高浓度钾抑制杜氏盐藻生长的生理机制

在含１ｍｏｌ／ＬＮａＣｌ的杜氏盐藻（Ｄｕｎａｌｉｅｌａｓａｌｉｎａ（Ｄｕｎａｌ）Ｔｅｏｄ．）培养液中加入５０ｍｍｏｌ／Ｌ以上的ＫＣｌ可观察到Ｋ＋对杜氏盐藻生长有明显的抑制作用,而当ＫＣｌ达１００ｍｍｏｌ／Ｌ时,杜氏盐藻的生长被完全抑制。另一方面,当培养液中缺乏Ｋ＋时,杜氏盐藻的生长也被显著抑制。在正常培养条件下,伴随着杜氏盐藻的生长,培养液的ｐＨ由８左右升高至１０左右,而高浓度Ｋ＋则显著抑制杜氏盐藻培养液ｐＨ的升高;而在培养液ｐＨ为７．０至１０．０的范围内,不同ｐＨ对杜氏盐藻的生长无明显影响。将杜氏盐藻在高浓度Ｋ＋条件下预处理１２ｈ以上,杜氏盐藻的光合放氧速率显著下降,光合速率下降的程度与Ｋ＋浓度的高低和预处理的时间长短呈正相关。高浓度Ｋ＋处理也引起杜氏盐藻叶绿素含量的显著下降。对经高浓度Ｋ＋预处理的杜氏盐藻的光合放氧速率与培养液中ｐＨ变化同时进行测定的结果表明,Ｋ＋抑制杜氏盐藻光合速率的同时也显著抑制了光照条件引起的培养液ｐＨ的上升。实验结果表明,Ｋ＋抑制杜氏盐藻光合作用以及抑制杜氏盐藻生长与Ｋ＋影响跨盐藻质膜的质子运输之间可能存在一定关系。     

Effects of High K+ and Alkaline pH on Ultrastructure of Dunaliella salina Chloroplasts

It was reported that the growth of Dunaliella salina Teod. cultured in medium containing 1 mol/L NaC1 was almost completely inhibited by the addition of 100 mmol/L KC1. The high K+ (100 mmol/L KC1) treatment also significantly inhibited the photosynthetic rate of D. salina and decreased chlorophyll contents in algae. This study focuses on possible effects of high K+ or alkaline pH on the ultrastructural change of chloroplasts in D. salina. After D. salina was cultured in a medium containing 100 n,anol/L KC1 or in a medium with alkaline pH for 8 to 10 days, dramatic ultrastructural changes occurred in the chloroplasts including thylakoid swelling, volume increase of chloroplast, and significant accumulation of starch grains in chloroplasts. The results are consistent with our previous report indicating that the ultrastmctuml changes in chloroplast under high K + or alkaline pH may lead to an inhibitory effects on photosynthesis and overall growth of D. salina.     

Amiloride-sensitive sodium transport in lamprey red blood cells: evidence for two distinct transport pathways

To determine Na+/H+ exchange in lamprey erythrocyte membranes, the cells were acidified to pH(i) 6.0 using the K+/H+ ionophore nigericin. Incubation of acidified erythrocytes in a NaCl medium at pH 8.0 caused a considerable rise in 22Na+ influx and H+ efflux during the first 1 min of exposure. In addition, exposure of acidified red cells to NaCl medium was associated with rapid elevation of intracellular Na+ content. The acid-induced changes in Na+ influx and H+ efflux were almost completely inhibited by amiloride and dimethylamiloride. In native lamprey erythrocytes, amiloride-sensitive Na+ influx progressively increased as the osmolality of incubation medium was increased by addition of 100, 200, or 300 mmol/l sucrose. Unexpectedly, the hypertonic stress induced a small, yet statistically significant decrease in intracellular Na+ content in these cells. The reduction in the cellular Na+ content increased with hypertonicity of the medium. The acid- and shrinkage-induced Na+ influxes were inhibited by both amiloride and 5-(N-ethyl-N-isopropyl)-amiloride (EIPA) in a dose-dependent manner. For both blockers, the half-maximal inhibitory values (IC50) were much greater for the shrinkage-induced (44 and 15 micromol/l for amiloride and EIPA, respectively) than for the acid-induced Na+ influx (5.1 and 3.3 micromol/l, respectively). The data obtained are the first demonstration of the presence of a Na+/H+ exchanger with high activity in acidified (pH(i) 6.0) lamprey red blood cells (on average, 512 +/- 56 mmol/l cells/h, n = 13). The amiloride-sensitive Na+ influxes produced by hypertonic cell shrinkage and acid load are likely to be mediated by distinct ion transporters in these cells.     

Effect of intracellular Na+ on platelet aggregation and Ca2+ mobilization

The effects of ionophores, which can carry alkali metal cations, on platelet aggregation were examined. At an alkaline extracellular pH, alkali metal cation/H+ exchanger nigericin accelerated aggregation in K+-enriched medium, whereas it rather inhibited aggregation in Na+-enriched medium, even though the intracellular pH was only slightly alkaline. The inhibitory effect of Na+ on platelet aggregation was more clearly shown with the alkali metal cation exchanger gramicidin D. The ionophore had no effect or a slightly accelerative effect on aggregation in K+-enriched medium, whereas it significantly inhibited aggregation induced by thrombin, ADP and platelet activating factor in Na+-enriched medium. Fluorescence studies on fura-2-labeled platelets revealed that in Na+-enriched medium gramicidin D inhibited agonist-induced Ca2+ mobilization both in the presence and absence of extracellular Ca2+. These results suggest that the intracellular Na+ inhibits platelet aggregation by inhibiting Ca2+ mobilization.     

The Physiological Basis of Growth Inhibition of Halophytes by Potassium

Salt-dilution halophyte Suaeda salsa, salt-secretory halophytes Atriplex centrMasiatica and Limonium bicolor were treated with different concentrations of KC1 and isosmotic NaC1. It was found that the organic dry weight and net photosynthetic rate of these plants were inhibited by 100--500 and 100--400 mmol/L KC1 respectively, indicating the existance of a relationship between the growth inhibition by K+ and the decrease of photosynthesis in halophytes. After treatment with 100--400 mmol/L KCI, the content of K+ increased while that of Na+ decreased, but there were no change in osmotic potential and ability of osmotic adjustment. It proved that growth inhibition by K+ was related to Na+ deficiency in osmotic adjustment and the toxic effect of high K+ on cells. Further there was no direct relation between the decrease of growth and photosynthesys and the content of sugars in leaves. Moreover, the inhibition of ATPase activity by high K+ concentration was probably related to growth inhibition.     

The Physiological Relevance of Na+-Coupled K+-Transport

Plant roots utilize at least two distinct pathways with high and low affinities to accumulate K+. The system for high-affinity K+ uptake, which takes place against the electrochemical K+ gradient, requires direct energization. Energization of K+ uptake via Na+ coupling has been observed in algae and was recently proposed as a mechanism for K+ uptake in wheat (Triticum aestivum L.). To investigate whether Na+ coupling has general physiological relevance in energizing K+ transport, we screened a number of species, including Arabidopsis thaliana L. Heynh. ecotype Columbia, wheat, and barley (Hordeum vulgare L.), for the presence of Na+-coupled K+ uptake. Rb+-flux analysis and electrophysiological K+-transport assays were performed in the presence and absence of Na+ and provided evidence for a coupling between K+ and Na+ transport in several aquatic species. However, all investigated terrestrial species were able to sustain growth and K+ uptake in the absence of Na+. Furthermore, the addition of Na+ was either without effect or inhibited K+ absorption. The latter characteristic was independent of growth conditions with respect to Na+ status and pH. Our results suggest that in terrestrial species Na+-coupled K+ transport has no or limited physiological relevance, whereas in certain aquatic angiosperms and algae this type of secondary transport energization plays a significant role.     

Effects of pH on the interaction of ligands with the (H+ + K+)-ATPase purified from pig gastric mucosa

The effects of K+, Na+ and ATP on the gastric (H+ + K+)-ATPase were investigated at various pH. The enzyme was phosphorylated by ATP with a pseudo-first-order rate constant of 3650 min-1 at pH 7.4. This rate constant increased to a maximal value of about 7900 min-1 when pH was decreased to 6.0. Alkalinization decreased the rate constant. At pH 8.0 it was 1290 min-1. Additions of 5 mM K+ or Na+, did not change the rate constant at acidic pH, while at neutral or alkaline pH a decrease was observed. Dephosphorylation of phosphoenzyme in lyophilized vesicles was dependent on K+, but not on Na+. Alkaline pH increased the rate of dephosphorylation. K+ stimulated the ATPase and p-nitrophenylphosphatase activities. At high concentrations K+ was inhibitory. Below pH 7.0 Na+ had little or no effect on the ATPase and p-nitrophenylphosphatase, while at alkaline pH, Na+ inhibited both activities. The effect of extravesicular pH on transport of H+ was investigated. At pH 6.5 the apparent Km for ATP was 2.7 microM and increased little when K+ was added extravesicularly. At pH 7.5, millimolar concentrations of K+ increased the apparent Km for ATP. Extravesicular K+ and Na+ inhibited the transport of H+. The inhibition was strongest at alkaline pH and only slight at neutral or acidic pH, suggesting a competition between the alkali metal ions and hydrogen ions at a common binding site on the cytoplasmic side of the membrane. Two H+-producing reactions as possible candidates as physiological regulators of (H+ + K+)-ATPase were investigated. Firstly, the hydrolysis of ATP per se, and secondly, the hydration of CO2 and the subsequent formation of H+ and HCO3-. The amount of hydrogen ions formed in the ATPase reaction was highest at alkaline pH. The H+/ATP ratio was about 1 at pH 8.0. When CO2 was added to the reaction medium there was no change in the rate of hydrogen ion transport at pH 7.0, but at pH 8.0 the rate increased 4-times upon the addition of 0.4 mM CO2. The results indicate a possible co-operation in the production of acid between the H+ + K+-ATPase and a carbonic anhydrase associated with the vesicular membrane.     

Na+ for H+ exchange in rabbit erythrocytes

The effect of a transmembrane pH gradient on the ouabain, bumetanide, and phloretin resistant H+ efflux was studied in rabbit erythrocytes. Proton equilibration was reduced by the use of DIDS (125 microM) and acetazolamide (1 mM). H+ efflux from acid loaded erythrocytes (pHi = 6.1) was measured in a K+ (145 mM) medium, pH0 = 8.0, in the presence and absence of 60 microM 5,N,N-dimethyl-amiloride (DMA). The H+ efflux rate in a K+-containing medium was 116.38 +/- 4.5 mmol/l cell X hr. Substitution of Nao+ for Ko+ strongly stimulated H+ efflux to 177.89 +/- 7.9 mmol/l cell X hr. The transtimulation of H+ efflux by Nao+ was completely abolished by DMA falling to values not different from controls with an ID50 of about 8.6 X 10(-7) M. The sequence of substrate selectivities for the external transport site were Na greater than greater than greater than Li greater than choline, Cs, K, and Glucamine. The transport system has no specific anion requirement, but is inhibited by NO3-. The DMA sensitive H+ efflux was a saturable function of [Na+]o, with an apparent Km and Vmax of about 14.75 +/- 1.99 mM and 85.37 +/- 7.68 mmol/l cell X hr, respectively. However, the Nao+-dependent and DMA-sensitive H+ efflux was sigmoidally activated by [H+]i, suggesting that Hi+ interacts at both transport and modifier sites. An outwardly directed H+ gradient (pHi 6.1, pH = 8.0) also promoted DMA sensitive Na+ entry (61.2 +/- 3.0 mmol/l cell X hr) which was abolished when pHo was reduced to 6.0. The data is therefore consistent with the presence of a Na+/H+ exchange system in rabbit erythrocytes.     

Na+ requirement for growth, photosynthesis, and pH regulation in the alkalotolerant cyanobacterium Synechococcus leopoliensis

We have found that Na+ is required for the alkalotolerance of the cyanobacterium Synechococcus leopoliensis. Cell division did not occur at any pH in the absence of Na+, but cells inoculated into Na+-free growth medium at pH 6.8 did continue metabolic activity, and over a period of 48 h, the cells became twice their normal size. Many of these cells remained viable for at least 59 h and formed colonies on Na+ -containing medium. Cells grown in the presence of Na+ and inoculated into Na+ -free growth medium at pH 9.6 rapidly lost viability. An Na+ concentration of ca. 0.5 milliequivalents X liter-1 was required for sustained growth above pH 9.0. The Na+ requirement could be only partially met by Li+ and not at all by K+ or Rb+. Cells incubated in darkness in growth medium at pH 6.8 had an intracellular pH near neutrality in the presence or absence of Na+. When the external pH was shifted to 9.6, only cells in the presence of Na+ were able to maintain an intracellular pH near 7.0. The membrane potential, however, remained high (-120 mV) in the absence or presence of Na+ unless collapsed by the addition of gramicidin. Thus, the inability to maintain a neutral intracellular pH at pH 9.6 in the absence of Na+ was not due to a generalized disruption of membrane integrity.Even cells containing Na+ still required added Na+ to restore photosynthetic rates to normal after the cells had been washed in Na+ -free buffer at pH 9.6. This requirement was only partially met by Li+ and was not met at all by K+, Rb+, Cs+ Mg2+, or Ca2+. The restoration of photosynthesis by added Na+ occurred within 30 s and suggests a role for extracellular Na+. Part of our results can be explained in terms of the operation of an Na+/H+ antiporter activity in the plasma membrane, but some results would seem to require other mechanisms for Na+ action.     

Activation of Na<Superscript>+</Superscript>/H<Superscript>+</Superscript> exchange by protein phosphatase inhibitors in red blood cells of the frog<Emphasis Type="Italic"> Rana ridibunda</Emphasis>

The present study was designed to evaluate the role of protein phosphatases in regulation of sodium transport in the marsh frog erythrocytes using 22Na as a tracer. For this purpose the cells were treated with several known inhibitors of protein phosphatases. In standard isotonic medium, exposure of the cells to 10 mmol l(-1) NaF, 20 nmol l(-1) calyculin A or 0.1 mmol l(-1) cantharidin resulted in a significant (1.7-fold) increase in unidirectional ouabain-insensitive Na+ influx. The Na+ influx in frog red cells was progressively activated as the medium osmolality was increased by addition of 100, 200 or 300 mmol l(-1) sucrose to standard isotonic medium. The stimulatory effect of protein phosphatase blockers on Na+ influx was much higher in hypertonic medium containing 100 or 200 mmol l(-1) sucrose than that in isotonic medium. Stimulation of Na+ transport enhanced with increasing concentrations of calyculin A, and half-maximal activation (EC50) was obtained at 16 nmol l(-1). However, Na+ influx induced by strong hypertonic treatment (+300 mmol l(-1) sucrose) was not altered further in the presence of protein phosphatase inhibitors. The changes in Na+ influx evoked by protein phosphatase inhibitors and hypertonic treatment were associated with a rise in the intracellular Na+, but not K+, content. Enhancement in Na+ influx after addition of protein phosphatase blockers to cell suspension in isotonic or hypertonic media was almost completely inhibited by Na+/H+ exchange inhibitors, amiloride and ethyl-isopropyl-amiloride. The basal Na+ influx in frog erythrocytes in isotonic medium was relatively low (1.7 mmol/l cells/h) and not affected by 1 mmol l(-1) amiloride. Thus, the data obtained clearly indicate that Na+/H+ exchanger in the marsh frog red blood cells is under tight regulatory control, in all likelihood via protein phosphatases of types PP-1 and PP-2A.     

NaCl胁迫下高纬度移植桐花树幼苗的生理生态效应

通过砂培试验,研究了不同浓度NaCl(0、100、200、300和400 mmol·L-1)处理对高纬度移植桐花树幼苗生物量、离子吸收、碳氮代谢、叶片光合色素和叶片抗氧化系统的影响.结果表明:100 mmol·L-1NaCl处理对桐花树生长有轻微的促进作用,当NaCl浓度达到300mmol·L-1时,桐花树根、茎、叶器官的干鲜质量、根冠比、株高和基径均显著下降.高盐胁迫抑制了叶片抗氧化系统中超氧化物歧化酶(SOD)和过氧化物酶(POD)的活性,提高了丙二醛含量,降低了叶片中叶绿素、类胡萝卜素总量以及根茎叶的可溶性总糖和游离氨基酸总量.不同浓度NaCl胁迫下,桐花树根茎叶中Na+含量快速上升,K+含量相对下降,K+/Na+快速下降,导致各器官中离子平衡失调.当NaCl浓度高于300 mmol·L-1时,桐花树根茎叶的碳氮代谢运转失调,抑制了植株的正常生长,导致各器官的生物量显著下降.     

Effects of Salt Stress on Carbohydrate Metabolism in Desert Soil Alga Microcoleus vaginatus Gom.

The effects of salt stress on carbohydrate metabolism in Microcoleus vaginatus Gom., a cyanobacterium isolated from desert algal crusts, were investigated in the present study. Extracellular total carbohydrates and exopolysaccharides （EPS） in the culture medium produced by M. vaginatus increased significantly during the growth phase and reached a maximum during the stationary phase. The production of extracellular carbohydrates also significantly increased under higher salt concentrations, which was attributed to an increase in low molecular weight carbohydrates. In the presence of NaCI, the production of cellular total carbohydrates decreased and photosynthetic activity was impaired, whereas cellular reducing sugars, water-soluble sugars and sucrose content and sucrose phosphate synthase activity increased, reaching a maximum in the presence of 200 mmol/L NaCI. These parameters were restored to original levels when the algae were transferred to a non-saline medium. Sodium and K＋ concentrations of stressed cells decreased significantly and H＋-ATPase activity increased after the addition of exogenous sucrose or EPS. The results suggest that EPS and sucrose are synthesized to maintain the cellular osmOtic equilibrium between the intra-and extracellular environment, thus protecting algal cells from osmotic damage, which was attributed to the selective exclusion of cellular Na＋ and K＋ by H＋-ATPase.     

Relationship of Plasmalemma Redox Activity to K+ Uptake by Dunaliella salina Cells

The plasmalemma-bond redox system localized within the plasmalemma of unicellular green alga Dunaliella salina was studied. This system oxidized exogenous NADH, increased O2 consumption to 165 % and increased the pH of the external medium, while K+ influx was inhibited. With no NADH added, ferricyanide stimulated K+ uptake about 3 folds. In the presence of exogenous NADH, ferricyanide was rapidly reduced and the external medium was acidified, generating a greater electrochemical proton gradient across the plasmalemma, thus resulting an 6-fold increase of K+ influx. Typical inhibitors of plasmalemma H+-ATPase and redox system inhibited K+ uptake to different extent. That the inhibition of K+ uptake by vanadate could be resumed partly by addition of NADH and ferricyanide indicated that plasmalemma redox system operated in association with the H+-ATPase to exert an influence on K+ transportation. A model was presented in which the implication of two possible redox chains and H+-ATPase in generating an electrochemical potential gradient for protons (△uH+) was discussed.     

Early stimulation of ATP turnover by EGF + insulin. Relation to external pH and Na+/H+ exchange system

We previously shown a rapid increase in ATP turnover after addition of epidermal growth factor and insulin to quiescent 3T3 cell cultures. Here, the relationship between this increase in ATP turnover and the activation by growth factors of Na+/H+ and Na+/K+ exchange systems was studied. Our results show that alkalinization of the medium enhances ATP turnover but they do not support the assumption that stimulation by growth factors of the Na+/H+ exchange induces an increase in ATP turnover since this increase was not inhibited by amiloride. Conversely, when ATP synthesis was abolished, the increase, in intracellular pH, by growth factors, was significantly decreased.     

配体对(Na~++K~+)-ATP酶E_2→E_1转变的影响

 本研究确定了在0℃条件下,(Na~++K~+)-ATP酶纯化制备物与5mmol/L Na~+或Mg~(2+)在5mmol/L咪唑(pH7.4)环境中预保温30分钟,然后进行磷酸化,可以获得最高磷酸化水平,Na~+或Mg~(2+)的K_(0.5)值分别为0.29mmol/L或0.35mmol/L;以ADP代替Na~+和Mg~(2+)与酶预保温,对E_2向E_1转变无任何影响,而与Na~+、Mg~(2+)一起存在时则能加强Na~+及Mg~2的预保温效果。     

Effect of Ca2+ and K+ on the intracellular pH of an Escherichia coli L-form.

The L-form NC7, derived from Escherichia coli K12, grew in a complex medium containing 0.2 M-CaCl2 as osmotic stabilizer, but not at pH values above 7.8. The cessation of growth at alkaline pH was not due to cell death. In complex media containing K+ or Na+, the L-form grew ove a wide pH range. Growth at alkaline pH was inhibited by 1 mM-amiloride, indicating that Na+/H+ antiport activity was required for growth at alkaline pH. The internal pH (pHi) of the L-form in media containing K+, Na+ or Ca2+ was constant at about 7.8 to 8.0 at external pH (pHo) values of 7.2 and 8.2. The rates of O2 consumption by intact cells, lactate oxidation by membrane vesicles from cells grown in Ca(2+)-containing medium, and cell division were all strongly repressed under alkaline conditions.     

Polarized distribution of the Na+/H+ exchange system in a renal cell line (LLC-PK1) with characteristics of proximal tubular cells

Studies of Na+ and H+ transport by confluent monolayers of the epithelial cell line LLC-PK1 were performed to verify the presence of a Na+/H+ exchange system. The presence of an outwardly directed H+ gradient produced a large stimulation of Na+ influx measured under net flux conditions. Amiloride (10(-3) M) completely inhibited Na+ influx stimulated by the H+ gradient and part of the Na+ influx measured in the absence of a pH gradient. Half-maximal inhibition of the Na+ influx stimulated by a pH gradient at 143 mM Na was observed at 5 microM amiloride. The presence of an inwardly oriented proton gradient also stimulated Na+ efflux from Na+-loaded cells. The stimulation was completely inhibited by the presence of 10(-3) M amiloride in the washout medium. These results indicate that this system could operate in the opposite direction depending on the orientation of the Na+ and H+ gradient. Incubation in Na+-free medium or in the presence of 10(-3) M ouabain resulted in a dramatic decrease of H+ release from LLC-PK1 cells. This H+ release was largely, although not completely, inhibited by 10(-4) M amiloride. Neither chloride substitution by the impermeable anion isethionate nor incubation in the presence of the ionophore valinomycin in high K+ medium affected Na+ influx by stimulated by a pH gradient. Inhibition of the Na+ influx by amiloride occurred only from the apical side of the monolayer. These results indicate that the Na+/H+ exchange system in LLC-PK1 monolayers is specifically localized in the apical membrane of the epithelial cells.     

Purification and properties of a histone acetyltransferase from Artemia salina, highly efficient with H1 histone.

An histone acetyltransferase has been purified from nuclei of 40-h-old Artemia salina larvae. The enzyme is very unstable at 0 degrees C, requires free -SH groups for activity and is rapidly inactivated at 40 degrees C. The optimal pH for activity is 8.5 and the activity is half inhibited by millimolar concentrations of Mn2+, Ca2+ or Mg2+ or decimolar concentrations of Na+ and K+. The molecular weight of the enzyme, determined by gel filtration chromatography, changed with the ionic strength of the medium (280,000 in 10 mM Tris . HCl, 170,000 in 0.2 M KCl). The very-lysine-rich histone H1 is a better substrate acceptor than the arginine-rich histones H3 or H4. Under proper conditions, the enzyme can modify all the internal lysyl residues in histones H1 and H4. The acetylation of H1 is inhibited when all the other histone fractions are present in the assay mixture.     

Effect of the activity of primary proton pumps on the growth of Escherichia coli in the presence of acetate

Escherichia coli batch cultures were grown under aerobic and anaerobic conditions on glucose with the substrate addition at pH 7.0. The cultures accumulated acetate in the medium at concentrations sufficient to inhibit the growth. This inhibitory effect of acetate was mediated apparently via its action on the intracellular pH. The inhibition of E. coli growth by acetate increased when the redox proton pump was switched off in the course of transition from aerobiosis to anaerobiosis and when the regulation of K+ fluxes was disordered in the presence of valinomycin. H+-ATPase was not essentially involved in maintaining the high rate of E. coli growth in the presence of acetate under aerobic conditions. If the activity of H+-ATPase was inhibited under anaerobic conditions at pH 7.0, the growth ceased after the dissipation of ionic gradients on the membrane. When CCCP was added under aerobic conditions, the growth did not stop at once if the medium had a pH of 7.6, but ceased immediately at pHout 7.0 in the glucose-salt medium.     

The sodium/proton antiport system in a newly isolated alkalophilic Bacillus sp.

The pH homeostasis and the sodium/proton antiport system have been studied in the newly isolated alkalophilic Bacillus sp. strain N-6, which could grow on media in a pH range from 7 to 10, and in its nonalkalophilic mutant. After a quick shift in external pH from 8 to 10 by the addition of Na2CO3, the delta pH (inside acid) in the cells of strain N-6 was immediately established, and the pH homeostatic state was maintained for more than 20 min in an alkaline environment. However, under the same conditions, the pH homeostasis was not observed in the cells of nonalkalophilic mutant, and the cytoplasmic pH immediately rose to pH 10. On the other hand, the results of the rapid acidification from pH 9 to 7 showed that the internal pH was maintained as more basic than the external pH in a neutral medium in both strains. The Na+/H+ antiport system has been characterized by either the effect of Na+ on delta pH formation or 22Na+ efflux in Na+-loaded right-side-out membrane vesicles of strain N-6. Na+- or Li+-loaded vesicles exhibited a reversed delta pH (inside acid) after the addition of electron donors (ascorbate plus tetramethyl-p-phenylenediamine) at both pH 7 and 9, whereas choline-loaded vesicles generated delta pHs of the conventional orientation (inside alkaline). 22Na+ was actively extruded from 22Na+-loaded vesicles whose potential was negative at pH 7 and 9. The inclusion of carbonyl cyanide m-chlorophenylhydrazone inhibited 22Na+ efflux in the presence of electron donors. These results indicate that the Na+/H+ antiport system in this strain operates electrogenically over a range of external pHs from 7 to 10 and plays a role in pH homeostasis at the alkaline pH range. The pH homeostasis at neutral ph was studied in more detail. K+ -depleted cells showed no delta pH (acid out) in the neutral conditions in the absence of K+, whereas these cells generated a delta pH if K+ was present in the medium. This increase of internal pH was accompanied by K+ uptake from the medium. These results suggest that electrogenic K+ entry allows extrusion of H+ from cells by the primary proton pump at neutral pH.