Active Transport Of Sodium Ions Research Articles

Stabilization of Hypoxia Inducible Factor by Cobalt Chloride Can Alter Renal Epithelial Transport

Around 600,000 people in the United States alone have polycystic kidney disease (PKD) which is characterized by the formation of fluid‐filled pouch‐like structures, called cysts. These cysts tend to expand over the years, finally leading to kidney failure. Previously, kidney cysts were reported to be hypoxic [1]. Cyst expansion, stagnant fluid accumulation, and insufficient vascular supply can result in localized chronic ischemia‐hypoxia in cysts and as well as in normal epithelia adjacent to a cyst. We hypothesize that in normal epithelia near a cyst, the stabilization of Hypoxia Inducible Factor 1α (HIF1α), a major regulator of cellular response to hypoxia, can cause altered paracellular and transcellular transport in normal renal epithelia, transforming the normal absorptive epithelia to a secretory and leaky phenotype, leading to cyst expansion.Using cobalt chloride (CoCl2), HIF1α was stabilized in cellular nucleus of a mouse cortical collecting duct cell line (mCCD 1296 (d)), which resulted in an increased level of erythropoietin, an effector molecule of HIF1α. The mCCD monolayers have high transepithelial resistance (TER) value around 3000 Ω‐cm2 and around 95% amiloride sensitive voltage value. Equivalent current (Ieq) was calculated to compare active ion transport and FITC‐conjugated dextran molecules (3 and 70 kDa) were used to quantify transepithelial transport in terms of permeability values. MTT assay was used in order to make sure CoCl2 treated and untreated groups maintained same levels of metabolically active cells. Our results showed that TER values decreased significantly after 48 and 72 hours of HIF stabilization by CoCl2. The decrease of TER value was consistent with the increase in the permeability of large‐size 70 kDa FITC‐dextran molecules, supporting HIF stabilization actually altered paracellular transport. Western blot analysis showed HIF stabilization caused a significant decrease in the protein level of one of the major tight junction proteins, zonula occludin 1 (ZO1). Decrease in the ZO1 protein level was consistent with the decreased TER value and the increased paracellular permeability. HIF stabilization by CoCl2 for 48 hours caused almost complete loss of active sodium ion transport, and very interestingly, 72 hours of HIF stabilization caused a switch in the direction of active ion transport. Western blot analysis showed that HIF stabilization caused a decrease in the protein level of sodium‐potassium‐ATPase (Na+‐K+‐ATPase) α1 subunit, the catalytic subunit of the enzyme responsible for active sodium ion transport. Decreased level of Na+‐K+‐ATPase α1 subunit due to HIF stabilization was consistent with the loss of active sodium ion transport. Thus, HIF stabilization by CoCl2 can affect Na+‐K+‐ATPase α1 subunit and ZO1 protein levels, leading to alterations of transcellular and paracellular transport through normal kidney epithelial monolayers. Thus, our results indicate that localized ischemia‐hypoxia can transform a normal absorptive epithelia to a secretory and leaky epithelia and may contribute to cyst expansion.Support or Funding InformationThis research was supported by the Dr. John Vitullos Pilot and Bridge Funding Program at the Center for Gene Regulation in Health and Disease (GRHD), NIH DK092716 award (both to AR) and Cleveland State University Dissertation Research Award to SN.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Oppositely directed actions of different doses of arginine-vasotocin and 1-deamino-arginine-vasotocin on sodium ion transport in skin of the frog Rana temporaria

Active transport of sodium ions across the isolated abdominal skin of the frog Rana temporaria after application of arginine-vasotocin (AVT) and 1-deamino-arginine-vasotocin (1dAVT) was studied by measurement of the short-circuit current (SCC). The maximal increase in the SCC values (26 and 19 microA/cm2) was observed after addition of 10 nM AVT or 100 nM 1dAVT, respectively, to the frog skin basal surface. An increase of concentration of AVT to 100 nM and of IdAVT to 1 microM terminated the sodium transport in the frog skin. A preliminary addition of an antagonist of arginine-vasopressin V1a-receptors to the Ringer's solution at the frog skin basal surface led to a rise in the SCC values in response to administration of ineffective doses of AVT or 1dAVT. V2-receptor antagonists did not affect the frog skin reaction to administration of these doses of AVT or IdAVT.

Hans Henriksen Ussing. 30 December 1911 — 22 December 2000

Hans Ussing was born on 30 December 1911 at Sorø Academy in Denmark, where his father Dr Henrik Ussing was a lecturer and, as historian, a leading Danish folklorist. After his doctoral thesis in marine biology, Hans Ussing came to August Krogh's laboratory, where he studied protein turnover by using deuterium-labelled amino acids. After World War II, when radioactive isotopes of light elements became available for biological research, Ussing pioneered the development of epithelial physiology by introducing new concepts and theoretical tools, such as unidirectional fluxes, exchange diffusion, the flux-ratio equation, the shortcircuiting technique, solvent drag, anomalous solvent drag and the pre-steady-state flux ratio theorem. In studies on frog skin, combining electrophysiology and radioactive tracer technology, he provided the first unambiguous demonstration of active transport of sodium ions. His two-membrane hypothesis of active transport by frog skin initiated studies of epithelial transport at the cellular level in other organs and of the mechanisms of action of hormones and drugs. His discovery of paracellular ion transports bridged the physiology of high-resistance and low-resistance epithelia. With the Na + recirculation theory of isotonic transport he continued his studies of epithelial physiology until shortly before his death. Ussing's scientific research provided analytical methods and new insights of general applicability for the study of absorbing and secreting epithelia—of equal importance to biology and medicine. Hans Ussing died on 22 December 2000 after a short illness.

Mechanisms of pulmonary edema clearance during acute hypoxemic respiratory failure: role of the Na,K-ATPase.

Pulmonary edema is the hallmark of acute respiratory distress syndrome. It occurs when the permeability of the alveolar-capillary barrier is increased, causing alveolar flooding and impaired gas exchange. The mechanisms of alveolar fluid resorption are different from those of alveolar edema formation. Alveolar fluid resorption into the vessels is brought about mainly by active transport of sodium ions (Na+) out of the alveolar spaces with water following the osmotic gradient. Na+ transport across the alveolar epithelium, and thus alveolar fluid resorption, is regulated by apical Na+ channels, the basolateral sodium potassium-adenosine triphosphatase (Na,K-ATPase), and possibly chloride channels. The Na,K-ATPase has been localized to the alveolar epithelium and the importance of its role in contributing to lung edema clearance has been demonstrated. In models of lung injury, several reports have shown that catecholamines such as isoproterenol and dopamine up-regulate Na+ channels and the Na,K-ATPase giving rise to increased alveolar fluid resorption. Although recombinant gene technology is not yet a therapeutic option for the treatment of pulmonary edema, several experimental studies have reported that overexpression of Na,K-ATPase genes causes increased fluid resorption during hyperoxic lung injury. There is significant evidence that fluid clearance is impaired in patients with lung injury. Therapeutic strategies aimed at increasing the ability of alveolar epithelium to resorb the edema should lead to benefits for patients with acute respiratory distress syndrome.

Lung Lining Liquid – The Hidden Depths

The volume of liquid which lines the lumen of the lung is small and thus exists as an extremely thin layer. Nevertheless, it appears to have important functions which include contributing to the mechanical and chemical defences of the pulmonary epithelium and providing the correct ionic environment for surfactant function. Energy is expended by the epithelium to prevent excess liquid accumulating, a process involving active transport of sodium ions out of the lumen. Precisely how lung lining liquid (LLL) is formed and how its volume is controlled remain uncertain. Control of volume is important to prevent excess liquid interfering with gas exchange: yet too little liquid would inhibit efficient functioning of the airway ciliary escalator and thus removal of mucus, debris and pathogens. The chemical content of LLL is also controlled by the epithelium. LLL pH is acidic which favours calcium ionisation, important for surfactant function. Glucose is removed from LLL so decreasing the likelihood of bacterial growth and reducing extra-cellular glycosylation of proteins such as immunoglobulins and surfactant apoproteins which would destroy their function. LLL is rich in antioxidant species, protecting the lung from atmospheric oxygen and free radicals formed by inflammation. The physico-chemical properties of water itself may be important when the lining layer is very thin. The structure of water can be semi-crystalline under some circumstances which could impart another role to LLL – that of contributing to the structural integrity of the surfactant/LLL/epithelial complex.

Cell volume regulation in Claviceps fusiformis. An animal-type Na,K-ATPase operating in a fungus?

Claviceps fusiformis, an ergot pathogen of pearl millet, was equilibrated in media without and with 0.1 mM ouabain (g-strophantin). Highly significant effects of ouabain on the contents of sodium ions and water were found, suggesting that an active transport of sodium ions, similar to that in animal cells, regulates the cell volume of the fungus. The basic role of the sodium pump appears to be to decrease larger cell volumes and to increase the small ones; for this purpose it can even reverse its direction.

Increase of lung sodium-potassium-ATPase activity during recovery from high-permeability pulmonary edema.

Previous studies have suggested that recovery from pulmonary edema may be dependent on active sodium ion transport. Most of the data supporting this concept came from work done in isolated type II cells, isolated lung preparations, or in models of alveolar flooding. There is a limited amount of information regarding the role of active sodium ion transport in vivo. Furthermore, most of this information was obtained in one model of pulmonary edema, the hyperoxic lung injury model. The purpose of these experiments was then to measure the activity of the sodium-potassium-adenosinetriphosphatase (Na(+)-K(+)-ATPase), the active component of the sodium transport process and an indirect marker of active sodium transport, during recovery from thiourea-induced pulmonary edema in rats. Na(+)-K(+)-ATPase activity was significantly increased during recovery from lung edema. This increase could not be accounted for by the Na(+)-K(+)-ATPase activity present in inflammatory cells recruited in the lung by the injury process or by a direct impact of thiourea on the enzyme. Alveolar flooding, induced by instillation of a protein-containing solution into the airways of ventilated rats also increased the activity of Na(+)-K(+)-ATPase, suggesting that activation of the enzyme is probably secondary to either the presence of edema or the physiological consequences associated with edema. The quantity of lung Na(+)-K(+)-ATPase protein was also elevated during edema resolution, indicating that augmented synthesis of this enzyme underlies the increased enzyme activity observed. The quantity of Na(+)-K(+)-ATPase protein in alveolar type II cells was also significantly enhanced during recovery from edema, suggesting that these cells contribute to active sodium transport in vivo. The results of this study suggest that active sodium transport could participate in the resolution of pulmonary edema.

Effect of cyclic aromatics on sodium active transport in frog skin.

A modified glass Ussing-chamber was used to mount the skin. The electrical potential difference (PD) was measured by two 3% agar-frog Ringer's bridges. Current (i.e. short-circuit current, or ISC) was passed by Ag-AgCl electrodes placed so that current density was uniform across the skin. Ringer's solution, bathing each side of the frog skin, was stirred and aerated by gas-lift pumps. The effect of toxicants on the ISC was determined by using the 15 min prior to toxicant administration as a control period, then calculating the change in ISC during the toxicant period as a percent of the control ISC. Phenol and benzene are components of crude oil and crude oil waste. These hydrocarbons and phenanthrene were tested for their effect on frog skin. The results show that the effect of organics on sodium active transport of an epithelium is to alter the active transport of sodium ions. 5 refs., 3 figs., 1 tab.

Active Transport of Sodium Ions through Interpolymer Carboxylic Membranes

Abstract The active transport of sodium ions in the system NaOH/membrane/NaCl-HCl has been investigated. The membranes used were carboxylic membranes of the interpolymer type polyethylene/poly(methacrylic acid-co-divinylbenzene), PE/poly(MA-co-DVB), containing ∼32 wt% of poly(MA-co-DVB) and 1.8–9.6 wt% of DVB. The best transport characteristics were achieved for membranes with 1.8–3.0 DVB wt% and 0.1 M HCl. However, the observed Na+ fluxes and selectivities were rather small. To explain the observed effects, a three-layer laminate model of the carboxylic membrane has been postulated where the working membrane can be viewed as consisting of three parallel layers. The first layer, contacting alkaline solution, is regarded as an ideal carboxylic ion-exchange membrane. The second layer, contacting acidic solution, is regarded as a nonionic gel. The third layer is the intermediate part localized between the above-defined layers. It has the properties of an ion-exchange membrane to some extent but it is relatively nonselective and contains coions. It has been stated that for the most effective action of the interpolymer membrane, the thickness of the second membrane layer should be as small as possible during the whole process. This can be achieved at low HCl concentration. On the other hand, low HCl concentration cannot secure the required driving force. Moreover, the net effect of a DVB content increase in the membrane was a flux decline. Therefore, the carboxylic membranes are not effective for the active transport of Na+ ions in the system studied.

Characterization of sodium transport in Acholeplasma laidlawii B cells and in lipid vesicles containing purified A. laidlawii (Na+-Mg2+)-ATPase by using nuclear magnetic resonance spectroscopy and 22Na tracer techniques.

The active transport of sodium ions in live Acholeplasma laidlawii B cells and in lipid vesicles containing the (Na+-Mg2+)-ATPase from the plasma membrane of this microorganism was studied by 23Na nuclear magnetic resonance spectroscopic and 22Na tracer techniques, respectively. In live A. laidlawii B cells, the transport of sodium was an active process in which metabolic energy was harnessed for the extrusion of sodium ions against a concentration gradient. The process was inhibited by low temperatures and by the formation of gel state lipid in the plasma membrane of this organism. In reconstituted proteoliposomes containing the purified (Na+-Mg2+)-ATPase, the hydrolysis of ATP was accompanied by the transport of sodium ions into the lipid vesicles, and the transport process was impaired by reagents known to inhibit ATPase activity. At the normal growth temperature (37 degrees C), this transport process required a maximum of 1 mol of ATP per mol of sodium ion transported. Together, these results provide direct experimental evidence that the (Na+-Mg2+)-ATPase of the Acholeplasma laidlawii B membrane is the cation pump which maintains the low levels of intracellular sodium characteristic of this microorganism.

Effects of sodium-transport inhibitors on diuresis and mid-gut (Na + + K +)-ATPase in the tsetse fly Glossina morsitans

The effects of four inhibitors of specific sodium-transport mechanisms on diuresis in the tsetse fly Glossina morsitans, have been determined. Ouabain (1.0, 0.1 mM) and ethacrynic acid (1.0, 0.2 mM) reduced the rate of water loss, whereas amiloride (1.0 mM) and furosemide (1.0 mM) did not. The effects of ouabain, ethacrynic acid and meal size upon the anterior mid-gut (Na + + K +)-ATPase activity were also determined. For ouabain, the negative logarithm causing 50% inhibition of (Na + + K +)-ATPase (pI 50) was 6.0, whilst ethacrynic acid together with meal size did not affect the activity of this enzyme. These results show that diuresis in this insect involves the active transport of sodium ions by both electrogenic and Na + K + exchange pumps.

血管条細胞内電位に関する実験的研究 か牛中央階外側壁構造内における直流電位分布

It is important to learn the generating mechanism of the EP, and the distribution of DC potentials in the stria vascularis of the guinea pig cochlea was investigated by inserting a recording microelectrode into the scara media through the stria vascularis.The microelectrode was driven at 3 microns per second by a special hydraulic autodrive micromanipulator and the position of the electrode tip was identified by the electrophoretic dye-injection (marking method).Several positive DC steps were always recorded from various parts of the stria vascularis, and the experimental results obtained by marking methods suggested that the source of these positive DC steps was the intracellular potential of the strial cell. The data substantiated that the intracellular potentials of the strial cells are positive, whereas it is common knowledge that the intracellular potential should be negative. In most cases, 3 steps of the positive DC potential could be recorded corresponding to the layers of the strial cells. It must be presumed that the positive DC steps were recorded at the moment of pushing through the strial cell membrane by a recording microelectrode, and the largest 3rd positive step arose from the marginal cell membrane facing the intermediate cell. In the marginal cells, the positivity of the intracellular DC potential reached as high as +80mV, which is still a few millivolts less than the positivity of the EP.It has been well known that the negative potential is generally recorded from inside the cell, as the membrane is much more permeable to K-ions than to Na-ions are extruded from inside the cell by active sodium ion transport. Suppose that the positive component of the EP is the positive secretion potential generated by the electrogenic potassium ion pump located in the strial cell membrane as having been postulated by Kuijpers, it could be assumed that the strial cell membrane is selectively permeable to Na-ions and not to K-ions. On this assumption, it might be expected that the positive intracellular DC potentials in the strial cells are composed of both the sodium ion concentration gradient between the interior and the exterior of the strial cell and the electrogenic active transport of K-ions in the strial cell membrane.

Absorption of Fluid by the Midgut of Rhodnius

ABSTRACT Isolated midguts of 5th-instar Rhodnius prolixus will transport fluid from the lumen that is close to iso-osmotic with the luminal contents. The transported fluid contains sodium and chloride ions as its major constituents. Fluid transport can be attributed to active transport of sodium ions from the lumen. The haemolymph side of the epithelium, towards which transport is directed, is at a potential positive with respect to the lumen; this potential difference is greatly increased if the lumen contains only impermeant anions, and the rate of fluid transport is strongly dependent on the concentration of sodium ions in the luminal fluid. The rate of fluid transport is increased approximately six times by treatment with 5-hydroxytryptamine (2 × 10−7 M) or cyclic AMP (2 × 10−3 M). The transepithelial potential is increased by such treatment but the major effects are on the short-circuit current, which increases by about five times, and on the electrical resistance of the epithelium, which falls to about a quarter of its earlier value.

Cellular localization of Na+ K+-dependent ATPase and of the enzymes of sulpholipid metabolism in rabbit kidney.

In mammals, cerebroside 3-sulphate is known to occur preferentially in brain and kidney (1). Although considerable information is available on the metabolism of cerebroside 3-sulphate in animal tissues (2,3,4,5), their role in mammalian metabolism is still speculative. There are several lines of evidence to indicate that in myelin membrane cerebroside 3-sulphate is linked to the basic myelin proteins by ionic interactions and this sulpholipid probably acts as a structural stabilizer for the myelin sheaths of the nerve cell (6,7). Further, the occurence of cerebroside 3-sulphate in tissues rich in Na+, K+-dependent ATPase involved in active sodium ion transport has led Karlsson et al.(8,9) to suggest that cerebroside 3-sulphate is someway related to the transport of sodium ions.KeywordsBasic Myelin ProteinAdenosine TriphosphataseRabbit KidneyDependent ATPaseFers SodiumThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Fluid secretion by the malpighian tubules of the tsetse fly Glossina morsitans: the effects of ouabain, ethacrynic acid and amiloride.

The effects of three inhibitors of sodium transport on the secretion of fluid by the Malpighian tubules of Glossina morsitans have been observed. The cardiac glycoside, ouabain, affects neither the rate of secretion nor the sodium concentration of the fluid secreted when isolated tubules are bathed by solutions containing a range of sodium and potassium concentrations. Secretion is inhibited, however, by ethacrynic acid and amiloride. The results confirm that fluid secretion by the Malpighian tubules of this insect is dependent on the active transport of sodium ions and show that Na+/k+ exchange pumps are not involved in this process.

Quantitative relationship between active sodium transport, expansion of endoplasmic reticulum and specialized vacuoles ("scalloped sacs") in the outermost living cell layer of the frog skin epithelium (Rana temporaria)

When an isolated frog skin (Rana temporaria) is exposed to a hydrostatic pressure difference between inside and outside bathing solutions (inside pressure higher than outside) of 20-50 cm of H2O and if under these conditions the skin is short-circuited electrically, small "vacuoles" appear light-microscopically in the outermost living cell layer in the epithelium. The number of such "vacuoles" shows a linear dependency on the rate of active sodium transport as measured by the short-circuit current. Electron-microscopically, the "vacuoles" are interpreted as previously undescribed organelles, the "scalloped sacs" which are about 0.5 mu in diameter, with a wrinkled surface and bounded by a unit membrane. This organelle is in intimate contact with sacs and tubules of smooth endoplasmic reticulum. The observed increase in the number of scalloped sacs usually is accompanied by a significant expansion of the whole system of endoplasmic reticulum. Some of the "vacuoles" seen light-microscopically must indeed be expanded cisternae of endoplasmic reticulum. The findings are discussed in light of the possibility that the scalloped sacs and the endoplasmic reticulum may be involved in active transport of sodium ions.

Current-fluctuation noise in toad urinary bladder during active transport of sodium ions

The voltage-fluctuation noise spectrum appearing across the excised urinary bladder of the female Dominican toad Bufo marinus during active transport of sodium is identified as arising from random current fluctuations in the sodium-ion active transport current. The noise is observed to disappear with the sodium-ion active transport blocked when the bladder is bathed in choline Ringer solution. The noise magnitude with the bladder open-circuited and with the bladder short-circuited is proportional to the sodium-ion active transport current. The open-circuit active transport current is taken to be equal to the Ohmic current through the bladder resistance at the resting potential. The short-circuit active transport current is taken to be equal to the external current that must be injected into the bladder to drive the transbladder potential to zero. The bladder resistance is determined from the measurement of the frequency-dependent impedance of the bladder. Interpretation of the voltage-fluctuation spectral density according to the theoretical expressions for current-fluctuation noise indicates that the fluctuations are produced by the correlated movement through the bladder of a large number of sodium ions, carrying approximately 5000 electronic charges in a time of 0.02 sec.

Regulation of protein phosphorylation and sodium transport in toad bladder.

It is well established that active sodium-ion transport and water flow across isolated toad bladder are increased by antidiuretic hormone (ADH) and by cAMP. These agents were also observed in previous studies to cause changes in the amount of radioactive phosphate in a specific protein in the toad bladder. This protein, found by SDS-polyacrylamide gel electrophoresis of toad bladder epithelial preparations, had an apparent molecular weight of 49,000 daltons. In the present study, a correlation was found between the ability of a variety of substances to affect the amount of radioactive phosphate in this 40,000-dalton protein and their ability to alter the rate of sodium transport. Thus several agents (ADH, cAMP, theophylline, adenine, prostaglandin E1, and Mn Cl-2) caused a decrease in the amount of radioactive phosphate in the 49,000-dalton protein and also stimulated active sodium transport across the bladder. Conversely, ZnCl-2 produced an increase in the amount of radioactive phosphate in this protein and an inhibition of sodium transport. With each of these agents, the time-course of change in phosphorylation of this protein was, in general, similar to that for sodium transport. A second phosphoprotein, with an apparent molecular weight of about 42,000 daltons, showed changes in parallel with, but less extensive than, those observed in the 49,000 dalton protein. There was no consistent relationship between changes in level of phosphorylation of either in the 49,000- or 42,000- dalton protein and changes in osmotic water permeability. The results are compatible with the possibility that regulation by ADH and by cAMP of sodium transport in the toad bladder epithelium may be mediated through regulation of the amount of phosphate in a specific protein.

On facilitation of transmitter release at the toad neuromuscular junction.

1. The time dependence of the increase in amplitude (facilitation) of a second end-plate potential (e.p.p.) elicited within 10-100 msec of a preceding e.p.p. was examined at neuromuscular junctions in sartorius muscles of toads. Facilitation was defined by two characteristics, initial facilitation and the time constant of its exponential decay.2. The time constant of decay of facilitation was longer at lower temperatures and the Q(10) was 4.3 in the range 10-25 degrees C. There was no significant effect of temperature on initial facilitation.3. Ouabain (10(-4)-10(-3)M), lithium substitution for sodium, sodium azide (5 mM) and N-ethylmaleimide (NEM, 0.1 mM) initially had no effect on initial facilitation or the decay of facilitation. After some time, they all caused a longer time constant of decay of facilitation and a depression of initial facilitation.4. It was concluded that the decay of facilitation is not directly dependent on active transport of sodium ions, calcium efflux, ATP-dependent movements of calcium or mitochondrial uptake of calcium following an action potential.5. Ouabain, lithium, sodium azide, and NEM all caused an increase in transmitter release. This effect, and the late effects on facilitation, were thought to be due to an increase in intracellular calcium concentration in nerve terminals.6. No relationship was found between the quantal content of e.p.p.s (range, 0.8-100) and initial facilitation, or the time constant of decay of facilitation.7. Substitution of strontium for calcium ions caused a marked prolongation of the time constant of decay of facilitation, and a depression of initial facilitation.8. The results were consistent with the hypothesis that the time constant of decay of facilitation is related to the rate of disappearance of an ;active' complex of calcium (CaA) which, of itself, is not sufficient for transmitter release. It is suggested that an action potential produces CaA which decays with the time constant of facilitation and CaS, a short-life complex of calcium which decays with the time constant of the phasic release of transmitter. The release of transmitter is proportional to some function of [CaA] and [CaS].

The interactions of potassium, sodium and strophanthidin during active transport of sodium ions in frog muscle cells

1. 1. After the addition of 1·10 −5 M strophanthidin to a Na +-free medium, about 15 min are required for the K +-stimulated Na + efflux from muscle fibers to diminish. The strophanthidin-sensitive Na +-stimulated Na + efflux is diminished immediately after the addition of 1·10 −5 M strophanthidin. 2. 2. The half-time of inhibition by 5·10 −5 M strophanthidin is 30 min in Na +-free, 5 mM K + Ringer solution, 16 min in 80 mM Na +, 5 mM K + Ringer solution and 9 min in K +-free, 105 mM Na + Ringer solution. 3. 3. The quantitative findings suggest that strophanthidin molecules react with two different membrane-located catalytic particles that are interconvertible and in equilibrium with one another. The equilibrium is sensitive to external K + and Na +. 4. 4. K +-activated Na + extrusion in Na +-free media follows Michaelis-Menten kinetics in the absence of strophanthidin and in the presence of submaximally inhibiting concentrations of strophanthidin. 5. 5. The maximal stimulation of strophanthidin-sensitive Na + loss by K + is depressed by strophanthidin. The non-competitive kinetics observed indicate that K + and strophanthidin molecules react at different membrane sites. Also, the kinetics indicate that one strophanthidin molecule is bound per one functional catalytic site. 6. 6. In the presence of external Na +, activation of Na + extrusion by external K + follows kinetics with a marked sigmoidal character. The kinetics, under these conditions, remain sigmoidal when fixed submaximal concentrations of strophanthidin are present in the medium. A synergistic action of external Na + and strophanthidin is apparent. 7. 7. The activation of energy of K +-activated Na + extrusion in muscle fibers is 22 kcal/mole and that for strophanthidin-sensitive Na +-activated Na + efflux is 10.2 kcal/mole. 8. 8. Experimental findings are interpreted on the basis of an allosteric model in which strophanthidin molecules react with two different forms of a membrane transport enzyme.