxy-2,1-phenylene)] bis [N-[25-[(acetyloxy) methoxy]-2-oxoethyl

  10. Study on the effect of doxorubicin on expressions of genes encoding myocardial sarcoplasmic reticulum Ca2+ transport proteins and the effect of taurine on myocardial protection in rabbits

    Institute of Scientific and Technical Information of China (English)

    黄先玫; 朱卫华; 康曼丽

    2003-01-01

    To investigate the effect of doxorubicin(DOX) on gene expression of the myocardial sarcoplasmic reticulum (SR)Ca2+ transport proteins and the mechanism of taurine(Tau) protecting cardiac muscle cells, 9 rabbits were injected with DOX , 8 rabbits with DOX and Tau, and 9 rabbits with normal saline. Cardiac function , concentration of calcium in cardiomyocytes (Myo[Ca2+]i), activity of SR Ca2+-ATPase(SERCA2a), level of SERCA2a mRNA and Ca2+ released channels(RYR2)mRNA were detected. The left ventricle tissues were observed by electron microscopy. The results showed that cardiac index, left ventricular systolic pressure, activity of SR Ca2+-ATPase and level of SERCA2a mRNA decreased , while Myo[Ca2+]i increased in DOX-treated rabbits. DOX could not affect the level of RYR2 mRNA. Tau intervention could alleviate the increase of left ventricular diastolic pressure, Myo[Ca2+]i and the decrease of SERCA2a mRNA induced by doxorubicin. The results suggested that downregulation of SERCA2a gene expression was an important mechanism of DOX-induced cardiomyopathy and that Tau could partially improve the heart function by reducing calcium overload and alleviating downregulation of SERCA2a mRNA.

  11. Hydrogen Storage Properties of Ca3-x Mg2+xNi13 Alloys%Ca3-xMg2+xNi13合金的储氢性能

    Institute of Scientific and Technical Information of China (English)

    张庆安; 赵刚; 斯庭智; 庞刚

    2009-01-01

    为了弄清Mg含量对Ca3Mg2Ni13型化合物结构参数和储氢性能的影响,利用X射线衍射研究了Ca3-xMg2+x,Ni13(x=0.5,1.0和1.5)合金的相结构,并采用Sieverts型设备测量了其P-C-T曲线.研究表明,Mg在Ca3Mg2Ni13型化合物中的最大固溶度接近于Ca1.5MgNi13合金中的Mg含量.固溶的Mg含量增加导致化合物点阵常数减小,这可以有效地改善吸放氢热力学性能,其中Ca2Mg3Ni13吸、放氢的焓变分别为-28,30 kJ/mol H2.此外,Ca2Mg3Ni13在吸放氢循环过程中不发生氢致非晶化和氢致分解,因而具有良好的循环稳定性.%To understand the effects of Mg content on the structural parameters and hydrogen storage properties of Ca3Mg2Ni13-type compound, the phase structures of the Ca3-xMg2+xNi13 (x =0.5, 1.0 and 1.5 ) alloys were investigated by X-ray diffraction (XRD) and their pressure-composition isotherms (P-C-T curves) were measured with a Sieverts-type apparatus. The results indicate that the maximum solid solubility of Mg in the Ca3Mg2Ni13-type compound is close to the Mg content of Ca1.5 Mg3.5 Ni13 alloy. The increase of Mg content leads to the decrease in the lattice parameters of Ca3 Mg2Ni13-type compound, which may effectively improve the thermodynamics of hydrogen absorption-desorption. The enthalpy changes for the hydrogen absorption and desorption of Ca2Mg3Ni13 are -28 and 30 kJ/mol H2, respectively. Moreover, Ca2Mg3Ni13 shows good cycling stability because the hydrogen-induced amorphization and decomposition do not occur during hydrogen absorption-desorption cycles.

  12. Glutamate receptor activation in cultured cerebellar granule cells increases cytosolic free Ca2+ by mobilization of cellular Ca2+ and activation of Ca2+ influx

    DEFF Research Database (Denmark)

    Bouchelouche, P; Belhage, B; Frandsen, A;

    1989-01-01

    The Ca2+ sensitive fluorescent probe, fura-2 has been used to monitor cytosolic free calcium levels in mature primary cultures of cerebellar granule cells during exposure to L-glutamate and other excitatory amino acids: quisqualate (QA) kainate (KA) and N-methyl-D-aspartate (NMDA). Glutamate...... at micromolar concentrations produced a prompt and dose-related increase in the intracellular concentration of free Ca2+, ([Ca2+]i), whereas QA, KA and NMDA had no effect. This increase was also seen in the absence of extracellular Ca2+, suggesting that L-glutamate promotes mobilization of Ca2+ from...

  13. Signaling between intracellular Ca2+ stores and depletion-activated Ca2+ channels generates [Ca2+]i oscillations in T lymphocytes

    OpenAIRE

    1994-01-01

    Stimulation through the antigen receptor (TCR) of T lymphocytes triggers cytosolic calcium ([Ca2+]i) oscillations that are critically dependent on Ca2+ entry across the plasma membrane. We have investigated the roles of Ca2+ influx and depletion of intracellular Ca2+ stores in the oscillation mechanism, using single-cell Ca2+ imaging techniques and agents that deplete the stores. Thapsigargin (TG; 5-25 nM), cyclopiazonic acid (CPA; 5-20 microM), and tert- butylhydroquinone (tBHQ; 80-200 micro...

  14. Ca2+-Regulated Photoproteins: Effective Immunoassay Reporters

    Directory of Open Access Journals (Sweden)

    Ludmila A. Frank

    2010-12-01

    Full Text Available Ca2+-regulated photoproteins of luminous marine coelenterates are of interest and a challenge for researchers as a unique bioluminescent system and as a promising analytical instrument for both in vivo and in vitro applications. The proteins are comprehensively studied as to biochemical properties, tertiary structures, bioluminescence mechanism, etc. This knowledge, along with available recombinant proteins serves the basis for development of unique bioluminescent detection systems that are “self-contained”, triggerable, fast, highly sensitive, and non-hazardous. In the paper, we focus on the use of photoproteins as reporters in binding assays based on immunological recognition element—bioluminescent immunoassay and hybridization immunoassay, their advantages and prospects.

  15. Clustered Conserved Cysteines in Hyaluronan Synthase Mediate Cooperative Activation by Mg(2+) Ions and Severe Inhibitory Effects of Divalent Cations.

    Science.gov (United States)

    Tlapak-Simmons, Valarie L; Medina, Andria P; Baggenstoss, Bruce A; Nguyen, Long; Baron, Christina A; Weigel, Paul H

    2011-11-15

    Hyaluronan synthase (HAS) uses UDP-GlcUA and UDP-GlcNAc to make hyaluronan (HA). Streptococcus equisimilis HAS (SeHAS) contains four conserved cysteines clustered near the membrane, and requires phospholipids and Mg(2+) for activity. Activity of membrane-bound or purified enzyme displayed a sigmoidal saturation profile for Mg(2+) with a Hill coefficient of 2. To assess if Cys residues are important for cooperativity we examined the Mg(2+) dependence of mutants with various combinations of Cys-to-Ala mutations. All Cys-mutants lost the cooperative response to Mg(2+). In the presence of Mg(2+), other divalent cations inhibited SeHAS with different potencies (Cu(2+)~Zn(2+) >Co(2+) >Ni(2+) >Mn(2+) >Ba(2+) Sr(2+) Ca(2+)). Some divalent metal ions likely inhibit by displacement of Mg(2+)-UDP-Sugar complexes (e.g. Ca(2+), Sr(2+) and Ba(2+) had apparent Ki values of 2-5 mM). In contrast, Zn(2+) and Cu(2+) inhibited more potently (apparent Ki ≤ 0.2 mM). Inhibition of Cys-null SeHAS by Cu(2+), but not Zn(2+), was greatly attenuated compared to wildtype. Double and triple Cys-mutants showed differing sensitivities to Zn(2+) or Cu(2+). Wildtype SeHAS allowed to make HA prior to exposure to Zn(2+) or Cu(2+) was protected from inhibition, indicating that access of metal ions to sensitive functional groups was hindered in processively acting HA•HAS complexes. We conclude that clustered Cys residues mediate cooperative interactions with Mg(2+) and that transition metal ions inhibit SeHAS very potently by interacting with one or more of these -SH groups.

  16. Ca2+ signals regulate mitochondrial metabolism by stimulating CREB-mediated expression of the mitochondrial Ca2+ uniporter gene MCU.

    Science.gov (United States)

    Shanmughapriya, Santhanam; Rajan, Sudarsan; Hoffman, Nicholas E; Zhang, Xueqian; Guo, Shuchi; Kolesar, Jill E; Hines, Kevin J; Ragheb, Jonathan; Jog, Neelakshi R; Caricchio, Roberto; Baba, Yoshihiro; Zhou, Yandong; Kaufman, Brett A; Cheung, Joseph Y; Kurosaki, Tomohiro; Gill, Donald L; Madesh, Muniswamy

    2015-03-03

    Cytosolic Ca2+ signals, generated through the coordinated translocation of Ca2+ across the plasma membrane (PM) and endoplasmic reticulum (ER) membrane, mediate diverse cellular responses. Mitochondrial Ca2+ is important for mitochondrial function, and when cytosolic Ca2+ concentration becomes too high, mitochondria function as cellular Ca2+ sinks. By measuring mitochondrial Ca2+ currents, we found that mitochondrial Ca2+ uptake was reduced in chicken DT40 B lymphocytes lacking either the ER-localized inositol trisphosphate receptor (IP3R), which releases Ca2+ from the ER, or Orai1 or STIM1, components of the PM-localized Ca2+ -permeable channel complex that mediates store-operated calcium entry (SOCE) in response to depletion of ER Ca2+ stores. The abundance of MCU, the pore-forming subunit of the mitochondrial Ca2+ uniporter, was reduced in cells deficient in IP3R, STIM1, or Orai1. Chromatin immunoprecipitation and promoter reporter analyses revealed that the Ca2+ -regulated transcription factor CREB (cyclic adenosine monophosphate response element-binding protein) directly bound the MCU promoter and stimulated expression. Lymphocytes deficient in IP3R, STIM1, or Orai1 exhibited altered mitochondrial metabolism, indicating that Ca2+ released from the ER and SOCE-mediated signals modulates mitochondrial function. Thus, our results showed that a transcriptional regulatory circuit involving Ca2+ -dependent activation of CREB controls the Ca2+ uptake capability of mitochondria and hence regulates mitochondrial metabolism.

  17. Ca2+ dialogue between acidic vesicles and ER.

    Science.gov (United States)

    Morgan, Anthony J

    2016-04-15

    Extracellular stimuli evoke the synthesis of intracellular second messengers, several of which couple to the release of Ca(2+)from Ca(2+)-storing organelles via activation of cognate organellar Ca(2+)-channel complexes. The archetype is the inositol 1,4,5-trisphosphate (IP3) and IP3receptor (IP3R) on the endoplasmic reticulum (ER). A less understood, parallel Ca(2+)signalling cascade is that involving the messenger nicotinic acid adenine dinucleotide phosphate (NAADP) that couples to Ca(2+)release from acidic Ca(2+)stores [e.g. endo-lysosomes, secretory vesicles, lysosome-related organelles (LROs)]. NAADP-induced Ca(2+)release absolutely requires organellar TPCs (two-pore channels). This review discusses how ER and acidic Ca(2+)stores physically and functionally interact to generate and shape global and local Ca(2+)signals, with particular emphasis on the two-way dialogue between these two organelles.

  18. DA-6034 Induces [Ca(2+)]i Increase in Epithelial Cells.

    Science.gov (United States)

    Yang, Yu-Mi; Park, Soonhong; Ji, Hyewon; Kim, Tae-Im; Kim, Eung Kweon; Kang, Kyung Koo; Shin, Dong Min

    2014-04-01

    DA-6034, a eupatilin derivative of flavonoid, has shown potent effects on the protection of gastric mucosa and induced the increases in fluid and glycoprotein secretion in human and rat corneal and conjunctival cells, suggesting that it might be considered as a drug for the treatment of dry eye. However, whether DA-6034 induces Ca(2+) signaling and its underlying mechanism in epithelial cells are not known. In the present study, we investigated the mechanism for actions of DA-6034 in Ca(2+) signaling pathways of the epithelial cells (conjunctival and corneal cells) from human donor eyes and mouse salivary gland epithelial cells. DA-6034 activated Ca(2+)-activated Cl(-) channels (CaCCs) and increased intracellular calcium concentrations ([Ca(2+)]i) in primary cultured human conjunctival cells. DA-6034 also increased [Ca(2+)]i in mouse salivary gland cells and human corneal epithelial cells. [Ca(2+)]i increase of DA-6034 was dependent on the Ca(2+) entry from extracellular and Ca(2+) release from internal Ca(2+) stores. Interestingly, these effects of DA-6034 were related to ryanodine receptors (RyRs) but not phospholipase C/inositol 1,4,5-triphosphate (IP3) pathway and lysosomal Ca(2+) stores. These results suggest that DA-6034 induces Ca(2+) signaling via extracellular Ca(2+) entry and RyRs-sensitive Ca(2+) release from internal Ca(2+) stores in epithelial cells.

  19. Dual Effect of Phosphate Transport on Mitochondrial Ca2+ Dynamics.

    Science.gov (United States)

    Wei, An-Chi; Liu, Ting; O'Rourke, Brian

    2015-06-26

    The large inner membrane electrochemical driving force and restricted volume of the matrix confer unique constraints on mitochondrial ion transport. Cation uptake along with anion and water movement induces swelling if not compensated by other processes. For mitochondrial Ca(2+) uptake, these include activation of countertransporters (Na(+)/Ca(2+) exchanger and Na(+)/H(+) exchanger) coupled to the proton gradient, ultimately maintained by the proton pumps of the respiratory chain, and Ca(2+) binding to matrix buffers. Inorganic phosphate (Pi) is known to affect both the Ca(2+) uptake rate and the buffering reaction, but the role of anion transport in determining mitochondrial Ca(2+) dynamics is poorly understood. Here we simultaneously monitor extra- and intra-mitochondrial Ca(2+) and mitochondrial membrane potential (ΔΨm) to examine the effects of anion transport on mitochondrial Ca(2+) flux and buffering in Pi-depleted guinea pig cardiac mitochondria. Mitochondrial Ca(2+) uptake proceeded slowly in the absence of Pi but matrix free Ca(2+) ([Ca(2+)]mito) still rose to ~50 μm. Pi (0.001-1 mm) accelerated Ca(2+) uptake but decreased [Ca(2+)]mito by almost 50% while restoring ΔΨm. Pi-dependent effects on Ca(2+) were blocked by inhibiting the phosphate carrier. Mitochondrial Ca(2+) uptake rate was also increased by vanadate (Vi), acetate, ATP, or a non-hydrolyzable ATP analog (AMP-PNP), with differential effects on matrix Ca(2+) buffering and ΔΨm recovery. Interestingly, ATP or AMP-PNP prevented the effects of Pi on Ca(2+) uptake. The results show that anion transport imposes an upper limit on mitochondrial Ca(2+) uptake and modifies the [Ca(2+)]mito response in a complex manner. PMID:25963147

  20. Identification and characterization of a novel mammalian Mg2+ transporter with channel-like properties

    Directory of Open Access Journals (Sweden)

    Quamme Gary A

    2005-04-01

    Full Text Available Abstract Background Intracellular magnesium is abundant, highly regulated and plays an important role in biochemical functions. Despite the extensive evidence for unique mammalian Mg2+ transporters, few proteins have been biochemically identified to date that fulfill this role. We have shown that epithelial magnesium conservation is controlled, in part, by differential gene expression leading to regulation of Mg2+ transport. We used this knowledge to identify a novel gene that is regulated by magnesium. Results Oligonucleotide microarray analysis was used to identify a novel human gene that encodes a protein involved with Mg2+-evoked transport. We have designated this magnesium transporter (MagT1 protein. MagT1 is a novel protein with no amino acid sequence identity to other known transporters. The corresponding cDNA comprises an open reading frame of 1005 base pairs encoding a protein of 335 amino acids. It possesses five putative transmembrane (TM regions with a cleavage site, a N-glycosylation site, and a number of phosphorylation sites. Based on Northern analysis of mouse tissues, a 2.4 kilobase transcript is present in many tissues. When expressed in Xenopus laevis oocytes, MagT1 mediates saturable Mg2+ uptake with a Michaelis constant of 0.23 mM. Transport of Mg2+ by MagT1 is rheogenic, voltage-dependent, does not display any time-dependent inactivation. Transport is very specific to Mg2+ as other divalent cations did not evoke currents. Large external concentrations of some cations inhibited Mg2+ transport (Ni2+, Zn2+, Mn2+ in MagT1-expressing oocytes. Ca2+and Fe2+ were without effect. Real-time reverse transcription polymerase chain reaction and Western blot analysis using a specific antibody demonstrated that MagT1 mRNA and protein is increased by about 2.1-fold and 32%, respectively, in kidney epithelial cells cultured in low magnesium media relative to normal media and in kidney cortex of mice maintained on low magnesium diets

  1. Spectral components of cytosolic [Ca2+] spiking in neurons

    DEFF Research Database (Denmark)

    Kardos, J; Szilágyi, N; Juhász, G;

    1998-01-01

    We show here, by means of evolutionary spectral analysis and synthesis of cytosolic Ca2+ ([Ca2+]c) spiking observed at the single cell level using digital imaging fluorescence microscopy of fura-2-loaded mouse cerebellar granule cells in culture, that [Ca2+]c spiking can be resolved into evolutio......We show here, by means of evolutionary spectral analysis and synthesis of cytosolic Ca2+ ([Ca2+]c) spiking observed at the single cell level using digital imaging fluorescence microscopy of fura-2-loaded mouse cerebellar granule cells in culture, that [Ca2+]c spiking can be resolved into...

  2. Endo-lysosomal TRP mucolipin-1 channels trigger global ER Ca2+ release and Ca2+ influx

    Science.gov (United States)

    Kilpatrick, Bethan S.; Yates, Elizabeth; Grimm, Christian; Schapira, Anthony H.

    2016-01-01

    ABSTRACT Transient receptor potential (TRP) mucolipins (TRPMLs), encoded by the MCOLN genes, are patho-physiologically relevant endo-lysosomal ion channels crucial for membrane trafficking. Several lines of evidence suggest that TRPMLs mediate localised Ca2+ release but their role in Ca2+ signalling is not clear. Here, we show that activation of endogenous and recombinant TRPMLs with synthetic agonists evoked global Ca2+ signals in human cells. These signals were blocked by a dominant-negative TRPML1 construct and a TRPML antagonist. We further show that, despite a predominant lysosomal localisation, TRPML1 supports both Ca2+ release and Ca2+ entry. Ca2+ release required lysosomal and ER Ca2+ stores suggesting that TRPMLs, like other endo-lysosomal Ca2+ channels, are capable of ‘chatter’ with ER Ca2+ channels. Our data identify new modalities for TRPML1 action. PMID:27577094

  3. Photolysis of caged compounds: studying Ca(2+) signaling and activation of Ca(2+)-dependent ion channels.

    Science.gov (United States)

    Almassy, Janos; Yule, David I

    2013-01-01

    A wide variety of signaling molecules have been chemically modified by conjugation to a photolabile chromophore to render the substance temporarily biologically inert. Subsequent exposure to ultraviolet (UV) light can release the active moiety from the "caged" precursor in an experimentally controlled manner. This allows the concentration of active molecule to be precisely manipulated in both time and space. These techniques are particularly useful in experimental protocols designed to investigate the mechanisms underlying Ca(2+) signaling and the activation of Ca(2+)-dependent effectors.

  4. Arabidopsis transcriptional response to extracellular Ca2þ depletion involves a transient rise in cytosolic Ca2þ

    Institute of Scientific and Technical Information of China (English)

    Zhi Qi

    2015-01-01

    Ecological evidence indicates a worldwide trend of dramatical y decreased soil Ca2þ levels caused by increased acid deposition and massive timber harvesting. Little is known about the genetic and cel ular mechanism of plants’ responses to Ca2þ depletion. In this study, transcriptional profiling analysis helped identify multiple extracel ular Ca2þ ([Ca2þ]ext) depletion‐respon-sive genes in Arabidopsis thaliana L., many of which are involved in response to other environmental stresses. Interestingly, a group of genes encoding putative cytosolic Ca2þ ([Ca2þ]cyt) sensors were significantly upregulated, implying that [Ca2þ]cyt has a role in sensing [Ca2þ]ext depletion. Consistent with this observation, [Ca2þ]ext depletion stimulated a transient rise in [Ca2þ]cyt that was negatively influenced by [Kþ]ext, suggesting the involvement of a membrane potential‐sensitive component. The [Ca2þ]cyt response to [Ca2þ]ext depletion was significantly desensitized after the initial treatment, which is typical of a receptor‐mediated signaling event. The response was insensi-tive to an animal Ca2þ sensor antagonist, but was suppressed by neomycin, an inhibitor of phospholipase C. Gd3þ, an inhibitor of Ca2þ channels, suppressed the [Ca2þ]ext‐triggered rise in [Ca2þ]cyt and downstream changes in gene expression. Taken together, this study demonstrates that [Ca2þ]cyt plays an important role in the putative receptor‐mediated cel ular and transcriptional response to [Ca2þ]ext depletion of plant cel s.

  5. Interference with Ca2+ release activated Ca2+ (CRAC) channel function delays T-cell arrest in vivo

    OpenAIRE

    Waite, Janelle C.; Vardhana, Santosh; Shaw, Patrick J.; Jang, Jung-Eun; McCarl, Christie-Ann; Cameron, Thomas O; Feske, Stefan; Dustin, Michael L

    2013-01-01

    Entry of lymphocytes into secondary lymphoid organs (SLOs) involves intravascular arrest and intracellular calcium ion ([Ca2+]i) elevation. TCR activation triggers increased [Ca2+]i and can arrest T-cell motility in vitro. However the requirement for [Ca2+]i elevation in arresting T cells in vivo has not been tested. Here, we have manipulated the Ca2+ release-activated Ca2+ (CRAC) channel pathway required for [Ca2+]i elevation in T cells through genetic deletion of stromal interaction molecul...

  6. Review: P4-ATPases as Phospholipid Flippases-Structure, Function, and Enigmas

    DEFF Research Database (Denmark)

    Andersen, Jens P; Vestergaard, Anna L; Mikkelsen, Stine A;

    2016-01-01

    coagulation, apoptosis, bile and cholesterol homeostasis, and neuronal cell survival. Some P4-ATPases transport phosphatidylserine and phosphatidylethanolamine across the plasma membrane or intracellular membranes whereas other P4-ATPases are specific for phosphatidylcholine. The importance of P4-ATPases...... of a peripheral hydrophobic gate pathway between transmembrane helices M1, M3, M4, and M6. This pathway, which partially overlaps with the suggested pathway for migration of Ca(2+) in the opposite direction in the Ca(2+)-ATPase, is wider than the latter, thereby accommodating the phospholipid head group. The head...... similar to the mechanism of these ion pumps, where the glutamate translocates the ions by moving like a pump rod. The accessory subunit CDC50 may be located in close association with the exoplasmic entrance of the suggested pathway, and possibly promotes the binding of the lipid substrate. This review...

  7. Removal of Mg2+, K+, SO4-2 Ions from Seawater by Precipitation Method

    Directory of Open Access Journals (Sweden)

    Pujiastuti Caecilia

    2016-01-01

    Full Text Available Removal of Mg2+, K+, and SO4-2 ions in seawater has been successfully done by precipitation in a mixing tank method. This study aims to remove the content of magnesium ions (Mg+, potassium (K+ and sulfate (SO4-2 in sea water with the addition of chemicals disodium phosphate (1.2 % volume, calcium chloride (2 % volume and sodium hydroxide (2% volume. Stirring is performed at 100 rpm and the pH solution is adjusted to 9. Disodium phosphate serves to bind magnesium ions and potassium, CaCl2 serves to bind the sulfate ion, while sodium hydroxide is used to adjust the pH of the solution mixture and also reacted with magnesium ions. In total, the removal efficiencies of Mg2+, K+ and SO4-2 ions in seawater were 97%, 96%, and 92%, respectively. The precipitated solids contains component of PO4- (14.5%, Mg2+(13.8%, SO4-2 (28.2%, Ca2+ (24.1% and K+ (1.9% ions.

  8. The contractile vacuole in Ca2+-regulation in Dictyostelium: its essential function for cAMP-induced Ca2+-influx

    Directory of Open Access Journals (Sweden)

    Schlatterer Christina

    2006-06-01

    Full Text Available Abstract Background cAMP-induced Ca2+-influx in Dictyostelium is controlled by at least two non-mitochondrial Ca2+-stores: acidic stores and the endoplasmic reticulum (ER. The acidic stores may comprise the contractile vacuole network (CV, the endosomal compartment and acidocalcisomes. Here the role of CV in respect to function as a potential Ca2+-store was investigated. Results Dajumin-GFP labeled contractile vacuoles were purified 7-fold by anti-GFP-antibodies in a magnetic field. The purified CV were shown for the first time to accumulate and release Ca2+. Release of Ca2+ was elicited by arachidonic acid or the calmodulin antagonist W7, the latter due to inhibition of the pump. The characteristics of Ca2+-transport and Ca2+-release of CV were compared to similarly purified vesicles of the ER labeled by calnexin-GFP. Since the CV proved to be a highly efficient Ca2+-compartment we wanted to know whether or not it takes part in cAMP-induced Ca2+-influx. We made use of the LvsA--mutant expected to display reduced Ca2+-transport due to loss of calmodulin. We found a severe reduction of cAMP-induced Ca2+-influx into whole cells. Conclusion The contractile vacuoles in Dictyostelium represent a highly efficient acidic Ca2+-store that is required for cAMP-induced Ca2+-influx.

  9. TRPV5, the gateway to Ca2+ homeostasis.

    NARCIS (Netherlands)

    Mensenkamp, A.R.; Hoenderop, J.G.J.; Bindels, R.J.M.

    2007-01-01

    Ca2+ homeostasis in the body is tightly controlled, and is a balance between absorption in the intestine, excretion via the urine, and exchange from bone. Recently, the epithelial Ca2+ channel (TRPV5) has been identified as the gene responsible for the Ca2+ influx in epithelial cells of the renal di

  10. Optical properties and electronic band structure of BiMg2PO6, BiMg2VO6, BiMg2VO6:Pr3+ and BiMg2VO6:Eu3+

    Science.gov (United States)

    Barros, A.; Deloncle, R.; Deschamp, J.; Boutinaud, P.; Chadeyron, G.; Mahiou, R.; Cavalli, E.; Brik, M. G.

    2014-08-01

    The luminescence properties of the yellow pigment BiMg2VO6 are revisited and those of BiMg2PO6, BiMg2VO6:Pr3+ and BiMg2VO6:Eu3+ are described. It is shown that the undoped systems exhibit broad band emission in the green or orange spectral regions, but only upon UV or near UV excitation. In contradiction with a previous report, we found that the blue, host absorbed, photons are lost non-radiatively and do not contribute to the luminescence processes in BiMg2VO6. To understand these experimental results, the optical properties of BiMg2VO6 and BiMg2PO6 are theoretically analysed on the basis of electronic structure diagrams calculated by the DFT method. It is found that the optical transitions are mostly localised within [VO4]3- units or non-regular Bi3+ ions and occur in the UV or near UV regions. The luminescence of the trivalent lanthanide dopants is weak (Eu3+) or unobserved (Pr3+) in BiMg2VO6 which is explained by inefficient energy migration in the host lattice to the impurity sites.

  11. Acceleration of Ca(2+) repletion in the junctional sarcoplasmic reticulum and alternation of the Ca(2+)-induced Ca(2+)-release mechanism in hypertensive rat (SHR) cardiac muscle.

    Science.gov (United States)

    Tanaka, Midori; Tameyasu, Tsukasa

    2008-04-01

    We estimated the time taken for a repletion of the junctional sarcoplasmic reticulum (JSR) Ca(2+) stores from a family of mechanical restitution curves after twitches of various magnitudes in the cardiac muscle of hypertensive rats (SHR), using a method described previously (Tameyasu et al. Jpn J Physiol. 2004;54:209-19), to evaluate abnormality in Ca(2+) handling by cardiac JSR in hypertension. We found no differences in contractility or in the time course of mechanical restitution between SHR and the controls (WKY) at 3 weeks of age. In comparison to WKY, 7- and 20-week-old SHR showed a greater rested state contraction (RST) and similar or smaller rapid cooling contracture, suggesting that their JSR contains a similar amount of Ca(2+) at saturation, but releases more Ca(2+) upon stimulation. The adult SHR and WKY showed similar mechanical restitution time courses, but the adults had longer pretwitch latencies. The function G(t) representing the time course of JSR Ca(2+) store repletion in adult SHR exceeded the WKY value at t JSR [Ca(2+)] change corresponding to the mechanical restitution after RST was smaller in the adult SHR at t JSR Ca(2+) store repletion and an alternation of the Ca(2+)-induced release of Ca(2+ )from the JSR in young adult SHR. PMID:18312741

  12. Cloning, expression, and functional characterization of a Ca(2+)-dependent endoplasmic reticulum nucleoside diphosphatase.

    Science.gov (United States)

    Failer, Bernd U; Braun, Norbert; Zimmermann, Herbert

    2002-10-01

    We have isolated and characterized the cDNA encoding a Ca(2+)-dependent nucleoside diphosphatase (EC ) related to two secreted ATP- and ADP-hydrolyzing apyrases of the bloodsucking insects, Cimex lectularius and Phlebotomus papatasi. The rat brain-derived cDNA has an open reading frame of 1209 bp encoding a protein of 403 amino acids and a calculated molecular mass of 45.7 kDa. The mRNA was expressed in all tissues investigated, revealing two major transcripts with varying preponderance. The immunohistochemical analysis of the Myc-His-tagged enzyme expressed in Chinese hamster ovary cells revealed its association with the endoplasmic reticulum and also with pre-Golgi intermediates. Ca(2+)-dependent nucleoside diphosphatase is a membrane protein with its catalytic site facing the organelle lumen. It hydrolyzes nucleoside 5'-diphosphates in the order UDP >GDP = IDP >CDP but not ADP. Nucleoside 5'-triphosphates were hydrolyzed to a minor extent, and no hydrolysis of nucleoside 5'-monophosphates was observed. The enzyme was strongly activated by Ca(2+), insensitive to Mg(2+), and had a K(m) for UDP of 216 microm. Ca(2+)-dependent nucleoside diphosphatase may support glycosylation reactions related to quality control in the endoplasmic reticulum. PMID:12167635

  13. Upregulation of the SERCA-type Ca2+ pump activity in response to endoplasmic reticulum stress in PC12 cells

    Directory of Open Access Journals (Sweden)

    Frandsen Aase

    2001-04-01

    Full Text Available Abstract Background Ca2+-ATPases of endoplasmic reticulum (SERCAs are responsible for maintenance of the micro- to millimolar Ca2+ ion concentrations within the endoplasmic reticulum (ER of eukaryotic cells. This intralumenal Ca2+ storage is important for the generation of Ca2+ signals as well as for the correct folding and posttranslational processing of proteins entering ER after synthesis. ER perturbations such as depletion of Ca2+ or abolishing the oxidative potential, inhibition of glycosylation, or block of secretory pathway, activate the Unfolded Protein Response, consisting of an upregulation of a number of ER-resident chaperones/stress proteins in an effort to boost the impaired folding capacity. Results We show here that in PC12 cells, depletion of ER Ca2+ by EGTA, as well as inhibition of disulphide bridge formation within the ER by dithiotreitol or inhibition of N-glycosylation by tunicamycin, led to a 2- to 3-fold increase of the SERCA-mediated 45Ca2+ transport to microsomes isolated from cells exposed to these stress agents. The time course of this response corresponded to that for transcriptional upregulation of ER stress proteins, as well as to the increase in the SERCA2b mRNA, as we recently observed in an independent study. Conclusions These findings provide the first functional evidence for the increase of SERCA pumping capacity in cells subjected to the ER stress. Since at least three different and unrelated mechanisms of eliciting the ER stress response were found to cause this functional upregulation of Ca2+ transport into the ER, these results support the existence of a coupling between the induction of the UPR pathway in general, and the regulation of expression of at least one of the SERCA pump isoforms.

  14. Deletion of PdMit1, a homolog of yeast Csg1, affects growth and Ca(2+) sensitivity of the fungus Penicillium digitatum, but does not alter virulence.

    Science.gov (United States)

    Zhu, Congyi; Wang, Weili; Wang, Mingshuang; Ruan, Ruoxin; Sun, Xuepeng; He, Meixian; Mao, Cungui; Li, Hongye

    2015-04-01

    GDP-mannose:inositol-phosphorylceramide (MIPC) and its derivatives are important for Ca(2+) sensitization of Saccharomyces cerevisiae and for the virulence of Candida albicans, but its role in the virulence of plant fungal pathogens remains unclear. In this study, we report the identification and functional characterization of PdMit1, the gene encoding MIPC synthase in Penicillium digitatum, one of the most important pathogens of postharvest citrus fruits. To understand the function of PdMit1, a PdMit1 deletion mutant was generated. Compared to its wild-type control, the PdMit1 deletion mutant exhibited slow radial growth, decreased conidia production and delayed conidial germination, suggesting that PdMit1 is important for the growth of mycelium, sporulation and conidial germination. The PdMit1 deletion mutant also showed hypersensitivity to Ca(2+). Treatment with 250 mmol/l Ca(2+) induced vacuole fusion in the wild-type strain, but not in the PdMit1 deletion mutant. Treatment with 250mmol/lCaCl2 upregulated three Ca(2+)-ATPase genes in the wild-type strain, and this was significantly inhibited in the PdMit1 deletion mutant. These results suggest that PdMit1 may have a role in regulating vacuole fusion and expression of Ca(2+)-ATPase genes by controlling biosynthesis of MIPC, and thereby imparts P. digitatum Ca(2+) tolerance. However, we found that PdMit1 is dispensable for virulence of P. digitatum.

  15. P-type ATPases.

    Science.gov (United States)

    Palmgren, Michael G; Nissen, Poul

    2011-01-01

    P-type ATPases form a large superfamily of cation and lipid pumps. They are remarkably simple with only a single catalytic subunit and carry out large domain motions during transport. The atomic structure of P-type ATPases in different conformations, together with ample mutagenesis evidence, has provided detailed insights into the pumping mechanism by these biological nanomachines. Phylogenetically, P-type ATPases are divided into five subfamilies, P1-P5. These subfamilies differ with respect to transported ligands and the way they are regulated. PMID:21351879

  16. Identification of store-independent and store-operated Ca2+ conductances in Caenorhabditis elegans intestinal epithelial cells.

    Science.gov (United States)

    Estevez, Ana Y; Roberts, Randolph K; Strange, Kevin

    2003-08-01

    The nematode Caenorhabditis elegans offers significant experimental advantages for defining the genetic basis of diverse biological processes. Genetic and physiological analyses have demonstrated that inositol-1,4,5-trisphosphate (IP3)-dependent Ca2+ oscillations in intestinal epithelial cells play a central role in regulating the nematode defecation cycle, an ultradian rhythm with a periodicity of 45-50 s. Patch clamp studies combined with behavioral assays and forward and reverse genetic screening would provide a powerful approach for defining the molecular details of oscillatory Ca2+ signaling. However, electrophysiological characterization of the intestinal epithelium has not been possible because of its relative inaccessibility. We developed primary intestinal epithelial cell cultures that circumvent this problem. Intestinal cells express two highly Ca2+-selective, voltage-independent conductances. One conductance, IORCa, is constitutively active, exhibits strong outward rectification, is 60-70-fold more selective for Ca2+ than Na+, is inhibited by intracellular Mg2+ with a K1/2 of 692 microM, and is insensitive to Ca2+ store depletion. Inhibition of IORCa with high intracellular Mg2+ concentrations revealed the presence of a small amplitude conductance that was activated by passive depletion of intracellular Ca2+ stores. Active depletion of Ca2+ stores with IP3 or ionomycin increased the rate of current activation approximately 8- and approximately 22-fold compared with passive store depletion. The store-operated conductance, ISOC, exhibits strong inward rectification, and the channel is highly selective for Ca2+ over monovalent cations with a divalent cation selectivity sequence of Ca2+ > Ba2+ approximately Sr2+. Reversal potentials for ISOC could not be detected accurately between 0 and +80 mV, suggesting that PCa/PNa of the channel may exceed 1,000:1. Lanthanum, SKF 96365, and 2-APB inhibit both IORCa and ISOC reversibly. Our studies provide the first

  17. General requirement for harvesting antennae at Ca2+ and H+ channels and transporters

    Directory of Open Access Journals (Sweden)

    Cristián Martínez

    2010-09-01

    Full Text Available The production and dissipation of energy in cells is intimately linked to the movement of small molecules in and out of enzymes, channels and transporters. An analytical model of diffusion was described previously, which was used to estimate local effects of these proteins acting as molecular sources. The present article describes a simple but more general model, which can be used to estimate the local impact of proteins acting as molecular sinks. The results show that the enzymes, transporters and channels, whose substrates are present at relatively high concentrations like ATP, Na+, glucose, lactate and pyruvate, do not operate fast enough to deplete their vicinity to a meaningful extent, supporting the notion that for these molecules the cytosol is a well-mixed compartment. One specific consequence of this analysis is that the well-documented cross-talk existing between the Na+/K+ ATPase and the glycolytic machinery should not be explained by putative changes in local ATP concentration. In contrast, Ca2+ and H+ transporters like the Na+/Ca2+ exchanger NCX and the Na+/H+ exchanger NHE, show experimental rates of transport that are two to three orders of magnitude faster than the rates at which the aqueous phase may possibly feed their binding sites. This paradoxical result implies that Ca2+ and H+ transporters do not extract their substrates directly from the bulk cytosol, but from an intermediate “harvesting” compartment located between the aqueous phase and the transport site.

  18. Ca(2+) homeostasis in the budding yeast Saccharomyces cerevisiae: Impact of ER/Golgi Ca(2+) storage.

    Science.gov (United States)

    D'hooge, Petra; Coun, Catherina; Van Eyck, Vincent; Faes, Liesbeth; Ghillebert, Ruben; Mariën, Lore; Winderickx, Joris; Callewaert, Geert

    2015-08-01

    Yeast has proven to be a powerful tool to elucidate the molecular aspects of several biological processes in higher eukaryotes. As in mammalian cells, yeast intracellular Ca(2+) signalling is crucial for a myriad of biological processes. Yeast cells also bear homologs of the major components of the Ca(2+) signalling toolkit in mammalian cells, including channels, co-transporters and pumps. Using yeast single- and multiple-gene deletion strains of various plasma membrane and organellar Ca(2+) transporters, combined with manipulations to estimate intracellular Ca(2+) storage, we evaluated the contribution of individual transport systems to intracellular Ca(2+) homeostasis. Yeast strains lacking Pmr1 and/or Cod1, two ion pumps implicated in ER/Golgi Ca(2+) homeostasis, displayed a fragmented vacuolar phenotype and showed increased vacuolar Ca(2+) uptake and Ca(2+) influx across the plasma membrane. In the pmr1Δ strain, these effects were insensitive to calcineurin activity, independent of Cch1/Mid1 Ca(2+) channels and Pmc1 but required Vcx1. By contrast, in the cod1Δ strain increased vacuolar Ca(2+) uptake was not affected by Vcx1 deletion but was largely dependent on Pmc1 activity. Our analysis further corroborates the distinct roles of Vcx1 and Pmc1 in vacuolar Ca(2+) uptake and point to the existence of not-yet identified Ca(2+) influx pathways.

  19. Different response of osteoblastic cells to Mg(2+), Zn(2+) and Sr(2+) doped calcium silicate coatings.

    Science.gov (United States)

    Hu, Dandan; Li, Kai; Xie, Youtao; Pan, Houhua; Zhao, Jun; Huang, Liping; Zheng, Xuebin

    2016-03-01

    Mg(2+), Zn(2+) and Sr(2+) substitution for Ca(2+) in plasma sprayed calcium silicate (Ca-Si) coatings have been reported to impede their degradation in physiological environment and, more importantly, to improve their biological performance. The reason for the improved biological performance is still elusive and, especially, the contribution of the dopant ions is lack of obvious and direct evidence. In this study, we aim to identify the effect of Mg(2+), Zn(2+) and Sr(2+) incorporation on the osteogenic ability of Ca-Si based coatings (Ca2MgSi2O7, Ca2ZnSi2O7 and Sr-CaSiO3) by minimizing the influence of Ca and Si ions release and surface physical properties. Similar surface morphology, crystallinity and roughness were achieved for all samples by optimizing the spray parameters. As expected, Ca and Si ions release from all the coatings showed the comparable concentration with immersing time. The response of MC3T3-E1 cells onto Mg(2+), Zn(2+) and Sr(2+) doped Ca-Si coatings were studied in terms of osteoblastic adhesion, proliferation, differentiation and mineralization. The results showed that the level of cell adhesion and proliferation increased the most on the surface of Mg-modified coating. Gene expressions of early markers of osteoblast differentiation (COL-I and ALP mRNA) were obviously improved on Zn-modified coating. Gene expressions of later markers for osteoblast differentiation (OPN and OC mRNA) and mineralized nodules formation were obviously accelerated on the surface of Sr-modified coating. Since Mg(2+), Zn(2+) and Sr(2+) play a regulatory role in different stages of osteogenesis, it may be possible to utilize this in the development of new coating materials for orthopedic application. PMID:26787488

  20. P4-ATPases

    DEFF Research Database (Denmark)

    Lopez Marques, Rosa Laura; Theorin, Lisa; Palmgren, Michael Broberg;

    2014-01-01

    Cellular membranes, notably eukaryotic plasma membranes, are equipped with special proteins that actively translocate lipids from one leaflet to the other and thereby help generate membrane lipid asymmetry. Among these ATP-driven transporters, the P4 subfamily of P-type ATPases (P4-ATPases......) comprises lipid flippases that catalyze the translocation of phospholipids from the exoplasmic to the cytosolic leaflet of cell membranes. While initially characterized as aminophospholipid translocases, recent studies of individual P4-ATPase family members from fungi, plants, and animals show that P4...... to include the regulation of membrane traffic, cytoskeletal dynamics, cell division, lipid metabolism, and lipid signaling. In this review, we will summarize the basic features of P4-ATPases and the physiological implications of their lipid transport activity in the cell. © 2013 The Author(s)....

  1. Crystal structure of the high-affinity Na+,K+-ATPase–ouabain complex with Mg2+ bound in the cation binding site

    DEFF Research Database (Denmark)

    Laursen, Mette; Yatime, Laure; Nissen, Poul;

    2013-01-01

    The Na+,K+-ATPase maintains electrochemical gradients for Na+ and K+ that are critical for animal cells. Cardiotonic steroids (CTSs), widely used in the clinic and recently assigned a role as endogenous regulators of intracellular processes, are highly specific inhibitors of the Na+,K+-ATPase. Here......325 toward the cation site and thereby hindering deep ouabain binding. Thus, the structural data establish a basis for the interpretation of the biochemical evidence pointing at direct K+–Mg2+ competition and explain the well-known antagonistic effect of K+ on CTS binding....

  2. Secondary structure of the intact H+,K+ -ATPase and of its membrane-embedded region. An attenuated total reflection infrared spectroscopy, circular dichroism and Raman spectroscopy study

    NARCIS (Netherlands)

    Raussens, V.; Jongh, H. de; Pézolet, M.; Ruysschaert, J.-M.; Goormaghtigh, E.

    1998-01-01

    Models of P-type ATPase predict that membrane-embedded fragments represent about 20% of the protein and adopt an all-α-helical structure. While this prediction was confirmed for the Ca2+ -ATPase [Corbalan-Garcia, S., Teruel, J., Villalain, J. and Gomez-Fernandez, J. (1994) Biochemistry 33, 8247-8254

  3. Crystal structure of a copper-transporting PIB-type ATPase

    DEFF Research Database (Denmark)

    Gourdon, Pontus Emanuel; Liu, Xiang-Yu; Skjørringe, Tina;

    2011-01-01

    (+)-ATPase, in a copper-free form, as determined by X-ray crystallography at 3.2 Å resolution. The structure indicates a three-stage copper transport pathway involving several conserved residues. A PIB-specific transmembrane helix kinks at a double-glycine motif displaying an amphipathic helix that lines a putative...... copper entry point at the intracellular interface. Comparisons to Ca(2+)-ATPase suggest an ATPase-coupled copper release mechanism from the binding sites in the membrane via an extracellular exit site. The structure also provides a framework to analyse missense mutations in the human ATP7A and ATP7B...

  4. Quantal release of Ca2+ from intracellular stores by InsP3: tests of the concept of control of Ca2+ release by intraluminal Ca2+.

    Science.gov (United States)

    Tregear, R T; Dawson, A P; Irvine, R F

    1991-03-22

    A possible mechanism for the generation of 'quantal' release of intracellular Ca2+ by InsP3 (Muallem et al., J. biol. Chem. 264, 205-212 (1989)) has been put forward in which intraluminal Ca2+ levels modulate InsP3 receptor structure (Irvine, FEBS Lett. 263, 5-9 (1990)). Here we have modelled such a steady-state mechanism, with an InsP3-sensitive store plus an InsP3-insensitive one, to test its ability to mimic published data. We have also performed experiments on InsP3-stimulated rat liver microsomes to test whether the model is consistent with one-way Ca2+ fluxes at a steady state. The model can simulate quantal release, in that InsP3 produces a release of part of the stored Ca2+ which is initially rapid relative to the one-way flux. In the original form of the model, in which InsP3-modulated Ca2+ binding to the intraluminal site opens the Ca2+ channel, the range of InsP3 concentrations needed to release Ca2+ is greater than that observed. When the model is changed so that Ca2(+)-modulated InsP3 binding opens the channels, the effective InsP3 range is shortened, but the quantal release effect is reduced. Other published data on one-way fluxes, and our own data on microsomes, can be simulated when leakage from the InsP3-insensitive store is adjusted to fit the observations; these data therefore do not test the existence of a steady state in the InsP3-sensitive store. We conclude that sensitivity of Ca2+ release to intraluminal Ca2+ provides a steady-state explanation of most, but not all, current quantal release observations.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1675803

  5. Reporting a new siderophore based Ca(2+) selective chemosensor that works as a staining agent in the live organism Artemia.

    Science.gov (United States)

    Raju, M; Nair, Ratish R; Raval, Ishan H; Haldar, Soumya; Chatterjee, Pabitra B

    2015-11-21

    A Ca(2+)-specific chemosensor involving acyclic non-ether and non-carboxylato-type metal chelating ligands is rare. The tetradentate OONO artificial receptor, HL, possessing a sulfur-containing intermediate siderophore aeruginic acid, tethered to a rhodamine 6G based signalling unit in a single molecule has been synthesized. The fluoroionophore required excitation in the visible wavelength (510 nm) and showed highly selective and sensitive detection of Ca(2+) ions in 100% water solution in HEPES buffer at physiological pH (7.4). The probe HL, with LOD as low as 70 nM, behaves reversibly and showed nearly 17-fold enhanced selectivity for Ca(2+) over other cell abundant alkali and alkaline metal ions such as Na(+), K(+), Li(+), and Mg(2+) without any intervention. Job's plot, (1)H NMR titration and ESI-MS data provided corroborative evidence in support of 1 : 1 association between HL and Ca(2+). From a wide range of transition and heavy metal ions series, HL also binds Cu(2+). However, the use of l-cysteine removes the interference from Cu(2+) and results in highly selective detection specificity of HL for Ca(2+). As a reversible "off-on-off" fluorescent chemosensor, it is possible to detect Ca(2+) at as low as 5 μM in the midgut region of the gastrointestinal tract of the live animal Artemia, a brine shrimp. PMID:26460620

  6. The mitochondrial Na+/Ca2+ exchanger upregulates glucose dependent Ca2+ signalling linked to insulin secretion.

    Directory of Open Access Journals (Sweden)

    Iulia I Nita

    Full Text Available Mitochondria mediate dual metabolic and Ca(2+ shuttling activities. While the former is required for Ca(2+ signalling linked to insulin secretion, the role of the latter in β cell function has not been well understood, primarily because the molecular identity of the mitochondrial Ca(2+ transporters were elusive and the selectivity of their inhibitors was questionable. This study focuses on NCLX, the recently discovered mitochondrial Na(+/Ca(2+ exchanger that is linked to Ca(2+ signalling in MIN6 and primary β cells. Suppression either of NCLX expression, using a siRNA construct (siNCLX or of its activity, by a dominant negative construct (dnNCLX, enhanced mitochondrial Ca(2+ influx and blocked efflux induced by glucose or by cell depolarization. In addition, NCLX regulated basal, but not glucose-dependent changes, in metabolic rate, mitochondrial membrane potential and mitochondrial resting Ca(2+. Importantly, NCLX controlled the rate and amplitude of cytosolic Ca(2+ changes induced by depolarization or high glucose, indicating that NCLX is a critical and rate limiting component in the cross talk between mitochondrial and plasma membrane Ca(2+ signalling. Finally, knockdown of NCLX expression was followed by a delay in glucose-dependent insulin secretion. These findings suggest that the mitochondrial Na(+/Ca(2+ exchanger, NCLX, shapes glucose-dependent mitochondrial and cytosolic Ca(2+ signals thereby regulating the temporal pattern of insulin secretion in β cells.

  7. Activation of the skeletal muscle Ca2+ release channel by the triazine dyes cibacron blue F3A-G and reactive red 120

    International Nuclear Information System (INIS)

    Vesicle-45Ca2+ ion flux and planar lipid bilayer single-channel measurements have shown that the Ca2+ release channel of skeletal muscle sarcoplasmic reticulum (SR) is activated by micromolar concentrations of Cibacron Blue F3A-G (Reactive Blue 2) and Reactive Red 120. Cibacron Blue increased the 45Ca2+ efflux rate from heavy SR vesicles by apparently interacting with both the adenine nucleotide and caffeine activating sites of the channel. Dye-induced 45Ca2+ release was inhibited by Mg2+ and ruthenium red. In single channel recordings with the purified channel protein complex, Cibacron Blue increased the open time of the Ca2+ release channel without an apparent change in the conductance of the main and subconductance states of the channel

  8. Basolateral Mg2+ extrusion via CNNM4 mediates transcellular Mg2+ transport across epithelia: a mouse model.

    Directory of Open Access Journals (Sweden)

    Daisuke Yamazaki

    Full Text Available Transcellular Mg(2+ transport across epithelia, involving both apical entry and basolateral extrusion, is essential for magnesium homeostasis, but molecules involved in basolateral extrusion have not yet been identified. Here, we show that CNNM4 is the basolaterally located Mg(2+ extrusion molecule. CNNM4 is strongly expressed in intestinal epithelia and localizes to their basolateral membrane. CNNM4-knockout mice showed hypomagnesemia due to the intestinal malabsorption of magnesium, suggesting its role in Mg(2+ extrusion to the inner parts of body. Imaging analyses revealed that CNNM4 can extrude Mg(2+ by exchanging intracellular Mg(2+ with extracellular Na(+. Furthermore, CNNM4 mutations cause Jalili syndrome, characterized by recessive amelogenesis imperfecta with cone-rod dystrophy. CNNM4-knockout mice showed defective amelogenesis, and CNNM4 again localizes to the basolateral membrane of ameloblasts, the enamel-forming epithelial cells. Missense point mutations associated with the disease abolish the Mg(2+ extrusion activity. These results demonstrate the crucial importance of Mg(2+ extrusion by CNNM4 in organismal and topical regulation of magnesium.

  9. Osmotically induced cytosolic free Ca(2+) changes in human neutrophils.

    Science.gov (United States)

    Morris, M R; Doull, I J; Hallett, M B

    2001-02-01

    Cytosolic free Ca(2+) concentration in neutrophils was measured by ratiometric fluorometry of intracellular fura2. Increasing the extracellular osmolarity, by either NaCl (300-600 mM) or sucrose (600-1200 mM), caused a rise in cytosolic free Ca(2+) (Delta(max) approximately equal to 600 nM). This was not due to cell lysis as the cytosolic free Ca(2+) concentration was reversed by restoration of isotonicity and a second rise in cytosolic free Ca(2+) could be provoked by repeating the change in extracellular osmolarity. Furthermore, the rise in cytosolic free Ca(2+) concentration occurred in the absence of extracellular Ca(2+), demonstrating that release of intracellular fura2 into the external medium did not occur. The osmotically-induced rise in cytosolic free Ca(2+) was not inhibited by either the phospholipase C-inhibitor U73122, or the microfilament inhibitor cytochalasin B, suggesting that neither signalling via inositol tris-phosphate or the cytoskeletal system were involved. However, the rise in cytosolic free Ca(2+) may have resulted from a reduction in neutrophil water volume in hyperosmotic conditions. As these rises in cytosolic Ca(2+) (Delta(max) approximately equal to 600 nM) were large enough to provoke changes in neutrophil activity, we propose that conditions which removes cell water may similarly elevate cytosolic free Ca(2+) to physiologically important levels. PMID:11341979

  10. Spontaneous Ca2+ oscillations in subcellular compartments of vascular smooth muscle cells rely on different Ca2+ pools

    Institute of Scientific and Technical Information of China (English)

    2004-01-01

    Spontaneous Ca2+ oscillations in vascular smooth muscle cells have been modeled using a single Ca2+ pool. This report describes spontaneous Ca2+ oscillations dependent on two separate Ca2+ sources for the nuclear versus cytoplasmic compartments. Changes in free intracellular Ca2+ were monitored with ratiometric Ca2+- fluorophores using confocal microscopy. On average, spontaneous oscillations developed in 79% of rat aortic smooth muscle cells that were synchronous between the cytoplasm and nucleus. Reduction of extracellular Ca2+ (< 1 μM) decreased the frequency and amplitude of the cytoplasmic oscillations with 48% of the oscillations asynchronous between the nuclear and cytoplasmic compartments. Similar results were obtained with the Ca2+ channel blockers, nimodipine and diltiazem.Arg-vasopressin (AVP) induced a rapid release of intracellular Ca2+ stores that was greater in the nuclear compartment (4.20 ± 0.23 ratio units, n = 56) than cytoplasm (2.54 ± 0.28) in cells that had spontaneously developed prior oscillations.Conversely, cells in the same conditions lacking oscillations had a greater AVP-induced Ca2+ transient in the cytoplasm (4.99 ± 0.66, n = 17) than in the nucleus (2.67 ± 0.29). Pre-treatment with Ca2+ channel blockers depressed the AVP responses in both compartments with the cytoplasmic Ca2+ most diminished. Depletion of internal Ca2+ stores prior to AVP exposure blunted the nuclear response, mimicking the response of cells that lacked prior oscillations. Spontaneous oscillating cells had a greater sarcoplasmic reticulum network than cells that did not oscillate. We propose that spontaneous nuclear oscillations rely on perinuclear sarcoplasmic reticulum stores, while the cytoplasmic oscillations rely on Ca2+ influx.

  11. Effect of vanadate and of removal of extracellular Ca2+ and Na+ on tension development and 45Ca efflux in rat and frog myocardium

    DEFF Research Database (Denmark)

    Gesser, H; Bonefeld-Jørgensen, Eva Cecilie

    1983-01-01

    Vanadate in the range 0-5 mM has positive inotropic effects on myocardial strips of frog and to a lesser extent on those of rat. Inhibiting the sarcolemmal Na+, Ca2+ exchange by a solution free of Ca2+ and Na+ caused a drop in 45Ca efflux and a transient increase in resting tension. These effects...... were more expressed for the frog than for the rat myocardium, which suggests that the Na+ for Ca2+ exchange across the cell membrane is more important in the frog than in the rat myocardium. A subsequent addition of vanadate at 2 or 5 mM had no effect on 45Ca efflux, while it increased the resting...... tension. This increase was higher for the frog than for the rat myocardium. These results suggest that the inotropic effects of vanadate may be due to an effect on membrane-bound Ca2+-ATPase....

  12. The other side of cardiac Ca2+ signaling: transcriptional control

    Directory of Open Access Journals (Sweden)

    Alejandro eDomínguez-Rodríquez

    2012-11-01

    Full Text Available Ca2+ is probably the most versatile signal transduction element used by all cell types. In the heart, it is essential to activate cellular contraction in each heartbeat. Nevertheless Ca2+ is not only a key element in excitation-contraction coupling (EC coupling, but it is also a pivotal second messenger in cardiac signal transduction, being able to control processes such as excitability, metabolism, and transcriptional regulation. Regarding the latter, Ca2+ activates Ca2+-dependent transcription factors by a process called excitation-transcription coupling (ET coupling. ET coupling is an integrated process by which the common signaling pathways that regulate EC coupling activate transcription factors. Although ET coupling has been extensively studied in neurons and other cell types, less is known in cardiac muscle. Some hints have been found in studies on the development of cardiac hypertrophy, where two Ca2+-dependent enzymes are key actors: Ca2+/Calmodulin kinase II (CaMKII and phosphatase calcineurin, both of which are activated by the complex Ca2+/ /Calmodulin. The question now is how ET coupling occurs in cardiomyocytes, where intracellular Ca2+ is continuously oscillating. In this focused review, we will draw attention to location of Ca2+ signaling: intranuclear ([Ca2+]n or cytoplasmic ([Ca2+]c, and the specific ionic channels involved in the activation of cardiac ET coupling. Specifically, we will highlight the role of the 1,4,5 inositol triphosphate receptors (IP3Rs in the elevation of [Ca2+]n levels, which are important to locally activate CaMKII, and the role of transient receptor potential channels canonical (TRPCs in [Ca2+]c, needed to activate calcineurin.

  13. External bioenergy-induced increases in intracellular free calcium concentrations are mediated by Na+/Ca2+ exchanger and L-type calcium channel.

    Science.gov (United States)

    Kiang, Juliann G; Ives, John A; Jonas, Wayne B

    2005-03-01

    External bioenergy (EBE, energy emitted from a human body) has been shown to increase intracellular calcium concentration ([Ca2+]i, an important factor in signal transduction) and regulate the cellular response to heat stress in cultured human lymphoid Jurkat T cells. In this study, we wanted to elucidate the underlying mechanisms. A bioenergy specialist emitted bioenergy sequentially toward tubes of cultured Jurkat T cells for one 15-minute period in buffers containing different ion compositions or different concentrations of inhibitors. [Ca2+], was measured spectrofluorometrically using the fluorescent probe fura-2. The resting [Ca2+]i in Jurkat T cells was 70 +/- 3 nM (n = 130) in the normal buffer. Removal of external calcium decreased the resting [Ca2+]i to 52 +/- 2 nM (n = 23), indicating that Ca2+ entry from the external source is important for maintaining the basal level of [Ca2+]i. Treatment of Jurkat T cells with EBE for 15 min increased [Ca2+]i by 30 +/- 5% (P EBE did not attenuate [Ca2+]i responsiveness to EBE. Removal of external Ca2+ or Na+, but not Mg2+, inhibited the EBE-induced increase in [Ca2+]i. Dichlorobenzamil, an inhibitor of Na+/Ca2+ exchangers, also inhibited the EBE-induced increase in [Ca2+]i in a concentration-dependent manner with an IC50 of 0.11 +/- 0.02 nM. When external [K+] was increased from 4.5 mM to 25 mM, EBE decreased [Ca2+]i. The EBE-induced increase was also blocked by verapamil, an L-type voltage-gated Ca2+ channel blocker. These results suggest that the EBE-induced [Ca2+]i increase may serve as an objective means for assessing and validating bioenergy effects and those specialists claiming bioenergy capability. The increase in [Ca2+]i is mediated by activation of Na+/Ca2+ exchangers and opening of L-type voltage-gated Ca2+ channels.

  14. The Role of Extracellular Ca2+ Influx,Intracellular Ca2+ Release and Calmodulin in Mouse Egg Fertilization

    Institute of Scientific and Technical Information of China (English)

    SunQing-yuan; TanJing-he; 等

    1999-01-01

    The effects of various Ca2+-modifying drugs on moue egg fertilization were studied.Ca2+ chelator,ethylen glycol-bis-(2-aminoethyl)-tetracetic acid(EGTA),and calmodulin(CaM) antagonist,trifluoperzaine (TFP),inhibited fertilization in a dose-dependent manner,whild Ca2+ channel bolcker,verspamil,did not have any effect.When intracellular Ca2+ release was blocked by 8-(N,N-diethylamino) octy 1-3,4,5-trimethoxy-benzonate(TME-8) or the Ca2+ oscillations were inhibited by an inhibitor of endoplasmic reticulum Ca2+-At-Pase,thapsigargin,the second polar body emission and pronuclear formation were significantly decreased.In contrast,inhibition of intracellular Ca2+ release via bolckage of inositol 1,4,5-triphosphate (IP3) production by neomycin or lithium did not affect fertilization.The results sugest that both extracellular influx,intracellular Ca2+ release and CaM activation are required for mormal fertilization.However,extracellular influx through voltage-gated Ca2+ channel and intracellular release induced by IP3 and not the only pathways for producing Ca2+ transients in moue eggs.

  15. The Role of Extracellular Ca2+Influx, Intracellular Ca2+ Release and Calmodulin in Mouse Egg Fertilization

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    The effects of various Ca2+-modifying drugs on moue egg fertilization were studied. Ca2+ chelator, ethylen glycol-bis-(2-aminoethyl)-tetracetic acid (EGTA) ,and calmodulin (CaM) antagonist,trifluoperzaine (TFP) ,inhibited fertilization in a dose-dependent manner,whild Ca2+ channel bolcker,verapamil ,did not have any effect. When intracellular Ca2+ release was blocked by 8-(N, N-diethylamino) octy1-3,4,5-trimethoxy- benzonate (TMB-8) or the Ca2+ oscillations were inhibited by an inhibitor of endoplasmic reticulum Ca2+-AT- Pase,thapsigargin,the second polar body emission and pronuclear formation were significantly decreased. In contrast,inhibition of intracellular Ca2+ release via bolckage of inositol 1,4,5-triphosphate (IP3) production by neomycin or lithium did not affect fertilization. The results sugest that both extracellular influx,intracellu- lar Ca2+ release and CaM activation are required for normal fertilization. However ,extracellular influx through voltage-gated Ca2+ channel and intracellular release induced by IP3 are not the only pathways for producing Ca2+ transients in moue eggs.

  16. Ca2+-induced activation and irreversible inactivation of chloride channels in the perfused plasmalemma of Nitellopsis obtusa.

    Science.gov (United States)

    Kataev, A A; Zherelova, O M; Berestovsky, G N

    1984-12-01

    Experiments were carried out on the algal cells with removed tonoplast using both continuous intracellular perfusion and voltage clamp on plasmalemma. The transient plasmalemma current induced by depolarization disappeared upon perfusion with the Ca2+-chelating agent, EGTA, since the voltage-dependent calcium channels lost their ability to activate. Subsequent replacement of the perfusion medium containing EGTA by another with Ca2+ for clamped plasmalemma (-100 mV) induced an inward C1- current which showed both activation and inactivation. The maximal amplitude of the current at [C1-]in = 15 mmol/l (which is similar to that in native cells) was approximately twice that in electrically excited cell in vivo. The inactivation of C1 channels in the presence of internal Ca2+ was irreversible and had a time constant of 1-3 min. This supports our earlier suggestion (Lunevsky et al. 1983) that the inactivation of C1 channels in an intact cell (with a time constant of 1-3 s) is due to a decrease in Ca2+ concentration rather than to the activity of their own inactivation mechanism. The C1 channel selectivity sequence was following: C1- much greater than CH3SO-4 approximately equal to K+ much greater than SO2-4 (PK/PSO4 approximately 10). Activation of one half the channels occurs at a Ca2+ concentration of 2 X 10(-5) mol/l. Sr2+ also (though to a lesser extent) activated C1 channels but had to be present in a much more higher concentration than Ca2+. Mg2+ and Ba2+ appeared ineffective. Ca2+ activation did not, apparently, require participation of water-soluble intermediator including ATP. Thus, C1 channel functioning is controlled by Ca2+-, Sr2+-sensitive elements of the subplasmalemma cytoskeleton. PMID:6099298

  17. Local Aqueous Solvation Structure Around Ca2+ During Ca2+---Cl– Pair Formation

    Energy Technology Data Exchange (ETDEWEB)

    Baer, Marcel D.; Mundy, Christopher J.

    2016-03-03

    The molecular details of single ion solvation around Ca2+ and ion-pairing of Ca2--Cl- are investigated using ab initio molecular dynamics. The use of empirical dispersion corrections to the BLYP functional are investigated by comparison to experimentally available extended X-ray absorption fine structure (EXAFS) measurements, which probes the first solvation shell in great detail. Besides finding differences in the free-energy for both ion-pairing and the coordination number of ion solvation between the quantum and classical descriptions of interaction, there were important differences found between dispersion corrected and uncorrected density functional theory (DFT). Specifically, we show significantly different free-energy landscapes for both coordination number of Ca2+ and its ion-pairing with Cl- depending on the DFT simulation protocol. Our findings produce a self-consistent treatment of short-range solvent response to the ion and the intermediate to long-range collective response of the electrostatics of the ion-ion interaction to produce a detailed picture of ion-pairing that is consistent with experiment. MDB is supported by MS3 (Materials Synthesis and Simulation Across Scales) Initiative at Pacific Northwest National Laboratory. It was conducted under the Laboratory Directed Research and Development Program at PNNL, a multiprogram national laboratory operated by Battelle for the U.S. Department of Energy. CJM acknowledges support from US Department of Energy, Office of Science, Office of Basic Energy Sciences, Division of Chemical Sciences, Geosciences & Biosciences. This research used resources of the National Energy Research Scientific Computing Center, a DOE Office of Science User Facility supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC02-05CH11231. Additional computing resources were generously allocated by PNNL's Institutional Computing program. The authors thank Prof. Tom Beck for discussions

  18. TRPV5-mediated Ca2+ Reabsorption and Hypercalciuria

    Science.gov (United States)

    Renkema, Kirsten Y.; Hoenderop, Joost G. J.; Bindels, René J. M.

    2007-04-01

    The concerted action of the intestine, kidney and bone results in the maintenance of a normal Ca2+ balance, a mechanism that is tightly controlled by the calciotropic hormones vitamin D, parathyroid hormone and calcitonin. Disturbances in the Ca2+ balance have been linked to diverse pathophysiological disorders like urolithiasis, hypertension, electroencephalogram abnormalities and rickets. Importantly, the final amount of Ca2+ that is released from the body is determined in the distal part of the nephron, where active Ca2+ reabsorption occurs. Here, Transient Receptor Potential Vanilloid member 5 (TRPV5), a highly Ca2+-selective channel, has been recognized as the gatekeeper of active Ca2+ reabsorption. The in vivo relevance of TRPV5 has been further investigated by the characterization of TRPV5 knockout (TRPV5-/-) mice, which exhibit severe disturbances in renal Ca2+ handling, such as profound hypercalciuria, intestinal Ca2+ hyperabsorption and reduced bone thickness. Hypercalciuria increases the risk of kidney stone formation in these mice. This review highlights our current knowledge about TRPV5-mediated Ca2+ reabsorption and emphasizes the physiological relevance and the clinical implications related to the TRPV5-/- mice model.

  19. New approaches to the study of mitochondrial Ca2+ dynamic

    OpenAIRE

    Fuente Pérez, Sergio de la

    2014-01-01

    A pesar de la importancia del Ca2+ como segundo mensajero intracelular, aun existen discrepancias en la literatura sobre los niveles de Ca2+ que se alcanzan en el interior de la mitocondria tras un estímulo fisiológico. En esta tesis se ha realizado una comparativa detallada de los diferentes métodos de medida del Ca2+mitocondrial y además se han desarrollado nuevos métodos de medida para dicho Ca2+. Gracias a estas nuevas técnicas hemos podido realizar un estudio exhaustivo de los flujos de ...

  20. Intracellular Ca(2+) release as irreversible Markov process.

    OpenAIRE

    Rengifo, Juliana; Rosales, Rafael; González, Adom; Cheng, Heping; Stern, Michael D.; Ríos, Eduardo

    2002-01-01

    In striated muscles, intracellular Ca(2+) release is tightly controlled by the membrane voltage sensor. Ca(2+) ions are necessary mediators of this control in cardiac but not in skeletal muscle, where their role is ill-understood. An intrinsic gating oscillation of Ca(2+) release-not involving the voltage sensor-is demonstrated in frog skeletal muscle fibers under voltage clamp. A Markov model of the Ca(2+) release units is shown to reproduce the oscillations, and it is demonstrated that for ...

  1. Revisiting the mechanisms of copper toxicity to rainbow trout: Time course, influence of calcium, unidirectional Na(+) fluxes, and branchial Na(+), K(+) ATPase and V-type H(+) ATPase activities.

    Science.gov (United States)

    Chowdhury, M Jasim; Girgis, Mina; Wood, Chris M

    2016-08-01

    In order to resolve uncertainties as to the mechanisms of toxic action of Cu and the protective effects of water [Ca], juvenile rainbow trout were acclimated to baseline soft water (SW, [Na(+)]=0.07, [Ca(2+)]=0.15, [Mg(2+)]=0.05mmolL(-1)) and then exposed to Cu with or without elevated [Ca] but at constant titratable alkalinity (0.27mmolL(-1)). The 96-h LC50 was 7-fold higher (63.8 versus 9.2μgCuL(-1); 1.00 versus 0.14μmolCuL(-1)) at [Ca]=3.0 versus 0.15mmolL(-1). Gill Cu burden increased with exposure concentration, and higher [Ca] attenuated this accumulation. At 24h, the gill Cu load (LA50≈0.58μgCug(-1); 9.13nmolCug(-1)) predictive of 50% mortality by 96h was independent of [Ca], in accord with Biotic Ligand Model (BLM) theory. Cu exposure induced net Na(+) losses (J(Na)net) by increasing unidirectional Na(+) efflux rates (J(Na)out) and inhibiting unidirectional Na(+) uptake rates (J(Na)in). The effect on J(Na)out was virtually immediate, whereas the effect on J(Na)in developed progressively over 24h and was associated with an inhibition of branchial Na(+), K(+) ATPase activity. The J(Na)in inhibition was eventually significant at a lower Cu threshold concentration (15μgCuL(-1)) than the J(Na)out stimulation (100μg Cu L(-1)). Elevated Ca protected against both effects, as well as against the inhibition of Na(+), K(+) ATPase activity. Branchial V-type H(+) ATPase activity was also inhibited by Cu exposure (100μgCuL(-1)), but only after 24h at high [Ca] (3.0mmolL(-1)). These novel results therefore reinforce the applicability of BLM theory to Cu, clarify that whether Na(+) influx or efflux is more sensitive depends on the duration of Cu exposure, show that elevated water [Ca], independent of alkalinity, is protective against both mechanisms of Cu toxicity, and identify V-type H(+)ATPase as a new Cu target for future investigation. PMID:27262060

  2. Ca2+-Clock-Dependent Pacemaking in the Sinus Node Is Impaired in Mice with a Cardiac Specific Reduction in SERCA2 Abundance

    OpenAIRE

    Logantha, Sunil Jit R.J.; Stokke, Mathis K.; Atkinson, Andrew J.; Kharche, Sanjay R.; Parveen, Sajida; Saeed, Yawer; Sjaastad, Ivar; Sejersted, Ole M; Dobrzynski, Halina

    2016-01-01

    Background: The sarcoplasmic reticulum Ca2+-ATPase (SERCA2) pump is an important component of the Ca2+-clock pacemaker mechanism that provides robustness and flexibility to sinus node pacemaking. We have developed transgenic mice with reduced cardiac SERCA2 abundance (Serca2 KO) as a model for investigating SERCA2's role in sinus node pacemaking. Methods and Results: In Serca2 KO mice, ventricular SERCA2a protein content measured by Western blotting was 75% (P < 0.05) lower than that in contr...

  3. Ca(2+)-activated chloride channel activity during Ca(2+) alternans in ventricular myocytes.

    Science.gov (United States)

    Kanaporis, Giedrius; Blatter, Lothar A

    2016-11-01

    Cardiac alternans, defined beat-to-beat alternations in contraction, action potential (AP) morphology or cytosolic Ca transient (CaT) amplitude, is a high risk indicator for cardiac arrhythmias. We investigated mechanisms of cardiac alternans in single rabbit ventricular myocytes. CaTs were monitored simultaneously with membrane currents or APs recorded with the patch clamp technique. A strong correlation between beat-to-beat alternations of AP morphology and CaT alternans was observed. During CaT alternans application of voltage clamp protocols in form of pre-recorded APs revealed a prominent Ca(2+)-dependent membrane current consisting of a large outward component coinciding with AP phases 1 and 2, followed by an inward current during AP repolarization. Approximately 85% of the initial outward current was blocked by Cl(-) channel blocker DIDS or lowering external Cl(-) concentration identifying it as a Ca(2+)-activated Cl(-) current (ICaCC). The data suggest that ICaCC plays a critical role in shaping beat-to-beat alternations in AP morphology during alternans.

  4. Effects of Siraitia grosvenori Leaves Flavonoid and Swim Training on Metabolism of ATPase in Quadriceps of Exhaustive Rats%罗汉果叶黄酮及游泳训练对力竭大鼠股四头肌组织ATP酶代谢的影响

    Institute of Scientific and Technical Information of China (English)

    邓启烈; 陈梅; 莫伟彬; 杨永亮; 杨峰

    2013-01-01

    activities in administration exhaustive exercise group and training exhaustive exercise group were significantly higher than that of quiet exhaustive exercise group ( P < 0. 05 , P < 0. 01 ) , Ca ATPase activities were appeared upward trend, though these data had no statistically significance. At the point of immediately after exhaustive exercise, the activity of total ATPase was significantly decreased, though the total ATPase in administration exhaustive exercise group and training exhaustive exercise group were significantly higher than that of quiet exhaustive exercise group ( P < 0. 01 ) , and it in training-administration exhaustive exercise group was significantly higher than that of quiet-administration exhaustive exercise group and training exhaustive exercise group (P < 0. 01). Conclusion:Na+/ K+ -ATPase, Ca2+/Mg2+ -ATPase and total ATPase in quadriceps of swimming exhaustive rats could increased by administration of S. grosvenori leaves flavonoid. This data indicated that Siraitia Grosvenori leaves flavonoid has important significance to maintain the osmotic pressure of cell membrane and the balance of transmembrane potential. And the 5. grosvenori leaves flavonoid could also maintain the balance of Ca2 + /Mg2+ to ensure higher energy supply, improve the capacity of anti-oxidant, and delay the onset of exercise-induced fatigue.%目的:通过建立大强度耐力训练及一次力竭性游泳运动损伤模型,研究罗汉果叶黄酮抗疲劳能力以及维持细胞膜内外的渗透压及跨膜电位平衡中的作用.方法:选用健康雄性SD大鼠40只,2~3月龄,体重180 ~ 220 g,随机分成5组,每组8只,即:安静对照组、安静力竭组、安静给药力竭组、训练力竭组、训练+给药力竭组.给药量以生理盐水配成含罗汉果黄酮提取物200 mg·kg-1·d-1(20 g·L-1的溶液,按10.0 mL·kg-1·d-1 ig给药),对照组ig等量的生理盐水.在递增负荷游泳训练4周后,各组进行3%的负重力竭性游泳训练,准确

  5. A yeast expression system for functional and pharmacological studies of the malaria parasite Ca2+/H+ antiporter

    Directory of Open Access Journals (Sweden)

    Salcedo-Sora J

    2012-08-01

    Full Text Available Abstract Background Calcium (Ca2+ signalling is fundamental for host cell invasion, motility, in vivo synchronicity and sexual differentiation of the malaria parasite. Consequently, cytoplasmic free Ca2+ is tightly regulated through the co-ordinated action of primary and secondary Ca2+ transporters. Identifying selective inhibitors of Ca2+ transporters is key towards understanding their physiological role as well as having therapeutic potential, therefore screening systems to facilitate the search for potential inhibitors are a priority. Here, the methodology for the expression of a Calcium membrane transporter that can be scaled to high throughputs in yeast is presented. Methods The Plasmodium falciparum Ca2+/H+ antiporter (PfCHA was expressed in the yeast Saccharomyces cerevisiae and its activity monitored by the bioluminescence from apoaequorin triggered by divalent cations, such as calcium, magnesium and manganese. Results Bioluminescence assays demonstrated that PfCHA effectively suppressed induced cytoplasmic peaks of Ca2+, Mg2+ and Mn2+ in yeast mutants lacking the homologue yeast antiporter Vcx1p. In the scalable format of 96-well culture plates pharmacological assays with a cation antiporter inhibitor allowed the measurement of inhibition of the Ca2+ transport activity of PfCHA conveniently translated to the familiar concept of fractional inhibitory concentrations. Furthermore, the cytolocalization of this antiporter in the yeast cells showed that whilst PfCHA seems to locate to the mitochondrion of P. falciparum, in yeast PfCHA is sorted to the vacuole. This facilitates the real-time Ca2+-loading assays for further functional and pharmacological studies. Discussion The functional expression of PfCHA in S. cerevisiae and luminescence-based detection of cytoplasmic cations as presented here offer a tractable system that facilitates functional and pharmacological studies in a high-throughput format. PfCHA is shown to behave as a divalent

  6. Ca2+ improves organization of single-stranded DNA bases in human Rad51 filament, explaining stimulatory effect on gene recombination.

    KAUST Repository

    Fornander, Louise H

    2012-02-22

    Human RAD51 protein (HsRad51) catalyses the DNA strand exchange reaction for homologous recombination. To clarify the molecular mechanism of the reaction in vitro being more effective in the presence of Ca(2+) than of Mg(2+), we have investigated the effect of these ions on the structure of HsRad51 filament complexes with single- and double-stranded DNA, the reaction intermediates. Flow linear dichroism spectroscopy shows that the two ionic conditions induce significantly different structures in the HsRad51/single-stranded DNA complex, while the HsRad51/double-stranded DNA complex does not demonstrate this ionic dependence. In the HsRad51/single-stranded DNA filament, the primary intermediate of the strand exchange reaction, ATP/Ca(2+) induces an ordered conformation of DNA, with preferentially perpendicular orientation of nucleobases relative to the filament axis, while the presence of ATP/Mg(2+), ADP/Mg(2+) or ADP/Ca(2+) does not. A high strand exchange activity is observed for the filament formed with ATP/Ca(2+), whereas the other filaments exhibit lower activity. Molecular modelling suggests that the structural variation is caused by the divalent cation interfering with the L2 loop close to the DNA-binding site. It is proposed that the larger Ca(2+) stabilizes the loop conformation and thereby the protein-DNA interaction. A tight binding of DNA, with bases perpendicularly oriented, could facilitate strand exchange.

  7. Glucose-stimulated oscillations in free cytosolic ATP concentration imaged in single islet beta-cells: evidence for a Ca2+-dependent mechanism.

    Science.gov (United States)

    Ainscow, Edward K; Rutter, Guy A

    2002-02-01

    Normal glucose-stimulated insulin secretion is pulsatile, but the molecular mechanisms underlying this pulsatility are poorly understood. Oscillations in the intracellular free [ATP]/[ADP] ratio represent one possible mechanism because they would be expected to cause fluctuations in ATP-sensitive K(+) channel activity and hence oscillatory Ca(2+) influx. After imaging recombinant firefly luciferase, expressed via an adenoviral vector in single human or mouse islet beta-cells, we report here that cytosolic free ATP concentrations oscillate and that these oscillations are affected by glucose. In human beta-cells, oscillations were observed at both 3 and 15 mmol/l glucose, but the oscillations were of a longer wavelength at the higher glucose concentration (167 vs. 66 s). Mouse beta-cells displayed oscillations in both cytosolic free [Ca(2+)] and [ATP] only at elevated glucose concentrations, both with a period of 120 s. To explore the causal relationship between [Ca(2+)] and [ATP] oscillations, the regulation of each was further investigated in populations of MIN6 beta-cells. Incubation in Ca(2+)-free medium lowered cytosolic [Ca(2+)] but increased [ATP] in MIN6 cells at both 3 and 30 mmol/l glucose. Removal of external Ca(2+) increased [ATP], possibly by decreasing ATP consumption by endoplasmic reticulum Ca(2+)-ATPases. These results allow a model to be constructed of the beta-cell metabolic oscillator that drives nutrient-induced insulin secretion.

  8. Store-Operated Ca2+ Release-Activated Ca2+ Channels Regulate PAR2-Activated Ca2+ Signaling and Cytokine Production in Airway Epithelial Cells.

    Science.gov (United States)

    Jairaman, Amit; Yamashita, Megumi; Schleimer, Robert P; Prakriya, Murali

    2015-09-01

    The G-protein-coupled protease-activated receptor 2 (PAR2) plays an important role in the pathogenesis of various inflammatory and auto-immune disorders. In airway epithelial cells (AECs), stimulation of PAR2 by allergens and proteases triggers the release of a host of inflammatory mediators to regulate bronchomotor tone and immune cell recruitment. Activation of PAR2 turns on several cell signaling pathways of which the mobilization of cytosolic Ca(2+) is likely a critical but poorly understood event. In this study, we show that Ca(2+) release-activated Ca(2+) (CRAC) channels encoded by stromal interaction molecule 1 and Orai1 are a major route of Ca(2+) entry in primary human AECs and drive the Ca(2+) elevations seen in response to PAR2 activation. Activation of CRAC channels induces the production of several key inflammatory mediators from AECs including thymic stromal lymphopoietin, IL-6, and PGE2, in part through stimulation of gene expression via nuclear factor of activated T cells (NFAT). Furthermore, PAR2 stimulation induces the production of many key inflammatory mediators including PGE2, IL-6, IL-8, and GM-CSF in a CRAC channel-dependent manner. These findings indicate that CRAC channels are the primary mechanism for Ca(2+) influx in AECs and a vital checkpoint for the induction of PAR2-induced proinflammatory cytokines. PMID:26238490

  9. RESPONSE SURFACE METHODOLOGY AS OPTIMIZATION TOOL IN STUDY OF COMPETITIVE EFFECT OF Ca2+ AND Mg2+ IONS IN SORPTION PROCESS OF Co2+ BY DRIED ACTIVATED SLUDGE

    Directory of Open Access Journals (Sweden)

    Lucia Remenárová

    2012-04-01

    Full Text Available The aim of present study was to investigate biosorption of cobalt ions onto dried activated sludge (DAS from ternary solution Co-Ca-Mg, as a function of pH and concentration of cobalt, calcium and magnesium via Box-Behnken design under Response surface methodology (RSM. The four parameters namely cobalt initial concentration of 2000 µmol/L, calcium initial concentration of 2000 µmol/L, magnesium initial concentration of 2000 µmol/L and pH of 5.5 were chosen as center points from the previous study of metal sorption process. The experimental data on sorption capacity Qexp of DAS for Co2+ ions were obtained by 60Co gamaspectrometry and fitted into a quadratic polynomial model using multiple regression analysis. The experimental Box-Behnken design revealed that biosorption of cobalt ions from ternary system increase with increasing pH of the system, initial cobalt concentration and magnesium concentration. Results showed also drastic inhibitory effect of increasing calcium concentration in sorption process of cobalt ions. The present study provides valuable information about combined effect of independent variables on sorption process from multivariable system and enables us to minimize number of needed experiments from 60 to 29.

  10. Development of a method for calculating the equilibrium and kinetics of ion exchange on a weak acid resin in a ternary system H+-Ca2+-Mg2+

    International Nuclear Information System (INIS)

    In technical applications ion exchange resins are applied in filters. The breakthrough behaviour of such filters can be calculated using mathematical relationships for equilibrium and kinetics. An according method has been developed for a ternary ion exchage problem on a weak acid resin. Theoretical results are verified by means of experimental data. (orig.)

  11. Synthesis of layered double hydroxides containing Mg2+, Zn2+, Ca2+ and Al3+ layer cations by co-precipitation methods-A review

    Science.gov (United States)

    Theiss, Frederick L.; Ayoko, Godwin A.; Frost, Ray L.

    2016-10-01

    Co-precipitation is a common method for the preparation of layered double hydroxides (LDHs) and related materials. This review article is aimed at providing newcomers to the field with some examples of the types of co-precipitation reactions that have been reported previously and to briefly investigate some of the properties of the products of these reactions. Due to the sheer volume of literature on the subject, the authors have had to limit this article to the synthesis of Mg/Al, Zn/Al and Ca/Al LDHs by co-precipitation and directly related methods. LDHs have been synthesised from various reagents including metal salts, oxides and hydroxides. Co-precipitation is also useful for the direct synthesis of LDHs with a wide range of interlayer anions and various bases have been successfully employed to prepare LDHs. Examples of other synthesis techniques including the urea method, hydrothermal synthesis and various mechanochemical methods that are undoubtedly related to co-precipitation have also been included in this review. The effect of post synthesis hydrothermal has also been summarised.

  12. Cell surface topology creates high Ca2+ signalling microdomains

    DEFF Research Database (Denmark)

    Brasen, Jens Christian; Olsen, Lars Folke; Hallett, Maurice B

    2010-01-01

    It has long been speculated that cellular microdomains are important for many cellular processes, especially those involving Ca2+ signalling. Measurements of cytosolic Ca2+ report maximum concentrations of less than few micromolar, yet several cytosolic enzymes require concentrations of more than...

  13. Hierarchic stochastic modelling applied to intracellular Ca(2+ signals.

    Directory of Open Access Journals (Sweden)

    Gregor Moenke

    Full Text Available Important biological processes like cell signalling and gene expression have noisy components and are very complex at the same time. Mathematical analysis of such systems has often been limited to the study of isolated subsystems, or approximations are used that are difficult to justify. Here we extend a recently published method (Thurley and Falcke, PNAS 2011 which is formulated in observable system configurations instead of molecular transitions. This reduces the number of system states by several orders of magnitude and avoids fitting of kinetic parameters. The method is applied to Ca(2+ signalling. Ca(2+ is a ubiquitous second messenger transmitting information by stochastic sequences of concentration spikes, which arise by coupling of subcellular Ca(2+ release events (puffs. We derive analytical expressions for a mechanistic Ca(2+ model, based on recent data from live cell imaging, and calculate Ca(2+ spike statistics in dependence on cellular parameters like stimulus strength or number of Ca(2+ channels. The new approach substantiates a generic Ca(2+ model, which is a very convenient way to simulate Ca(2+ spike sequences with correct spiking statistics.

  14. Spectral components of cytosolic [Ca2+] spiking in neurons

    DEFF Research Database (Denmark)

    Kardos, J; Szilágyi, N; Juhász, G;

    1998-01-01

    . Delayed complex responses of large [Ca2+]c spiking observed in cells from a different set of cultures were synthesized by a set of frequencies within the range 0.018-0.117 Hz. Differential frequency patterns are suggested as characteristics of the [Ca2+]c spiking responses of neurons under different...

  15. The NA+/K+-ATPase controls gap junctions via membrane microdomain interactions in rat smooth muscles.

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Nilsson, Holger; Aalkjær, Christian

    The Na+/K+-ATPase is known to interact with many membrane and cytosolic proteins by organizing various signaling complexes. These interactions were suggested to be important in regulation of various cellular responses. Pumping activity of the Na+/K+-ATPase is suggested to be essential for some...... in rat mesenteric small arteries. Paired cultured rat smooth muscle cells (A7r5) were used as a model for electrical coupling of SMC by measuring membrane capacitance (Cm). PCR, Western blotting and immunohistochemistry were used to identify the membrane transporters. SMCs were uncoupled (evaluated...... in regulation of the intercellular communication. We have here shown that gap junctions between SMCs are regulated through an interaction between the Na+/K+-ATPase and the Na+/Ca2+-exchanger leading to an increase in [Ca2+]i in discrete areas near the plasma membrane. We have also suggested that this Na...

  16. Glutamate excitotoxicity and Ca(2+)-regulation of respiration: Role of the Ca(2+) activated mitochondrial transporters (CaMCs).

    Science.gov (United States)

    Rueda, Carlos B; Llorente-Folch, Irene; Traba, Javier; Amigo, Ignacio; Gonzalez-Sanchez, Paloma; Contreras, Laura; Juaristi, Inés; Martinez-Valero, Paula; Pardo, Beatriz; Del Arco, Araceli; Satrustegui, Jorgina

    2016-08-01

    Glutamate elicits Ca(2+) signals and workloads that regulate neuronal fate both in physiological and pathological circumstances. Oxidative phosphorylation is required in order to respond to the metabolic challenge caused by glutamate. In response to physiological glutamate signals, cytosolic Ca(2+) activates respiration by stimulation of the NADH malate-aspartate shuttle through Ca(2+)-binding to the mitochondrial aspartate/glutamate carrier (Aralar/AGC1/Slc25a12), and by stimulation of adenine nucleotide uptake through Ca(2+) binding to the mitochondrial ATP-Mg/Pi carrier (SCaMC-3/Slc25a23). In addition, after Ca(2+) entry into the matrix through the mitochondrial Ca(2+) uniporter (MCU), it activates mitochondrial dehydrogenases. In response to pathological glutamate stimulation during excitotoxicity, Ca(2+) overload, reactive oxygen species (ROS), mitochondrial dysfunction and delayed Ca(2+) deregulation (DCD) lead to neuronal death. Glutamate-induced respiratory stimulation is rapidly inactivated through a mechanism involving Poly (ADP-ribose) Polymerase-1 (PARP-1) activation, consumption of cytosolic NAD(+), a decrease in matrix ATP and restricted substrate supply. Glutamate-induced Ca(2+)-activation of SCaMC-3 imports adenine nucleotides into mitochondria, counteracting the depletion of matrix ATP and the impaired respiration, while Aralar-dependent lactate metabolism prevents substrate exhaustion. A second mechanism induced by excitotoxic glutamate is permeability transition pore (PTP) opening, which critically depends on ROS production and matrix Ca(2+) entry through the MCU. By increasing matrix content of adenine nucleotides, SCaMC-3 activity protects against glutamate-induced PTP opening and lowers matrix free Ca(2+), resulting in protracted appearance of DCD and protection against excitotoxicity in vitro and in vivo, while the lack of lactate protection during in vivo excitotoxicity explains increased vulnerability to kainite-induced toxicity in Aralar

  17. Glutamate excitotoxicity and Ca2+-regulation of respiration: Role of the Ca2+ activated mitochondrial transporters (CaMCs).

    Science.gov (United States)

    Rueda, Carlos B; Llorente-Folch, Irene; Traba, Javier; Amigo, Ignacio; Gonzalez-Sanchez, Paloma; Contreras, Laura; Juaristi, Inés; Martinez-Valero, Paula; Pardo, Beatriz; Del Arco, Araceli; Satrustegui, Jorgina

    2016-08-01

    Glutamate elicits Ca(2+) signals and workloads that regulate neuronal fate both in physiological and pathological circumstances. Oxidative phosphorylation is required in order to respond to the metabolic challenge caused by glutamate. In response to physiological glutamate signals, cytosolic Ca(2+) activates respiration by stimulation of the NADH malate-aspartate shuttle through Ca(2+)-binding to the mitochondrial aspartate/glutamate carrier (Aralar/AGC1/Slc25a12), and by stimulation of adenine nucleotide uptake through Ca(2+) binding to the mitochondrial ATP-Mg/Pi carrier (SCaMC-3/Slc25a23). In addition, after Ca(2+) entry into the matrix through the mitochondrial Ca(2+) uniporter (MCU), it activates mitochondrial dehydrogenases. In response to pathological glutamate stimulation during excitotoxicity, Ca(2+) overload, reactive oxygen species (ROS), mitochondrial dysfunction and delayed Ca(2+) deregulation (DCD) lead to neuronal death. Glutamate-induced respiratory stimulation is rapidly inactivated through a mechanism involving Poly (ADP-ribose) Polymerase-1 (PARP-1) activation, consumption of cytosolic NAD(+), a decrease in matrix ATP and restricted substrate supply. Glutamate-induced Ca(2+)-activation of SCaMC-3 imports adenine nucleotides into mitochondria, counteracting the depletion of matrix ATP and the impaired respiration, while Aralar-dependent lactate metabolism prevents substrate exhaustion. A second mechanism induced by excitotoxic glutamate is permeability transition pore (PTP) opening, which critically depends on ROS production and matrix Ca(2+) entry through the MCU. By increasing matrix content of adenine nucleotides, SCaMC-3 activity protects against glutamate-induced PTP opening and lowers matrix free Ca(2+), resulting in protracted appearance of DCD and protection against excitotoxicity in vitro and in vivo, while the lack of lactate protection during in vivo excitotoxicity explains increased vulnerability to kainite-induced toxicity in Aralar

  18. Modulation of the matrix redox signaling by mitochondrial Ca2+

    Institute of Scientific and Technical Information of China (English)

    Jaime; Santo-Domingo; Andreas; Wiederkehr; Umberto; De; Marchi

    2015-01-01

    Mitochondria sense,shape and integrate signals,and thus function as central players in cellular signal transduction. Ca2+ waves and redox reactions are two such intracellular signals modulated by mitochondria. Mitochondrial Ca2+ transport is of utmost physio-pathological relevance with a strong impact on metabolism and cell fate. Despite its importance,the molecular nature of the proteins involvedin mitochondrial Ca2+ transport has been revealed only recently. Mitochondrial Ca2+ promotes energy metabolism through the activation of matrix dehydrogenases and downstream stimulation of the respiratory chain. These changes also alter the mitochondrial NAD(P)H/NAD(P)+ ratio,but at the same time will increase reactive oxygen species(ROS) production. Reducing equivalents and ROS are having opposite effects on the mitochondrial redox state,which are hard to dissect. With the recent development of genetically encoded mitochondrial-targeted redoxsensitive sensors,real-time monitoring of matrix thiol redox dynamics has become possible. The discoveries of the molecular nature of mitochondrial transporters of Ca2+ combined with the utilization of the novel redox sensors is shedding light on the complex relation between mitochondrial Ca2+ and redox signals and their impact on cell function. In this review,we describe mitochondrial Ca2+ handling,focusing on a number of newly identified proteins involved in mitochondrial Ca2+ uptake and release. We further discuss our recent findings,revealing how mitochondrial Ca2+ influences the matrix redox state. As a result,mitochondrial Ca2+ is able to modulate the many mitochondrial redox-regulated processes linked to normal physiology and disease.

  19. Isoprenaline enhances local Ca2+ release in cardiac myocytes

    Institute of Scientific and Technical Information of China (English)

    Jian-xin SHEN

    2006-01-01

    Aim: Contraction of cardiac myocytes is controlled by the generation and amplification of intracellular Ca2+ signals. The key step of this process is the coupling between sarcolemma L-type Ca2+ channels (LCCs) and ryanodine receptors (RyRs) in the sarcoplasmic reticulum (SR). β-Adrenergic stimulation is an important regulatory mechanism for this coupling process. But the details underlied the global level, which require local Ca2+ release study are still unclear. The present study is to explore the effects of β-adrenergic stimulation on local Ca2+ release. Methods: Using confocal microscopy combined with loose-seal patch-clamp approaches, effects of isoprenaline (1 μmol·L-1), a β-adrenergic agonist, on local SR Ca2+ release triggered by Ca2+ influx through LCCs in intact rat cardiac myocytes were investigated. Results: Isoprenaline increased the intensity of ensemble averaged local Ca2+ transients, the peak of which displayed a typical bell-shaped voltage-dependence over the membrane voltages ranging from ~-40mV to ~+35mV. Further analysis showed that this enhancement could be explained by the increased coupling fidelity (which refers the increased probability of RyRs activation upon depolarization), and the increased amplitude of evoked Ca2+ sparks (due to more Ca2+ releases through local RyRs). In addition, isoprenaline decreased the first latency, which displayed a typical "U"-shaped voltage-dependence, showing the available acceleration and synchronization of β-adrenergic stimulation on intracellular calcium release. Conclusions: Isoprenaline enhances local Ca2+ release in cardiac myocytes. These results underscore the importance of regulation of β-adrenergic stimulation on local intermolecular signals between LCCs and RyRs in heart cells.

  20. Cannabinoid receptors in submandibular acinar cells: functional coupling between saliva fluid and electrolytes secretion and Ca2+ signalling.

    Science.gov (United States)

    Kopach, Olga; Vats, Juliana; Netsyk, Olga; Voitenko, Nana; Irving, Andrew; Fedirko, Nataliya

    2012-04-15

    Cannabinoid receptors (CBRs) belong to the G protein-coupled receptor superfamily, and activation of CBRs in salivary cells inhibits agonist-stimulated salivation and modifies saliva content. However, the role of different CBR subtypes in acinar cell physiology and in intracellular signalling remains unclear. Here, we uncover functional CB(1)Rs and CB(2)Rs in acinar cells of rat submandibular gland and their essential role in saliva secretion. Pharmacological activation of CB(1)Rs and CB(2)Rs in the submandibular gland suppressed saliva outflow and modified saliva content produced by the submandibular gland in vivo. Using Na(+)-selective microelectrodes to record secretory Na(+) responses in the lumen of acini, we observed a reduction in Na(+) transport following the activation of CBRs, which was counteracted by the selective CB(1)R antagonist AM251. In addition, activation of CB(1)Rs or CB Rs caused inhibition of Na(+)-K(+) 2 -ATPase activity in microsomes derived from the gland tissue as well as in isolated acinar cells. Using a Ca(2+) imaging technique, we showed that activation of CB(1)Rs and CB(2)Rs alters [Ca(2+)](cyt) signalling in acinar cells by distinct pathways, involving Ca(2+) release from the endoplasmic reticulum (ER) and store-operated Ca(2+) entry (SOCE), respectively. Our data demonstrate the expression of CB(1)Rs and CB(2)Rs in acinar cells, and their involvement in the regulation of salivary gland functioning.

  1. Inhibition of Brain Swelling after Ischemia-Reperfusion by β-Adrenergic Antagonists: Correlation with Increased K+ and Decreased Ca2+ Concentrations in Extracellular Fluid

    Directory of Open Access Journals (Sweden)

    Dan Song

    2014-01-01

    Full Text Available Infarct size and brain edema following ischemia/reperfusion are reduced by inhibitors of the Na+, K+, 2Cl−, and water cotransporter NKCC1 and by β1-adrenoceptor antagonists. NKCC1 is a secondary active transporter, mainly localized in astrocytes, driven by transmembrane Na+/K+ gradients generated by the Na+,K+-ATPase. The astrocytic Na+,K+-ATPase is stimulated by small increases in extracellular K+ concentration and by the β-adrenergic agonist isoproterenol. Larger K+ increases, as occurring during ischemia, also stimulate NKCC1, creating cell swelling. This study showed no edema after 3 hr medial cerebral artery occlusion but pronounced edema after 8 hr reperfusion. The edema was abolished by inhibitors of specifically β1-adrenergic pathways, indicating failure of K+-mediated, but not β1-adrenoceptor-mediated, stimulation of Na+,K+-ATPase/NKCC1 transport during reoxygenation. Ninety percent reduction of extracellular Ca2+ concentration occurs in ischemia. Ca2+ omission abolished K+ uptake in normoxic cultures of astrocytes after addition of 5 mM KCl. A large decrease in ouabain potency on K+ uptake in cultured astrocytes was also demonstrated in Ca2+-depleted media, and endogenous ouabains are needed for astrocytic K+ uptake. Thus, among the ionic changes induced by ischemia, the decrease in extracellular Ca2+ causes failure of the high-K+-stimulated Na+,K+-ATPase/NKCC1 ion/water uptake, making β1-adrenergic activation the only stimulus and its inhibition effective against edema.

  2. Mg2(Si,Sn)-based thermoelectric materials and devices

    Science.gov (United States)

    Gao, Peng

    Thermoelectric effects are phenomena found in materials that can achieve direct conversion between heat flow and electricity. One important application of thermoelectric effects is thermoelectric generators, which can generate electricity when a temperature gradient is applied. Thermoelectric generators make use of various sources of heat and it is considered a promising solution for waste heat recovery. The conversion efficiency of thermoelectric generators depends on the materials used in the devices. Significant improvement in the performance of thermoelectric materials has been made in the past few decades. However, most of the good thermoelectric materials being investigated have limitations, such as the high materials cost, high materials density and toxicity of the constituent elements. The Mg2(Si,Sn)-based materials studied in this work are promising candidates for thermoelectric generators in the mid-temperature range and have drawn increasing research interest in recent years because these materials are high performance thermoelectrics that are low cost, low-density and non-toxic. In this work, systematic studies were performed on the Mg2(Si,Sn) thermoelectric materials. Thermal phase stability was studied for different compositions of Mg2Si1-xSnx and Mg2Si0.4Sn 0.6 was used as base material for further optimization. Both n-type and p-type samples were obtained by doping the materials with different elements. Peak ZT ˜ 1.5 for the n-type and ZT ˜ 0.7 for the p-type materials were obtained, both of which are among the best reported results so far. Experimental work was also done to study the techniques to develop the Mg2Si 0.4Sn0.6 materials into working devices. Different electrode materials were tested in bonding experiment for this compound, and copper was found to be the best electrode material for Mg2Si 0.4Sn0.6. Preliminary work was done to demonstrate the possibility of fabricating a Mg2Si0.4Sn0.6-based thermoelectric generator and the result is

  3. Genetic Background Influences Adaptation To Cardiac Hypertrophy and Ca2+ Handling Gene Expression

    Directory of Open Access Journals (Sweden)

    Steve B Waters

    2013-03-01

    Full Text Available Genetic variability has a profound effect on the development of cardiac hypertrophy in response to stress. Consequently, using a variety of inbred mouse strains with known genetic profiles may be powerful models for studying the response to cardiovascular stress. To explore this approach we looked at male C57BL/6J and 129/SvJ mice. Hemodynamic analyses of left ventricular pressures indicated significant differences in 129/SvJ and C57BL/6J mice that implied altered Ca2+ handling. Specifically, 129/SvJ mice demonstrated reduced rates of relaxation and insensitivity to dobutamine(Db. We hypothesized that altered expression of genes controlling the influx and efflux of Ca2+ from the sarcoplasmic reticulum was responsible and investigated the expression of several genes involved in maintaining the intracellular and sarcoluminal Ca2+ concentration using quantitative real-time PCR analyses (qRT-PCR. We observed significant differences in baseline gene expression as well as different responses in expression to isoproterenol (ISO challenge. In untreated control animals, 129/SvJ mice expressed 1.68x more ryanodine recptor 2(Ryr2 mRNA than C57BL/6J mice but only 0.37x as much calsequestrin 2(Casq2. After treatment with ISO, sarco(endoplasmic reticulum Ca2+-ATPase(Serca2 expression was reduced nearly two-fold in 129/SvJ while expression in C57BL/6J was stable. Interestingly, β(1 adrenergic receptor(Adrb1 expression was lower in 129/SvJ compared to C57BL/6J at baseline and lower in both strains after treatment. Metabolically, the brain isoform of creatine kinase(Ckb was up-regulated in response to ISO in C57BL/6J but not in 129/SvJ. These data suggest that the two strains of mice regulate Ca2+ homeostasis via different mechanisms and may be useful in developing personalized therapies in human patients.

  4. Genetic background influences adaptation to cardiac hypertrophy and Ca(2+) handling gene expression.

    Science.gov (United States)

    Waters, Steve B; Diak, Douglass M; Zuckermann, Matthew; Goldspink, Paul H; Leoni, Lara; Roman, Brian B

    2013-01-01

    Genetic variability has a profound effect on the development of cardiac hypertrophy in response to stress. Consequently, using a variety of inbred mouse strains with known genetic profiles may be powerful models for studying the response to cardiovascular stress. To explore this approach we looked at male C57BL/6J and 129/SvJ mice. Hemodynamic analyses of left ventricular pressures (LVPs) indicated significant differences in 129/SvJ and C57BL/6J mice that implied altered Ca(2+) handling. Specifically, 129/SvJ mice demonstrated reduced rates of relaxation and insensitivity to dobutamine (Db). We hypothesized that altered expression of genes controlling the influx and efflux of Ca(2+) from the sarcoplasmic reticulum (SR) was responsible and investigated the expression of several genes involved in maintaining the intracellular and sarcoluminal Ca(2+) concentration using quantitative real-time PCR analyses (qRT-PCR). We observed significant differences in baseline gene expression as well as different responses in expression to isoproterenol (ISO) challenge. In untreated control animals, 129/SvJ mice expressed 1.68× more ryanodine receptor 2(Ryr2) mRNA than C57BL/6J mice but only 0.37× as much calsequestrin 2 (Casq2). After treatment with ISO, sarco(endo)plasmic reticulum Ca(2+)-ATPase(Serca2) expression was reduced nearly two-fold in 129/SvJ while expression in C57BL/6J was stable. Interestingly, β (1) adrenergic receptor(Adrb1) expression was lower in 129/SvJ compared to C57BL/6J at baseline and lower in both strains after treatment. Metabolically, the brain isoform of creatine kinase (Ckb) was up-regulated in response to ISO in C57BL/6J but not in 129/SvJ. These data suggest that the two strains of mice regulate Ca(2+) homeostasis via different mechanisms and may be useful in developing personalized therapies in human patients.

  5. Echinacea-induced cytosolic Ca2+ elevation in HEK293

    Directory of Open Access Journals (Sweden)

    Nikolau Basil J

    2010-11-01

    Full Text Available Abstract Background With a traditional medical use for treatment of various ailments, herbal preparations of Echinacea are now popularly used to improve immune responses. One likely mode of action is that alkamides from Echinacea bind to cannabinoid type 2 (CB2 receptors and induce a transient increase in intracellular Ca2+. Here, we show that unidentified compounds from Echinacea purpurea induce cytosolic Ca2+ elevation in non-immune-related cells, which lack CB2 receptors and that the Ca2+ elevation is not influenced by alkamides. Methods A non-immune human cell line, HEK293, was chosen to evaluate E. purpurea root extracts and constituents as potential regulators of intracellular Ca2+ levels. Changes in cytosolic Ca2+ levels were monitored and visualized by intracellular calcium imaging. U73122, a phospholipase C inhibitor, and 2-aminoethoxydiphenyl borate (2-APB, an antagonist of inositol-1,4,5-trisphosphate (IP3 receptor, were tested to determine the mechanism of this Ca2+ signaling pathway. E. purpurea root ethanol extracts were fractionated by preparative HPLC, screened for bioactivity on HEK293 cells and by GC-MS for potential constituent(s responsible for this bioactivity. Results A rapid transient increase in cytosolic Ca2+ levels occurs when E. purpurea extracts are applied to HEK293 cells. These stimulatory effects are phospholipase C and IP3 receptor dependent. Echinacea-evoked responses could not be blocked by SR 144528, a specific CB2 receptor antagonist, indicating that CB2 is not involved. Ca2+ elevation is sustained after the Echinacea-induced Ca2+ release from intracellular Ca2+ stores; this longer-term effect is abolished by 2-APB, indicating a possible store operated calcium entry involvement. Of 28 HPLC fractions from E. purpurea root extracts, six induce cytosolic Ca2+ increase. Interestingly, GC-MS analysis of these fractions, as well as treatment of HEK293 cells with known individual and combined chemicals, indicates the

  6. Development of Ca2+ hotspots between Lymnaea neurons during synaptogenesis.

    Science.gov (United States)

    Feng, Zhong-Ping; Grigoriev, Nikita; Munno, David; Lukowiak, Ken; MacVicar, Brian A; Goldberg, Jeffrey I; Syed, Naweed I

    2002-02-15

    Calcium (Ca2+) channel clustering at specific presynaptic sites is a hallmark of mature synapses. However, the spatial distribution patterns of Ca2+ channels at newly formed synapses have not yet been demonstrated. Similarly, it is unclear whether Ca2+ 'hotspots' often observed at the presynaptic sites are indeed target cell contact specific and represent a specialized mechanism by which Ca2+ channels are targeted to select synaptic sites. Utilizing both soma-soma paired (synapsed) and single neurons from the mollusk Lymnaea, we have tested the hypothesis that differential gradients of voltage-dependent Ca2+ signals develop in presynaptic neuron at its contact point with the postsynaptic neuron; and that these Ca2+ hotspots are target cell contact specific. Fura-2 imaging, or two-photon laser scanning microscopy of Calcium Green, was coupled with electrophysiological techniques to demonstrate that voltage-induced Ca2+ gradients (hotspots) develop in the presynaptic cell at its contact point with the postsynaptic neuron, but not in unpaired single cells. The incidence of Ca2+ hotspots coincided with the appearance of synaptic transmission between the paired cells, and these gradients were target cell contact specific. In contrast, the voltage-induced Ca2+ signal in unpaired neurons was uniformly distributed throughout the somata; a similar pattern of Ca2+ gradient was observed in the presynaptic neuron when it was soma-soma paired with a non-synaptic partner cell. Moreover, voltage clamp recording techniques, in conjunction with a fast, optical differential perfusion system, were used to demonstrate that the total whole-cell Ca2+ (or Ba2+) current density in single and paired cells was not significantly different. However, the amplitude of Ba2+ current was significantly higher in the presynaptic cell at its contact side with the postsynaptic neurons, compared with non-contacted regions. In summary, this study demonstrates that voltage-induced Ca2+ hotspots develop

  7. Short and long range functions of amino acids in the transmembrane region of the sarcoplasmic reticulum ATPase. A mutational study.

    Science.gov (United States)

    Chen, L; Sumbilla, C; Lewis, D; Zhong, L; Strock, C; Kirtley, M E; Inesi, G

    1996-05-01

    Mutational analysis of several amino acids in the transmembrane region of the sarcoplasmic reticulum ATPase was performed by expressing wild type ATPase and 32 site-directed mutants in COS-1 cells followed by functional characterization of the microsomal fraction. Four different phenotype characteristics were observed in the mutants: (a) functions similar to those sustained by the wild type ATPase; (b) Ca2+ transport inhibited to a greater extent than ATPase hydrolytic activity; (c) inhibition of transport and hydrolytic activity in the presence of high levels of phosphorylated enzyme intermediate; and (d) total inhibition of ATP utilization by the enzyme while retaining the ability to form phosphoenzyme by utilization of P(i). Analysis of experimental observations and molecular models revealed short and long range functions of several amino acids within the transmembrane region. Short range functions include: (a) direct involvement of five amino acids in Ca2+ binding within a channel formed by clustered transmembrane helices M4, M5, M6, and M8; (b) roles of several amino acids in structural stabilization of the helical cluster for optimal channel function; and (c) a specific role of Lys297 in sealing the distal end of the channel, suggesting that the M4 helix rotates to allow vectorial flux of Ca2+ upon enzyme phosphorylation. Long range functions are related to the influence of several transmembrane amino acids on phosphorylation reactions with ATP or P(i), transmitted to the extramembranous region of the ATPase in the presence or in the absence of Ca2+.

  8. Study on the effect of doxorubicin on expressions of genes encoding myocardial sarcoplasmic reticulum Ca2+ transport proteins and the effect of taurine on myocardial protection in rabbits

    Institute of Scientific and Technical Information of China (English)

    黄先玫; 朱卫华; 康曼丽

    2003-01-01

    To investigate the effect of doxorubicin(DOX) on gene expression of the myocardial sarcoplasmic reticulum (SR)Ca2+ transport proteins and the mechanism of taurine(Tau) protecting cardiac muscle cells, 9 rabbits were injected with DOX , 8 rabbits with DOX and Tau, and 9 rabbits with normal saline. Cardiac function , concentration of calcium in cardiomyocytes ( Myo [ Ca2+ ]i ), activity of SR Ca2+ -ATPase (SERCA2a) , level of SERCA2a mRNA and Ca2+ released channels(RYR2) mRNA were detected. The left ventricle tissues were observed by electron microscopy. The results showed that cardiac index, left ventricular systolic pressure, activity of SR Ca2+ -ATPase and level of SERCA2a mRNA decreased , while Myo[ Ca2+ ]i increased in DOX-treated rabbits. DOX could not affect the level of RYR2 mRNA. Tau intervention could alleviate the increase of left ventricular diastolic pressure, Myo[ Ca2+ ] i and the decrease of SERCA2a mRNA induced by doxorubicin. Tile results suggested that downregulation of SERCA2a gene expression was an important mechanism of DOX-induced cardiomyopathy and that Tau could partially improve the heart function by reducing calcium overload and alleviating downregulation of SERCA2a mRNA.

  9. Modulation of intracellular Ca2+ levels by Scorpaenidae venoms.

    Science.gov (United States)

    Church, Jarrod E; Moldrich, Randal X; Beart, Philip M; Hodgson, Wayne C

    2003-05-01

    The crude venoms of the soldierfish (Gymnapistes marmoratus), the lionfish (Pterois volitans) and the stonefish (Synanceia trachynis) display pronounced neuromuscular activity. Since [Ca(2+)](i) is a key regulator in many aspects of neuromuscular function we sought to determine its involvement in the neuromuscular actions of the venoms. In the chick biventer cervicis muscle, all three venoms produced a sustained contraction (approx 20-30% of 1mM acetylcholine). Blockade of nicotinic receptors with tubocurarine (10 micro M) failed to attenuate the contractile response to either G. marmoratus venom or P. volitans venom, but produced slight inhibition of the response to S. trachynis venom. All three venoms produced a rise in intracellular Ca(2+) (approx. 200-300% of basal) in cultured murine cortical neurons. The Ca(2+)-channel blockers omega-conotoxin MVIIC, omega-conotoxin GVIA, omega-agatoxin IVa and nifedipine (each at 1 micro M) potentiated the increase in [Ca(2+)](i) in response to G. marmoratus venom and P. volitans venom, while attenuating the response to S. trachynis venom. Removal of extracellular Ca(2+), replacement of Ca(2+) with La(3+) (0.5mM), or addition of stonefish antivenom (3units/ml) inhibited both the venom-induced increase in [Ca(2+)](i) in cultured neurones and contraction in chick biventer cervicis muscle. Venom-induced increases in [Ca(2+)](i) correlated with an increased cell death of cultured neurones as measured using propidium iodide (1 micro g/ml). Morphological analysis revealed cellular swelling and neurite loss consistent with necrosis. These data indicate that the effects of all three venoms are due in part to an increase in intracellular Ca(2+), possibly via the formation of pores in the cellular membrane which, under certain conditions, can lead to necrosis. PMID:12727272

  10. Hypervitaminosis D mediates compensatory Ca2+ hyperabsorption in TRPV5 knockout mice.

    NARCIS (Netherlands)

    Renkema, K.Y.R.; Nijenhuis, T.; Eerden, B.C. van der; Kemp, J.W.C.M. van der; Weinans, H.; Leeuwen, J.P.P.M. van; Bindels, R.J.M.; Hoenderop, J.G.J.

    2005-01-01

    Vitamin D plays an important role in Ca(2+) homeostasis by controlling Ca(2+) (re)absorption in intestine, kidney, and bone. The epithelial Ca(2+) channel TRPV5 mediates the Ca(2+) entry step in active Ca(2+) reabsorption. TRPV5 knockout (TRPV5(-/-)) mice show impaired Ca(2+) reabsorption, hypercalc

  11. A calcium-dependent protein kinase can inhibit a calmodulin-stimulated Ca2+ pump (ACA2) located in the endoplasmic reticulum of Arabidopsis

    Science.gov (United States)

    Hwang, I.; Sze, H.; Harper, J. F.; Evans, M. L. (Principal Investigator)

    2000-01-01

    The magnitude and duration of a cytosolic Ca(2+) release can potentially be altered by changing the rate of Ca(2+) efflux. In plant cells, Ca(2+) efflux from the cytoplasm is mediated by H(+)/Ca(2+)-antiporters and two types of Ca(2+)-ATPases. ACA2 was recently identified as a calmodulin-regulated Ca(2+)-pump located in the endoplasmic reticulum. Here, we show that phosphorylation of its N-terminal regulatory domain by a Ca(2+)-dependent protein kinase (CDPK isoform CPK1), inhibits both basal activity ( approximately 10%) and calmodulin stimulation ( approximately 75%), as shown by Ca(2+)-transport assays with recombinant enzyme expressed in yeast. A CDPK phosphorylation site was mapped to Ser(45) near a calmodulin binding site, using a fusion protein containing the N-terminal domain as an in vitro substrate for a recombinant CPK1. In a full-length enzyme, an Ala substitution for Ser(45) (S45/A) completely blocked the observed CDPK inhibition of both basal and calmodulin-stimulated activities. An Asp substitution (S45/D) mimicked phosphoinhibition, indicating that a negative charge at this position is sufficient to account for phosphoinhibition. Interestingly, prior binding of calmodulin blocked phosphorylation. This suggests that, once ACA2 binds calmodulin, its activation state becomes resistant to phosphoinhibition. These results support the hypothesis that ACA2 activity is regulated as the balance between the initial kinetics of calmodulin stimulation and CDPK inhibition, providing an example in plants for a potential point of crosstalk between two different Ca(2+)-signaling pathways.

  12. Effects of Calcium on ATPase Activity and Lipid Composition of Plasma Membranes from Wheat Roots Under Aluminum Stress

    Institute of Scientific and Technical Information of China (English)

    HE Long-fei; SHEN Zhen-guo; LIU You-liang

    2003-01-01

    Effects of calcium on ATPase activities, lipid contents, and fatty acid compositions of plasma membrane from wheat roots were assayed under aluminum stress. The results showed that the increase of calcium concentration in the nutrient solution increased the activity of H+-ATPase and the phospholipid content, decreased the activity of Ca2+-ATPase and the galactolipid of plasma membrane. Owing to the decrease of linolenic acid content, the index of unsaturated fatty acid (IUFA) and index of double bond (DBI) decreased in Altas66. The IUFA and DBI of plasma membrane from Scout66 roots increased because its linolenic acid content increased obviously and its palmitic acid content decreased apparently.

  13. Anandamide reduces intracellular Ca2+ concentration through suppression of Na+/Ca2+ exchanger current in rat cardiac myocytes.

    Directory of Open Access Journals (Sweden)

    Qian Li

    Full Text Available PURPOSE: Anandamide, one of the endocannabinoids, has been reported to exhibit cardioprotective properties, particularly in its ability to limit the damage produced by ischemia reperfusion injury. However, the mechanisms underlying the effect are not well known. This study is to investigate whether anandamide alter Na(+/Ca(2+ exchanger and the intracellular free Ca(2+ concentration ([Ca(2+]i. METHODS: Na(+/Ca(2+ exchanger current (I(NCX was recorded and analysed by using whole-cell patch-clamp technique and [Ca(2+]i was measured by loading myocytes with the fluorescent Ca(2+ indicator Fura-2/AM. RESULTS: We found that I(NCX was enhanced significantly after perfusion with simulated ischemic external solution; [Ca(2+]i was also significantly increased by simulated ischemic solution. The reversal potential of I(NCX was shifted to negative potentials in simulated ischemic external solution. Anandamide (1-100 nM failed to affect I(NCX and [Ca(2+]i in normal solution. However, anandamide (1-100 nM suppressed the increase in INCX in simulated ischemic external solution concentration-dependently and normalized INCX reversal potential. Furthermore, anandamide (100 nM significantly attenuated the increase in [Ca(2+]i in simulated ischemic solution. Blocking CB1 receptors with the specific antagonist AM251 (500 nM failed to affect the effects of anandamide on I(NCX and [Ca(2+]i in simulated ischemic solution. CB2 receptor antagonist AM630 (100 nM eliminated the effects of anandamide on I(NCX and [Ca(2+]i in simulated ischemic solution, and CB2 receptor agonist JWH133 (100 nM simulated the effects of anandamide that suppressed the increase in I(NCX and [Ca(2+]i in simulated ischemic solution. In addition, pretreatment with the Gi/o-specific inhibitor pertussis toxin (PTX, 500 ng/ml eliminated the effects of anandamide and JWH133 on I(NCX in simulated ischemic solution. CONCLUSIONS: Collectively, these findings suggest that anandamide suppresses calcium

  14. Interaction of heavy metals with membrane Ca2+ channels

    Institute of Scientific and Technical Information of China (English)

    PengSQ; HajelRK

    2002-01-01

    The objective of our study was to determine if specific types of high voltage-activated Ca2+ channels,typically found in neurons were affected differentially by MeHg,Hg2+ and Pb2+.Expression cDNA clones of α1C,α1B or α1E subunits coding for neuronal L-,N- and R- subtypes respectively,were combined with α2b δ and β3 Ca2+ channel subunits of human neuronal origin to transfect HEK293 cells.Current was measured using whole cell voltage clamp recording techniques.It the present studies,we conclude: (1)neurotoxic heavy metals such as MeHg,Hg2+ and Pb impair the function of voltage-gated Ca2+ channels at low μmolar to sub-μmolar concentrations-concentrations in the range of which are pathologically and environmentally relevant; (2)a particular metal,i.e.Pb2+,may inhibit function of phenotypically distince Ca2+ channels with variable potency; (3)different metals have differing “orders of potency” at inhibiting defined populations of Ca2+ channels; (4)for “susceptible populations” of patients with either underlying diseases or genetic alter ations of Ca2+ channel function,these metals may have heightened effectiveness.As such,for these populations,environmental toxic metals could produce a more dominant neurotoxicity.

  15. Signal integration by Ca2+ regulates intestinal stem cell activity

    Science.gov (United States)

    Deng, Hansong; Gerencser, Akos A.; Jasper, Heinrich

    2015-01-01

    Summary Somatic stem cells (SCs) maintain tissue homeostasis by dynamically adjusting proliferation and differentiation in response to stress and metabolic cues. Here, we identify Ca2+ signaling as a central regulator of intestinal SC (ISC) activity in Drosophila. We find that dietary L-glutamate stimulates ISC division and gut growth. The metabotropic glutamate receptor (mGluR) is required in ISCs for this response and for an associated modulation of cytosolic Ca2+ oscillations that results in sustained high cytosolic Ca2+ concentrations. High cytosolic Ca2+ induces ISC proliferation by regulating Calcineurin and CREB - regulated transcriptional co-activator (CRTC). In response to a wide range of dietary and stress stimuli, ISCs reversibly transition between Ca2+ oscillation states that represent poised or activated modes of proliferation, respectively. We propose that the dynamic regulation of intracellular Ca2+ levels allows effective integration of diverse mitogenic signals in ISCs to tailor their proliferative activity to the needs of the tissue. PMID:26633624

  16. Glutamate receptor activation in cultured cerebellar granule cells increases cytosolic free Ca2+ by mobilization of cellular Ca2+ and activation of Ca2+ influx

    DEFF Research Database (Denmark)

    Bouchelouche, P; Belhage, B; Frandsen, A;

    1989-01-01

    The Ca2+ sensitive fluorescent probe, fura-2 has been used to monitor cytosolic free calcium levels in mature primary cultures of cerebellar granule cells during exposure to L-glutamate and other excitatory amino acids: quisqualate (QA) kainate (KA) and N-methyl-D-aspartate (NMDA). Glutamate...

  17. NITRIC OXIDE INHIBITS A RISE OF ATP-INTRODUCEDCYTOSOLIC FREE Ca2+ CONCENTRATION AND RELEASE FROM INTRACELLULAR STORED Ca2

    Institute of Scientific and Technical Information of China (English)

    于德洁; 鲍光宏; 王泽君; 邓艳春

    2000-01-01

    Object. The effects of ATP-introduced a rise in cytosolic free Ca2+ concentration and inhibition of nitric oxide were investigated.Method. Measurement of free Ca2+ ([Ca2+]i) of cultured rat tail arterial smooth muscle cells using Fura-2/AM dual excitation wavelength spectrofluorometer.Results. There are two components of [Ca2+]i can be evoked by ATP. One part is Ca2+ entry fixxn Ca2+ channel and formed a plateau. The another is a peak that rdeased from Ca2+ store. Both of them can be inhibited by NO.Conclusion. The ATP induced [ Ca2+ ]i rise that release Ca2+ from both Insp3 and ryanochine receptors and Ca2+ entry through calcium channels. The inhibition of NO on ATP induced [ Ca2+ ]i rise that was mediated by cGMP.

  18. Characterization of Ca2+-Dependent Protein-Protein Interactions within the Ca2+ Release Units of Cardiac Sarcoplasmic Reticulum

    Science.gov (United States)

    Rani, Shilpa; Park, Chang Sik; Sreenivasaiah, Pradeep Kumar; Kim, Do Han

    2016-01-01

    In the heart, excitation-contraction (E-C) coupling is mediated by Ca2+ release from sarcoplasmic reticulum (SR) through the interactions of proteins forming the Ca2+ release unit (CRU). Among them, calsequestrin (CSQ) and histidine-rich Ca2+ binding protein (HRC) are known to bind the charged luminal region of triadin (TRN) and thus directly or indirectly regulate ryanodine receptor 2 (RyR2) activity. However, the mechanisms of CSQ and HRC mediated regulation of RyR2 activity through TRN have remained unclear. We first examined the minimal KEKE motif of TRN involved in the interactions with CSQ2, HRC and RyR2 using TRN deletion mutants and in vitro binding assays. The results showed that CSQ2, HRC and RyR2 share the same KEKE motif region on the distal part of TRN (aa 202–231). Second, in vitro binding assays were conducted to examine the Ca2+ dependence of protein-protein interactions (PPI). The results showed that TRN-HRC interaction had a bell-shaped Ca2+ dependence, which peaked at pCa4, whereas TRN-CSQ2 or TRN-RyR2 interaction did not show such Ca2+ dependence pattern. Third, competitive binding was conducted to examine whether CSQ2, HRC, or RyR2 affects the TRN-HRC or TRN-CSQ2 binding at pCa4. Among them, only CSQ2 or RyR2 competitively inhibited TRN-HRC binding, suggesting that HRC can confer functional refractoriness to CRU, which could be beneficial for reloading of Ca2+ into SR at intermediate Ca2+ concentrations. PMID:26674963

  19. Ivermectin is a nonselective inhibitor of mammalian P-type ATPases.

    Science.gov (United States)

    Pimenta, Paulo Henrique Cotrim; Silva, Claudia Lucia Martins; Noël, François

    2010-02-01

    Ivermectin is a large spectrum antiparasitic drug that is very safe at the doses actually used. However, as it is being studied for new applications that would require higher doses, we should pay attention to its effects at high concentrations. As micromolar concentrations of ivermectin have been reported to inhibit the sarco-endoplasmic reticulum Ca(2+)-ATPase (SERCA), we decided to investigate its putative inhibitory effect on other two important P-type ATPases, namely the Na(+) , K(+)-ATPase and H(+)/K(+)-ATPase. We first extended the data on SERCA, using preparations from rat enriched in SERCA1a (extensor digitorum longus) and 1b (heart) isoforms. Secondly, we tested the effect of ivermectin in two preparations of rat Na(+), K(+)-ATPase in order to appreciate its putative selectivity towards the alpha(1) isoform (kidney) and the alpha(2)/alpha(3) isoforms (brain), and in an H(+)/K(+)-ATPase preparation from rat stomach. Ivermectin inhibited all these ATPases with similar IC(50) values (6-17 microM). With respect to the inhibition of the Na(+), K(+)-ATPase, ivermectin acts by a mechanism different from the classical cardiac glycosides, based on selectivity towards the isoforms, sensibility to the antagonistic effect of K(+) and to ionic conditions favoring different conformations of the enzyme. We conclude that ivermectin is a nonselective inhibitor of three important mammalian P-type ATPases, which is indicative of putative important adverse effects if this drug were used at high doses. As a consequence, we propose that novel analogs of ivermectin should be developed and tested both for their parasitic activity and in vitro effects on P-type ATPases.

  20. Tight coupling of Na+/K+-ATPase with glycolysis demonstrated in permeabilized rat cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Mervi Sepp

    Full Text Available The effective integrated organization of processes in cardiac cells is achieved, in part, by the functional compartmentation of energy transfer processes. Earlier, using permeabilized cardiomyocytes, we demonstrated the existence of tight coupling between some of cardiomyocyte ATPases and glycolysis in rat. In this work, we studied contribution of two membrane ATPases and whether they are coupled to glycolysis--sarcoplasmic reticulum Ca2+ ATPase (SERCA and plasmalemma Na+/K+-ATPase (NKA. While SERCA activity was minor in this preparation in the absence of calcium, major role of NKA was revealed accounting to ∼30% of the total ATPase activity which demonstrates that permeabilized cell preparation can be used to study this pump. To elucidate the contribution of NKA in the pool of ATPases, a series of kinetic measurements was performed in cells where NKA had been inhibited by 2 mM ouabain. In these cells, we recorded: ADP- and ATP-kinetics of respiration, competition for ADP between mitochondria and pyruvate kinase (PK, ADP-kinetics of endogenous PK, and ATP-kinetics of total ATPases. The experimental data was analyzed using a series of mathematical models with varying compartmentation levels. The results show that NKA is tightly coupled to glycolysis with undetectable flux of ATP between mitochondria and NKA. Such tight coupling of NKA to PK is in line with its increased importance in the pathological states of the heart when the substrate preference shifts to glucose.

  1. Demethoxycurcumin Is A Potent Inhibitor of P-Type ATPases from Diverse Kingdoms of Life

    Science.gov (United States)

    Dao, Trong Tuan; Sehgal, Pankaj; Tung, Truong Thanh; Møller, Jesper Vuust; Nielsen, John; Palmgren, Michael; Christensen, Søren Brøgger

    2016-01-01

    P-type ATPases catalyze the active transport of cations and phospholipids across biological membranes. Members of this large family are involved in a range of fundamental cellular processes. To date, a substantial number of P-type ATPase inhibitors have been characterized, some of which are used as drugs. In this work a library of natural compounds was screened and we first identified curcuminoids as plasma membrane H+-ATPases inhibitors in plant and fungal cells. We also found that some of the commercial curcumins contain several curcuminoids. Three of these were purified and, among the curcuminoids, demethoxycurcumin was the most potent inhibitor of all tested P-type ATPases from fungal (Pma1p; H+-ATPase), plant (AHA2; H+-ATPase) and animal (SERCA; Ca2+-ATPase) cells. All three curcuminoids acted as non-competitive antagonist to ATP and hence may bind to a highly conserved allosteric site of these pumps. Future research on biological effects of commercial preparations of curcumin should consider the heterogeneity of the material. PMID:27644036

  2. Mitochondrial Ca(2+) uniporter (MCU)-dependent and MCU-independent Ca(2+) channels coexist in the inner mitochondrial membrane.

    Science.gov (United States)

    Bondarenko, Alexander I; Jean-Quartier, Claire; Parichatikanond, Warisara; Alam, Muhammad Rizwan; Waldeck-Weiermair, Markus; Malli, Roland; Graier, Wolfgang F

    2014-07-01

    A protein referred to as CCDC109A and then renamed to mitochondrial calcium uniporter (MCU) has recently been shown to accomplish mitochondrial Ca(2+) uptake in different cell types. In this study, we investigated whole-mitoplast inward cation currents and single Ca(2+) channel activities in mitoplasts prepared from stable MCU knockdown HeLa cells using the patch-clamp technique. In whole-mitoplast configuration, diminution of MCU considerably reduced inward Ca(2+) and Na(+) currents. This was accompanied by a decrease in occurrence of single channel activity of the intermediate conductance mitochondrial Ca(2+) current (i-MCC). However, ablation of MCU yielded a compensatory 2.3-fold elevation in the occurrence of the extra large conductance mitochondrial Ca(2+) current (xl-MCC), while the occurrence of bursting currents (b-MCC) remained unaltered. These data reveal i-MCC as MCU-dependent current while xl-MCC and b-MCC seem to be rather MCU-independent, thus, pointing to the engagement of at least two molecularly distinct mitochondrial Ca(2+) channels.

  3. Inhibition of ruthenium red-induced Ca2+ efflux from liver mitochondria by the antibiotic X-537A.

    Science.gov (United States)

    Pereira da Silva, L; Bernardes, C F; Vercesi, A E

    1984-10-15

    It has been reported (Becker, G.L., Fiskum, G. and Lehninger, A.L. (1980) J. Biol. Chem. 255, 9009-9012) that respiring rat liver mitochondria suspended in KC1 medium containing ATP, Mg2+ and phosphate, maintain a steady state extramitochondrial free Ca2+ concentration of about 0.5 microM. The results reported here show that the addition of the antibiotic X-537A, at concentrations far below those required for ionophorous activity, caused a perturbation in this steady state, lowering the extramitochondrial free Ca2+ concentration by about 0.20 microM. This shift in steady state was clarified by a study of X-537A inhibition of the Ca2+ efflux induced by ruthenium red; a half-maximum effect was observed at approximately 25 nM X-537A. No effect on Ca2+ transport through the influx uniporter was observed. The possibility of a generalized stabilizing action of the antibiotic on the mitochondrial membrane seems to be ruled out by its effectiveness at very low concentrations. PMID:6208904

  4. Cardiac protective role of a novel erythrocyte-derived depressing factor on rats and its Ca2+ mechanism

    Institute of Scientific and Technical Information of China (English)

    WANG Yutang; WEN Yunyi; MA Ning; SHI Lei

    2003-01-01

    The cardiac protective role of a novel erythrocyte-derived depressing factor (EDDF) on spontaneous hypertensive rats (SHR), calcium overload (CaO) rats and Wistar rats and its mechanism was evaluated. Mean artery pressure (MAP), heart rate (HR) and LVdp/dtmax were measured by physiological recorder. The effect of EDDF on the Ca2+-ATPase activity in myocardial sarcoplasmic reticulum (SR) of CaO rats was determined by inorganic phosphate assay. Calcium transport in myocytes was measured by 45Ca2+ radioactive isotope measurement. The phosphorylation levels of extracellular signal-regulated protein kinases (ERK1/2) in myocardial tissue of SHR and CaO rats were measured by Western blot method. And the ultrastructures of cardiac muscle cells were observed with the transmission electron microscope. The results indicated that EDDF could significantly decrease MAP, HR and LVdp/dtmax in a dose dependent manner (P < 0.05). It seems that the mechanism might relate with activating the Ca2+-APTase, enhancing the uptake and release of Ca2+ from SR (P < 0.05), decreasing the phosphorylation levels of ERK1/2 of myocytes (P < 0.01) and lightening the ultrastructural lesion of cardiac muscle cells. In CaO rats, the Ca2+-ATPase activity decreased clearly compared to control (64.99 ± 7.16 vs 94.48 ± 7.68 nmol·min-1·mg-1 protein, P < 0.01), while EDDF (100 μg/mL) could significantly increase the activity (87.93 ± 9.54 vs 64.99 ± 7.16, P < 0.05, n = 7). Both uptake and release rate of Ca2+ (μmol 45Ca2+/g protein/min) from myocardial SR of CaO rats remarkably decreased compared to control (32.40 ± 2.70 and 15.46 ± 1.49 vs 61.09 ± 10.89 and 25.47 ± 4.29, P < 0.05); EDDF (100 μg/mL) could significantly stimulate their activities (50.48 ± 6.76 and 21.76 ± 2.75 vs 32.40 ± 2.70 and 15.46 ± 1.49, P < 0.05). EDDF could evidently down-regulate the phosphorylation of ERK1/2 in myocardial tissue from SHR and CaO rats (P < 0.01), lighten the ultrastructural lesion of cardiac muscle

  5. The polycystin complex mediates Wnt/Ca(2+) signalling.

    Science.gov (United States)

    Kim, Seokho; Nie, Hongguang; Nesin, Vasyl; Tran, Uyen; Outeda, Patricia; Bai, Chang-Xi; Keeling, Jacob; Maskey, Dipak; Watnick, Terry; Wessely, Oliver; Tsiokas, Leonidas

    2016-07-01

    WNT ligands induce Ca(2+) signalling on target cells. PKD1 (polycystin 1) is considered an orphan, atypical G-protein-coupled receptor complexed with TRPP2 (polycystin 2 or PKD2), a Ca(2+)-permeable ion channel. Inactivating mutations in their genes cause autosomal dominant polycystic kidney disease (ADPKD), one of the most common genetic diseases. Here, we show that WNTs bind to the extracellular domain of PKD1 and induce whole-cell currents and Ca(2+) influx dependent on TRPP2. Pathogenic PKD1 or PKD2 mutations that abrogate complex formation, compromise cell surface expression of PKD1, or reduce TRPP2 channel activity suppress activation by WNTs. Pkd2(-/-) fibroblasts lack WNT-induced Ca(2+) currents and are unable to polarize during directed cell migration. In Xenopus embryos, pkd1, Dishevelled 2 (dvl2) and wnt9a act within the same pathway to preserve normal tubulogenesis. These data define PKD1 as a WNT (co)receptor and implicate defective WNT/Ca(2+) signalling as one of the causes of ADPKD. PMID:27214281

  6. Delta(9)-tetrahydrocannabinol activates [Ca2+], increases partly sensitive to capacitative store refilling

    NARCIS (Netherlands)

    Filipeanu, CM; deZeeuw, D; Nelemans, SA

    1997-01-01

    Delta(9)-Tetrahydrocannabinol induces [Ca2+](i) increases in DDT1MF-2 smooth muscle cells. Both Ca2+ entry and release from intracellular Ca2+ stores were concentration dependently activated. The Ca2+ entry component contributed most to the increases in [Ca2+](i). Stimulation with Delta(9)-tetrahydr

  7. Calbindin-D28K dynamically controls TRPV5-mediated Ca2+ transport.

    NARCIS (Netherlands)

    Lambers, T.T.; Mahieu, F.; Oancea, E.; Hoofd, L.J.C.; Lange, F. de; Mensenkamp, A.R.; Voets, T.; Nilius, B.; Clapham, D.E.; Hoenderop, J.G.J.; Bindels, R.J.M.

    2006-01-01

    In Ca(2+)-transporting epithelia, calbindin-D(28K) (CaBP(28K)) facilitates Ca(2+) diffusion from the luminal Ca(2+) entry side of the cell to the basolateral side, where Ca(2+) is extruded into the extracellular compartment. Simultaneously, CaBP(28K) provides protection against toxic high Ca(2+) lev

  8. Caffeine-induced Ca2+ transients and exocytosis in Paramecium cells. A correlated Ca2+ imaging and quenched-flow/freeze-fracture analysis.

    Science.gov (United States)

    Klauke, N; Plattner, H

    1998-01-01

    Caffeine causes a [Ca2+]i increase in the cortex of Paramecium cells, followed by spillover with considerable attenuation, into central cell regions. From [Ca2+]resti approximately 50 to 80 nm, [Ca2+]acti rises within /=2 sec. Chelation of Ca2+o considerably attenuated [Ca2+]i increase. Therefore, caffeine may primarily mobilize cortical Ca2+ pools, superimposed by Ca2+ influx and spillover (particularly in tl cells with empty trichocyst docking sites). In nd cells, caffeine caused trichocyst contents to decondense internally (Ca2+-dependent stretching, normally occurring only after membrane fusion). With 7S cells this usually occurred only to a small extent, but with increasing frequency as [Ca2+]i signals were reduced by [Ca2+]o chelation. In this case, quenched-flow and ultrathin section or freeze-fracture analysis revealed dispersal of membrane components (without fusion) subsequent to internal contents decondensation, opposite to normal membrane fusion when a full [Ca2+]i signal was generated by caffeine stimulation (with Ca2+i and Ca2+o available). We conclude the following. (i) Caffeine can mobilize Ca2+ from cortical stores independent of the presence of Ca2+o. (ii) To yield adequate signals for normal exocytosis, Ca2+ release and Ca2+ influx both have to occur during caffeine stimulation. (iii) Insufficient [Ca2+]i increase entails caffeine-mediated access of Ca2+ to the secretory contents, thus causing their decondensation before membrane fusion can occur. (iv) Trichocyst decondensation in turn gives a signal for an unusual dissociation of docking/fusion components at the cell membrane. These observations imply different threshold [Ca2+]i-values for membrane fusion and contents discharge.

  9. Curcumin modulation of Na,K-ATPase: phosphoenzyme accumulation, decreased K+ occlusion, and inhibition of hydrolytic activity

    DEFF Research Database (Denmark)

    Mahmmoud, Yasser Ahmed

    2005-01-01

    Curcumin, the major constitute of tumeric, is an important nutraceutical that has been shown to be useful in the treatment of many diseases. As an inhibitor of the sarcoplasmic reticulum Ca2+-ATPase, curcumin was shown to correct cystic fibrosis (CF) defects in some model systems, whereas others...

  10. Ca2+ signaling in pancreatic acinar cells: physiology and pathophysiology

    Directory of Open Access Journals (Sweden)

    O.H. Petersen

    2009-01-01

    Full Text Available The pancreatic acinar cell is a classical model for studies of secretion and signal transduction mechanisms. Because of the extensive endoplasmic reticulum and the large granular compartment, it has been possible - by direct measurements - to obtain considerable insights into intracellular Ca2+ handling under both normal and pathological conditions. Recent studies have also revealed important characteristics of stimulus-secretion coupling mechanisms in isolated human pancreatic acinar cells. The acinar cells are potentially dangerous because of the high intra-granular concentration of proteases, which become inappropriately activated in the human disease acute pancreatitis. This disease is due to toxic Ca2+ signals generated by excessive liberation of Ca2+ from both the endoplasmic reticulum and the secretory granules.

  11. Scallop DMT functions as a Ca2+ transporter.

    Science.gov (United States)

    Toyohara, Haruhiko; Yamamoto, Sayuri; Hosoi, Masatomi; Takagi, Masaya; Hayashi, Isao; Nakao, Kenji; Kaneko, Shuji

    2005-05-01

    We identified a DMT (divalent metal transporter) homologous protein that functions as a Ca(2+) transporter. Scallop DMT cDNA encodes a 539-amino-acid protein with 12 putative membrane-spanning domains and has a consensus transport motif in the fourth extracellular loop. Since its mRNA is significantly expressed in the gill and intestine, it is assumed that scallop DMT transports Ca(2+) from seawater by the gill and from food by the intestine. Scallop DMT lacks the iron-responsive element commonly found in iron-regulatory proteins, suggesting that it is free of the post-transcriptional regulation from intracellular Fe(2+) concentration. Scallop DMT distinctly functions as a Ca(2+) transporter unlike other DMTs, however, it also transports Fe(2+) and Cd(2+) similar to them.

  12. Hippocampal Area CA2: An Overlooked but Promising Therapeutic Target.

    Science.gov (United States)

    Chevaleyre, Vivien; Piskorowski, Rebecca A

    2016-08-01

    While the hippocampus has long been recognized as a brain structure specialized in mapping 'space' in rodents, human studies and now recent data from rodents have shown that its function extends well beyond spatial coding. Recently, an overlooked area of the hippocampus, CA2, has emerged as a critical region for social memory. This area is also uniquely altered during several pathologies such as schizophrenia and age-related dementia. Because of its singular connectivity, we propose that area CA2 resides at the interface between emotional brain activity and higher cognitive function. Furthermore, because of the unique expression of multiple neuromodulator receptors in area CA2, we posit that this region may represent a fruitful therapeutic target for diseases where social dysfunction occurs. PMID:27372610

  13. Annexins and Ca2+ handling in the heart.

    Science.gov (United States)

    Camors, Emmanuel; Monceau, Virginie; Charlemagne, Daniéle

    2005-03-01

    Annexins are a family of 13 proteins known to bind phospholipids (PL) in a Ca(2+)-dependent way. They are ubiquitous proteins and share a similar structure characterized by a conserved C-terminal domain with Ca(2+) binding sites and a variable N-terminal domain. Depending on Ca(2+) concentration, they have been reported to participate in a variety of membrane-related events such as exocytosis, endocytosis, apoptosis and binding to cytoskeletal proteins. They have also been reported to regulate protein activities. This review will focus on annexins in the heart, and particularly on annexins A2, A5, A6 and A7. Annexin A2 has been found in endothelial cells and reported to play a central role in control of plasmin-mediated processes. Annexin A5 is mainly localized in cardiomyocytes. However, it could be relocated to interstitial tissue in ischemic and failing hearts or it could be externalized and exhibit a proapoptotic effect in cardiomyocytes. Annexin A6 is the most abundant annexin in the heart, and has been localized in various cell types including myocytes. Overexpression of annexin A6 has underlined physiological alterations in contractile mechanics leading to dilated cardiomyopathy, whereas knockout has been found to induce faster changes in Ca(2+) transient and increased contractility, suggesting a negative inotropic role for annexin A6. Annexin A7 is expressed in heart and skeletal muscle. In annexin A7 null mutant mice decreases in the force-frequency relationship were observed in adult cardiomyocytes, consistent with regulation of Ca(2+) handling. In conclusion, while annexin A2 was involved in regulation of fibrin homeostasis, alterations in expression and activity of annexins A5, A6 and A7 have been associated with regulation of Ca(2+) handling in the heart, but the target of each annexin has not yet been identified. PMID:15721859

  14. Meiosis, egg activation, and nuclear envelope breakdown are differentially reliant on Ca2+, whereas germinal vesicle breakdown is Ca2+ independent in the mouse oocyte

    Science.gov (United States)

    Tombes, R. M.; Simerly, C.; Borisy, G. G.; Schatten, G.

    1992-01-01

    During early development, intracellular Ca2+ mobilization is not only essential for fertilization, but has also been implicated during other meiotic and mitotic events, such as germinal vesicle breakdown (GVBD) and nuclear envelope breakdown (NEBD). In this study, the roles of intracellular and extracellular Ca2+ were examined during meiotic maturation and reinitiation at parthenogenetic activation and during first mitosis in a single species using the same methodologies. Cumulus-free metaphase II mouse oocytes immediately resumed anaphase upon the induction of a large, transient Ca2+ elevation. This resumption of meiosis and associated events, such as cortical granule discharge, were not sensitive to extracellular Ca2+ removal, but were blocked by intracellular Ca2+ chelators. In contrast, meiosis I was dependent on external Ca2+; in its absence, the formation and function of the first meiotic spindle was delayed, the first polar body did not form and an interphase-like state was induced. GVBD was not dependent on external Ca2+ and showed no associated Ca2+ changes. NEBD at first mitosis in fertilized eggs, on the other hand, was frequently, but not always associated with a brief Ca2+ transient and was dependent on Ca2+ mobilization. We conclude that GVBD is Ca2+ independent, but that the dependence of NEBD on Ca2+ suggests regulation by more than one pathway. As cells develop from Ca(2+)-independent germinal vesicle oocytes to internal Ca(2+)-dependent pronuclear eggs, internal Ca2+ pools increase by approximately fourfold.

  15. Calcineurin homologous protein: a multifunctional Ca2+-binding protein family

    OpenAIRE

    Di Sole, Francesca; Vadnagara, Komal; MOE, ORSON W.; Babich, Victor

    2012-01-01

    The calcineurin homologous protein (CHP) belongs to an evolutionarily conserved Ca2+-binding protein subfamily. The CHP subfamily is composed of CHP1, CHP2, and CHP3, which in vertebrates share significant homology at the protein level with each other and between other Ca2+-binding proteins. The CHP structure consists of two globular domains containing from one to four EF-hand structural motifs (calcium-binding regions composed of two helixes, E and F, joined by a loop), the myristoylation, a...

  16. Effects of Time Delay on Intracellular Ca2+ Concentration Oscillations

    Institute of Scientific and Technical Information of China (English)

    YING Yang-Jun; HUANG Zu-Qia

    2001-01-01

    Based on the SS-model [Somogyi R and Stucki J W J. Biol. Chem. 266 (1991) 11 068] for the generation of intracellular Ca2+ concentration oscillations, we consider a time delay for the binding kinetics of the Ca2+ channel and find a significant phenomenon that the oscillation takes two quite different modes when a parameter of the system crosses a threshold. One is a quick oscillation mode and the other is a slow oscillation mode. The oscillation frequencies of these modes differ from each other by more than ten times. The change of oscillation form with parameters and its critical behaviour are illustrated by numerical simulation results.

  17. Enhancing the potency of lithospermate B for inhibiting Na+/K+-ATPase activity by forming transition metal ion complexes

    OpenAIRE

    Lin, Nan-Hei; Chung, Tse-yu; Li, Feng-yin; Chen, Hsin-An; Tzen, Jason TC

    2013-01-01

    Aim: To determine whether replacing Mg2+ in magnesium lithospermate B (Mg-LSB) isolated from danshen (Salvia miltiorrhiza) with other metal ions could affect its potency in inhibition of Na+/K+-ATPase activity. Methods: Eight metal ions (Na+, K+, Mg2+, Cr3+, Mn2+, Co2+, Ni2+, and Zn2+) were used to form complexes with LSB. The activity of Na+/K+-ATPase was determined by measuring the amount of inorganic phosphate (Pi) liberated from ATP. Human adrenergic neuroblastoma cell line SH-SY5Y was us...

  18. Ionophore 4-BrA23187 transports Zn2+ and Mn2+ with high selectivity over Ca2+.

    Science.gov (United States)

    Erdahl, W L; Chapman, C J; Wang, E; Taylor, R W; Pfeiffer, D R

    1996-10-29

    The cation transport selectivities of the Ca2+ ionophores A23187, Ionomycin, and 4-BrA23187 have been determined using a model system comprised of phospholipid vesicles loaded with the chelator/indicator Quin-2. At pH 7.00 and a 100 microM concentration of the cations, A23187 displays the transport selectivity sequence Zn2+ > Mn2+ > Ca2+ > Co2+ > Ni2+ > Sr2+, with the absolute rates of transport spanning approximately 3 orders of magnitude. Similar data are obtained with Ionomycin, although the relative transport rates of Zn2+ and Mn2+ are equivalent, and the range of absolute rates is decreased by a factor of approximately 3. When values are normalized to those of Ca2+, transport selectivity is seen to be only weakly related to complexation or extraction selectivity. It is also seen that, when used to manipulate Ca2+ (or Mg2+), both ionophores can be expected to alter the distribution of additional divalent cations which have known biological activities. 4-BrA23187 is a low-activity ionophore for Ca2+, compared to A23187 and Ionomycin, while retaining comparable activities as an ionophore for the other cations. As a consequence, 4-BrA23187 is highly selective for the transport of Zn2+ and Mn2+, compared to Ca2+, with selectivity ratios approaching that of valinomycin for K+ over Na+ when conditions are optimal. Plots of the log of the rate of cation transport vs the log of the ionophore concentration indicate that Ca2+ is transported primarily as a 2:1 complex by A23187 and 4-BrA23187, but Zn2+ and Mn2+ are transported, in part, as 1:1 complexes. These findings, together with a postulated low stability of 2:1, compared to 1:1 complexes between 4-BrA23187 and divalent cations, partially explain the novel transport selectivity of this compound. Unlike A23187 or Ionomycin, 4-BrA23187 may be useful for investigating cell regulation by Zn2+ and Mn2+, without interference by regulatory mechanisms which respond to Ca2+. PMID:8901524

  19. CGP37157, an inhibitor of the mitochondrial Na+/Ca2+ exchanger, protects neurons from excitotoxicity by blocking voltage-gated Ca2+ channels.

    Science.gov (United States)

    Ruiz, A; Alberdi, E; Matute, C

    2014-04-10

    Inhibition of the mitochondrial Na(+)/Ca(2+) exchanger (NCLX) by CGP37157 is protective in models of neuronal injury that involve disruption of intracellular Ca(2+) homeostasis. However, the Ca(2+) signaling pathways and stores underlying neuroprotection by that inhibitor are not well defined. In the present study, we analyzed how intracellular Ca(2+) levels are modulated by CGP37157 (10 μM) during NMDA insults in primary cultures of rat cortical neurons. We initially assessed the presence of NCLX in mitochondria of cultured neurons by immunolabeling, and subsequently, we analyzed the effects of CGP37157 on neuronal Ca(2+) homeostasis using cameleon-based mitochondrial Ca(2+) and cytosolic Ca(2+) ([Ca(2+)]i) live imaging. We observed that NCLX-driven mitochondrial Ca(2+) exchange occurs in cortical neurons under basal conditions as CGP37157 induced a decrease in [Ca(2)]i concomitant with a Ca(2+) accumulation inside the mitochondria. In turn, CGP37157 also inhibited mitochondrial Ca(2+) efflux after the stimulation of acetylcholine receptors. In contrast, CGP37157 strongly prevented depolarization-induced [Ca(2+)]i increase by blocking voltage-gated Ca(2+) channels (VGCCs), whereas it did not induce depletion of ER Ca(2+) stores. Moreover, mitochondrial Ca(2+) overload was reduced as a consequence of diminished Ca(2+) entry through VGCCs. The decrease in cytosolic and mitochondrial Ca(2+) overload by CGP37157 resulted in a reduction of excitotoxic mitochondrial damage, characterized here by a reduction in mitochondrial membrane depolarization, oxidative stress and calpain activation. In summary, our results provide evidence that during excitotoxicity CGP37157 modulates cytosolic and mitochondrial Ca(2+) dynamics that leads to attenuation of NMDA-induced mitochondrial dysfunction and neuronal cell death by blocking VGCCs.

  20. Cinética de absorção e eficiência nutricional de K+, Ca2+ e Mg2+ em plantas jovens de quatro clones de eucalipto Uptake kinetics and nutritional efficiency for K+, Ca2+ and Mg2+ in four eucalypt clones seedlings

    OpenAIRE

    Augusto Miguel Nascimento Lima; Júlio César Lima Neves; Ivo Ribeiro Silva; Fernando Palha Leite

    2005-01-01

    Modelos mecanísticos baseados em princípios de transporte de solutos pode ser de grande utilidade para prever os impactos do cultivo de florestas plantadas, por exemplo, com eucalipto sobre o capital e fluxo de nutrientes no solo. Dentre as variáveis de entrada demandadas por tais modelos, tem-se os valores das constantes Vmax, Km e Cmin da cinética de absorção iônica. Assim, os objetivos do presente trabalho foram determinar os valores de Vmax, Km e Cmin para K, Ca e Mg, bem como avaliar as ...

  1. 酸性条件下剩余污泥中Ca2+和Mg2+溶出对磷回收的影响%Impacts of Ca2+ and Mg2+ release on phosphorus recovery in excess sludge under acidic condition

    Institute of Scientific and Technical Information of China (English)

    苑宏英; 员建; 陈轶; 吴丽杰

    2011-01-01

    为了在酸性条件下实现剩余污泥中磷的高效回收,对pH=3时剩余污泥水解酸化过程中氨氮、正磷酸盐和钙镁离子的溶出现象以及磷回收进行了研究分析。结果表明:当pH=3时,所溶出的氨氮、镁离子和钙离子与磷酸盐的摩尔比均大于1,能满足采用鸟粪石沉淀法或者羟磷灰石沉淀法回收磷的要求;但所溶出的钙镁离子的摩尔比大于1,会对鸟粪石沉淀法回收磷的顺利进行有较大影响;有无外加镁剂对磷回收率影响不大。采用改型后的镁型强酸性阳离子交换树脂进行离子交换可以得到较高纯度的鸟粪石沉淀产品,通过XRD检测其纯度为95%以上。%To study the impact of release of calcium and magnesium ions in excess sludge under acidic con- dition on phosphorus recovery, release process of ammonia-nitrogen, phosphate and calcium and magnesium ions at pH = 3 and phosphorus recovery process were discussed. The results shows that at pH = 3, all the molar ratios of ammonia, magnesium and calcium ions dissolved to phosphate were greater than 1, which could meet the re- quirements of phosphorus recovery by means of struvite precipitation or hydroxyapatite precipitation. Molar ratio of calcium and magnesium ions dissolved was also greater than 1, which would have a serious influence on phos- phorus recovery by the method of struvite precipitation. Whether additional magnesium existed or not had little effect on the percent recovery of phosphorus. It is feasible to choose strongly acidic cation exchange resin modi- fied with magnesium to ion exchange for improving the quality of the precipitation products. It is determined that precipitation products contain more than 95% of struvite by chemical analysis and XRD detection.

  2. Cinética de absorção e eficiência nutricional de K+, Ca2+ e Mg2+ em plantas jovens de quatro clones de eucalipto Uptake kinetics and nutritional efficiency for K+, Ca2+ and Mg2+ in four eucalypt clones seedlings

    Directory of Open Access Journals (Sweden)

    Augusto Miguel Nascimento Lima

    2005-12-01

    Full Text Available Modelos mecanísticos baseados em princípios de transporte de solutos pode ser de grande utilidade para prever os impactos do cultivo de florestas plantadas, por exemplo, com eucalipto sobre o capital e fluxo de nutrientes no solo. Dentre as variáveis de entrada demandadas por tais modelos, tem-se os valores das constantes Vmax, Km e Cmin da cinética de absorção iônica. Assim, os objetivos do presente trabalho foram determinar os valores de Vmax, Km e Cmin para K, Ca e Mg, bem como avaliar as respectivas eficiências nutricionais de clones de eucalipto. O estudo consistiu de três ensaios em solução nutritiva (um para cada cátion, sendo utilizadas mudas propagadas vegetativamente de um híbrido de E. grandis x E. urophylla (clone 1213 e de três híbridos de E. grandis (clones 7074, 57 e 129. Com base nos teores de cada um desses nutrientes nas soluções de depleção, em cada tempo de amostragem, no volume inicial e final de solução nos vasos e no peso de matéria fresca de raízes, foram obtidos os valores das constantes cinéticas. Para K, o clone 7074 apresentou o menor valor de Vmax em relação aos demais clones, os quais não diferiram entre si, e em relação ao Km e Cmin, os clones não diferiram estatisticamente. Para Ca, os clones estudados diferiram quanto ao valor de Vmax e Km, não diferindo, entretanto, para o Cmin. Os menores valores de Km para Mg foram verificados para os clones 57 e 7074, ou seja, as proteínas transportadoras de Mg na membrana plasmática das células radiculares apresentaram maior afinidade para esse nutriente. Contudo, os valores de Vmax e Cmin não diferiram entre os clones estudados. Diferenças na eficiência nutricional dos clones estudados quanto a K e a Ca foram devidas às diferenças na eficiência de absorção, e para Mg às diferenças na eficiência de absorção e de utilização.The use of mechanistic models based on principles of solute transport may be of great utility to estimate, for example, the impact of eucalypt forest cultivation on the pools and fluxes of nutrient in soils. Among the parameters required by such models are the values of the ion absorption kinetics constants Vmax, Km and Cmin. Thus, the objective of this study was to determine the values of Vmax, Km and Cmin for K, Ca and Mg, as well as to evaluate the nutritional efficiency of eucalypt clones to these nutrients. The present study consisted of three experiments in solution culture (one for each cation. It was employed four genetic materials (clones 7074, 57, 1213 and 129 originated from clonal propagation. Clone 1213 is an Eucalyptus grandis x Eucalyptus urophylla hybrid, whereas the others are natural hybrids of E. grandis. The concentration of each nutrient being studied in the depletion solution at specific sampling times, initial and final pot solution volume and fresh root weight were used in order to obtain the value of the kinetics constants. For K, clone 7074 showed the lowest Vmax value in relation to the other clones, which did not differ among themselves. All clones did not differ statistically in relation to Km and Cmin for K. For Ca, the studied clones differed in relation to the values of Vmax and Km but, they did not differ for Cmin. The lower values of Km for Mg were observed for the clones 57 and 7074, that is, the carrier proteins at the root cells plasma membrane have a high affinity for this nutrient. Nevertheless, the values of Vmax and Cmin did not differ among the studied clones. Differences in nutritional efficiency of the studied clones in relation to K and Ca were due to differences in the absorption efficiency and for Mg were related with differences in the nutrient use efficiency and absorption efficiency.

  3. Modeling effects of L-type Ca2+ current and Na+-Ca2+ exchanger on Ca2+ trigger flux in rabbit myocytes with realistic t-tubule geometries

    Directory of Open Access Journals (Sweden)

    Peter M Kekenes-Huskey

    2012-09-01

    Full Text Available The transverse tubular system of rabbit ventricular myocytes consists of cell membrane invaginations (t-tubules that are essential for efficient cardiac excitation-contraction coupling. In this study, we investigate how t-tubule micro-anatomy, L-type Ca2+ channel clustering and allosteric activation of Na+/Ca2+ exchanger by L-type Ca2+ current affects intracellular Ca2+ dynamics. Our model includes a realistic 3D geometry of a single t-tubule and its surrounding half-sarcomeres for rabbit ventricular myocytes. The effects of spatially-distributed membrane ion-transporters (L-type Ca2+ channel, Na+/Ca2+ exchanger, sarcolemmal Ca2+ pump, sarcolemmal Ca2+ leak, and stationary and mobile Ca2+ buffers (troponin C, ATP, calmodulin, and Fluo-3 are also considered. We used a coupled reaction-diffusion system to describe the spatio-temporal concentration profiles of free and buffered intracellular Ca2+. We obtained parameters from voltage-clamp protocols of L-type Ca2+ current and line-scan recordings of Ca2+ concentration profiles in rabbit cells, in which the sarcoplasmic reticulum is disabled. Our model results agree with experimental measurements of global Ca2+ transient in myocytes loaded with 50 µM Fluo-3. We found that local Ca2+ concentrations within the cytosol and sub-sarcolemma, as well as the local trigger fluxes of Ca2+ crossing the cell membrane, are sensitive to details of t-tubule micro-structure and membrane Ca2+ flux distribution. The model additionally predicts that local Ca2+ trigger fluxes are at least 3-fold to 8-fold higher than the whole-cell Ca2+ trigger flux. We found also that the activation of allosteric Ca2+-binding sites on the Na+/Ca2+ exchanger could provide a mechanism for regulating global and local Ca2+ trigger fluxes in vivo. Our studies indicate that improved structural and functional models could improve our understanding of the contributions of L-type and Na+/Ca2+ exchanger fluxes to intracellular Ca2+ dynamics.

  4. Alteration of complex sphingolipid composition and its physiological significance in yeast Saccharomyces cerevisiae lacking vacuolar ATPase.

    Science.gov (United States)

    Tani, Motohiro; Toume, Moeko

    2015-12-01

    In the yeast Saccharomyces cerevisiae, complex sphingolipids have three types of polar head group and five types of ceramide; however, the physiological significance of the structural diversity is not fully understood. Here, we report that deletion of vacuolar H+-ATPase (V-ATPase) in yeast causes dramatic alteration of the complex sphingolipid composition, which includes decreases in hydroxylation at the C-4 position of long-chain bases and the C-2 position of fatty acids in the ceramide moiety, decreases in inositol phosphorylceramide (IPC) levels, and increases in mannosylinositol phosphorylceramide (MIPC) and mannosyldiinositol phosphorylceramide [M(IP)2C] levels. V-ATPase-deleted cells exhibited slow growth at pH 7.2, whereas the increase in MIPC levels was significantly enhanced when V-ATPase-deleted cells were incubated at pH 7.2. The protein expression levels of MIPC and M(IP)2C synthases were significantly increased in V-ATPase-deleted cells incubated at pH 7.2. Loss of MIPC synthesis or an increase in the hydroxylation level of the ceramide moiety of sphingolipids on overexpression of Scs7 and Sur2 sphingolipid hydroxylases enhanced the growth defect of V-ATPase-deleted cells at pH 7.2. On the contrary, the growth rate of V-ATPase-deleted cells was moderately increased on the deletion of SCS7 and SUR2. In addition, supersensitivities to Ca2+, Zn2+ and H2O2, which are typical phenotypes of V-ATPase-deleted cells, were enhanced by the loss of MIPC synthesis. These results indicate the possibility that alteration of the complex sphingolipid composition is an adaptation mechanism for a defect of V-ATPase.

  5. Computational approaches for classification and prediction of P-type ATPase substrate specificity in Arabidopsis.

    Science.gov (United States)

    Zinati, Zahra; Alemzadeh, Abbas; KayvanJoo, Amir Hossein

    2016-01-01

    As an extended gamut of integral membrane (extrinsic) proteins, and based on their transporting specificities, P-type ATPases include five subfamilies in Arabidopsis, inter alia, P4ATPases (phospholipid-transporting ATPase), P3AATPases (plasma membrane H(+) pumps), P2A and P2BATPases (Ca(2+) pumps) and P1B ATPases (heavy metal pumps). Although, many different computational methods have been developed to predict substrate specificity of unknown proteins, further investigation needs to improve the efficiency and performance of the predicators. In this study, various attribute weighting and supervised clustering algorithms were employed to identify the main amino acid composition attributes, which can influence the substrate specificity of ATPase pumps, classify protein pumps and predict the substrate specificity of uncharacterized ATPase pumps. The results of this study indicate that both non-reduced coefficients pertaining to absorption and Cys extinction within 280 nm, the frequencies of hydrogen, Ala, Val, carbon, hydrophilic residues, the counts of Val, Asn, Ser, Arg, Phe, Tyr, hydrophilic residues, Phe-Phe, Ala-Ile, Phe-Leu, Val-Ala and length are specified as the most important amino acid attributes through applying the whole attribute weighting models. Here, learning algorithms engineered in a predictive machine (Naive Bays) is proposed to foresee the Q9LVV1 and O22180 substrate specificities (P-type ATPase like proteins) with 100 % prediction confidence. For the first time, our analysis demonstrated promising application of bioinformatics algorithms in classifying ATPases pumps. Moreover, we suggest the predictive systems that can assist towards the prediction of the substrate specificity of any new ATPase pumps with the maximum possible prediction confidence. PMID:27186030

  6. NITRIC OXIDE INHIBITS A RISE OF ATP-INTRODUCED CYTOSOLIC FREE Ca2 + CONCENTRATION AND RELEASE FROM INTRACELLULAR STORED Ca2+

    Institute of Scientific and Technical Information of China (English)

    王泽君; 邓艳春; 于德洁; 鲍光宏

    2000-01-01

    Object. The effects of ATP-introduced a rise in cytosolic free Ca2 + concentration and inhibition of nitric oxide were investigated. Method. Measurement of f ree Ca2+ ([Ca2+ ]i)of cultured rat tail arterial smooth muscle cells using Fura-2/AM dual excitation wavelength spoctrofluorometer. Results. There are two components of [ Ca2 + ]i can be evoked by ATP. One part is Ca2 + ertry from Ca2 + channel and formed a plateau. The another part is a peak that released f rom Ca2 + store. Both of them can be inhibited by NO. Conclusion. The ATP induced [ Ga2 + ]i rise that release Ca2 + from both InsP3 and ryanochine receptors and Ga2 + entry through calcium channels. The inhibition of NO on ATP induced [ Ca2 + ]i rise that was mediated by cGMP.

  7. Ca2+Entry Through TRPC1 Channels Contributes to Intraceilular Ca2+ Dynamics and Consequent Glutamate Release from Rat Astrocytes

    Institute of Scientific and Technical Information of China (English)

    ERIK B. MALARKEY; YINGCHUN NI; VLADIMIR PARPURA

    2008-01-01

    Astrocytes can respond to a variety of stimuli by elevating their cytoplasmic Ca2+ concentration and can in turn release glutamate to signal adjacent neurons. The majority of this Ca2+ is derived from internal stores while a portion also comes from outside of the cell. Astrocytes use Ca2+ entry through store-operated Ca channels to refill their internal stores. Therefore, we investigated what role this store-operated Ca2+ entry plays in astrocyticCa2+ responses and subsequent glutamate release. Astrocytes express canonical transient receptor potential (TRPC)channels that have been implicated in mediating store-operated Ca2+ entry. Here, we show that astroeytes in culture and freshly isolated as trocytes from visual cortex express TRPC1, TRPC4, and TRPCS. Indirect immunocy to-chemistry reveals that these proteins are present throughout the cell; the predominant expression of functionally test-ed TRPC1, however, is on the plasma membrane. Labeling in freshly isolated astrocytes reveals changes in TRP Cexpression throughout development. Using an antibody against TRPC1 we were able to block the function of TR-PC1 channels and determine their involvement in mechanically and agonist-evoked Ca2+ entry in cultured astrocytes.Blocking TRPC1 was also found to reduce mechanically induced Ca2+-dependent glutamate release. These data indi-cate that Ca2+entry through TRPC1 channels contributes to Ca2+signaling in astrocytes and the consequent gluta-mate release from these cells.%各种不同的刺激作用于星型胶质细胞,可以导致胞浆内Ca2+浓度增加,进而释放更多谷氨酸作用于周边的神经元.大部分Ca2+来源于细胞内,小部分来源于细胞外.Ca2+内流是通过钙池操纵Ca2+通道(SOC)实现的.因此,作者观察在星型胶质细胞内Ca2+激活与谷氨酸释放过程中钙池操纵Ca2+通道(SOC)发挥了什么样的作用.已有研究显示星型胶质细胞所表达的TRPC通道(Ca2+通过瞬时受体电位通道相关蛋白)介导了钙池操纵Ca

  8. Wnt/Ca2+ signaling pathway: a brief overview

    Institute of Scientific and Technical Information of China (English)

    Antara De

    2011-01-01

    The non-canonical Wnt/Ca2+ signaling cascade is less characterized than their canonical counterpart,the Wnt/β-catenin pathway.The non-canonical Wnt signaling pathways are diverse,defined as planer cell polarity pathway,Wnt-RAP1 signaling pathway,Wnt-Ror2 signaling pathway,Wnt-PKA pathway,Wnt-GSK3MT pathway,Wnt-aPKC pathway,Wnt-RYK pathway,Wnt-mTOR pathway,and Wnt/calcium signaling pathway.All these pathways exhibit a considerable degree of overlap between them.The Wnt/Ca2+ signaling pathway was deciphered as a crucial mediator in development.However,now there is substantial evidence that the signaling cascade is involved in many other molecular phenomena.Many aspects of Wnt/Ca2+ pathway are yet enigmatic.This review will give a brief overview of the fundamental and evolving concepts of the Wnt/Ca2+ signaling pathway.

  9. Thalamic T-type Ca2+ channels and NREM sleep

    OpenAIRE

    Crunelli, Vincenzo; Cope, David W.; Hughes, Stuart W.

    2006-01-01

    T-type Ca2+ channels play a number of different and pivotal roles in almost every type of neuronal oscillation expressed by thalamic neurones during non-Rapid Eye Movement (NREM) sleep, including those underlying sleep theta waves, the K-complex and the slow (

  10. Plasmalemmal and mitochondrial Na(+) -Ca(2+) exchange in neuroglia.

    Science.gov (United States)

    Parpura, Vladimir; Sekler, Israel; Fern, Robert

    2016-10-01

    In the absence of the electrical signaling for which neurons are so highly specialized, GLIA rely on the slow propagation of ionic signals to mediate network events such as Ca(2+) and Na(+) waves. Glia differ from neurons in another important way, they are replete with a high density of ionic-transport proteins that are essential for them to fulfil their basic functions as guardians of the intra and extra-cellular milieux. Both the signaling and the homeostatic properties of glial cells are therefore particularly dependent upon the regulation of the two principle physiological metal cations, Ca(2+) and Na(+) . For both ions, glia express high-affinity/low capacity ATP-fuelled pumps that can rapidly move small numbers of ions against an electro-chemical gradient. For both Ca(2+) and Na(+) regulation, a single transporter family, the Na(+) -Ca(2+) exchanger (NCX), is used to maintain cellular ion homeostasis over the longer term and under conditions of prolonged or acute ionic dysregulation in astrocytes, oligodendroglia and microglia. Our understanding of glial NCX, both plasmalemmal and mitochondrial, is undergoing the kind of transformation that our understanding of glial cells, in general, has undergone in recent decades. These exchange proteins are becoming increasingly recognized for their essential roles in intracellular homeostasis while their signaling functions are starting to come to light. This review summarizes these key aspects and highlights the many areas where work has yet to begin in this rapidly evolving field. GLIA 2016;64:1646-1654. PMID:27143128

  11. Effects of cadmium alone and in combination with low molecular weight chitosan on metallothionein, glutathione-S-transferase, acid phosphatase, and ATPase of freshwater crab Sinopotamon yangtsekiense.

    Science.gov (United States)

    Li, Ruijin; Zhou, Yanying; Wang, Lan; Ren, Guorui; Zou, Enmin

    2014-03-01

    Cadmium (Cd) is an environmental contaminant showing a variety of deleterious effects, including the potential threat for the ecological environment and human health via food chains. Low molecular weight chitosan (LMWC) has been demonstrated to be an effective antioxidant. Metallothionein (MT) mRNA levels and activities of glutathione-S-transferase (GST), superoxide dismutase (SOD), acid phosphatase (ACP), Na(+),K(+)-ATPase, and Ca(2+)-ATPase as well as malondialdehyde (MDA) contents in the gills of the freshwater crab Sinopotamon yangtsekiense were analyzed in vivo in order to determine the injury of Cd exposure on the gill tissues as well as the protective effect of LMWC against this injury. The results showed that there was an apparent accumulation of Cd in the gills, which was lessened by the presence of LMWC. Moreover, Cd(2+) significantly increased the gill MT mRNA levels, ACP activity and MDA content while decreasing the activities of SOD, GST, Na(+),K(+)-ATPase, and Ca(2+)-ATPase in the crabs relative to the control. Cotreatment with LMWC reduced the levels of MT mRNA and ACP but raised the activities of GST, Na(+),K(+)-ATPase, and Ca(2+)-ATPase in gill tissues compared with the crabs exposed to Cd(2+) alone. These results suggest that LMWC may exert its protective effect through chelating Cd(2+) to form LMWC-Cd(2+) complex, elevating the antioxidative activities of GST, Na(+),K(+)-ATPase, and Ca(2+)-ATPase as well as alleviating the stress pressure on MT and ACP, consequently protecting the cell from the adverse effects of Cd.

  12. Dynamics of matrix-free Ca2+ in cardiac mitochondria: two components of Ca2+ uptake and role of phosphate buffering.

    Science.gov (United States)

    Wei, An-Chi; Liu, Ting; Winslow, Raimond L; O'Rourke, Brian

    2012-06-01

    Mitochondrial Ca(2+) uptake is thought to provide an important signal to increase energy production to meet demand but, in excess, can also trigger cell death. The mechanisms defining the relationship between total Ca(2+) uptake, changes in mitochondrial matrix free Ca(2+), and the activation of the mitochondrial permeability transition pore (PTP) are not well understood. We quantitatively measure changes in [Ca(2+)](out) and [Ca(2+)](mito) during Ca(2+) uptake in isolated cardiac mitochondria and identify two components of Ca(2+) influx. [Ca(2+)](mito) recordings revealed that the first, MCU(mode1), required at least 1 µM Ru360 to be completely inhibited, and responded to small Ca(2+) additions in the range of 0.1 to 2 µM with rapid and large changes in [Ca(2+)](mito). The second component, MCU(mode2), was blocked by 100 nM Ru360 and was responsible for the bulk of total Ca(2+) uptake for large Ca(2+) additions in the range of 2 to 10 µM; however, it had little effect on steady-state [Ca(2+)](mito). MCU(mode1) mediates changes in [Ca(2+)](mito) of 10s of μM, even in the presence of 100 nM Ru360, indicating that there is a finite degree of Ca(2+) buffering in the matrix associated with this pathway. In contrast, the much higher Ca(2+) loads evoked by MCU(mode2) activate a secondary dynamic Ca(2+) buffering system consistent with calcium-phosphate complex formation. Increasing P(i) potentiated [Ca(2+)](mito) increases via MCU(mode1) but suppressed [Ca(2+)](mito) changes via MCU(mode2). The results suggest that the role of MCU(mode1) might be to modulate oxidative phosphorylation in response to intracellular Ca(2+) signaling, whereas MCU(mode2) and the dynamic high-capacity Ca(2+) buffering system constitute a Ca(2+) sink function. Interestingly, the trigger for PTP activation is unlikely to be [Ca(2+)](mito) itself but rather a downstream byproduct of total mitochondrial Ca(2+) loading.

  13. Safrole-induced cellular Ca2+ increases and death in human osteosarcoma cells.

    Science.gov (United States)

    Lin, Hsueh-Chi; Cheng, He-Hsiung; Huang, Chun-Jen; Chen, Wei-Chuan; Chen, I-Shu; Liu, Shiuh-Inn; Hsu, Shu-Shong; Chang, Hong-Tai; Huang, Jong-Khing; Chen, Jin-Shyr; Lu, Yih-Chau; Jan, Chung-Ren

    2006-08-01

    The effect of the carcinogen safrole on intracellular Ca2+ movement has not been explored in osteoblast-like cells. This study examined whether safrole could alter Ca2+ handling and viability in MG63 human osteosarcoma cells. Cytosolic free Ca2+ levels ([Ca2+]i) in populations of cells were measured using fura-2 as a fluorescent Ca2+ probe. Safrole at concentrations above 130 microM increased [Ca2+]i in a concentration-dependent manner with an EC50 value of 450 microM. The Ca2+ signal was reduced by 30% by removing extracellular Ca2+. Addition of Ca2+ after safrole had depleted intracellular Ca2+ induced Ca2+ influx, suggesting that safrole caused Ca2+ entry. In Ca2+-free medium, after pretreatment with 650 microM safrole, 1 microM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor) failed to release more Ca2+; and pretreatment with thapsigargin inhibited most of the safrole-induced [Ca2+]i increases. Inhibition of phospholipase C with U73122 did not affect safrole-induced Ca2+ release; whereas activation of protein kinase C with phorbol ester enhanced safrole-induced [Ca2+]i increase. Trypan exclusion assays revealed that incubation with 65 microM safrole for 30 min did not kill cells, but incubation with 650 microM safrole for 10-30 min nearly killed all cells. Flow cytometry demonstrated that safrole evoked apoptosis in a concentration-dependent manner. Safrole-induced cytotoxicity was not reversed by chelation of Ca2+ with BAPTA. Collectively, the data suggest that in MG63 cells, safrole induced a [Ca2+]i increase by causing Ca2+ release mainly from the endoplasmic reticulum in a phospholipase C-independent manner. The safrole response involved Ca2+ influx and is modulated by protein kinase C. Furthermore, safrole can cause apoptosis in a Ca2+-independent manner.

  14. Ca2+ paradox injury mediated through TRPC channels in mouse ventricular myocytes.

    Science.gov (United States)

    Kojima, Akiko; Kitagawa, Hirotoshi; Omatsu-Kanbe, Mariko; Matsuura, Hiroshi; Nosaka, Shuichi

    2010-12-01

    BACKGROUND AND PURPOSE The Ca(2+) paradox is an important phenomenon associated with Ca(2+) overload-mediated cellular injury in myocardium. The present study was undertaken to elucidate molecular and cellular mechanisms for the development of the Ca(2+) paradox. EXPERIMENTAL APPROACH Fluorescence imaging was performed on fluo-3 loaded quiescent mouse ventricular myocytes using confocal laser scanning microscope. KEY RESULTS The Ca(2+) paradox was readily evoked by restoration of the extracellular Ca(2+) following 10-20 min of nominally Ca(2+)-free superfusion. The Ca(2+) paradox was significantly reduced by blockers of transient receptor potential canonical (TRPC) channels (2-aminoethoxydiphenyl borate, Gd(3+), La(3+)) and anti-TRPC1 antibody. The sarcoplasmic reticulum (SR) Ca(2+) content, assessed by caffeine application, gradually declined during Ca(2+)-free superfusion, which was further accelerated by metabolic inhibition. Block of SR Ca(2+) leak by tetracaine prevented Ca(2+) paradox. The Na(+) /Ca(2+) exchange (NCX) blocker KB-R7943 significantly inhibited Ca(2+) paradox when applied throughout superfusion period, but had little effect when added for a period of 3 min before and during Ca(2+) restoration. The SR Ca(2+) content was better preserved during Ca(2+) depletion by KB-R7943. Immunocytochemistry confirmed the expression of TRPC1, in addition to TRPC3 and TRPC4, in mouse ventricular myocytes. CONCLUSIONS AND IMPLICATIONS These results provide evidence that (i) the Ca(2+) paradox is primarily mediated by Ca(2+) entry through TRPC (probably TRPC1) channels that are presumably activated by SR Ca(2+) depletion; and (ii) reverse mode NCX contributes little to the Ca(2+) paradox, whereas inhibition of NCX during Ca(2+) depletion improves SR Ca(2+) loading, and is associated with reduced incidence of Ca(2+) paradox in mouse ventricular myocytes.

  15. Rapid recycling of Ca2+ between IP3-sensitive stores and lysosomes.

    Directory of Open Access Journals (Sweden)

    Cristina I López Sanjurjo

    Full Text Available Inositol 1,4,5-trisphosphate (IP3 evokes release of Ca2+ from the endoplasmic reticulum (ER, but the resulting Ca2+ signals are shaped by interactions with additional intracellular organelles. Bafilomycin A1, which prevents lysosomal Ca2+ uptake by inhibiting H+ pumping into lysosomes, increased the amplitude of the initial Ca2+ signals evoked by carbachol in human embryonic kidney (HEK cells. Carbachol alone and carbachol in combination with parathyroid hormone (PTH evoke Ca2+ release from distinct IP3-sensitive Ca2+ stores in HEK cells stably expressing human type 1 PTH receptors. Bafilomycin A1 similarly exaggerated the Ca2+ signals evoked by carbachol or carbachol with PTH, indicating that Ca2+ released from distinct IP3-sensitive Ca2+ stores is sequestered by lysosomes. The Ca2+ signals resulting from store-operated Ca2+ entry, whether evoked by thapsigargin or carbachol, were unaffected by bafilomycin A1. Using Gd3+ (1 mM to inhibit both Ca2+ entry and Ca2+ extrusion, HEK cells were repetitively stimulated with carbachol to assess the effectiveness of Ca2+ recycling to the ER after IP3-evoked Ca2+ release. Blocking lysosomal Ca2+ uptake with bafilomycin A1 increased the amplitude of each carbachol-evoked Ca2+ signal without affecting the rate of Ca2+ recycling to the ER. This suggests that Ca2+ accumulated by lysosomes is rapidly returned to the ER. We conclude that lysosomes rapidly, reversibly and selectively accumulate the Ca2+ released by IP3 receptors residing within distinct Ca2+ stores, but not the Ca2+ entering cells via receptor-regulated, store-operated Ca2+ entry pathways.

  16. Optimisation of recombinant production of active human cardiac SERCA2a ATPase

    OpenAIRE

    Antaloae, Ana V.; Cédric Montigny; Marc le Maire; Watson, Kimberly A.; Thomas L-M Sørensen

    2013-01-01

    Methods for recombinant production of eukaryotic membrane proteins, yielding sufficient quantity and quality of protein for structural biology, remain a challenge. We describe here, expression and purification optimisation of the human SERCA2a cardiac isoform of Ca(2+) translocating ATPase, using Saccharomyces cerevisiae as the heterologous expression system of choice. Two different expression vectors were utilised, allowing expression of C-terminal fusion proteins with a biotinylation domain...

  17. High extracellular Ca2+ stimulates Ca2+-activated Cl- currents in frog parathyroid cells through the mediation of arachidonic acid cascade.

    Directory of Open Access Journals (Sweden)

    Yukio Okada

    Full Text Available Elevation of extracellular Ca(2+ concentration induces intracellular Ca(2+ signaling in parathyroid cells. The response is due to stimulation of the phospholipase C/Ca(2+ pathways, but the direct mechanism responsible for the rise of intracellular Ca(2+ concentration has remained elusive. Here, we describe the electrophysiological property associated with intracellular Ca(2+ signaling in frog parathyroid cells and show that Ca(2+-activated Cl(- channels are activated by intracellular Ca(2+ increase through an inositol 1,4,5-trisphophate (IP(3-independent pathway. High extracellular Ca(2+ induced an outwardly-rectifying conductance in a dose-dependent manner (EC(50 ∼6 mM. The conductance was composed of an instantaneous time-independent component and a slowly activating time-dependent component and displayed a deactivating inward tail current. Extracellular Ca(2+-induced and Ca(2+ dialysis-induced currents reversed at the equilibrium potential of Cl(- and were inhibited by niflumic acid (a specific blocker of Ca(2+-activated Cl(- channel. Gramicidin-perforated whole-cell recording displayed the shift of the reversal potential in extracellular Ca(2+-induced current, suggesting the change of intracellular Cl(- concentration in a few minutes. Extracellular Ca(2+-induced currents displayed a moderate dependency on guanosine triphosphate (GTP. All blockers for phospholipase C, diacylglycerol (DAG lipase, monoacylglycerol (MAG lipase and lipoxygenase inhibited extracellular Ca(2+-induced current. IP(3 dialysis failed to induce conductance increase, but 2-arachidonoylglycerol (2-AG, arachidonic acid and 12S-hydroperoxy-5Z,8Z,10E,14Z-eicosatetraenoic acid (12(S-HPETE dialysis increased the conductance identical to extracellular Ca(2+-induced conductance. These results indicate that high extracellular Ca(2+ raises intracellular Ca(2+ concentration through the DAG lipase/lipoxygenase pathway, resulting in the activation of Cl(- conductance.

  18. 补骨脂、生地黄对正常大鼠体温及ATP酶活性的影响%Effect of Psoraleae Fructus and Rehmanniae Radix on Body Temperature and ATPase Activity in Rats

    Institute of Scientific and Technical Information of China (English)

    宋晓玲; 李峰; 崔光志

    2013-01-01

    目的:探讨补骨脂、生地黄对正常大鼠体温及ATP酶活性的影响.方法:将大鼠随机分为空白对照组、附子热性对照组、大黄寒性对照组、补骨脂组与生地黄组,附子组6.75 g·kg-1,大黄组13.5 g·kg-1,补骨脂组4.05 g·kg-1,生地黄组6.75 g·kg-1,按10 mL·kg-1ig,1次/d,每周给药6d,停药1d,共给药28 d,空白组ig等量(10 mL·kg-1)生理盐水.给药28 d后测量大鼠趾温、肛温,大鼠处死后,取出肝脏,制备组织匀浆,测定三磷酸腺苷(ATP)酶活性.结果:与空白对照组相比,补骨脂组大鼠趾温、肛温均明显升高(P<0.05),生地黄组大鼠趾温、肛温均明显降低(P<0.05).与空白对照组相比,补骨脂组肝组织中Na+-K+-ATP酶、Ca2+-Mg2+-ATP酶活性均明显升高(P<0.05);生地黄组Na+-K+-ATP酶,Ca2+-Mg2+-ATP酶活性均明显降低(P<0.05).结论:补骨脂、生地黄对正常大鼠体温及ATP酶活性影响明显,提示寒、热药性不同的药物与对体内ATP酶活性的影响不同可能是对体温产生不同的作用趋势的原因之一.%Objective: To discuss the effect of Psoraleae Fructus and Rehmanniae Radix on body temperature and ATPase activity in rats. Method: Fifty rats were randomly divided into the blank control group, Aconiti Lateralis Radix Praeparata control group, Rhei Radix et Rhizoma control group, psoraleae fructus group and Rehmanniae Radix group, the dosage of aconiti Lateralis Radix Praeparata control group was 6. 75 g ·kg-1; the dosage of Rhei Radix et Rhfizoma control group was 13. 5 g·kg-1; the dosage of Psoraleae Fructus group was 4. 05 g ·kg-1 ; the dosage of Rehmanniae Radix group was 6. 75 g ·kg-1. The rats were fed with the amount of 10 mL · kg-1 for 28 days ( the blank group was fed with equal volume of physiological saline irrigation ) , 1 time a day, lasting 6 days, stopping 1 day. The toe temperature and Rectal Temperature of the rats were measured 28 days after administration. The liver was removed and

  19. Glucose-induced Ca2 + signals in rat pancreatic β cells

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Using microfluorometry to assay intracellular Ca2+ , the influences of varied factors on glucose induced Ca22+ signals, such as glucose-induced initial decline phase (GIDP), Ca2+ oscillation, and Ca2+ release from internal stores, were investigated in single rat pancreatic β cells. Glucose was able to evoke GIDP even at non-stimulus concentration (5 mol/L), which is insufficient to induce Ca2+ spikes. GIDP was dependent on neither membrane depo larization nor extraeellular Ca2+ . However, GIDP was inhibited by thapsigargin, indicating a dependence on Ca2+ up take by Ca22+ stores. The glucose-induced calcium oscillation was inhibited when external Ca2+ was removed. However, thapsigargin could not block the Ca2+ oscillation. These results suggest that maintenance of Ca22+ oscillation requires ex tracellular Ca2+ but not Ca2+ stores. Glucose was able to evoke Ca2+ signals even in the absence of external Ca2+ . The glucose-induced Ca2+ release from intracellular Ca2+ stores was blocked by TTX. However, TTX had no effect on high K--induced Ca2+ store release, suggesting that membrane depolarization can directly release Ca2+ from some internal Ca2+ stores in β cells.

  20. The permeability transition pore as a Ca2+ release channel: New answers to an old question

    Science.gov (United States)

    Bernardi, Paolo; von Stockum, Sophia

    2012-01-01

    Mitochondria possess a sophisticated array of Ca2+ transport systems reflecting their key role in physiological Ca2+ homeostasis. With the exception of most yeast strains, energized organelles are endowed with a very fast and efficient mechanism for Ca2+ uptake, the ruthenium red (RR)-sensitive mitochondrial Ca2+ uniporter (MCU); and one main mechanism for Ca2+ release, the RR-insensitive 3Na+–Ca2+ antiporter. An additional mechanism for Ca2+ release is provided by a Na+ and RR-insensitive release mechanism, the putative 3H+–Ca2+ antiporter. A potential kinetic imbalance is present, however, because the Vmax of the MCU is of the order of 1400 nmol Ca2+ mg−1 protein min−1 while the combined Vmax of the efflux pathways is about 20 nmol Ca2+ mg−1 protein min−1. This arrangement exposes mitochondria to the hazards of Ca2+ overload when the rate of Ca2+ uptake exceeds that of the combined efflux pathways, e.g. for sharp increases of cytosolic [Ca2+]. In this short review we discuss the hypothesis that transient opening of the Ca2+-dependent permeability transition pore may provide mitocondria with a fast Ca2+ release channel preventing Ca2+ overload. We also address the relevance of a mitochondrial Ca2+ release channel recently discovered in Drosophila melanogaster, which possesses intermediate features between the permeability transition pore of yeast and mammals. PMID:22513364

  1. The permeability transition pore as a Ca(2+) release channel: new answers to an old question.

    Science.gov (United States)

    Bernardi, Paolo; von Stockum, Sophia

    2012-07-01

    Mitochondria possess a sophisticated array of Ca(2+) transport systems reflecting their key role in physiological Ca(2+) homeostasis. With the exception of most yeast strains, energized organelles are endowed with a very fast and efficient mechanism for Ca(2+) uptake, the ruthenium red (RR)-sensitive mitochondrial Ca(2+) uniporter (MCU); and one main mechanism for Ca(2+) release, the RR-insensitive 3Na(+)-Ca(2+) antiporter. An additional mechanism for Ca(2+) release is provided by a Na(+) and RR-insensitive release mechanism, the putative 3H(+)-Ca(2+) antiporter. A potential kinetic imbalance is present, however, because the V(max) of the MCU is of the order of 1400nmol Ca(2+)mg(-1) proteinmin(-1) while the combined V(max) of the efflux pathways is about 20nmol Ca(2+)mg(-1) proteinmin(-1). This arrangement exposes mitochondria to the hazards of Ca(2+) overload when the rate of Ca(2+) uptake exceeds that of the combined efflux pathways, e.g. for sharp increases of cytosolic [Ca(2+)]. In this short review we discuss the hypothesis that transient opening of the Ca(2+)-dependent permeability transition pore may provide mitocondria with a fast Ca(2+) release channel preventing Ca(2+) overload. We also address the relevance of a mitochondrial Ca(2+) release channel recently discovered in Drosophila melanogaster, which possesses intermediate features between the permeability transition pore of yeast and mammals. PMID:22513364

  2. Crosstalk between mitochondrial and sarcoplasmic reticulum Ca2+ cycling modulates cardiac pacemaker cell automaticity.

    Directory of Open Access Journals (Sweden)

    Yael Yaniv

    Full Text Available BACKGROUND: Mitochondria dynamically buffer cytosolic Ca(2+ in cardiac ventricular cells and this affects the Ca(2+ load of the sarcoplasmic reticulum (SR. In sinoatrial-node cells (SANC the SR generates periodic local, subsarcolemmal Ca(2+ releases (LCRs that depend upon the SR load and are involved in SANC automaticity: LCRs activate an inward Na(+-Ca(2+ exchange current to accelerate the diastolic depolarization, prompting the ensemble of surface membrane ion channels to generate the next action potential (AP. OBJECTIVE: To determine if mitochondrial Ca(2+ (Ca(2+ (m, cytosolic Ca(2+ (Ca(2+ (c-SR-Ca(2+ crosstalk occurs in single rabbit SANC, and how this may relate to SANC normal automaticity. RESULTS: Inhibition of mitochondrial Ca(2+ influx into (Ru360 or Ca(2+ efflux from (CGP-37157 decreased [Ca(2+](m to 80 ± 8% control or increased [Ca(2+](m to 119 ± 7% control, respectively. Concurrent with inhibition of mitochondrial Ca(2+ influx or efflux, the SR Ca(2+ load, and LCR size, duration, amplitude and period (imaged via confocal linescan significantly increased or decreased, respectively. Changes in total ensemble LCR Ca(2+ signal were highly correlated with the change in the SR Ca(2+ load (r(2 = 0.97. Changes in the spontaneous AP cycle length (Ru360, 111 ± 1% control; CGP-37157, 89 ± 2% control in response to changes in [Ca(2+](m were predicted by concurrent changes in LCR period (r(2 = 0.84. CONCLUSION: A change in SANC Ca(2+ (m flux translates into a change in the AP firing rate by effecting changes in Ca(2+ (c and SR Ca(2+ loading, which affects the characteristics of spontaneous SR Ca(2+ release.

  3. Modeling a dehalogenase fold into the 8-A density map for Ca(2+)-ATPase defines a new domain structure.

    OpenAIRE

    Stokes, D.L.; Green, N M

    2000-01-01

    Members of the large family of P-type pumps use active transport to maintain gradients of a wide variety of cations across cellular membranes. Recent structures of two P-type pumps at 8-A resolution have revealed the arrangement of transmembrane helices but were insufficient to reveal the architecture of the cytoplasmic domains. However, recent proposals of a structural homology with a superfamily of hydrolases offer a new basis for modeling these domains. In the current work, we have extende...

  4. A tomato ER-type Ca2+-ATPase, LCA1, has a low thapsigargin-sensitivity and can transport manganese

    DEFF Research Database (Denmark)

    Johnson, Neil A.; Liu, F; Weeks, P. D.;

    2008-01-01

    and directly via in vitro ATP hydrolysis. We found LCA1 to be approximately 300-fold less sensitive to thapsigargin than animal SERCAs, whereas ECA1 was thapsigargin-resistant. LCA1 showed typical pharmacological sensitivities to cyclopiazonic acid, vanadate, and eosin, consistent with it being a P...

  5. Hyperbaric oxygen treatment induces dynamic ATPase activity changes in the rat brain following transient global cerebral ischemia-reperfusion

    Institute of Scientific and Technical Information of China (English)

    Shiming Xu; Hongjuan Wang; Tongnan Gu; Xiuyan Zhou; Rui Chen

    2008-01-01

    BACKGROUND: Energy depletion, induced by ischemia or hypoxia, is one of the first events in neuronal injury. OBJECTIVE: To investigate the dynamic changes of Na+-K+-ATPase and Ca2+-ATPase activity in the rat brain following transient global cerebral ischemia-reperfusion (IR), as well as the effects of hyperbaric oxygen (HBO) treatment. DESIGN, TIME AND SETTING: A randomized and controlled animal study was performed in the Department of Biochemistry and Molecular Biology, Capital Medical University between February and December 2006. MATERIALS: Clean-grade, female, Sprague Dawley rats were provided by the Animal Research Department of Capital Medical University (License number: SYXK11-00-0047). Na+-K+-ATPase and Ca2+-ATPase kits were provided by Nanjing Jiancheng Bioengineering Institute (Nanjing, China). A hyperbaric oxygen chamber (DWC150-300) was supplied by Shanghai 701 Medical Oxygen Chamber Factory (Shanghai, China). METHODS: Sixty-three rats were randomly divided into nine groups: sham operated group (sham-O) as control, groups of IR, and groups treated with hyperbaric oxygen (HBO) after IR. Animal from the IR and HBO groups were sacrificed after four different survival intervals of 6, 24, 48 and 96 hours, respectively. Each group consisted of seven rats. The rats of HBO groups were placed into the hyperbaric chamber. The HBO chamber was flushed with pure oxygen for 5 minutes, followed by a gradual rise in pressure over 5 minutes and stabilization at 0.2 MPa. Then, pure oxygen was supplied for 45 minutes in stabilized pressure, followed by gradually reduced pressure over 15 minutes. The rats of the 6-h HBO group were placed into the HBO chamber following reperfusion for 3 hours on the first day, which was repeated on three consecutive days, always at the same time. Rats in the sham-O group and IR group remained under normal atmospheric pressure. MAIN OUTCOME MEASURES: The Na+-K+-ATPase and Ca2+-ATPase activity in rat brain homogenate was detected by the

  6. Diglyceride kinase activity of microtubules. Characterization and comparison with the protein kinase and ATPase activities associated with vinblastine-isolated tubulin of chick embryonic muscles.

    Science.gov (United States)

    Daleo, G R; Piras, M M; Piras, R

    1976-09-15

    Vinblastine-isolated microtubule protein from chick embryonic muscles has an enzymatic activity which catalyzes the formation of phosphatidic acid from diglycerides and ATP. The pH optimum (6.4), sedimentation on sucrose gradients (Mr = 85 000), and sensitivity to ions of this diglyceride kinase activity are different to those of a similar enzymatic activity present in 150 000 X g supernatants of chick embryonic muscle homogenates, suggesting that it is a different species which is associated specifically with the microtubules. The reaction requires a divalent ion (e.g. 0.4 mM Mg2+ gives half-maximal stimulation), and GTP can replace ATP rather effectively, especially at nucleotide concentrations lower than 50 muM. The sedimentation of the diglyceride kinase on sucrose gradients coincides with that of the microtubules-associated protein kinase (Mr = 75 000); the heat-stability and sensivitity to proteolysis of both activities are also very similar. Stimulation of one reaction by the addition of the corresponding exogenous substrate does not impair the phosphorylation of the other, and no radioactivity is lost from phosphatidic acid or the protein moiety upon incubation of pre-labelled microtubules with a large excess of unlabelled ATP or GTP. In addition to diglyceride and protein kinase activities (0.2 and 0.3 nmol 32P-transferred X min-1 X mg-1 microtubular protein, respectively), microtubules also contain an associated ATPase (2.8 nmol X min-1 X mg-1), which requires either Mg2+ or Ca2+, can hydrolyze GTP quite effectively, and sediments with a molecular weight of 95000. The results obtained are discussed in connection with the possible relationships existing among these enzymatic activities, as well as their probable role in microtubular functions.

  7. Chloroplast-Specific in Vivo Ca2+ Imaging Using Yellow Cameleon Fluorescent Protein Sensors Reveals Organelle-Autonomous Ca2+ Signatures in the Stroma.

    Science.gov (United States)

    Loro, Giovanna; Wagner, Stephan; Doccula, Fabrizio Gandolfo; Behera, Smrutisanjita; Weinl, Stefan; Kudla, Joerg; Schwarzländer, Markus; Costa, Alex; Zottini, Michela

    2016-08-01

    In eukaryotes, subcellular compartments such as mitochondria, the endoplasmic reticulum, lysosomes, and vacuoles have the capacity for Ca(2+) transport across their membranes to modulate the activity of compartmentalized enzymes or to convey specific cellular signaling events. In plants, it has been suggested that chloroplasts also display Ca(2+) regulation. So far, monitoring of stromal Ca(2+) dynamics in vivo has exclusively relied on using the luminescent Ca(2+) probe aequorin. However, this technique is limited in resolution and can only provide a readout averaged over chloroplast populations from different cells and tissues. Here, we present a toolkit of Arabidopsis (Arabidopsis thaliana) Ca(2+) sensor lines expressing plastid-targeted FRET-based Yellow Cameleon (YC) sensors. We demonstrate that the probes reliably report in vivo Ca(2+) dynamics in the stroma of root plastids in response to extracellular ATP and of leaf mesophyll and guard cell chloroplasts during light-to-low-intensity blue light illumination transition. Applying YC sensing of stromal Ca(2+) dynamics to single chloroplasts, we confirm findings of gradual, sustained stromal Ca(2+) increases at the tissue level after light-to-low-intensity blue light illumination transitions, but monitor transient Ca(2+) spiking as a distinct and previously unknown component of stromal Ca(2+) signatures. Spiking was dependent on the availability of cytosolic Ca(2+) but not synchronized between the chloroplasts of a cell. In contrast, the gradual sustained Ca(2+) increase occurred independent of cytosolic Ca(2+), suggesting intraorganellar Ca(2+) release. We demonstrate the capacity of the YC sensor toolkit to identify novel, fundamental facets of chloroplast Ca(2+) dynamics and to refine the understanding of plastidial Ca(2+) regulation. PMID:27252306

  8. Bursting Ca2+ Oscillations and Synchronization in Coupled Cells

    Institute of Scientific and Technical Information of China (English)

    JI Quan-Bao; LU Qi-Shao; Yang Zhuo-Qin; Duan Li-Xia

    2008-01-01

    A mathematical model proposed by Grubelnk et al. [Biophys. Chem. 94 (2001) 59] is employed to study the physiological role of mitochondria and the cytosolic proteins in generating complex Ca2+ oscillations. Intracellular bursting calcium oscillations of point-point, point cycle and two-folded limit cycle types are observed and explanations are given based on the fast/slow dynamical analysis, especially for point-cycle and two-folded limit cycle types, which have not been reported before. Furthermore, synchronization of coupled bursters of Ca2+oscillations via gap junctions and the effect of bursting types on synchronization of coupled cells are studied. It is argued that bursting oscillations of point-point type may be superior to achieve synchronization than that of point-cycle type.

  9. Antifungal Effect of Chitosan as Ca(2+) Channel Blocker.

    Science.gov (United States)

    Lee, Choon Geun; Koo, Ja Choon; Park, Jae Kweon

    2016-06-01

    The aim of this study was to investigate antifungal activity of a range of different molecular weight (MW) chitosan against Penicillium italicum. Our results demonstrate that the antifungal activity was dependent both the MW and concentration of the chitosan. Among a series of chitosan derived from the hydrolysis of high MW chitosan, the fractions containing various sizes of chitosan ranging from 3 to 15 glucosamine units named as chitooligomers-F2 (CO-F2) was found to show the highest antifungal activity against P. italicum. Furthermore, the effect of CO-F2 toward this fungus was significantly reduced in the presence of Ca(2+), whereas its effect was recovered by ethylenediaminetetraacetic acid, suggesting that the CO-F2 acts via disruption of Ca(2+) gradient required for survival of the fungus. Our results suggest that CO-F2 may serve as potential compounds to develop alternatives to synthetic fungicides for the control of the postharvest diseases. PMID:27298599

  10. Antifungal Effect of Chitosan as Ca2+ Channel Blocker

    Science.gov (United States)

    Lee, Choon Geun; Koo, Ja Choon; Park, Jae Kweon

    2016-01-01

    The aim of this study was to investigate antifungal activity of a range of different molecular weight (MW) chitosan against Penicillium italicum. Our results demonstrate that the antifungal activity was dependent both the MW and concentration of the chitosan. Among a series of chitosan derived from the hydrolysis of high MW chitosan, the fractions containing various sizes of chitosan ranging from 3 to 15 glucosamine units named as chitooligomers-F2 (CO-F2) was found to show the highest antifungal activity against P. italicum. Furthermore, the effect of CO-F2 toward this fungus was significantly reduced in the presence of Ca2+, whereas its effect was recovered by ethylenediaminetetraacetic acid, suggesting that the CO-F2 acts via disruption of Ca2+ gradient required for survival of the fungus. Our results suggest that CO-F2 may serve as potential compounds to develop alternatives to synthetic fungicides for the control of the postharvest diseases. PMID:27298599

  11. 中等强度持续训练对大鼠骨骼肌线粒体抗氧化能力及ATP酶活性的影响%Effects of moderate-intensity continuous training on mitochondrial antioxidant capacity and activity of ATPase in skeletal muscle:experiment with rats

    Institute of Scientific and Technical Information of China (English)

    谢红; 陈立军; 史娜

    2014-01-01

    目的:观察中等强度持续训练后骨骼肌线粒体抗氧化能力和ATP酶活性的变化。方法 SD大鼠24只,8只为正常对照组(A组),16只采用递增负荷训练建立中等强度持续运动跑台训练模型,又均分为运动2周及6周组(B及C组)。B及C组大鼠按Bradford等的方法分别训练2和6周,每周训练6 d。训练结束后大鼠处死,用差速离心法提取腓肠肌线粒体,紫外-可见分光光度计检测锰超氧化物歧化酶(Mn-SOD)活性;脂质过氧化产物丙二醛(MDA)含量采用TBA(硫代巴比妥酸)法测定;Na+K+-ATP酶和Ca2+Mg2+-ATP酶活性采用酶促反应定磷法测定。结果B组的骨骼肌线粒体Mn-SOD水平与A组相比差异无统计学意义,P>0.05,而MDA含量显著高于对照组,P0.05, whereas the content of MDA of Group C was significantly higher than that of Group A ,P<0.05. The Mn-SOD level of Group C was significantly higher than that of Group A,P<0.05, however, the content of MDA of Group C was signiuficantly lower than that of Groupo A ,P<0.05. The activities of Na+K+-ATPase and Ca2+Mg2+-ATPase of Group B were both significantly lower than those of Group A,bothP<0.05, however, those of Group C were both significantly higher than that of Group A ,P <0.05. Conclusion A certain period of moderate-intensity continuous training increases the activity of Mn-SOD, decreases the level of lipid peroxidation, reduces the injury of free radical, enhances the activity of ATPase, and improves the level of the energy metabolism in skeletal muscle mitochondria .

  12. Effects of Aluminum on ATPase Activity and Lipid Composition of Plasma Membranes from Wheat Roots

    Institute of Scientific and Technical Information of China (English)

    HE Long-fei; LIU You-liang; SHEN Zhen-guo; WANG Ai-qin

    2002-01-01

    The effects of aluminum on ATPase activity and lipid composition of the plasma membranes isolated from root tips of Al-tolerant (Altas 66) or Al-sensitive (Scout 66) cultivar of Triticum aestivum L.was assayed. The results showed that both cultivars had similar changes in H+ -ATPase and Ca2+ -ATPase activities after aluminum treatment. Exposure of both cultivars to 20 and 100 (mol/L aluminum for 5 d significantly decreased the activities of Ca2+ -ATPase of plasma membranes. The activities of H+-ATPasc in plasma membrane increased under 20 μmol/L aluminum and decreased at 100 μmol/L aluminum. With aluminum treatment, the PL content of plasma membrane decreased, but GL content increased. The ratio of PL to GL decreased more distinctly in Scout 66 than that in Altas 66. Treated with 20 and 100 μmol/L aluminum, linolenic acid content and the index of unsaturated fatty acids decreaced greatly in Scout 66, but the index of unsaturated fatty acids in Altas 66 increased slightly.

  13. Ketamine inhibits human sperm function by Ca(2+)-related mechanism.

    Science.gov (United States)

    He, Yuanqiao; Zou, Qianxing; Li, Bingda; Chen, Houyang; Du, Xiaohong; Weng, Shiqi; Luo, Tao; Zeng, Xuhui

    2016-09-01

    Ketamine, a dissociative anesthetic, which was widely used in human and animal medicine, has become a popular recreational drug, as it can induce hallucinatory effects. Ketamine abuse can cause serious damage to many aspects of the organism, mainly reflected in the nervous system and urinary system. It has also been reported that ketamine can impair the male genital system. However, the detailed effect of ketamine on human spermatozoa remains unclear. Thus, we investigated the in vitro effects of ketamine on human sperm functions, to elucidate the underlying mechanism. Human sperm were treated in vitro with different concentrations of ketamine (0, 0.125, 0.25, 0.5, 1 g/L). The results showed that 0.25-1 g/L ketamine inhibited sperm total motility, progressive motility and linear velocity, in a dose-dependent manner. In addition, the sperm's ability to penetrate viscous medium and the progesterone-induced acrosome reaction were significantly inhibited by ketamine. Ketamine did not affect sperm viability, capacitation and spontaneous acrosome reaction. The intracellular calcium concentration ([Ca(2+)]i), which is a central factor in the regulation of human sperm function, was decreased by ketamine (0.125-1 g/L) in a dose-dependent manner. Furthermore, the currents of the sperm-specific Ca(2+) channel, CatSper, which modulates Ca(2+) influx in sperm, were inhibited by ketamine (0.125-1 g/L) in a dose-dependent manner. Our findings suggest that ketamine induces its toxic effects on human sperm functions by reducing sperm [Ca(2+)]i through inhibition of CatSper channel. PMID:27143628

  14. Ca2+influx insensitive to organic Ca2+entry blockers contributes to noradrenaline-induced contractions of the isolated guinea pig aorta

    NARCIS (Netherlands)

    Gouw, M.A.M.; Wilffert, B.; Wermelskirchen, D.; Van Zwieten, P.A.

    1990-01-01

    We determined the contribution of intracellular Ca2+to the noradrenaline (NA, 3 x 10-5mmol/l)-induced contraction of the isolated guinea pig aorta. Since only about 55% of the NA-induced contraction could be attributed to intracellular Ca2+release, we assumed that a Ca2+influx component contributes

  15. Plasma membrane Ca2+ pumping plays a prominent role in adenosine A(1) receptor mediated changes in [Ca2+](i) in DDT1 MF-2 cells

    NARCIS (Netherlands)

    Sipma, H; Fredholm, BB; DenHertog, A; Nelemans, A

    1996-01-01

    Adenosine A(1) receptor mediated formation of inositol 1,4,5-trisphosphate (Ins(1,4,5)P-3) and accumulation of cytoplasmic Ca2+ ([Ca2+](i)) were investigated in DDT1 MF-2 smooth muscle cells. A strong reduction of the adenosine and N-6-cyclopentyladenosine (CPA) induced rise in [Ca2+](i) was observe

  16. Ca2+-Doped CeBr3 Scintillating Materials

    Energy Technology Data Exchange (ETDEWEB)

    Guss, Paul [NSTec; Foster, Michael E. [SNL; Wong, Bryan M. [SNL; Doty, F. Patrick [SNL; Shah, Kanai [RMD; Squillante, Michael R. [RMD; Shirwadkar, Urmila [RMD; Hawrami, Rastgo [RMD; Tower, Josh [RMD; Yuan, Ding [NSTec

    2014-01-01

    Despite the outstanding scintillation performance characteristics of cerium tribromide (CeBr3) and cerium-activated lanthanum tribromide, their commercial availability and application are limited due to the difficulties of growing large, crack-free single crystals from these fragile materials. This investigation employed aliovalent doping to increase crystal strength while maintaining the optical properties of the crystal. One divalent dopant (Ca2+) was used as a dopant to strengthen CeBr3 without negatively impacting scintillation performance. Ingots containing nominal concentrations of 1.9% of the Ca2+ dopant were grown. Preliminary scintillation measurements are presented for this aliovalently doped scintillator. Ca2+-doped CeBr3 exhibited little or no change in the peak fluorescence emission for 371 nm optical excitation for CeBr3. The structural, electronic, and optical properties of CeBr3 crystals were studied using the density functional theory within the generalized gradient approximation. The calculated lattice parameters are in good agreement with the experimental data. The energy band structures and density of states were obtained. The optical properties of CeBr3, including the dielectric function, were calculated.

  17. Sympathetically evoked Ca2+ signaling in arterial smooth muscle

    Institute of Scientific and Technical Information of China (English)

    Wei-jin ZANG; Joseph ZACHARIA; Christine LAMONT; Withrow Gil WIER

    2006-01-01

    The sympathetic nervous system plays an essential role in the control of total peripheral vascular resistance and blood flow, by controlling the contraction of small arteries. Perivascular sympathetic nerves release ATP, norepinephrine (NE) and neuropeptide Y. This review summarizes our knowledge of the intracellular Ca2+ signals that are activated by ATP and NE, acting respectively on P2X1 and α1 adrenoceptors in arterial smooth muscle. Each neurotransmitter produces a unique type of post-synaptic Ca2+ signal and associated contraction. The neural release of ATP and NE is thought to vary markedly with the pattern of nerve activity, probably reflecting both pre- and post-synaptic mechanisms. Finally, we show that Ca2+ signaling during neurogenic contractions activated by trains of sympathetic nerve fiber action potentials are in fact significantly different from that elicited by simple bath application of exogenous neurotransmitters to isolated arteries (a common experimental technique), and end by identifying important questions remaining in our understanding of sympathetic neurotransmission and the physiological regulation of contraction of small arteries.

  18. Apical Scaffolding Protein NHERF2 Modulates the Localization of Alternatively Spliced Plasma Membrane Ca2+ Pump 2B Variants in Polarized Epithelial Cells*

    OpenAIRE

    Padányi, Rita; Xiong, Yuning; Antalffy, Géza; Lór, Krisztina; Pászty, Katalin; STREHLER, EMANUEL E.; Enyedi, Ágnes

    2010-01-01

    The membrane localization of the plasma membrane Ca2+-ATPase isoform 2 (PMCA2) in polarized cells is determined by alternative splicing; the PMCA2w/b splice variant shows apical localization, whereas the PMCA2z/b and PMCA2x/b variants are mostly basolateral. We previously reported that PMCA2b interacts with the PDZ protein Na+/H+ exchanger regulatory factor 2 (NHERF2), but the role of this interaction for the specific membrane localization of PMCA2 is not known. Here we show that co-expressio...

  19. The permeability transition pore as a Ca2+ release channel: New answers to an old question

    OpenAIRE

    Bernardi, Paolo; von Stockum, Sophia

    2012-01-01

    Mitochondria possess a sophisticated array of Ca2+ transport systems reflecting their key role in physiological Ca2+ homeostasis. With the exception of most yeast strains, energized organelles are endowed with a very fast and efficient mechanism for Ca2+ uptake, the ruthenium red (RR)-sensitive mitochondrial Ca2+ uniporter (MCU); and one main mechanism for Ca2+ release, the RR-insensitive 3Na+–Ca2+ antiporter. An additional mechanism for Ca2+ release is provided by a Na+ and RR-insensitive re...

  20. Modeling of the Modulation by Buffers of Ca2+ Release through Clusters of IP3 Receptors

    OpenAIRE

    Zeller, S.; Rüdiger, S.; H. Engel; Sneyd, J.; Warnecke, G.; Parker, I; Falcke, M.

    2009-01-01

    Intracellular Ca2+ release is a versatile second messenger system. It is modeled here by reaction-diffusion equations for the free Ca2+ and Ca2+ buffers, with spatially discrete clusters of stochastic IP3 receptor channels (IP3Rs) controlling the release of Ca2+ from the endoplasmic reticulum. IP3Rs are activated by a small rise of the cytosolic Ca2+ concentration and inhibited by large concentrations. Buffering of cytosolic Ca2+ shapes global Ca2+ transients. Here we use a model to investiga...

  1. Acrosome Reaction and Ca2+ Imaging in Single Human Spermatozoa: New Regulatory Roles of [Ca2+]i1

    OpenAIRE

    Sánchez-Cárdenas, Claudia; Servín-Vences, Martha Rocio; José, Omar; Treviño, Claudia Lydia; Hernández-Cruz, Arturo; Darszon, Alberto

    2014-01-01

    The spermatozoa acrosome reaction (AR) is essential for mammalian fertilization. Few methods allow visualization of AR in real time together with Ca2+ imaging. Here, we show that FM4-64, a fluorescent dye used to follow exocytosis, reliably reports AR progression induced by ionomycin and progesterone in human spermatozoa. FM4-64 clearly delimits the spermatozoa contour and reports morphological cell changes before, during, and after AR. This strategy unveiled the formation of moving tubular a...

  2. Phospholipase C-η1 is activated by intracellular Ca(2+) mobilization and enhances GPCRs/PLC/Ca(2+) signaling.

    Science.gov (United States)

    Kim, Jung Kuk; Choi, Jung Woong; Lim, Seyoung; Kwon, Ohman; Seo, Jeong Kon; Ryu, Sung Ho; Suh, Pann-Ghill

    2011-06-01

    Phospholipase C-η1 (PLC-η1) is the most recently identified PLC isotype and is primarily expressed in nerve tissue. However, its functional role is unclear. In the present study, we report for the first time that PLC-η1 acts as a signal amplifier in G protein-coupled receptor (GPCR)-mediated PLC and Ca(2+) signaling. Short-hairpin RNA (shRNA)-mediated knockdown of endogenous PLC-η1 reduced lysophosphatidic acid (LPA)-, bradykinin (BK)-, and PACAP-induced PLC activity in mouse neuroblastoma Neuro2A (N2A) cells, indicating that PLC-η1 participates in GPCR-mediated PLC activation. Interestingly, ionomycin-induced PLC activity was significantly decreased by PLC-η1, but not PLC-η2, knockdown. In addition, we found that intracellular Ca(2+) source is enough for PLC-η1 activation. Furthermore, the IP(3) receptor inhibitor, 2-APB, inhibited LPA-induced PLC activity in control N2A cells, whereas this effect was not observed in PLC-η1 knockdown N2A cells, suggesting a pivotal role of intracellular Ca(2+) mobilization in PLC-η1 activation. Finally, we found that LPA-induced ERK1/2 phosphorylation and expression of the downstream target gene, krox-24, were significantly decreased by PLC-η1 knockdown, and these knockdown effects were abolished by 2-APB. Taken together, our results strongly suggest that PLC-η1 is activated via intracellular Ca(2+) mobilization from the ER, and therefore amplifies GPCR-mediated signaling.

  3. Reversibility of gelsolin/actin interaction in macrophages. Evidence of Ca2+-dependent and Ca2+-independent pathways

    OpenAIRE

    1987-01-01

    We have developed an immunoadsorption technique for quantitating EGTA- resistant gelsolin/actin complexes in macrophages extracted with Triton X-100. We report here that the proportion of gelsolin complexed irreversibly to actin is low in freshly harvested macrophages. The amount of the EGTA-resistant complex increases spontaneously during incubation of the cells in suspension at 37 degrees C, or after exposure to the Ca2+ ionophore ionomycin. On the other hand, exposure of suspended cells to...

  4. CA2+-DEPENDENT AND CA2+-INDEPENDENT MECHANISM OF CYCLIC-AMP REDUCTION - MEDIATION BY BRADYKININ B-2 RECEPTORS

    NARCIS (Netherlands)

    SIPMA, H; DENHERTOG, A; NELEMANS, A

    1995-01-01

    1 Bradykinin caused a transient reduction of about 25% in the cyclic AMP level in forskolin prestimulated DDT1 MF-2 smooth muscle cells (IC50: 36.4 +/- 4.9 nM) and a pronounced, sustained inhibition (40%) of the isoprenaline-stimulated cyclic AMP level (IC50: 37.5 +/- 1.1 nM). 2 The Ca2+ ionophore,

  5. The solution structure of the Mg2+ form of soybean calmodulin isoform 4 reveals unique features of plant calmodulins in resting cells

    OpenAIRE

    Huang, Hao; Ishida, Hiroaki; Vogel, Hans J.

    2010-01-01

    Soybean calmodulin isoform 4 (sCaM4) is a plant calcium-binding protein, regulating cellular responses to the second messenger Ca2+. We have found that the metal ion free (apo-) form of sCaM4 possesses a half unfolded structure, with the N-terminal domain unfolded and the C-terminal domain folded. This result was unexpected as the apo-forms of both soybean calmodulin isoform 1 (sCaM1) and mammalian CaM (mCaM) are fully folded. Because of the fact that free Mg2+ ions are always present at high...

  6. The carcinogen safrole increases intracellular free Ca2+ levels and causes death in MDCK cells.

    Science.gov (United States)

    Chen, Wei-Chuan; Cheng, He-Hsiung; Huang, Chun-Jen; Lu, Yih-Chau; Chen, I-Shu; Liu, Shiuh-Inn; Hsu, Shu-Shong; Chang, Hong-Tai; Huang, Jong-Khing; Chen, Jin-Shyr; Jan, Chung-Ren

    2007-02-28

    The effect of the carcinogen safrole on intracellular Ca2+ movement in renal tubular cells has not been explored previously. The present study examined whether safrole could alter Ca2+ handling in Madin-Darby canine kidney (MDCK) cells. Cytosolic free Ca2+ levels ([Ca2+]i) in populations of cells were measured using fura-2 as a fluorescent Ca2+ probe. Safrole at concentrations above 33 microM increased [Ca2+]i in a concentration-dependent manner with an EC50 value of 400 microM. The Ca2+ signal was reduced by 90% by removing extracellular Ca2+, but was not affected by nifedipine, verapamil, or diltiazem. Addition of Ca2+ after safrole had depleted intracellular Ca(2+)-induced dramatic Ca2+ influx, suggesting that safrole caused store-operated Ca2+ entry. In Ca(2+)-free medium, after pretreatment with 650 microM safrole, 1 microM thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor) failed to release more Ca 2+. Inhibition of phospholipase C with 2 microM U73122 did not affect safrole-induced Ca2+ release. Trypan blue exclusion assays revealed that incubation with 650 microM safrole for 30 min did not kill cells, but killed 70% of cells after incubation for 60 min. Collectively, the data suggest that in MDCK cells, safrole induced a [Ca2+] increase by causing Ca2+ release from the endoplasmic reticulum in a phospholipase C-independent fashion, and by inducing Ca2+ influx via store-operated Ca2+ entry. Furthermore, safrole can cause acute toxicity to MDCK cells.

  7. Voltage-dependent Ca2+ channels, not ryanodine receptors, activate Ca2+-dependent BK potassium channels in human retinal pigment epithelial cells

    OpenAIRE

    Wimmers, Sönke; Halsband, Claire; Seyler, Sebastian; Milenkovic, Vladimir; Strauß, Olaf

    2008-01-01

    Purpose In different tissues the activation of large conductance Ca2+-activated (BK) potassium channels has been shown to be coupled to voltage-gated Ca2+ channels as well as ryanodine receptors. As activation of BK channels leads to hyperpolarization of the cell, these channels provide a negative feedback mechanism for Ca2+-induced functions. Many cellular functions of the retinal pigment epithelium (RPE) are coupled to changes in [Ca2+]i. The aim of this study was to identify which Ca2+-ent...

  8. Mitochondrial Ca2+ uniporter is critical for store-operated Ca2+ entry-dependent breast cancer cell migration

    International Nuclear Information System (INIS)

    Metastasis of cancer cells is a complicated multistep process requiring extensive and continuous cytosolic calcium modulation. Mitochondrial Ca2+ uniporter (MCU), a regulator of mitochondrial Ca2+ uptake, has been implicated in energy metabolism and various cellular signaling processes. However, whether MCU contributes to cancer cell migration has not been established. Here we examined the expression of MCU mRNA in the Oncomine database and found that MCU is correlated to metastasis and invasive breast cancer. MCU inhibition by ruthenium red (RuR) or MCU silencing by siRNA abolished serum-induced migration in MDA-MB-231 breast cancer cells and reduced serum- or thapsigargin (TG)-induced store-operated Ca2+ entry (SOCE). Serum-induced migrations in MDA-MB-231 cells were blocked by SOCE inhibitors. Our results demonstrate that MCU plays a critical role in breast cancer cell migration by regulating SOCE. - Highlights: • MCU is correlated to metastasis and invasive breast cancer. • MCU inhibition abolished serum-induced migration in MDA-MB-231 breast cancer cells and reduced serum- or TG-induced SOCE.