Further observations on the inhibitory effect of extracellular potassium ions on glycine uptake by mouse ascites-tumour cells

1. The initial rate of uptake of glycine by the tumour cells was measured as a function of the Na(+) and K(+) concentrations in the solution in which the cells were suspended. When [Gly] was 1mm or 12mm, the rate in the absence of Na(+) was independent of [K(+)] and about 3% or 10% respectively of the rate when [Na(+)] was 150m-equiv./l. 2. The Na(+)-dependent glycine entry rate, v, at a given value of [Na(+)] was successively lowered when [K(+)] was increased from 8 to 47 to 96m-equiv./l. A kinetic analysis indicated that K(+) competitively inhibited the action of Na(+). The results were in fair agreement with previous determinations of the kinetic parameters. 3. The presence of 2mm-sodium cyanide and 10mm-2-deoxyglucose lowered the cellular ATP content to less than 3% of the value in the respiring cells. Although v was then about 50% smaller, the relative effects of K(+) and Na(+) on the system were similar to those observed during respiration. 4. A theoretical analysis indicated that the variation of v with [K(+)] is not a reliable guide to the extent to which the K(+) gradient between the cells and their environment may contribute to the net transport of glycine.     

The effects of varying the cellular and extracellular concentrations of sodium and potassium ions on the uptake of glycine by mouse ascites-tumour cells in the presence and absence of sodium cyanide

1. Tumour cells were starved to deplete them of ATP and transferred to 0.9mm-glycine in Ringer solutions containing 2mm-sodium cyanide and various Na(+) and K(+) concentrations. The uptake of glycine then usually reached a peak by about 10min. 2. When cellular [Na(+)] and extracellular [Na(+)] were each about 30m-equiv./l., the maximum amount of glycine absorbed increased between 1.2- and 3.0-fold on lowering extracellular [K(+)] from 128 to 10m-equiv./l. 3. When extracellular [Na(+)] was 150m-equiv./l., the ratio, R, of the cellular to extracellular glycine concentrations increased progressively, from near 1 to about 9, when cellular [Na(+)] was lowered from 120 to 40m-equiv./l. 4. When cellular [Na(+)] was almost constant, either at 45 or 70m-equiv./l., R fell about 14-fold when extracellular [Na(+)] varied from 150 to 16m-equiv./l. 5. Values of R near 0.2 were found when cellular [Na(+)] was about four times as large as extracellular [Na(+)]. 6. R fell about threefold when the cells were put with 12mm- instead of 0.9mm-glycine. 7. The results were taken to imply that, under these conditions, the spontaneous movements of both Na(+) and K(+) across the cell membrane, down their respective concentration gradients, served to concentrate the glycine in the tumour cells (Christensen's hypothesis).     

Ionophore-mediated coupling between ion fluxes and amino acid absorption in mouse ascites-tumour cells. Restoration of the physiological gradients of methionine by valinomycin in the absence of adenosine triphosphate

1. Preparations of mouse ascites-tumour cells depleted of ATP and Na(+) ions accumulated l-methionine, in the presence of cyanide and deoxyglucose, from a 1mm solution containing 80mequiv. of Na(+)/l and about 5mequiv. of K(+)/l. Valinomycin increased, from about 4 to 16, the maximum value of the ratio of the cellular to extracellular concentrations of methionine formed under these conditions without markedly affecting the distributions of Na(+) and of K(+). Similar observations were made with 2-aminoisobutyrate, glycine and l-leucine. Increasing the extracellular concentration of K(+) progressively decreased the accumulation of methionine in the presence of valinomycin. Over the physiological range of ionic gradients, the system behaved as though the absorption of methionine with Na(+) was closely coupled to the electrogenic efflux of K(+) through the ionophore. The process was insensitive to ouabain and so the sodium pump was probably not involved. 2. The amount of methionine accumulated during energy metabolism was similar to the optimal accumulation in the presence of valinomycin when ATP was lacking. It was also similarly affected by increasing the methionine concentration. 3. A mixture of nigericin and tetrachlorosalicylanilide mimicked the action of valinomycin. The anilide derivative inhibited the absorption of 2-aminoisobutyrate in the presence of valinomycin, but not in its absence. 4. Gramicidin inhibited methionine absorption and caused the preparations to absorb Na(+) and lose K(+). 5. The observations appear to verify the principle underlying the gradient hypothesis by showing that the tumour cells can efficiently couple the electrochemical gradient of Na(+) to the amino acid gradient.     

Interactions between potassium ions and glycine transport in the yeast Saccharomyces carlsbergensis

A study has been made of the effects of both varying the pH and extracellular [K(+)] on the initial rate of uptake of glycine (v) by a strain of Saccharomyces carlsbergensis that concentrated the amino acid, with respect to the extracellular phase, by up to 1400 times. When no other substrate than glycine was provided and [glycine] was relatively small (相似文献   

A net gain of sodium ions and a net loss of potassium ions accompanying the uptake of glycine by mouse ascites-tumour cells in the presence of sodium cyanide

1. The tumour cells were starved in a solution lacking Na(+) and then transferred to a Ringer solution containing 2mm-sodium cyanide, 150m-equiv. of Na(+)/l. and 10m-equiv. of K(+)/l. Such cells were depleted of ATP and contained an endogenous pool of various amino acids equivalent to a 26mm solution. 2. At 4min. after the transfer the cellular Na(+) content had increased by about 100% and roughly an equivalent amount of K(+) had left the cells. 3. Under these conditions [(14)C]glycine was absorbed from an 11mm solution and reached the same cellular concentration by about 4min. The pool size increased by approximately the same amount (DeltaGly), so glycine did not simply exchange with the endogenous components. 4. After 4min. with glycine, the cells contained about 20% more Na(+) (DeltaNa(+)) than the control and about 10% less K(+) (DeltaK(+)). The mean values of DeltaNa(+)/DeltaGly and DeltaK(+)/DeltaGly from five experiments were respectively 0.90+/-0.11 and 0.62+/-0.11equiv./mole. 5. A further indication that these two ratios were not equal was that the cells absorbed more water than the movement of glycine itself required. The excess of water was osmotically equivalent to 0.95+/-0.16equiv. of solute/mole of glycine absorbed. 6. The variation of DeltaNa(+)/DeltaGly with the duration of the incubation was consistent with the stimulated uptake of Na(+) being linked to the actual transport of glycine. The same may apply to the movement of K(+), though the time-dependence was not examined in that case. 7. The observations were analysed in terms of a model in which both K(+) and Na(+) moved with a glycine-carrier system without ATP being involved. The analysis supported the idea that the spontaneous movements of the ions through the system might concentrate glycine in the cells significantly by purely physical means (Christensen's hypothesis).     

A sodium ion concentration gradient formed during the absorption of glycine by mouse ascites-tumour cells

1. To deplete them of ATP the tumour cells were starved at 37 degrees in a Ringer solution containing 33m-equiv. of Na(+)/l., 131m-equiv. of Li(+)/l., 2mM-sodium cyanide and 0.1mm-ouabain. The cellular content of K(+) was largely replaced by Li(+), but cellular [Na(+)] remained near 33m-equiv./l. 2. The addition of 12mm-glycine to the system caused cellular [Na(+)] to increase, during the next 4min., by about 4m-equiv./l., so that it slightly exceeded extracellular [Na(+)]. This occurred in parallel with the absorption of glycine. 3. The cellular K(+) content fell by an amount representing about 10% of the amount of Na(+) absorbed. 4. The results provide a clear demonstration that the flow of glycine into the cells is linked to a parallel movement of Na(+); K(+) appears to play a facultative role in the carrier system, whereas Li(+) is almost inert. 5. The effects produced by glycine were not reproduced by l-arabinose.     

Cation specificity of osmosensing by the betaine carrier BetP of Corynebacterium glutamicum

The Na(+)/betaine carrier BetP from Corynebacterium glutamicum was purified and reconstituted in Escherichia coli phospholipid liposomes and its osmosensory properties were studied with respect to the cation specificity of osmotic activation. To dissect the influence of the co-substrate Na(+) on the energetics of uptake from its possible role as a putative trigger of osmolality-dependent BetP activation, the internal Na(+) concentration was varied without changing DeltapNa(+). Studying betaine uptake at increasing luminal Na(+) or K(+) revealed that BetP activity was triggered by Na(+) only to a negligible extent compared to activation by K(+). We conclude that activation of BetP in proteoliposomes depends solely on K(+), both in mechanistic and in physiological terms.     

Sodium and Potassium Requirements for Active Transport of Glutamate by Escherichia coli K-12

Active transport of glutamate by Escherichia coli K-12 requires both Na(+) and K(+) ions. Increasing the concentration of Na(+) in the medium results in a decrease in the K(m) of the uptake system for glutamate; the capacity is not affected. Glutamate uptake by untreated cells is not stimulated by K(+). K(+)-depleted cells show a greatly reduced capacity for glutamate uptake. Preincubation of such cells in the presence of K(+) fully restores their capacity for glutamate uptake when Na(+) ions are also present in the uptake medium. Addition of either K(+) or Na(+) alone restores glutamate uptake to only about 20% of its maximum capacity in the presence of both cations. Changes in K(+) concentration affect the capacity for glutamate uptake but have no effect on the K(m) of the glutamate transport system. Ouabain does not inhibit the (Na(+)-K(+))-stimulated glutamate uptake by intact cells or spheroplasts of E. coli K-12.     

GerN, an endospore germination protein of Bacillus cereus, is an Na(+)/H(+)-K(+) antiporter

GerN, a Bacillus cereus spore germination protein, exhibits homology to a widely distributed group of putative cation transporters or channel proteins. GerN complemented the Na(+)-sensitive phenotype of an Escherichia coli mutant that is deficient in Na(+)/H(+) antiport activity (strain KNabc). GerN also reduced the concentration of K(+) required to support growth of an E. coli mutant deficient in K(+) uptake (strain TK2420). In a fluorescence-based assay of everted E. coli KNabc membrane vesicles, GerN exhibited robust Na(+)/H(+) antiport activity, with a K(m) for Na(+) estimated at 1.5 mM at pH 8.0 and 25 mM at pH 7.0. Li(+), but not K(+), served as a substrate. GerN-mediated Na(+)/H(+) antiport was further demonstrated in everted vesicles as energy-dependent accumulation of (22)Na(+). GerN also used K(+) as a coupling ion without completely replacing H(+), as indicated by partial inhibition by K(+) of H(+) uptake into right-side-out vesicles loaded with Na(+). K(+) translocation as part of the antiport was supported by the stimulatory effect of intravesicular K(+) on (22)Na(+) uptake by everted vesicles and the dependence of GerN-mediated (86)Rb(+) efflux on the presence of Na(+) in trans. The inhibitory patterns of protonophore and thiocyanate were most consistent with an electrogenic Na(+)/H(+)-K(+) antiport. GerN-mediated Na(+)/H(+)-K(+) antiport was much more rapid than GerN-mediated Na(+)/H(+) antiport.     

The influence of external sodium ions on the sodium pump in erythrocytes

1. A study has been made of the interaction between Na(+) and K(+) on the adenosine triphosphatase activity of erythrocyte ;ghosts', and on the K(+) influx and Na(+) efflux of intact erythrocytes. The adenosine triphosphatase activity and the ion movements were greater at a low external K(+) concentration in the absence of Na(+) than they were in the presence of 150mm-Na(+). The inhibition by external Na(+) of K(+) influx had an inhibitory constant of 5-10mm. 2. Activation by K(+) of kidney microsomal adenosine triphosphatase was retarded by Na(+), and activation by Na(+) was retarded by K(+). Fragmented erythrocyte membranes behaved similarly. 3. These observations suggest that there is competition between Na(+) and K(+) at the K(+)-sensitive site of the membrane.     

The effects of potassium ions and denervation on protein synthesis and the transport of amino acids in muscle

1. The effects of varying concentrations of K(+) during incubation, of denervation and of various drugs on the accumulation of (14)C-labelled amino acids, their incorporation into protein and the stimulation of these processes by insulin in rat diaphragm preparations were studied. 2. The accumulation of glycine and aminoisobutyrate and incorporation of glycine into protein was less in tissue incubated in K(+)-free buffer or 20mm-K(+) than with 5-10mm-K(+). Incorporation of leucine was unaffected. 3. Incorporation into protein of amino acids by diaphragm that had been denervated 3 days previously was elevated. Accumulation of both glycine and aminoisobutyrate was also raised but that of phenylalanine was unaffected. 4. Accumulation of glycine by diaphragm and extensor digitorum longus muscle was decreased by a number of agents including cocaine and mepyramine. 5. The stimulation of incorporation by insulin was unaffected by changes in K(+) or in the presence of cocaine and mepyramine. Denervated tissue was markedly less responsive to insulin than its control. 6. The results are discussed in the context of the relation of amino acid accumulation to operation of the Na(+) pump and the influence of insulin thereon.     

The effect of electrical stimulation and ouabain on the uptake and efflux of l-[U-14C]valine in chopped tissue from guinea-pig cerebral cortex

1. Chopped tissue from guinea-pig cerebral cortex carried out an energy-dependent accumulation of l-[(14)C]valine. 2. The uptake was dependent on the extracellular concentration of Na(+) and was markedly inhibited by ouabain (20mum). The extent of the inhibition of uptake by ouabain was also Na(+)-dependent. 3. The accumulation of labelled valine was not directly dependent on the ATP and creatine phosphate contents of the slices. 4. Electrical stimulation increased the rate of [(14)C]valine uptake at first but ultimately led to a net loss of the label so that the amount of label present in the tissue was lower than in the controls. 5. The rate of loss of label during prolonged stimulation was dependent on the extracellular concentration of Na(+). 6. The efflux of labelled valine from slices preloaded at 164mm-Na(+) was studied at 164, 80 and 40mm-Na(+) with and without electrical stimulation or ouabain. 7. Lowering the Na(+) concentration or adding ouabain increased the rate of efflux. 8. Electrical stimulation had little effect on the rate of efflux at first but ultimately led to a more complete loss of label from the tissue than occurred in the control. A kinetic analysis of the efflux curves was attempted.     

The Wheat cDNA LCT1 Generates Hypersensitivity to Sodium in a Salt-Sensitive Yeast Strain

Salinity affects large areas of agricultural land, and all major crop species are intolerant to high levels of sodium ions. The principal route for Na(+) uptake into plant cells remains to be identified. Non-selective ion channels and high-affinity potassium transporters have emerged as potential pathways for Na(+) entry. A third candidate for Na(+) transport into plant cells is a low-affinity cation transporter represented by the wheat protein LCT1, which is known to be permeable for a wide range of cations when expressed in yeast (Saccharomyces cerevisiae). To investigate the role of LCT1 in salt tolerance we have used the yeast strain G19, which is disrupted in the genes encoding Na(+) export pumps and as a result displays salt sensitivity comparable with wheat. After transformation with LCT1, G19 cells became hypersensitive to NaCl. We show that LCT1 expression results in a strong decrease of intracellular K(+)/Na(+) ratio in G19 cells due to the combined effect of enhanced Na(+) accumulation and loss of intracellular K(+). Na(+) uptake through LCT1 was inhibited by K(+) and Ca(2+) at high concentrations and the addition of these ions rescued growth of LCT1-transformed G19 on saline medium. LCT1 was also shown to mediate the uptake of Li(+) and Cs(+). Expression of two mutant LCT1 cDNAs with N-terminal truncations resulted in decreased Ca(2+) uptake and increased Na(+) tolerance compared with expression of the full-length LCT1. Our findings strongly suggest that LCT1 represents a molecular link between Ca(2+) and Na(+) uptake into plant cells.     

Extrusion of Sodium and Hydrogen Ions as the Primary Process in Potassium Ion Accumulation by Streptococcus faecalis

Glycolyzing cells of Streptococcus faecalis accumulate K(+) with concurrent extrusion of equivalent amounts of H(+) and Na(+). An attempt was made to clarify the retionship between the movements of Na(+) and K(+). Sodium was displaced from cells glycolyzing in the presence of ammonia, diethylamine, tris(hydroxymethyl)aminomethane, and other nitrogenous cations; by contrast, K(+) was completely retained. Accumulation of K(+) by heterologous exchange for Na(+) was not inhibited by antibiotics which facilitate diffusion of K(+) across the membrane, but was blocked by proton conductors. The results indicate that extrusion of Na(+) and H(+) from the cells is a primary, energy-linked process which generates an electrical potential (interior negative); K(+) accumulation occurs in response to this potential. Two mutants deficient in K(+) accumulation and retention were examined in terms of this model. One mutant is apparently defective in exchange of K(+) for H(+). In the other mutant, exchange of K(+) for Na(+) is impaired.     

阳离子对豚鼠Ⅱ型前庭毛细胞ACh-敏感性电流的调制

本文旨在探讨哺乳动物前庭胆碱能传出神经系统的作用机制,应用全细胞膜片钳技术研究新鲜分离的豚鼠Ⅱ型前庭毛细胞ACh-敏感性电流的特性以及细胞内外的阳离子对ACh-敏感性电流的调制作用。结果显示,Ⅱ型前庭毛细胞对细胞外ACh敏感,ACh激活缓慢持久的外向性电流,室温下此电流的再次完全激活时间约为(60±10)s。ACh-敏感性电流的反转电位为(-66±8)mV,提示此电流主要由K+参与形成,其直接作用是使毛细胞超极化。ACh-敏感性电流对较高浓度的四乙胺(tetraethylammonium chloride,TEA)敏感,提示细胞外ACh激活钙依赖性钾电流。进一步检测细胞内外阳离子对 ACh-敏感性电流的调制作用发现,细胞外Na+和细胞内Ca2+释放不参与此电流的激活过程,而细胞外K+、细胞外Ca2+和细胞内Mg2+对ACh-敏感性电流具有重要的调制作用。进一步提示,ACh是哺乳动物前庭传出神经系统重要的神经递质。 Na+不参与ACh-敏感性电流的激活过程提示,ACh-敏感性电流可能由非α9-N型胆碱能受体(α9-nAChR)介导。ACh诱导的Ⅱ型前庭毛细胞超极化作用受细胞外Ca2+浓度和细胞内Mg2+浓度调制。     

Amino acid absorption by mouse ascites-tumour cells depleted of both endogenous amino acids and adenosine triphosphate

1. Despite the depletion of both their content of exchangeable endogenous amino acids and reserves of ATP, starved hypo-osmotically shocked preparations of the tumour cells accumulated relatively large amounts of (14)C-labelled 2-aminoisobutyrate, l-alanine, glycine, l-leucine, l-methionine, l-phenylalanine and l-serine, against their respective concentration gradients, by a process apparently driven by the spontaneous flow of Na(+) ions into the cellular phase. Dependent on (a) which compound was used, (b) its concentration and (c) the direction of the Na(+) ion gradient, the peak value of the ratio of the cellular to extracellular amino acid concentration varied from about 0.4 to 7. 2. The extent to which ATP increased the ratio was defined for l-methionine. 3. Chemical analysis of the cellular amino acid content showed that this increased in parallel with the absorption of (14)C. 4. The accumulation of l-methionine and of glycine, against their own concentration gradients, continued in the presence of either 0.3mm-ouabain or 10mug of oligomycin/ml. Thus the sodium pump was probably not involved in the process when ATP was lacking. 5. l-Leucine caused 0.72+/-0.12 (s.e.m.; 6) extra equivalents of Na(+) to enter the shocked starved tumour cells in parallel with the uptake of leucine itself. Only a small loss of K(+) was induced. 6. The influx and efflux of l-methionine in preparations depleted of ATP were both markedly accelerated by the presence of Na(+) ions. 7. The observations provide further examples of the application of the ion-gradient hypothesis, according to which Na(+) ions act as co-substrates of the amino acid pump. The quantitative importance of parallel Na(+)-independent systems was studied with a new mathematical model.     

Dynamics of forward and reverse transport by the glial glycine transporter, glyt1b

Glycine is a coagonist at the N-methyl-D-aspartate receptor. Changes in extracellular glycine concentration may modulate N-methyl-D-aspartate receptor function and excitatory synaptic transmission. The GLYT1 glycine transporter is present in glia surrounding excitatory synapses, and plays a key role in regulating extracellular glycine concentration. We investigated the kinetic and other biophysical properties of GLYT1b, stably expressed in CHO cells, using whole-cell patch-clamp techniques. Application of glycine produced an inward current, which decayed within a few seconds to a steady-state level. When glycine was removed, a transient outward current was observed, consistent with reverse transport of accumulated glycine. The outward current was enhanced by elevating intracellular or lowering extracellular [Na(+)], and was modulated by changes in extracellular [glycine] and time of glycine application. We developed a model of GLYT1b function, which accurately describes the time course of the transporter current under a range of experimental conditions. The model predicts that glial uptake of glycine will decay toward zero during a sustained period of elevated glycine concentration. This property of GLYT1b may permit spillover from glycinergic terminals to nearby excitatory terminals during a prolonged burst of inhibitory activity, and reverse transport may extend the period of elevated glycine concentration beyond the end of the inhibitory burst.     

Cloning, functional expression and primary characterization of Vibrio parahaemolyticus K+/H+ antiporter genes in Escherichia coli

The regulation of internal Na(+) and K(+) concentrations is important for bacterial cells, which, in the absence of Na(+) extrusion systems, cannot grow in the presence of high external Na(+). Likewise, bacteria require K(+) uptake systems when the external K(+) concentration becomes too low to support growth. At present, we have little knowledge of K(+) toxicity and bacterial outward-directed K(+) transport systems. We report here that high external concentrations of K(+) at alkaline pH are toxic and that bacteria require K(+) efflux and/or extrusion systems to avoid excessive K(+) accumulation. We have identified the first example of a bacterial K(+)(specific)/H(+) antiporter, Vp-NhaP2, from Vibrio parahaemolyticus. This protein, a member of the cation : proton antiporter-1 (CPA1) family, was able to mediate K(+) extrusion from the cell to provide tolerance to high concentrations of external KCl at alkaline pH. We also report the discovery of two V. parahaemolyticus Na(+)/H(+) antiporters, Vp-NhaA and Vp-NhaB, which also exhibit a novel ion specificity toward K(+), implying that they work as Na(+)(K(+))/H(+) exchangers. Furthermore, under specific conditions, Escherichia coli was able to mediate K(+) extrusion against a K(+) chemical gradient, indicating that E. coli also possesses an unidentified K(+) extrusion system(s).     

Presence of a Na+-stimulated P-type ATPase in the plasma membrane of the alkaliphilic halotolerant cyanobacterium Aphanothece halophytica

Aphanothece cells could take up Na(+) and this uptake was strongly inhibited by the protonophore, carbonyl cyanide m-chlorophenylhydrazone (CCCP). Cells preloaded with Na(+) exhibited Na(+) extrusion ability upon energizing with glucose. Na(+) was also taken up by the plasma membranes supplied with ATP and the uptake was abolished by gramicidin D, monensin or Na(+)-ionophore. Orthovanadate and CCCP strongly inhibited Na(+) uptake, whereas N, N'-dicyclohexylcarbodiimide (DCCD) slightly inhibited the uptake. Plasma membranes could hydrolyse ATP in the presence of Na(+) but not with K(+), Ca(2+) and Li(+). The K(m) values for ATP and Na(+) were 1.66+/-0.12 and 25.0+/-1.8 mM, respectively, whereas the V(max) value was 0.66+/-0.05 mumol min(-1) mg(-1). Mg(2+) was required for ATPase activity whose optimal pH was 7.5. The ATPase was insensitive to N-ethylmaleimide, nitrate, thiocyanate, azide and ouabain, but was substantially inhibited by orthovanadate and DCCD. Amiloride, a Na(+)/H(+) antiporter inhibitor, and CCCP showed little or no effect. Gramicidin D and monensin stimulated ATPase activity. All these results suggest the existence of a P-type Na(+)-stimulated ATPase in Aphanothece halophytica. Plasma membranes from cells grown under salt stress condition showed higher ATPase activity than those from cells grown under nonstress condition.     

Potassium Transport and the Relationship Between Intracellular Potassium Concentration and Amino Acid Uptake by Cells of a Marine Pseudomonad

Transport of K(+) by K(+)-depleted cells of marine pseudomonad B-16 (ATCC 19855) exhibited saturation kinetics. Rb(+) inhibited both K(+) transport and the K(+)-dependent transport of alpha-aminoisobutyric acid (AIB) into K(+)-depleted cells of the organism in proportion to the concentration of Rb(+) in the suspending medium. Inhibition of the K(+)-dependent uptake of AIB into K(+)-depleted cells by Rb(+) could be overcome by increasing the concentration of K(+) in the medium. When AIB and K(+) were added simultaneously to a suspension of K(+)-depleted cells, the uptake of K(+) occurred immediately and rapidly, whereas the accumulation of AIB occurred only after a lag. The initial uptake rate of AIB was directly proportional to the intracellular K(+) concentration. The intracellular concentration of K(+) and AIB at their steady-state levels increased to a maximum as the Na(+) concentration in the suspending medium was increased. At Na(+) concentrations between 0.2 and 0.3 M, the molar ratio of K(+) to AIB at their intracellular steady-state concentrations was constant at 1.6. At external Na(+) concentrations less than 0.2 M, the cells maintained a relatively higher K(+) intracellular steady-state level than AIB.