Influence of chronic alterations of salt intake and aging on the kinetic of red cell Na+ and K+ transport in Sprague-Dawley rats

The kinetics of Na+ and K+ (Rb)+ transport mediated by the Na(+)-K+ pump and Na(+)-K+ cotransport system (assessed as a function of Rb+o and Na+i) as well as the magnitude of cation leaks were determined in red cells of young male rats subjected to chronic salt deprivation or salt loading (0.1% and 8% NaCl diet). These salt intake alterations induced moderate kinetic changes of the Na(+)-K+ pump which did not result in significant changes of ouabain-sensitive (OS) Rb+ uptake or Na+ net extrusion at in vivo Na+i and K+o concentrations because a decreased affinity for Na+i in salt-loaded animals was compensated by an increased maximal transport rate. High furosemide-sensitive (FS) Rb+ uptake in red cells of salt-deprived rats was caused by an increase of both the maximal transport rate and the affinity for Rb+o. Cation leaks were also higher in salt-deprived than in salt-loaded rats. In three age groups of rats fed a 1% NaCl diet FS Rb+ uptake (but not FS Na+ net uptake) rose with age due to an increasing maximal transport rate whereas the affinity of the cotransport system for Rbo+ did not change. The age-dependent changes in the kinetics of the Na(+)-K+ pump resulted in a slight decrease of OS Rb+ uptake with age that was not paralleled by corresponding Na+ net extrusion. No major age-related changes of cation leaks were found. Thus some intrinsic properties of red cell transport systems can be altered by salt intake and aging.     

Ion transport in erythrocytes of Prague hypertensive rat

The Prague hypertensive rat is a unique strain exhibiting genetic hypertension in which a hypertensive line (PHR) was bred in parallel with a normotensive one (Prague normotensive rat--PNR) from the same parental pair. Sodium efflux from Na(+)-loaded erythrocytes into Mg2(+)-sucrose medium was measured in these two strains as well as in spontaneously hypertensive rats (Okamoto-Aoki, SHR) and in normotensive outbred Wistar rats. Kinetic parameters--maximal velocity and apparent dissociation constant (reflecting the affinity for internal sodium)--were calculated. It was found that PHR as well as SHR had a higher Na+ leak and a decreased activity of the ouabain-sensitive Na+ transport as compared to Wistar rats. Furosemide-sensitive Na+ transport was substantially lower in erythrocytes of both hypertensive strains (PHR and SHR) than in the respective normotensive strains (PNR and Wistar).     

Evidence for a raised concentration of a circulating sodium transport inhibitor in essential hypertension.

A cytochemical technique that measures the ability of plasma to stimulate guinea-pig renal glucose-6-phosphate dehydrogenase (G6PD) activity in vitro, which is a marker of its ability to inhibit Na+-K+-adenosine-triphosphatase (Na+-K+-ATPase), was used in 19 patients with essential hypertension and 23 normotensive, healthy subjects. The ability of plasma to stimulate G6PD was significantly greater in the hypertensive patients when they were taking their normal sodium diet than in the normotensive subjects, and was significantly correlated with blood pressure. The ability of plasma to stimulate G6PD was inversely correlated with plasma renin activity in the hypertensive patients and increased with age and sodium intake in the normotensive subjects. These results support the hypothesis that essential hypertension, and also perhaps the increase in blood pressure with age in communities that consume large quantities of salt, is in part due to an increase in a circulating concentration of an inhibitor of Na+-N+-ATPase.     

Na+/H+ exchange activity in the plasma membrane of Arabidopsis

In plants, Na+/H+ exchangers in the plasma membrane are critical for growth in high levels of salt, removing toxic Na+ from the cytoplasm by transport out of the cell. The molecular identity of a plasma membrane Na+/H+ exchanger in Arabidopsis (SOS1) has recently been determined. In this study, immunological analysis provided evidence that SOS1 localizes to the plasma membrane of leaves and roots. To characterize the transport activity of this protein, purified plasma membrane vesicles were isolated from leaves of Arabidopsis. Na+/H+ exchange activity, monitored as the ability of Na to dissipate an established pH gradient, was absent in plants grown without salt. However, exchange activity was induced when plants were grown in 250 mm NaCl and increased with prolonged salt exposure up to 8 d. H+-coupled exchange was specific for Na, because chloride salts of other monovalent cations did not dissipate the pH gradient. Na+/H+ exchange activity was dependent on Na (substrate) concentration, and kinetic analysis indicated that the affinity (apparent Km) of the transporter for Na+ is 22.8 mm. Data from two experimental approaches supports electroneutral exchange (one Na+ exchanged for one proton): (a) no change in membrane potential was measured during the exchange reaction, and (b) Na+/H+ exchange was unaffected by the presence or absence of a membrane potential. Results from this research provide a framework for future studies into the regulation of the plant plasma membrane Na+/H+ exchanger and its relative contribution to the maintenance of cellular Na+ homeostasis during plant growth in salt.     

Physiological basis for the high salt tolerance of Debaryomyces hansenii.

The effects of KCl, NaCl, and LiCl on the growth of Debaryomyces hansenii, usually considered a halotolerant yeast, and Saccharomyces cerevisiae were compared. KCl and NaCl had similar effects on D. hansenii, indicating that NaCl created only osmotic stress, while LiCl had a specific inhibitory effect, although relatively weaker than in S. cerevisiae. In media with low K+, Na+ was able to substitute for K+, restoring the specific growth rate and the final biomass of the culture. The intracellular concentration of Na+ reached values up to 800 mM, suggesting that metabolism is not affected by rather high concentrations of salt. The ability of D. hansenii to extrude Na+ and Li+ was similar to that described for S. cerevisiae, suggesting that this mechanism is not responsible for the increased halotolerance. Also, the kinetic parameters of Rb+ uptake in D. hansenii (Vmax, 4.2 nmol mg [dry weight]-1 min-1; K(m), 7.4 mM) indicate that the transport system was not more efficient than in S. cerevisiae. Sodium (50 mM) activated the transport of Rb+ by increasing the affinity for the substrate in D. hansenii, while the effect was opposite in S. cerevisiae. Lithium inhibited Rb+ uptake in D. hansenii. We propose that the metabolism of D. hansenii is less sensitive to intracellular Na+ than is that of S. cerevisiae, that Na+ substitutes for K+ when K+ is scarce, and that the transport of K+ is favored by the presence of Na+. In low K+ environments, D. hansenii behaved as a halophilic yeast.     

Effects of sodium chloride stress and calcium supply on growth, potassium uptake, and internal chloride and sodium levels of winter wheat seedlings

The effects of saline conditions on the K+ (86Rb), Na+ and Cl- uptake and growth of 6-day-old wheat (Triticum aestivum L. cv. GK Szeged) seedlings were studied in the absence and presence of Ca2+. It was found that on direct NaCl treatment the K+ uptake of the roots in the absence of Ca2+ declined significantly with increasing salinity. The reverse was true, however, in the case of NaCl pretreatment: seedlings grown under highly saline conditions (50 mM NaCl) absorbed more K+ than those pretreated with low levels of NaCl (1 or 10 mM NaCl). The data indicate a definite Na(+)-induced K+ uptake inhibition and/or feed-back regulation in the K+ uptake of roots under the above-mentioned growth conditions. As regards the Ca2+ effect, it was established that supplemental Ca2+ counteracts the unfavourable effect of saline conditions as concerns both the K+ uptake of the roots and the dry matter yield of the seedlings. The internal concentrations of Na+ and Cl- in the seedlings increased in proportion to increasing salinity. Marked differences were experienced, however, in the internal concentrations of Na+ and Cl- in the roots and shoots, respectively. It was concluded that under these experimental conditions the salt tolerance of wheat could be related to its capability of restricting the transport of Na+ at low and moderate levels to the shoots, where it is highly toxic.     

The excretion of salt load by the developing chick embryo

Salt loads (0.17 or 0.34 mmol Na+; 6 M NaCl solution labelled with 24Na) were administered into the amnion of 7-day-old chick embryos. The 24Na distribution in embryonic blood, amniotic and allantoic fluids was measured in 1, 2, 4, 8, 12 and 24 h intervals to assess the kinetics of salt load movements in particular egg compartments. The aim was to estimate the efficiency of the embryonic homeostatic apparatus to maintain ionic balance in the internal environment of the embryonic body. The Na+ concentration in amniotic fluid was expected to rise after salt loading by about 275 and 400 mM, respectively. More than 10% of the salt dose per ml appeared in the embryonic blood 2 h after salt load administration while only 0.2% were found in the urine (collected as allantonic fluid). The maximal rise of 24Na activity in the blood of salt-loaded embryos reached 11%-12% of the dose which corresponded to an increase of Na+ concentration by 19 and 41 mM, respectively. The maximum of 24Na activity appeared in the allantoic fluid with a delay of several hours and indicated an increment of Na+ concentration by 6% and 9% of the dose per ml in the case of salt-loaded embryos. The Na+ concentration in the allantoic fluid (urine) never exceeded that in the blood. The final Na+ activity (estimated in the blood 24 h after salt loading) was equal to 5% of the dose per ml in both cases, indicating a persistent elevation of Na+ concentration by 8.6 and 17.2 mM, respectively.     

盐胁迫对药用蒲公英生长特性及有效成分积累与离子吸收分配效应的影响

以药用蒲公英(Taraxacum officinale)为试材,研究不同浓度盐胁迫对其生长特性、有效成分积累和离子吸收分配的影响。结果表明,低盐胁迫(0.1%NaCl)对药用蒲公英生长和菊苣酸含量无显著影响,叶中Na⁺含量与对照无显著差异,K⁺含量及K⁺/Na⁺显著升高;高盐胁迫(≥0.2%NaCl)下其生长受到显著抑制,菊苣酸含量显著降低,类囊体膜结构随着盐胁迫加剧趋于紊乱,光合能力减弱,叶片Na⁺含量显著上升,而K⁺、Ca²⁺和Mg²⁺含量下降,K⁺/Na⁺、Ca²⁺/Na⁺和Mg²⁺/Na⁺显著降低。离子运输选择性系数(S_Ca,Na、S_Mg,Na、S_K,Na)随着盐胁迫加剧呈先升后降趋势。相关性分析表明,盐胁迫下...     

On the interaction of NH+4 and Na+ fluxes in the isolated trout gill.

1. Sodium influx was measured in isolated, previously perfused gill arches of rainbow trout, Salmo gairdneri, by measuring incorporation of 22Na into gill tissue following timed exposure to a 1 mM 22NaCl medium. Transport rates approximated those estimated for intact fish and were linear for at least one min. 2. NH4Cl-containing perfusates at pH 7 and 8 stimulated Na+ influx equally, indicating that only ionized ammonia is important in the transport process. A Na+/NH4+ exchange at basal and/or lateral membranes of the transporting cells is suggested. 3. Low-sodium Ringer perfusate augmented Na+ influx; in one group of gills the transport rate was more than double that of NaCl Ringer controls. The increase in transport induced by internal NH4+ was not additive with the low sodium augmentation. A reduction in intracellular (Na+) is postulated as the mechanism operating in both cases. 4. Ouabain had no appreciable effect on Na+ influx, either with or without NH4+ in the perfusate. Diamox partially blocked the augmented Na+ influx induced by NH4+. Amiloride completely inhibited Na+ influx, both with and without NH4+ in the perfusate.     

Modifications in choline transport activity as a function of membrane potential and the sodium gradient

Advantage was taken of a preparation of proteoliposomes made using Torpedo presynaptic membranes in which both the internal and external media can be controlled to investigate the effects of membrane potential and the Na+ gradient on choline transport activity. Under control conditions, Na+ outside and K+ inside, choline was concentrated by proteoliposomes and this phenomenon was sensitive to hemicholinium-3 and high levels of external choline. While proteoliposomes showed no permeability towards K+ spontaneously, in the presence of valinomycin a transmembrane potential was developed. The rate of transport was higher, the greater the inside negative potential. Both the affinity and the maximal velocity of high affinity transport rose in the presence of a potential. Likewise, the affinity and velocity of this transporter increased with increasing external Na+. Increasing internal Na+, on the other hand, caused a decrease in affinity and had little effect on the maximal velocity. The low affinity component was much less, if at all, affected by these changes. These results are consistent with a model of high affinity choline transport in which Na+ binds before choline and the carrier-Na+-choline complex is positively charged. However, these results do not provide a direct explanation for choline transport activation by nerve activity, underlining the need to study the effects of parameters other than membrane potential and the Na+ gradient on choline transport activity.     

NMR studies on Na+ transport in Synechococcus PCC 6311.

The freshwater cyanobacterium Synechococcus PCC 6311 is able to adapt to grow after sudden exposure to salt (NaCl) stress. We have investigated the mechanism of Na+ transport in these cells during adaptation to high salinity. Na+ influx under dark aerobic conditions occurred independently of delta pH or delta psi across the cytoplasmic membrane, ATPase activity, and respiratory electron transport. These findings are consistent with the existence of Na+/monovalent anion cotransport or simultaneous Na+/H+ +anion/OH- exchange. Na+ influx was dependent on Cl-, Br-, NO3-, or NO2-. No Na+ uptake occurred after addition of NaI, NaHCO3, or Na2SO4. Na+ extrusion was absolutely dependent on delta pH and on an ATPase activity and/or on respiratory electron transport. This indicates that Na+ extrusion via Na+/H+ exchange is driven by primary H+ pumps in the cytoplasmic membrane. Cells grown for 4 days in 0.5 m NaCl medium, "salt-grown cells," differ from control cells by a lower maximum velocity of Na+ influx and by lower steady-state ratios of [Na+]in/[Na+]out. These results indicate that cells grown in high-salt medium increase their capacity to extrude Na+. During salt adaptation Na+ extrusion driven by respiratory electron transport increased from about 15 to 50%.     

Erythrocyte membrane permeability for univalent cations (Na+, K+) and their transformation in patients with hypertension and symptomatic (renal) hypertension

The form and surface architectonic of erythrocytes studied in 18 teenagers and men with hypertensive disease of I stage (HD) and in 7 men with symptomatic (renal) hypertension (SH). Simultaneously permeability of erythrocyte membranes for Na+ and K+ was studies. The change in the form and surface architectonics was found in the erythrocytes of the patients with hypertensive disease, I stage. The same was true for the patients with renal hypertension but the difference was not so prominent. Rapid shift of correlation of erythrocyte morphological varieties has been revealed during the study of Na+ and K+ ions' transport rate after the treatment by p-chloromercuribenzoate acid. Increase of irreversible transformation of erythrocytes was discovered in patients with HD. Besides in some cases there was decrease in the size of echinocytes. The transformation of erythrocytes into echinocytes is more prominent in healthy subjects and in patients with SH. Our data suggest that the change in erythrocyte form is related to the change of erythrocyte membrane permeability to Na+ and K+ ions and the alteration of membrane structure is the basis for these disturbances.     

Sodium/calcium exchange in smooth-muscle microsomal fractions.

The existence of Na+ -dependent Ca2+ transport was investigated in microsomal fractions from the longitudinal smooth muscle of the guinea-pig ileum and from the rat aorta, and its activity was compared with that of the plasmalemmal ATP-dependent Ca2+ pump previously identified in these preparations. The rate of Ca2+ release from plasmalemmal vesicles previously loaded with Ca2+ through the ATP-dependent Ca2+ pump was transiently faster in the presence of 150 mM-NaCl in the medium than in the presence of 150 mM-KCl or -LiCl or 300 mM-sucrose. Na+-loaded vesicles took up Ca2+ when an outwardly directed Na+ gradient was formed across the membrane. The Ca ionophore A23187 induced a rapid release of 85% of the sequestered Ca2+, whereas only 15% was displaced by La3+. Ca2+ accumulated by the Na+-induced Ca2+ transport was released by the addition of NaCl, but not KCl, to the medium. Ca2+ uptake in Na+-loaded vesicles was inhibited in the presence of increasing NaCl concentration in the medium. Half-maximum inhibition was observed with 28 mM-NaCl. Data fitted the Hill equation, with a Hill coefficient (h) of 1.9. Na+-induced Ca2+ uptake was a saturable function of Ca2+ concentration in the medium. Half-maximum activity was obtained with 18 microM-Ca2+ in intestinal-smooth-muscle microsomal fraction and with 50 microM-Ca2+ in aortic microsomal fraction. The results suggest that in these membrane preparations a transmembrane movement of Ca2+ can be driven by a Na+ gradient. However, the Na+-induced Ca2+ transport had a lower capacity, a lower affinity and a slower rate than the ATP-dependent Ca2+ pump.     

Effects of NaCl concentration on adenylate cyclase regulation by prostaglandins and guanine nucleotides

Na+ has been implicated as a requirement for the inhibition of adenylate cyclase by hormones and neurotransmitters. This study examines effects of salt concentration on neuroblastoma plasma membranes that occur in the absence of an inhibitory hormone. The adenylate cyclase response to stimulatory agonists (GTP plus PGE1 (3), PGI2 or PGE2) was influenced by NaCl. As the [NaCl] increased to 150 mM, an increase in maximal activity and a decrease in apparent affinity was observed. At concentrations above 150 mM, NaCl decreased prostaglandin affinity and progressively decreased maximal activation. The GTP requirement was not altered by 30 or 150 mM NaCl in the presence of PGE1 or PGI2. The rate of Gpp(NH)p stimulated activity increased as the [NaCl] was increased in the assay. This increased rate was conserved when membranes activated in the presence of Gpp(NH)p and NaCl were reassayed in the absence of guanine nucleotide or salt. The salt evoked rate increase was proportionally greater at submaximal MgCl2 concentrations. The concentration requirement for Mg2+ was reduced by salt for adenylate cyclase in the presence of GTP or Gpp(NH)p. However, the enzyme stimulated by hormone exhibited a Mg2+ requirement that was low in the absence of salt and could not be further reduced by increased [NaCl]. Alternative monovalent cations (150 mM Li+, K+, Cs+, but not choline or tetramethylammonium) and anions (SO4=) substituted for NaCl. The observed effects were reversible upon washing the membranes and neither ouabain nor tetrodotoxin altered the response. These effects may result from a conformational alteration of a protein particularly sensitive to neutral salts in the assay.     

转HAL1基因番茄的耐盐性

利用农杆菌介导的叶盘法,把HAL1 基因转入番茄,Southern杂交检测得到转基因植株.耐盐实验表明, T1代转基因番茄在150 mmol/L的NaCl胁迫下仍有43%的发芽率,200 mmol/L的NaCl胁迫下发芽率为6%,而对照种子在100和150 mmol/L的NaCl胁迫下发芽率分别为11.0%和0.转基因番茄的电解质相对外渗率小于对照,而根冠比和叶绿素含量大于对照,转HAL1基因显著提高了番茄的耐盐性.盐胁迫下Na 、K 的累积状况表明,转基因番茄根、茎、叶的K /Na 均有所提高,根系的SK/Na增大,茎、叶的RSK/Na和RLK/Na减小,说明根系对K /Na 离子的选择吸收和运输能力加强.不但选择吸收K /Na ,而且表现出整株水平上的有利于耐盐的K /Na 区域化分配.     

Function of transport H+-ATPases in plant cell plasma and vacuolar membranes of maize under salt stress conditions and effect of adaptogenic preparations

Participations of electrogenic H+-pumps of plasma and vacuolar membranes represented by E1-E2 and V-type H+-ATPases in plant cell adaptation to salt stress conditions has been studied by determination of their transport activities. Experiments were carried out on corn seedlings exposed during 1 or 10 days at 0.1 M NaCl. Preparations Methyure and Ivine were used by seed soaking at 10(-7) M. Plasma and vacuolar membrane fractions were isolated from corn seedling roots. In variants without NaCl a hydrolytical activity of plasma membrane H+-ATPase was increased with seedling age and its transport one was changed insignificantly, wherease the response of the weaker vacuolar H+-ATPase was opposite. NaCl exposition decreased hydrolytical activities of both H+-ATPases and increased their transport ones. These results demonstrated amplification of H+-pumps function especially represented by vacuolar H+-ATPase. Both preparations, Methyure mainly, caused a further increase of transport activity which was more expressed in NaCl variants. Obtained results showed the important role of these H+-pumps in plant adaptation under salt stress conditions realized by energetical maintenance of the secondary active Na+/H+ -antiporters which remove Na+ from cytoplasm.     

Active phosphate ion transport in plasma membrane vesicles isolated from mouse fibroblasts.

Inorganic phosphate accumulated 8-fold in plasma membrane vesicles derived from simian virus 40-transformed 3T3 mouse fibroblasts when a NaCl gradient (external greater than internal) was artificially imposed across the membrane. Preincubation with Na+ or addition of monensin markedly reduced phosphate accumulation. Na+-stimulated phosphate transport was not affected by addition of either dicarboxylic acids, antimycin A, or ouabain and persisted after addition of proton ionophores. The coupling of phosphate transport to Na+ gradients was pH-dependent, with maximal stimulation by Na+ below pH 7. These findings suggest that monovalent phosphate anion moves across the plasma membrane in co-transport with sodium ion.     

Protein phosphatase type 2B (calcineurin)-mediated, FK506-sensitive regulation of intracellular ions in yeast is an important determinant for adaptation to high salt stress conditions.

To assess the physiological function of Ca(2+)-dependent protein phosphatase (PP2B) in the yeast Saccharomyces cerevisiae, the phenotypes of PP2B-deficient mutants were investigated. Although PP2B was dispensable for growth under normal conditions, the mutations did, however, cause growth inhibition under certain stress circumstances. The growth of the mutants was inhibited by NaCl and LiCl, but not by KCl, CaCl2, MgCl2 or nonspecific osmotic stresses. Upon shift to high NaCl medium, intracellular Na+ levels of both wild type yeast and the mutants initially increased at a comparable rate. However, internal Na+ in wild type cells started to decline more rapidly than the mutant cells during cultivation in high NaCl medium, indicating that PP2B is important in maintaining a gradient across the membrane. The protection against salt stress was achieved, at least in part, by the stimulation of Na+ export. The maintenance of a high level of internal K+ in high NaCl medium was also PP2B-dependent. In the presence of the immunosuppressant FK506, the growth behaviour and intracellular Na+ and K+ of wild type cells in high NaCl medium became very similar to those of the PP2B-deficient mutant in a manner dependent on the presence of the FK506 binding protein.     

Evaluation of intra-erythrocyte Na+ activity in persons at high risk for essential arterial hypertension and as an aid in differential diagnosis from secondary forms of hypertension

The intracellular "Na+ activity" was measured in erythrocytes of normotensive subjects (46), in essential hypertensive patients (18), in their children (20) and in patients with secondary hypertension (8). In normotensive subjects without a genetic trait of hypertension intracellular "Na+ activity" was 7.3 +/- 0.8 mmol/l, in secondary hypertensive patients was 7.5 +/- 0.6 mmol/l, in essential hypertensive patients was 10.9 +/- 1.1 mmol/l and in their children was 8.6 +/- 2.1 mmol/l. In this group (children) it was possible to differentiate between 2 population, the 1 degree with height intracellular "Na+ activity" (8); the 2 degrees with normal intracellular "Na+ activity".     

Modulation of erythrocyte Na transport pathway(s) by excess Na intake

Different Na transport pathways were studied in the erythrocytes of 10 normotensive subjects who received 240 meq/day of Na in excess of their usual diet. In most of these subjects the maximal rate (Vmax) of the Na,K pump and the Na,K-cotransport system was markedly decreased on the first day of the diet. In some of these subjects, excess Na intake induced an increase in the apparent affinity for internal Na for the Na,K pump and the Na,K-cotransport system. The decrease in the Na,K pump fluxes was not concomitant to that of the co-transport system and not accompanied with an increase in blood pressure or cation concentration in the plasma. Interestingly, the apparent affinity for internal Li of the Li-Na exchange was markedly increased without alteration of the Vmax. The passive permeability for Na and the cellular Na content were not altered by excess Na intake. Ouabain and bumetanide at low concentrations respectively induced an increase in the apparent affinity for internal Na of the Na,K pump and the Na,K- cotransport system. These results are similar to those observed after excess Na intake. These later agree with the hypothesis that Na homeostasis regulates some endogenous factors with ouabain-like and furosemide-like properties that might contribute to the regulation of cellular Na handling.