Reversible stimulation of the Na+/K+ pump by monensin in cultures of vascular smooth muscle

Two ionophores, monensin and salinomycin, increased total cell Na⁺ and ouabain-sensitive ⁸⁶Rb⁺ uptake in cultures of smooth muscle cells from rat aorta. Monensin was used to produced graded increases in cell Na⁺ in order to assess the Na⁺ dependence of the Na⁺/K⁺ pump in the intact cell. The relationship between internal Na⁺ and ouabain-sensitive ⁸⁶Rb⁺ uptake was hyperbolic (K¹_{Na = 3 mM)}. Monensin did not stimulate ⁸⁶Rb⁺ uptake in the absence of external Na⁺. Loading the cells with Na⁺ by exposing cultures to a K⁺-free medium for 3 hr maximally increased cell Na⁺ and ouabain-sensitive ⁸⁶Rb⁺ uptake to the same extent as monensin. Total cell Na⁺ and pump activity in monensin-treated cells returned to the initial values after removing the ionophore. Monensin was then able to increase total cell Na⁺ and ouabain-sensitive ⁸⁶Rb⁺ uptake to the same extent as the initial treatment with the ionophore.     

Adaptation of potassium metabolism and restoration of mitosis during prolonged treatment of mouse lymphoblasts with ouabain

The effects of ouabain on the growth of murine lymphoblasts in vitro have been studied. Exposure of cells to ouabain (0.1 mM) initially inhibited ⁸⁶Rb⁺ uptake rate, reduced the intracellular potassium concentration, and decreased population growth rates. Continued exposure to the same ouabain concentration resulted in an increase of ⁸⁶Rb⁺ uptake rate, intracellular potassium content and population growth rates to control values (adaptation). When treated cells were resuspended in medium free of ouabain after 12 to 15 hours of ouabain treatment, ⁸⁶Rb⁺ uptake rates and intracellular potassium levels exceeded those of untreated cells. Adaptation was inhibited by cycloheximide (3 μg/ml) and by actinomycin D (0.05 μg/ml). Kinetic analysis of transport suggested that while the total capacity of the Na⁺, K⁺ transport system increased, the affinity for both the cation (⁸⁶Rb⁺) and ouabain decreased.     

Insulin action on cardiac glucose transport Studies on the role of the Na+/K+ pump

Isolated muscle cells from adult rat heart have been used to study the relationship between myocardial glucose transport and the activity of the Na⁺/K⁺ pump. ⁸⁶Rb⁺-uptake by cardiac cells was found to be linear up to 2 min with a steady-state reached by 40–60 min, and was used to monitor the activity of the Na⁺/K⁺ pump. Ouabain (10^?3 mol/I) inhibited the steady-state uptake of ⁸⁶Rb⁺ by more than 90%. Both, the ouabain-sensitive and ouabain-insensitive ⁸⁶Rb⁺-uptake by cardiac cells were found to be unaffected by insulin treatment under conditions where a significant stimulation of 3-O-methylglucose transport occurred. ⁸⁶Rb⁺-uptake was markedly reduced by the presence of calcium and/or magnesium, but remained unresponsive towards insulin treatment. Inhibition of the Na⁺/K⁺ pump activity by ouabain and a concomitant shift in the intracellular Na⁺:K⁺ ratio did not affect basal or insulin stimulated rates of 3-O-methylglucose transport in cardiac myocytes. The data argue against a functional relationship between the myocardial Na⁺/K⁺ pump and the glucose transport system.     

Na/K-ATPase assay in the intact guinea pig liver submitted to in situ perfusion

We describe an assay for the enzyme Na/K-ATPase in intact guinea pig livers perfused through the portal vein with modified Hank’s solution. The model uses the measurement of non-radioactive rubidium ion incorporation by liver cells, both in the absence and in the presence of the specific Na/K-ATPase inhibitor ouabain, followed by a rinsing procedure with cold saline. The concentration of Rb⁺ in acid-digested liver lobes was measured by atomic emission spectrometry and Na/K pump activity was calculated by the difference between the incorporation of Rb⁺ in the absence and in the presence of ouabain. The optimal conditions for Rb⁺ incorporation were: perfusion flow rate, 3 ml/min per liver; perfusion time at 37 °C, 60 min; rinsing time with cold saline, 5-10 min; and concentration of ouabain, 3 mM. The calculated ouabain IC₅₀ was 100 μM. The major advantage of this model is the possibility of testing experimental drugs affecting this enzyme in conditions close to those in the intact organ.     

Identification of a third isoform of Na+, K+-ATPase activity in rat brain synaptosomes

³H-ouabain binding and ouabain-inhibitable ⁸⁶Rb⁺ (K⁺) uptake were investigated as a means to identify a third isoform of Na⁺, K⁺-ATPase in crude synaptosome preparations. The specific binding of low concentrations (10 nM and 1 uM) of ³H-ouabain, in crude synaptosome preparations, was markedly inhibited by K⁺ (0.5–5 mM). Accordingly, ⁸⁶Rb⁺ (K⁺) uptake, in the presence of 5 mM K⁺ was not sensitive to inhibition by low concentrations (10⁻¹¹–10⁻⁷ M) of ouabain. Higher concentrations (10⁻⁶–10^−2.6 M) of ouabain resulted in a biphasic inhibition of K⁺ uptake, which distinguished the activities of the presumed alpha 2 and alpha 1 isozymes of Na⁺, K⁺-ATPase. Reduction of K⁺ (1.25 mM and 0.5 mM) in the incubation, resulted in the observation of a third component of ouabain- sensitive K⁺ uptake. This Na⁺, K⁺-ATPase activity, which was defined, pharmacologically, as very sensitive (VS) to ouabain, exhibited IC₅₀s of 3.6 nM and 92 nM at 1.25 mM K⁺ and 0.5 mM K⁺, respectively. Inhibition of ouabain binding and VS-dependent K⁺ uptake, at a high, physiological cocentration (5 mM) of K⁺, suggests that VS may be an inactive isoform of brain Na⁺, K⁺-ATPase under resting conditions.     

Inhibition of Na+/K+-ATPase may be one mechanism contributing to potassium efflux and cell shrinkage in CD95-induced apoptosis

To investigate the involvement of K⁺ efflux in apoptotic cell shrinkage, we monitored efflux of the K⁺ congener,⁸⁶ Rb⁺, and cell volume during CD95-mediated apoptosis in Jurkat cells. An anti-CD95 antibody caused apoptosis associated with intracellular GSH depletion, a significant increase in ⁸⁶Rb⁺ efflux, and a decrease in cell volume compared with control cells. Preincubating Jurkat cells with Val-Ala-Asp-chloromethylketone (VAD-cmk), an inhibitor of caspase proteases, prevented the observed ⁸⁶Rb⁺ efflux and cell shrinkage induced by the anti- CD95 antibody. A wide range of inhibitors against most types of K⁺ channels could not inhibit CD95-mediated efflux of⁸⁶ Rb⁺, however, the uptake of⁸⁶ Rb⁺ by Jurkat cells was severely compromised when treated with anti-CD95 antibody. Uptake of⁸⁶ Rb⁺ in Jurkat cells was sensitive to ouabain (a specific Na⁺/K⁺-ATPase inhibitor), demonstrating Na⁺/K⁺-ATPase dependent K⁺ uptake. Ouabain induced significant⁸⁶ Rb⁺ efflux in untreated cells, as well as it seemed to compete with⁸⁶ Rb⁺ efflux induced by the anti-CD95 antibody, supporting a role for Na⁺/K⁺-ATPase in the CD95-mediated⁸⁶ Rb⁺ efflux. Ouabain treatment of Jurkat cells did not cause a reduction in cell volume, although together with the anti-CD95 antibody, ouabain potentiated CD95-mediated cell shrinkage. This suggests that the observed inhibition of Na⁺+/K⁺-ATPase during apoptosis may also facilitate apoptotic cell shrinkage.     

Influence of Bcl-2 overexpression on Na+/K+-ATPase pump activity: Correlation with radiation-induced programmed cell death

Bcl-2 overexpression in transfected PW cells is associated with inhibition of radiation-induced programmed cell death (PCD). We have previously reported that there is a relationship between inhibition of radiation-induced PCD and membrane hyperpolarization in these cells. In this article, we report that Na⁺/K⁺-ATPase pump activity, as measured by the uptake of Rubidium-86 (⁸⁶Rb⁺), is significantly higher in Bcl-2 overexpressing PW cells than in control PW cells, and that pump activity following irradiation with doses ≥ 500 cGy was reduced to a lesser extent in the Bcl-2 transfectants than in the control cells. When PW-Bcl-2 cells were incubated with a dose of ouabain (1 μM) that decreased pump activity significantly, but did not induce PCD, the previously reported protection from radiation-induced PCD associated with overexpression of Bcl-2 no longer existed. In order to demonstrate that reactive oxygen species (ROS) affected Na⁺/K⁺-ATPase pump activity, cells were incubated with N-acetyl cysteine (NAC) prior to irradiation, or treated with the ROS generating drug buthionine sulphoxamine (BSO). ⁸⁶Rb⁺uptake was significantly higher in irradiated cells incubated with NAC compared to cells irradiated in the absence of NAC, while BSO resulted in lower levels of ⁸⁶Rb⁺uptake, suggesting that the effects of radiation on the Na⁺/K⁺-ATPase pump were due to ROS. Furthermore, the resting cell membrane potential of cells exposed to NAC were slightly hyperpolarized compared to control PW cells, whereas cells exposed to BSO were depolarized in comparison to control PW cells. In summary, this data suggests that Bcl-2 affects Na⁺/K⁺-ATPase pump activity, which is associated with the resting membrane potential and the level of susceptibility to radiation-induced PCD. J. Cell. Physiol. 171:299–304, 1997. © 1997 Wiley-Liss, Inc.     

Regulation of Na+, K+-ATPase activity in MDCK kidney epithelial cell cultures: Role of growth state,cyclic AMP,and chemical inducers of dome formation and differentiation

Na⁺,K⁺-ATPase activity was monitored in MDCK kidney epithelial cell monolayers and in cell extracts as a function of cell density, cAMP elevation, and exposure to hexamethylene bisacetamide (HMBA) and dimethylsulfoxide (Me₂SO). Ouabain-sensitive Na⁺,K⁺-ATPase and ⁸⁶Rb⁺ uptake activities, and the number of [³H]-ouabain binding sites were maximal in subconfluent cultures and decreased accompanying the development of a confluent monolayer. A sodium pump density of 8 × 10⁷ pumps/cell was estimated for subconfluent cultures, declining to 9 × 10⁵ pumps/cell at confluence. Previous studies have shown that dibutyryl cyclic AMP (Bt₂cAMP), 1-methyl-3-isobutylxanthine (IBMX), or the differentiation inducers HMBA and Me₂SO, which also caused cAMP elevation, all stimulated dome formation, a visible manifestation of active transepithelial Na⁺ and water transport (Lever, 1979). In the present study, all of these inducers were found to elevate intracellular Na⁺ content, implicating this variable in control of induction of dome formation. Operationally, inducers could be divided into two classes. HMBA and Me₂SO partially inhibited ouabain-sensitive ⁸⁶Rb⁺ influx. Ouabain, at a concentration that caused partial sodium pump inhibition and increased intracellular Na⁺ content, was also effective as an inducer. The second class, exemplified by IBMX and Bt₂cAMP caused a furosemide-sensitive increase in intracellular Na⁺ content. This class of inducers stimulated ouabain-sensitive ⁸⁶Rb⁺ uptake, presumably by substrate effects due to increased Na⁺ levels. The Na⁺ or ATP activation of Na⁺,K⁺-ATPase activity assayed in cell-free extracts, the affinity of the transport system for Rb⁺ in intact cells and intracellular ATP levels were unchanged by inducer treatment. Elevation of intracellular Na⁺ concentration, either by cAMP-stimulated, furosemide-sensitive mechanisms or by partial inhibition of the sodium pump may stimulate the induction of dome formation in MDCK cells.     

Na,K-ATPase mutations in familial hemiplegic migraine lead to functional inactivation

The Na,K-ATPase is an ion-translocating transmembrane protein that actively maintains the electrochemical gradients for Na⁺ and K⁺ across the plasma membrane. The functional protein is a heterodimer comprising a catalytic α-subunit (four isoforms) and an ancillary β-subunit (three isoforms). Mutations in the α₂-subunit have recently been implicated in familial hemiplegic migraine type 2, but almost no thorough studies of the functional consequences of these mutations have been provided. We investigated the functional properties of the mutations L764P and W887R in the human Na,K-ATPase α₂-subunit upon heterologous expression in Xenopus oocytes. No Na,K-ATPase-specific pump currents could be detected in cells expressing these mutants. The binding of radiolabelled [³H]ouabain to intact cells suggested that this could be due to a lack of plasma membrane expression. However, plasma membrane isolation showed that the mutated pumps are well expressed at the plasma membrane. ⁸⁶Rb⁺-flux and ATPase activity measurements demonstrated that the mutants are inactive. Therefore, the primary disease-causing mechanism is loss-of-function of the Na,K-ATPase α₂-isoform.     

Role of the sodium pump and the background K+ channel in passive K+(Rb+) uptake by isolated cardiac sarcolemmal vesicles

Summary A simple procedure was developed for the isolation of a sarcolemma-enriched membrane preparation from homogenates of bullfrog (Rana catesbeiana) heart. Crude microsomes obtained by differential centrifugation were fractionated in Hypaque density gradients. The fraction enriched in surface membrane markers consisted of 87% tightly sealed vesicles. The uptake of⁸⁶Rb⁺ by the preparation was measured in the presence of an opposing K⁺ gradient using a rapid ion exchange technique. At low extravesicular Rb⁺ concentrations, at least 50% of the uptake was blocked by addition of 1mm ouabain to the assay medium. Orthovanadate (50 m), ADP (2.5mm), or Mg (1mm) were also partial inhibitors of Rb⁺ uptake under these conditions, and produced a complete block of Rb⁺ influx in the presence of 1mm ouabain. When⁸⁶Rb⁺ was used as a tracer of extravesicular K⁺ (Rb ₀ ⁺ 40 m K ₀ ⁺ =0.1–5mm) a distinct uptake pathway emerged, as detected by its inhibition by 1mm Ba²⁺ (K _0.5=20 m). At a constant internal K⁺ concentration (K _in ⁺ =50mm) the magnitude of the Ba²⁺-sensitive K⁺ uptake was found to depend on K ₀ ⁺ in a manner that closely resembles the K⁺ concentration dependence of the background K⁺ conductance (I _Kl) observed electrophysiologically in intact cardiac cells. We conclude that K⁺ permeates passively this preparation through two distinct pathways, the sodium pump and a system identifiable as the background potassium channel.     

Modulation of β-Cell Ouabain-Sensitive 86Rb+ Influx (Na+/K+ Pump) by D-Glucose,Glibenclamide or Diazoxide

The activity of the β-cell Na⁺/K⁺ pump was studied by using ouabain-sensitive (lmM ouabain) ⁸⁶Rb⁺ influx in β-cell-rich islets of Umeå-ob/ob mice as an indicator of the pump function. The present results show that the stimulatory effect of glucose on ouabain-sensitive ⁸⁶Rb⁺ influx reached its approximate maximum at 5mM glucose. Pre-treatment of the islets with 20mM glucose for 60 min strongly reduced the glucose-induced stimulation of the Na⁺/K⁺ pump. Pre-treatment (60 or 180 min) of islets at 0mM glucose, on the other hand, did not affect the magnitude of the glucose-induced stimulation of ⁸⁶Rb⁺ influx dunng the subsequent 5-min incubation. Glibenclamide stimulated the ouabain-sensitive ⁸⁶Rb⁺ uptake in the same manner as glucose. The stimulatory effect, showed its apparent maximum at 0.5μM. Pre-treatment (60 min) of islets with 1μM glibenclamide did not reduce the subsequent stimulation of the ouabain-sensitive ⁸⁶Rb⁺ influx. The stimulatory effect of glibenclamide and D-glucose were not .additive, suggesting that they may have the same mechanism of action. No direct effect of glibenclamide (0.01-1μM) was observed on the Na⁺/K⁺ ATPase activity in homogenates of islets. Diazoxide (0.4mM) inhibited the Na⁺/K⁺ pump. This effect was sustained even after 60 min of pre-treatment of islets with 0.4mM diazoxide. The stimulatory effect of glibenclamide and D-glucose were abolished by diazoxide. It is concluded that nutrient as well as non-nutrient insulin secretagogues activate the Na⁺/K⁺ pump, probably as part of the membrane repolarisation process.     

Closure of the yeast mitochondria unspecific channel (YMUC) unmasks a Mg2+ and quinine sensitive K+ uptake pathway in Saccharomyces cerevisiae

The K⁺ uptake pathways in yeast mitochondria are still undefined. Nonetheless, the K⁺-mediated mitochondrial swelling observed in the absence of phosphate (PO₄) and in the presence of a respiratory substrate has led to propose that large K⁺ movements occur in yeast mitochondria. Thus, the uptake of K⁺ by isolated yeast mitochondria was evaluated. Two parallel experiments were conducted to evaluate K⁺ transport; these were mitochondrial swelling and the uptake of the radioactive K⁺ analog ⁸⁶Rb⁺. The opening of the yeast mitochondrial unspecific channel (YMUC) was regulated by different PO₄ concentrations. The high protein concentrations used to measure ⁸⁶Rb⁺ uptake resulted in a slight stabilization of the transmembrane potential at 0.4 mM PO₄ but not at 0 or 4 mM PO₄. At 4 mM PO₄ swelling was inhibited while, in contrast, ⁸⁶Rb⁺ uptake was still observed. The results suggest that an energy-dependent K⁺ uptake mechanism was unmasked when the YMUC was closed. To further analyze the properties of this K⁺ uptake system, the Mg²⁺ and quinine sensitivity of both swelling and ⁸⁶Rb⁺ uptake were evaluated. Under the conditions where the unspecific pore was closed, K⁺ transport sensitivity to Mg²⁺ and quinine increased. In addition, when Zn²⁺ was added as an antiport inhibitor, uptake of ⁸⁶Rb⁺ increased. It is suggested that in yeast mitochondria, the K⁺ concentration is highly regulated by the equilibrium of uptake and exit of this cation through two specific transporters.     

Alkylguanidines as inhibitors of k transport in isolated barley roots

It has been shown that plants can accumulate K⁺ through an energy-dependent process. The effect of alkylguanidines, in particular octylguanidine on the uptake of ⁸⁶Rb⁺ by excised barley roots (Hordeum vulgare var. Apizaco LV-72), has been studied. ⁸⁶Rb⁺ was used as tracer of K⁺. The uptake of ⁸⁶Rb⁺ which is linear with time and shows saturation kinetics is inhibited by octylguanidine. Half-maximal inhibition of ⁸⁶Rb⁺ uptake is attained at 50 μM octylguanidine. Octylguanidine induces a decrease in the V_max of the process and increases the Km of the system for Rb⁺. When the effects of various alkylguanidines were studied, the following order of effectiveness was encountered; octylguanidine = hexilguanidine > butylguanidine > ethylguanidine > guanidine. This suggests that guanidines inhibit Rb⁺ uptake by interacting through its positively charged guanidinium group with a Rb⁺ carrier while the alkyl chain interacts with the hydrophobic milieu of the membrane.     

The Na+,K+,2Cl−-cotransport system in HeLa cells: Aspects of its physiological regulation

We have previously reported on the biochemical properties of a Na⁺,K⁺,2Cl^?-cotransport in HeLa cells and here we deal with aspects of its physiological regulation. Na⁺,K⁺,2Cl^?-cotransport in HeLa cells was studied by ⁸⁶Rb⁺ influx and ⁸⁶Rb⁺/²²Na⁺ efflux measurements. The effects of rat atrial natriuretic peptide (ANP), isoproterenol, and amino acids on ⁸⁶Rb⁺ flux, mediated by the bumet-anide-sensitive Na⁺, K⁺, 2Cl^?-cotransport system and the ouabain-sensitive Na⁺/K⁺-pump, were investigated. ANP reduced bumetanide-sensitive ⁸⁶Rb⁺ influx under isotonic as well as under hypertonic conditions. Similar decrease of bumetanide-sensitive ⁸⁶Rb⁺ influx was observed in the presence of 8-bromo-cGMP, while neither isoproterenol as a β-receptor agonist nor 8-bromo-cAMP-could alter bumetanide-sensitive ⁸⁶Rb⁺ influx. Furthermore, efflux of ⁸⁶Rb⁺ and ²²Na⁺ was greatly reduced in the presence of bumetanide and ANP. Together with our recent findings, showing functionally active, high affinity receptors for ANP on HeLa cells (Kort and Koch, Biochim. Biophys. Res. Commun. 168:148–154, 1990), this study indicates that ANP participates in the regulation of the Na⁺, K⁺, 2Cl^?-cotransport system in HeLa cells. Further measurements revealed that amino acids as present in the growth medium (Joklik's minimal essential medium) and the amino acid derivative α-methyl-aminoisobutyric acid (metAlB, 1 and 5 mM, respectively) also reduced Na⁺, K⁺, 2Cl^?-cotransport-mediated ⁸⁶Rb⁺ uptake and diminished the stimulatory effect of hypertonicity on the cotransporter. In addition, the Na⁺/K⁺-pump was markedly stimulated in the presence of amino acids, while neither ANP and 8-Br-cGMP nor isoproterenol and 8-Br-cAMP had a significant effect on the activity of the Na⁺/K⁺-pump.     

22Na+ and 86Rb+ fluxes in spermatozoa and oocytes of arbacia punctulata

The fluxes of ²²Na⁺ and ⁸⁶Rb⁺ in Arbacia sperm and oocytes were studied in order to determine how these cells carry out cation exchange with the sea environment. The uptake of these ions by serum followed a pattern of early rapid influx (initial 0.5 min) and subsequent efflux (1–3 min) followed by a gradual uptake (after 3 min). Neither the uptake nor the efflux of these cations by Arbacia sperm were affected by ouabain, suggesting that influx and efflux of ²²Na⁺ and ⁸⁶Rb⁺ in Arbacia sperm occur predominantly by passive transport. The ²²Na⁺ uptake by Arbacia oocytes showed a steady increase after an initial rapid uptake. A slight but significant inhibition of ²²Na⁺ uptake was observed with ouabain. However, ⁸⁶Rb⁺ uptake by the oocytes reached an early equilibrium and was not affected by ouabain. The uptake of Rb⁺ by Arbacia oocyte is by passive transport while that of Na⁺ is both by passive and active transport.     

Cell Cycle activation and ouabain-sensitive ion movements of 3T3 and C3H-10T½ fibroblasts

The introduction of either PGF_2α (10^?7 M) or TPA (10^?7 M) stimulated, ouabain-sensitive ⁸⁶Rb⁺ influx at 30 min in postconfluent 3T3-4 mouse fibroblast cultures by 117% and 124%, respectively. Both TPA and PGF_2α at these concentrations stimulated the incorporation of ³H-TdR into DNA. TPA had the greatest stimulatory effect, which was similar to that obtained with 10% fetal calf serum. In accord with the idea that modulation of membrane processes such as Na⁺/K⁺ pump activity in fibroblasts may reflect important events related to the initiation of DNA synthesis, it was observed that in both 3T3-4 and C3H-1 0T½ cells there were parallel increases in ³H-TdR incorporation and ouabain-sensitive ⁸⁶Rb⁺ influxes with 10^?7 M TPA, whereas PGF_2α stimulated a significant increase in ³H-TdR incorporation in 3T3-4 but not C3H-10T½ cells and only marginal increases in ouabain-sensitive ⁸⁶Rb⁺ influx in both. Therefore, although there appears to be a close correlation between Na⁺/K⁺ pump activation and subsequent S-phase entry following TPA stimulation, a similar correlation for PGF_2α cannot be confirmed.     

Sodium-pump activity and its inhibition by extracellular calcium in cardiac myocytes of guinea pigs

Myocardial sodium-pump activity was examined from ouabain-sensitive ⁸⁶Rb⁺ uptake using myocytes isolated from guinea-pig heart. Either sodium loading or the sodium ionophore, monensin, increased ⁸⁶Rb⁺ uptake by over 400%, indicating that the amount of Na⁺ available to the pump is the primary determinant of its activity, and that the sodium pump has a substantial reserve capacity in quiescent myocytes. Moreover, the degree of the above stimulation is markedly higher than corresponding values reported with multicellular preparations, suggesting that diffusion barriers make it impossible to observe the capacity of the sodium pump in the latter preparations. Removal of extracellular Ca²⁺ increased ouabain-sensitive ⁸⁶Rb⁺ uptake, probably by enhancing turnover of the sodium pump rather than increasing availability of Na⁺ to the pump.     

Cell growth and ouabain-sensitive Rb uptake and (Na + K)-ATPase activity in 3T3 and SV40 transformed 3T3 fibroblasts

The uptake of ouabain-sensitive ⁸⁶Rb⁺ uptake measured at 5 min and the uptake measured at 60 min was 4.5- and 2.7-fold greater respectively for SV40 transformed 3T3 cells compared to 3T3 cells during the late log phase of growth. This uptake, however, varied markedly with cell growth. Ouabain-sensitive ⁸⁶Rb⁺ uptake was found to be a sensitive indicator of protein synthesis as measured by total protein content. Cessation of cell growth as measured by total protein content was associated with a decline in ouabain-sensitive ⁸⁶Rb⁺ uptake in both cell types. This increased ouabain-sensitive cation transport was reflected in increased levels of (Na⁺ + K⁺)-ATPase activity for SV40 3T3 cells, which showed a 2.5-fold increase V but the same K_rmm as 3T3 cells.These results are compared with the results of related work. Possible mechanisms for these effects are discussed and how changes in cation transport might be related to alterations in cell growth.     

Conditions controlling relative uptake of potassium and rubidium by plants from soils

Earlier studies have demonstrated close inverse relationships between Rb⁺ concentrations in plants and pH or base (including K⁺) saturation of soils. This study aims at elucidating conditions in soils influencing plant uptake of Rb⁺. Growth experiments with Carex pilulifera L. were performed, modifying the acidity and K⁺ supply of acid soils and solutions. We were unable to assess any reduction in Rb⁺ uptake by adding precipitated CaCO₃ to acid soil unless pH was raised to near neutrality. Though not fully compensating the loss of soil solution K⁺and exchangeable K⁺ from uptake by the growing plants, soil treated with 0.5 mM K⁺ (as KCl) reduced the Rb⁺ concentration in the shoots by 40% without measurably changing soil pH. Experiments varying the pH and K⁺ concentration of a nutrient solution (20% Hoagland), spiked with 6 uM Rb⁺, clearly demonstrated that plant uptake of Rb⁺ and K⁺ was unaffected by acidity in the pH range 3.6–5.0 tested, whereas Rb⁺ uptake was reduced by ca. 50%, when K⁺ concentration was increased from 1.2 to 3.6 mM. The sensitivity of this reaction indicates that shortage or low availability of K⁺ controls Rb⁺ uptake from acid soils, being probably more important than soil acidity per se. Secondary effects of high soil acidity, such as leaching losses of K⁺, might also be of importance in accounting for the high uptake of Rb⁺ from such soils. It is suggested that leaf analysis of Rb⁺ may be used as a method to assess early stages of K⁺ deficiency in plants on acid soils.