Angiotensin increases microsomal (Na−K-ATPase activity in several tissues

Angiotensin increased microsomal (Na⁺K⁺)-activated ATPase from rat hypothalamus, mucosa of colon and bovine adrenal cortex but not brain cortex. Angiotensin did not change significantly Mg²⁺-ATPase in these tissues. Enhancement by angiotensin was evident at concentrations of 10⁻⁸ − 10⁻¹² M. In bovine adrenal cortex angiotensin increased ATPase activity in the outer and not in the inner layer. Angiotensin increased ATPase activity with Na⁺ above 20 mM and K⁺ above 5 mM present simultaneously, but not with either ion alone.     

Relationship of Na+-K+-activated ATPase to fluid production by Malpighian tubules of Locusta migratoria.

The presence of a Na⁺K⁺-activated, Mg²⁺-dependent ATPase (E.C. 3.6.1.3) has been demonstrated in microsomal preparations from the Malpighian tubules of Locusta. The effects of sodium and potassium ions, and different concentrations of ouabain, have been studied in relation to the activity of this enzyme and the ability of in vitro Malpighian tubule preparations to secrete fluid. From these studies it seems highly likely that a Na⁺K⁺ activated ATPase ‘pump’ is involved in fluid transport across the walls of the tubules.     

Ageing in the honey-bee, Apis mellifera, as related to brain ATPases and their DDT sensitivity

The adenosine triphosphatase (ATPase) system in worker honey-bee brains showed an increased activity of 57 per cent in Na⁺K⁺ATPase and 63 per cent in Mg²⁺ATPase from adult emergence to 7 days post-emergence. Mg²⁺ATPase activity remained about the same throughout the remainder of adult life, while Na⁺K⁺ATPase remained the same until the sixth week, when a decline occurred. The percentage mortality of the bees exceeded 90 per cent at the time of decline of Na⁺K⁺ATPase. The in vitro inhibition of Mg²⁺ATPase and Na⁺K⁺ATPase by 10 μM DDT was between 40 and 50 per cent and about 20 per cent, respectively. A somewhat greater sensitivity to DDT was determined in brains of older honey-bees.     

Properties of (Na+ + K+)-activated ATPase in rat liver plasma membranes enriched with bile canaliculi

Liver plasma membranes enriched in bile canaliculi were isolated from rat liver by a modification of the technique of Song et al. (J. Cell Biol. (1969) 41, 124–132) in order to study the possible role of ATPase in bile secretion. Optimum conditions for assaying (Na⁺ + K⁺)-activated ATPase in this membrane fraction were defined using male rats averaging 220 g in weight. (Na⁺ + K⁺)-activated ATPase activity was documented by demonstrating specific cation requirements for Na⁺ and K⁺, while the divalent cation, Ca²⁺, and the cardiac glycosides, ouabain and scillaren, were inhibitory. (Na⁺ + K⁺)-activated ATPase activity averaged 10.07 ± 2.80 μmol P_i/mg protei per h compared to 50.03 ± 11.41 for Mg²⁺-activated ATPase and 58.66 ± 10.07 for 5′-nucleotidase. Concentrations of ouabain and scillaren which previously inhibited canalicular bile secretion in the isolated perfused rat liver produced complete inhibition of (Na⁺ + K⁺)-activated ATPase without any effect on Mg²⁺-activated ATPase. Both (Na⁺ + K⁺)-activated ATPase and Mg²⁺-activated ATPase demonstrated temperature dependence but differed in temperature optima. Temperature induced changes in specific activity of (Na⁺ + K⁺)-activated ATPase directly paralleled previously demonstrated temperature optima for bile secretion. These studies indicate that (Na⁺ + K⁺)-activated ATPase is present in fractions of rat liver plasma membranes that are highly enriched in bile canaliculi and provide a model for further study of the effects of various physiological and chemical modifiers of bile secretion and cholestasis.     

Sensitivity of ATPase activities to fire ant venom and abdomen preparations.

The sensitivity of fire ant, $\text{Solenopsis richteri}$ (Forel), head homogenate ATPase to its venom and to a cyclohexane extract of whole fire ants were investigated. Na⁺K⁺ and oligomycin-sensitive Mg²⁺ ATPase activities were inhibited by both preparations. Oligomycin-insensitive Mg²⁺ ATPase activity was inhibited by low concentrations but showed strong stimulation at high concentrations of the venom preparations. Lineweaver-Burk plots of enzyme data in the presence or absence of inhibitor indicated that the inhibitor action was non-competitive with ATP for Na⁺K⁺ and oligomycin-sensitive Mg²⁺ ATPase activities. However, the oligomycin-insensitive Mg²⁺ ATPase activity showed a mixed type response to the inhibitor. Tests on pure samples of known venom components indicate that they cause the observed effects on the ATPase activities.     

Purification and properties of adenosinetriphosphatase from Zea mays seedling microsomes

A simple method was developed for selective solubilization of membrane ATPase from etiolated corn seedlings using 0.01% Triton X100 and 0.01% deoxycholate containing 200 mM KI. An 81-fold enriched enzyme preparation, with specific activity of 133 μmol Pi/mg protein/hr, was obtained. The enzyme stored in 25 mM Tris-HCl buffer (pH 7.5) at 4° showed rapid loss of activity. The enzyme was stabilized by 1 mM EDTA with addition of 1.2 mM Mg²⁺°. Mg²⁺ and Ca²⁺ (1.2 mM) increased enzymatic activity by 12 and 10.8% respectively, whereas Na⁺ and K⁺ brought about a 20% increase in ATP-hydrolysis. The effect of combined mono- and di-valent ions was neither synergistic nor additive. Ouabain exerted no effect on enzyme activity. The enzyme showed two pH optima (6.0 and 7.5) in the presence of Na⁺ and K⁺, and one optimum at pH 6.5 in the absence of these ions. On polyacrylamide gel the enzyme was resolved into two protein bands, both exhibiting ATPase activity. It is suggested that the soluble enzyme from the microsomal fraction of corn seedlings contains two ATP-hydrolyzing enzymes, one of them being stimulated by Na⁺ and K⁺ ions.     

ATPases in rabbit erythrocytes: stimulation by HCO3-anion and by Na-cation-plus-K-cation.

A Mg²⁺Na⁺K⁺ATPase was found in a ghost preparation from rabbit erythrocytes, a finding in conflict with previous reports, but in agreement with the known kinetics of cation movements in these cells. However the Mg²⁺Na⁺K⁺ATPase was not inhibited by 10⁻⁴M ouabain, nor by 10⁻⁴M Ca²⁺. The physiological status of this enzyme is discussed. The basic Mg²⁺-ATPase activity in this preparation is also stimulated by HCO₃⁻; it is suggested that the HCO₃⁻-stimulated ATPases reported in a variety of other preparations are not necessarily due to mitochondrial contamination but could well originate from the plasma membrane.     

Variable ATPase composition of human tumor plasma membranes

Purified plasma membranes from several transplantable human tumors exhibit very high Mg²⁺-dependent ATPase activities. Three types of Mg²⁺-dependent ATPases can be demonstrated: (1) an ouabain sensitive Na⁺, K⁺-ATPase, which is a minor component of the tumor plasma membrane ATPase, (2) a Mg²⁺-activated ATPase, which is a non-specific nucleoside triphosphatase, and (3) an ATPase activity stimulated by Na⁺ (or K⁺) alone. In three human melanomas, only the first two activities are found. In an astrocytoma and an oat cell carcinoma, all three activities are found. In the same two tumors, the plasma membrane Mg²⁺-ATPase is also stimulated by Con A. The relationship of these ATPases are discussed.     

Adenosine Triphosphatases Bound to Plasmalemma, Mitochondria, and Microsomes or Present in the Cytoplasmic Phase from Potato Tubers

Between pH 4–10, basal ATPase activity, measured in the absence of mineral ions, was 10 to 100 times higher in the final cytoplasmic supernatant from potato tuber homogenates than in the membraneous fractions (purified plasmalemma, purified mitochondria and microsomes). The soluble ATPase was slightly inhibited, whereas the membrane-bound ATPases were all stimulated by Mg²⁺ ions. A further stimulation by Na⁺ or K⁺ ions was only observed in purified plasmalemma or mitochondria, at alkaline pH (7.5–9.5). At a fixed (Na⁺+ K⁺) concentrations (80 mM), this last stimulation was much greater in purified mitochondria (350%) than in plasmalemma (33%); it also increased with (Na⁺+ K⁺) concentrations up to 200 mM in mitochondria whereas, in plasmalemma, it was roughly constant for monovalent ion concentrations between 20 and 200 mM. General properties of the plasma membrane-bound ATPase have been determined, i.e. substrate specificity, activity variations with quantity of substrate, temperature, pH, etc. Divalent cations stimulated strongly the ATPase in the following order: Mn²⁺ > Mg²⁺ > Ca²⁺. The maximum ATP hydrolysis velocity for that part of ATPase activity which is strictly dependent on Mg²⁺ ions was 3.85 μmol × mg^?1 protein × h^?1. This plasma membrane ATPase was not sensitive to ouabaïn or to oligomycin.     

Properties and nature of (Na+ + Mg2+)-dependent adenosine triphosphatase in the gills of the eel, angvilla anguilla (L.)

The specific activity of (Na⁺ + Mg²⁺)-dependent ATPase is three times greater in the microsomes of sea-water eels than in freshwater eels; the specific activity is one quarter of that of (Na⁺ + K⁺ + Mg²⁺)-dependent ATPase in both cases.(Na⁺ + Mg²⁺)-dependent ATPase is optimally active in a medium containing 8 mM NaCl, 4 mM MgCI₂, 4 mM ATP, pH 8.8 and at 30 °C; the enzyme is inhibited by ouabain, by NaCl concentrations > 100 mM and by treatment with urea.It is concluded that the (Na⁺ + Mg²⁺)-dependent ATPase activity of gills arises from the presence of a (Na⁺ + K⁺ + Mg²⁺)-dependent ATPase.     

Effects of some volatile anesthetic agents on rat heart sarcolemma.

The effects of ether, chloroform and halothane on rat heart sarcolemmal ATP hydrolyzing and calcium binding activities were studied. Sarcolemmal Na⁺ ? K⁺ ATPase activity was inhibited by halothane (1.8 – 18 mM) and stimulated by ether (7.1 – 42.6 mM) and chloroform (7.5 – 45 mM). Higher concentrations of ether (56.8 – 71 mM) and chloroform (60 – 75 mM) depressed the Na⁺ ? K⁺ ATPase activity. Chloroform (7.5 – 75 mM) and halothane (1.8 – 18 mM) were found to decrease Mg²⁺ ATPase and Ca²⁺ ATPase activities, whereas e0her (42.6 – 71 mM) depressed only the Mg²⁺ ATPase activity. Sarcolemmal calcium binding was depressed by ether (42.6 – 71 mM), chloroform (45 – 75 mM) and halothane (10.8 – 18 mM). These results suggest that the anesthetic - induced cardiac depression may partly be due to decreased sarcolemmal activities.     

Quantitative aspects of the interaction between ouabain and (Na + K)-activated ATPase in vitro

The inhibitory effect of ouabain on (Na⁺ + K⁺)-activated ATPase (Mg²⁺-dependent, (Na⁺ + K⁺)-activated ATP phosphohydrolase, EC 3.6.1.3) obtained from rat brain microsomal fraction was re-examined using a modified method to estimate the inhibited reaction velocity. This method involves a preincubation of a ouabain-enzyme mixture in the presence of Na⁺, Mg²⁺ and ATP to bring the ouabain-enzyme reaction to near equilibrium. The (Na⁺ + K⁺)-activated ATPase reaction was subsequently started by the addition of a KCl solution.     

Stimulation of (Na+K+)-ATPase of rat liver plasma membrane by amino acids

The effect of twelve l-amino acids on the activity of liver plasma membrane (Na⁺K⁺)-ATPase has been tested. Histidine and arginine significantly enhanced the activity. The activtion by histidine showed saturation kinetics with an apparent K_a of about 8 mM, and was evident over a wide range of Na⁺ concentrations. The same amino acid did not significantly affect the Mg²⁺-dependent ATPase activity.     

Cyclic AMP and calcium modulated ATPase activity in the salivary glands of the lone star tick Amblyomma americanum (L.)

Na⁺,K⁺-activated ATPase activity in tick salivary glands increases during the rapid stage of tick feeding paralleling similar increases in dopamine and cAMP-stimulated fluid secretion. High concentrations of cyclic AMP increase Na⁺,K⁺-ATPase activity in a plasma membrane-enriched fraction from the salivary glands of rapidly feeding ticks. Cyclic AMP-dependent protein kinase inhibitor protein blocks activation of Na⁺,K⁺-ATPase activity at low but not high concentrations of cAMP indicating that both activator and inhibitor modulator phosphoproteins of Na⁺,K⁺-ATPase activity exist in the plasma membrane-enriched fraction.ATPase activity in the plasma membrane-enriched fraction is not measurable in the absence of Mg²⁺, Ca²⁺ and Na⁺. Ca-stimulated nucleotidase activity is highest with ATP serving as the preferred substrate in a series including CTP, UTP, GTP and ADP. Calcium, Mg²⁺ stimulated ATPase activity is activated further by calmodulin and partially inhibited by low concentration of vanadate, trifluoperazine and oligomycin. Results suggest that the plasma membrane-enriched fraction of tick salivary glands contains both Ca²⁺-ATPase activity and oligomycin-sensitive Ca²⁺, Mg²⁺-ATPase activities, the latter likely from a small amount of mitochondria in the partially purified organelle fraction.     

Partial characterization of an endogenous factor which modulates the effect of catecholamines on synaptosomal Na+, K+-ATPase

We have previously presented evidence for the existence of a brain soluble factor which mediates the stimulation of synaptosomal ATPases by catecholamines. The stimulation of synaptosomal ATPases by dopamine plus brain soluble fraction was not modified if the soluble fraction was heated for 5 min at 95°C. One day after preparation, the soluble factor inhibited the Na⁺, K⁺-ATPase, but not the Mg²⁺-ATPase activity, and subsequent addition of noradrenaline stimulated the ATPases activities. The inhibitory effect of a 24 h soluble fraction disappeared if the soluble fraction was dialyzed; in this case, noradrenaline did not activate the enzyme activities. Gel filtration in Sephadex G-50 permitted separating a subfraction which inhibited ATPase activity (peak II) from another which stimulated ATPase activity (peak I). Peak I stimulated both Na⁺, K⁺, and Mg²⁺ ATPases. Peak II inhibited only Na⁺, K⁺-ATPase, and when stored acidified, it mediated ATPases stimulation by noradrenaline.Special Issue dedicated to Prof. Eduardo De Robertis.     

Fluid transport, ATP level and ATPase activities in isolated rabbit corneal endothelium

The effect of certain biochemical parameters on transendothelial fluid transport has been studied. Cellular ATP level and (Na⁺ + K⁺)-activated as well as Mg²⁺-activated ATPase activities were measured by ultramicrotechniques using individual rabbit corneal endothelium after they had been subjected to in vitro perfusion with solutions fully supplemented or deficient singly or severally in glucose, adenosine and glutathione (GSH). With the complete medium, the transport system operates in vitro for approx. 6 h. Deletion of glucose alone, glucose and adenosine or glucose, adenosine and GSH brings about a cessation of fluid transport after 3.5 h, 2 to 2.5 h and 0.5 to 1 h, respectively. A marked decrease (62%) of the endothelial ATP level, however, occurs only when all metabolites are omitted. The favorable effect of GSH on transport activity is attributable to its capacity to sustain cellular ATP rather than to protect the functionality of (Na⁺ + K⁺)-activated ATPase. Adenosine, in the presence of GSH, maintains normal ATP levels and, additionally, exerts a protective effect on Mg²⁺-activated ATPase and possibly also on (Na⁺ + K⁺)-activated ATPase.     

Direct stimulation of human erythrocyte membrane (Na+ + k+)-mg ATPase activity in vitro by physiological concentrations of d-aldosterone.

The effect of d-aldosterone on human erythrocyte ghost (Na⁺ + K⁺)-Mg ATPase has been studied. Aldosterone at 3.225 × 10^?10M caused a 450% activation of (Na⁺ + K⁺)-Mg ATPase activity whilst inhibiting (Na⁺ + Na⁺)-Mg ATPase activity. Aldosterone acts by reducing the affinity of the external K⁺ site of (Na⁺ + K⁺)Mg ATPase for Na⁺ thereby resulting in improved efficiency of Na⁺ ? K⁺ transfer. Aldosterone was additionally found to modify both the Na⁺ and K⁺ activation of (Na⁺ + K⁺)Mg ATPase incubated in the presence of commercial ATP containing orthovanadate. Aldosterone was found to reverse the inhibitory effects of orthovanadate at high Na⁺ and K⁺ concentrations. The physiological significance of orthovanadate and aldosterone are discussed.     

Effect of concanavalin A on membrane-bound enzymes from mouse lymphocytes

The ionic influence and ouabain sensitivity of lymphocyte Mg²⁺-ATPase and Mg²⁺-(Na⁺ + K⁺)-activated ATPase were studied in intact cells, microsomal fraction and isolated plasma membranes. The active site of 5′-nucleotidase and Mg²⁺-ATPase seemed to be localized on the external side of the plasma membrane whereas the ATP binding site of (Na⁺ + K⁺)-ATPase was located inside the membrane.Concanavalin A induced an early stimulation of Mg²⁺-ATPase and (Na⁺ + K⁺)-ATPase both on intact cells and purified plasma membranes. In contrast, 5′-nucleotidase activity was not affected by the mitogen. Although the thymocyte Mg²⁺-ATPase activity was 3–5 times lower than in spleen lymphocytes, it was much more stimulated in the former cells (about 40 versus 20 %). (Na⁺ + K⁺)-ATPase activity was undetectable in thymocytes. However, in spleen lymphocytes (Na⁺ + K⁺)-ATPase activity can be detected and was 30 % increased by concanavalin A. Several aspects of this enzymic stimulation had also characteristic features of blast transformation induced by concanavalin A, suggesting a possible role of these enzymes, especially Mg²⁺-ATPase, in lymphocyte stimulation.     

Effects of delta9-tetrahydrocannabinol on ATPases in mouse brain subcellular fractions.

The effect of (?)-Δ⁹-THC on the activities of Mg²⁺?, Na⁺?K⁺? and Mg²⁺Ca²⁺-ATPases were studied in mouse brain subcellular fractions. $\text{In vitro}$treatment with Δ⁹-THC produced a dose dependent stimulation of Mg²⁺ ATPase in the crude mitochondrial fraction and its subfractions and a dose-related inhibition of this activity in the microsomal fraction. Na⁺-K⁺- and Mg²⁺-Ca²⁺-ATPase activities were inhibited in a dose-related manner in all subcellular fractions studied.