Adenosine effect on lactate and pyruvate production in the hypoxic guineapig heart

• 1.1. It has been shown that adenosine stimulates glycolysis in some cells and this ability of adenosine was tested in the hypoxic guineapig heart.
• 2.2. Adenosine (10 μM) activated lactate production in the isolated perfused guineapig heart under conditions of normoxia but did not under hypoxia.
• 3.3. Despite this, the nucleoside favorably influenced the energy metabolism of the hypoxic heart as revealed by the better posthypoxic functional recovery (98%) compared to the control without adenosine (78%).
• 4.4. Our findings suggest a role for the glycolytic pathway in this effect of the nucleoside as long as other cardiac energy-yielding pathways are strictly aerobic.

     

Proteolytic cleavage of band 3 protein in relation to anion transport in fish (Oncorhynchus mykiss) red blood cells

• 1.1. The effects of trypsin and chymotrypsin on HCO₃⁻/Cl⁻ exchange through red blood cell membranes of humans and trout were studied.
• 2.2. To measure the anion exchange we used a right-angle light-scattering technique by applying the Jacobs-Stewart cycle in ammonium solution and the osmotiration method at constant cell volume.
• 3.3. The Cl⁻ flux in human red blood cells remained unaltered after treatment with external trypsin and chymotrypsin while in trout red blood cells the flux decreased.
• 4.4. This partial inhibition of anion transport in fish, ranging from 30 to 40%,suggest that one or several of the cleavage sites in band 3 protein, essential for anion transport function, are exposed in fish red blood cells.
• 5.5. In human red blood cells the fragments of band 3 which are affected by proteolytic digestion, retain their tertiary structure because there is no influence on anion transport.

     

Nucleoside monophosphoramidate hydrolase from rat liver: Purification and characterization

• 1.1. Adenosine 5'-phosphoramidate hydrolase of 29 kDa was isolated from rat liver cytosol.
• 2.2. It consisted of two subunits of 14 kDa.
• 3.3. It hydrolyzed nucleoside 5'-monophosphoramidates into nucleoside 5'-monophosphates and ammonia, while it did not hydrolyze adenylyl phosphoramidate, adenylyl imidodiphosphate and N-phosphorylated compounds like phosphocreatine, N^ω-phosphoarginine, 6-phospholysine and 3-phosphohistidine.
• 4.4. Divalent cations and cyclic AMP had no effect on the hydrolytic activity.

     

Ion transport in crab gills: A new method using isolated half platelets of Eriocheir gills in an ussing-type chamber

• 1.1. A half platelet preparation from Chinese crab (Eriocheir sinensis) gill is described which allows electrophysiological investigations of ion transport by gill epithelial monolayer when mounted in a modified Ussing chamber.
• 2.2. The resistance of these preparations equals half that of complete gill platelets (containing the gill epithelium and cuticle twice) indicating that cell damage during preparation of half platelets is negligible.
• 3.3. The transepithelial resistance (resistance of cuticle subtracted previously) was determined to be about 140 Ω cm² when both sides are bathed with identical salines.
• 4.4. Similarities to the results obtained with perfused complete gills demonstrates the reliability of this preparation.
• 5.5. When identical salines are applied on both sides of the epithelium an outside positive transepithelial potential difference (PD_te) up to 40 mV was measured.
• 6.6. The occurrence of such a high PD_te under symmetric conditions and its sensitivity to CN⁻ suggests the PD_te to be generated by active transport processes.

     

Sodium transport in red blood cells of lamprey LAmpetra fluviatilis

• 1.1. Unidirectional Na⁺ influx in lamprey red blood cells was determined using ²²Na as a tracer.
• 2.2. Total Na⁺ uptake and amiloride-inhibitable Na⁺ influx increased in a saturable fashion as a function of external Na⁺ concentration (Na_e).
• 3.3. At 141 mM Na_e, the average value of net Na⁺ influx was 13 ± 1.1 and the amiloride-sensitive Na⁺ influx was 5.3±1.1 mmol/l cells per hr (±SE).
• 4.4. The amiloride-sensitive component of Na⁺ influx was significantly activated by 10⁻⁵ M isoproterenol, by 2 × 10⁻⁵ M DNP, and by cell shrinkage.
• 5.5. Furosemide (1 mM) had no effect on the Na⁺ transport in red cells.
• 6.6. The residual amiloride-insensitive component of Na⁺ transport was a linear function of Na_e in the range of 5–141 mM. This transport seems to be accounted for by simple diffusion.

     

The effects of heavy metals and metabolic inhibitors on calcium uptake by gills and scales of Fundulus heteroclitus in vitro

• 1.1. Cadmium (Cd) and zinc (Zn) were inhibitory to calcium uptake by isolated gills of Fundulus heteroclitus in vitro. The metals appeared to act by displacing Ca²⁺ ions from protein carriers involved in facilitated diffusion.
• 2.2. In saltwater fish, transport of calcium across the serosal membrane of gill chloride cells is partly energy dependent and is likely mediated by Ca²⁺-ATPase. However, much of the calcium transport through the gill epithelium appears to occur by passive processes.
• 3.3. Cd (10⁻⁵M—10⁻³M) and Zn (10⁻⁷M—10⁻³ M) inhibited calcium uptake by isolated scale patches incubated in a physiological saline.
• 4.4. Cyanide, oubain, and quercetin treatment of scale patches produced results similar to those of the Cd and Zn treatments suggesting that metal-induced inhibition of ATPases may be responsible for reduced calcium transport by scale osteoblasts.

     

Functional properties of the two major hemoglobin components from Leporinus friderici (Pisces)

• 1.1. The hemoglobins of Leporinus friderici were separated by liquid chromatography on DEAE-Sepharose in order to isolate the two major electrophoretic components.
• 2.2. The chromatographic fraction I (electrophoretically slow anodic) showed no Bohr effect and no nucleoside triphosphate modulation.
• 3.3. The chromatographic fraction III (electrophoretically fast anodic) showed a normal Bohr effect and addition of nucleoside triphosphate decreased oxygen affinity but did not alter the Bohr effect.
• 4.4. The whole hemolysate showed a normal Bohr effect and phosphate modulation altered both Bohr effect and oxygen affinity.
• 5.5. No or little difference between the effect of adenosine or guanosine triphosphates on hemoglobin function was observed.

     

The effect of triiodothyronine on glucose utilization in adipocytes from obese rats

• 1.1. In the presence of insulin, 10⁻⁵ M 3,3',5-triiodothyronine (T₃) treatment for 1/2 hr decreased fatty acid synthesis 35% only in adipocytes from lean rats, whereas at 10⁻¹¹ M through 10⁻⁷M T₃ the obese adipocytes had nearly a 20% increase in fatty acid synthesis.
• 2.2. A 2 hr pretreatment of adipocytes with 10⁻⁹ and 10⁻⁷ M T₃ decreased insulin-stimulated fatty acid synthesis by nearly 20% in both lean and obese adipocytes.
• 3.3. In the absence of insulin, the 2 hr pretreatment with 10⁻⁹ M T₃ resulted in a 45% increase in lean adipocyte fatty acid synthesis, though the obese adipocytes required at least 10⁻⁷ M T₃ for 2 hr to increase the non-insulin-stimulated fatty acid synthesis by 50%.
• 4.4. At 10⁻⁹M T₃ concentrations non-insulin-stimulated fatty acid synthesis was increased by 200% in lean adipose tissue explants, but obese adipose expiants were not significantly affected under these conditions.
• 5.5. The addition of 10⁻⁹ M T₃ plus insulin to the explant media decreased fatty acid synthesis by 35% in both the lean and obese tissues.
• 6.6. The results also imply that the low T₃ status of the obese rat may be contributory to the elevated fatty acid synthesis observed in obese adipocytes.

     

Combined effect of adenosine,alpha adrenergic and adenosine antagonists on serum insulin and insulin secretion from rat pancreatic islets

• 1.1. The effect of adenosine separately or in combination with alpha-1 adrenergic antagonist prazosin and alpha-2 adrenergic antagonist yohimbine as well as adenosine antagonists 8-phenyltheophylline and xanthine amine conjugate on glucose-induced insulin secretion from isolated rat pancreatic islets was studied.
• 2.2. Their in vivo effects on serum glucose and insulin levels were also investigated. Adenosine at 10 and 100 μM inhibited significantly, insulin secretion from the isolated islets whereas at 10 mM slightly increased the secretion of insulin.
• 3.3. Prazosin used at 100 μM inhibited insulin secretion. When it combined with adenosine (10 μM) it augmented the inhibitory effect of adenosine.
• 4.4. In vivo prazosin (21 mg/kg bodywt) caused a hyperglycaemia which was accompanied by hypoinsulinaemia.
• 5.5. Concurrent administration of this drug with adenosine neither affect the hyperglycaemic nor the hypoinsulinaemic effects of adenosine.
• 6.6. On the other hand, yohimbine (100 μM) has no effect neither separately nor in combination with adenosine (10 μM) in modulating the inhibitory effect of adenosine on insulin secretion.
• 7.7. When Yohimbine administered at 19.5 mg/kg body wt it did not alter serum glucose but it markedly increased the serum insulin level. Its combined administration with adenosine reduced the hyperglycaemic effect of adenosine with a remarkable increase in serum insulin.
• 8.8. Both adenosine-antagonists were ineffective in alteration of insulin secretion.
• 9.9. However, combination of 8-phenyltheophylline with adenosine (10 μM) totally blocked the inhibitory effect of adenosine on insulin secretion while xanthine amine conjugate failed to prevent this effect of adenosine.
• 10.10. These results indicate that the inhibitory effect of adenosine on insulin secretion is neither mediated via alpha-1 nor alpha-2 adrenoceptors. It might be via activation of specific adenosine receptors on rat islets which are sensitive to blockade by 8-phenyltheophylline.

     

Purification and some properties of a Mg2+-activated acid phosphatase from rat testis

• 1.1. The acid phosphatase (AcPase, EC 3.1.3.2) IV from rat testicular tissue was purified to apparent homogeneity.
• 2.2. The enzyme displays a native molecular weight of 70 kDa determined on gel permeation chromatography on a Sephadex G-100 column and 68 kDa using linear 5–20% sucrose density gradient centrifugation. The subunit molecular weight on SDS-PAGE analysis is 67 kDa, suggesting that the enzyme is a monomeric protein.
• 3.3. The enzyme does not bind to Concanavaline A-Sepharose 4B column, indicating that it is not a glycoprotein.
• 4.4. The rat testis AcPase IV is a metal activated enzyme in which Mg²⁺ is the metal activating agent with a K_a, = 0.88 × 10⁻³ M. The Michaelis constant for p-nitrophenylphosphate, in the presence of saturating concentrations of Mg²⁺ ions, is 0.23 × 10⁻³ M.
• 5.5. The enzyme preferentially hydrolizes p-nitrophenylphosphate, phenylphosphate and ATP.

     

Interaction with functional membrane proteins—A common mechanism of toxicity for lipophilic environmental chemicals?

• 1.1. The effects of several phenols, anilines and aliphatic alcohols on yeast plasma membrane H⁺-ATPase and purine transport system as well as on Na⁺, K⁺-ATPase and adenosine uptake by Chinese hamster ovary cells (CHO) were investigated.
• 2.2. In all cases an inhibition was observed, which could be correlated with the octanol/water partition coefficients of the substances tested, thus making quantitative structure-activity predictions possible.
• 3.3. The observed effects correlated well with the influence of the chemicals on cell growth.
• 4.4. The results suggest a common mechanism of toxicity by the action of hydrophobic xenobiotics on biomembranes.

     

Driving forces for the uphill transport of amino acids into epidermal brush border membrane vesicles of the sea anemone,Anemonia sulcata (Cnidaria,anthozoa)

• 1.1. The transport of amino acids into membrane vesicles prepared from epidermal tentacle tissue of the sea anemone, Anemonia sulcata, depends on an electrochemical potential difference caused, e.g. by sodium chloride gradients.
• 2.2. Potassium or choline chloride gradients energized the transport less effectively than sodium chloride gradients. Both Na⁺-ions and Cl⁻-ions were required for the amino acid transport.
• 3.3. The uphill transport of amino acids along the downhill movement of driver ions (sodium chloride gradient conditions) was characterized by an overshoot; under sodium chloride equilibrium conditions, however, an accumulation of amino acids within the vesicles could not be measured.
• 4.4. Potassium diffusion potentials in combination with valinomycin indicated that hyperpolarization (vesicle inside negative) and hypopolarization (vesicle inside positive) enhanced or depressed the accumulation of amino acids within the vesicles.
• 5.5. Being at the phylogenetic base of the Eumetazoa, cnidarians show characteristics for the transmembrane transport of amino acids comparable to those established for vertebrates.

     

Intestinal l-methionine transport in the cultured gilthead bream (Sparus aurata)

• 1.1. The influx and transepithelial movements of l-methionine and its effects on the electrophysiology and Na-Cl-transport in upper and lower intestine of the cultured fish, Spanis aurata, were measured.
• 2.2. The K_m and V_max of l-methionine influx into the tissues were higher in lower intestine than in upper intestine. A prominent diffusion-like transport component was also measured in both segments during influx experiments.
• 3.3. Net transepithelial fluxes of l-methionine (1 mM) were observed in both upper and lower intestine, this transport being Na⁺-dependent.
• 4.4. The two intestinal segments exhibited an electrical potential difference (PD) and a short circuit current (I_sc) serosa negative or near zero. Tissue conductance (G_t) was higher in posterior than in lower intestine.
• 5.5. Addition of l-methionine to the mucosal side of lower or upper intestine did not induce changes in PD in either part.
• 6.6. Isotopic fluxes of Cl⁻ or Na⁺ measurements under short circuit conditions showed that there were no net Cl⁻ or Na⁺ transport in either part.
• 7.7. l-Methionine additions to the mucosa did not induce changes in unidirectional fluxes of Cl⁻ or Na⁺ or in the (I_sc) in either the anterior or posterior intestine.

     

Peritubular Na-K exchange ion pump in maleate-treated frog kidney proximal tubular cells

• 1.1. After perfusion of isolated frog kidneys for 1 hr with 10⁻³ or 10⁻² M maleate Ringer, the peritubular membrane potential gradually declined in a dose-dependent manner.
• 2.2. The ouabain-like effects of maleate on cell Na and K activities were dose-dependent and smaller than the effects of zero K or 10⁻⁴M ouabain. Intracellular pH was not altered in the presence of 10⁻²M maleate.
• 3.3. The driving force for Na entry into the cell was reduced, respectively, to 81.4 and 58.4% (of control) in the presence of 10⁻³ and 10⁻² M maleate.
• 4.4. There was no histochemically detectable inhibition of proximal tubule Na-K ATPase activity during 3 hr of perfusion with 10⁻² M maleate.

     

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.

     

Effects of the nonsteroidal anti-inflammatory drug piroxicam on energy metabolism in the perfused rat liver

• 1.1. The actions of piroxicam, a nonsteroidal and noncarboxylic anti-inflammatory drug, on the metabolism of the isolated perfused rat liver were investigated. The main purpose was to verify if piroxicam is also active on glycogenolysis and energy metabolism, as demonstrated for several carboxylic nonsteroidal anti-inflammatories.
• 2.2. Piroxicam increased oxygen consumption in livers from both fed and fasted rats.
• 3.3. Piroxicam increased glucose release and glycolysis from endogenous glycogen (glycogenolysis).
• 4.4. Gluconeogenesis from lactate plus pyruvate was inhibited.
• 5.5. The action of piroxicam on oxygen consumption was blocked by antimycin A, but not by atractyloside.
• 6.6. The action of piroxicam in the perfused rat liver metabolism seems to be a consequence of its action on mitochondria.
• 7.7. It can be concluded that inhibition of energy metabolism and stimulation of glycogenolysis are not specific properties of carboxylic nonsteroidal anti-inflammatory drugs.

     

Sea water drinking as a homeostatic response to dehydration in hatchling loggerhead turtles Caretta caretta

• 1.1. Hatching Caretta caretta may lose up to 12% of their initial hatched weight from water loss during emergence from the nest.
• 2.2. After subsequent osmotic and excretory water loss in sea water, hatchlings will drink sea water (166 μl 100 g⁻¹ hr⁻¹) and return to their initial weight within 10–15 days, without feeding.
• 3.3. There were no significant changes in plasma osmolarity or sodium levels over this period.
• 4.4. This osmoregulatory strategy is in marked contrast to that seen in the estuarine crocodile, Crocodylus porosus.

     

The possibility that Ca2+-ATPase from the plasma membrane-rich fraction of bovine parotid gland is ecto-Ca2+-dependent nucleoside triphosphatase

• 1.1. Parotid plasma membrane nonpump low-affinity Ca²⁺-ATPase, which possesses high-affinity (Ca²⁺ + Mg²⁺ )-ATPase activity, was characterized.
• 2.2. Purified Ca²⁺-ATPase hydrolyzed the nucleoside triphosphates, GTP, ITP, CTP, UTP, TTP (67–93% of ATP) and nucleoside diphosphates, ADP. GDP, IDP, CDP, TDP (12–40% of ATP) but not AMP and p-NPP.
• 3.3. The maximum activities of Ca²⁺- and (Ca²⁺ +Mg²⁺ )-ATPases were obtained in the presence of 1 mM and 0.13 μ M Ca²⁺, respectively.
• 4.4. The K_m values for Ca²⁺ in Ca²⁺- and (Ca²⁺+ Mg²⁺ )-ATPases were 0.2 mM and 22 nM. respectively.
• 5.5. The activities of both Ca²⁺- and (Ca²⁺ + Mg²⁺ )-ATPases were found in the right-side-out-vesicles obtained from the plasma membrane-rich fraction.
• 6.6. These features suggest that Ca²⁺-ATPase is an ecto-Ca²⁺-dependent nucleoside triphosphatase.

     

2-Deoxyglucose transport by the frog sartorius: Effects of electrical stimulation and N-carbobenzoxy-glycyl-l-phenylalaninamide

• 1.1. Studies characterizing glucose transport in the frog sartorius were performed.
• 2.2. For nonstimulated and stimulated muscles, intracellular 2-deoxyglucose exceeded 2-deoxyglucose-6-phosphate at 15 min, showed little further increase, and was maintained below the extracellular concentration for 2 hr.
• 3.3. Accumulated 2-deoxyglucose-6-phosphate did not inhibit glucose transport.
• 4.4. Unlike in adipocytes, basal and stimulated 2-deoxyglucose transport showed no difference in sensitivity to N-carbobenzoxy-glycyl-l-phenylalaninamide.
• 5.5. Phenylarsine oxide blocked contraction-enhanced 2-deoxyglucose uptake.
• 6.6. These results suggest that the glucose transporter of the sartorius exhibits auto-regulation, and that basal transport is not regulated by the same process as in adipocytes.

     

Nacl flux in the flounder (Pseudopleuronectes americanus) intestine: Effects of ph and transport inhibitors

• 1.1. The effects of extracellular pH on Na⁺ and Cl⁻ absorption were studied in vitro in the small intestine of the winter flounder, Pseudopleuronectes americanus.
• 2.2. Reductions in bathing solution pH inhibited J_ms^na (mucosal-to-serosal flux) and J_net^na (net flux) (r = 0.90) and J_net^Cl (r = 0.92) [due to an increase in J_sm^Cl, (serosal-to-mucosal)] and decreased short circuit current (Isc).
• 3.3. Luminal bumetanide (0.1 mM) and amiloride (1 mM) inhibited Na⁺ and Cl⁻ absorption by reducing J_ms.
• 4.4. Luminal barium (5mM) and luminal copper (100 μM) decreased J_ms^Cl and increased J_sm^Cl.
• 5.5. We conclude that reductions in extracellular pH inhibit a luminal membrane NaCl absorptive process (Na⁺-K⁺-2Cl⁻) and stimulate an electrogenic Cl⁻ secretory process.