Pathophysiology of cestode infections: Effect of Hymenolepis diminuta on oxygen tensions,pH and gastrointestinal function

Studies on the normal and parasitized rat intestine were used to investigate the effect of the tapeworm, Hymenolepis diminuta, on in vivo intestinal lumenal oxygen tensions, acid-base balance and mucosal absorption and accumulation of fluid and glucose.The lumenal bulk aqueous phase is considerable, well mixed and aerobic with an oxygen tension of 40–50 mm Hg. Neither the unstirred layers adjacent to the brush border membrane nor the area adjacent to the mucosa (“paramucosal lumen”) are significant barriers to the diffusion of oxygen from the blood to the intestinal lumen. In the uninfected distal ileum and colon anoxic conditions may occur in the central lumen, but, in the parasitized intestine fluid absorption is reduced and anoxic conditions do not occur. Increased H⁺ ion concentration in the parasitized intestine plays a role in increasing the availability of oxygen to intestinal helminths. Concomitant with the lower pH, the pCO₂ in the lumen of the parasitized intestine was twice as high as that found in normal animals. The total CO₂ in the parasitized intestine steadily decreased over a 3-h perfusion period, while in the normal intestine the total CO₂ content increased after an initial fall during the first 30 min of perfusion. When the worms were removed, the ability of the intestine to restore normal acid-base balance was restored. Glucose and fluid absorption in both the infected and uninfected intestine were reduced by an increase in H⁺ ion concentration; both parameters were lower in the parasitized intestine than in the normal animals. Low pH increased fluid and glucose transport by H. diminuta.While the dry weights of both the parasitized and uninfected total small intestine and of the intestinal mucosa were the same, the wet weights were considerably different, indicating defective fluid balance in the infected intestine. Accumulation of glucose by the parasitized mucosa was greater than in control animals and decreased with an increase in H⁺ ion concentration. The glucose transport system in the parasitized gut was therefore affected at two levels, one at the brush border, where transport into the mucosa was decreased by lowering the pH, and secondly at the level of the basal and lateral membranes, where transport out of the mucosal tissue into the circulatory system was also reduced.The above results are discussed in terms of current widely accepted but erroneous concepts relating to the intestinal ‘microcosm’.     

Insulin action on cardiac glucose transport Studies on the role of the Na+/K+ pump

Isolated muscle cells from adult rat heart have been used to study the relationship between myocardial glucose transport and the activity of the Na⁺/K⁺ pump. ⁸⁶Rb⁺-uptake by cardiac cells was found to be linear up to 2 min with a steady-state reached by 40–60 min, and was used to monitor the activity of the Na⁺/K⁺ pump. Ouabain (10^?3 mol/I) inhibited the steady-state uptake of ⁸⁶Rb⁺ by more than 90%. Both, the ouabain-sensitive and ouabain-insensitive ⁸⁶Rb⁺-uptake by cardiac cells were found to be unaffected by insulin treatment under conditions where a significant stimulation of 3-O-methylglucose transport occurred. ⁸⁶Rb⁺-uptake was markedly reduced by the presence of calcium and/or magnesium, but remained unresponsive towards insulin treatment. Inhibition of the Na⁺/K⁺ pump activity by ouabain and a concomitant shift in the intracellular Na⁺:K⁺ ratio did not affect basal or insulin stimulated rates of 3-O-methylglucose transport in cardiac myocytes. The data argue against a functional relationship between the myocardial Na⁺/K⁺ pump and the glucose transport system.     

Multiplicity of carriers for free glucalogues in hamster small intestine

The transport of free glucalogues across the mucosal pole of hamster small intestine is carried out by (at least) two transport systems, one of which is common to glucose, galactose, 6-deoxyglucose and 3-methylglucose, whereas the order is essentially specific for glucose and galactose, among the monosaccharides tested. This was shown by:

1. 1. The dependence of the unidirectional medium-mucosa flux on the substrate concentration which is described by a single horizontal hyperbola in the cases of 6-deoxyglucose and of 3-O-methylglucose, but by the sum of two such hyperbolas in the cases of both glucose and galactose.

2. 2. The mutual inhibition among glucalogues and the inhibition by arbutin, which are best described by the existence of two carriers, as given above:

3. 3. The difference in the absorption of glucose and galactose in baby hamsters as compared with adult hamsters.

Both systems are Na⁺-dependent and (to a different degree) phlorizin-sensitive. Neither is identical with the fructose transport system.     

Regulation of the intracellular potassium concentration in Escherichia coli B 525

The mutant Escherichia coli B 525 requires histidine, leucine and methionine and an elevated extracellular K⁺ concentration for growth, and is unable to retain K⁺ tightly inside the cells when incubated in media supplemented with glucose, arabinose, galactose or lactose as the sole energy and carbon source. The loss of K⁺ from the cells of B 525 can be prevented by adding histidine and leucine, which react specifically and only in combination. In media supplemented with glycerol as the substrate, with glucose and NH₄⁺, or with glucose under anaerobic conditions, a stationary level of K⁺ inside the cells can be obtained without the addition of histidine-leucine.On the addition of ribose to glycerol-adapted cells of B 525 preincubated in glycerol media, the intracellular K⁺ decreased immediately and markedly. This decrease can be overcome by the addition of histidine-leucine.     

Sarcolemmal glucose transport in Ca2+-tolerant myocytes from adult rat heart. Calcium dependence of insulin action

Cardiac myocytes were isolated from adult rat ventricles by a method which preserves their functional integrity, including long survival in physiological concentrations of Ca²⁺. Sarcolemmal glucose transport was assessed by measuring linear initial uptake rates of the nonmetabolized glucose analog3-O-methyl-d-glucose. Transport was saturable and showed competition byd-glucose and other features of chemical and stereo-selectivity. Transport was stimulated by insulin in a dose-dependent manner, resulting in an almost 5-fold increase inV_max, with little change inK_m. Stimulation of 3-methylglucose transport by insulin was largely Ca²⁺ -dependent. Omission of Ca²⁺ from the incubation medium caused a minor rise in basal 3-methylglucose uptake but the insulin-stimulated rise inV_max was only 30%. The Ca²⁺ antagonist D600 also antagonized stimulation of hexose transport by insulin. In all the above respects, 3-methylglucose transport in myocytes is identical to that in intact heart muscle. In addition, the decrease in insulin response by Ca²⁺ emission was partially reversed by subsequent return to a Ca²⁺ -containing medium. ATP levels remained stable in the absence of Ca²⁺, showing that the Ca²⁺ dependence did not reflect nonspecific cell damage.     

The inhibition of hexose transport and metabolism by small amounts of adenosine 5′-triphosphate in isolated rat adipocytes

The effects of ATP on glucose transport and metabolism were studied in rat adipocytes. Over a concentration range of 10–250 μm, ATP was found to inhibit several aspects of adipocyte glucose metabolism, particularly when stimulated by insulin. Much of the effect of ATP on glucose metabolism appeared related to impairment of glucose transport, reflected by inhibition of both basal and insulin-stimulated rates of 3-O-methylglucose transport. ATP inhibited the V of insulin-stimulated 3-O-methylglucose transport, but had no effect on the K_m. The inhibitory effects of ATP were much less apparent when cells were preincubated with insulin, suggesting that ATP inhibited only the components of hexose transport not yet activated by the hormone. At very high medium glucose concentrations, where transport was no longer rate limiting for metabolism, there was no inhibition of glucose oxidation by 250 μm ATP. However, when hexose transport was blocked with cytochalasin B (50 μm), a small inhibitory effect of ATP persisted on basal and insulin-stimulated glucose and fructose oxidation, suggesting that intracellular metabolism was impaired. The mechanism of the intracellular effect did not appear to be caused by uptake of exogenous ATP. These studies provide further evidence that energy metabolism may play an important role in the regulation of facilitated glucose transport.     

Solute Transport Process in Intestinal Epithelial Cells

In rat small intestine, the active transport of organic solutes results in significant depolarization of the membrane potential measured in an epithelial cell with respect to a grounded mucosal solution and in an increase in the transepithelial potential difference. According to the analysis with an equivalent circuit model for the epithelium, the changes in emf's of mucosal and serosal membranes induced by active solute transport were calculated using the measured conductive parameters. The result indicates that the mucosal cell membrane depolarizes while the serosal cell membrane remarkably hyperpolarizes on the active solute transport. Corresponding results are derived from the calculations of emf's in a variety of intestines, using the data that have hitherto been reported. The hyperpolarization of serosal membrane induced by the active solute transport might be ascribed to activation of the serosal electrogenic sodium pump. In an attempt to determine the causative factors in mucosal membrane depolarization during active solute transport, cell water contents and ion concentrations were measured. The cell water content remarkably increased and, at the same time, intracellular monovalent ion concentrations significantly decreased with glucose transport. Net gain of glucose within the cell was estimated from the restraint of osmotic balance between intracellular and extracellular fluids. In contrast to the apparent decreases in intracellular Na⁺ and K⁺ concentrations, significant gains of Na⁺ and K⁺ occurred with glucose transport. The quantitative relationships among net gains of Na⁺, K⁺ and glucose during active glucose transport suggest that the coupling ratio between glucose and Na⁺ entry by the carrier mechanism on the mucosal membrane is approximately 1:1 and the coupling ratio between Na⁺-efflux and K⁺-influx of the serosal electrogenic sodium pump is approximately 4:3 in rat small intestine. In addition to the electrogenic ternary complex inflow across the mucosal cell membrane, the decreases in intracellular monovalent ion concentrations, the temporary formation of an osmotic pressure gradient across the cell membrane and the streaming potential induced by water inflow through negatively charged pores of the cell membrane in the course of an active solute transport in intestinal epithelial cells are apparently all possible causes of mucosal membrane depolarization.     

Transport of glucose and galactose in kidney-cortex cells

1. The aerobic transport of d-glucose and d-galactose in rabbit kidney tissue at 25° was studied. 2. In slices forming glucose from added substrates an accumulation of glucose against its concentration gradient was found. The apparent ratio of intracellular ([S]_i) and extracellular ([S]_o) glucose concentrations was increased by 0·4mm-phlorrhizin and 0·3mm-ouabain. 3. Slices and isolated renal tubules actively accumulated glucose from the saline; the apparent [S]_i/[S]_o fell below 1·0 only at [S]_o higher than 0·5mm. 4. The rate of glucose oxidation by slices was characterized by the following parameters: K_m 1·16mm; V_max. 4·5μmoles/g. wet wt./hr. 5. The active accumulation of glucose from the saline was decreased by 0·1mm-2,4-dinitrophenol, 0·4mm-phlorrhizin and by the absence of external Na⁺. 6. The kinetic parameters of galactose entry into the cells were: K_m 1·5mm; V_max 10μmoles/g. wet wt./hr. 7. The efflux kinetics from slices indicated two intracellular compartments for d-galactose. The galactose efflux was greatly diminished at 0°, was inhibited by 0·4mm-phlorrhizin, but was insensitive to ouabain. 8. The following mechanism of glucose and galactose transport in renal tubular cells is suggested: (a) at the tubular membrane, these sugars are actively transported into the cells by a metabolically- and Na⁺-dependent phlorrhizin-sensitive mechanism; (b) at the basal cell membrane, these sugars are transported in accordance with their concentration gradient by a phlorrhizin-sensitive Na⁺-independent facilitated diffusion. The steady-state intracellular sugar concentration is determined by the kinetic parameters of active entry, passive outflow and intracellular utilization.     

Glucose transport across the intestinal wall of the freshwater snail Biomphalaria glabrata

• 1.1. Isolated midguts of the freshwater snail Biomphalaria glabrata were mounted in an incubation chamber in saline containing 2 mM glucose and perfused with the same solution. External and internal media were continuously gassed with carbogen gas (95% O₂, 5% CO₂). In order to measure the flux rates of glucose [¹⁴C]glucose was applied in the perfusion medium or in the incubation medium. Net fluxes of glucose were calculated as the differences between unidirectional in- and effluxes.
• 2.2. A directed net flux from the mucosal to the serosal side of the intestine was demonstrated (mucosal to serosal = 50 ± 10 nmol cm⁻²hr⁻¹(N = 6) serosal to mucosal 7 ± 1 nmol cm⁻²hr⁻¹ (N = 6), net flux = 43 nmol cm⁻²hr⁻¹).r
• 3.3. The active transport of glucose was reduced by the presence of metabolic inhibitors, cyanide (1 mM) and dinitrophenol (1 mM) on the mucosal as well as on the serosal side. Ouabain (1 mM) inhibited the transport rate only when it was added on the serosal side. Amiloride (1 mM) had no effect on the transport rate whether added on the mucosal or on the serosal side.
• 4.4. Inhibition of glucose transport by oubain, a specific inhibitor of Na⁺/K⁺-ATPase, suggests that glucose transport is secondary active and coupled to Na⁺-transport.

     

X-ray microanalysis of elements in frozen-hydrated sections of an electrogenic K+ transport system: The posterior midgut of tobacco hornworm (Manduca sexta) in vivo andin vitro

Summary The lepidopteran midgut is a model for the oxygendependent, electrogenic K⁺ transport found in both alimentary and sensory tissues of many economically important insects. Structural and biochemical evidence places the K⁺ pump on the portasome-studded apical plasma membrane which borders the extracellular goblet cavity. However, electrochemical evidence implies that the goblet cell K⁺ concentration is less than 50mm. We used electron probe X-ray microanalysis of frozenhydrated cryosections to measure the concentration of Na, Mg, P, S, Cl, K, Ca and H₂O in several subcellular sites in the larval midgut ofManduca sexta under several experimental regimes. Na is undetectable at any site. K is at least 100mm in the cytoplasm of all cells. Typicalin vivo values (mm) for K were: blood, 25; goblet and columnar cytoplasm, 120; goblet cavity, 190; and gut lumen, 180. The high K concentration in the apically located goblet cavity declined by 100mm under anoxia. Both cavity and gut fluid are Cl deficient, but fixed negative charges may be present in the cavity. We conclude that the K⁺ pump is sited on the goblet cell apical membrane and that K⁺ follows a nonmixing pathway via only part of the goblet cell cytoplasm. The cavity appears to be electrically isolated in alimentary tissues, as it is in sensory sensilla, thereby allowing a PD exceeding 180 mV (lumen positive) to develop across the apical plasma membrane. This PD appears to couple K⁺ pump energy to nutrient absorption and pH regulation.     

Calcium inhibition of potassium absorption in corn roots

Calcium (or magnesium) sulfate or chloride was found to inhibit energy dependent potassium transport in excised corn roots. This Ca²⁺ inhibition of K⁺ transport was most pronounced during the initial phases of transport. As the absorption periods were lengthened the effect of Ca²⁺ gradually changed from an inhibition to a typical promotion (after about 30-45 mins) of K⁺ transport. Kinetic analysis indicated the inhibition to be of a non-competitive nature.

Identical experiments with excised barley roots showed that CaSO₄ had no effect on K⁺ absorption whereas CaCl₂ had a typical stimulatory effect on K⁺ absorption. Kinetic analysis indicated that both corn and barley have efficient K⁺ transporting systems but barley roots are approximately 5 times more active (on a fr wt basis) than corn roots.

These results illustrate the hazards involved in applying results obtained with 1 (or even several) plant species to all species.

     

Relationship between coupled Na+−K+−Cl− transport and water absorption across the seawater eel intestine

Summary Simultaneous measurements of net ion and water fluxes were made in the stripped intestine of the seawater eel, and the relationship between Na⁺, K⁺, Cl^– and water transport were examined in the presence of mucosal KCl and serosal NaCl Ringer (standard condition). When Cl^– was removed from both sides of the intestine, net K⁺ flux from mucosa to serosa was reduced, accompanied by complete blockage of water absorption. Since it has been shown that net Cl^– and water fluxes depend on K⁺ transport under the standard condition (Ando 1983), the interdependence of K⁺ and Cl^– transport suggests the existence of a coupled KCl transport system, while the parallelism between the net Cl^– and water fluxes suggests that water absorption is linked to the coupled KCl transport. The coupled KCl and water transport were inhibited by treatment with ouabain or with Na⁺-free Ringer solutions, suggesting the existence of a Na⁺-dependent KCl transport system and linkage of water absorption to the coupled Na⁺–K⁺–Cl^– transport. Since ouabain blocked the active Na⁺–K⁺–Cl^– transport almost completely, the permeability coefficients for K⁺ and Na⁺ through the paracellular shunt pathway were estimated as P_K=0.076 and P_Na=0.058 cm/h, and P_Cl was calculated as 0.005 cm/h. Although Na⁺-independent K⁺ and Cl^tt- fluxes were observed again in the present study, these fluxes were not inhibited by CN^–, ouabain or diuretics, and evoked even after blocking the Na⁺–K⁺–Cl^– transport completely with ouabain. These results indicate that the Na⁺-independent K⁺ and Cl^– fluxes are distinct from the active Na⁺–K⁺–Cl^– transport and are not themselves active.     

Transport of protons and potassium ions across the membranes of bacteria <Emphasis Type="Italic">Enterococcus hirae</Emphasis> dependent on ATP and nicotinamide adenine dinucleotides

The transport of protons and potassium ions across the membranes of the bacteria Enterococcus hirae growing in an alkaline medium (pH 8.0) or under experimental conditions (pH 7.5) during glucose fermentation accomplished by a KtrI system of absorption of potassium ions, which can interact with F₀F₁-ATPase to form at H⁺-K⁺-pump, has been studied. It was found on cells with a high membrane permeability that the administration of nicotinamide adenine dinucleotides results in the potassium absorption which is insensitive to the inhibitor of F₀F₁-ATPase N,N′-dicyclohexylcarbodiimide. It is assumed that, along with the KtrI system which interacts with F₀F₁-ATPase, a separate KtrI or another K⁺ absorption system operates in these bacteria under particular conditions, which is dependent on NAD⁺ +NADH. Presumably, these interact with this system, changing its conformational state required for the transition to the “active” form.     

Sugar transport and potassium permeability in yeast plasma membrane vesicles

Plasma membrane vesicles were isolated from homogenised yeast cells by filtration, differential centrifugation and aggregation of the mitochondrial vesicles at pH 4. As judged by biochemical, cell electrophoretic and electron microscopic criteria a pure plasma membrane vesicle preparation was obtained.The surface charge density of the plasma membrane vesicles is similar to that of intact yeast cells with an isoelectric point below pH 3. The mitochondrial vesicles have a higher negative surface charge density in the alkaline pH range. Their isoelectric point is near pH 4.5, where aggregation is maximal.The yield of vesicles sealed to K⁺ was maximal at pH 4 and accounted for about one third of the total vesicle volume.The plasma membrane vesicles demonstrate osmotic behaviour, they shrink in NaCl solutions when loosing K⁺.As in intact yeast cells the entry and exit of sugars like glucose or galactose in plasma membrane vesicles is inhibited by UO₂²⁺.Counter transport in plasma membrane vesicles with glucose and mannose and iso-counter transport with glucose suggests that a mobile carrier for sugar transport exists in the plasma membrane.After galactose pathway induction in the yeast cells and subsequent preparation of plasma membrane vesicles the uptake of galactose into the vesicles increased by almost 100% over the control value without galactose induction. This increase is explained by the formation of a specific galactose carrier in the plasma membrane.     

Does an electroneutral K<Superscript>+</Superscript>/Cl<Superscript>−</Superscript> antiport occur in cardiomyocyte during acute ischemia?

Electron probe microanalysis was applied to determine cytoplasmic elemental (K, Na, Cl) concentrations in cardiac cells of the rat (Wistar). Potassium, sodium and chlorine contents were measured in papillary muscle myocytes of the rat heart perfused by the Langendorff procedure. Ischemic depletion was created by perfusion with deeply deoxygenated Tyrode’s solution in the absence of glucose. It was found that the initial phase of acute ischemia is characterized by the potassium deficiency and the accumulation of sodium and chlorine in cardiac myocytes. It should be noted that changes in the total charge of the main intracellular cations (K⁺, Na⁺) do not compensate for the increased chlorine concentration. This result can be accounted for by the appearance of ionic (K⁺ and Cl⁻) transport coupled with the removal of lactate anions produced in cardiomyocytes during anaerobic glycolysis.     

The Cellular Transport of Magnesium in Rat Liver

The bidirectional transport of Mg in rat liver was studied using slices labeled with ²⁸Mg in a closed two-compartment system under steady-state conditions. The influx (K_bs) and efflux (K_sb) transfer coefficients governing transport between the extracellular phase and a rapidly exchanging cell fraction were 0.074 and 0.019 per min, respectively. An increased extracellular concentration of Mg⁺⁺ caused a 30% decrease in K_bs and a 31% increase in K_sb. A decreased extracellular Mg⁺⁺ had an opposite effect. At 0°C, both transfer coefficients were reduced by 65%. Increased pH and NaCN increased transport, whereas Ca⁺⁺ reduced transport. Reduced pH, altered Na⁺:K⁺ ratio, Sr⁺⁺, glucose deletion, iodoacetate, ethanol, and lactate had no significant influence. Dinitrophenol reduced K_sb but had no effect on K_bs. These data support the thesis that the intracellular concentration of Mg is in part regulated by a reciprocal change in the influx transfer coefficient and a parallel change in the efflux transfer coefficient in response to altered extracellular concentrations of Mg⁺⁺. The qualitative and quantitative similarities of Mg and Ca transport in this system suggest that Mg and Ca share a common transport mechanism which is primarily dependent upon the binding of these divalent cations to macromolecular ligands within the cell membrane or within the cell.     

Potassium activation of Na+-dependent leucine transport in brush-border membrane vesicles from rat jejunum

Na⁺-dependent leucine uptake was greater in potassium loaded brush-border membrane vesicles compared with controls. This effect was not mediated by an electrical potential difference, since it was still present in voltage-clamped conditions. Inhibition experiments indicate the same Na⁺-dependent leucine transport activity in the presence or in the absence of potassium. The affinity of sodium for the cotransporter was identical at 10 or 100 mM potassium. Leucine kinetics at different potassium concentrations showed a maximum 2.4-fold increase in V_max, while K_m was unaffected. The secondary plots of the kinetic results were not linear. This kinetic behaviour suggests that K⁺ acts as a non-essential activator of Na⁺-dependent leucine cotransport. A charge compensation of sodium-leucine influx is most probably a component of the potassium effect in the presence of valinomycin.