Membrane-Bound Ferric Hemoglobin in Nucleated Erythrocytes of the Black Scorpionfish Scorpaena porcus, Linnaeus 1758.

The content of membrane-bound methemoglobin (MtHb) in nucleated erythrocytes was studied in the black scorpionfish Scorpaena porcus (Linnaeus, 1758) in vitro. Spectral characteristics were determined for a whole hemolysate, a hemolysate obtained by stroma precipitation (a clarified hemolysate), and a resuspended stroma. The MtHb proportion in the erythrocyte stroma was found to exceed 80% (6.20 ± 0.59 µM). Clarified hemolysates were nearly free of MtHb (0.5 ± 0.2 µM). Membrane-bound ferric hemoglobin did not affect the erythrocyte resistance to osmotic shock. The osmotic fragility range was determined using a LaSca-TM laser microparticle analyzer (BioMedSystems, Russia) to be 102-136 mOsm/kg, much the same as in other bony fish species. A nitrite load (10 mg/L) significantly increased the MtHb content in the blood. However, the membrane-bound ferric hemoglobin content did not change significantly, amounting to 6.34 ± 1.09 µM (approximately 95%). The finding suggested a functional importance for MtHb present in the plasma membrane of nucleated erythrocytes. Membrane-bound MtHb was assumed to neutralize the external oxidative load and the toxic effect of hydrogen sulfide in bottom water layers, where the species lives.

Effect of hypersalinic stress on hemocyte morphology and hemolymph cellular composition of the ark clam (Anadarakagoshimensis).

Bivalve mollusks as typical osmoconformers are unable to maintain a constant level of internal osmolarity in conditions of salinity stress. Adaptation to fluctuations of environmental salinity is achieved through cellular osmoregulatory responses, which are accompanied with a substantial shift in functional state of cells. In the present work we investigated the effect of hypersalinity stress on hemolymph cellular composition and morphology of the ark clam (Anadara kagoshimensis) hemocytes. Ark clams were subjected to a gradual increase of environmental salinity from 18‰ to 35‰ and 45‰ and maintained at those conditions for two days. Exposure to hypersalinity 35‰ induced changes in erythrocyte morphology and led to a decrease of their diameter. At salinity 45‰ a substantial increase of hemocyte average diameter was observed, whereas the shape of cells did not change (18‰). Hyperosmotic stress was not associated with changes in hemocyte viability as well as changes in hemolymph cellular composition. The results of the present work demonstrate high tolerance of A. kagoshimensis to short-time exposure to hypersalinic conditions.

Monocyclicity in the Function of the Erythroid Hematopoietic Lineage in Teleost Fish Exemplified by Platichthys Flesus (Linnaeus, 1758).

The erythroid lineage was studied in the flounder-gloss (Platichthys flesus Linnaeus, 1758) during the annual cycle. The erythrocyte count in the blood was determined along with the contents of immature erythroid forms (basophilic and polychromatophilic normoblasts) in the head kidney (pronephros) and the blood. Cell proliferative activity was inferred from the [3H]thymidine inclusion in circulating immature erythrocytes. Irregularity was observed in erythropoiesis occurring in flounder-gloss hematopoietic tissue. Intense production of erythroid mass was mainly associated with a post-spawning period. This was evident from an increase in the contents of immature erythroid forms in the pronephros and circulating blood and an increase in their proliferative activity. The changes were associated with peculiarities of the erythroid system organization, which precludes regular production of erythropoietin in the kidney in teleost fish.

Superfluid ß phase of

It is known that in low magnetic fields the superfluid transition of ^{3}He in nematic aerogel occurs into the polar phase. Using a vibrating aerogel resonator, we observe that in high magnetic fields this transition splits into two discrete transitions, occurring at different temperatures. According to theoretical models, a new superfluid phase-the ß phase-should be realized between these two transitions. The temperature range of existence of the new phase is measured as a function of magnetic field. The results are well consistent with theoretical expectations for the ß phase.

Morphologic, cytometric and functional characterisation of Anadara kagoshimensis hemocytes.

Circulating hemocytes of ark clam, Anadara kagoshimensis, were investigated using light microscopy and flow cytometry. Hemolymph contained 3 morphotype of cells, amebocytes, erythrocytes and intermediate type cells, which formed two distinct subpopulations on flow cytometric dot plots. Large cells (intermediate morphotype and erythrocytes) amounted 85.6 ± 2.8% total cells in hemolymph. Erythrocytes were hemoglobin-containing cells with up to 40 granules presented in the cytoplasm. All hemocyte types, observed in the ark clam hemolymph demonstrated equal capacity to spontaneous production of reactive oxygen species.

Erythrocyte profile of circulating blood of Neogobius melanostomus (Pallas, 1814) under conditions of experimental hypothermia.

The influence of hypothermia on erythrocyte profile of thermophile teleost species round goby, Neogobius melanostomus (Pallas, 1814), has been studied. Fish were acclimated to temperature 1-2оС, 15-16оС and 19-20оС (control group) and held at given conditions for 5 days. The number of red blood cell precursors (pronormoblasts, basophilic and polychromatophilic normoblasts) in circulating blood has been estimated. Also, the number of abnormal erythrocytes, i.e. cells with micronuclei, nuclei invaginations, red blood cell shades, dacryocytes and cells undergoing amitosis has been determined on smears. The number of immature erythrocytes increased more than two times (p < 0,001) at 1-2оС. The number of low-differentiated precursors, pronormoblasts and early basophilic normoblasts, increased for the most part. The number of abnormal erythrocytes did not change substantially, The changes in cellular blood composition were accompanied with the increase of plasma lactate concentration, indicating hypoxic state of fish. The results of the present work indicate that hematopoietic tissue remains sensitive to controlling factors at hypothermia, such as hypoxia, and may enhance proliferation and differentiation of erythroid cells.

Gene Encoding a Novel Enzyme of LDH2/MDH2 Family is Lost in Plant and Animal Genomes During Transition to Land.

L-Lactate/malate dehydrogenases (LDH/MDH) and type 2 L-lactate/malate dehydrogenases (LDH2/MDH2) belong to NADH/NADPH-dependent oxidoreductases (anaerobic dehydrogenases). They form a large protein superfamily with multiple enzyme homologs found in all branches of life: from bacteria and archaea to eukaryotes, and play an essential role in metabolism. Here, we describe the gene encoding a new enzyme of LDH2/MDH2 oxidoreductase family. This gene is found in genomes of all studied groups/classes of bacteria and fungi. In the plant kingdom, this gene was observed only in algae, but not in bryophyta or spermatophyta. This gene is present in all taxonomic groups of animal kingdom beginning with protozoa, but is lost in lungfishes and other, higher taxa of vertebrates (amphibians, reptiles, avians and mammals). Since the gene encoding the new enzyme is found only in taxa associated with the aquatic environment, we named it AqE (aquatic enzyme). We demonstrated that AqE gene is convergently lost in different independent lineages of animals and plants. Interestingly, the loss of the gene is consistently associated with transition from aquatic to terrestrial life forms, which suggests that this enzyme is essential in aquatic environment, but redundant or even detrimental in terrestrial organisms.

Hemoglobin deoxygenation and methemoglobinemia prevent regulatory volume decrease in crucian carp (Carassius carassius) red blood cells.

Fish red blood cells (RBCs) exhibit an oxygen-dependent regulatory volume decrease (RVD) in hypoosmotic environment. In higher vertebrates, membrane-associated hemoglobin is involved in the regulation of osmotic ion movements across the cellular membrane. However, whether the hemoglobin conformational state plays a role in the regulation of osmotic responses in fish red blood cells is still not fully understood. We found that changes in hemoglobin conformation influence the pattern of the regulatory volume decrease in Carassius carassius red blood cells. In oxygenated cells (96.4 ± 3.7% oxygenated hemoglobin), the volume recovery was completed within 125 min. Deoxygenation of hemoglobin (96.5 ± 2.7% of deoxygenated hemoglobin) inhibited the volume decrease in hyposmotically swollen red blood cells. Reoxygenation restored regulatory volume decrease in cells within 5 min. Induced methemoglobinemia (48.4 ± 1.8% of methemoglobin and 41.3 ± 2.3% of deoxygenated hemoglobin) blocked the process of volume recovery and significantly decreased osmotic stability of red blood cells.

Effect of Magnetic Boundary Conditions on Superfluid

We report results of experiments with superfluid ^{3}He confined in aerogels with parallel strands which lead to anisotropic scattering of ^{3}He quasiparticles. We vary boundary conditions for the scattering by covering the strands with different numbers of atomic ^{4}He layers and observe that the superfluid phase diagram and the nature of superfluid phases strongly depend on the coverage. We assume the main reason for these phenomena is a magnetic channel of the scattering which becomes important at low coverages and can be essential in other Fermi systems with triplet pairing.

Bose-Einstein Condensation of Magnons and Spin Superfluidity in the Polar Phase of

The polar phase of ^{3}He, which is topological spin-triplet superfluid with the Dirac nodal line in the spectrum of Bogoliubov quasiparticles, has been recently stabilized in a nanoconfined geometry. We pump magnetic excitations (magnons) into the sample of polar phase and observe how they form a Bose-Einstein condensate, revealed by coherent precession of the magnetization of the sample. Spin superfluidity, which supports this coherence, is associated with the spontaneous breaking of U(1) symmetry by the phase of precession. We observe the corresponding Nambu-Goldstone boson and measure its mass emerging when applied rf field violates the U(1) symmetry explicitly. We suggest that the magnon BEC in the polar phase is a powerful probe for topological objects such as vortices and solitons and topological nodes in the fermionic spectrum.

Observation of Half-Quantum Vortices in Topological Superfluid

One of the most sought-after objects in topological quantum-matter systems is a vortex carrying half a quantum of circulation. They were originally predicted to exist in superfluid ^{3}He-A but have never been resolved there. Here we report an observation of half-quantum vortices (HQVs) in the polar phase of superfluid ^{3}He. The vortices are created with rotation or by the Kibble-Zurek mechanism and identified based on their nuclear magnetic resonance signature. This discovery provides a pathway for studies of unpaired Majorana modes bound to the HQV cores in the polar-distorted A phase.

[FUNCTIONAL MORPHOLOGY OF BLOOD ERYTHROID CELLS IN NEOGOBIUS MELANOSTOMUS P. DURING CELL DIFFERENTIATION].

The morphometric characteristics of immature erythroid cells in circulating blood of the round goby (Neogobius melanostomus P) were studied: late basophilic normoblasts (BN), polychromatic normoblasts (PN), normocytes (mature erythrocytes). The linear sizes of the blood cells were assessed on photographs in the computer program ImageJ 1.44p. The sizes of the longitudinal and transverse axes of the cell and of its nucleus were determined. Using the appropriate algorithm, the following parameters were calculated: form index (MS), volume (Vc), area (Sc), thickness (h) and the specific surface area (SSc) of cells and nuc- lei as well as the nuclear-plasma ratio (NCR). The major changes occurred at the stage: PN - normocy- tes. They were aimed at improving the respiratory characteristics of cells. Besides accumulating hemoglobin in the cytoplasm and suppressing the functional activity of the nucleus, a significant increase in the diffusion surface of blood corpuscles was noted. As compared to BNs, S, and SS, of normocytes increased by 40 and 17 %, respectively, and the cells became ellipse-shaped. The processes underlying the formati- on of mature erythrocyte cytoskeleton are discussed.

Polar Phase of Superfluid (3)He in Anisotropic Aerogel.

We report the first observation of the polar phase of superfluid (3)He. This phase appears in (3)He confined in a new type of aerogel with a nearly parallel arrangement of strands which play the role of ordered impurities. Our experiments qualitatively agree with theoretical predictions and suggest that in other systems with unconventional Cooper pairing (e.g., in unconventional superconductors) similar phenomena may be found in the presence of anisotropic impurities.

[FUNCTIONAL STATE OF SCORPION FISH SCORPAENA PORCUS L. NUCLEATED ERYTHROCYTES UNDER DOSED HYPOXIA].

Hypoxia is a wide spread phenomena throughout the world's aquatic systems. This paper investigates the functional state of scorpion fish red blood cells under acute hypoxia between 0.57-8.17 mg O2/l (in vitro) by assessing of the fluorescence of Rhodamine 123 (R123) and Fluorescein diacetate (FDA). Acute hypoxia induced a significant increase in the fluorescence of both fluorochromes (R123 - 12-60, FDA - 30-184%). This reaction is based on a probable decrease in erythrocyte membrane permeability. The reduction of functional ionic channels in red blood cells delays FDA in the cytoplasm and lowers the energy costs required for transmembrane cationic gradient maintenance, which reflects an intensification of R123 fluorescence.

[The activity of transaminases in the tissues of the Black-Sea mollusk Mytilus galloprovincialis].

The activities of aspartate aminotransferase (AST, E. C. 2.6.1.1) and alanine aminotransferase (ALT, E. C. 2.6.1.2) were studied in the tissues of the bivalve mollusks differing in shell color. Shell color patterns were assessed by photographing and computer processing in Adobe Photoshop CS-3. The activity of transaminases was determined by the Reitman-Frankel procedure. Four color groups of mollusks were singled out: black, intermediate, dark- and light brown. The AST and ALT activities were found to increase as shells lose color probably reflecting different tolerance of mollusks to habitats with different concentration of oxygen. The ALT distribution pattern differs appreciably in the foot of M. galloprovincialis where the highest values were found in the black-colored mussels, evidently due to the change of the predominant enzyme function. In this tissue, the reaction of transamination replenishes the pool of free amino acids required for the synthesis of byssus threads.

[Hemoglobin system of golden mullet (Liza aurata, Risso) at adaptation to conditions of outer hypoxia].

Under conditions of experiment, effect of outer hypoxia on the hemoglobin heterogeneous system and respiratory blood characteristics were studied in the golden mullet (Liza aurata R). The control fish groups was maintained at oxygen pressure of 158-162 gPa, the experimental group at 54-55 gPa. Exposure--15 days, water temperature--15 degrees C. Under conditions of oxygen deficit in the mullet body there is noted development of complex of compensatory reactions directed to maintenance of the oxygen blood capacity. Initially (the 1st-2nd day), the release of erythrocytes from spleen is noted, which increases the number of red blood cells and the hemoglobin concentration in blood (emergency adaptation). Then (the 10th-15th day), quantitative readjustments occur at the level of the hemoglobin system, which lead to an increase of blood affinity to oxygen and to a decrease of its sensitivity to pH (long-term adaptation). The revealed changes take place on the background of a decrease of volume of circulating erythrocytes, which reflects the character of change of erythrocytic (MCV, MCHC) and seem to be a consequence of partial dehydration of blood plasma.

[Functional states of the antioxidant enzymatic complex of tissues of Mytilus galloprovincialis Lam. under conditions of oxidative stress].

There are generalized materials on reaction of enzymatic antioxidant (AO) complex of tissues of molluscs of the genus Mytilus to oxidative loading of different intensity. It is shown that with increase of level of oxidative stress, a rise of activity of glutathione peroxidase (GP) and glutathione reductase (GR) is observed, as well as the rate of glutathione (GSH) in tissues also increases. Then the toxic loading is compensated by a rise of activities of enzymes of low affinity to hydroperoxides, specifically of catalase. In the glutathione system, it is possible to identify several relatively steady states: of low, moderate, and highly intensive processes. In several tissues, functioning of the AO system seems to be directed not to rendering harmless, but to generation of reactive oxygen species (O2-, superoxide dismutase), which is likely to be determined by specificity of their running processes (destruction of damaged byssus theads). The molluscan AO complex is characterized by high lability and sensitivity both to physiological states (spawning) and to action of factors of anthropogenic nature (cationic detergents). Reactions of AO complex of the mussel to the natural states and to toxic action are comparable between each other, which is necessary to be taken into account at diagnostics of the aquatic medium.

[Coupling of membranous and metabolic functions in nucleated erythrocytes of Scorpaena porcus L. in hypoxia (experiments in vivo and in vitro)].

Effect of hypoxia (diapason of 0.57-8.17 mg O2 l(-1)) on functional characteristics of nucleated erythrocytes of the benthonic marine fish Scorpaena porcus L. was studied under conditions in vivo and in vitro. It has been established that characteristic of cellular system of this species is a balanced unhibition of membranous and metabolic functions under conditions of external deficit of oxygen (experiments in vivo). This is expressed in the ability of cells to maintain within norm the intracellular ATP concentration and transmembrane gradients for Na+ and K+ with low activities of Na+, K(+)-ATPase and hexokinase. This phenomenon seems to be based on the process of a decrease of the number of functioning ion channel at the level of the cell cytoplasmic membrane; this process reduces energy expenditure for maintenance of cationic gradients (the phenomenon of metabolic arrest). The same is also indicated by an increase of intensity of fluorescence of FDA and R123 in the scorpaena erythrocytic suspensions in hypoxia (experiments in vitro). Mechanisms underlying this phenomenon are discussed.

[Composition and content of carotenoids in body of the Black Sea gastropod mollusc Rapana venosa (Valenciennes, 1846)].

There were studied content and composition of carotenoids in body of the Black Sea gastropod mollusc Rapana venosa. To separate and identify this group of compounds, methods of thin layer and high performance liquid chromatography, mass-spectra and spectra in UV-VIS range were used. There were identified 84-87% of carotenoids. The main proportion belonged to mytiloxanthine--28-30%. The rest--pectenolon, diatoxanthin, alloxanthin--accounted for 10-15% of the total content except for beta-carotene--3-4%. The fraction is also isolated which contains complex esters of pectenolon, diatoxanthin, alloxanthin, and mytiloxanthin. It amounts to 17-20 % of pigments of carotenoid line. Minor components were not determined. Pathways of metabolic transformation of carotenoids in tissues of R. venosa are considered on the basis of study of the trophic system "mollusc-filtrator--mollusc-predator" by the example of Mytilus galloprovincialis and R. venosa.

[Qualitative composition of carotenoids, catalase and superoxide dismutase activities in tissues of bivalve mollusc Anadara inaequivalvis (Bruguiere, 1789)].

By using high-performance liquid chromatography, UV-VIS-spectra and mass spectra (FAB MS) in tissues of bivalve mollusc Anadara inaequivalvis (Bruguiere, 1789) there are identified seven kinds of carotenoids: trans- and cis-pectenolon, alloxanthin, pectenol A, beta-carotene, zeaxanthin, and diatoxanthin. Their quantitative ratio in hepatopancreas, gills, and foot of the animals was determined. There was revealed negative correlation (R2 about 0.9) between content of several carotenoids (trans- and cis-pectenolon, zeaxanthin, alloxanthin, and diatoxanthin) in tissues and activities of antioxidant enzymes (catalase and superoxide dismutase). The presence of competitive relations between these molecular systems is assumed and their underlying causes are discussed.