Mass-transfer, utilization, and diffusion of oxygen in skeletal muscles of the stenohaline goby Gobius cobitus pallas under conditions of hypoosmotic medium

Effect of hypoosmotic conditions of medium on oxygen regime of skeletal muscles of the stenohalin goby Gobius cobitus Pallas was studied under conditions of experiment. The control fish group was maintained at 12–14‰, the experimental one—at 4.8–5.6‰. Duration of the experiment—44–45 days, water temperature—15 ± 1°C, photoperiod—12 day/12 night. It was established that under conditions of external hypoosmia there occurred hydration of the goby skeletal muscles and a decrease of their diffusion capability with respect to oxygen. The latter was accompanied by the tissue {ie215-1} decrease, which is indicated by low values of {ie215-2} in the venous blood outflowing from muscles. For the first 14–16 days of adaptation to the hypoosmotic medium there were restricted processes of mass transfer and oxygen utilization, which was associated with a decrease of the voluminous tissue blood flow and the blood oxygen concentration. These changes occurred on the background of the blood plasma hydration and a decrease of the number of circulated erythrocytes, and then they were completely compensated.     

The functional morphology of erythrocytes of the black scorpion fish <Emphasis Type="Italic">Scorpaena porcus</Emphasis> (Linnaeus, 1758) (scorpaeniformes: scorpaenidae) during hypoxia

The influence of hypoxia on the morphological characteristics of circulating erythrocytes of the scorpion fish Scorpaena porcus (Linnaeus, 1758) has been investigated in an in vivo experiment. Under a 4-h adaptation of the fish to the conditions of ranked hypoxia their erythrocytes demonstrated a number of consecutive reactions. The volume and the surface area of the red blood cells was reduced by 4–5% (p < 0.001) at an oxygen concentration of 2.6 mg/L (30% saturation of water with oxygen) and increased by 4% (p < 0.001) at a concentration of 1.3 mg/L (15% saturation), relative to the control values (normoxia: 7–8 mg/L). The observed reaction of erythrocytes coincided quantitatively and qualitatively (the order of events) with the results of the experiments we performed previously in vitro. Our study has shown that the physiology of the black scorpion fish is tolerant to hypoxia and allows autonomous functioning of red blood cells under conditions of oxygen deficit.     

Oxygen binding by marine fish blood under hypothermic experimental conditions

Influence of temperature in the range of 1-15 degrees C on oxygen binding properties of blood of thermophilic--golden mullet (Liza aurata), anchovy (Engraulis encrasicolus), and cold-tolerant--sardelle (Clupeonella cultriventris) fishes has been investigated under experimental conditions. Heat dependence of oxygenation reaction in thermophilic fish blood at temperature below 10 degrees C considerably increases, which is evidenced by high deltaH values. That is accompanied by a substantial increase of blood oxygen affinity and complicates blood deoxygenation at the tissue level. This reaction is apparently determined by the change of hemoglobin interaction with intraerythrocyte medium. The concentration of NTP in erythrocytes increases, that partially compensates negative changes of blood oxygen affinity (parameter P50 is raised) under long-term maintenance of fishes at 5 degrees C. However this reaction is not observed at low temperatures (1-2 degrees C).     

The influence of oxygen barotherapy on erythropoiesis in the recuperative period of hemorrhagic collapse]

Under the effect of high-pressure oxygen therapy (2 atm of O2 for 40 min) in anemic rats (blood loss - 2,8% of body weight in the course of 30 min) erythropoiesis in the bone marrow was more intense and led to effective restoration of erythrocytes and hemoglobin in the peripheral blood. Biphasic character of accumulation of erythropoientins in the blood serum of rats was seen on the 1st and the 10th days after acute blood loss. Erythropoiesis inhibitor was revealed in the blood serum of rats on the 15th day. A conclusion was drawn on the importance of erythropoietins and high pressure oxygen therapy in the mechanism of blood regeneration following the loss of blood.     

Studies on avian erythrocyte metabolism. Effect of organic phosphates on oxygen affinity of embryonic and adult-type hemoglobins of the chick embryo.

The effects of 2,3 diphosphoglyceric acid (2,3-DPG), adenosine triphosphate (ATP), and inositol hexaphosphate (IHP) on the oxygen affinity of whole “stripped” hemoglobin (WSH), hemoglobin H (Hb-H), hemoglobin A (Hb-A) and hemoglobin D (Hb-D) isolated from 18-day chick embryo blood have been determined. The effect of the three organic phosphates upon the oxygen dissociation curves is similar and the following order of decreasing oxygen affinity of the organic phosphates was observed for each hemoglobin: 2,3-DPG < ATP < IHP. 2,3-DPG appears to have a slightly greater effect upon the P₅₀ of Hb-H than upon that of either of the two adult-type hemoglobins. However, this effect seems insufficient to suggest a preferential interaction of 2,3-DPG with Hb-H which would account for either the large amounts of 2,3-DPG in the erythrocytes of embryos or the higher oxygen affinity of the whole blood. The effects of the organic phosphates upon the Hill constant of the purified hemoglobins are variable. It is concluded that since the distribution of hemoglobins H, A, and D in the erythrocytes during the developmental period from 18-day embryos to 6-day chicks remains fairly constant, the previously described progressive decrease in oxygen affinity of the whole blood during this period results from changes in the total amount and distribution of the intraerythrocytic organic phosphates.²     

Red blood of cosmonauts during missions aboard the international space station (ISS)

Morphological and biochemical investigations of red blood in cosmonauts on board the International Space Station (ISS) (from the 6th to the 12th expeditions) in the space experiment program “Hematology” were carried out 30 days before the space mission (SM), at the initial (days 6–10) and final (days 160–190) stages of SM, and after the SM (immediately after SM (day 0) and on the 7th and 15th days of the adaptation period to earth conditions). A reduction of the concentration of hemoglobin after a prolonged influence of SM factors has been found, which is probably related not only to the intensity of erythropoiesis but also with the possible early removal of a part of low-quality (probably, old) erythrocytes from the bloodstream, which is confirmed by the results on the metabolism of red blood cells and the state of the cell membrane. Stimulation of erythropoiesis (increase in erythropoietin, decreased level of iron in blood, removal of low-quality and old erythrocytes) in the period of readaptation to conditions on earth is aimed at maintenance of the optimal level of red blood cells required for increased oxygen demand in tissues under the conditions of earth gravitation and enhancement of muscular load.     

Morphofunctional peculiarities of erythrocytes in wild and farmed Coregonid fishes from Lake Baikal

Structure and cytometric indices of red blood cells (RBC) and hemoglobin content (Hb) and oxygen capacity of the blood (OCB) of omul, whitefish, and hybrids thereof captured in Lake Baikal (wild) and incubated and grown in a freshwater aquarium complex (FAC) (farmed) have been analyzed. Cytometric parameters of red blood cells of wild omul, whitefish, and hybrids thereof exceed those of the cells of fish reared in aquariums under identical conditions. The effect of aquarium rearing on the shape of red blood cells is the least pronounced in Siberian whitefish and F1 progeny of Siberian whitefish females and omul males (f Sw x m Om). The erythrocyte size in hybrids of female Lacustrine whitefish and male omul (f Lw x m Om) is determined by the size of these cells in female parents, since female Lacustrine whitefish have the largest erythrocytes. Cytometric parameters of erythrocytes of all Coregonid fishes investigated are higher in fish reared in warm aquaria than in conspecifics reared in aquaria with cold water. Erythrocyte nuclei are smaller in artificially propagated hybrids than in parent fish captured in the wild or in whitefish and omul reared in aquaria under the same conditions. A distinct pool of erythrocytes from whitefish captured in the wild have a 20–30% higher content of functionally active mitochondria than erythrocytes of whitefish reared in aquaria; a disrupted mitochondrial structure is also observed in erythrocytes from the latter population of fish. The results show that distinctive features of metabolism related to oxygen transport in the Baikal coregonid fish that were investigated are determined by adaptation to the conditions of the ecological niches occupied by the fish.     

Hemolysis and ATP Release from Human and Rat Erythrocytes under Conditions of Hypoxia: A Comparative Study

Red blood cells are involved not only in transportation of oxygen and carbon dioxide but also in autoregulation of vascular tone by ATP release in hypoxic conditions. Molecular mechanisms of the ATP release from red blood cells in response to a decrease in partial oxygen pressure still remain to be elucidated. In this work we have studied effects of hypoxia on red blood cell hemolysis in humans and rats and compared the effects of inhibitors of ecto-ATPase and pannexin on the release of ATP and hemoglobin from rat erythrocytes. The 20-min hypoxia at 37°C increased hemolysis of red blood cells in humans and rats 1.5- and 2.5-fold, respectively. In rat erythrocytes a significant increase in hypoxia-induced extracellular ATP level was found only in the presence of ecto-ATPase inhibitor ARL 67156. In these conditions we observed a positive correlation (R² = 0.5003) between the increase in free hemoglobin concentration and the ATP release. Neither carbenoxolon nor probenecid, the inhibitors of low-selectivity pannexin channels, altered the hypoxia-induced ATP release from rat erythrocytes. The obtained results indicate a key role of hemolysis in the ATP release from red blood cells.     

Structural changes in the erythrocyte membrane of the trout Salmo irideus at seasonal acclimatization

Differences of thermostability were studied in red blood cells of the trout Salmo irideus differing in sex and age, as well as structural-dynamic characteristics of erythrocyte membrane proteins at seasonal acclimatization in the interval of reservoir water temperature of 0–19°C. An increase of resistance of erythrocytes to temperature lysis with elevation of the environmental temperature was revealed to be accompanied by a rise of the proteins segmental mobility and a decrease of intermolecular interactions in spectrin-actin cytoskeleton from the data of the ESR spin labeling method. Regulation of erythrocyte stability during acclimation was concluded to occur both changes of the fatty acids chain package at the variations of lipid composition and by changes of the cytoskeleton structural lability. Thereby this provides an increase of the bilayer firmness, on the one hand, while, on the other hand, a rise of elasticity and expansibility of the membrane on the whole, which increases resistance of cells to colloidal-osmotic hemolysis. Changes of concentration of oxygen dissolved in water, which are caused by temperature fluctuations, do not deem to be of crucial importance for structural stability of erythrocytes, as it can be compensated by another mechanism, specifically by changes of affinity of hemoglobin to oxygen.     

The red blood cells in growing guinea pigs. (Cavia cobaya). II. Communication: erythropoiesis and red blood cells in the postnatal development period

Bone-marrow smears of 175 guinea pigs aged 1-27 days and venous blood samples of 351 animals aged 1-25 days were prepared for cell counting. A significant increase of erythroblasts were found between life day 1 and 2; normoblasts increased in number synchronously with a decrease of erythroblasts after the 5th day. The percentage of the erythroid bone marrow increased from 10 to 14 during the developmental period. Beyond the perinatal period the red blood picture is characterized by the following changes: a decrease of erythrocyte count, hematocrit, hemoglobin, mean corpuscular volume, and mean corpuscular hemoglobin; a constant mean corpuscular hemoglobin concentration; an increase of the reticulocyte count. The decrease of the red cell count is compensated by a decreasing oxygen affinity attained by an important increase of 2,3-DPG. Nevertheless, the stimulus for a raising erythropoiesis remains constant which can be shown by the growing percentage of erythroid cells and reticulocytes. The difference between the human postnatal development and that of the guinea pig becomes obvious. Cell counts in dependence of body masses in postnatally growing guinea pigs, veil the perinatal finding of the increase in erythrocytes up to the 5th day and the decrease of the mean corpuscular volume after the 3rd day.     

Morphological and functional features of the blood in humans during a nine-day stay in an oxyargon environment with various oxygen contents

Biochemical parameters of the red blood and oxygen transportation of hemoglobin by erythrocytes were studied in subjects who volunteered for chamber experiments (0.15 MPa) with normal and hypoxic oxygen-nitrogen-argon (O₂-N₂-Ar) gas environments. Erythrocyte metabolism, lipid and phospholipid spectra, and the efficiency of oxygen releasing and retention by hemoglobin were studied in the period of collecting baseline data, on the sixth day of stay in a hyperbaric normoxic O₂-N₂-Ar gas environment (13.7% O₂), on the third day of stay in a hyperbaric hypoxic O₂-N₂-Ar environment (9.9% O₂), and on the first and the tenth days of a postexperimental rehabilitation period. The high pressure, hypoxia, and the subsequent decompression resulted in a decrease in the ATP levels, which is probably due to the changes within the membrane and increased G6PD activity associated with cell repair. Changes in the lipid and phospholipid compositions of the membrane indicate an altered phase state of the plasma membrane, i.e., an increased viscosity, which may be related to possible changes in the membrane’s permeability. As a rule, hyperbaric conditions lead to a decrease in oxygen transportation in the blood of the subjects. Hypoxia against the background of hyperbaria had different effects on the subjects, namely, the absence of changes, a decrease, or an increase in the efficiency of oxygen transportation, which can be explained by the selectivity and individual sensitivity of the subjects to the experimental factors. A decrease in the efficiency of oxygen transportation by hemoglobin is most likely to be related to an increase in the viscosity of the plasma membrane, which may affect the conformation of hemoporphyrin of the membrane-bound hemoglobin and obstruct oxygen transportation through the membrane.     

The fatty acid composition of phospholipids and absorption spectra of lipid extracts from lamprey,frog, and rat erythrocytes

The work studies the content and fatty acid composition of phospholipids as well as the absorption spectra of lipid extracts from red blood cells of poikilothermal and homoiothermal animals at different evolutionary levels. The objects of study include two poikilothermal species, the river lamprey (Lampetra fluviatilis) that uses oxygen dissolved in water, and the common frog (Rana temporaria) that consumes oxygen both from water and from air. A homoithermal animal is the white rat (Rattus rattus) that inhabits the terrestrial-aerial environment. The animals are studied in winter and spring. The phospholipid content in lamprey blood plasma is found to be twice higher than that in its erythrocytes. In the frog and the rat, the ratio is reverse. Determination of the fatty acid lipid composition of red blood cell phospholipids suggests that membranes in the lamprey are denser than in the frog. As for the fatty acids in the erythrocyte fraction of rat blood, they appear to be less diverse, with a double prevalence of saturated acids over unsaturated ones and devoid of long chain (C22) ω3 fatty acids. All of this results in a lower degree of unsaturation and a denser packing of fatty acids in the membrane structures of rat erythrocytes. The mechanism of reversible binding of O₂ molecules to hemoglobin in erythrocytes is discussed. Presumably, the mechanism of interaction between molecules of O₂ and molecules of water prevents the exchange interaction of electrons of the hemoglobin iron atoms with an oxygen molecule. This is confirmed by our obtained absorption spectra, which show that in the lipid extract almost totally devoid of water the heme isolated from erythrocytes is converted to hemin.     

Orally administrated cerium chloride induces the conformational changes of rat hemoglobin, the hydrolysis of 2,3-DPG and the oxidation of heme-Fe(II), leading to changes of oxygen affinity

The structure and oxygen affinity of hemoglobin from erythrocytes of CeCl(3) fed Wistar rats in the dose range of 0.2-20.0 mg/kg body weight/day were investigated by means of various spectroscopic methods. The changes in oxygen saturation curves of hemoglobin are dependent upon both feeding dose and feeding time. After 40 days feeding with 20 mg CeCl(3)/kg body weight/day, the curve changed to a double sigmoid shape and the oxygen affinity in low oxygen pressure increases. It regained the sigmoid form after 80 days feeding, but the degree of oxygen saturation in higher oxygen pressure became higher than that in the control. These results indicate that CeCl(3) can increase the oxygen affinity of hemoglobin of rat erythrocytes. This effect is further demonstrated by the analysis of M?ssbauer spectra of erythrocytes. Increase of hemoglobin content in erythrocytes was found in rats fed with CeCl(3). It might be the offset response to the poor oxygen-releasing capability of the hemoglobin. CD and FT-IR deconvoluted spectra indicate that secondary structures of hemoglobin have remarkable changes, characterized by a gradual decrease of alpha-helix content, in a dose- and feeding time-dependent fashion. Meanwhile, the 31P NMR spectra demonstrate that the level of 2,3-diphosphoglyceric acid (2,3-DPG) in erythrocytes, an allosteric regulator of oxygen release from hemoglobin, decreases due to its hydrolysis. In addition, the M?ssbauer and ESR spectra show clearly that a fraction of the heme-iron changes from Fe (II) to Fe (III) in CeCl(3) fed rats. The results indicate that the oral administration of CeCl(3) leads to a microenvironment changes of heme in intracellular hemoglobin. Oxygen affinity changes might be attributed to a series of events triggered by the binding of Ce (III) to hemoglobin and 2,3-DPG, including conformational changes of hemoglobin and 2,3-DPG hydrolysis, respectively and also the partial transformation from heme-Fe (II) to heme-Fe (III).     

Release of erythrocytes from the spleen during exercise and splenic constriction by adrenaline infusion in the rainbow trout

Erythrocyte-supplying function of the spleen was examined in the rainbow troutSalmo gairdneri under exercise. The spleen showed remarkable reduction, about 70% in weight and about 85% in hemoglobin content, after forced exercise of 15 min. The amount of erythrocytes released from the spleen was 2.33 ml/kg body, and this amount corresponds to about 20% of the total volume of circulating erythrocytes in resting condition. No damage was observed at the spleen, splenic artery and splenic vein after the exercise. Examination of the vascular system by a corrosion casting method showed that no place other than the venous circulation exists for the erythrocytes released from the contracted spleen. The spleen was strongly constricted by infusion of adrenaline into the organ. These facts imply that the fish spleen supplies stored hemoglobin into the circulating blood in response to an increased demand of oxygen during exercise, under the control of the sympathetic nervous system.     

Serum glucose level in severe acute exogenous normobaric hypoxia in humans at rest

The study of the effect of acute normobaric hypoxia, which was simulated by inhalation of the oxygen-nitrogen mixture containing 8% of oxygen, which corresponds to its partial pressure at an altitude of 7000 m above sea level, was conducted in a group of apparently healthy men (aged 19–23 years, n = 10). The biochemical analysis during the test included determining the glucose, pyruvate, and lactate levels in venous blood; the hemoglobin content, pH, hematocrit, partial oxygen and carbon dioxide pressures, and hemoglobin saturation with oxygen. It was shown that, at the fifth minute of hypoxia, the serum glucose level decreased significantly (p < 0.05) compared to the background. However, on the whole, the maximal glucose level decrease was 0.76 mM, and the lowest individual parameter values did not decrease below 4.0 mM. The serum glucose level was restored to the background values at the tenth minute of testing. It was suggested that the syncopal form of altitude hypoxia in humans is unlikely to be linked to the development of hypoglycemia.     

The respiratory function of blood in elderly and old age and the factors that determine it

Indices of pulmonary gas exchange, blood gases, the oxyhemoglobin dissociation curve, and intraerythrocytic metabolic parameters were analyzed in 62 apparently healthy elderly and senile subjects (60–92 years old) and 18 young healthy subjects (19–30 years old). PaO₂ was found to decrease in elderly and senile subjects. Arterial hypoxemia in old age is caused by an increase in the alveoloarterial PO₂ gradient, primarily as a result of the malcoordination of pulmonary ventilation and blood flow. A rightward compensatory shift of the oxyhemoglobin dissociation curve was observed, which was due to facilitated oxygen release in tissues owing to a pH decrease in erythrocytes (the Bohr effect). However, the facilitated oxygen release by oxyhemoglobin cannot compensate for the effect of factors deteriorating oxygen supply delivery to tissues, observed with aging, which is confirmed by the decrease in the partial pressure of oxygen in the venous blood of elderly and senile people, reflecting PO₂ in tissues.     

Morphofunctional properties of red blood cells in humans during seven-day “Dry” immersion

Healthy male volunteers were subjected to seven-day “dry” immersion. After that, morphological and biochemical features of erythrocytes, erythropoiesis intensity, including the indices of iron metabolism and erythropoietin, lipid and phospholipid spectrum of the plasma membrane of erythrocytes, and the efficiency of binding and release of oxygen by hemoglobin were studied. The studies were performed before immersion, at the last seventh day of immersion, and on the 7th and 15th days of the recovery period. We found that seven-day “dry” immersion tended to change morphological composition of red blood, erythropoiesis intensity, and metabolic indices in erythrocytes. Seven-day simulated microgravity resulted in significant changes in the indices of oxygen-transporting function of erythrocytes, probably, due to changes at the membrane level and, particularly, in phospholipid fractions. These changes have no clinical importance, because all of them returned to the baseline after the 15-day recovery period. Substantial variability of data is related to an individual response of the body to stress induced by experimental conditions.     

Changes of gas exchange parameters and of functional-biochemical properties of erythrocytes in dynamics of experimental anemia in rats

In experiments on male Wistar rats, in a specially constructed computerized installation, O₂ consumption by the animals in comparison with changes of hematological, biochemical, and rheological blood properties is studied after anemization—acute blood loss (12–15% of the total blood mass). An increase of the O₂ consumption by the organism and tissues by 18–28% has been revealed for the first 7 days after the blood loss, in spite of a pronounced decrease of hematocrit and of the amount of erythrocytes and hemoglobin in peripheral blood by 20–25% of the initial level. There was a 5–10-fold increase of the content of immature erythrocyte forms—reticulocytes and a progressive rise of cell acidic resistance, which is characteristic of young erythrocyte forms. An increase of O₂ consumption at a decrease of the blood oxygen capacity (a low hemoglobin level) seems to be due to the more efficient transport and yield of O₂ to tissues. At the 3rd and 7th day after the blood loss, activity of Na,K-ATPase has been found to increase by 60% and 20%, respectively. Analysis of the erythrocyte rheological properties has shown that the maximal firmness of aggregates (U_q) and the aggregation rate (1/T) decrease progressively beginning from 3 days after the blood loss; index of deformability (I_max) turned out to be elevated by 7–11%, probably due to an increase of the cell membrane elasticity. The conclusion is made that changes of erythrocyte rheological properties are interconnected with changes of the Na,K-ATPase activity and are directed at optimization of blood circulation in large vessels and the capillary network.     

The osmometric behavior of human erythrocytes.

A reexamination of the data of Farrant and Woolgar (Cryobiology, 9, 9–15 (1972)) demonstrates that human erythrocytes exhibit osmometric behavior as predicted from the Boyle-Van't Hoff law when suspended in sodium chloride solutions as high as 3.5 osmolal. A Boyle-Van't Hoff plot is linear with r = 0.998. However, 24% of the water in isotonic red blood cells appears to be osmotically-inactive, although nearly 75% of this apparent non-osmotic water can be accounted for by the water of hydration of hemoglobin. These facts have direct significance to hypotheses concerning hemolysis as well as to the modeling of osmometric events which occur at sub-zero temperatures.     

The rate of uptake of carbon monoxide and of nitric oxide by normal human erythrocytes and experimentally produced spherocytes

The uptake of gases such as oxygen, carbon monoxide, or nitric oxide by the erythrocyte involves: (a) diffusion across the cellular membrane, (b) intraerythrocytic diffusion, and (c) chemical combination with hemoglobin. The aim of this investigation was to obtain data which would permit an analysis of each of these factors in limiting the rate of gas uptake. The initial over-all rate of uptake of gases which combine chemically with hemoglobin to produce a color change can be measured by a modified version of the Hartridge-Roughton-Millikan constant flow, rapid reaction apparatus. If nitric oxide is the reactant gas, only (a) and (b) are measured since the chemical combination of this gas with hemoglobin is extremely rapid. Our studies have shown that human biconcave discoidal erythrocytes at 38 and 48°C., have the same initial rate of carbon monoxide and nitric oxide uptake as the same cells converted into spherocytes of equal volume. Similarly there was no difference between discs and cells sphered with a 30 per cent increase in volume. Shrunken erythrocytes showed a marked decrease in rate of gas uptake. This suggests that surface area and maximum linear distance for intracellular diffusion of this magnitude do not measurably retard gas uptake. In the shrunken cells, a change in the orientation and concentration of intraerythrocytic hemoglobin and/or of the membrane components may have impeded gas diffusion.