Light Stimulation of Active Transport in Hydrodictyon africanum

The mechanism of light stimulation of active K and Cl influx and active Na efflux, in Hydrodictyon africanum has been investigated using different wavelengths of red light and different gas mixtures, and the inhibitors DCMU and CCCP. The active Cl influx requires photosystem 2, since its relative quantal efficiency falls with increasing wavelength of red light, and it is as sensitive to the inhibitor DCMU as is photosynthesis; it is relatively insensitive to the uncoupler CCCP. The active K influx and active Na efflux are inhibited by CCCP, but the relative quantal efficiency of these processes increases with increasing wavelength of red light, and they are relatively insensitive to DCMU. These cation fluxes can be supported by cyclic photophosphorylation, whereas Cl influx needs photosystem 2 but probably not ATP.     

The Linkage of Light-stimulated Cl Influx to K and Na Influxes in Hydrodictyon africanum

There are reciprocal stimulations of Cl influx by K and Na,and of K and Na influx by Cl, in the light in Hydrodictyon africanum.The component of the K influx which stimulates, and is stimulatedby, Cl, is independent of the ouabainsensitive mechanism forK influx also found in H. africanum. The concentration dependenceof the cation effects on Cl influx and on the Cl-stimulatedportion of their own influxes are similar. The stimulation withK saturates at about 0.3 mM K; that with Na saturates at about2 mM Na. The Cl-dependent portions of the K and Na influxeshave similar responses to changes in photo-synthetic metabolism(far-red illumination, CDMU, and CCCP) as does the light-stimulatedCl influx. This suggests that Cl influx, and the Cl-stimulatedportions of K and Na influxes are both dependent on photosystem2 of photosynthesis, and are less sensitive to the uncouplerCCCP than is ¹⁴CO₂ fixation or the K-Na pump. It would thusappear that the Cl-dependent portions of the K and Na influxesin the light are linked to the cation-stimulated portion ofthe Cl influx. There is no very great change in the electricalcomponent of the inwardly directed passive driving force oncations under conditions in which Cl is being pumped comparedwith those under which it is not. It is not clear whether suchincrease in this driving force as do occur could account quantitativelyfor the increase in the cation influxes associated with Cl transport,or whether chemical coupling must be invoked. In addition tothe Cl-stimulated portions of the cation influxes, there arealso light-stimulated portions of K and Na influx which areindependent of Cl, not associated with the cation regulatingmechanism, and which seem to have a similar linkage to photosynthesisas does the Cl-K-Na pump. Since the light-stimulated portionof the K efflux appears to be similar to this portion of theK influx, these Cl-independent light-stimulated portions ofK and Na influxes are tentatively related to light-induced changesin cation permeability.     

Furosemide-sensitive Na and K fluxes in human red cells. Net uphill Na extrusion and equilibrium properties

This paper reports experiments designed to find the concentrations of internal and external Na and K at which inward and outward furosemide-sensitive (FS) Na and K fluxes are equal, so that there is no net FS movement of Na and K. The red cell cation content was modified by using the ionophore nystatin, varying cell Na (Nai) from 0 to 34 mM (K substitution, high-K cells) and cell K (Ki) from 0 to 30 mM (Na substitution, high-Na cells). All incubation media contained NaCl (Nao = 130 or 120 nM), and KCl (Ko = 0-30 mM). In high-K cells, incubated in the absence of Ko, there was net extrusion of Na through the FS pathway. The net FS Na extrusion increased when Nai was increased. Low concentrations of Ko (0-6 mM) slightly stimulated, whereas higher concentrations of Ko inhibited, FS Na efflux. Increasing Ko stimulated the FS Na influx (K0.5 = 4 mM). Under conditions similar to those that occur in vivo (Nai = 10, Ki = 130, Nao = 130, Ko = 4 mM, Cli/Clo = 0.7), net extrusion of Na occurs through the FS pathway (180-250 mumol/liter cell X h). The concentration of Ko at which the FS Na influx and efflux and the FS K influx and efflux become equal increased when Nai increased in high-K cells and when Ki was increased in high-Na cells. The net FS Na and K fluxes both approached zero at similar internal and external Na and K concentrations. In high-K cells, under conditions when net Na and K fluxes were near zero, the ratio of FS Na to FS K unidirectional flux was found to be 2:3. In high-K cells, the empirical expression (Nai/Nao)2(Ki/Ko)3 remained at constant value (apparent equilibrium constant, Kappeq +/- SEM = 22 +/- 2) for each set of internal and external cation concentrations at which there was no net Na flux. These results indicate that in the physiological region of concentrations of internal and external Na, K, and Cl, the stoichiometry of the FS Na and K fluxes is 2 Na:3 K. In high-Na cells under conditions when net FS Na and K fluxes were near zero, the ratio of FS Na to FS K unidirectional fluxes was 3:2 (1).(ABSTRACT TRUNCATED AT 400 WORDS)     

Coupling of a loop diuretic-sensitive Na+ influx with the net loop diuretic-sensitive K+ efflux in mouse NIH 3T3 cells

Mouse 3T3 fibroblasts have a loop diuretic sensitive Na+ transport system, responsible for more than 50% of the total Na+ influx. This transport system is dependent on the simultaneous presence of all three ions; Na+, K+, (Rb+) and Cl- in the extracellular medium. The same requirement for these three ions was also found for the loop diuretic-sensitive K+ efflux. In addition, the sensitivities of Na+ influx and Rb+ efflux for the two loop diuretics, furosemide and bumetanide were found to be similar. The similar ionic requirement and sensitivity towards loop diuretics of the two fluxes, support the hypothesis, that this loop diuretic-sensitive Na+ influx in mouse 3T3 cells, is accompanied by the net loop diuretic-sensitive K+ efflux.     

Responses of frog skin Na(+) and Cl(-) transport to guanylin

A possible role for the peptide hormone guanylin was investigated in frog skin (Rana pipiens) epithelium. Sodium and chloride fluxes in response to this peptide were evaluated in Ussing-type chambers. Net and unidirectional Na(+) fluxes were measured by using (22)Na(+) and atomic absorption analysis of total [Na(+)], whereas net Cl(-) fluxes were measured by using electrometric titration for [Cl(-)]. Mucosal application of guanylin (0.5-2.0 micromol/l) caused marked increases in serosal to mucosal net flux and efflux of Na(+). Serosal application of guanylin over the same dose range caused similar large increases in net serosal to mucosal (S-->M) Na(+) and Cl(-) flux as well as Na(+) efflux. Responses of Na(+) influx were small and inconsistent. When frog skin was bathed on the serosal side with Cl(-)-free Ringer's solution mucosal application of guanylin stimulated large efflux and S-->M net fluxes of Na(+). Serosal treatment yielded large Na(+) effluxes and S-->M Na(+) and Cl(-) net fluxes. When frog skin serosal surfaces were bathed with Na(+)- free Ringer's solution mucosal guanylin treatment had no effect but serosal treatment produced large S-->M Cl(-) net fluxes.     

Regulatory interaction of ATP Na+ and Cl- in the turnover cycle of the NaK2Cl cotransporter

To probe the mechanism by which intracellular ATP, Na+, and Cl- influence the activity of the NaK2Cl cotransporter, we measured bumetanide-sensitive (BS) 86Rb fluxes in the osteosarcoma cell line UMR- 106-01. Under physiological gradients of Na+, K+, and Cl-, depleting cellular ATP by incubation with deoxyglucose and antimycin A (DOG/AA) for 20 min at 37 degrees C reduced BS 86Rb uptake from 6 to 1 nmol/mg protein per min. Similar incubation with 0.5 mM ouabain to inhibit the Na+ pump had no effect on the uptake, excluding the possibility that DOG/AA inhibited the uptake by modifying the cellular Na+ and K+ gradients. Loading the cells with Na+ and depleting them of K+ by a 2-3- h incubation with ouabain or DOG/AA increased the rate of BS 86Rb uptake to approximately 12 nmol/mg protein per min. The unidirectional BS 86Rb influx into control cells was approximately 10 times faster than the unidirectional BS 86Rb efflux. On the other hand, at steady state the unidirectional BS 86Rb influx and efflux in ouabain-treated cells were similar, suggesting that most of the BS 86Rb uptake into the ouabain-treated cells is due to K+/K+ exchange. The entire BS 86Rb uptake into ouabain-treated cells was insensitive to depletion of cellular ATP. However, the influx could be converted to ATP-sensitive influx by reducing cellular Cl- and/or Na+ in ouabain-treated cells to impose conditions for net uptake of the ions. The BS 86Rb uptake in ouabain-treated cells required the presence of Na+, K+, and Cl- in the extracellular medium. Thus, loading the cells with Na+ induced rapid 86Rb (K+) influx and efflux which, unlike net uptake, were insensitive to cellular ATP. Therefore, we suggest that ATP regulates a step in the turnover cycle of the cotransporter that is required for net but not K+/K+ exchange fluxes. Depleting control cells of Cl- increased BS 86Rb uptake from medium-containing physiological Na+ and K+ concentrations from 6 to approximately 15 nmol/mg protein per min. The uptake was blocked by depletion of cellular ATP with DOG/AA and required the presence of all three ions in the external medium. Thus, intracellular Cl- appears to influence net uptake by the cotransporter. Depletion of intracellular Na+ was as effective as depletion of Cl- in stimulating BS 86Rb uptake.(ABSTRACT TRUNCATED AT 400 WORDS)     

The Control of Na Uptake into Nitella translucens

Movement of Na into cells of Nitella translucens is a ‘downhill’process; the ions move across the plasmalemma down an electrochemicalpotential gradient. Nevertheless, measurements of Na influxesunder a wide range of experimental conditions have shown thatthere must be links between Na uptake and processes controlledby metabolism. When Ca ions are present in the bathing solution,Na influxes are greatly increased by light under conditionswhere photosynthesis can proceed (i.e. when both photosystemsare active). In the presence of Ca, the influx of Na increasesonly slightly when the external Na concentration is raised above1 mM, and the light-promoted Na influx is considerably inhibitedwhen Cl is removed from the bathing solution. When the Cl concentrationis kept constant, the Na influx in light is determined by theconcentrations of other cations present in solution (K, Ca,or NH₄). In the absence of Ca from the cell wall and solution,the influx is stil enhanced by light, but does not saturatewhen the external Na concentration is raised above 1 mM. Itis suggested that the Na influx in light is partly linked tothe inward Cl pump, but there is also a separate (Cl-independent)effect of light on the permeability of the plasmalemma to Na.Links between Na and Cl uptake could be maintained by effectsof Cl on electrochemical driving forces controlling Na entry;alternatively, chemical coupling between the two processes maybe involved.     

The Effect of Dio-9 on Photosynthesis and Ion Transport in Nitella, Tolypella, and Hydrodictyon

The effect of Dio-9, on photosynthesis and active and passiveion transport at the plasmalemma has been investigated in Nitellatranslucens, Tolypella intricata, and Hydrodictyon africanum.The active K influx and the coupled active Na efflux were moreinhibited by this inhibitor of energy transfer in phosphorylationthan was photosynthesis. The active Cl influx, the associateddownhill cation influxes, and passive ion fluxes were inhibitedby Dio-9, to a smaller extent than was photosynthesis. Cl influxin the light was often stimulated at concentrations of Dio-9,which inhibited photosynthesis. It was concluded that the activeK influx and Na efflux require ATP, while the active Cl influxdoes not. Possible links between the Cl influx and electrontransport and intermediates of photophosphorylation, and possibleinhibition by Dio-9 of the transport ATPase in the plasmalemmaare discussed.     

Temperature Adaptation of Active Sodium-Potassium Transport and of Passive Permeability in Erythrocytes of Ground Squirrels

Unidirectional active and passive fluxes of ⁴²K and ²⁴Na were measured in red blood cells of ground squirrels (hibernators) and guinea pigs (nonhibernators). As temperature is lowered, "active" (ouabain-sensitive) K influx and Na efflux were more greatly diminished in guinea pig cells than in those of ground squirrels. The fraction of total K influx which is ouabain sensitive in red blood cells of ground squirrels was virtually constant at all temperatures, whereas it decreased abruptly in guinea pig cells as temperature was lowered. All the passive fluxes (i.e., Na influx, K efflux, and ouabain-insensitive K influx and Na efflux) decreased logarithmically with decrease in temperature in both species, but in ground squirrels the temperature dependence (Q₁₀ 2.5–3.0) was greater than in guinea pig (Q₁₀ 1.6–1.9). Thus, red blood cells of ground squirrel are able to resist loss of K and gain of Na at low temperature both because of relatively greater Na-K transport (than in cells of nonhibernators) and because of reduced passive leakage of ions.     

Chloride and sodium influx: a coupled uptake mechanism in the squid giant axon

The squid giant axon was internally dialyzed while the unidirectional fluxes of either Cl or Na were measured. The effects of varying the internal or external concentration of either Na or Cl were studied. Chloride influx was directly proportional to the external Na concentration whereas Cl efflux was unaffected by changes of the external Na concentration between 0 and 425 mM. Neither Cl influx nor efflux were affected by changes of internal Na concentration over the range of 8-158 mM. After ouabain and TTX treatment a portion of the remaining Na influx was directly dependent on the extracellular Cl concentration. Furthermore, when the internal Cl concentration was increased from 0 to 150 mM, the influxes of Cl and Na were decreased by 14 and 11 pmol/cm2.s, respectively. The influx of both ions could be substantially reduced when the axon was depleted of ATP. The influxes of both ions were inhibited by furosemide but unaffected by ouabain. It is concluded that the squid axolemma has an ATP-dependent coupled Na-Cl co-transport uptake mechanism.     

Sodium and potassium fluxes and membrane potential of human neutrophils: evidence for an electrogenic sodium pump

Sodium and potassium ion contents and fluxes of isolated resting human peripheral polymorphonuclear leukocytes were measured. In cells kept at 37 degrees C, [Na]i was 25 mM and [K]i was 120 mM; both ions were completely exchangeable with extracellular isotopes. One-way Na and K fluxes, measured with 22Na and 42K, were all approximately 0.9 meq/liter cell water . min. Ouabain had no effect on Na influx or K efflux, but inhibited 95 +/- 7% of Na efflux and 63% of K influx. Cells kept at 0 degree C gained sodium in exchange for potassium ([Na]i nearly tripled in 3 h); upon rewarming, ouabain-sensitive K influx into such cells was strongly enhanced. External K stimulated Na efflux (Km approximately 1.5 mM in 140-mM Na medium). The PNa/PK permeability ratio, estimated from ouabain insensitive fluxes, was 0.10. Valinomycin (1 microM) approximately doubled PK. Membrane potential (Vm) was estimated using the potentiometric indicator diS-C3(5); calibration was based on the assumption of constant-field behavior. External K, but not Cl, affected Vm. Ouabain caused a depolarization whose magnitude dependent on [Na]i. Sodium-depleted cells became hyperpolarized when exposed to the neutral exchange carrier monensin; this hyperpolarization was abolished by ouabain. We conclude that the sodium pump of human peripheral neutrophils is electrogenic, and that the size of the pump-induced hyperpolarization is consistent with the membrane conductance (3.7-4.0 microseconds/cm2) computed from the individual K and Na conductances.     

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

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

Arterial effects of aldosterone and antimineralocorticoid compounds mechanism of action

The aim of our work was to study the mechanism of action of aldosterone and antialdosterone compounds on Na+ and K+ fluxes in vascular smooth muscle. In the long term, regulation of salt metabolism depends on aldosterone effects on Na+, K+, H+ and H2O transport by the renal tubules. Furthermore, it has been shown that aldosterone modifies several epithelial transports, inducing a positive sodium balance. The chronic in vivo administration of aldosterone modifies transmembrane ionic fluxes in vascular smooth muscle. Garwitz and Jones suggested that aldosterone may enhance net Na+ transport through the stimulation of the sodium pump. The results obtained in our laboratory indicate that aldosterone has a direct stimulatory action on ouabain-dependent and on ouabain-independent Na efflux. Furthermore, the mineralocorticoid enhances passive K permeability, as well as the Na pump dependent K influx. Both effects are blocked by antimineralocorticoid compounds. Recent experiments have shown that vasopressin potentiates some of the in vivo effects of aldosterone.     

Analysis of the monovalent ion fluxes in U937 cells under the balanced ion distribution: recognition of ion transporters responsible for changes in cell ion and water balance during apoptosis

Unidirectional (22)Na, Li(+) and Rb(+) fluxes and net fluxes of Na(+) and K(+) were measured in U937 human leukemic cells before and after induction of apoptosis by staurosporine (1 microM, 4 h) to answer the question which ion transporter(s) are responsible for changes in cell ion and water balance at apoptosis. The original version of the mathematical model of cell ion and water balance was used for analysis of the unidirectional ion fluxes under the balanced distribution of major monovalent ions across the cell membrane. The values of all major components of the Na(+) and K(+) efflux and influx, i.e. fluxes via the Na(+),K(+)-ATPase pump, Na(+) channels, K(+) channels, Na/Na exchanger and Na-Cl symport were determined. It is concluded that apoptotic cell shrinkage and changes in Na(+) and K(+) fluxes typical of apoptosis in U937 cells induced by staurosporine are caused by a complex decrease in the pump activity, Na-Cl symport and integral Na(+) channel permeability.     

Na(+) current through KATP channels: consequences for Na(+) and K(+) fluxes during early myocardial ischemia

During early myocardial ischemia, the myocytes are loaded with Na(+), which in turn leads to Ca(2+) overload and cell death. The pathway of the Na(+) influx has not been fully elucidated. The aim of the study was to quantify the Na(+) inward current through sarcolemmal KATP channels (IKATP,Na) in anoxic isolated cardiomyocytes at the actual reversal potential (Vrev) and to estimate the contribution of this current to the Na(+) influx in the ischemic myocardium. IKATP,Na was determined in excised single channel patches of mouse ventricular myocytes and macropatches of Xenopus laevis oocytes expressing SUR2A/Kir6.2 channels. In the presence of K+ ions, the respective permeability ratios for Na(+) to K(+) ions, PNa/PK, were close to 0.01. Only in the presence of Na(+) ions on both sides of the membrane was IKATP,Na similarly large to that calculated from the permeability ratio PNa/PK, indicative of a Na(+) influx that is largely independent of the K+ efflux at Vrev. With the use of a peak KATP channel conductance in anoxic cardiomyocytes of 410 nS, model simulations for a myocyte within the ischemic myocardium showed that the amplitude of the Na(+) influx and K(+) efflux is even larger than the respective fluxes by the Na(+) - K(+) pump and all other background fluxes. These results suggest that during early ischemia the Na(+) influx through KATP channels essentially contributes to the total Na+ influx and that it also balances the K(+) efflux through KATP channels.     

Some Ionic and Bioelectric Properties of the Ameba Chaos chaos

Ionic relationships in the giant ameba Chaos chaos were studied by analyzing bulk preparations of ground cytoplasm for K, Na, and Cl. Ion levels under normal conditions were compared with the levels in cells exposed to varying concentrations of different ions, for varying times and at different temperatures. By standard intracellular electrode techniques, the bioelectric potential, electrical resistance, and rectifying properties of the plasmalemma were studied on intact cells in media of different composition. The results obtained, when related to evidence from other studies on ion fluxes and osmotic relationships, suggest the following concept of ionic regulation in Chaos chaos. In the absence of active membrane uptake, the plasmalemma is essentially impermeable to anions but permeable to both K and Na, which enter passively. In the cold the cell does not discriminate between K and Na, the cytoplasmic level of K + Na is determined by a Donnan distribution, and osmotic imbalance leads to slow swelling. At normal temperatures active processes are added: Na and water are pumped out by the contractile vacuole system; Cl is accumulated, along with the colloid components of the cytoplasm, only during feeding and growth, which depend upon membrane uptake and intracellular membrane transformations. There is no evidence for active transport of any ion species directly across the plasmalemma.     

吲哚乙酸对向日葵下胚轴生长区单向K~+(~(86)Rb~+)通量的影响

植物生长素在刺激某些植物组织生长的同时促进K~+吸收和H~+分泌(Hager等1971,Cleland1975,赖寿鹏和倪晋山1983),可能是由于生长素刺激位于植物细胞质膜上的H~+泵ATP酶(Scherer 1984,赖寿鹏和倪晋山1985)。但以往的文献中只测定了生长素促进的植物组织对K~+的净吸收速率,即组织中K~+的累积速率或吸收溶液中K~+的减少速率。自从MacRobbie和Dainty(1958)首先运用放射性     

Sodium extrusion by the sea-water-acclimated fiddler crab Uca pugilator: comparison with other marine crustacea and marine teleost fish.

1. Measurements of the blood Na concentration and transepithelial electrical potential (T.E.P.) across Uca pugilator acclimated to sea water indicate that Na is maintained out of electrochemical equilibrium with sea water. The resulting net Na influx as well as the sodium gain due to ingestion of the medium must be balanced by extrarenal Na extrusion. 2. The small T.E.P. (-0.7 mV) and the 'transport numbers' of Na and Cl indicate that the permeability to these ions is equivalent. 3. Removal of external K results in a significant stimulation of unidirectional Na efflux that is dependent upon external Na but is not inhibited by ouabain. 4. Transfer of Uca to K and Na-free sea water results in a 54% decline in unidirectional efflux, which is not due to T.E.P. changes. Readdition of 25mM-K stimulates Na efflux much more than can be accounted for by changes in the T.E.P. Readdition of 25mM-Na to potassium-free sea water does not change the Na efflux. 5. The results indicate that Na extrusion by Uca is via a Na/K exchange mechanism which partially inhibits Na/Na exchange. Cessation of Na/K exchange (in K-free sea water) removes this inhibition and allows rapid Na/Na exchange. It is not known whether Na/K and Na/Na exchange are via the same or parallel carrier systems.     

Ion transport studies and determination of the cell wall elastic modulus in the marine alga Halicystis parvula

Using cultured cells of the marine alga, Halicystis parvula, we measured the concentrations of 11 inorganic ions in the vacuolar sap and the electrical potential difference (PD) between the vacuole and the external solution. In normal cells under steady-state conditions a comparison of the electrochemical equilibrium (Nernst) potential for each ion with the PD of -82 mV (inside negative) indicates that Na+ and K+ are actively transported out of the vacuole whereas all anions are pumped into the cell. Although the [K+] in the vacuole is only 9 mM, the cytoplasmic [K+] is about 420 mM, which suggests that the outwardly directed pump is at the tonoplast. Using large Halicystis cells we perfused the vacuole with an artificial seawater and conducted a short-circuit analysis of ion transport. The short-circuit current (SCC) of 299 peq - cm-2-s-1 is not significantly different from the net influx of Cl-. There is a small, but statistically significant net efflux of K+ (less than 1 pmol-cm-2.-1), while the influx and efflux of Na+ are not significantly different. Therefore, the SCC is a good measure of the activity of the Cl- pump. Finally, we measured the volumetric elastic modulus (epsilon) of the cell wall by measuring the change in cell volume when the internal hydrostatic pressure was altered. The value of epsilon at applied pressures between 0 and 0.4 atm is about 0.6 atm, which is at least 100-fold lower than the values of epsilon for all other algae which have been studied.     

Ion transport in Nitellopsis obtusa

The distribution and rates of exchange of the ions sodium, potassium, and chloride in single internodal cells of the ecorticate characean, Nitellopsis obtusa, have been studied. In tracer experiments three kinetic compartments were found, the outermost "free space" of the cell, a compartment we have called "protoplasmic non-free space", and the cell sap. The concentrations in the vacuole were 54 mM Na(+), 113 mM K(+), and 206 mM Cl(-). The steady state fluxes across the vacuolar membrane were 0.4 pmole Na(+)/cm.(2) sec., 0.25 pmole K(+)/cm.(2) sec., and 0.5 pmole Cl(-)/cm.(2) sec. The protoplasmic Na/K ratio is equal to that in the vacuole but protoplasmic chloride is relatively much lower. Osmotic considerations suggest a layer 4 to 6 micro thick with sodium and potassium concentrations close to those in the vacuole. The fluxes between protoplasm and external solution were of the order of 8 pmoles Na(+)/cm.(2) sec. and 4 pmoles K(+)/cm.(2) sec. We suggest that the protoplasm is separated from the cell wall by an outer protoplasmic membrane at which an outward sodium transport maintains the high K/Na ratio of the cell interior, and from the vacuole by the tonoplast at which an inward chloride transport maintains the high vacuolar chloride. The tonoplast appears to be the site of the principal diffusion resistance of the cell, but the outer protoplasmic membrane probably of the main part of the potential.