Na-K ATPase Activity Research Articles

The sublethal effects of zinc at different water temperatures on selected haematological variables in Oreochromis mossambicus

Trace metals such as zinc play an important role in the normal metabolic functioning of all organisms. However, metals can become toxic if background concentrations are exceeded in the environment. This study investigated the sublethal effects of zinc on the haematology of Oreochromis mossambicus at different water temperatures. Fish were exposed to sublethal zinc (Zn) concentrations of 40μg/l (mean Zn concentration measured from the Mhlathuze River) for a period of 96h at different water temperatures representing the seasonal temperatures in the Mhlathuze River (18°C, 24°C, 28°C and 32°C). Exposure to zinc at 28°C and 32°C resulted in the most pronounced haematological changes in O. mossambicus. The erythrocytotic conditions found following exposure to Zn could be attributed to damage to the gill surface. The ensuing hypoxic conditions are alleviated through the release of large amounts of immature RBC into the circulatory system. The leucocytotic conditions support the findings of damage to the gill surface. The increased glucose concentrations were attributed to a general adaptation response whereby metabolites are mobilised to meet increased energy demands during periods of stress. The changes in Na+ and K+ levels in the plasma of O. mossambicus can be attributed to a combination of stimulation of Na-K ATPase activity, reduced membrane permeability in the gill tissue and leakage from cells into the blood, respectively.

Cell coupling and impulse propagation in the failing heart.

Cell coupling and impulse propagation were investigated in the ventricle of cardiomyopathic hamsters at an advanced stage of heart failure. An appreciable decline in junctional conductance was found, a phenomenon in part related to activation of the plasma and cardiac renin-angiotensin systems. Decreased expression of connexin43 or an alteration of junctional proteins also might be implicated in the decreased cell coupling. Morphologic abnormalities such as fibrosis, necrosis, and rupture of cell contacts contribute to the decline of conduction velocity or to the blockade of impulse propagation in some areas of the ventricle, creating the conditions for anisotropic conduction and cardiac arrhythmias. The decrease in membrane potential found in myopathic cells is related in part to depression of Na-KATPase activity, and the lack of action of beta-adrenergic agonists on junctional conductance is explained by down-regulation of beta receptors and an abnormality of adenyl cyclase.

Insulin stimulates transepithelial sodium transport by activation of a protein phosphatase that increases Na-K ATPase activity in endometrial epithelial cells.

The objective of this study was to investigate the effects of insulin and insulin-like growth factor I on transepithelial Na(+) transport across porcine glandular endometrial epithelial cells grown in primary culture. Insulin and insulin-like growth factor I acutely stimulated Na(+) transport two- to threefold by increasing Na(+)-K(+) ATPase transport activity and basolateral membrane K(+) conductance without increasing the apical membrane amiloride-sensitive Na(+) conductance. Long-term exposure to insulin for 4 d resulted in enhanced Na(+) absorption with a further increase in Na(+)-K(+) ATPase transport activity and an increase in apical membrane amiloride-sensitive Na(+) conductance. The effect of insulin on the Na(+)-K(+) ATPase was the result of an increase in V(max) for extracellular K(+) and intracellular Na(+), and an increase in affinity of the pump for Na(+). Immunohistochemical localization along with Western blot analysis of cultured porcine endometrial epithelial cells revealed the presence of alpha-1 and alpha-2 isoforms, but not the alpha-3 isoform of Na(+)-K(+) ATPase, which did not change in the presence of insulin. Insulin-stimulated Na(+) transport was inhibited by hydroxy-2-naphthalenylmethylphosphonic acid tris-acetoxymethyl ester [HNMPA-(AM)(3)], a specific inhibitor of insulin receptor tyrosine kinase activity, suggesting that the regulation of Na(+) transport by insulin involves receptor autophosphorylation. Pretreatment with wortmannin, a specific inhibitor of phosphatidylinositol 3-kinase as well as okadaic acid and calyculin A, inhibitors of protein phosphatase activity, also blocked the insulin-stimulated increase in short circuit and pump currents, suggesting that activation of phosphatidylinositol 3-kinase and subsequent stimulation of a protein phosphatase mediates the action of insulin on Na(+)-K(+) ATPase activation.

Norepinephrine induced alpha-adrenoceptor mediated increase in rat brain Na-K ATPase activity is dependent on calcium ion

It has been reported that norepinephrine increases Na-K ATPase activity by acting on α-1 adrenoceptors. The mechanism of such an increase was investigated. The norepinephrine induced increase in synaptosomal Na-K ATPase activity was prevented by pretreating the rat brain homogenate with either EDTA, a divalent cation chelator or prazosin, an α-1 adrenoceptor blocker. The norepinephrine and EGTA increased the Na-K ATPase activity in the synaptosome prepared from rat brain homogenate untreated with EDTA. The EGTA was ineffective in stimulating the enzyme activity if the synaptosome was prepared from homogenate treated with norepinephrine. However, the EGTA was effective in increasing the enzyme activity if the synaptosome was prepared from the homogenate treated with norepinephrine in the presence of prazosin. Thus, norepinephrine did not increase the Na-K ATPase activity in the presence of EDTA or α-1 adrenoceptor blocker. Similarly, the Ca ++ chelator, EGTA, could not increase the enzyme activity if the homogenate was pretreated with norepinephrine alone. However, if norpeinephrine action was blocked by α-1 antagonist prazosin, EGTA increased the enzyme activity possibly by chelation of Ca ++. Further, chlorotetracycline fluorescence study showed that norepinephrine removes membrane bound Ca ++. Thus, it is likely that norepinephrine acts on adrenoceptors and removes membrane bound Ca ++ and thereby increases the Na-K ATPase activity in the synaptosome.

Rapid communication Comparison of Na-K ATPase activity in rat brain synaptosome under various conditions

Rapid eye movement (REM) sleep deprivation alters neuronal excitability possibly by increasing Na-K ATPase activity. The enzyme activity is known to be affected by norepinephrine as well as calcium (Ca ++) and both are affected by REM sleep deprivation. Before studying its molecular mechanism of action, synaptosomal Na-K ATPase activity was estimated under various conditions. The enzyme activity in synaptosome increased after lysis and in the presence of EDTA. The increase in the lysed preparation was possibly because almost all the active sites of the enzyme molecules were exposed to express their activity, unlike unlysed preparation where half are likely to be inside out. EDTA possibly increased the enzyme activity by chelating the Ca ++ which is known to have an inhibitory effect on the enzyme activity. Also, the REM sleep deprivation induced increase in the enzyme activity was observed in lysed preparations and in the presence of EDTA only. These observations fit with the existing knowledge, however, the molecular mechanism of the increase needs to be investigated.

Intracellular accumulation of thallium as a marker of cisplatin cytotoxicity in nonsmall cell lung carcinoma: an application of inductively coupled plasma mass spectrometry.

Thallium-201 (201Tl) scintigraphy has been used to detect malignant pulmonary disease. The mechanism of Tl influx in tumor cells is believed to be similar to that of cisplatin (CDDP) mediated by sodium- and potassium-activated adenosine triphosphatase (Na-K ATPase), and the Na-K ATPase activity may determine the cellular CDDP accumulation and sensitivity to CDDP. The objective of this study was to determine the accumulation of CDDP and Tl in vitro by using inductively coupled plasma mass spectrometry (ICP-MS), a new analytic technique for detecting ultra trace elements, and to evaluate the correlations between cellular CDDP and Tl accumulation, between CDDP 50% inhibitory concentration (IC50) values and cellular CDDP accumulation, and between CDDP IC50 values and cellular Tl accumulation. Eight nonsmall cell lung carcinoma (NSCLC) cell lines were used (five adenocarcinomas and three squamous cell carcinomas). The cell lines were exposed to CDDP or Tl for 1 hour, and the resulting cellular accumulation of platinum and Tl was determined by ICP-MS. CDDP IC50 values were determined by a soluble tetrazolium/formazan assay. The authors were able to measure cellular CDDP and Tl accumulation precisely, and heterogeneity in the cellular accumulation of CDDP and Tl existed among the NSCLC cell lines. A significant inverse correlation was observed between CDDP IC50 values and the cellular accumulation of both CDDP and Tl. ICP-MS is suitable for the determination of cellular CDDP and Tl accumulation in NSCLC cell lines. Cellular Tl accumulation determined by ICP-MS may reflect CDDP cytotoxicity rather than cellular CDDP accumulation.

The Effect of Different Doses of Epidermal Growth Factor on Liver Ornithine Decarboxylase and Na-K ATPase Activities in Newborn Rats

1. Ornithine decarboxylase and Na-K ATPase activities were studied in rat livers that were treated with different doses of epidermal growth factor (EGF). 2. The ornithine decarboxylase activities were studied with spectrophotometry, and results were expressed as micromoles of putrescine per hour per milligram of protein. Na-K ATPase activities were studied on the basis of the principle of measuring the amount of inorganic phosphates released by the hydrolysis of ATP, and the results were expressed as micromoles of inorganic phosphate per hour per milligram of protein. 3. When compared with the controls, although the Na-K ATPase activities were decreased at low doses of EGF, their activities were found to be increased at high doses of EGF. On the other hand, there was a positive correlation between ornithine decarboxylase activities and EGF doses. 4. The results of this study suggest that, whereas the decrease in Na-K ATPase activities at low doses of EGF can be due to the utilization of the enzyme, the increase in Na-K ATPase activities at high doses of EGF can be attributed to its enhanced synthesis.

The mechanism of positive scintigraphy with 99mTc-pertechnetate in adenolymphomas of the parotid grand

In order to clarify the mechanism of positive scintigraphy with Tc pertechnetate in adenolymphomas of the parotid gland, 10 cases of adenolymphoma were studied morphologically and from a functional view point. Salivary tissue obtained from normal parotid glands were also investigated for comparison with the tumor. In morphological studies, rapid tissue fixation by microwave irradiation produced very good results for examining mitochondria by high resolution light microscopy. A number of mitochondria were found both in the epithelial cells of adenolymphoma and in the striated duct cells of the parotid gland. In functional studies, the one-step method of enzymatic histochemistry was used for detecting Na-K ATPase. Na-K ATPase was found on the baso-lateral side of the epithelial cells of adenolymphoma and the striated duct cells of the parotid gland, and this activity of Na-K ATPase may enhance the accumulation of Tc pertechnetate in these cells. These observations may provide evidence for the bi-cellular theory which indicates that adenolymphoma originated from immature intercalated duct reserve cells.

Effect of Norepinephrine on Ouabain-sensitive, K+-dependent p-Nitrophenylphosphatase Activity in Strial Marginal Cells of the Cochlea in Normal and Reserpinized Guinea Pigs

On the basolateral infoldings of the strial marginal cells in the cochlea, Na-K ATPase activity is abundant. To clarify the humoral control by norepinephrine, K-NPPase activity of strial marginal cells in the cochlea was investigated in normal, reserpine, norepinephrine (NE), reserpine plus NE-treated guinea pigs using a cerium-based method. K-NPPase activity was almost completely decreased 3-20 days after reserpine administration. At 10 days after reserpinization and following NE repeated treatment, enzyme activity was detectable. These results suggested that norepinephrine might restore and regulate strial K-NPPase activity.

Timing of parr-smolt transformation in Atlantic salmon ( Salmo salar): effects of changes in temperature and photoperiod

Water temperature modified the development of smolt characteristics and their duration after an abrupt increase in daylength. A temperature change alone was not a sufficient environmental cue to induce a similar development. To complete parr-smolt transformation after an abrupt increase in daylength, a thermal sum of approximately 400 degree-days (°d) seemed to be necessary. Three groups of Atlantic salmon (1 +) were reared under continuous light (LL) from hatching and then exposed to a 7-week period of short days (LD 8.15:15.45) which ended April 2. In parallel, three groups were exposed to continuous light alone. All six groups were then split into two subgroups of equal numbers. A subgroup from the short-day regime were then mixed with a subgroup from the continuous light regime, to make it possible to continue the experiment with only six tanks and still use duplicates in each combination of light and temperature regime. Thereafter, three temperature regimes were applied: 7.7°C (mean) (4.5–10.8) (range), 10.7°C (7.6–13.7) or 13.7°C (10.5–17.3). Mean temperature from January 31 to April 2 was 9.1°C. The medium temperature regime included no abrupt temperature shift. The short-day regime caused a lower thermal-growth coefficient (TGC) than the continuous regime until the increase in daylength. After this increase and in the rest of the experiment, the TGC was still lowest ( P<0.05) in the groups previously exposed to short days. The short-day regime caused a reduction in the condition factor ( P<0.001) and an increase in gill Na–K-ATPase ( P<0.001) after the increase in daylength in contrast to the continuous light regime. Silvery colour and the ability to regulate plasma Cl − level in a 24-h seawater challenge test developed under both light regimes. The decrease in condition factor was not significantly affected by temperature. Silvering and Na–K-ATPase activity developed more rapidly at the high temperature regime ( P<0.001), but no difference was found between the medium and low temperatures. An elevated gill Na–K-ATPase activity and low plasma Cl − levels in the 24-h seawater challenge test were found for a shorter period at higher temperatures. Plasma Cl − levels below 150 mM were found for approximately 1, 2 or 6 weeks in the high, medium and low temperature regime, respectively. In the continuous light regime, the temperature regime had no significant effect on the development of silvering, condition factor or gill Na–K-ATPase activity, but the ability to regulate plasma Cl − was affected by temperature ( P<0.05).

Role of Digitalis-Like Substance in the Hypertension of Streptozotocin-Induced Diabetes and Simulated Weightlessness in Rats

We have examined the role of plasma Na+-K+pump inhibitor (SPI) in the hypertension of streptozotocin induced insulin dependent diabetes (IDDM) in reduced renal mass rats. The increase in blood pressure (BP) was associated with an increase in extracellular fluid volume (ECFV), and SPI and a decrease in myocardial Na+,K+ATPase (NKA) activity, suggesting that increased SPI, which inhibits cardiovascular muscle (CVM) cell NKA activity, may be involved in the mechanism of IDDM-hypertension. In a second study, using prolonged suspension resulted in a decrease in cardiac NKA activity, suggesting that cardiovascular deconditioning following space flight might in part result from insufficient SPI

Hypertension related adducin polymorphism affects Na-KATPase activity

Hypertension related adducin polymorphism affects Na-KATPase activity

Short-Term vs. Sustained Inhibition of Proximal Tubule Na, K-ATPase Activity by Dopamine: Cellular Mechanisms

Dopamine (DA) produces a natriuresis attributed in part to inhibition of Na, K-ATPase activity (NKA) in the proximal tubule (PCT), and impairment in this inhibition has been linked to several forms of hypertension in animals. Here we examined whether the intracellular signaling mechanisms involved are the same in the early and late phases of this phenomenon. DA (1 μM) inhibited NKA similarly after 15 min (by 38%) or 180 min (by 36%) incubation, taken to represent short-term (ST) and sustained (Sd) pump regulation, respectively. Calphostin C, a specific inhibitor of protein kinase C (PKC), completely blocked the ST action of DA on NKA, whereas IP20, a specific inhibitor of protein kinase (PKA), had no effect. In contrast, IP20 completely abolished the Sd (180 min) inhibition by DA, whereas calphostin C had only a partial or variable effect. The DA-1 agonist fenoldopam (which does not activate PKC but increases cAMP) alone failed to inhibit the pump at 180 min (as it does also in the short-term in PCT), suggesting that ST inhibition is required for the Sd effect to occur. Furthermore, PTH1,34, a known ST inhibitor of NKA suppressed the pump at 180 min (by 46%), but unlike in the short-term, this effect was completely prevented by 1P2@ In contrast, PTH, which does not stimulate adenylyl cyclase or activate PKA, caused only a small (19%) and variable Sd inhibition. In conclusion, short-term inhibition of the PCT pump by dopamine is mediated via PKC, whereas the sustained inhibition requires the PKA pathway in addition to the ongoing PKC-mediated effect.

Regulation of Sodium Transport by Endogenous Dopamine Production in Proximal Tubular and OK Cells

Inhibition of AADC for several hours increases the activity and mass of NaKATPase in proximal tubular basolateral membranes and reduces phosphate (Pi) and citrate excretion, but has only a small effect on Na excretion. The reduction in citrate excretion is consistent with the observed increase in brush border Na+/H+ exchange. Thus, in Na replete rats, endogenous D inhibits Na entry into proximal tubular cells, through cotransport with Pi and exchange with H+ and inhibits exit through the Na pump. Tonic D inhibition of NaKATPase and Na+/H+ antiporter activity is not found in the SH rats’ kidneys, which have defective linkage of proximal tubular D receptors to adenylate cyclase. The inhibitory action of endogenous D on Pi reabsorption is retained in SHR kidneys. This suggests that different signaling systems are responsible for the effects of D on NaPi transport and Na+/H+ exchange. In OK cells D inhibits NaPi cotransport (Ki 0.2 μM). The D effect is not blocked by cAMP, adenylate cyclase, PKA or PKC inhibitors. Thus it appears that D regulates NaPi transport by a non-cAMP, non-PKC mechanism and is a homeostatic regulator of phosphate reabsorption by SHR.

Mechanism Responsible for Oligomycin-induced Occlusion of Na+ within Na/K-ATPase

The mechanism whereby oligomycin occludes Na+ within Na/K-ATPase was investigated to study Na+ and K+ transport mechanisms. Oligomycin stimulated Na+ binding to Na/K-ATPase but inhibited Na-K and Na-Na exchange. The oligomycin concentration required to stimulate Na+ binding to half-maximal was 4.5 microM, which was close to the concentration that reduced Na-Na and Na-K exchange and ATPase activity to half-maximal, suggesting that Na/K-ATPase possesses an oligomycin binding site responsible for stimulating Na+ binding and reducing ion exchange and ATPase activity. In contrast, neither K+ binding nor K+ transport was affected by oligomycin. Limited tryptic digestion of Na/K-ATPase showed that, unlike Na+, K+, and ouabain, oligomycin treatment did not result in a specific digestion pattern. Oligomycin appeared to inhibit ouabain binding in a noncompetitive manner, whereas it did not affect ATP binding. Na/K-ATPase isoforms with low and high sensitivities to ouabain were equally sensitive to oligomycin. These results suggest that the oligomycin binding site is located on the extracellular side of Na/K-ATPase, at a different position from the ouabain binding site, and this antibiotic did not induce a conformational change of Na/K-ATPase. We propose that oligomycin interacts with the Na+ occlusion site from the extracellular side of Na/K-ATPase, which delays Na+ release to the extracellular side without inducing a conformational change, suggesting that the pathways responsible for Na+ and K+ transport differ.

The effects of 2-chloroadenosine and deoxycoformycin on the ATP level, Na-K ATPase activity in experimental brain ischemia of gerbil.

The effect of 2-chloroadenosine, stable adenosine analog, and deoxycoformycin, adenosine deaminase inhibitor on brain ATP level and Na-K ATPase activity in ischemia were studied. The brain ATP level was increased after we administered both 2-chloroadenosine and deoxycoformycin, but Na-K ATPase activity did not change after deoxycoformycin. The results suggest that 2-chloroadenosine treatment influenced both the ATP production and membrane permeability due to cerebral ischemia. Deoxycoformycin did not protect the membrane permeability, although it increased the ATP production.

Regulation of NaK-ATPase by platelet-derived growth factors in cultured rat thoracic aortic smooth muscle cells

Regulation of (Na(+) + K(+))-adenosine triphosphatase (NaK-ATPase) by platelet-derived growth factor (PDGF) in cultured rat thoracic aortic smooth muscle cells (SMC) was examined. PDGF-BB enhances SMC proliferation and NaK-ATPase activity. Ouabain, an inhibitor of NaK-ATPase activity, prevents PDGF-BB-induced SMC proliferation. As shown by Western blot and immunochemiluminescence analysis, PDGF-BB also enhances alpha(1), truncated alpha(1), and beta(1) NaK-ATPase subunit levels. PDGF-AA and PDGF-AB show no effect on alpha(1) and truncated alpha(1) levels in slot blot analysis. Induction of NaK-ATPase subunit levels by PDGF-BB could be one of the initial events in vascular SMC proliferation. Copyright 1996 S. Karger AG, Basel

Effect of human serum and a new monocolonal anti-digoxin antibody on Na-K and Mg atpase activity

Effect of human serum and a new monocolonal anti-digoxin antibody on Na-K and Mg atpase activity

Insulin downregulates diabetic-enhanced intestinal glucose transport rapidly in ileum and slowly in jejunum.

The effect of insulin on intestinal sodium-dependent glucose transport and brush border membrane surface area was examined in male Sprague-Dawley rats following a 30-day period of nontreated streptozocin-induced diabetes. Nontreated diabetic rats were hyperglycemic and demonstrated increased jejunal and ileal Na-dependent glucose transport Jmax (maximal transport capacity) and Na-K ATPase activity compared with controls. Daily administration of insulin resulted in a steady decline in blood glucose levels over a period of 6 days. Jejunal Jmax was normalized after 2 days of insulin therapy, while ileal Jmax was normalized 12 h following a single insulin injection. The normalization of Na-K ATPase activity lagged behind Na-dependent glucose transport regulation by 24 h in both jejunum and ileum. Brush border surface area was increased in the ileum of diabetic rats as a result of an increase in microvillus height. Insulin treatment resulted in a decrease in ileal microvillus height to control values by 12 h, which correlated directly with the decrease in Na-dependent glucose transport. Insulin had no effect on jejunal brush border surface area. In conclusion, these findings indicate that the jejunum and ileum respond differentially to experimentally induced diabetes, and these regions also utilize different adaptive mechanisms to regulate Na-dependent glucose transport.

Ouabain induces inhibition of the progression phase in human T-cell proliferation.

Ouabain, a specific inhibitor of the Na-K ATPase, has been shown to exert immunosuppressive effects. The goals of this study were to define the stage of the proliferative response which is sensitive to ouabain and to correlate the inhibitory action of ouabain on cell proliferation with its effect on Na-K ATPase activity. We found that ouabain inhibited T-cell proliferation in a dose-dependent manner and this inhibition was similar in CD4+ and CD8+ T cells. To define the role of the Na-K ATPase in early activation of T lymphocytes, we examined the effects of ouabain on the induction of competence (acquisition of responsiveness to interleukin (IL)-2 or IL-4) by phytohemagglutinin (PHA) or the combination of phorbol dibutyrate/ionomycin. Ouabain, at concentrations that completely inhibited the enzyme activity, did not interfere with the induction of competence, suggesting that although activated cells express increased activity of Na-K ATPase, this enzyme activity does not play a role in early activation pathways. In contrast, ouabain inhibited the progression phase to DNA synthesis in a dose-dependent manner even at concentrations that had little or no effect on Na-K ATPase activity. This inhibition was not due to a decrease in the production of IL-2 but rather to an inhibition of the expression of the p55 and p75 subunits of the IL-2 receptor (IL-2R). The inhibition of p55 appeared to occur at the mRNA level. These results indicate that the activity of the Na-K ATPase is not essential for the induction of competence or early activation. On the other hand, inhibition of cell proliferation and transcription of IL-2R subunits by low concentrations of ouabain may be related to changes in intracellular K+ concentrations or to inhibition of membranal phospholipid metabolism secondary to alteration in Na-K ATPase activity.