UV-Stimulated K Efflux from Rose Cells: Counterion and Inhibitor Studies

Irradiation of a washed suspension of cultured rose (Rosa damascena var. Gloire de Guilan) cells with about 1,680 joules per square meter of short wave ultraviolet (UV) light (254 nanometers) caused K⁺ to appear in the external medium. Short-term tracer (⁸⁶Rb⁺) experiments confirmed the earlier suggestion (Wright, Murphy 1978 Plant Physiol 61: 434-436) that UV increases the efflux of K⁺; there was also a small decrease in influx of K⁺. There was a partial recovery of fluxes from the effects of UV radiation, but no net accumulation of K⁺ within 16 to 18 hours after the irradiation. The K⁺ appearing in the medium was matched by an equivalent amount of HCO₃⁻; it was suggested that HCO₃⁻ was the principal counterion for the K⁺ flux induced by UV. Inhibitors of ATP synthesis (10⁻⁵ molar carbonyl cyanide m-chlorophenyl hydrazone; 0.05 millimolar KCN plus 0.75 millimolar salicylhydroxamic acid) strongly reduced the UV-stimulated K⁺ leakage, suggesting that the leakage was dependent in some way on ATP concentration inside the cells. The UV-induced K⁺ leakage was also dependent on temperature and the presence of Ca²⁺ in the external medium.     

Transmembrane potential-mediated coupling between H+ pump operation and K+ fluxes in Elodea densa leaves hyperpolarized by fusicoccin, light or acid load

In isolated Elodea densa leaves, the relationships between H⁺ extrusion (-ΔH⁺), K⁺ fluxes and membrane potential (E_m) were investigated for two different conditions of activation of the ATP-dependent H⁺ pump. The ‘basal condition’ (darkness, no pump activator present) was characterized by low values of-ΔH⁺ and K⁺ uptake (ΔK⁺), wide variability of the ?ΔH⁺/ΔK⁺ ratio, relatively low membrane polarization and E_m values more positive than EK for external K⁺ concentrations (|K⁺]_o of up to 2mol m^?3. A net K⁺ uptake was seen already at [K⁺]_o below 1 mol m^?3, suggesting that K⁺ influx in this condition was a thermodynamically uphill process involving an active mechanism. When the H⁺ pump was stimulated by fusicoccin (FC), by cytosol acidification, or by light (the ‘high polarization condition’), K⁺ influx largely dominated K⁺ and C^? efflux, and the ?ΔH⁺/ΔK⁺ ratio approached unity. In the range 50 mmol m^?3?5 mol m^?3 [K⁺]₀, E_m was consistently more negative than E_K. The curve of K⁺ influx at [K⁺]₀ ranging from 50 to 5000mmol m^?3 fitted a monophasic, hyperbolic curve, with an apparent half saturation value = 0–2 mol m^?3. Increasing |K⁺]₀ progressively depolarized E_m, counteracting the strong hyperpolarizing effect of FC. The effects of K⁺ in depolarizing E_m were well correlated with the effects on both K⁺ influx and ?ΔH⁺, suggesting a cause-effect chain: K⁺₀ influx → depolarization → activation of H⁺ extrusion. Cs⁺ competitively inhibited K⁺ influx much more strongly in the ‘high polarization’ than in the ‘basal’ condition (50% inhibition at [Cs⁺]/[K⁺]₀ ratios of 1:14 and 1:2, respectively) thus confirming the involvement of different K⁺ uptake systems in the two conditions. These results suggest that in E. densa leaves two distinct modes of interactions rule the relationships between H⁺ pump, membrane polarization and K⁺ transport. At low membrane polarization, corresponding to a low state of activation of the PM H⁺-ATP^ase and to E_m values more positive than E_K, K⁺ influx would mainly     

CFTR-mediated anion secretion in parathyroid hormone-treated Caco-2 cells is associated with PKA and PI3K phosphorylation but not intracellular pH changes or Na+/K+-ATPase abundance

Parathyroid hormone (PTH) has previously been shown to enhance the transepithelial secretion of Cl^? and HCO₃^? across the intestinal epithelia including Caco-2 monolayer, but the underlying cellular mechanisms are not completely understood. Herein, we identified the major signaling pathways that possibly mediated the PTH action to its known target anion channel, i.e., cystic fibrosis transmembrane conductance regulator anion channel (CFTR). Specifically, PTH was able to induce phosphorylation of protein kinase A and phosphoinositide 3-kinase. Since the apical HCO₃^? efflux through CFTR often required the intracellular H⁺/HCO₃^? production and/or the Na⁺-dependent basolateral HCO₃^? uptake, the intracellular pH (pH_i) balance might be disturbed, especially as a consequence of increased endogenous H⁺ and HCO₃^? production. However, measurement of pH_i by a pH-sensitive dye suggested that the PTH-exposed Caco-2 cells were able to maintain normal pH despite robust HCO₃^? transport. In addition, although the plasma membrane Na⁺/K⁺-ATPase (NKA) is normally essential for basolateral HCO₃^? uptake and other transporters (e.g., NHE1), PTH did not induce insertion of new NKA molecules into the basolateral membrane as determined by membrane protein biotinylation technique. Thus, together with our previous data, we concluded that the PTH action on Caco-2 cells is dependent on PKA and PI3K with no detectable change in pH_i or NKA abundance on cell membrane.     

Bicarbonate-stimulated ATPase in the renal proximal tubule luminal (brush border) memebrane.

Brush border membranes of the rabbit renal tubule have an ATPase which was stimulated 60% by 50 mm HCO₃^?. The K_a for HCO₃^? was 36 mm. Kinetic studies of the “HCO₃^?-ATPase” indicate that HCO₃^? had no effect on the K_m for ATP and ATP did not alter the K_a for HCO₃^?. Several anions, notably SO₃^2?, also accelerated the rate of dephosphorylation of ATP. The V for “SO₃^2?-ATPase” was fivefold greater than that for “HCO₃^?-ATPase.” The K_a for SO₃^2? was 0.78 mm. Other anions including Cl^? and phosphates, did not enhance ATPase activity. Thus, of the anions present in the glomerular filtrate in appreciable concentrations only HCO₃^? stimulated the luminal membrane enzyme. The anion-stimulated ATPase activity increased sharply from pH 6.1 to 7.1 and moderately with higher pH. The renal ATPase was not inhibited by SCN^? nor methyl sulfonyl chloride and was relatively insensitive to oligomycin and quercetin. Carbonyl cyanide p-trifluoromethoxy phenylhydrazone increased the basal rate of the membranal ATPase, suggesting that the ATPase activity is limited by transmembrane H⁺ flux. Carbonic anhydrase significantly increased the HCO₃^?-stimulated ATPase activity. This increment was blocked by Diamox. These findings provide evidence consistent with the hypothesis that the brush border membrane ATPase is involved in the extrusion of H⁺ from tubular cell to lumen and support suggested interrelationships between HCO₃^?-stimulated ATPase, H⁺ secretion, and bicarbonate transport in the kidney.     

Potential difference responses due to K+, Na+ and Cl− changes in Bullfrog antrum with and without HCO3−

The effect of changing [K⁺], [Na⁺] and [Cl^?] in nutrient solution was studied in bullfrog antrum with and without HCO₃^? in nutrient. In 25 mM HCO₃^? (95% O₂/5% CO₂) and in zero HCO₃^? (100% O₂), nutrient pH was maintained at 7.3. Changing from 4 to 40 mM K⁺ or from 81 to 8.1 mM Cl^? gave a decrease 10 min later in transmucosal PD (nutrient became more negative) — a normal response. These responses were less in zero than in 25 mM HCO₃^?. A decrease from 102 to 8 mM Na⁺ decreased PD (anomalous response of electrogenic NaCl symport). This effect was attenuated or eliminated in zero HCO₃^?. In contrast, change from 4 to 40 mM K⁺ gave initial anomalous PD response and change from 102 to 8 mM Na⁺, initial normal PD response with either zero or 25 mM HCO₃^?. Both responses were associated with (Na⁺ + K⁺)-ATPase pump and were greater in zero than in 25 mM HCO₃^?. Initial PD increases in zero HCO₃^? are explained as due to increase in the resistance of passive conductance and/or NaCl symport pathways. Thus, removal of HCO₃^? modifies conductance pathways of nutrient membrane.     

Early Effects of Penconazole on H+ and K+ Transport,Electrolyte Leakage and Transmembrane Electrical Potential in Egeria densa Leaves

The early effects of penconazole (PCZ) at relatively high concentration (10^?4 to 5 × 10^?4 M) on changes in pH and in titratable acidity of the medium, transmembrane electrical potential difference (Em), electrolyte leakage and cell morphology were investigated in Egeria densa leaves. At the lowest (10^?4 M) concentration and in the presence of a very low (10 μM) K⁺ concentration, triazole induced an early, moderate hyperpolarization of Em, associated with a decrease of net K⁺ uptake, suggesting some increase in the passive permeability to K⁺. This Em hyperpolarization was no longer detectable at high (2 mM) K⁺_out concentration. At high PCZ concentrations (3 × 10^?4 M and 5 × 10^?4 M) the early hyperpolarization detectable in the presence of a low K⁺_out concentration became transient, and was followed by a marked depolarization. PCZ, at these concentrations, suppressed acidification of the medium, stimulated electrolyte leakage and, in the mesophyll cells, induced some shrinking of the cytoplasm and its disconnection from the cell walls. These results are interpreted as due to an early effect of this triazole leading to the disorganization of the plasma membrane.     

Effects of Extracellular pH on UV-Induced K Efflux from Cultured Rose Cells

Ultraviolet (UV) light causes a specific leakage of K⁺ from cultured rose cells (Rosa damascena). During K⁺ efflux, there is also an increase in extracellular HCO₃⁻ and acidification of the cell interior. We hypothesized that the HCO₃⁻ originated from intracellular hydration of respiratory CO₂ and served as a charge balancing mechanism during K⁺ efflux, the K⁺ and HCO₃⁻ being cotransported out of the cell through specific channels. An alternative hypothesis which would yield similar results would be the countertransport of K⁺ and H⁺. To test these hypotheses, we studied the effect of a range of external pH values (pH 5-9), regulated by various methods (pH-stat, 100 millimolar Tris-Mes buffer, or CO₂ partial pressure), on the UV-induced K⁺ efflux. Both UV-C (<290 nanometers) and UV-B (290-310 nanometers) induced K⁺ efflux with a minimum at about pH 6 to 7, and greater efflux at pH values of 5, 8, and 9. Since pH values of 8 and 9 increased instead of reduced the efflux of K⁺, these data are not consistent with the notion that the efflux of K⁺ is dependent on an influx of H⁺, a process that would be sensitive to external H⁺ concentration. We suggest that the effect of pH on K⁺ efflux may be mediated through the titration of specific K⁺-transporting proteins or channels in the plasma membrane. Since we could not detect the presence of carbonic anhydrase activity in cell extracts, we could not use the location of this enzyme to aid in our interpretation regarding the site of hydration of CO₂.     

Effects of Cr on proton extrusion, potassium uptake and transmembrane electric potential in maize root segments

Abstract Proton extrusion of maize root Zea mays segments, was inhibited by the presence of Cr (o.n. + 6; present in solution as CrO₄^2-, Cr₂O₇^2-) in the incubation medium: the minimum inhibiting concentration was 2 × 10^?3 mol m^?3 and the inhibition progressively increased with Cr concentration. Cr inhibited proton extrusion. Also, when this activity was stimulated by the presence of K⁺ or fusicoccin (FC) in the incubation medium, the K⁺ and FC stimulating effect was still present when proton extrusion was inhibited by Cr. In addition, Cr inhibited K⁺ uptake. This inhibition was higher (50%) at K⁺ concentrations up to 1 mol m^?3 lower (15%) at higher K⁺ concentrations. This result indicates that the system responsible for K⁺ uptake operating at low K⁺ concentrations is more sensitive to Cr inhibition. Cr had no effect on transmembrane electric potential (PD). The depolarizing and hyper-polarizing effect of K+ and FC, respectively, were not affected by Cr; but Cr enhances the depolarizing effect of the uncoupler carbonylcyanide m-chlorophenylhydrazone (CCP). These results indicate that Cr inhibited the proton translocating mechanism coupled with K⁺ uptake, but did not change the net transport of charges through the plasmalemma. The Cr effect is discussed, taking into account the possibility of a direct effect of Cr at the membrane level or, alternatively, of an effect on some metabolic processes controlling membrane function.     

Taurine Stimulation of Isolated Hamster Brain Na+, K+-ATPase: Activation Kinetics and Chemical Specificity

Epileptic foci are associated with locally reduced taurine (2-aminoethanesulfonic acid) concentration and Na⁺, K⁺-ATPase (EC 3.6.1.3) specific activity. Topically applied and intraperitoneally administered taurine can prevent the development and/or spread of foci in many animal models. Taurine has been implicated as a possible cytosolic modulator of monovalent ion distribution, cytosolic “free” calcium activity, and neuronal excitability. Taurine may act in part by modulating Na⁺, K⁺-ATPase activity of neuronal and glial cells. We characterized the requirements for in vitro modulation of Na⁺, K⁺-ATPase by taurine. Normal whole brain homogenate Na⁺, K⁺-ATPase activity is 5.1 ± 0.4 (4) μmol P_i± h^?1± mg^?1 Lowry protein. Partial purification of the plasma membrane fraction to remove cytosolic proteins and extrinsic proteins and to uncouple cholinergic receptors yields a membrane-bound Na⁺, K⁺-ATPase activity of 204.6 ± 5.8 (4) mol P_i± h^?1± mg^?1 Lowry protein. Taurine activates the Na⁺, K⁺-ATPase at all levels of purification. The concentration dependence of activation follows normal saturation kinetics (K_1/2= 39 mM taurine, activation maximum =+87%). The activation exhibits chemical specificity among the taurine analogues and metabolites: taurine = isethionic acid > hypotaurine > no activation =β-alanine = methionine = choline = leucine. Taurine can act as an endogenous activator/modulator of Na⁺, K⁺-ATPase. Its action is mediated by a membrane-bound protein.     

The K+- and HCO3−-stimulated phosphatase activities in renal microsomes of rats

The K⁺-stimulated phosphatase activity of microsomes from rat kidney was not inhibited by l-phenylalanine, but the HCO₃^?-stimulated phosphatase activity was markedly inhibited by l-phenylalanine. Valinomycin enhanced the HCO₃^?-stimulated phosphatase activity, but did not enhance the K⁺-stimulated phosphatase activity. Ouabain did not inhibit the HCO₃^?-stimulated phosphatase activity, but inhibited the K⁺-stimulated phosphatase activity.The renal K⁺-stimulated phosphatase activity was suppressed to 40% of the control values by adrenalectomy, but the renal HCO₃^?-stimulated phosphatase activity was little suppressed by adrenalectomy. The renal K⁺-stimulated phosphatase activity in intact and adrenalectomized rats was found to be significantly elevated, in a manner similar to the elevation of the renal (Na⁺ + K⁺)-ATPase activity by aldosterone treatment (P < 0.02).     

Potassium-Induced Stimulation of Glutamate Uptake in Mouse Cerebral Astrocytes: The Role of Intracellular pH

Abstract: The Na⁺-glutamate cotransporters are believed to countertransport OH^? and K⁺. Previous evidence that the velocity of glutamate uptake can exceed the acid extrusion capacity of astrocytes raised the question of whether intracellular pH can become rate limiting for glutamate uptake. Cytoplasmic buffering capacity and acid extrusion in astrocytes are partially HCO₃^? dependent. Also, it was reported recently that raising extracellular [K⁺] alkalinizes astrocyte cytoplasm by an HCO₃^?-dependent mechanism. Here, we have compared glutamate uptake in HCO₃^?-buffered and HCO₃^?-depleted solutions at varying [K⁺]. We observed a pronounced stimulation of glutamate uptake by extracellular K⁺ (3–24 mM) that was substantially HCO₃^? dependent and affected preferentially the uptake of high concentrations (>25 µM) of glutamate. Stimulation of uptake by low extracellular [K⁺] (1.5–3 mM) was less dependent on HCO₃^?. Potassium-induced stimulation of uptake was weaker in rat astrocyte cultures than in mouse. The effects of Ba²⁺ and amiloride on glutamate uptake, as well as the HCO₃^?-dependent stimulatory effects of K⁺ and the species difference, all related consistently to effects on intracellular pH. The effects on uptake, however, were much larger than predicted by the associated changes in electrochemical gradient of OH^?. A “bimodal” scheme for glutamate transport can account qualitatively for the observed correlation between intracellular pH and velocity of glutamate uptake.     

Rubidium transport in the cyanobacterium Synechococcus R-2 (Anacystis nidulans, S. leopoliensis) PCC 7942

Synechococcus R-2 is a unicellular blue-green alga. The cells will grow on Rb⁺ as a substitute for K⁺ but at a slower rate (t₂～ 15 h versus 12 h). Potassium is not, strictly speaking, an essential element for Synechococcus. Rubidium duxes (using ⁸⁶Rb⁺) are much slower than those of potassium, about 1 nmol m^?2 s^?1 in the light (0.35 mol m^?3 Rb⁺). ⁸⁶Rb⁺ fluxes in the dark are about 0.1 nmol m^?2 s^?1. These fluxes are very slow compared to those of Na⁺ and other ions. Isotopic influx of Rb⁺ can supply sufficient Rb⁺ to keep up with the demands for growth, but the net dux needed to keep up with growth in the light is a large proportion of the total observed dux. Kinetic studies of Rb⁺ uptake versus [Rb⁺] show two uptake phases consistent with a high-affinity and a low-affinity system. Both systems appear to be light-activated. Transport of Rb⁺ appears to be passive at pH_o 10 in the light and dark. There is no case for active transport of Rb⁺ at pH_o 7.5 in the light, but a marginal case for active uptake in the dark (about 3 kJ mol^?1). There is only a small effect of Na⁺ upon Rb⁺ transport. ⁸⁶Rb⁺ should not be used in place of ⁴²K⁺ in K⁺ nutrition studies as the details of Rb⁺ transport are different to those of K⁺ transport.     

Turgor regulation in Lamprothamnium papulosum. I. I/V analysis and pharmacological dissection of the hypotonic effect

Current-voltage (I/V) analysis and pharmacological dissection were applied to membranes of Lamprothamnium at the time of hypotonic stress. At least three types of process were found to be involved in the response to this stress.

• 1 The first 10min of exposure to hypotonic medium resulted in a depolarization of about 50mV accompanied by a decrease or no change in conductance. This depolarization occurred with either K⁺ or Ca²⁺ (and consequently C^? channels inactivated.
• 2 The CI^? channels opened mainly in the first 15min of the hypotonic stress, increasing the membrane conductance by about an order of magnitude.
• 3 The K⁺ conductance rose as the Cl^? conductance started to diminish and reached a maximum after about 40 min.

Both types of channel were strongly potential-dependent with a conductance peak between -150 and 0mV. An inactivation of K⁺or CI^? channels resulted in moving the membrane potential away from the conductance maximum toward either E_K or E_CI, diminishing the ion efflux (and turgor regulation). The time courses of the conductance increases remained the same, suggesting that the conductance changes are not driven by feedback to some preset turgor level. The electrophysiology of the Lamprothamnium transporters is compared to that of salt-sensitive charophytes.     

Characterization of a K+-ATPase from Lactobacillus helveticus ATCC 15009

Lactobacillus helveticus ATCC 15009 (wild-type) membrane preparations hydrolyzed Mg²⁺-ATP as a function of K⁺ concentration (2–200 mM). Mg²⁺-ATP hydrolysis by L. helveticus membranes was strongly inhibited in the absence of exogenous K⁺, while it amounted to 6 nmol ATP hydrolyzed min^–1 (mg membrane protein)^–1 at 50 mM KCl (saturating conditions) and pH 7.2. The K⁺-dependent ATPase of L. helveticus displayed a relatively high affinity for potassium ions (K _m = 800 μM) and was not affected by pretreatment of membranes with N,N’-dicyclohexylcarbodiimide. Membrane preparations were subjected to hypotonic shock to obtain a maximum yield of open profiles. The formation of a maximum level of enzyme-phosphate complex with a molecular mass of approximately 82 kDa was induced upon treatment of L. helveticus membrane preparations with low concentrations of [γ-³²P]ATP in the presence of K⁺ and La³⁺ ions and was visualized by acidic SDS-PAGE. It was concluded that L. helveticus membranes contain an inwardly directed K⁺ pump whose presence is discussed in terms of its putative role in cytoplasmic pH regulation. Received: 16 December 1996 / Accepted: 14 May 1997     

Differential effects of Na+, Mg2+, K+, Ca2+ and osmotic stress on the wild type and the NaCl-tolerant mutants stl1 and stl2 of Ceratopteris richardii

In order to identify physiological components that contribute to salinity tolerance, we compared the effects of Na⁺, Mg²⁺ and K⁺ salts (NaCl, Na₂SO₄, MgCl₂, MgSO₄, KCl and K₂SO₄), Ca₂₊ (CaSO₄), mannitol and melibiose on the wild type and the single-gene NaCl-tolerant mutants stl1 and stl2 of Ceratopteris richardii. Compared with gametophytic growth of the wild type, stl2 showed a low level of tolerance that was restricted to Na⁺ salts and osmotic stress. stl2 exhibited high tolerance to both Na⁺ and Mg²⁺ salts, as well as to osmotic stress. In response to short-term exposure (3 d) to NaCl, accumulation of K⁺ and Na⁺ was similar in the wild type and stl1. In contrast, stl2 accumulated higher levels of K⁺ and lower levels of Na⁺. Ca²⁺ supplementation (1.0 mol m^?3) ameliorated growth inhibition by Na⁺ and Mg²⁺ stress in wild type and stll, but not in stl2. In addition, under Na⁺ stress (175 mol m^?3) wild-type, stll and stl2 gametopbytes maintained higher tissue levels of K⁺ and lower levels of Na⁺ when supplemented with Ca²⁺ (1.0 mol m^?3). stl2 gametophytes were extremely sensitive to K⁺ supplementation. Growth of stl2 was greater than or equal to that of the wild type at trace concentrations of K⁺ but decreased substantially with increasing K⁺ concentration. Supplementation with K⁺ from 0 to 1.85 mol m^?3 alleviated some of the inhibition by 75 mol m^?3 NaCl in the wild type and in stl1. In stl2, growth at 75 mol m^?3 NaCl was similar at 0 and 1.85 mol m^?3 K⁺ supplementation. Although K⁺ supplementation above 1.85 mol m^?3 did not alleviate inhibition of growth by Na⁺ in any genotype, stl2 maintained greater relative tolerance to NaCl at all K⁺ concentrations tested.     

The effect of CO2 on potassium transport by Chlorella fusca

Abstract. The effect of CO₂ on net K⁺ uptake by Chlorella fusca grown on high CO₂ levels was examined by passing 1.5% CO₂ through algal suspensions gassed previously with air or CO₂-free air Addition of CO₂ in the light caused a large net uptake of K⁺ (initial velocity 4.2–9.2 mmol s^?1 m^?3 cells) which decreased the concentration of K⁺ in the supernatant from 0.1–0.2 mol m^?3 to 3–10 mmol m^?3. In the dark and in the presence of 30 mmol m^?3 DCMU, no effects were found. Measurement or the unidirectional K⁺ fluxes by using ⁸⁶Rb⁺ as a label showed that in the presence of 1.5% CO₂, influx of K⁺ was increased by a factor of 2–4 while efflux was inhibited completely. CO₂ hyperpolarized the membrane potential (determined through TPP⁺ uptake) from –120mV to –130 mV which could not explain the more than 15,000-fold K⁺ accumulations. In the light, CO₂ lowered the intracellular pH (determined with DMO) by 0.5 units. In the dark and in the presence of DCMU only, a small acidification of 0.1 units was found. During the first 15 min after addition of CO₂ the malate content of the cells increased from 0.7 to 1.5 mol m^?3 packed cells. On the basis of these and earlier results, CO₂-induced net K⁺ uptake is interpreted as a stimulation of an electroneutral ATP-dependent K⁺/H⁺ exchange at the plasmalemma. This exchange acts as a ‘pHstat’ by reducing the intracellular acidification caused by production of acidic assimilation products.     

Endothelin 1 Stimulates Na+,K+-ATPase and Na+-K+-Cl− Cotransport Through ETA Receptors and Protein Kinase C-Dependent Pathway in Cerebral Capillary Endothelium

Abstract: The effect of endothelins (ET-1 and ET-3) on ⁸⁶Rb⁺ uptake as a measure of K⁺ uptake was investigated in cultured rat brain capillary endothelium. ET-1 or ET-3 dose-dependently enhanced K⁺ uptake (EC₅₀ = 0.60 ± 0.15 and 21.5 ± 4.1 nM, respectively), which was inhibited by the selective ET_A receptor antagonist BQ 123 (cyclo-d -Trp-d -Asp-Pro-d -Val-Leu). Neither the selective ET_B agonists IRL 1620 [N-succinyl-(Glu⁹,-Ala^11,15)-ET-1] and sarafotoxin S6c, nor the ET_B receptor antagonist IRL 1038 [(Cys¹¹,Cys¹⁵)-ET-1] had any effect on K⁺ uptake. Ouabain (inhibitor of Na⁺,K⁺-ATPase) and bumetanide (inhibitor of Na⁺-K⁺-Cl^? cotransport) reduced (up to 40% and up to 70%, respectively) the ET-1-stimulated K⁺ uptake. Complete inhibition was seen with both agents. Phorbol 12-myristate 13-acetate (PMA), activator of protein kinase C (PKC), stimulated Na⁺,K⁺-ATPase and Na⁺-K⁺-Cl^? cotransport. ET-1-but not PMA-stimulated K⁺ uptake was inhibited by 5-(N-ethyl-N-isopropyl)amiloride (inhibitor of Na⁺/H⁺ exchange system), suggesting a linkage of Na⁺/H⁺ exchange with ET-1-stimulated Na⁺,K⁺-ATPase and Na⁺-K⁺-Cl^? cotransport activity that is not mediated by PKC.     

Sodium Transport in Capillaries Isolated from Rat Brain

Abstract: Brain capillary endothelial cells form a bloodbrain barrier (BBB) that appears to play a role in fluid and ion homeostasis in brain. One important transport system that may be involved in this regulatory function is the Na⁺,K⁺-ATPase that was previously demonstrated to be present in isolated brain capillaries. The goal of the present study was to identify additional Na⁺ transport systems in brain capillaries that might contribute to BBB function. Microvessels were isolated from rat brains and ²²Na ⁺ uptake by and efflux from the cells were studied. Total ²²Na ⁺ uptake was increased and the rate of ²²Na ⁺ efflux was decreased by ouabain, confirming the presence of Na⁺,K⁺-ATPase in capillary cells. After inhibition of Na⁺,K⁺-ATPase activity, another saturable Na ⁺ transport mechanism became apparent. Capillary uptake of ²²Na ⁺ was stimulated by an elevated concentration of Na ⁺or H⁺ inside the cells and inhibited by extracellular Na⁺, H⁺, Li⁺, and NH₄⁺. Amiloride inhibited ²²Na ⁺ uptake with a K_i between 10^?5 and 10^?6M but there was no effect of 1 mM furosemide on ²²Na⁺ uptake by the isolated microvessels. These results indicate the presence in brain capillaries of a transport system capable of mediating Na ⁺/ Na ⁺ and Na ⁺/H ⁺ exchange. As a similar transport system does not appear to be present on the luminal membrane of the brain capillary endothelial cell, it is proposed that Na ⁺/H ⁺ exchange occurs primarily across the antiluminal membrane.     

Neutral Red as a Redox Dye Induces K+ Efflux and Current-Voltage Changes in Eremosphaera,Lemna, and Guard Cells*

Membrane effects of the redox and pH indicator neutral red were studied with the chlorococcal alga Eremosphaera viridis, with Lemna gibba, and with “isolated” guard cells in epidermal peels of Valerianella locusta. Neutral red was extracellularly reduced and caused transmembrane current-voltage changes, an increase in membrane conductance by about 14 nS, an apparent K⁺ net efflux of up to 120 μmol g^?1 FW in 5 min, and an intracellular acidification by up to 0.7 pH units. Neutral red-triggered K⁺ net efflux was most pronounced at low pH, at an E_o more positive than ?200 mV, and without extracellular Ca²⁺. From the experimental data it is concluded that, due to the redox function of the phenazine molecule, extracellular neutral red triggers a trans-plasmalemma e^? transfer, leading to strong membrane depolarization and charge compensating K⁺ net efflux, in addition to some unspecific ion release. As a consequence the intracellular concentration of strong cations relative to strong anions (SID) decreases, resulting in intracellular acidification.     

Interactions between K+ and NH4+: effects on ion uptake by rice roots

Interactive effects of K⁺ and N (principally NH₄⁺) on plant growth and ion uptake were investigated using hydroponically grown rice (Oryza sativa L. cv. M202) seedlings by varying the availability of NH₄⁺ or NO₃^? and K⁺ during an 18d growth period, a 3d pretreatment period and during flux measurements. Plants grew best in media containing 100 mmol m^?3 NH₄⁺ and 200mmolm^?3 K⁺ (N100/K200), followed by N2/K200 < N100/K2 < N2/K2. ⁸⁶Rb⁺(K⁺) fluxes were increased by exposure to N during the 18 d growth period and the 3 d of pretreatment, but decreased by the presence of NH₄⁺ during flux measurements. This inhibition was a function of prior N/K provision and the [NH₄⁺]₀ present during flux determinations. NH₄⁺ was least inhibitory to 86Rb⁺(K⁺) influx in high-N/low-K plants. Pretreatments with K⁺ failed to stimulate NH₄⁺ uptake, and the presence of K⁺ in the uptake solutions reduced NH₄⁺ fluxes only in high-N/low-K plants.