Ion Absorption and Retention by Chlorella pyrenoidosa. III. Selective Accumulation of Rubidium, Potassium and Sodium

The selective preference of Chlorella pyrenoidosa for alkali metal cations was found to have the order Rb > K > > > Na. It was demonstrated that a cation of higher preference can replace in the cell cations of lower preference by an ion interchange process.

The replacement of Na from the cell by K or Rb occurred against high external Na concentrations up to 450 meq/1 Na.

It is suggested that the structural selectivity of the Chlorella cell may be amplified by a chromatographic type repetitive selection process, driven by metabolically dependent unsymmetric shape changes of cellular membranes.

     

Cation flux in the ehrlich ascites tumor cell. Evidence for Na+-for-Na+ and K+-for-K+ exchange diffusion.

In a previous study, evidence was presented for an external Na+-dependent, ouabain-insensitive component of Na+ efflux and an external K+-dependent component of K+ efflux in the Ehrlich ascites tumor cell. Evidence is now presented that these components are inhibited by the diuretic furosemide and that under conditions of normal extracellular Na+ and K+ they represent Na+-for-Na+ and K-+for-K+ exchange mechanisms. Using 86Rb to monitor K+ movements, furosemide is shown to inhibit an ouabain-insensitive component of Rb+ influx and a component of Rb+ efflux, both representing approx. 30 percent of the total flux. Inhibition of Rb+ efflux is greatly reduced by removal of extracellular K+. Furosemide does not alter steady-state levels of intracellular K+ and it does not prevent cells depleted of K+ by incubation in the cold from regaining K+ upon warming. Using 22Na to monitor Na+ movements, furosemide is shown to inhibit an ouabain-insensitive component of unidirectional Na+ efflux which represents approx. 22 percent of total Na+ efflux. Furosemide does not alter steady-state levels of intracellular Na+ and does not prevent removal of intracellular Na+ upon warming from cells loaded with Na+ by preincubation in the cold. The ability of furosemide to affect unidirectional Na+ and K+ fluxes but not net fluxes is consistent with the conclusion that these components of cation movement across the cell membrane represent one-for-one exchange mechanisms. Data are also presented which demonstrate that the uptake of alpha-aminoisobutyrate is not affected by furosemide. This indicates that these components of cation flux are not directly involved in the Na+-dependent amino acid transport system A.     

Efficiency of Nitrogen Assimilation by N(2)-Fixing and Nitrate-Grown Soybean Plants (Glycine max [L.] Merr.)

Nodulated and non-nodulated (not inoculated) soybeans (Glycine max [L.] Merr. cv Wells) were grown in controlled environments with N₂ or nonlimiting levels of NO₃⁻, respectively, serving as sole source of nitrogen. The efficiency of the N₂-fixing plants was compared with that of the nitrate-supplied plants on the basis of both plant age and plant size. Efficiency evaluations of the plants were expressed as the ratio of moles of carbon respired by the whole plant to the moles of nitrogen incorporated into plant material.

Continuous 24-hour CO₂ exchange measurements on shoot and root systems made at the beginning of flowering (28 days after planting) indicated that N₂-fixing plants respired 8.28 moles of carbon per mole of N, fixed from dinitrogen, while nitrate-supplied plants respired only 4.99 moles of carbon per mole of nitrate reduced. Twenty-one-day-old nitrate-supplied plants were even more efficient, respiring only 3.18 moles of carbon per mole of nitrate reduced. The decreased efficiency of the N₂-fixing plants was not due to plant size since, on a dry weight basis, the 28-day-old N₂-fixing plants were intermediate between the 28- and 21-day-old nitrate-supplied plants.

The calculated efficiencies were predominantly a reflection of root-system respiration. N₂-fixing plants lost 25% of their daily net photosynthetic input of carbon through root-system respiration, compared with 16% for 28-day-old nitrate-supplied plants and 12% for 21-day-old nitrate-supplied plants. Shoot dark respiration was similar for all three plant groups, varying between 7.9% and 9.0% of the apparent photosynthate.

The increased respiratory loss by the roots of the N₂-fixing plants was not compensated for by increased net photosynthetic effectiveness. Canopy photosynthesis expressed on a leaf area basis was similar for 28-day-old N₂-fixing plants (15.5 milligrams CO₂ square decimeter per hour) and 21-day-old nitrate-supplied plants (14.5 milligrams CO₂ square decimeter per hour). Both were similar in total canopy leaf area. The larger nitrate-supplied plants (28-day-old) had lower photosynthetic rates (12.5 milligrams CO₂ square decimeter per hour), presumably due to self-shading of the leaves.

These data indicate that, during the early stages of plant development, dependence solely on N₂-fixation is an expensive process compared to nitrate reduction in nitrate-supplied plants, since the N₂-fixing plants retained 8% to 12% less of their photosynthate as dry matter.

     

Crassulacean acid metabolism (CAM) in an epiphytic ant-plant, Myrmecodia beccarii Hook.f. (Rubiaceae)

This study demonstrates unequivocally the presence of crassulacean acid metabolism (CAM) in a species of the Rubiaceae, the fourth largest angiosperm plant family. The tropical Australian endemic epiphytic ant-plant, Myrmecodia beccarii Hook.f., exhibits net CO₂ uptake in the dark and a concomitant accumulation of titratable acidity in plants in the field and in cultivation. Plants growing near Cardwell, in a north Queensland coastal seasonally dry forest of Melaleuca viridiflora Sol. ex Gaertn., accumulated ~50 % of their 24 h carbon gain in the dark during the warm wet season. During the transition from the wet season to the dry season, 24 h carbon gain was reduced whilst the proportion of carbon accumulated during the dark increased. By mid dry season many plants exhibited zero net carbon uptake over 24 h, but CO₂ uptake in the dark was observed in some plants following localised rainfall. In a shade-house experiment, droughted plants in which CO₂ uptake in the light was absent and dark CO₂ uptake was reduced, were able to return to relatively high rates of CO₂ uptake in the light and dark within 12 h of rewatering.     

Effects of calcium on the permeability of isolated adult rat heart cells to sodium

The permeability of rat heart myocytes to Na increases when extracellular Ca (Ca₀) is decreased. This increased permeability is reflected in elevated $\text{Na}\text{K}$ ratios in nonenergized myocytes and in increased ouabain-sensitive lactate production in anaerobic myocytes supplemented with glucose. Myocytes treated with ethylene glycol bis(β-aminoethyl ether)N,N′-tetraacetic acid (1 mm) maintain low $\text{Na}\text{K}$ ratios, but expend considerable glycolytic ATP in ouabain-sensitive cation cycling. The data suggest that Ca₀ bound to the sarcolemma can restrict transmembrane movement of Na via pathways that are not yet defined. The lack of significant net accumulation of Ca argues against the explanation that Ca₀ maintains low internal Na levels as a result of NaCa exchange. Both the increased uptake of Na and increased utilization of ATP in the absence of Ca₀ may be relevant to the phenomenon of “Ca-paradox” in situ.     

Effects of Light/Dark on Cation Fluxes in Guard Cells of Commelina communis L.

The effects of light/dark on cation fluxes in isolated guardcells of Commelina communis L. have been studied, using ⁸⁶RbCland ²²NaCl. Transfer to the dark has no effect on ⁸⁶Rb influx,but produces a marked transient stimulation of ⁸⁶Rb efflux,similar to that seen previously on adding ABA. The ⁸⁶Rb effluxfalls on return to light only during the period of stimulatedflux; after the transient, return to light has no effect onefflux. The ability to produce this transient stimulation ontransfer to the dark is recovered in a subsequent light period.In general, in Na-loaded cells, the stimulated efflux is notseen. and the cells do not close in the dark. The results arenot consistent with a simple permeability or potential change,but suggest a specific ion excretion activated by the transferto the dark. Key words: Commelina communis L., Light/dark effects, Cation flux, Guard cells     

Potassium Fluxes in Desheathed Frog Sciatic Nerve

Desheathed frog (R. pipiens) sciatic nerves were soaked in Na-deficient solutions, and measurements were made of their Na and K contents and of the movements of K⁴². When a nerve is in Ringer's solution, the Na fluxes are equal to the K fluxes, and about 75 per cent of the K influx is due to active transport. The Na content and the Na efflux are linearly related to the Na concentration of the bathing solution, while the K content and the K fluxes are not so related. When a nerve is in a solution in which 75 per cent of the NaCl has been replaced by choline chloride or sucrose, the active K influx exceeds the active Na efflux, and the K content is maintained. When a nerve is soaked in a solution that contains Li, the K⁴² uptake is inhibited, and the nerve loses K and gains Li. When a Li-loaded nerve recovers in a Li-free solution, K is taken up in exchange for Li. This uptake of K requires Na in the external solution. It is concluded that the active transports of K and of Na may be due to different processes, that an accumulation of K occurs only in exchange for an intracellular cation, which need not be Na, and that Na plays a specific, but unknown, role in K transport.     

Effect of N-source on soybean leaf sucrose phosphate synthase, starch formation, and whole plant growth

Soybeans (Glycine max L. Merr. cv Tracy and Ransom) were grown under N₂-dependent or NO₃⁻-supplied conditions, and the partitioning of photosynthate and dry matter was characterized. Although no treatment effects on photosynthetic rates were observed, NO₃⁻-supplied plants in both cultivars had lower starch accumulation rates than N₂-dependent plants. Leaf extracts of NO₃⁻-supplied plants had higher activities of sucrose phosphate synthase (SPS) and cytoplasmic fructose-1,6-bisphosphatase (FBPase) than N₂-dependent plants. The variation in starch accumulation was correlated negatively with the activity of SPS, but not the activity of FBPase, UDP-glucose pyrophosphorylase, or ADP-glucose pyrophosphorylase. These results suggested that starch accumulation is biochemically controlled, in part, by the activity of SPS. Leaf starch content at the beginning of the photoperiod was lower in NO₃⁻-supplied plants than N₂-dependent plants in both cultivars which suggested that net starch utilization as well as accumulation was affected by N source.

Total dry matter accumulation and dry matter distribution was affected by N source in both cultivars, but the cultivars differed in how dry matter was partitioned between the shoot and root as well as within the shoot. The activity of SPS was correlated positively with total dry matter accumulation which suggested that SPS activity is related to plant growth rate. The results suggested that photosynthate partitioning is an important but not an exclusive factor which determines whole plant dry matter distribution.

     

Hemisodium, a novel selective Na ionophore. Effect on normal human erythrocytes.

Hemisodium is a novel Na ionophore that belongs to the class of compounds called cryptands. These compounds possess an electron-rich cavity for binding of cations and are conformationally organized during synthesis to favor the selective binding of one cation over another. In media containing 145 mM NaCl and 5 mM KCl, hemisodium (10(-5) M) increased erythrocyte Na content from 23 to 345 mmol/kg.dry cell solid (dcs) over 4 h and increased water content from 1.8 to 3.5 liter/kg.dcs over the same period. K content decreased somewhat over the same time period, but this fall in K content was prevented entirely by incubation in either low Na media (to prevent net Na entry) or in Cl free media. Thus, the decrease in K content in high NaCl media was due to cell swelling, which activated KCl cotransport, and not due to a direct action of hemisodium on K permeability. Hemisodium-mediated Na transport was conductive, because erythrocyte membrane potential (Vm), determined by diS-C3-5 fluorescence, changed from -9 to +22 mV in high Na media in the presence of hemisodium and DIDS. In cells equilibrated with sulfamate, an anion with low conductive permeability, Vm changed 54 mV per 10-fold change in external Na concentration with the addition of hemisodium. In contrast, a 10-fold change in the external concentration of K, Rb, Cs, or T1 failed to alter Vm in the presence of hemisodium, suggesting a high Na specificity of the ionophore. Na conductance determined from net fluxes increased from 0.04 to 5.2 microS/cm2 with 10 microM hemisodium, and with that concentration the ratio of Na to K conductance was 45:1. Among the Na ionophores available so far, hemisodium appears to have the greatest specificity. Hemisodium may be a valuable tool in membrane transport studies.     

Changes in k, rb, and na transport to shoots after anoxia

The effect of anoxia on subsequent uptake and transport of K, Rb, and Na was examined with seedlings of barley (Hordeum vulgare L.), corn (Zea mays L.), and tall fescue (Lolium × Festuca hybrid derivative) to further our understanding of xylem loading. Roots were incubated in solutions depleted of O₂ by flushing with N₂ gas. After 1 hour exposure, plants were returned to aerated solutions for 16 hours prior to measuring uptake and transport. For each species, anoxia pretreatment significantly enhanced Na transport to the shoot. The rate of Na accumulation into roots, however, was not affected. There was no enhancement of either K or Rb accumulation in shoots, indicating specificity for Na transport. A minimum exposure to anoxia of 30 minutes and a minimum of 12 hours elapsed time was necessary to achieve the maximum rate of Na transport to the shoot in barley seedlings. Accumulation of Na in the shoot of both the control and anoxia pretreated barley plants was inhibited by anoxia and by addition of the proline analog, l-azetidine-2-carboxylic acid, during the uptake period. Enhancement of Na transport was associated with a proportional increase in the rate of synthesis of a membrane bound protein with a molecular weight of 78,000 daltons.     

Cation transport in vascular endothelial cells and aging

Summary To understand the generation and maintenance of Na and K gradients in cultured vascular endothelial cells, net Na and K movements were studied. Ouabain-sensitive (OS) net Na gain and K loss were estimated as the difference between the cation content in the presence of ouabain and that in the control. Ouabain-and furosemide-resistant (OFR) fluxes were determined in the presence of the two inhibitors. When the normal medium bicarbonate and phosphate buffers were replaced by N-2-hydroxyethylpiperazine-N'-2-ethane sulfonic acid both the OS ans OFR fluxes decreased more than 50%. Ouabain-sensitive and ouabain-and furosemide-resistant fluxes decreased with increasing cellular age (passage number) an effect not observed when the cation movements were studied in the absence of bicarbonate and phosphate. These results suggest that cultured vascular endothelial cells possess bicarbonate-and phosphate-dependent Na and K pathways which account for a significant portion of their passive movements. Furthermore, the behavior of cation permeabilities with passage number suggests that these modulations may be related to the cellular aging process.     

Sodium Extrusion and Potassium Uptake in Guinea Pig Kidney Cortex Slices

Slices from the cortex corticis of the guinea pig kidney were immersed in a chilled solution without K and then reimmersed in warmer solutions. The Na and K concentrations and the membrane potential V_m were then studied as a function of the Na and K concentrations of the reimmersion fluid. It was found that Na is extruded from the cells against a large electrochemical potential gradient. Q₁₀ for net Na outflux was ∼2.5. At bath K concentrations larger than 8 mM the behavior of K was largely passive. At the outset of reimmersion (V_m > E_K) K influx seemed secondary to Na extrusion. Na extrusion would promote K entrance, being limited and requiring the presence of K in the bathing fluid. At bath K concentrations below 8 mM, K influx was up an electrochemical potential gradient. Thus a parallel active K uptake is apparent. Q₁₀ for net K influx was ∼2.0. Dinitrophenol inhibited net Na outflux and net K influx, Q₁₀ became <1.1 for both fluxes. The ratio between these fluxes varied. Thus at the outset of reimmersion the net Na outflux to net K influx ratio was >1. After 8 minutes it was <1.     

Cation Transport in Escherichia coli : I. Intracellular Na and K concentrations and net cation movement

Methods have been developed to study the intracellular Na and K concentrations in E. coli, strain K-12. These intracellular cation concentrations have been shown to be functions of the extracellular cation concentrations and the age of the bacterial culture. During the early logarithmic phase of growth, the intracellular K concentration greatly exceeds that of the external medium, whereas the intracellular Na concentration is lower than that of the growth medium. As the age of the culture increases, the intracellular K concentration falls and the intracellular Na concentration rises, changes which are related to the fall in the pH of the medium and to the accumulation of the products of bacterial metabolism. When stationary phase cells, which are rich in Na and poor in K, are resuspended in fresh growth medium, there is a rapid reaccumulation of K and extrusion of Na. These processes represent oppositely directed net ion movements against concentration gradients, and have been shown to be dependent upon the presence of an intact metabolic energy supply.     

Deoxygenation-induced cation fluxes in sickle cells: relationship between net potassium efflux and net sodium influx

Deoxygenation-induced cation fluxes in sickle cells were studied by measuring net cation movements in ouabain-treated cells. These deoxy cation fluxes were highly dependent on pH, showing inhibition at pH less than 7 and greater than 8 and a maximum at 7.4-7.5. Activation occurred at oxygen tensions around 40-50 torr and fluxes rose sharply as PO2 fell lower. Deoxy K efflux paralleled deoxy Na influx at pH values between 7 and 8, and at all oxygen tensions. Sickle cells were separated by density on Percol-Stractan gradients. Dense cells had lower deoxy cation fluxes of both Na and K than did lighter cell fractions, but in none of the fractionated populations did deoxy K efflux exceed deoxy Na influx. These data demonstrate that deoxy cation fluxes are activated at physiological pH and oxygen tensions and that there are no conditions of pH and PO2 and no cell populations in which cation fluxes induced by deoxygenation contribute directly to net cation loss in sickle cells. Chloride replacement (with nitrate) did not alter deoxy cation fluxes, and deoxy K efflux did not require the presence of external Na (tetramethylammonium replacement). Thus, deoxy cation fluxes do not have the characteristics of a cation-chloride cotransport or cation countertransport system.     

The Effect of Abscisic Acid on the Uptake and Distribution of Ions in Intact Seedlings of Phaseolus vulgaris cv. Redland Pioneer

Abscisic acid (ABA) was found to increase the accumulation of ³⁶Cl, total Cl, ²²Na and total Na⁺ in roots of intact bean seedlings. After an initial promotion. ABA inhibited longdistance transport of these ions from the root to the shoot. However, it consistently inhibited both uptake and transport of ⁴²K and total K⁺ in intact bean seedlings. A promotion of net ³⁶Cl influx (ψ_oc) and its accumulation in the root (Q*_v) concomitant decrease in transport index (long-distance transport as percentage of total influx) showed that ABA stimulates -³⁶Cl transport at the tonoplast. It inhibited H⁴ extrusion and net ⁸⁶Rb influx which agrees with a cation exchange theory K⁺/Rb⁺ transport.     

Components of Sodium and Chloride Flux Across Toad Bladder

The effect of transepithelial potential difference (ψ) on Na and Cl flux across toad bladder was assessed by measuring isotopic flux between identical media at various values of ψ. The contribution of edge damage to ionic permeability was eliminated, resulting in relatively high spontaneous ψ (-97 ±4 mv) and low electrical conductance g. Bidirectional Na fluxes were measured simultaneously. Unidirectional Cl fluxes were measured in paired hemibladders at ψ = 0 mv or -97 mv. Net Na flux J_Na, at ψ = 0 mv, was slightly less than short-circuit current (SCC). At ψ = -97 mv, J_Na averaged 17% of SCC, and was sometimes zero. ΔJ_Na/Δψ (= g₊) averaged 60% of g between -97 mv and +75 mv; at -150 mv, g₊ fell, indicating rectification. Analysis of unidirectional Na fluxes indicates low passive conductance (1.5 μmho/mg wet weight), a bidirectional, electrically neutral flux of approximately 0.13 μa/mg, and relatively large conductance of the active transport path at ψ ≥ -97 mv. The absence of appreciable transstimulation of serosal (S)-to-mucosal (M) Na flux (in response to increasing mucosal Na concentration) indicates that the electrically neutral flux is not exchange diffusion in the usual sense. Analysis of Cl fluxes indicates similar values for passive conductance and neutral flux, suggesting linked neutral flux of Na and Cl. Either the electromotive force of the Na pump E, its conductance g_a, or both are strong functions of ψ. The product of these two quantities, Eg_a, is a measure of the “transport capacity” at any given value of ψ, independent of the direct effect of ψ on J_Na through the pump path. Eg_a varies with ψ. Hence estimation of the net Na flux or current at any one value of ψ, including ψ = 0, fails to reveal the maximal transport capacity of the pump, its resting electromotive force (when J_Na = 0 through the pump), or the dependence of transport capacity on potential.     

Ba2+-sensitive K+ channels in luminal-membrane vesicles from pars convoluta of rabbit proximal tubule

This paper describes properties of 86Rb+ fluxes through a novel K+ channel in luminal-membrane vesicles isolated from pars convoluta of rabbit proximal tubule. The uptake of 86Rb+ into potassium salt loaded vesicles was specifically inhibited by Ba2+. The isotope accumulation is driven by an electrical diffusion potential as shown in experiments using these membrane vesicles loaded with anions of different membrane permeability and was as follows: gluconate greater than SO4(2-) greater than Cl-. Furthermore, the vesicles containing the channels show a cation selectivity with the order K+ greater than Rb+ greater than Li+ greater than Na+ = choline+.     

Growth, Pigment Synthesis, and Ultrastructural Responses of Phaseolus vulgaris L. cv. Blue Lake to Intermittent and Flashing Light

Growing bean plants (Phaseolus vulgaris L. cv. Blue Lake) on cycles of 1 minute light-1 minute dark or 5 minutes light-5 minutes dark, providing an integrated 12 hours light-12 hours dark per day for each set of plants, led to production after 21 days of new leaves low or lacking in chloroplast pigments. Subsequently, dry weight increase was sharply cut. Leaf area was affected by the light regimes after the second week of growth. By the fourth week, plants on the 1 minute light-1 minute dark cycle showed about one-half the leaf area of the controls. Shoot growth was favored over root growth to the greatest degree on the 1 minute light-1 minute dark regimes. Chlorophyll a/b ratios were close to 3.0 in all of the intermittent light regimes, but the total amounts of chlorophyll in milligrams per primary leaf were higher from day 9 to day 23 for the 12 hour light-12 hours dark controls than for other plants.

Although they produced chlorophyll, the plants receiving 1 or 2 milliseconds per second of light continued to lose weight at the same rate as the dark controls; thus, it is assumed there was no net photosynthesis. Plants receiving flashing light allocated significantly more food reserves from the seed to roots than did dark controls. Total chlorophyll formation was significantly accelerated by 2 milliseconds per second light. With 1 millisecond per second light, it took 5 days longer to achieve the same level of chlorophyll. After the 18th day, there was a steady decline in chlorophyll, b degrading more rapidly than a.

It is thought that several light-driven reactions are involved in the observed pigment synthesis, photosynthesis, food allocation, and growth of bean. Some of these reactions may be cyclic and others linear. Collectively, they must reach a harmonic point for normal metabolism and development to occur. Because time courses for each of these reactions are different, the intermittent and flashing light technique offers the possibility of individually studying some of the key light-driven reactions.

     

Carbon Dioxide Fixation in Roots and Nodules of Alnus glutinosa: I. Role of Phosphoenolpyruvate Carboxylase and Carbamyl Phosphate Synthetase in Dark CO(2) Fixation, Citrulline Synthesis, and N(2) Fixation

Detached roots and nodules of the N₂-fixing species, Albus glutinosa (European black alder), actively assimilate CO₂. The maximum rates of dark CO₂ fixation observed for detached nodules and roots were 15 and 3 micromoles CO₂ fixed per gram dry weight per hour, respectively. The net incorporation of CO₂ in these tissues was catalyzed by phosphoenolpyruvate carboxylase which produces organic acids, some of which are used in the synthesis of the amino acids, aspartate, glutamate, and citrulline and by carbamyl phosphate synthetase. The latter accounts for approximately 30 to 40% of the CO₂ fixed and provides carbamyl phosphate for the synthesis of citrulline. Results of labeling studies suggest that there are multiple pools of malate present in nodules. The major pool is apparently metabolically inactive and of unknown function while the smaller pool is rapidly utilized in the synthesis of amino acids. Dark CO₂ fixation and N₂ fixation in nodules decreased after treatment of nodulated plants with nitrate while the percentage of the total ¹⁴C incorporated into organic acids increased. Phosphoenolpyruvate carboxylase and carbamyl phosphate synthetase play key roles in the synthesis of amino acids including citrulline and in the metabolism of N₂-fixing nodules and roots of alder.     

Effects of Fusicoccin in 'Isolated' Guard Cells of Commelina communis L

The effects of 3 ? 10^–2 mol m^–3 FC on rubidium fluxesand contents in isolated guard cells of Commelina communis L.have been studied using ⁸⁶RbCl. Fusicoccin causes a marked stimulationof influx and an immediate, apparently irreversible, decreasein efflux of ⁸⁶Rb. The effect on influx is usually more importantin determining the new net flux of Rb. Observed fluxes differmarkedly from those predicted by the Goldman-Hodgkin-Katz equation,suggesting that FC does not act solely via an effect upon theplasmalemma potential. Fusicoccin appears to have a more directeffect upon the ion movements associated with changes in stomatalaperture than either ABA or transfer to the dark. Observed changesin Rb content cannot account fully for the osmotic changes associatedwith aperture increase. Key words: Fusicoccin, Guard cells, Ion fluxes, Commelina communis