Accumulation of sodium and potassium ions in oocytes of the river lamprey Lampetra fluviatilis during prespawning period

Accumulation of Na+ and K+ ions in oocytes of the river lamprey Lampetra fluviatilis and their transport across the plasma membrane is realized by two main mechanisms--Na,K-pump and Na,K,Cl-cotransport. At the prespawning period from December to May the intracellular Na+ concentration was observed to increase from 10 to 25 mM and the K+ concentration--from 28 to 45 mM. Results obtained on isolated oocytes with aid of 204Tl radioactive label have shown that contributions of the Na,K-pump and Na,K,Cl-cotransport to potassium accumulations were close until March. In spring, the total K+ inflow almost doubled owing to activation of the Na,K-pump, whereas contribution of Na,K,Cl-cotransport did not change. It seems that an increase of the intracellular content of the main inorganic cations in oocytes resulted in parallel activation of the Na,K-pump and probably of Na/H-exchange. The biological significance of activation of these mechanisms of ion transport at the prespawning period might be due to a necessity of accumulation of Na+ and K+ ions in concentrations optimal for subsequent embryonic development.     

Accumulation of sodium and potassium ions in oocytes of the river lamprey <Emphasis Type="Italic">Lampetra fluviatilis</Emphasis> at the prespawning period

Accumulation of Na⁺ and K⁺ ions in oocytes of the river lamprey Lampetra fluviatilis and their transport across the plasma membrane is realized by two main mechanisms-Na,K-pump and Na,K,Cl-cotransport. At the prespawning period from December to May, the intracellular Na⁺ concentration was observed to increase from 10 to 25 mM and the K⁺ concentration-from 28 to 45 mM. Results obtained on isolated oocytes with aid of radioactive labels ²²Na and ²⁰⁴Tl have shown that contributions of the Na,K-pump and Na,K,Cl-cotransport to potassium accumulations were close until March. In spring, the total K⁺ inflow almost doubled owing to activation of the Na,K-pump, whereas contribution of Na,K,Cl-cotransport did not change. It seems that an increase of the intracellular content of the main inorganic cations in oocytes resulted in parallel activation of the Na,K-pump and probably of Na/H-exchange. The biological significance of activation of these mechanisms of ion transport at the prespawning period might be due to a necessity of accumulation of Na⁺ and K⁺ ions in concentrations optimal for subsequent embryonic development.     

Different Permeability of Potassium Salts across the Blood-Brain Barrier Follows the Hofmeister Series

The passage of ions across biological membranes is regulated by passive and active mechanisms. Passive ion diffusion into organs depends on the ion-pairing properties of salts present in the serum. Potassium ions could affect brain activity by crossing the blood-brain barrier (BBB) and its accumulation in the extracellular cerebral space could precipitate seizures. In the present study, we analyze passive diffusion of a series of potassium salts in the in vitro isolated guinea pig brain preparation. Different potassium counter-anions confer ion-pairing and lipophilicity properties that modulate membrane diffusion of the salt. Extracellular recordings in different cortical areas demonstrated the presence of epileptiform activities that strongly relate to anion identity, following the qualitative order of the Hofmeister series. Indeed, highly lipophilic salts that easily cross the BBB enhanced extracellular potassium concentration measured by ion-selective electrodes and were the most effective pro-epileptic species. This study constitutes a novel contribution for the understanding of the potential epileptogenicity of potassium salts and, more generally, of the role of counter-anions in the passive passage of salts through biological membranes.     

Mechanism of radiation injury in the ion transport systems of nervous tissue

The biological effect of ionizing radiation (IR) in lethal and sublethal doses on the sodium-potassium transport systems in the fractions, enriched of neuron and glial cells and in cortex slices from rat brain was investigated. It was shown that IR leads to marked disturbances in the activity of Na,K-ATPase both in neuron and in glial cells. Some phasic character of alterations may be noted, which is expressed in different degree for various cellular elements of the brain. Using the surviving brain slices we have shown that IR causes essential phasic changes in potassium ion reaccumulation in different times after exposure. The mechanisms of the disturbance of Na,K-pump function in nervous tissue after irradiation are under discussion.     

Relationship between ionic surroundings and insulin actions on glucose transport and Na,K-pump in muscles

1. It is well known that insulin has various effects on glucose transport and the Na,K-pump in muscles. It is also known to have some effects on the membrane potential--in general, insulin induces a hyperpolarization of the membrane in muscles. Furthermore, it is suggested that the actions of insulin are modified by changes in ionic surroundings. 2. In this review article, the actions of ionic surroundings and insulin on glucose transport in muscles are discussed; in particular, the effects of changes in extracellular and/or intracellular concentrations of Na, K, Ca and H ions will be mentioned. 3. The actions of ionic surroundings and insulin on the Na,K-pump in muscles are discussed; in particular, the effects of changes in extracellular an/or intracellular concentrations of Na, K, Ca and H ions will be examined. 4. The relationship between the actions of ionic surroundings and insulin are discussed. 5. In particular, the effects of changes in ionic surroundings on the insulin-induced hyperpolarization of the membrane are discussed by relating it to the Na,K-pump function. The relationship between the insulin-induced change in membrane potential and glucose transport will be also mentioned.     

Active transport of thallous ions by Streptococcus lactis.

Tl+ ions have been shown to mimic or compete with K+ in a number of membrane systems. We confirmed that in starved, valinomycin-treated cells of Streptococcus lactis 7962, Tl+ ions distributed themselves across the bacterial membrane in response to the potassium diffusion potential. In glucose-energized cells, however, Tl+ was taken up by a system specifically stimulated by sodium salts. The intracellular levels of Tl+ exceeded those attained by [3H]triphenylmethylphosphonium ion, a lipophilic cation which accumulates in response to the membrane potential. The uptake of Tl+ by (Na+ and glucose)-stimulated cells was strongly inhibited by potassium salts. These experiments suggest that metabolic energy is coupled to Tl+ transport by means of a high energy phosphate compound and that Tl+ ions are actively transported by a membrane carrier whose normal substrate is K+. The uptake of Tl+ is not a valid method for determining the streptococcal membrane potential.     

Na and K Fluxes in Nitella clavata

Na and K influxes and effluxes, membrane potentials, and cell ion concentrations of Nitella clavata were measured as functions of external NaCl concentrations and time. It appears necessary to conclude that active K transport into the cells as well as active Na extrusion is present, although the latter is of small magnitude and possibly is explicable as exchange-diffusion. An attempt has been made to account for the capacity of the cells to discriminate between K and Na ions and yet have fairly independent passive ion movements. This is done by proposing a model in which the permeation areas are "slit-pores" rather than cylindrical pores. The slit-pores would permit rather independent movements of ions within them so long as the pores do not tend to become saturated from both or either side. In the latter case one way movement results. The experimental results are in fair agreement with this suggested model.     

The causal theory of the resting potential of cells

In this pedagogical article the causal theory of the resting potential of cells is presented, which for given extracellular ion concentrations predicts the intracellular ones simultaneously with the resting potential. In addition to the Na, K-pump, fixed charges on the membrane surfaces are taken into account. The equation determining the resting potential in the causal theory suggests a new explanation of the genesis of the resting potential. The usual criterion for an ion pump to be electrogenic is not relevant for the whole of the resting potential, and may therefore be misleading. The physical meaning of the Goldman-Hodgkin-Katz formula for the membrane potential as a diffusion potential is also explained and tested with numbers for the giant axon of the squid. A significant discrepancy between theory and experiment is found which calls for an experimental re-examination of the constitutive equations for passive potassium and sodium currents.     

Ion leakage through transient water pores in protein-free lipid membranes driven by transmembrane ionic charge imbalance

We have employed atomic-scale molecular dynamics simulations to address ion leakage through transient water pores in protein-free phospholipid membranes. Our results for phospholipid membranes in aqueous solution with NaCl and KCl salts show that the formation of transient water pores and the consequent ion leakage can be induced and be driven by a transmembrane ionic charge imbalance, an inherent feature in living cells. These processes take place if the gradient is large enough to develop a sufficiently significant potential difference across the membrane. The transport of cations and anions through the water pores is then seen; it discharges the transmembrane potential, considerably reduces the size of a water pore, and makes the water pore metastable, leading eventually to its sealing. The ion transport is found to be sensitive to the type of ions. It turns out that Na(+) and Cl(-) ions leak through a membrane at approximately the same ratio despite the fact that Na(+) ions are expected to experience a lower potential barrier for the permeation through the pore. This is because of strong interactions of sodium ions with the carbonyl region of a phospholipid membrane as well as with lipid headgroups forming pore "walls," considerably slowing down the permeation of sodium ions. In contrast, we observed a pronounced selectivity of a phospholipid membrane to the permeation of potassium ions as compared to chloride ions: Potassium ions, being larger than sodium ions, interact only weakly with phospholipid headgroups, so that these interactions are not able to compensate for a large difference in free-energy barriers for permeation of K(+) and Cl(-) ions. These findings are found to be robust to a choice of force-field parameters for ions (tested by Gromacs and Charmm force-fields for ions). What is more, a potassium ion is found to be able to permeate a membrane along an alternate, "water-defect-mediated" pathway without actual formation of a pore. The "water-defect-mediated" leakage involves formation of a single water defect only and is found to be at least one order of magnitude faster than the pore-mediated ion leakage.     

Is flexoelectricity the coupling factor between chemical energy and osmotic work in the pump? A model of pump

The following pump model is proposed. A gate is responsible for pump specificity. The actual driving force of the transport of ions against the electrochemical potential gradient is the electric field originating from an altered curvature of the phospholipid bilayer around the pump. The physical origin of this curvature-induced electric field arises from a basic liquid crystal property of lipid bilayers called flexoelectricity. Alterations occurring in phospholipid bilayer arrangement are due to changed conformation of protein; the main energy source of this change is ATP. Consequently, the energy of ATP is transformed, in our pump model, into osmotic work in following steps: ATP + protein (conformation I)----protein (conformation II)----alterations in phospholipid bilayer arrangement----electric field----active transport of ions. This model is the most simple one. In Na, K-pump there is a bidirectional ion transport. In our model of Na, K-pump three conformational states of pump proteins and two different electric fields formed sequentially in opposite directions are supposed.     

The transport of Na+ and K+ ions through phospholipid bilayers mediated by the antibiotics salinomycin and narasin studied by 23Na- and 39K-NMR spectroscopy

Addition of the ionophoric antibiotics salinomycin or narasin to preparations of large unilamellar vesicles made from egg yolk phosphatidylcholine in sodium or potassium chloride solutions gives rise to dynamic effects in the 23Na- and 39K-NMR spectra. The dynamic spectra arise from the ionophore-mediated transport of the metal ions through the membrane. The kinetics of the transport are followed as a function of the concentrations of ionophore and the metal ion and are compatible in all cases with a model in which one ionophore molecule transports one metal ion. For both ionophores the transport of potassium ions is appreciably faster than that of sodium and in both cases the rate-limiting step for sodium transport is dissociation of the ionophore-metal complex. Assuming dissociation to be rate limiting in all four cases it is shown that the transport rate differences between the pairs of complexes of each metal arise solely from differences in the rates of formation. The stability constants for ionophore-metal complex formation in the membrane/water interface are evaluated.     

Functional expression of N-terminal truncated alpha-subunits of Na,K-ATPase in Xenopus laevis oocytes.

N-terminal deletion mutants of Na,K-ATPase alpha 1 isoforms initiating translation at Met34 (alpha 1T1) or at Met43 (alpha 1T2) were expressed in X. laevis oocytes. Compared to beta 3 cRNA injected controls, the co-expression of alpha 1wt, alpha 1T1, alpha 1T2 with beta 3 subunits results in a 2- to 3-fold increase of ouabain binding sites, parallelled by a concomitant increase in Na,K-pump current. The apparent K1/2 for potassium activation of the alpha 1T2/beta 3 Na,K-pumps is significantly higher than that of the alpha 1wt/beta 3 or alpha 1T1/beta 3 Na,K-pumps expressed at the cell surface. Total deletion of the lysine-rich N-terminal domain thus allows the expression of active Na,K-pump but with distinct cation transport properties.     

D2O as a modifier of ionic specificity of Na, K-ATPase

Effect of heavy water D2O on the rate of hydrolysis of ATP and pNPP by Na,K-ATPase was studied. Heavy water of high concentration inhibits the rate of ATPase reaction in all the studied ratios of the ions Na/K at constant ionic strength 150 mM. Activation of the enzyme was observed in the solution with low concentration of heavy water (less than 5%). The value of isotope effects depended on the ratio between sodium and potassium ion concentrations in the medium. At low temperature no activation of the enzyme with heavy water in low concentration was observed. Substitution of usual water for the heavy one was accompanied by a decrease of apparent constants of enzyme activation with sodium and potassium ions. During pNPP hydrolysis with Na,K-ATPase an increase of reaction rate in the medium with heavy water was observed. Substitution of potassium ions by cesium resulted in an increase of isotope effects during ATP and pNPP hydrolysis. Analysis of isotope effects in terms of the molecular model of sodium pump proposed permits a conclusion that the isotope effects of heavy water are explained by its influence as a solvent, the binding centres of potassium and sodium ions are localized in different regions of the enzyme differing in physico-chemical properties. The structure of sodium centres is controlled by hydrogen bonds, and of potassium ones--by hydrophobic interactions; the transport of ions by the enzyme is accompanied by dehydration of ions.     

Na,K-ATPase in excitation-contraction coupling of vascular smooth muscle from cattle.

Lowering the extracellular K+ content from 6 to 0.6 mM causes a rise, and elevation from 6 to 8.5 mM a fall of 45Ca++ efflux from the vascular smooth muscle cells of the arteria carotis communis of cattle. In contrast, a level of 17 mM K+ has no influence. Removal of extracellular calcium does not block these effects. 10(-4) M ouabain also induces a rise in Ca++ efflux, additional potassium reduction then being without effect; 10(-9) M ouabain is of no influence. The 45Ca++ efflux kinetics correlates with the activity of the isolated Na,K-ATPase. Tonus increases of the vascular strips by 10(-4) M ouabain and potassium deficiency cannot be blocked by 4 mM lanthanum or removal of extracellular calcium. Unlike sodium, potassium stimulates the active Ca++ binding and the activity of the Ca-ATPase of the microsomal fraction. The ative Ca++ binding of the mitochondria is stimulated by both ions. It is postulated that the activity of the plasma membrane Na,K-pump is able to regulate the tonus of big arteries through alteration of Ca++ storage processes.     

Resolution of Pump and Leak Components of Sodium and Potassium Ion Transport in Human Erythrocytes

Further support for the pump-leak concept was obtained. Net transport was resolved into pump and leak components with the cardiac glycoside, ouabain. The specificity of ouabain as a pump inhibitor was demonstrated by its ineffectiveness when the pump was already inhibited by lack of one of the three pump substrates, sodium ion, potassium ion, or adenosine triphosphate. In the presence of ouabain the rates of passive transport of sodium and potassium ions changed almost in proportion to changes in their extracellular concentrations when one ion was exchanged for the other. In the presence of ouabain and at the extracellular concentrations which produced zero net transport, the ratio of potassium ions to sodium ions was 1.2-fold higher inside the cells than outside. This finding was attributed to a residual pump activity of less than 2% of capacity. The permeability to potassium ions was 10% greater than the permeability to sodium ions. A test was made of the independence of pump and leak. Conditions were chosen to change the rate through each pathway separately or in combination. When both pathways were active, net transport was the sum of the rates observed when each acted separately. A ratio of three sodium ions pumped outward per two potassium ions pumped inward was confirmed.     

Changes in the neuronal membrane potential of the mollusk Planorbarius corneus under the action of cardiac glycosides

In experiments on isolated neurones from the gastropod mollusc P. corneus, strophantin and digoxin in low concentrations produce slow hyperpolarization, in higher ones--depolarization; at concentrations about 1 mM, hyperpolarization was more evident. In all cases, the decrease in membrane resistance was observed. Presumably, membrane permeability for potassium ions increases. During application of the drugs in concentrations 10-100 microM, hyperpolarization may be masked by depolarization due to block of Na,K-pump. Higher concentrations, increasing potassium permeability of the membrane, may result in substitution of depolarization by hyperpolarization.     

The electrical potential profile of gallbladder epithelium.

In this study the relative ionic permeabilities of the cell membranes of Necturus gallbladder epithelium have been determined by means of simultaneous measurement of transmural and transmucosal membrane potential differences (PD) and by ionic substitution experiments with sodium, potassium and chloride ions. It is shown that the mucosal membrane is permeable to sodium and to potassium ions. The baso-lateral membrane PD is only sensitive to potassium ions. In both membranes chloride conductance is negligible or absent. The ratio of the resistances of the mucosal and baso-lateral membranes, RM/RS, increases upon reducing the sodium concentration in the mucosal solution. The same ratio decreases when sodium is replaced by potassium which implies a greater potassium than sodium conductance in the mucosal membrane. The relative permeability of the shunt for potassium, sodium and chloride ions is: PK/PNa/PCl=1.81:1.00:0.32. From the results obtained in this study a value for the PK/PNa ratio of the mucosal membrane could be evaluated. This ratio is 2.7. From the same data the magnitude of the electromotive forces generated across the cell membranes could be calculated. The EMF's are -15mV across the mucosal membrane and -81mV across the baso-lateral one. Due to the presence of the low resistance shunt the transmucosal membrane PD is -53.2mV (cell inside negative) and the transmural PD is +2.6mV (serosal side positive). The change in potential profile brought about by the low resistance shunt favors passive entry of Na ions into the cell across the mucosal membrane. Calculations show that this passive Na influx is maximally 64% of the net Na flux estimated from fluid transport measurements. The C-1 conductive of the baso-lateral membrane is too small to allow electrogenic coupling of C1 with Na transport across this membrane. Experiments with rabbit gallbladder epithelium indicate that the membrane properties in this tissue are qualitatively similar to those of Necturus gallbladder epithelium.     

Electrical potential differences across salt transporting membranes

The electrical potential differences across membranes where active transport of ions occurs has been examined using the formalism of linear non-equilibrium thermodynamics, and can be represented as the arithmetic sum of a resistive term, a term directly dependent on metabolism (i.e. electrogenic) and terms appropriate for describing a diffusion potential. The Hittorf transport number for each ion in the latter terms is the ratio of the partial conductances of the membrane to that ion to the total membrane conductance, and the conductance to an ion consists of the arithmetic sum of conductance of active and passive pathways providing these are independent. The conductances of active transport mechanisms arise from variation of the rate of transport with the electrochemical potentials against which they operate. The electrogenic term arises from imbalance between anion and cation transport. If an ion is transported by an obligatorily electrically neutral exchange for some other ion such transport gives rise to no electrogenic effect. A membrane will transport salt most efficiently if there is no imbalance between anion and cation transport, when it will not be electrogenic, but modest deviations from this condition will not degrade the efficiency of active transport markedly.     

CHIF, a member of the FXYD protein family, is a regulator of Na,K-ATPase distinct from the gamma-subunit

The biological role of small membrane proteins of the new FXYD family is largely unknown. The best characterized FXYD protein is the gamma-subunit of the Na,K-ATPase (NKA) that modulates the Na,K-pump function in the kidney. Here, we report that, similarly to gamma(a) and gamma(b) splice variants, the FXYD protein CHIF (corticosteroid-induced factor) is a type I membrane protein which is associated with NKA in renal tissue, and modulates the Na,K-pump transport when expressed in Xenopus oocytes. In contrast to gamma(a) and gamma(b), which both decrease the apparent Na+ affinity of the Na,K-pump, CHIF significantly increases the Na+ affinity and decreases the apparent K+ affinity due to an increased Na+ competition at external binding sites. The extracytoplasmic FXYD motif is required for stable gamma-subunit and CHIF interaction with NKA, while cytoplasmic, positively charged residues are necessary for the gamma-subunit's association efficiency and for CHIF's functional effects. These data document that CHIF is a new tissue-specific regulator of NKA which probably plays a crucial role in aldosterone-responsive tissues responsible for the maintenance of body Na+ and K+ homeostasis.