Active Transport Of Monovalent Cations Research Articles

Individual and combined haematotoxic effects of fumonisin B1 and T-2 mycotoxins in rabbits

Weaned rabbits were fed diets contaminated with 2mg/kg diet T-2 toxin alone, or 10mg/kg diet fumonisin B1 (FB1) alone, and both toxins in combination (2+10mg/kg, resp.), as compared to a toxin free control. Samplings were performed after 2 and 4weeks. Bodyweight of the T-2 fed group was lower after 4weeks; the liver weight increased dramatically. Red blood cell (RBC) Na+/K+ ATPase activity decreased after 4weeks in the T-2 group, it increased in the FB1 group and antagonism was found by the combined treatment. The RBC membrane fatty acid profile was modified by both toxins similarly during the entire feeding. After 4weeks T-2 alone and in combination (with FB1) was found to increase mean cell volume (MCV). The time-dependent alterations in the T-2 group were significant for MCV (increase) and the mean cell haemoglobin (increase). The active monovalent cation transport was altered by both mycotoxins. Most probably FB1 exerts its sodium pump activity modification via an altered ceramide metabolism (behenic acid decrease in the RBC membrane), while for T-2 toxin a moderate membrane disruption and enzyme (protein) synthesis inhibition was supposed (ca. 75% decrease of the sodium pump activity).

All-Russian symposium on comparative endocrinology dedicated to the memory of professor L G Leĭbson (on his 100th birthday)

To study role of glycolysis and oxidative metabolism in providing active transport of monovalent cations, isolated erythrocytes of the lamprey Lampetra fluviatlis were incubated at 20°C in the presence of various metabolic inhibitors. The active (ouabain-sensitive) K+ (86Rb) influx into erythrocytes did not change after cell incubation for 1–2 h in the absence of glucose or in the presence of 10 mM deoxy-D-glucose or 1 mM monoiodoacetate. Inhibitors of oxidative phosphorylation (antimycin A, rotenone, sodium azide, cyanide) produced a significant decrease (on average, by 74% ) in the active K+ transport in the lamprey erythrocytes. All blockers of oxidative phosphorylation produced the same degree of inhibition of the K+ transport after the cell pre-incubation with them for 30 and 60 min. In experiments with rotenone, the K+ influx was reduced statistically significantly as early as in 5 min of cell incubation and reached a maximal effect after 10–20 min. The intracellular ATP content in erythrocytes decreased by 17, 37, and 45% after 5, 10, and 20 min of cell incubation with rotenone, respectively. The active K+ transport in the lamprey erythrocytes is most likely to be closely associated with the intracellular ATP concentration. The data obtained indicate that the energy supply of the Na,K-pump in the lamprey erythrocytes is due exclusively to oxidative phosphorylation processes.

Early Enhancement of Passive Potassium Efflux from Rat Liver by Thyroid Hormone: Relation to Induction of Na,K-ATPase*

The effect of thyroid hormone treatment on the passive efflux of 42K+ from rat liver slices was studied to determine whether an increase in permeability might play a role in the known enhancement of active monovalent cation transport and Na,K-ATPase activity induced by thyroid hormone. Initial studies showed that the sc injection of 50 micrograms T3/100 g BW on 3 alternate days resulted in an 82% increase in the 42K+ passive efflux rate constant in liver slices and a 23% increase in the Na,K-ATPase activity of liver homogenates. Subsequent investigation of the relative time courses of these two hormonal effects showed that after a single injection of T3 the rate constant for 42K+ efflux increased within 6 h and reached a plateau between 24 and 48 h. In contrast, Na,K-ATPase activity was not augmented until 24 h and continued to rise between 24 and 48 h. These results suggest that an early increase in cation permeability (with resultant cation leak) may account for the known stimulatory effect of thyroid hormone on active monovalent cation transport in rat liver. The relatively late increase in Na,K-ATPase activity might then be regarded as an adaptive cellular response to increased passive cation fluxes rather than a direct effect of thyroid hormone. The magnitude and early onset of the effect of thyroid hormone treatment on cellular 42K+ efflux raise the possibility that an increase in passive cation permeability may be a proximal event in the mediation of thyroid hormone action.

In vitro studies of skeletal muscle membranes. Effects of denervation on the macromolecular components of cation transport in red and white skeletal muscle.

The effects of denervation on the macromolecular components of active monovalent cation transport in skeletal muscle have been studied using purified sarcolemma membranes. A comparison of membrane activities of fast-twitch, slow-twitch, and mixed-fiber muscles was made to determine what role, if any, the motor nerve has in regulating this important aspect of muscle metabolism. A dramatic increase in the basal sarcolemmal Mg++ ATPase activity (three- to fourfold) was found for both major muscle types. An increase in the ouabain-inhibitable (Na+ + K+)-stimulated enzyme was also found, but the effect was substantially less (1.5- to twofold). [3H]-ouabain binding, as an index of glycoside receptor sites, also increased (two- to threefold) midway in the course of denervation. On the other hand, the phosphorylated intermediate activity, a functional component of the transport system, clearly decreased over the same time course and remained below control values for the remainder of the course. This resulted in a two- to threefold increase in the turnover number, suggesting that active transport of cations should increase dramatically with denervation. The membrane protein patterns on SDS gels were less obvious than the changes observed in the functional components. The major effects appeared after only one week and seemed to be restricted to high molecular weight membrane proteins, especially in the 100,000 to 250,000 daltons range. This effect was more prominent in slow-twitch membranes with an apparent semiquantitative decrease in stain at 240,000 daltons. In gels of membranes from fast-twitch muscles a decreased stain in the range of 100,000 to 110,000 daltons occurred, and this became more obvious with longer periods of denervation. The results suggest that considerable influence on the macromolecular components of active cation transport in skeletal muscle is exerted by the motor nerve. No appreciable difference was found in this effect when the two major types of skeletal muscle, fast-twitch and slow-twitch, were compared, suggesting that motor nerve regulation of this membrane property is qualitatively the same.

Resistance of active monovalent cation transport to pronase digestion of intact human erythrocytes

Human erythrocytes were treated with pronase under conditions which gave complete cleavage of the pronase-sensitive major protein and glycoprotein components of the membrane. 86Rb + influx of these modified cells did not differ significantly from that of untreated cells, and showed identical sensitivity to inhibition by ouabain. Pronase treatment of porous erythrocyte ghost membranes resulted in complete loss of (Na + + K +)-ATPase activity. These results indicate that the cation-transport complex is not readily accessible from the outer surface of native intact membranes.

Cation permeability induced by spermine and Polybrene in rat liver mitochondria.

The effect of the polybasic substances Polybrene and spermine on the passive and active transport of monovalent cations in mitochondria was studied. These agents were found to stimulate the low amplitude swelling of mitochondria. Volume oscillations were induced by addition of substrate in the presence of spermine. In conditions where weak oscillations were obtained without these substances, oscillations were stimulated and their frequencies increased in the presence of Polybrene and spermine. Their effects were maximal with 100–300 μmoles spermine per litre and 3–5 mg Polybrene per litre. These results are discussed in relation to an interaction of the agents studied with membrane negative charges which may be important regulators of ion transport.

Conformational model of active transport.

A model of active transport of monovalent cations in mitochondria is developed. The model is based on the coupling of electron transfer to the generation of a metastable protein conformation which in turn leads to the generation of an asymmetric surface charge, a membrane potential, and a redistribution of diffusible ions across the inner mitochondrial membrane. The ions at all times move spontaneously down an electrochemical potential gradient in this model so that there is no need to invoke the concept of an ion pump. It is shown that a wide variety of experimental facts can be rationalized in terms of the present model.

FURTHER OBSERVATIONS OF CATION‐STIMULATED STOMATAL OPENING IN ISOLATED EPIDERMIS

SummaryMeasurements of apparent osmotic pressures of guard cells in isolated epidermis of Commelina communis indicate a considerable rise (up to 26.3 bars) when K+ ions are added to the incubation medium. This rise in apparent osmotic pressure appears to be the major cause of the stomatal opening that is stimulated by K+ ions, but reduction in turgor of the epidermal cells, owing to the solutes in the incubation medium, may also contribute to the wide stomatal apertures. Histochemical tests on epidermis taken from leaves with stomata that were closed in darkness, or that were open in light, suggest that K+ accumulates in guard cells when stomata open in response to light. No movements of K+ or Na+ from epidermis to mesophyll could be detected and it is concluded that ion movements associated with changes in guard cell turgor are probably confined to the epidermis in C. communis. Ion uptake (indicated by 86Rb and 24Na) into epidermis was stimulated by light but not by a reduction in CO2 concentration, whereas stomatal apertures were affected by these environmental factors to approximately the same extent. It is tentatively concluded that active transport of monovalent cations into guard cells might explain the CO2‐independent light responses of stomata, but not their light‐independent CO2 responses.

Potassium dependent phosphatase in bovine parotid gland

Naf, K+-dependent adenosine triphosphatase (Na+, Kf-dependent ATPase) [ATP phosphohydrolase, EC 3.6.1.31 may play an important role in active transport of monovalent cations. Hydrolysis of ATP by the Na+, K+-dependent ATPase is thought to be a two-staged reaction. The first step is the Na+-dependent phosphorylation of an intermediate, and second step is the K+-dependent dephosphorylation of the intermediate. It was reported that hydrolysis of p-nitrophenyl phosphate is catalyaed by an enzyme in the membrane fraction of various tissues and it is stimulated by the presence of potassium (JUDAH, AHMED and MCLEAN, 1962; AHMED and JUDAH, 1964). It was also suggested that, since p-nitrophenyl phosphate is known to serve as a substrate for phosphoprotein phosphatase [EC 3.1.3.161, it may be assumed that the action of K+-dependent phosphatase on p-nitrophenyl phosphate is representative of the dephosphorylation process in the Na+, K+-dependent ATPase reaction (NAGAI, IZUMI and YOSHIDA, 1965). Many investigators have demonstrated the presence and discussed the biological meaning of Na +, K+-dependent ATPase in salivary glands (SCHWARZ and MATSUI, 1967). However, K+-dependent phosphatase in salivary glands has not been reported. Therefore, in the present work the presence and some properties of K+-dependent phosphatase in bovine parotid gland were investigated. Female bovine parotid gland was homogenized in 4 volumes of 0 * 32 M sucrose. After removal of debris by centrifugation at 800 g for 10 min, the supernatant was centrifuged at 15,OOOg for 20 min to obtain the mitochondrial fraction. Then, microsomal and soluble fraction were separated by centrifugation of the supernatant at 105,000 g for 60 min. Each fraction was washed once, suspended in 0.32 M sucrose and used for enzyme assay. K+-dependent phosphatase activity was determined as follows. Two ml of the reaction mixture, containing SOpmoles of Tris buffer (pH 7*8), 10 pmoles of p-nitrophenyl phosphate disodium salt, 10 pmoles of MgC12, 20 pmoles of KC1 and a certain amount of the enzymes were incubated at 25°C for 20 min. The reaction was stopped by addition of 4 ml of O-5 M NaOH and, after centrifugation, an absorbance of the supernatant was estimated at 410 rnp in a spectrophotometer. The activity of the K+-dependent phosphatase was expressed as the difference between the activity in the presence and absence of K+ in the reaction mixture. The amount of protein was determined with Folin-Ciocalteu’s reagent (LOWRY et al., 1951).

Effect of monovalent cations on the adenosinetriphosphatase of a skeletal muscle microsomal preparation

Summary 1. The possible involvement of Mg2+-dependent adenosinetriphosphatase (EC 3.6.1.4) of skeletal muscle microsomes in the process of active transport of monovalent cations was studied. 2. The adenosinetriphosphatase activity of a light microsomal preparation of rabbit skeletal muscle was stimulated by either Na+, K+, NH4+ or Rb+. Cs+ and Li+ inhibited the enzymic activity both in the presence and absenpe of stimulating cations. 3. The simultaneous presence of Na+ and K+ did not cause any additional activation beyond that obtained by either ion alone. Also, ouabain had no effect on the enzymic activity. It was, therefore, concluded that a correlation between the adenosinetriphosphatase and active transport of Na+ and K+ was not apparent. 4. Na+ and K+ were found to inhibit the adenosinetriphosphate-dependent Ca2+-binding by the light microsomes. A possible relation of Na+, K+-activated adenosinetriphosphatase to Na+, K+-inhibited Ca2+-binding was suggested.