Angiotensin II inhibits Na+/K+ATPase activity in pulmonary artery smooth muscle cells via glutathionylation and with the involvement of a 15.6 kDa inhibitor protein.

The role of angiotensin II in regulating Na+/K+-ATPase activity has been investigated in bovine pulmonary artery smooth muscle cells (BPASMCs). Our study reveals that angiotensin II inhibits the Na+/K+ATPase activity via glutathionylation of the pump with the involvement of an increase in NADPH oxidase-derived O2.-. Additionally, angiotensin II treatment to the cells increases the inhibitory potency of the 15.6 kDa inhibitor towards the Na+/K+ATPase activity.

Synergic actions of polyphenols and cyanogens of peanut seed coat (Arachis hypogaea) on cytological, biochemical and functional changes in thyroid.

In animals, long-term feeding with peanut (Arachis hypogaea) seed coats causes hypertrophy and hyperplasia of the thyroid gland. However, to date there have been no detailed studies. Here, we explored the thyroidal effects of dietary peanut seed coats (PSC) in rats. The PSC has high levels of pro-goitrogenic substances including phenolic and other cyanogenic constituents. The PSC was mixed with a standard diet and fed to rats for 30 and 60 days, respectively. Animals fed with the PSC-supplemented diet showed a significant increase in urinary excretion of thiocyanate and iodine, thyroid enlargement, and hypertrophy and/or hyperplasia of thyroid follicles. In addition, there was inhibition of thyroid peroxidase (TPO) activity, 5’-deiodinase-I (DIO1) activity, and (Na+-K+)-ATPase activity in the experimental groups of rats as compared to controls. Furthermore, the PSC fed animals exhibited decreased serum circulating total T4 and T3 levels, severe in the group treated for longer duration. These data indicate that PSC could be a novel disruptor of thyroid function, due to synergistic actions of phenolic as well as cyanogenic constituents.

A QSAR study on a series of simplified digitalis-like compounds acting on Na+,K+-ATPase

Among the cardiotonics (agents against congestive heart failure), the most important group is of the digitalis cardiac glycosides, but since these compounds suffer from a low therapeutic index, attention has been paid to investigating safer cardiotonic agents through the inhibition of Na+,K+-ATPase, the mechanism by which the digitalis cardiac glycosides elicit their action. Recently, a series of perhydroindenes were studied for their Na+,K+-ATPase inhibition activity. We report here a QSAR study on them to investigate the physicochemical and structural properties of the molecules that govern their activity in order to rationalize the structural modification to have more potent drugs. A multiple regression analysis reveals a significant correlation between the Na+,K+-ATPase inhibition activity of the compounds and Kier’s first order valence molecular connectivity index of their R5-substituents and some indicator parameters, suggesting that the R5-substituents of the compounds containing atoms with low valence and high saturation and the R1-substituents having =N−O− moiety will be conducive to the activity.

A low concentration of ouabain (0.18 µg/kg) enhances hypertension in spontaneously hypertensive rats by inhibiting the Na+ pump and activating the renin-angiotensin system

We investigated the effects of low ouabain concentrations on systolic (SAP) and diastolic (DAP) arterial pressures and on pressor reactivity in 3-month-old male spontaneously hypertensive rats (SHR). Arterial blood pressure (BP) and pressor reactivity to phenylephrine (PHE) were investigated before and after 0.18 ìg/kg ouabain administration (N = 6). The influence of hexamethonium (N = 6), canrenone (N = 6), enalapril (N = 6), and losartan (N = 6) on ouabain actions was evaluated. Ouabain increased BP (SAP: 137 ± 5.1 to 150 ± 4.7; DAP: 93.7 ± 7.7 to 116 ± 3.5 mmHg; P<0.05) but did not change PHE pressor reactivity. Hexamethonium reduced basal BP in control but not in ouabain-treated rats. However, hexamethonium + ouabain increased DAP sensitivity to PHE. Canrenone did not affect basal BP but blocked ouabain effects on SAP. However, after canrenone + ouabain administration, DAP pressor reactivity to PHE still increased. Enalapril and losartan reduced BP and abolished SAP and DAP responses to ouabain. Enalapril + ouabain reduced DAP reactivity to PHE, while losartan + ouabain reduced SAP and DAP reactivity to PHE. In conclusion, a small dose of ouabain administered to SHR increased BP without altering PHE pressor reactivity. Although the renin-angiotensin system (RAS), Na+ pump and autonomic reflexes are involved in the effects of ouabain on PHE reactivity, central mechanisms might blunt the actions of ouabain on PHE pressor reactivity. The effect of ouabain on SAP seems to depend on the inhibition of both Na+ pump and RAS, whereas the effect on DAP seems to depend only on RAS.

Characterization of arylsulphatase A in a 70 kDa protein isolated from goat spermatozoa having Na+, K+-ATPase inhibitory activity.

A protein having inhibitory effect on Na+, K+-ATPase as well as showing arylsulphatase A activity (ASA) was isolated from the cytosolic fraction of goat spermatozoa and characterized biochemically. The molecular mass of the protein was found to be 70 kDa (P70) on 10% SDS-PAGE after 35% ammonium sulphate precipitation, followed by hydroxyapatite column chromatographic separation. The isoelectric point (pI) of the protein was found to be 4.9. The sequencing results of first ten N-terminal amino acid residues of protein showed 100%, 90%, and 80% homology with N-terminal 18-27 amino acid residues of mice, pig and human testicular ASA, respectively. The optimum pH, temperature and incubation time for maximum ASA activity of the protein was 5.5, 37°C and 30 min respectively. The ASA activity of protein and AS from a commercial source was studied with respect to the sensitivity to different metal ions, vanadate, carbonyl compounds and ascorbate. Inhibition of AS activity of P70 by silver nitrate suggested that it was related to ASA. Comparable effects of different polyunsaturated fatty acids (eicosapentaenoic and docosahexaenoic acids) and purified anti P70-antibody on P70 and AS from commercial source were observed. The findings suggested that protein was novel in nature, having both regulatory and catalytic functions and showed similarities with the ASA reported from different sources.

Effect of methacrylonitrile on membrane bound enzymes of rat brain.

Methacrylonitrile (MeAN) is a plastic monomer. Its effect on membrane bound enzymes like Na+K+ -ATPase, Ca2+ -ATPase, Mg2+ -ATPase, NADH dehydrogenase, alkaline phosphatase (ALP) and various elements like sodium (Na+), potassium (K+), and calcium (Ca2+) in rat brain were studied. Administration of 50 mg/kg body weight/day (0.25 LD50) and 100 mg/kg body weight/day (0.5 LD50) by gavage to rats for 7 days resulted in a significant decrease in activities of Na+K+ -ATPase, Ca2+ -ATPase, Mg2+ -ATPase, and NADH dehydrogenase. A significant reduction in calcium content, potassium content and a significant increase in sodium content and alkaline phosphatase activity in MeAN treated animals were observed. Inhibition of membrane bound enzymes occurred due to either direct effect of MeAN or indirect effect of changes in ionic homeostasis in MeAN treated animals.

Amelioration of altered antioxidant status and membrane linked functions by vanadium and Trigonella in alloxan diabetic rat brains.

Trigonella foenum graecum seed powder (TSP) and sodium orthovanadate (SOV) have been reported to have antidiabetic effects. However, SOV exerts hypoglycemic effects at relatively high doses with several toxic effects. We used low doses of vanadate in combination with TSP and evaluated their antidiabetic effects on anti-oxidant enzymes and membrane-linked functions in diabetic rat brains. In rats, diabetes was induced by alloxan monohydrate (15 mg/100 g body wt.) and they were treated with 2 IU insulin, 0.6 mg/ml SOV, 5% TSP and a combination of 0.2 mg/ml SOV with 5% TSP for 21 days. Blood glucose levels, activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), Na+/K+ ATPase, membrane lipid peroxidation and fluidity were determined in different fractions of whole brain after 21 days of treatment. Diabetic rats showed high blood glucose (P less than 0.001), decreased activities of SOD, catalase and Na+/K+ ATPase (P less than 0.01, P less than 0.001 and P less than 0.01), increased levels of GPx and MDA (P less than 0.01 and P less than 0.001) and decreased membrane fluidity (P less than 0.01). Treatment with different antidiabetic compounds restored the above-altered parameters. Combined dose of Trigonella and vanadate was found to be the most effective treatment in normalizing these alterations. Lower doses of vanadate could be used in combination with TSP to effectively counter diabetic alterations without any toxic effects.

Inhibition of membrane Na(+)-K+ Atpase of the brain, liver and RBC in rats administered di(2-ethyl hexyl) phthalate (DEHP) a plasticizer used in polyvinyl chloride (PVC) blood storage bags.

Significant amounts of di(2-ethylhexyl) phthalate (DEHP) leach out into blood stored in DEHP plasticized polyvinyl chloride (PVC) bags resulting in the exposure of recipients of blood transfusion to this compound. The aim of this study was to find out whether DEHP at these low levels has any effect on the activity of membrane Na(+)-K+ ATPase, since a decrease in this enzyme activity has been reported to take place in a number of disorders like neurodegenerative and psychiatric disorders, coronary artery disease and stroke, syndrome-X, tumours etc. DEHP was administered (ip) at a low dose of 750 microg/100 g body weight to rats and the activity of membrane Na(+)-K+ ATPase in liver, brain and RBC was estimated. Histopathology of brain, activity of HMG CoA reductase (a major rate limiting enzyme in the isoprenoid pathway of which digoxin, the physiological inhibitor of Na(+)-K+ ATPase is a product), intracellular concentration of Ca2+ and Mg2+ in RBC (which is altered as a result of inhibition of Na(+)-K+ ATPase) were also studied. (In the light of the observation of increase of intracellular Ca2+ load and intracellular depletion of Mg2+ when Na(+)-K+ ATPase is inhibited). Histopathology of brain revealed areas of degeneration in the rats administered DEHP. There was significant inhibition of membrane Na(+)-K+ ATPase in brain, liver and RBC. Intracellular Ca2+ increased in the RBC while intracellular Mg2+ decreased. However activity of hepatic HMG CoA reductase decreased. Activity of Na(+)-K+ ATPase and HMG CoA reductase, however returned to normal levels within 7 days of stopping administration of DEHP. The inhibition of membrane Na(+)-K+ ATPase activity by DEHP may indicate the possibility of predisposing recipients of transfusion of blood or hemodialysis to the various disorders mentioned above. However since this effect is reversed when DEHP administration is stopped, it may not be a serious problem in the case of a few transfusion; but in patients receiving repeated blood transfusion as in thalassemia patients or patients undergoing hemodialysis, possibility of this risk has to be considered. This inhibition is a direct effect of DEHP or its metabolites, since activity of HMG CoA reductase, (an enzyme which catalyses a major rate limiting step in the isoprenoid pathway by which digoxin, the physiological inhibitor of Na(+)-K+ ATPase is synthesized) showed a decrease.

Effect of lead on Na+, K(+)-ATPase activity in Penaeus indicus postlarvae.

In vivo effect of lead on Na, K(+)-ATPase was studied in plasma membrane/mitochondrial fraction of P. indicus post-larvae (PL), exposed to 30 days to a sublethal concentration (1.44 ppm) of lead. A significant (P < 0.05) decrease in the enzyme activity was observed for exposed PL with respect to their controls at different intervals except 24hr. Further the substrate (ATP) and ion (Na+ and K+)-dependent kinetics of Na+, K(+)-ATPase was studied with the plasma membrane/mitochondrial fractions of control and 30 days exposed PL. The apparent KM and V(max). values were calculated to determine the nature of inhibition. Both the control and exposed PL showed almost the same apparent KM values in the presence of different substrate or ion concentrations indicating that lead interacts with the enzyme at a different binding site.

Endogenous sodium-potassium ATPase inhibition related biochemical cascade in trisomy 21 and Huntington's disease: neural regulation of genomic function.

The isoprenoid pathway related cascade was assessed in trisomy 21 and Huntington's disease. Membrane Na+-K+ ATPase activity, serum magnesium and ubiquinone were decreased while HMG CoA reductase activity, serum digoxin and dolichol levels were increased in both the disorders. There were increased levels of tryptophan catabolites (nicotine, strychnine, quinolinic acid and serotonin) and decreased levels of tyrosine catabolites (dopamine, noradrenaline and morphine) in both trisomy 21 and Huntington's disease. There was an increase in dolichol levels, carbohydrate residues of glycoproteins, glycolipids, total/individual GAG fractions and lysosomal enzymes in both disorders. Reduced levels of ubiquinone, reduced glutathione and free radical scavenging enzymes as well as increased lipid peroxidation products and nitric oxide were noticed in both the disorders. The role of hypothalamic digoxin and a disordered isoprenoid pathway in the pathogenesis of trisomy 21 and Huntington's disease is discussed.

Inhibition of membrane Na+-K+ ATPase activity: a common pathway in central nervous system disorders.

OBJECTIVES: The study was conducted to assess the role of hypothalamic digoxin in neuropsychiatric and systemic disorders. A hypothesis regarding the central role of hypothalamic digoxin in neuroimmunoendocrine integration is proposed. METHODOLOGY: Blood samples from patients of CNS glioma, multiple sclerosis, systemic lupus erythematosis, subacute sclerosing panencephalitis, primary generalized epilepsy, Parkinson's disease, Down syndrome, AIDS dementia with neuropsychiatric features, syndrome X with multiple lacunar state, senile dementia, familial group (a family with familial coexistence of schizophrenia, Parkinson's disease, primary generalized epilepsy, malignant neoplasia, rheumatoid arthritis and syndrome X over three generations), schizophrenia and manic depressive psychosis were analysed for RBC membrane Na+-K+ ATPase, levels of digoxin and Mg++. RESULTS: Inhibition of RBC membrane Na+-K+ ATPase activity was observed in most cases along with increase in the levels of serum digoxin and decrease in the level of serum Mg++. CONCLUSION: The decreased Na+-K+ ATPase activity can be due to increased digoxin, which is a potent inhibitor of this enzyme. The inhibition of Na+-K+ ATPase can contribute to increase in intracellular calcium and decrease in magnesium, which can result in 1) defective neurotransmitter transport mechanism, 2) neuronal degeneration and apoptosis, 3) mitochondrial dysfunction, 4) defective golgi body function and protein processing dysfunction, 5) immune dysfunction and oncogenesis. The mechanism of how increased intracellular calcium and decreased magnesium can contribute to the above effects is discussed.

Sodium pumps in the Malpighian tubule of Rhodnius sp

Malpighian tubule of Rhodnius sp. express two sodium pumps: the classical ouabain-sensitive (Na+ + K+)ATPase and an ouabain-insensitive, furosemide-sensitive Na+-ATPase. In insects, 5-hydroxitryptamine is a diuretic hormone released during meals. It inhibits the (Na+ + K+)ATPase and Na+ -ATPase activities indicating that these enzymes are involved in fluid secretion. Furthermore, in Rhodnius neglectus, proximal cells of Malpighian tubule exposed to hyperosmotic medium, regulate their volume through a mechanism called regulatory volume increase. This regulatory response involves inhibition of the (Na+ + K+)ATPase activity that could lead to accumulation of active osmotic solute inside the cell, influx of water and return to the normal cell volume. Adenosine, a compound produced in stress conditions, also inhibits the (Na+ + K+)ATPase activity. Taken together these data indicate that (Na+ + K+)ATPase is a target of the regulatory mechanisms of water and ions transport responsible for homeostasis in Rhodnius sp.

Biochemical profile of erythrocyte membrane of jaundiced neonates.

Studies in newborn humans have demonstrated alteration in the lipid, phospholipid and cholesterol content when compared with age-matched control. Membrane bound (Na+ + K+)ATPase activity is found to be significantly increased in jaundiced neonates. Alteration in membrane permeability characteristics in jaundiced neonates causes severe microenvironmental changes in red blood cell profile.

A perspective of smooth muscle contractility through actions of vanadium compounds.

Smooth muscle contraction has a characteristic step-response with successive additions of stimulating compounds, and instant reversal on withdrawing the stimulus, indicative of an equilibrium situation wherein continuous, rapid reactions are occurring. Vanadium compounds, ortho- and meta-vanadates, decavanadate and peroxovanadate, were found to contract a variety of smooth muscles. Their actions were analyzed with respect to activation of receptors, increase in the intracellular calcium concentration, and increase in calmodulin-dependent myosin light chain phosphorylation leading to contraction. A new perspective of smooth muscle contractility has emerged from the studies with vanadium compounds suggesting control mechanisms involving phosphorylation for contraction and redox for relaxation.

Intoxicaçäo digitálica / Digitalis intoxication

O fator mais importante na intoxicaçäo digitálica é o seu reconhecimento. A intoxicaçäo continua sendo comum e de difícil confirmaçao. Isto se deve às semelhanças existentes entre arritmias, sintomas geraos pela intoxicaçäo digitálica e doença cardíaca de base. Além disso, os níveis plasmáticos terapêuticos da digoxina apresentam alto grau de variabilidade e muitas vezes sobrepöm-se aos da toxicidade, näo servindo, portanto, como parâmetro diagnóstico, especialmente na emergência. A intoxicaçäo pela digital, complicaçäo potencialmente fatal, inclui também os pacientes que ingerem doses maciças na medicaçäo intencional ou acidentalmente. A terapia convencional continua sendo a reposiçäo de potássio sempre que necessário, objetivando manter nível plasmático igual ou superior a 4 mEq/l, o controle dos níveis do magnésio e cálcio, correçäo do balanço hídrico e ácido-básico, bem como a adminstraçäo de antiarrítmicos apropriados ou a instalaçäo de marcapasso temporário. A aquisiçäo terapêutica mais recente é o uso de anticorpo antidigoxina (Fab), que apesar do seu alto custo, quando bem indicado, constitui tratamento efetivo. O objetivo deste artigo é rever o mecanismo de açäo da digital, realçar as manifestaçöes cardíacas e extracardíacas da intoxicaçäo, rever a importância da determinaçäo dos níveis séricos da digital e as alternativas terapêuticas.

Isatin (2,3-dioxoindole): a competitive inhibitor of Na(+)-dependent lysine uptake in rat instestine.

Isatin (2,3-dioxoindole) competitively inhibited (27-40%) Na(+)-dependent L-lysine uptake in rat intestine. The value of Kt was increased from 3.04 mM in control to 5.88 mM in presence of 10 mM isatin. Effect of isatin on the Na(+)-independent amino acid uptake was insignificant (12-18%). The inhibitory constant (Ki) was 2.8 mM under these conditions. The observed inhibition was unaffected by -SH group reacting agents. Isatin (1-10 mM) inhibited Na+, K(+)-ATPase activity in intestine in vitro, the maximum inhibition (66%) being at 10 mM isatin concentration. But the drug had no effect on enzyme activity under in vivo conditions.

Mechanism of enhancement of renal (Na+ + K+) ATPase activity following chronic ethanol exposure

A method was devised to determine the nature of the mechanism of the increase in renal (NA++K+)-ATPase in rats fed dilute ethanol for ten weeks. Antiserum to (NA++K+)-ATPase obtained from rabbits was added to microssomal fractions of Kidney and the activities of (NA++K+)-ATPase and Mg2+ ATPase were determined. The addition of antiserum resulted in a same pattern of dose-related inhibition of (NA++K+)-ATPase activity in control and ethanol-fed rats, whereas mg2+-ATPase was not affected by the antiserum. These results suggest that the mechanism of ethanol-induced enhancement of renal (NA++K+)-ATPase activity could be explained through an increase in the number of catalytic units.

Alloxan diabetes in Swiss mice: activity of Na(+)-K(+)-ATPase and succinic dehydrogenase.

The activities of two enzymes viz: Na(+)-K(+)-ATPase and succinic dehydrogenase (SDH) in brain and liver of alloxan diabetic Swiss albino mice are reported. Alloxan diabetes caused significant decrease in the activity of Na(+)-K(+)-ATPase reflecting reduced glucose transport across the cell membrane. On the contrary, the observed enhanced activity of the enzyme SDH is attributed to increased supply of TCA cycle substrates from accelerated oxidation of fatty acids.

Lithium-induced inhibition of Na-K ATPase and Ca ATPase activities in rat brain synaptosome

To explore the action mechanism of lithium in the brain, the author investigated the effects of lithium on Na-K ATPase and Ca ATPase in rat brain synaptosomes prepared from forebrains by the method of Booth and Clark. The activities of Na-K ATPase and Ca ATPase were assayed by the level of inorganic phosphate liberated from the hydrolysis of ATP. Lithium at the optimum therapeutic concentration of 1 mM decreased the activity of Na-K ATPase from the control value of 19.08 +/- 0.29 to 18.27 +/- 0.10 micromoles Pi/mg protein/h and also reduced the activity of Ca ATPase from 6.38 +/- 0.12 to 5.64 +/- 0.12 micromoles Pi/mg protein/h. The decreased activity of Na-K ATPase will decrease the rate of Ca2+ efflux, probably via an Na-Ca exchange mechanism and will increase the rate of Ca2+ entry by the depolarization of nerve terminals. The reduced activity of Ca ATPase will result in the decreased efflux of Ca2+. As a Conclusion, it can be speculated that lithium elevates the intrasynaptosomal Ca2+ concentration via inhibition of the activities of Na-K ATPase and Ca ATPase, and this increased [Ca2+]i will cause the release of neurotransmitters and neurological effects of lithium.