The effects of adrenaline on the oxygen transport properties of Salmo gairdneri blood

1. An injection of adrenaline into the dorsal aorta of Salmo gairdneri caused an increase in the dorsal aortic O2 content at 7 and 14 degrees C. 2. At 7 degrees C the number of erythrocytes in the dorsal aorta increased as indicated by the increases in both the Hct value and the haemoglobin concentration. 3. At 14 degrees C the erythrocytes in the dorsal swelled owing to the injection of adrenaline; this shifts the O2 dissociation curve to the left and in this way increases the dorsal aortic O2 content.     

The blood oxygen binding properties of hypoxicSalmo gairdneri

Summary The blood oxygen binding properties of rainbow trout responded to environmental hypoxia (the oxygen saturation of water 30% at 11°C) in three ways. The quickest response was a moderate acidosis, leading to slightly lowered blood oxygen loading due to the Bohr effect. The second response, an increase of blood oxygen carrying capacity, was completed with 6 h from the onset of hypoxia. The speed of the response suggests that the formation of new haemoglobin played no practical role, the increase being caused either by a decrease of plasma volume or the liberation of erythrocytes from a storage organ. The slowest response, a 25% increase of the blood oxygen affinity within a week of hypoxia, was probably caused by the concurrent decrease of the erythrocyte ATP concentration from 4.45 to 2.51 mol/ml erythrocytes.     

An analysis of oxygen binding by trout (Salmo gairdneri) red cells

• 1.1. Oxygen equilibrium curves have been measured on ATP-depleted trout red cells, at pHs between approx. 7 and 9, at temperatures of 20 and 15°C.
• 2.2. Results were analysed according to the model of Di Cera E., Robert C. H. and Gill S.J. (Biochemistry, 1987 26, 4003–4008), using a non-linear least-squares method. This allosteric model, proposed for human HbA, assumes the existence of two quaternary states, T and R, plus large functional differences between the α and β chains within the T state.
• 3.3. For this material, the fit was found rather better than that previously observed with the original Monod-Wyman-Changeux model. This is in accord with the idea that important α-β chain differences, a demonstrated property of Root effect Hbs, would play an important role in oxygen binding by trout red cells, where the Root effect Hbs are dominant.
• 4.4. In comparison, the fit was found poor for oxygen binding by human red cells, suggesting that the hypothesis of important chain differences would not apply to this material.

     

The fundamental niche of blood oxygen binding in the pelagic ocean

Marine species ranging in size from microscopic zooplankton to large predatory fish move vertically in the ocean water column to forage for food and avoid predators. Oxygen and temperature decrease, often rapidly, from shallow to deeper depths, restricting the ability of species to use the vertical habitat. One physiological trait that determines the tolerance of organisms to low oxygen is the oxygen affinity of oxygen carrier proteins, hemoglobin and hemocyanin, in the blood. To quantify the range of oxygen affinities for marine organisms, we surveyed the literature for measurements of oxygen binding to blood at multiple temperatures to account for its temperature sensitivity. Oxygen affinity is mapped within the ocean environment using the depth at which oxygen pressure decreases to the point at which the blood is 50% oxygenated (P₅₀ depth) as organisms move from the surface to depth in the ocean water column. We find that vertical gradients in both temperature and oxygen impact the vertical position and areal extent of P₅₀ depths. Shifts in P₅₀ due to temperature cause physiological types with the same P₅₀ in the surface ocean to have different P₅₀ depths and physiological types with different P₅₀'s in the surface ocean to have the same P₅₀ depth. The vertical distances between P₅₀ depths are spatially variable, which may determine the frequency of ecological interactions, such as competition and predation. In summary, P₅₀ depth, which represents a key physiological transition point between dexoxygenated and oxygenated blood, provides mechanistic insight into organism function within the water column of the global ocean.     

The oxygen uptake of Sarotherodon niloticus L. and the oxygen binding properties of its blood and hemolysate (Pisces: Cichlidae)

The oxygen consumption of Sarotherodon niloticus L. was found to decline below a critical oxygen concentration of about 2 mg O2/l. An important influence of CO2 on the oxygen affinity of whole blood was observed at all temperatures between 20 and 35 degrees C for gas mixtures containing 5.6% CO2. Purified hemolysate showed extremely high oxygen affinities (p50 = 1.08 mmHg at pH 8.2 and 20 degrees C). Low cooperativity was observed at all temperatures from 20 to 35 degrees C, and pH values between 6.5 and 8.2. The Bohr effect proved to be important at pH values lower than pH 7.5 (phi = delta log P50/delta pH = -0.58 between pH 6.5 and 7.0 at 35 degrees C). The oxygen affinities show high thermal sensitivity without a marked pH influence (delta H value for overall oxygenation at pH was -71.7 kJ/mol). The obtained results are interpreted as adaptations to diurnal variations in ambient temperature and oxygen availability.     

Temperature acclimation and oxygen binding properties of blood of the European eel, Anguilla anguilla

Temperature acclimation of the European eel, Anguilla anguilla, resulted in red cell GTP/Hb molar ratios of 1.20, 1.77 and 0.80 at 2, 17 and 29 degrees C, respectively. A small increase in blood oxygen capacity was present in 29 degrees C acclimated eels. The CO2 Bohr effect and the shape of the oxygen binding curve (n-Hill) were invariant with both temperature and GTP/Hb. The significant differences in the GTP/Hb ratio corresponded with a strong enhancement of the temperature effect on blood oxygen affinity between 2 and 17 degrees C and a similarly strong compensation between 17 and 29 degrees C. Predicted in vivo P50 values were 3.0, 13.8 and 17.6 mmHg at 2 degrees C, 17 and 29 degrees C, respectively. The adaptational value of these findings are discussed in relation to standard metabolic rates at the various temperatures. A tentative hypothesis is proposed that the present study confirms and expands earlier work and supports the contention that adjustments in blood oxygen affinity of thermally acclimated teleosts serve to provide them with an unloading O2 tension for diffusion closely matching the standard oxygen requirements at the various temperatures.     

Effect of sodium nitroprusside on hemoglobin oxygen binding properties of the blood during hepatic ischemia-reperfusion in rabbits

The parameters of blood oxygen transport were determined in rabbits during the hepatic ischemia/reperfusion (HIR) with or without sodium nitroprusside (SNP) administration. Hepatic ischemia was induced for 30 min by a. hepatica propria clamping, reperfusion lasted 120 min. Indices of blood oxygen transport (hemoglobin-oxygen affinity index (p50), pO2, pH, pCO2, HCO3-, TCO2, ABE, etc.) and nitrite/nitrate (NO(x)) amounts were measured in blood during HIR. Animals were subdivided into two groups: 1st group--HIR; 2nd--HIR plus SNP infusion (SNP, Sigma, i.v. 10 mcmol/kg). The experiments had shown that HIR led to significant acidic changes in the acid-base balance and high blood p50. The SNP infusion in the 2nd group led to less changing in the p50 values during HIR which were accompanied with high NO(x) levels. We conclude that oxyhemoglobin dissociation curve shift leftwards after SNP administration promotes the maintenance liver during ischemia-reperfusion.     

Thermoacclimatory changes in blood oxygen binding properties and gill secondary lamellar structure ofSalmo gairdneri

Summary The blood oxygen binding properties and gill secondary lamellar structure of rainbow trout acclimated to several temperatures were studied. The blood oxygen carrying capacity decreased as acclimation temperature increased from 2 to 15 °C; the decrease was probably caused by an increase in plasma volume. Also the blood oxygen affinity decreased as the acclimation temperature increased from 2 to 15 °C. This change had no effect on the oxygen loading in gills, since the efferent arterial oxygen tension was adequate for approximately 100% erythrocytic O₂ saturation at all acclimation temperatures, but facilitated the oxygen unloading in tissues. At the highest acclimation temperature (18 °C) the oxygen loading in gills was facilitated by the changes in the secondary lamellar structure; the proportion of erythrocytes in the secondary lamellar capillaries was higher than at the other acclimation temperatures (2 and 10 °C).     

Effect of Cl- and H+ on the oxygen binding properties of glutaraldehyde-polymerized bovine hemoglobin-based blood substitutes

Bovine hemoglobin was cross-linked with glutaraldehyde, resulting in high oxygen affinity polymeric hemoglobin dispersions of varying molecular weight distributions. High oxygen affinity acellular oxygen carriers were designed in order to exhibit oxygen release profiles closer to that of human red blood cells (RBCs), without exhibiting the inherent increased vasoactivity that occurs with low oxygen affinity acellular oxygen carriers (1, 2). Oxygen dissociation curves were measured for polymerized hemoglobin dispersions at various pH values (7.0, 7.4, and 8.0) and chloride ion concentrations. Unmodified hemoglobin showed an increase in oxygen affinity with increased chloride ion concentration and a decrease in oxygen affinity with increased pH, as was previously demonstrated in the literature (3). For glutaraldehyde-polymerized hemoglobin dispersions, the ability of the oxygen affinity to respond to changes in Bohr H+ and Cl- concentration was weakened. However, at acidic physiological pH (pH = 7), the Bohr effect was still present at high Cl- concentrations. Thus, the Bohr effect maintained some dependency on the Cl- concentration.     

Effects of hyperchloremia on blood oxygen binding in healthy calves

Three different levels of hyperchloremia wereinduced in healthy Friesian calves to study the effects of chloride onblood oxygen transport. By infusion, the calves received either 5 ml/kg of 0.9% NaCl (low-level hyperchloremia; groupA), 5 ml/kg of 7.5% NaCl (moderate hyperchloremia;group B), or 7.5 ml/kg of 7.5% NaCl(high-level hyperchloremia; groupC). Blood was sampled from the jugular vein and thebrachial artery. Chloride concentration, hemoglobin content, arterialand venous pH, PCO₂, and PO₂ were determined. At each timepoint (0, 15, 30, 60, and 120 min), the whole blood oxygen equilibriumcurve (OEC) was measured under standard conditions. Ingroups B andC, hyperchloremia was accompanied by asustained rightward shift of the OEC, as indicated by the significantincrease in the standard PO₂ at 50%hemoglobin saturation. Infusion of hypertonic saline also inducedrelative acidosis. The arterial and venous OEC were calculated, withbody temperature, pH, and PCO₂ valuesin arterial and venous blood taken into account. The degree of blooddesaturation between the arterial and the venous compartments[O₂ exchange fraction(OEF%)] and the amount of oxygen released at tissue level by 100 ml of bovine blood (OEF vol%) were calculated from the arterial andvenous OEC combined with the PO₂ andhemoglobin concentration. The chloride-induced rightward shift of theOEC was reinforced by the relative acidosis, but the alteredPO₂ values combined with the lowerhemoglobin concentration explained the absence of any significantdifference in OEF (% and vol%). We conclude that infusion ofhypertonic saline induces hyperchloremia and acidemia, which canexplain the OEC rightward shift observed in arterial and peripheralvenous blood.

     

Effect of oxygen binding on the dielectric properties of hemoglobin

The dielectric properties of horse hemoglobin have been investigated in the frequency range for 100 kcps to 15 Mcps at varying degrees of oxygenation. A linear dependence of the specific increment on the degree of oxygenation was found under a variety of experimental conditions, the increment of oxygenated hemoglobin being about 10% larger than that of deoxygenated hemoglobin. A similar difference was obtained with human adult and fetal hemoglobin. No variation of the dielectric parameters as reported by Takashima and Lumry could be detected.     

Monomer-dimer equilibrium and oxygen binding properties of ferrous Vitreoscilla hemoglobin

The monomer-dimer equilibrium and the oxygen binding properties of ferrous recombinant Vitreoscilla hemoglobin (Vitreoscilla Hb) have been investigated. Sedimentation equilibrium data indicate that the ferrous deoxygenated and carbonylated derivatives display low values of equilibrium dimerization constants, 6 x 10(2) and 1 x 10(2) M(-1), respectively, at pH 7.0 and 10 degrees C. The behavior of the oxygenated species, as measured in sedimentation velocity experiments, is superimposable to that of the carbonylated derivative. The kinetics of O(2) combination, measured by laser photolysis at pH 7.0 and 20 degrees C, is characterized by a second-order rate constant of 2 x 10(8) M(-1) s(-1) whereas the kinetics of O(2) release at pH 7.0 is biphasic between 10 and 40 degrees C, becoming essentially monophasic below 10 degrees C. Values of the first-order rate constants (at 20 degrees C) and of the activation energies for the fast and slow phases of the Vitreoscilla Hb deoxygenation process are 4.2 s(-1) and 19.2 kcal mol(-1) and 0.15 s(-1) and 24.8 kcal mol(-1), respectively. Thus the biphasic kinetics of Vitreoscilla Hb deoxygenation is unrelated to the association state of the protein. The observed biphasic oxygen release may be accounted for by the presence of two different conformers in thermal equilibrium within the monomer. The two conformers may be assigned to a structure in which the heme-iron-bound ligand is stabilized by direct hydrogen bonding to TyrB10 and a structure in which such interaction is absent. The slow interconversion between the two conformers may reflect a very large conformational rearrangement in the disordered distal pocket segment connecting helices C and E.     

Thermodynamic studies on oxygen binding by human red blood cells

Oxygen equilibrium curves have been measured on human normal red blood cells, at the temperatures of 20, 25, 30, 37 and 41 degrees C, and at pHs ranging from 6.8 to 8.2. The thermodynamical parameters have been determined for the four successive steps of oxygenation and for overall oxygenation, according to the Adair and MWC models [Monod J, Wyman J, Changeux JP. On the nature of allosteric transitions: a plausible model. J Mol Biol 1965;12:88-118]. The heat release appears to be nearly equal for the four steps. At the first three steps, the delta H change is counterbalanced by a nearly equivalent change of delta S, resulting in a rather small delta G value. delta G is greater at the fourth step, because of diminution of this enthalpy-entropy compensation phenomenon. The four steps are both enthalpy and entropy driven. According to the MWC model, the T to R transition is endothermic, and allosteric quaternary transition occurs at binding of the third oxygen. The average heat release increases by 2.8 kcal/mol when pH raises from 7.4 to 8.2, but flattens below pH 7.4. After correction for the heat of solution of oxygen and for the heat of proton release (referred to intracellular pH), an intrinsic heat for oxygenation of the heme of approximately--13 kcal/mol is obtained for the successive steps of oxygenation (at pH 7.4, 37 degrees C). These results are compared with those previously obtained for pigeon and trout red blood cells.     

Influence of oxidation and crosslinking on oxygen binding properties of mouse erythrocytes

Different chemical treatments for mouse erythrocyte modification has been used. Oxidation treatments with Ascorbate/Fe(3+), a system able to react with intracellular proteins, produced a displacement of the O(2) binding equilibrium curve to a higher affinity behaviour with loss of the haemoglobin cooperativity for oxygen binding. Incubation of mouse erythrocytes with diamide showed that at low reagent concentration (0.8 mM) no modification on oxygen binding equilibrium curves was observed. At higher reagent concentration (2.0 mM), an increased affinity and a disappearance of the cooperative behaviour can be observed. Additionally, crosslinking reactions on mouse erythrocytes with band 3 crosslinkers seemed to affect oxygen binding properties when used at a crosslinker concentration of 5 mM. Oxyhaemoglobin levels in crosslinked and diamide-treated erythrocytes are similar to those found in control cells. In contrast, ascorbate/Fe(3+) treatments produced an increment in the proportion of methaemoglobin, decreasing the oxyhaemoglobin levels in these oxidized erythrocytes.     

Adrenergic involvement in blood oxygen transport and acid-base balance during hypercapnic acidosis in the Rainbow Trout,Salmo gairdneri

Summary Rainbow trout (Salmo gairdneri) were subjected to 12 h of external hypercapnia (1% CO₂ in air) during - and/or -adrenoceptor blockade in order to assess the importance of adrenergic responses in modulating blood oxygen transport and acid-base balance during an acute acidotic stress. External hypercapnia caused an elevation of blood carbon dioxide tension and a reciprocal decrease in whole blood pH. A gradual elevation of blood bicarbonate levels caused whole blood pH to increase toward pre-hypercapnic values throughout the hypercapnic period. Pre-treatment of fish with propranolol (a -adrenoceptor antagonist) or phentolamine (an -adrenoceptor antagonist) did not affect their ability to regulate extracellular acid-base status during hypercapnia. On the other hand, adrenergic responses were essential in the maintenance of arterial blood oxygen content during hypercapnia despite the severe extracellular acidosis and a marked Root effect in trout blood, in vitro. Important adrenergic responses included pronounced increases in haematocrit (an -adrenergic effect) and arterial oxygen tension (- and -adrenergic effects) as well as partial regulation of red blood cell pH (a -adrenergic effect). Although pre-treatment of fish with either propranolol or phentolamine caused a reduction in blood oxygen content during hypercapnia, fish died only during complete adrenoceptor blockade, presumably due to severe hypoxemia.Symbols and abbreviations total concentration of oxygen or carbon dioxide, respectively - hct haemotocrit - rbc red blood cell     

The reversible binding of oxygen to sulfhemoglobin.

The O2 binding properties of sulfhemoglobin were studied. The oxygen tension required for half-saturation of sulfhemoglobin is more than 2 orders of magnitude higher than that for hemoglobin A. The binding of O2 exhibits an alkaline Bohr effect larger than that observed for hemoglobin, yet the Hill number is unity. From the Bohr titration curve, 0.68 proton is released during O2 binding at 0 degrees C. Sulfhemoglobin prepared from carboxypeptidase A-treated hemoglobin has an affinity for O2 which is about the same as that of sulfhemoglobin at the theoretical limit of the Bohr titration curve. Like its carboxypeptidase A-treated hemoglobin precursor, this sulfhemoglobin does not bind O2 cooperatively. Thus, sulfhemoglobin appears to be in a high affinity form at alkaline pH and a low affinity form at acid pH, similar to hemoglobin A. These results demonstrate that the magnitude of the Hill number is not always an indicator of the interaction between oxygen binding and other functions in a hemoglobin.     

Oxygen uptake by rainbow trout blood, Salmo gairdneri

Oxygen uptake rate by rainbow trout red cells was measured in the temperature range 5° to 40° C. Between 5° and 20° C there was a steady increase in rate and at 20° C a maximum rate of 2.5 μl O₂/min/ml red cells occurred, followed by a decline in the rate as the temperature increased. A morphological investigation of the red cell was conducted and possible functions of the nucleus in fish red blood cells are discussed.