Transfer of trace elements in the ocean-atmosphere-continent system as a factor in the formation of the elemental composition of the Earth's soil cover.

The dynamics of global flows of matter in the biosphere has been studied. The general law of redistribution of average elemental compositions in the biosphere between the solid and liquid phases (lithosphere-hydrosphere) is established. In these processes, the main role is played by "living matter." The most active processes of redistribution of mean elemental compositions in the biosphere take place in the places of "life thickening" (biogeochemical barriers) such as ocean-atmosphere, river-sea and hydrosphere-lithosphere. The result of these processes is a general relative increase in the concentrations of trace elements in the environment of living organisms. A new methodological approach is proposed to study the role of the transfer of trace elements in the ocean-atmosphere-continent system as a factor in the formation of the elemental composition of the Earth's soil cover.

VENTRICULAR REPOLARIZATION MEASURES IN PROFESSIONAL AND AMATEUR ATHLETES WITH HIGH NORMAL ARTERIAL PRESSURE.

There is merely little data regarding changes in repolarization among professional athletes with prehypertension, especially in comparison with amateur athletes. It was previously shown that disturbances of repolarization may have an important role in the development of cardiac changes in athletes and be observed in a number of conditions, such as ventricular hypertrophy, channelopathies, and aortic aneurysm. Therefore, in this study, it has been aimed to compare the ventricular repolarization measures in professional and amateur athletes with high normal arterial pressure.Thirty professional and thirty amateur all-around track and field athletes were enrolled in this project. According to the level of sport activities and the presence of HNBP, all athletes were divided into 4 groups. The first group consisted of professional athletes with normal and optimal blood pressure (NOBP) values (<130/80 mmHg, total of 21 persons), the second group was formed by 9 professional athletes with HNBP (≥130/85 mmHg), the following 3rd (17 persons) and 4th (13 persons) groups were composed by amateur athletes with the level of arterial blood pressure up to 130/85 mmHg and higher respectively. We have indicated that professional athletes with upper normal values of blood pressure have an extension of the TpTe interval compared to other groups of athletes, the average value of this indicator comprised 86 ± 10.94 ms, which, however, was below the dangerous level of >100 ms. This group of people also showed an increase in the ratio of TpTe/QT values (p=0.001) in the absence of a significant difference in S-Tpc intervals (p=0.605). These changes indicate an increase in transmural dispersion of repolarization with the development of left ventricular remodeling of professional athletes with upper normal values of blood pressure. In order to establish a connection between the revealed features of repolarization and non-functional overreaching/overtraining and altered autonomic cardiovascular activity with sympathetic activation syndrome, other pathogenesis features, their clinical and prognostic value, further examinations are required.

[Neurulation continues: the parade commander is...apical constriction].

Neurulation is traditionally defined as the process of closure of the neural tube. New data have shown that the major driving forces ofneurulation continue to operate with the closure of the neural tube, at least until the central canal of the neural tube has formed. Owing to this, the paper proposes to distinguish two periods of neurulation. According to these notions, early neurulation corresponds to the period of closure of the neural tube, and late neurulation corresponds to the period of formation of the central canal. Examples of neural tube defects that affect late neurulation are discussed.

Myogenesis and molecules - insights from zebrafish Danio rerio.

Myogenesis is a fundamental process governing the formation of muscle in multicellular organisms. Recent studies in zebrafish Danio rerio have described the molecular events occurring during embryonic morphogenesis and have thus greatly clarified this process, helping to distinguish between the events that give rise to fast v. slow muscle. Coupled with the well-known Hedgehog signalling cascade and a wide variety of cellular processes during early development, the continual research on D. rerio slow muscle precursors has provided novel insights into their cellular behaviours in this organism. Similarly, analyses on fast muscle precursors have provided knowledge of the behaviour of a sub-set of epitheloid cells residing in the anterior domain of somites. Additionally, the findings by various groups on the roles of several molecules in somitic myogenesis have been clarified in the past year. In this study, the authors briefly review the current trends in the field of research of D. rerio trunk myogenesis.

[Search for tissue-specific regulatory elements using Tol2 transposon as an example of evolutionary synthesis of genomics and developmental biology].

The GFP gene controlled by a mini-promoter of the gene was cloned into a modified nonautonomous transposon Tol2 from the medaka genome. Such transposon cannot itself transpose, but external transposase allows it to integrate into the germ cell genome, where it remains after transposase is depleted. This gives rise to a transgenic zebrafish line stably maintained for many generations. This approach was used to obtain several tens of transgenic zebrafish lines, each of which demonstrates stably inherited pattern of GFP expression. This pattern depends on the enhancer activity in the vicinity of the transposon integration site. These lines can be used for the analysis of DNA sequences of enhancers, in vivo studies of complex cell morphogenesis, description of organs whose development has not been described in zebrafish, and analysis of activity of genes controlled by the identified promoters during development. Current data on the transgenic lines were integrated into the dedicated ZETRAP database. At the second stage of this project, the transposon was mobilized by a short-term exposure of two transgenic lines to transposase and left the initial chromosomal location. This allowed us to isolate about 20 transgenic lines with new expression patterns. This approach can be successfully used to study the zebrafish biology and anatomy, activity of the regulatory part of the genome, and development of new promising techniques in developmental biology and, in the foreseeable future, gene therapy.

Genetic control of early neuronal development in vertebrates.

The specification of neuronal fate starts with cell commitment and determination. These early events are accompanied by rearrangement and reshaping of presumptive neural cells. Later, the neural differentiation begins, and its course can be followed using specific molecular markers. Such events take place long before the cells acquire a typical neuronal phenotype. Primary neurons of lower vertebrates differ from secondary neurons by their size, position, timing of differentiation and length of axon. Primary neurons start to express early markers of neural differentiation at the end of gastrulation. Recent data indicate that in lower vertebrates the neural induction of primary neurons differs from the induction of secondary neurons; however, neural induction in higher vertebrates appears to be similar to the induction of secondary neurons in lower vertebrates.

A new splicing variant of a type III POU gene from zebrafish encodes a POU protein with a distinct C-terminal.

A zebrafish POU protein cDNA clone was isolated and sequenced. It appears to be a novel splicing variant of the previously reported zebrafish POU gene zp-12. There are four splicing variants and at least three of them lead to different C-terminal amino acid sequences. The four splicing variants are differentially regulated during development, indicating that they may be functionally diversified.

Developmental analysis of ceruloplasmin gene and liver formation in zebrafish.

Formation of the liver in zebrafish has been analyzed during normal embryogenesis using ceruloplasmin (Cp) as a specific marker. The asymmetric expression of Cp has been detected in dorsal endoderm at 16 hpf and later in the early hepatic cells in the yolk sac. The liver primordium can be detected after 32 hpf. In oep-/- mutant, which lacks dorsal endoderm, the liver fails to form. In the notochordless flh-/- mutant, the asymmetry of the liver has been lost. Therefore the notochord, dorsal endoderm and endoderm of the yolk sac play a role in liver formation in zebrafish.

Expression of a zebrafish iroquois homeobox gene, Ziro3, in the midline axial structures and central nervous system.

We describe a zebrafish gene, Ziro3, which is highly homologous to Xenopus and mouse iroquois3. Ziro3 expression starts during gastrulation in the dorsal axial mesoderm that develops into the notochord. Later, the expression is limited to the chordo-neural hinge in the tailbud. Ziro3 expression also occurs in the central nervous system (CNS), excluding the telencephalon. The level of Ziro3 expression differs in odd and even rhombomeres. In the midbrain-hindbrain boundary (MHB) and rhombomere 6, Ziro3 transcripts appear only after the formation of the cerebellum and otic vesicle, respectively.

Expression pattern of two zebrafish genes, cxcr4a and cxcr4b.

We cloned and mapped two novel zebrafish genes, cxcr4a and cxcr4b, which are closely related to mammalian CXCR4. Expression analysis by reverse transcription-polymerase chain reaction and in situ hybridization demonstrated that these two genes are expressed in most cell lineages known to express Cxcr4 in mammals. These genes are co-expressed in lateral mesoderm and posterior midbrain. The transcripts of cxcr4a were detected in interneurons and endoderm, whereas cxcr4b was specifically expressed in sensory neurons, motoneurons and cerebellum. In the lateral mesoderm, cxcr4b transcripts appeared earlier than those of cxcr4a. Thus, the function of mammalian CXCR4 could be split between the two zebrafish genes. These genes probably derived from the genome duplication event, which occurred during the evolution of teleosts. Similar pairs of Cxcr4 may exist in other species, where genome duplication has occurred.

Expression of two novel zebrafish iroquois homologues (ziro1 and ziro5) during early development of axial structures and central nervous system.

Previously, we reported a zebrafish iroquois gene, ziro3, and its expression during early embryogenesis (Mech. Dev. 87 (1999) 165). In the present study, we have isolated two novel zebrafish iroquois genes, ziro1 and ziro5, homologs of mouse Irx1 and mouse Irx5, respectively. The expression of both genes is initiated in dorsal neuroectoderm and mesoderm during gastrulation. Later, their expression appears in the central nervous system (CNS), excluding the telencephalon and most of the diencephalon. ziro1 expression is complementary to that of ziro3 in the notochord and later in the gut. In contrast, ziro5 expression mostly overlaps with that of ziro3. Interestingly, all three iroquois zebrafish genes are expressed in the notochord while only Irx3 is active in the mouse notochord. Their expression in later stages of embryogenesis was also compared.

fgfr3 and regionalization of anterior neural tube in zebrafish.

Here we describe the isolation of the zebrafish fgfr3 gene, its structure and chromosomal location. Expression in wild type embryos occurs in the axial mesoderm, the diencephalon, the anterior hindbrain and the anterior spinal cord. In the hindbrain, a differential expression of fgfr3 was detected at several levels of intensity, with the highest expression in the posterior rhombomere 1 that is morphologically distinct from the anterior part, which develops into the cerebellum. Further, analysis of fgfr3 expression in mutants deficient in the formation of the midbrain-hindbrain boundary (MHB), noi(-/-) and ace(-/-), demonstrated that in the absence of Pax2.1 and FGF8 activity, the expression domains of FGFR3 expand into the MHB, tegmentum, cerebellum and optic tectum, which are the affected structures in these mutants.

Expression of brain subtype creatine kinase in the zebrafish embryo.

Creatine kinases (CK) play crucial roles in intracellular energy transfer. We have isolated a cDNA from zebrafish embryos, which encodes a CK highly related to the mammalian brain subtype creatine kinase (BCK). The bck mRNA is expressed maternally in the zebrafish embryo and transcripts are distributed uniformly in blastula and gastrula stages. Expression becomes restricted to the prechordal plate and the nervous system during subsequent somitogenesis stages. bck transcripts are abundant in primary neurons in the developing central nervous system of the 1-day-old embryo. While some bck expression persists in the hindbrain, expression vanishes in the spinal cord of the 2-day-old embryo. In summary, the expression pattern of bck is highly dynamic and suggests a role for bck during gastrulation and neuronal differentiation.

A novel zebrafish bHLH gene, neurogenin3, is expressed in the hypothalamus.

Many basic helix-loop-helix transcriptional factors play important roles in vertebrate neurogenesis. Among them, Neurogenins act as determination factors and initiate the expression of differentiation genes such as neuroD and other neurogenic genes. Here we describe a zebrafish cDNA (neurogenin3 or ngn3) encoding a novel member of the Neurogenin family closest to mouse Ngn3 and human NGN3. Using a zebrafish radiation hybrid panel, ngn3 was mapped to zebrafish linkage group 13 and the region displayed a conserved synteny with the region of human chromosome 10 containing NGN3. As judged by RT-PCR and whole-mount in situ hybridization, ngn3 expression in zebrafish started much later than other neurogenin genes, at only around 24 h post-fertilization (hpf) and with a higher level of expression on the left side of the anterio-ventral diencephalon. Later at 48 hpf, ngn3 expression was detected in a small number of cells in the tuberal hypothalamus. Unlike Ngn3 in the mouse, zebrafish ngn3 mRNAs were not detected in developing pancreas and spinal cord. Genomic Southern blot hybridization suggested that a closely related sequence is present in the zebrafish genome and the hypothetical gene might result from the recent genome duplication in certain teleost lineage and share the function of the common ancestor with the currently characterized ngn3.

Use of an IRES bicistronic construct to trace expression of exogenously introduced mRNA in zebrafish embryos.

To understand gene function in developing vertebrate embryos, co-injection of an mRNA for a reporter protein and an mRNA for a testing factor is widely used. However, because of the mosaic segregation of injected nucleic acids during early embryogenesis, whether both mRNAs are translated in the same cell remains uncertain. In the present study, we tested a new system of tracing the expression of a testing gene in zebrafish using an internal ribosomal entry site (IRES) to express two proteins from the same mRNA template, thus eliminating the problem of independent translation observed in co-injection essays. A DNA construct was made for synthesizing bicistronic mRNA for NeuroD, a neurogenic transcription factor, and the enhanced green fluorescent protein (EGFP) reporter. When the bicistronic mRNA for NeuroD and EGFP was injected into zebrafish embryos at one cell stage, all EGFP-expressing embryos showed ectopic expression of neuroD mRNA and the mRNA of its potential downstream gene, islet-1. Thus, the IRES bicistronic mRNA construct might be a more convincing means of analyzing gene function in developing zebrafish embryos.

A class of neuroD-related basic helix-loop-helix transcription factors expressed in developing central nervous system in zebrafish.

Neuronal basic helix-loop-helix (bHLH) transcription factors such as neuroD and neurogenin have been shown to play important roles in neuronal development. In the present study, several distinct bHLH DNA fragments were isolated from the zebrafish genomic DNA by a pair of degenerate polymerase chain reaction (PCR) primers deduced from the conserved bHLH domains of neuroD and neurogenins. Based on the bHLH fragments, three complete neuroD-related cDNA clones, including complete coding regions, ndr1a, ndr1b, and ndr2 (ndr for neuroD related), were isolated and assembled by 5' and 3' rapid amplification of cDNA ends (RACE). A phylogenetic analysis indicated the presence of four groups of neuroD-related genes in the neuroD subfamily in vertebrates: neuroD, ndr1a/ndr1b/MATH-2, ndr2/NDRF, and neuroM/MATH3. Expression of the newly isolated neuroD-related genes was examined by reverse transcriptase (RT)-PCR and whole-mount in situ hybridization. Unlike neuroD, which was expressed broadly in primary neurons during early zebrafish development starting from 10 h postfertilization (hpf), expression of ndr1a and ndr1b started relatively late (around 22 hpf) and was restricted to the olfactory system: olfactory bulbs in the telecephalon (ndr1a and ndr1b) and olfactory organs (ndr1b) starting around 22 hpf. Although a faint ndr2 mRNA signal was detected by RT-PCR in early embryos, no ndr2 mRNA was detected by whole-mount in situ hybridization in embryos up to 72 hpf, suggesting that it is expressed rather late. Our observations suggest that the two novel neuroD-related genes, ndr1a and ndr1b, are involved in the development of the olfactory system and perhaps contribute to its functional complexity.

White spot syndrome virus (WSSV) infects specific hemocytes of the shrimp Penaeus merguiensis.

White spot syndrome virus (WSSV) was specifically detected by PCR in Penaeus merguiensis hemocytes, hemolymph and plasma. This suggested a close association between the shrimp hemolymph and the virus. Three types of hemocyte from shrimp were isolated using flow cytometry. Dynamic changes of the hemocyte subpopulations in P. merguiensis at different times after infection were observed, indicating that the WSSV infection selectively affected specific subpopulations. Immunofluorescence assay (IFA) and a Wright-Giemsa double staining study of hemocyte types further confirmed the cellular localization of the virus in the infected hemocytes. Electron microscopy revealed virus particles in both vacuoles and the nucleus of the semigranular cells (SGC), as well as in the vacuoles of the granular cells (GC). However, no virus could be detected in the hyaline cells (HC). Our results suggest that the virus infects 2 types of shrimp hemocytes--GCs and SGCs. The SGC type contains higher virus loads and exhibits faster infection rates, and is apparently more susceptible to WSSV infection.