Functional morphology of blood erythroid cells in <Emphasis Type="Italic">Neogobius melanostomus</Emphasis> P. during cell differentiation

The morphometric characteristics of the following 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 dimensions of the blood cells were evaluated on photographs in a computer program ImageJ 1.44p. The longitudinal and transverse axes of the cell and its nucleus were measured. Using appropriate algorithms, the following parameters were calculated: shape index (SI), volume (V_c), area (S_c), thickness (h), and specific surface area (SS_c) of cells and nuclei as well as the nuclear–cytoplasmic ratio (NCR). Major changes were found to occur at the stage of PN → normocytes, being aimed at improving the respiratory characteristics of cells. In addition to accumulating hemoglobin in the cytoplasm and suppressing functional activity of the nucleus, a significant increment in the diffusion surface of erythroid cells was noted. As compared to BNs, S_c and SS_c of normocytes increased by 40 and 17%, respectively, while the cells assumed an ellipsoid shape. The processes underlying the formation of the mature erythrocyte cytoskeleton are discussed.     

Formation of electron transport barriers under ECR control of the q(r) profile in the T-10 tokamak

Abstract-the formation of transport barriers under electron cyclotron resonance heating and current drive in the t-10 tokamak is studied. in regimes with off-axis co-eccd and q _L <4 at the limiter, a spontaneous transition to improved confinement accompanied by the formation of two electron transport barriers is observed. the improvement resembles an L-H transition. It manifests itself as density growth, a decrease in the D_α emission intensity, and an increase in the central electron and ion temperatures. Two deep wells on the potential profile (the first one at r/a _L ≈0.6, where a _L is the limiter radius, and the second one near the edge) arise during the transition. the internal barrier is formed when dq/dr～0 with q≈1 in the barrier region.     

Tissue specificity of metabolism in the bivalve mollusc <Emphasis Type="Italic">Anadara inaequivalvis</Emphasis> Br. under conditions of experimental anoxia

Peculiarities of the course of metabolic processes in tissues of the bivalve mollusc Anadara inaequivalvis Br. were studied under conditions of experimental anoxia. In the absence of oxygen, in gill and foot the protein catabolism processes were found to be enhanced; this led to a decrease of the protein content and to an increase of the free amino acid and urea levels. Predominantly hydrolyzed were low molecular peptides, which was indicated by a decrease of the cathepsin D activity on the background of a rise of the γ-glutamyltranspeptidase activity. Anoxia was accompanied by enhancement of the succinate thiokinase and fumarate reductase reactions controlled by alanine and aspartate aminotransferases. This prevented accumulation of toxic lactate in tissues and allowed obtaining an additional macroerg resource. Metabolic processes in the mollusc hepatopancreas were oriented to production of amino acids.     

Effect of Cavβ Subunits on Structural Organization of Cav1.2 Calcium Channels

Background

Voltage-gated Ca_v1.2 calcium channels play a crucial role in Ca²⁺ signaling. The pore-forming α_1C subunit is regulated by accessory Ca_vβ subunits, cytoplasmic proteins of various size encoded by four different genes (Ca_vβ₁ - β₄) and expressed in a tissue-specific manner.

Methods and Results

Here we investigated the effect of three major Ca_vβ types, β_1b, β_2d and β₃, on the structure of Ca_v1.2 in the plasma membrane of live cells. Total internal reflection fluorescence microscopy showed that the tendency of Ca_v1.2 to form clusters depends on the type of the Ca_vβ subunit present. The highest density of Ca_v1.2 clusters in the plasma membrane and the smallest cluster size were observed with neuronal/cardiac β_1b present. Ca_v1.2 channels containing β₃, the predominant Ca_vβ subunit of vascular smooth muscle cells, were organized in a significantly smaller number of larger clusters. The inter- and intramolecular distances between α_1C and Ca_vβ in the plasma membrane of live cells were measured by three-color FRET microscopy. The results confirm that the proximity of Ca_v1.2 channels in the plasma membrane depends on the Ca_vβ type. The presence of different Ca_vβ subunits does not result in significant differences in the intramolecular distance between the termini of α_1C, but significantly affects the distance between the termini of neighbor α_1C subunits, which varies from 67 Å with β_1b to 79 Å with β₃.

Conclusions

Thus, our results show that the structural organization of Ca_v1.2 channels in the plasma membrane depends on the type of Ca_vβ subunits present.     

Peculiarities of organization of tissue metabolism in molluscs with different tolerance to external hypoxia

Oxygen consumption, content of several carbohydrate metabolites, and activities of their coupled enzymes were studied in bivalve molluscs with different tolerance to oxygen deficit: Mytilus galloprovincialis Lam. (black morpha) and Anadara Inaequivalvis Br. It has been shown that under conditions of external normoxia the anadara resistance to hypoxia preserves anaerobic orientation of metabolism. Its tissues are distinguished by high activities of malate and lactate dehydrogenases with the decreased content of glucose and the increased level of lactate. In several organs the succinate thiokinase and fumarate reductase reactions are realized, which is indicated by elevated activities of the alanine and aspartate aminotransferases. The anaerobic orientation of protein metabolism is added by a high pool of free amino acids and the elevated urea content in the mollusc tissues. The total orientation of metabolism in the anadara tissues rules out a significant lactate accumulation and determines low requirement of its organism in oxygen.     

Voltage-gated rearrangements associated with differential beta-subunit modulation of the L-type Ca(2+) channel inactivation

Auxiliary beta-subunits bound to the cytoplasmic alpha(1)-interaction domain of the pore-forming alpha(1C)-subunit are important modulators of voltage-gated Ca(2+) channels. The underlying mechanisms are not yet well understood. We investigated correlations between differential modulation of inactivation by beta(1a)- and beta(2)- subunits and structural responses of the channel to transition into distinct functional states. The NH(2)-termini of the alpha(1C)- and beta-subunits were fused with cyan or yellow fluorescent proteins, and functionally coexpressed in COS1 cells. Fluorescence resonance energy transfer (FRET) between them or with membrane-trapped probes was measured in live cells under voltage clamp. It was found that in the resting state, the tagged NH(2)-termini of the alpha(1C)- and beta-subunit fluorophores are separated. Voltage-dependent inactivation generates strong FRET between alpha(1C) and beta(1a) suggesting mutual reorientation of the NH(2)-termini, but their distance vis-à-vis the plasma membrane is not appreciably changed. These voltage-gated rearrangements were substantially reduced when the beta(1a)-subunit was replaced by beta(2). Differential beta-subunit modulation of inactivation and of FRET between alpha(1C) and beta were eliminated by inhibition of the slow inactivation. Thus, differential beta-subunit modulation of inactivation correlates with the voltage-gated motion between the NH(2)-termini of alpha(1C)- and beta-subunits and targets the mechanism of slow voltage-dependent inactivation.     

Identification of signature and primers specific to genus <Emphasis Type="Italic">Pseudomonas</Emphasis> using mismatched patterns of 16S rDNA sequences

Background

Pseudomonas, a soil bacterium, has been observed as a dominant genus that survives in different habitats with wide hostile conditions. We had a basic assumption that the species level variation in 16S rDNA sequences of a bacterial genus is mainly due to substitutions rather than insertion or deletion of bases. Keeping this in view, the aim was to identify a region of 16S rDNA sequence and within that focus on substitution prone stretches indicating species level variation and to derive patterns from these stretches that are specific to the genus.

Results

Repeating elements that are highly conserved across different species of Pseudomonas were considered as guiding markers to locate a region within the 16S gene. Four repeating patterns showing more than 80% consistency across fifty different species of Pseudomonas were identified. The sub-sequences between the repeating patterns yielded a continuous region of 495 bases. The sub-sequences after alignment and using Shanon's entropy measure yielded a consensus pattern. A stretch of 24 base positions in this region, showing maximum variations across the sampled sequences was focused for possible genus specific patterns. Nine patterns in this stretch showed nearly 70% specificity to the target genus. These patterns were further used to obtain a signature that is highly specific to Pseudomonas. The signature region was used to design PCR primers, which yielded a PCR product of 150 bp whose specificity was validated through a sample experiment.

Conclusions

The developed approach was successfully applied to genus Pseudomonas. It could be tried in other bacterial genera to obtain respective signature patterns and thereby PCR primers, for their rapid tracking in the environmental samples.