Calmodulin and ATP dependence of the high and low calcium affinity p-nitrophenylphosphatase from human erythrocyte membranes

In calmodulin depleted membranes from human erythrocytes, the Ca2+-dependent phosphatase showed different sensitivity to calmodulin and ATP with variable affinity towards free calcium concentrations: a calmodulin-dependent activity with high calcium affinity, K1/2 = 1.2 X 10(-7) mol/l calcium, that was fully activated at submicromolar calcium concentrations, higher concentrations being rather inhibitory; an ATP-dependent activity with lower calcium affinity, K1/2 = 10(-6) mol/l calcium, that was fully activated at 10(-5) mol/l calcium in the presence of 50-200 mumol/l ATP and was insensitive to calmodulin, and a calcium dependent phosphatase that was active at a wider ranger of free calcium, 10(-8)-10(-5) mol/l, and required the presence of both calmodulin and ATP.     

The Effect of Calcium Nutrition of Ethylene-induced Abscission

The influence of calcium nutrition on ethylene-induced abscission was studied by growing cotton (Gossypium hirsutum L. cv. Stoneville 213) and bean (Phaseolus vulgaris L. cv. Resistant Black Valentine) plants for several weeks in nutrient solutions containing 2, 10 (normal level), 15, or 20 meq/l of calcium, and then treating the plants with ethylene. Increasing the calcium level of cotton from 2 to 20 meq/l resulted in a 9-fold increase in the calcium content of the abscission zone and a maximum reduction of 25% in the amount of leaf abscission induced by ethylene (9 μl/l). Bean plants grown on 10, 15, or 20 meq/l calcium solutions showed corresponding increases in the calcium content of the abscission zone but showed no significant differences in the rate of ethyleneinduced abscission. Only at the lowest calcium level of 2 meq/l, where deficiency symptoms became apparent, was a significant effect observed. These results suggest that under normal cultural practices calcium nutrition has little influence on the rate of ethylene-induced abscission.     

Enhancement of intracellular calcium concentration by extracellular ATP and UTP in Madin Darby Canine Kidney cells

Fura2 - fluorescence was utilized to test for the effect of extracellular nucleotides on intracellular calcium concentration of subconfluent Madin-Darby Canine Kidney (MDCK)-cells. Extracellular ATP (10 mumol/l) and UTP (10 mumol/l) lead to rapid (within seconds), sustained, and fully reversible enhancement of intracellular calcium concentration from 138 +/- 9 nmol/l (n = 27), to 1561 +/- 260 nmol/l (n = 10) and 3435 +/- 949 nmol/l (n = 5), respectively. Half maximal effects are observed at some 1 mumol/l. In the absence of extracellular calcium the effect of ATP is transient, pointing to release of intracellular calcium. The sustained effect in the presence of extracellular calcium indicates that the nucleotides in addition recruit calcium from extracellular space.     

Analysis of calcium concentration fluctuations in hepatopancreatic R cells of Marsupenaeus japonicus during the molting cycle

In this study we examined the fluctuations of the intracellular calcium concentration in isolated hepatopancreatic R cells during the four molting stages of the prawn Marsupenaeus japonicus. In addition, we used the Fura-2-AM fluorescence technique to investigate the release of calcium from mitochondria and ATP-sensitive calcium stores (endoplasmic reticulum (ER), Golgi, and nucleus) into cytoplasm during the molting cycle. Results demonstrate that both the cytosolic free calcium concentration and the total cell calcium (free, bound to calcium-binding proteins, and stored in amorphous form) in the R cells strictly depend upon the molting cycle. Interestingly, the total cell calcium was higher (approximately 10 mmol l(-1)) in postmolt than in premolt (approximately 1 mmol l(-1)) and intermolt (approximately 0.3 mmol l(-1)). The calcium released from mitochondria was higher during premolt than during postmolt and intermolt, but the amount of calcium released from ATP-sensitive calcium stores was similar during all four stages. All together, our results suggest that the mitochondria-ATP-sensitive calcium stores system does not play a key role in calcium storage during the molting cycle but that it is involved in transcellular calcium flux. We hypothesize that lysosome or membrane-clad concretion vacuoles could represent the main site of calcium storage in hepatopancreatic R cells.     

Endothelin-Stimulated Capacitative Calcium Entry in Enteric Glial Cells: Synergistic Effects of Protein Kinase C Activity and Nitric Oxide

Abstract: Depletion of intracellular calcium stores by agonist stimulation is coupled to calcium influx across the plasma membrane, a process termed capacitative calcium entry. Capacitative calcium entry was examined in cultured guinea pig enteric glial cells exposed to endothelin 3. Endothelin 3 (10 n M ) caused mobilization of intracellular calcium stores followed by influx of extracellular calcium. This capacitative calcium influx was inhibited by Ni²⁺ (89 ± 2%) and by La³⁺ (78 ± 2%) but was not affected by L-, N-, or P-type calcium channel blockers. Chelerythrine, a specific antagonist of protein kinase C, dose-dependently inhibited capacitative calcium entry. The nitric oxide synthase inhibitor N ^G-nitro- l -arginine decreased calcium influx in a dose-dependent manner. The combination of chelerythrine and N ^G-nitro- l -arginine produced synergistic inhibitory effects. Capacitative calcium entry occurs in enteric glial cells via lanthanum-inhibitable channels through a process regulated by protein kinase C and nitric oxide.     

Utilization of the coconut shell of babaçu (Orbignya sp.) to produce cement-bonded particleboard

The zinc biosorptive capacity of the brown seaweed Sargassum sp. (Phaeophyceae) was studied in the presence or absence of competing calcium ions, using a continuous system with tubular fixed-bed reactors. In order to detect the effect of calcium on zinc biosorption, a 130 mg/l zinc solution was used, and calcium was added at 50-340 mg/l. The potential zinc biosorptive capacity of the biomass was markedly influenced by the presence of ionic calcium. Zinc sorption decreased with increasing calcium concentrations, as expressed by zinc uptake rates. Calcium was effectively recovered only during the initial stages of the process, as expressed by the decrease in its uptake rates. Calcium uptake rates were also much higher than zinc uptake rates, indicating that calcium was preferentially recovered when compared to zinc.     

Arachidonic acid and calcium metabolism in rnelittin stimulated neutrophils

Melittin, the predominant fraction of bee venom proteins, was studied in an experimental model of human neutrophil granulocytes to reveal its influence on eicosanoid release, metabolism and receptor function in relation to intracellular calcium metabolism. Melittin (2 mumol/l) was as potent as the calcium ionophore A23187 (10 mumol/l) for activation of 5-lipoxygenase, releasing arachidonate only from phosphatidyl-choline and phosphatidyl-ethanolamine of cellular membranes, as judged from the decreases in radioactivity by 15.4% and 30.5%, respectively. The mechanism responsible for the release of arachidonate from cellular membranes is closely coupled to cellular calcium metabolism, and melittin was found to promote calcium entry through receptor gated calcium channels, probably due to an activation of phospholipase A(2). Furthermore, a down-regulation of leukotriene B(4) receptors was seen. The maximal number of binding sites per cell was reduced from a median of 1520 to 950 with melittin (1 mumol/l). The study has revealed some factors important for the inflammatory mechanisms mediated by melittin.     

Calcium export by sodium-calcium exchange in bovine chromaffin cells

Calcium efflux from bovine chromaffin cells in tissue culture has been examined after loading them with small amounts of Ca2+ by brief depolarization in media containing 20 mumol/l to 1 mmol/l Ca2+ and 45Ca2+ in trace amounts. In the presence of normal external Na+ and Ca2+ concentrations cells depolarized in media containing up to 200 mumol/l Ca2+ exported nearly 100% of their accumulated Ca2+ loads within 10 min and 20% within the first 5 s. In the absence of external Na+ and Ca2+ the proportion of a small (i.e., depolarization in 20 mumol/l calcium) Ca2+ load exported at any time point in the range to 10 min was approximately two thirds of the total efflux measured in their presence indicating that under these conditions the external Na+/Ca(2+)-dependent and Na+/Ca(2+)-independent mechanisms both contribute significantly to the export of calcium. At higher cellular loads of calcium (i.e., depolarization in 200 mumol/l to 1 mmol/l calcium) the Na+/Ca(2+)-dependent mechanism exported a progressively greater proportion of the accumulated Ca2+. Both sodium and calcium alone promoted a component of Ca2+ efflux; Ca2+ (i.e. calcium-calcium exchange) was as effective as Na+ (i.e. sodium-calcium exchange). The Km for Na+ stimulation of Ca(2+)-efflux (KNa) was approximately 65 mM. Increased external Mg2+ (from 1.2 to 10 mmol/l) increased the apparent KNa to 90 mM.(ABSTRACT TRUNCATED AT 250 WORDS)     

Electrogenic proton pump in Nitella and the effect of calcium

The hyperpolarisation of the membrane potential in Characeae above that of the diffusion potential is explained by the operation of the electrogenic proton pump. We studied the interaction of calcium with the functioning of the pump. The membrane potential was measured using the standard microelectrode technique. An increase in the calcium concentration resulted in depolarisation, its magnitude increasing with lower proton concentrations. Calcium-induced membrane potential changes, tested in the concentration range of 0.25 mmol/l to 25 mmol/l, were greatest at 0.25 mmol/l CaCl2 and decreased with the increasing calcium concentration. Light-induced initial changes in the membrane potential also showed a dependence on the presence of calcium in the external medium. We conclude that calcium has a role in the regulation of the proton pump in Nitella.     

Calcium malate overproduction by Penicillium viticola 152 using the medium containing corn steep liquor

In this study, after screening of eight fungal strains for their ability to produce calcium malate, it was found that Penicillium viticola 152 isolated from marine algae among them could produce the highest titer of calcium malate. At the same time, it was found that corn steep liquor (CSL) could stimulate calcium malate production and 0.5 % (v/v) CSL was the most suitable for calcium malate production. Under the optimal conditions, a titer of calcium malate in the supernatant was 132 g/l at flask level. During a 10-l fermentation, a titer of 168 g/l, a yield of 1.28 g/g of glucose, and a productivity of 1.75 g/l/h were reached within 96 h of the fermentation, and 93.4 % of the sugar was used for calcium malate production and cell growth, demonstrating that the titer, yield, and productivity of calcium malate by this fungal strain were very high and the fermentation period was very short. After analysis of the partially purified product with high-performance liquid chromatography, it was found that the main product was calcium malate. The results demonstrated that P. viticola 152 obtained in this study was the most suitable for developing a novel one-step fermentation process for calcium malate production from glucose on a large scale.     

Calcium as a limiting factor in the biology of Biomphalaria pfeifferi (Krauss), (Gastropoda: Planorbidae)

The minimum calcium requirements, relative importance of buffering and optimum ratio of calcium to magnesium, calcium to sodium, and calcium to potassium ions were determined for laboratory populations ofBiomphalaria pfeifferi and related to suggested limiting factors for the natural distribution of this species. Snails were reared in a range of concentrations of both calcium bicarbonate and unbuffered calcium sulphate from 0.5 to 20 mg/l as Ca⁺⁺ and also in a series of media with a constant concentration of 2 mg/l as Ca⁺⁺ but with a range of Ca/Mg, Ca/Na and Ca/K ratios of 4.0 to 0.1. Shell growth, survivorship, fecundity, egg fertility, and the net reproductive rate were compared. In calcium bicarbonate cultures a concentration of 2mg/l Ca⁺⁺ appeared to be the lower limit for the survival of laboratory populations but a concentration of 4 mg/l Ca⁺⁺ was needed for a population to thrive. The calcium sulphate salt gave much poorer results, emphasizing the importance of the bicarbonate buffer. In the cationic ratio experiments the low Ca/Mg ratios proved to have the most damaging effects on snail populations but the effects of very low Ca/Na and Ca/K ratios could also be measured. A parallel experiment on the hatching rate of snail eggs, using similar experimental solutions, gave comparable results. The significance of these findings to snail ecology is discussed.     

Effect of single high dose infusions of aminohydroxypropylidene diphosphonate on hypercalcaemia caused by cancer.

Single intravenous infusions of 30 mg aminohydroxypropylidene diphosphonate were given to 16 patients who had malignant hypercalcaemia to assess host tolerance and the effect on serum calcium concentration. Ten of these patients also received intravenous rehydration or corticosteroids, or both. The serum calcium concentrations decreased significantly after treatment with aminohydroxypropylidene diphosphonate. Ten patients became normocalcaemic (normal range, adjusted for serum albumin, 2.25-2.75 mmol/l), two became hypocalcaemic, three showed decreases in serum calcium concentrations of more than 0.75 mmol/l, and one showed a decrease of more than 0.55 mmol/l. Only one patient had a minimum concentration greater than 2.77 mmol/l. Aminohydroxypropylidene diphosphonate was effective in metastatic and non-metastatic hypercalcaemia, and its hypocalcaemic effect was prolonged in some cases. There were no appreciable side effects. Single high dose infusions of aminohydroxypropylidene diphosphonate could replace conventional daily lower dose infusions, but the optimum frequency of high dose infusions remains to be determined.     

Kinetics of ethanol production by immobilized Kluyveromyces marxianus cells at varying sugar concentrations of Jerusalem artichoke juice

Summary Kinetics of ethanol fermentation at varying sugar concentrations of Jerusalem artichoke tuber extract has been studied using Kluyveromyces marxianus cells immobilized in calcium alginate gel beads. A maximum ethanol concentration of 111 g/l was achieved at an initial sugar concentration of 260 g/l in 20 hours, when the immobilized cell concentration in the calcium alginate beads was 53.3 g dry wt./l bead volume. Ethanol yield remained almost unaffected by initial sugar concentration up to 250 g/l and was found to be about 88% of the theoretical. Maximum rate of ethanol production decreased from 22.5 g ethanol/l/h to 10.5 g ethanol/l/h while the maximum rate of total sugars utilization decreased from 74.9 g sugars/l/h to 28.5 g sugars/l/h as the initial substrate concentration was increased from 100 to 300 g/l. The concentration of free cells in the fermentation broth was low.     

Evidence on the role of three calcium pools in Ca-ionophore A23187-stimulated rat blood platelet aggregation.

The effect of Ca-ionophore A23187 on activation of rat blood platelets was investigated to elucidate the involvement of extracellular and intracellular Ca2+ ions. Platelet aggregation induced by 10 concentrations of the stimulus was studied in Ca-free medium as well as in the presence of EGTA and/or calcium. In Ca-free medium, A23187 induced platelet aggregation in a dose-dependent way; the mean effective concentration was 1.43 +/- 0.08 mumol/l. The stimulatory effect of ionophore was potentiated by addition of 0.01 and 0.1 mM calcium and inhibited when the calcium concentration was increased to 1 mmol/l. In the presence of EGTA, A23187-stimulated aggregation of isolated rat platelets was recorded only at a 10-times higher ionophore concentration and was then reduced to 30% in comparison with aggregation in Ca-free medium. The inhibitory effect of 1 mM EGTA was abolished by addition of 2 mM calcium. We suggest the participation of at least three calcium pools in the stimulation of rat platelets by A23187, i.e. the extracellular pool, the membrane-associated pool and the pool displacing calcium intracellularly.     

Calciotropic hormones and lipolysis of human adipose tissue: role of extracellular calcium as conditioning but not regulating factor

The influences of different calcium concentrations (0, 0.924 and 2.772 mMol/l) on lipolysis of in vitro incubated human adipose tissue slices or adipocytes were studied under the conditions of stimulation with isoproterenol and parathyroid hormone preparations or inhibition by insulin. Extractive bovine PTH (as well as synthetic PTH 1--34) stimulated glycerol release in a biphasic pattern similarly to isoproterenol; PTH was about half as potent as isoproterenol. The optimal conditions for lipolysis were observed using a calcium concentration of 0.924 mMol/l, whereas lipolysis was distinctly impaired at concentrations of 0 or 2.772 mMol/l; this was true for basal as well as isoproterenol- and PTH stimulated lipolysis or the inhibitory effect of insulin. In contrast to partially purified extractive calcitonin, pure synthetic calcitonin did not inhibit lipolysis. Isoproterenol- and PTH-administrations led to cAMP accumulation in the adipose tissue, this process was also diminished at the non-optimal calcium concentrations. The results suggest a conditioning, but not a regulating significance of extracellular calcium for lipolysis, whereas the importance of the lipolytic potency of PTH remains to be elucidated.     

Elevation of calcium in rat enterocytes byEscherichia coli heat-stable (STa) enterotoxin

The effect of STa enterotoxin on intracellular calcium in primary cultures of rat enterocytes was investigated. The enterocytes were loaded with a fluorescent calcium indicator, indo-1, and changes of intracellular calcium determined by flow cytometry. When the rat enterocytes were treated with EGTA, a calcium chelator, or ionomycin, a calcium ionophore, a decrease or increase, respectively, in free intracellular calcium was demonstrated. The addition of STa enterotoxin to rat enterocytes loaded with indo-1 caused an initial decrease, followed by a rapid increase, in free intracellular calcium. These effects on intracellular calcium occurred prior to or during an increase in the intracellular concentration of cyclic 3,5-guanosine monophosphate. The current results demonstrate that the activation of rat enterocytes by STa enterotoxin has an effect on transmembrane regulation of the receptor-dependent calcium signal and/or the release of calcium from intracellular storage sites.     

Inotropic effect of chlorpromazine on the frog atrium

The chlorpromazine, a calmodulin inhibitor, has been studied for its action on the contraction force and calcium current of the frog atrium fibres. Chlorpromazine (10(-5) mol/l) was observed to induce maximal increase of the contraction force that 30 min after the agent action amounted to (47.3 + 9.3)% of control. The high concentration of chlorpromazine (10(-4) mol/l) produced irreversible decrease in the contraction force. Chlorpromazine (10(-5) mol/l) increased the calcium current by (27.5 +/- 4.8)%. It is supposed that chlorpromazine increases contraction force and calcium current through the inhibition of calmodulin-dependent phosphodiesterase activity.     

Kinetics of repeated batch production of ethanol by immobilized growing yeast cells

Summary Kinetic and yield parameters for growth and ethanol production from sucrose (100 g/l) bySaccharomyces cerevisia entrapped in K-carrageenan and calcium alginate were identical to those of free cells. Cell leakage was minimum with calcium alginate gel. For the sixth batch, 4.51 g/lh ethanol productivity (94% conversion of sucrose) was obtained; 60.5 g/l of ethanol was obtained from 200 g/l sucrose with 83.2% conversion, indicating inhibition effects.     

Calcium and renin release: inhibition of low sodium-induced renin secretion by high calcium concentration in rat kidney perfusion

The effects of changes in calcium on renin secretion have been studied in the isolated perfused rat kidney. Perfusion with free calcium buffer significantly decreases renin secretion as compared with control experiments (Ca++: 2.5 mM/l). Other calcium concentrations (1.25 mM/l) and 5 mM/l) do not affect basal renin secretion. When the renin release is previously increased by low sodium concentration (Na+: 110 mM/l) however, perfusion with high calcium buffer (Ca++: 5 mM/l) significantly inhibits this stimulation.     

The activity of calcium ions in aqueous solutions of the lower calcium oligogalacturonates

The activity of calcium ions in aqueous solutions of calcium mono-, di-, tri-, and tetra-galacturonates was determined by a spectrophotometric method in which tetramethylmurexide was used as an auxiliary ligand. The concentration of the solutions was 3.00 mequiv./l with respect to [-COOCa_0.5]. In this system, the activity of calcium ions in the solution of calcium galacturonate was very close to that of calcium chloride, while the activity of the calcium ions decreased in a continuous manner with increasing degree of polymerisation of the oligogalacturonate anion. The results are interpreted as evidence that, in calcium pectate, an intramolecular chelate bond of calcium involving two consecutive galacturonic acid units is not very probable.