Interaction of phosphate with monovalent cation uptake in yeast

The uptake of monovalent cations by yeast via the monovalent cation uptake mechanism is inhibited by phosphate. The inhibition of Rb⁺ uptake shows saturation kinetics and the phosphate concentration at which halfmaximal inhibition is observed is equal to the K_m of phosphate for the sodiumindependent phosphate uptake mechanism. The kinetic coefficients of Rb⁺ and Tl⁺ uptake are affected by phosphate: the maximal rate of uptake is decreased and the apparent affinity constants for the translocation sites are increased.In the case of Na⁺ uptake, the inhibition by phosphate may be partly or completely compensated by stimulation of Na⁺ uptake via a sodium-phosphate cotransport mechanism.Phosphate effects a transient stimulation of the efflux of the lipophilic cation dibenzyldimenthylammonium from preloaded yeast cells and a transient inhibition of dibenzyldimethylammonium eptake. Possibly, the inhibition of monovalent cation uptake in yeast can be explained by a transient depolarization of the cell membrane by phosphate.     

Effect of ethidium bromide on Ca2+ uptake by yeast

Summary Ethidium bromide and other cationic dyes have been found to inhibit movalent cation uptake. This dye also produces in a K⁺-free medium an efflux of K⁺ which could be of the electrogenic type.The study of the effects of the same cationic dyes on Ca²⁺ uptake showed a large stimulation of the uptake rate of the divalent cation of more than tenfold.The analysis of the effects of one of the cationic dyes on Ca²⁺ uptake indicated that the efflux of K⁺ is of the electrogenic type and can drive the uptake of the divalent cation.Kinetic data on Ca²⁺ uptake indicate that, both under normal or under stimulated conditions, the divalent cation is taken up by the same transport system.The addition of ethidium bromide, besides, can stimulate the uptake of Mn²⁺ and¹⁴C-glycine and could be a good weapon to magnify and study some of the characteristics of ion transport systems in yeast.     

Studies on the mechanism of K+ transport in yeast.

The effect of uncouplers and diffusible acids on K⁺ transport was studied in yeast.Although the K⁺ transport system seems to depend on ATP to function, the effects of uncouplers are not due primarily to its action on the energy conserving systems of the cell.Other uncouplers with different structures to that of DNP showed also an inhibitory effect on K⁺ transport, which agrees with their reported ability to conduct protons through membranes.Uncouplers, besides inhibiting K⁺ uptake, produce an efflux of this cation; however, the rate of efflux produced is quantitatively important only when the cells have previously taken up the cation; there seems to exist a mechanism which prevents the loss of cations by yeast.In the absence of substrate, at pH 8.5, with 0.5 m KCl, TCS produces the efflux of H⁺, and when ⁸⁶Rb⁺ was used as a substitute for K⁺, an increase of the entrance of the cation could be detected in the presence of the uncoupler. It seems that the effect of the uncoupler depends on the direction of the combined H⁺ and K⁺ gradients, or the electrochemical potential of the cell.As reported by other authors, weak diffusible acids increase the uptake of K⁺ by yeast, and this effect is not due to changes in the metabolism, but to the magnitude of the entrance of the molecules to the yeast cell.It was found that the efflux of the acids (H₂CO₃), on the other hand, can produce an efflux of K⁺, which means that anions are important not only for the entrance of the cations, but for its permanence within the cell as well.The data seem to be in agreement with the hypothesis of the existence of a proton pump, responsible for the creation of an electrochemical potential, involved in K⁺ transport. At low pH, this pump seems to be activated by the transport of K⁺ into the cell.     

The role of intracellular pH in the regulation of cation exchanges in yeast

1. When yeast oxidizes propan-2-ol in the presence of KCl no uptake of K⁺ occurs. 2. When propionate is added to suspensions containing propan-2-ol, or if the suspensions are bubbled with CO₂, a considerable uptake of K⁺ occurs. 3. Maximum K⁺ uptake occurs at a propionate concentration of 2mm. 4. The addition of 20mm-propionate to the suspension lowers the intracellular pH of the yeast from a resting value in the region of 6.2 to approx. 5.6. 5. When K⁺ uptake is measured in the presence of 20mm-propionate, progressive changes in the rate of K⁺ uptake and intracellular pH occur. The optimum rate of K⁺ uptake occurs at an intracellular pH of 5.70. 6. The effect of both intra- and extra-cellular pH on K⁺–K⁺ exchange was studied and an optimum rate was found at an extracellular pH of 5.35, the corresponding intracellular pH being 6.44. 7. When a Na⁺-loaded yeast oxidizes propan-2-ol in the presence of KCl, a steady efflux of Na⁺ and influx of K⁺ occurs. The addition of 10mm-propionate to the suspension markedly inhibited the Na⁺ efflux but only slightly decreased the K⁺ influx. 8. The effect of both extra- and intra-cellular pH on Na⁺ efflux was studied with propan-2-ol and with glucose. The results can be best interpreted in terms of intracellular pH changes, and an optimum was obtained at approx. pH6.40.     

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% of the total fux. 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% 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 α-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.     

Relationship of Cation Influxes and Effluxes in Yeast

The Na⁺ efflux from Na⁺-rich yeast cells into a cation-free medium is largely balanced by the excretion of organic anions. In the presence of Rb⁺, K⁺, or high levels of H⁺ (pH 3–4), the Na⁺ efflux is increased and the organic anion excretion is suppressed so that stoichiometric cation exchanges occur. H⁺ participates in the exchanges, moving into or out of the cells depending on the external pH and on the concentration of external Rb⁺(K⁺). The total cation efflux is dependent on the external Rb⁺ concentration in a "saturation" relationship, but the individual cations in the efflux stream are not. The discrimination factor in the efflux pathway between H⁺ and Na⁺ is very large (of the order of 10,000), and between Na⁺ and K⁺ considerable (of the order of 50). For the latter pair, the recycling of K⁺ from the cell wall space is an important factor in the discrimination. In addition, the Na⁺ efflux as a function of Na⁺ content follows a sigmoidal curve so that the discrimination factor is increased at high levels of cellular Na⁺. Although the influx and efflux pathways behave as a tightly coupled system, the mechanism of coupling is not entirely clear. A single system with different cation specificities and kinetic behaviors on the inside and outside faces of the membrane could account for the data.     

Osmotic adaptation in halotolerant yeast, Debaryomyces nepalensis NCYC 3413: role of osmolytes and cation transport

Debaryomyces nepalensis NCYC 3413, a food spoiling yeast isolated from rotten apple, has been previously demonstrated as halotolerant yeast. In the present study, we assessed its growth, change in cell size, and measured the intracellular polyol and cations (Na⁺ or K⁺) accumulated during growth in the absence and presence of different concentrations of salts (NaCl and KCl). Cells could tolerate 2 M NaCl and KCl in defined medium. Scanning electron microscopic results showed linear decrease in mean cell diameter with increase in medium salinity. Cells accumulated high amounts of K⁺ during growth at high concentrations of KCl. However, it accumulated low amounts of Na⁺ and high amounts of K⁺ when grown in the presence of NaCl. Cells grown in the absence of salt showed rapid influx of Na⁺/K⁺ on incubation with high salt. On incubation with 2 M KCl, cells grown at 2 M NaCl showed an immediate efflux of Na⁺ and rapid uptake of K⁺ and vice versa. To withstand the salt stress, osmotic adjustment of intracellular cation was accompanied by intracellular accumulation of polyol (glycerol, arabitol, and sorbitol). Based on our result, we hypothesize that there exists a balanced efflux and synthesis of osmolytes when D. nepalensis was exposed to hypoosmotic and hyperosmotic stress conditions, respectively. Our findings suggest that D. nepalensis is an Na⁺ excluder yeast and it has an efficient transport system for sodium extrusion.     

Discrimination between Alkali Metal Cations by Yeast : II. Cation interactions in transport

K⁺ is a competitive inhibitor of the uptake of the other alkali metal cations by yeast. Rb⁺ is a competitive inhibitor of K⁺ uptake, but Li⁺, Na⁺, and Cs⁺ act like H⁺. At relatively low concentrations they behave as apparent noncompetitive inhibitors of K⁺ transport, but the inhibition is incomplete. At higher concentrations they inhibit the remaining K⁺ transport competitively. Ca⁺⁺ and Mg⁺⁺ in relatively low concentrations partially inhibit K⁺ transport in an apparently noncompetitive manner although their affinity for the transport site is very low. In each case, in concentrations that produce "noncompetitive" inhibition, very little of the inhibiting cation is transported into the cell. Competitive inhibition is accompanied by appreciable uptake of the inhibiting cation. The apparently noncompetitive effect of other cations is reversed by K⁺ concentrations much higher than those necessary to essentially "saturate" the transport system. A model is proposed which can account for the inhibition kinetics. This model is based on two cation-binding sites for which cations compete, a carrier or transporting site, and a second nontransporting (modifier) site with a different array of affinities for cations. The association of certain cations with the modifier site leads to a reduction in the turnover of the carrier, the degree of reduction depending on the cation bound to the modifier site and on the cation being transported.     

Alkali cation selectivity of the wheat root high-affinity potassium transporter HKT1

The wheat root high-affinity K⁺ transporter HKT1 functions as a sodium-coupled potassium co-uptake transporter. At toxic millimolar levels of sodium (Na⁺), HKT1 mediates low-affinity Na⁺ uptake while potassium (K⁺) uptake is blocked. In roots, low-affinity Na⁺ uptake and inhibition of K⁺ uptake contribute to Na⁺ toxicity. In the present study, the selectivity among alkali cations of HKT1 expressed in Xenopus oocytes and yeast was investigated under various ionic conditions at steady state. The data show that HKT1 is highly selective for uptake of the two physiologically significant alkali cations, K⁺ and Na⁺ over Rb⁺, Cs⁺ and Li⁺. In addition, Rb⁺ and Cs⁺, and an excess of extracellular K⁺ over Na⁺, are shown to partially reduce or block HKT1-mediated K⁺-Na⁺ uptake. Furthermore, K⁺, Rb⁺ and Cs⁺ also effectively reduce outward currents mediated by HKT1, thereby causing depolarizations. In yeast, HKT1 can produce high-affinity Rb⁺ uptake at approximately 15-fold lower rates than for K⁺. Rb⁺ influx in yeast can be mediated by the ability of the yeast plasma membrane proton pump to balance the 35-fold lower HKT1 conductance for Rb⁺. A model for HKT1 activity is presented involving a high-affinity K⁺ binding site and a high-affinity Na⁺ binding site, and competitive interactions of K⁺, Na⁺ and other alkali cations for binding to these two sites. Possible implications of the presented results for physiological K⁺ and Na⁺ uptake in plants are discussed.     

Relation between permeability to potassium and sodium ions and fusicoccin-stimulated hydrogen-ion efflux in barley roots

Summary Measurements are described of fusicoccin (FC)-stimulated H⁺ efflux in barley (Hordeum vulgare L.) roots when K⁺ and Na⁺ concentrations were varied. In low-salt roots H⁺ efflux was stimulated in both 5 mM KCl and NaCl. In salt-saturated roots H⁺ efflux was stimulated more effectively in KCl than in NaCl solution. The stimulation of H⁺ efflux thus is parallel with the selectivity of these different root preparations for K⁺ and Na⁺ and with estimates of permeability ratios (P _Na/P _K) determined from electrical measurements. It is suggested that the results support electrogenic coupling between FC-stimulated H⁺ efflux and cation uptake.     

Na+-dependent amino acid transport in preimplantation mouse embryos. II. Metabolic inhibitors and nature of the cation requirement.

Experiments were conducted in order to determine the energy source and nature of the cation dependency of [³H]methionine transport in preimplantation mouse embryos. The energy source of methionine transport was studied at the late four-cell and early blastocyst stages. The embryos, raised in vitro, were incubated for 1 hr in inhibitor(s) of energy metabolism and then transferred for 1 hr to medium that contained inhibitor(s) and ³H-methionine. These inhibitor studies suggest that respiration and glycolysis are needed to maintain uptake of methionine in early blastocysts. Late four-cell embryos seem to utilize respiration alone for transport.The cation dependency of methionine transport was studied at the late morula and early blastocyst stages. The kinetics of methionine uptake by early blastocysts in Na⁺-depleted media indicate a competitive type of inhibition. The uptake of methionine by early blastocysts is relatively resistant to ouabain and unaffected by K⁺-free medium. In contrast, methionine uptake by late morula-stage embryos is markedly inhibited by ouabain and K⁺-free medium in 1 hr. These results suggest that 1) Na⁺ serves to increase the affinity of methionine for the carrier in early blastocysts, 2) the cation gradients do not supply a major fraction of the energy required for methionine transport, and/or the gradients are difficult to perturb once the blastocyst has formed, and 3) putative Na⁺ pumps may be localized on the blastocoelic surface of the blastocysts.     

Activation of Rb+ and Na+ uptake into yeast by monovalent cations

⁸⁶Rb⁺ uptake by yeast was not only stimulated by Rb⁺ or K⁺ but also by Na⁺. The uptake of ²²Na⁺ was enhanced by both Rb⁺ and K⁺, but not by Na⁺, which was inhibitory at all concentrations applied. Inhibition of ²²Na⁺ uptake by inactive Na⁺ occurred in two phases: one phase refers to inhibition at low Na⁺ concentrations and the other to inhibition at high Na⁺ concentrations. Our results can be qualitatively described by a two-site transport mechanism, having two cation binding sites, which must be occupied with monovalent cations before transport can occur.     

Human Brain Capillary Endothelium: Modulation of K+ Efflux and K+, Ca2+ Uptake by Endothelin

This report describes K⁺ efflux, K⁺ and Ca²⁺ uptake responses to endothelins (ET-1 and ET-3) in cultured endothelium derived from capillaries of human brain (HBEC). ET-1 dose dependently increased K⁺ efflux, K⁺ and Ca²⁺ uptake in these cells. ET-1 stimulated K⁺ efflux occurred prior to that of K⁺ uptake. ET-3 was ineffective. The main contributor to the ET-1 induced K⁺ uptake was ouabain but not bumetanide-sensitive (Na⁺-K⁺-ATPase and Na⁺-K⁺-Cl^– cotransport activity, respectively). All tested paradigms of ET-1 effects in HBEC were inhibited by selective antagonist of ET_A but not ET_B receptors and inhibitors of phospholipase C and receptor-operated Ca²⁺ channels. Activation of protein kinase C (PKC) decreased whereas inhibition of PKC increased the ET-1 stimulated K⁺ efflux, K⁺ and Ca²⁺ uptake in HBEC. The results indicate that ET-1 affects the HBEC ionic transport systems through activation of ET_A receptors linked to PLC and modulated by intracellular Ca²⁺ mobilization and PKC.     

Alkali Cation/Sucrose Co-transport in the Root Sink of Sugar Beet

The mechanism of sucrose transport into the vacuole of root parenchyma cells of sugar beet was investigated using discs of intact tissue. Active sucrose uptake was evident only at the tonoplast. Sucrose caused a transient 8.3 millivolts depolarization of the membrane potential, suggesting an ion co-transport mechanism. Sucrose also stimulated net proton efflux. Active (net) uptake of sucrose was strongly affected by factors that influence the alkali cation and proton gradients across biological membranes. Alkali cations (Na⁺ and K⁺) at 95 millimolar activity stimulated active uptake of sucrose 2.1- to 4-fold, whereas membrane-permeating anions inhibited active sucrose uptake. The pH optima for uptake was between 6.5 and 7.0, pH values slightly higher than those of the vacuole. The ionophores valinomycin, gramicidin D, and carbonyl cyanide m-chlorophenylhydrazone at 10 micromolar concentrations strongly inhibited active sucrose uptake. These data are consistent with the hypothesis that an alkali cation influx/proton efflux reaction is coupled to the active uptake of sucrose into the vacuole of parenchyma cells in the root sink of sugar beets.     

The effects of alterations of transmembrane Na+ and K+ gradients by ionophores (nigericin,monensin) on serotonin transport in human blood platelets

Ionophores (monensin, nigericin) capable of transporting both Na⁺ and K⁺ across cell membranes down their concentration gradients reduce the rate and total magnitude of serotonin uptake by platelets. The effect of the ionophores was time dependent, so that inhibition increased progressively until eventually uptake ceased entirely. Nigericin and monensin produced loss of platelet K⁺ and an equivalent molar uptake of Na⁺ thereby abolishing the normal transmembrane Na⁺ and K⁺ gradients. The time course of these ionophore-induced cation shifts at 37° C corresponded to the rate at which inhibition of serotonin transport developed. The ionophores did not affect total ATP concentration of platelets nor the metabolic pool of ATP formed from [¹⁴C] adenine. Nigericin and monensin released about 80% of platelet ¹⁴C and endogenous serotonin over a 30 min period, without release of platelet adenine nucleotides, calcium or β-glucuronidase. The ionophores did not elicit platelet aggregation nor did they prevent maximal aggregation brought about by ADP, collagen or A23187. Replacement of Na⁺ in the medium by K⁺ abolished serotonin uptake but only 10–20% of endogenous serotonin was released. In KCl medium the Na⁺ gradient was initially reversed, but quickly dissipated as Na⁺ reequilibrated with the extracellular fluid. At 37° C the ionophores did not affect either the rate of Na⁺ reequilibration or the efflux of [¹⁴C] serotonin. Na⁺ reequilibration was slower at 20° C and the ionophores significantly increased platelet Na⁺ loss and strongly inhibited the efflux of [¹⁴C] serotonin. The data support a mechanism of serotonin transport due to a Na⁺-dependent carrier-mediated process which need not be directly dependent on metabolic energy, but which does require metabolic energy to maintain normal $\text{Na}{}^{\mathrm{+}}\text{+ K}{}^{\mathrm{+}}$ gradients.     

CHX14 is a plasma membrane K‐efflux transporter that regulates K+ redistribution in Arabidopsis thaliana

Potassium (K⁺) is essential for plant growth and development, yet the molecular identity of many K⁺ transporters remains elusive. Here we characterized cation/H⁺ exchanger (CHX) 14 as a plasma membrane K⁺ transporter. CHX14 expression was induced by elevated K⁺ and histochemical analysis of CHX14 promoter::GUS transgenic plants indicated that CHX14 was expressed in xylem parenchyma of root and shoot vascular tissues of seedlings. CHX14 knockout (chx14) and CHX14 overexpression seedlings displayed different growth phenotypes during K⁺ stress as compared with wild‐type seedlings. Roots of mutant seedlings displayed higher K⁺ uptake rates than wild‐type roots. CHX14 expression in yeast cells deficient in K⁺ uptake renders the mutant cells more sensitive to deficiencies of K⁺ in the medium. CHX14 mediates K⁺ efflux in yeast cells loaded with high K⁺. Uptake experiments using ⁸⁶Rb⁺ as a tracer for K⁺ with both yeast and plant mutants demonstrated that CHX14 expression in yeast and in planta mediated low‐affinity K⁺ efflux. Functional green fluorescent protein (GFP)‐tagged versions of CHX14 were localized to both the yeast and plant plasma membranes. Taken together, we suggest that CHX14 is a plasma membrane K⁺ efflux transporter involved in K⁺ homeostasis and K⁺ recirculation.     

Assessing the efficacy of vesicle fusion with planar membrane arrays using a mitochondrial porin as reporter

The gene for a putative cation calcium exchanger (CCX) from Arabidopsis thaliana, AtCCX5, was cloned and its function was analyzed in yeast. Green fluorescent protein-tagged AtCCX5 expressed in yeast was localized in the plasma membrane and nuclear periphery. The yeast transformants expressing AtCCX5 were created and their growth in the presence of various cations (K⁺, Na⁺, Ca²⁺, Mg²⁺, Fe²⁺, Cu²⁺, Co²⁺, Cd²⁺, Mn²⁺, Ba²⁺, Ni²⁺, Zn²⁺, and Li⁺) were analyzed. AtCCX5 expression was found to affect the response to K⁺ and Na⁺ in yeast. The AtCCX5 transformant also showed a little better growth to Zn²⁺. The yeast mutant 9.3 expressing AtCCX5 restored growth of the mutant on medium with low K⁺ (0.5 mM), and also suppressed its Na⁺ sensitivity. Ion uptake experiments showed that AtCCX5 mediated relatively high-affinity K⁺ uptake and was also involved in Na⁺ transport in yeast. Taken together, these findings suggest that the AtCCX5 is a novel transport protein involves in mediating high-affinity K⁺ uptake and Na⁺ transport in yeast.     

Effects of temperature and dinitrophenol on the uptake of potassium and sodium ions in Ricinus communis roots

Summary Detopped root systems of Ricinus communis plants were used for the study of the effects of temperature and DNP on the uptake of K and Na ions supplied as KNO₃ and NaNO₃.When K and Na ions were offered together in equivalent concentrations, the steady state uptake rates for K⁺ and Na⁺ at 23 to 25° gave a K⁺/Na⁺ ratio of 3. Increasing the Na⁺ concentration relative to K⁺ 3-fold did not alter the preferential uptake of K⁺. The uptake of K⁺ was more sensitive to temperature in the range 10 to 40° and to the application of DNP at 1.5x10^-4 M than was the uptake of Na⁺. When NaNO₃ was the only salt supplied Na⁺ uptake became more sensitive to DNP than when both K⁺ and Na⁺ nitrates were supplied. Prolonged application of DNP led to net K⁺ efflux from the roots, even when no K⁺ was being supplied to the roots. Net Na⁺ efflux under the influence of DNP occurred only in roots previously grown on Na-containing nutrient medium.The different responses of the K⁺ and Na⁺ uptake processes to temperature and DNP suggest the operation of different uptake mechanisms for K⁺ and Na⁺ These results have been considered in relation to the recent concept of dual mechanisms for the absorption of alkali cations by plant tissues.     

Effect of ouabain upon diuretic-sensitive K+ transport in cultured cells evidence for separate modes of operation of the transporter

(1) Unidirectional K⁺ (⁸⁶Rb) influx and efflux were measured in subconfluent layers of MDCK renal epithelial cells and HeLa carcinoma cells. (2) In both MDCK and HeLa cells, the furosemide-inhibitable and chloride-dependent component of K⁺ influx/efflux was stimulated 2-fold by a 30 min incubation in 1 · 10^?3 M ouabain. (3) Measurements of net K⁺ loss and Na⁺ gain in ouabain-treated cells at 1 h failed to show any diuretic sensitive component, confirming the exchange character of the diuretic-sensitive fluxes. (4) Prolonged incubations for 2.5 h in ouabain revealed a furosemide- and anion-dependent K⁺ (Cl^?) outward net flux uncoupled from net Na⁺ movement. Net K⁺ (Cl^?) outward flux was half-maximally inhibited by 2 μM furosemide. (5) After 2.5 h ouabain treatment, the anion and cation dependence of the diuretic-sensitive K⁺ influx/efflux were essentially unchanged when compared to untreated controls.     

Effect of sodium on potassium fluxes at the cell membrane and vacuole membrane of red beet

Slices of red beet (Beta vulgaris) washed for 5 to 6 days are known to accumulate Na⁺ in preference to K⁺ from solutions containing both ions. The present work, using ion concentrations of 1.0 mm or less, with Ca²⁺ added in some cases, shows that Na⁺ strongly inhibits K⁺ influx at the cell membrane (plasmalemma) while K⁺ efflux is increased to a lesser extent. This result from compartmental analysis is confirmed by short (15-minute) influx experiments, which indicate an immediate inhibitory effect of Na⁺ on K⁺ influx at the cell membrane. It is concluded that cation selectivity, even when Na⁺ is favored for uptake, is primarily determined at the cell membrane. Nevertheless, a high level of K⁺ in the cytoplasm is maintained during Na⁺ influx, by an inhibition of K⁺ transfer to the vacuole.