TRANSMERIDIAN JET LAG AND ITS PHARMACOLOGICAL CORRECTION.

The article deals with the development and correction of acute jet lag in a flight across several time zones. The investigation had the purpose to study dynamics of subjective and objective psychophysiological parameters and demonstrate methods of prophylaxis and correction of acute jet lag due to transmerdian flights. Subjects were 8 normal volunteers (experimental group) at the age of 26-55 years flying eastward over 7 times zones. The investigation included 3 stages: baseline, preparatory (21-d course of Euricoma longifolia extraction) and main (intake of donormil, cirkadin and artificial sleep during the flight). Functional diagnostics was performed on the baseline stage, on completion of the preparatory stage and on the next day after the flight (21-22 days from the beginning of the preparatory stage). Objective and subjective methods were used to evaluate the autonomic and cardiovascular systems and mental performance. In the control group (n = 4) functional diagnostics was performed on the same days. The investigations showed the benefit of preparation for transmeridian air travel and experimentally demonstrated positive effects of the proposed pharmacological correction of acute transmeridian jet lag.

Magnetic dipole radiation tailored by substrates: numerical investigation.

Nanoparticles of high refractive index materials can possess strong magnetic polarizabilities and give rise to artificial magnetism in the optical spectral range. While the response of individual dielectric or metal spherical particles can be described analytically via multipole decomposition in the Mie series, the influence of substrates, in many cases present in experimental observations, requires different approaches. Here, the comprehensive numerical studies of the influence of a substrate on the spectral response of high-index dielectric nanoparticles were performed. In particular, glass, perfect electric conductor, gold, and hyperbolic metamaterial substrates were investigated. Optical properties of nanoparticles were characterized via scattering cross-section spectra, electric field profiles, and induced electric and magnetic moments. The presence of substrates was shown to have significant impact on particle's magnetic resonances and resonant scattering cross-sections. Variation of substrate material provides an additional degree of freedom in tailoring optical properties of magnetic multipoles, important in many applications.

SLC13 family of Na⁺-coupled di- and tri-carboxylate/sulfate transporters.

The SLC13 family comprises five genes (SLC13A1, SLC13A2, SLC13A3, SLC13A4, and SLC13A5) encoding structurally related multi-spanning transporters (8-13 transmembrane domains) with orthologues found in prokaryotes and eukaryotes. Mammalian SLC13 members mediate the electrogenic Na(+)-coupled anion cotransport at the plasma membrane of epithelial cells (mainly kidney, small intestine, placenta and liver) or cells of the central nervous system. While the two SLC13 cotransporters NaS1 (SLC13A1) and NaS2 (SLC13A4) transport anions such sulfate, selenate and thiosulfate, the three other SLC13 members, NaDC1 (SLC13A2), NaCT (SLC13A5) and NaDC3 (SLC13A3), transport di- and tri-carboxylate Krebs cycle intermediates such as succinate, citrate and α-ketoglutarate. All these transporters play a variety of physiological and pathophysiological roles in the different organs. Thus, the purpose of this review is to summarize the roles of SLC13 members in human physiology and pathophysiology and what the therapeutic perspectives are. We have also described the most recent advances on the structure, expression, function and regulation of SLC13 transporters.

WITHDRAWN: NaS1 sulfate transporter is linked to hyposulfatemia and longevity.

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Reduced mucin sulfonation and impaired intestinal barrier function in the hyposulfataemic NaS1 null mouse.

OBJECTIVE: Sulfate (SO(4)(2-)) is an abundant component of intestinal mucins and its content is decreased in certain gastrointestinal diseases, including inflammatory bowel disease. In this study, the hyposulfataemic NaS1 sulfate transporter null (Nas1(-/-)) mice were used to investigate the physiological consequences of disturbed sulfate homeostasis on (1) intestinal sulfomucin content and mRNA expression; (2) intestinal permeability and proliferation; (3) dextran sulfate sodium (DSS)-induced colitis; and (4) intestinal barrier function against the bacterial pathogen, Campylobacter jejuni. METHODS: Intestinal sulfomucins and sialomucins were detected by high iron diamine staining, permeability was assessed by fluorescein isothiocyanate (FITC)-dextran uptake, and proliferation was assessed by 5-bromodeoxyuridine (BrdU) incorporation. Nas1(-/-) and wild-type (Nas1(+/+)) mice received DSS in drinking water, and intestinal damage was assessed by histological, clinical and haematological measurements. Mice were orally inoculated with C jejuni, and intestinal and systemic infection was assessed. Ileal mRNA expression profiles of Nas1(-/-) and Nas1(+/+) mice were determined by cDNA microarrays and validated by quantitative real-time PCR. RESULTS: Nas1(-/-) mice exhibited reduced intestinal sulfomucin content, enhanced intestinal permeability and DSS-induced colitis, and developed systemic infections when challenged orally with C jejuni. The transcriptional profile of 41 genes was altered in Nas1(-/-) mice, with the most upregulated gene being pancreatic lipase-related protein 2 and the most downregulated gene being carbonic anhydrase 1 (Car1). CONCLUSION: Sulfate homeostasis is essential for maintaining a normal intestinal metabolic state, and hyposulfataemia leads to reduced intestinal sulfomucin content, enhanced susceptibility to toxin-induced colitis and impaired intestinal barrier to bacterial infection.

Development of an HPLC-MS/MS method for the selective determination of paracetamol metabolites in mouse urine.

An HPLC-MS/MS method has been developed for the selective quantitative analysis of paracetamol and its two major metabolites. The use of tandem MS enabled the detection and quantitation of metabolites in small sample sizes with high sensitivity and selectivity. Isocratic elution using acetonitrile and water containing formic acid combined with electrospray-tandem MS enabled the separation and accurate quantitation of each analyte and the internal standard 3-acetamidophenol. The on-column limits of detection for paracetamol, paracetamol sulfate, and paracetamol glucuronide were 2.4, 1.2, and 1.2 pmol, respectively. The method was applied to quantitate paracetamol and its metabolites in mouse urine. It is highly specific, sensitive, and easily adaptable to measure these analytes in biological fluids of other animals.

Pathogenetics of the human SLC26 transporters.

Over the past decade, 11 human genes belonging to the solute linked carrier (SLC) 26 family of transporters, have been identified. The SLC26 proteins, which include SAT-1, DTDST, DRA/CLD, pendrin, prestin, PAT-1/CFEX and Tat-1, are structurally related and have been shown to transport one or more of the following substrates: sulfate, chloride, bicarbonate, iodide, oxalate, formate, hydroxyl or fructose. Special interest has focused on four members of the SLC26 family that are associated with distinct recessive diseases: (i) Mutations in SLC26A2 lead to four different chondrodysplasias (diastrophic dysplasia, atelosteogenesis type II, achondrogenesis type IB and multiple epiphyseal dysplasia); (ii) SLC26A3 is associated with congenital chloride diarrhea; (iii) SLC26A4 is associated with Pendred syndrome and non-syndromic deafness, DFNB4; and (iv) SLC26A5 is defective in non-syndromic hearing impairment. This review article summarizes current information on the pathophysiological consequences of mutations in the human SLC26A2 to A5 genes.

Purinergic inhibition of the epithelial Na+ transport via hydrolysis of PIP2.

Stimulation of purinergic receptors inhibits amiloride-sensitive Na+ transport in epithelial tissues by an unknown mechanism. Because previous studies excluded the role of intracellular Ca2+ or protein kinase C, we examined whether purinergic regulation of Na+ absorption occurs via hydrolysis of phospholipid such as phosphatidylinositol-bisphosphates (PIP2). Inhibition of amiloride-sensitive short-circuit currents (Isc-Amil) by adenine 5'-triphosphate (ATP) in native tracheal epithelia and M1 collecting duct cells was suppressed by binding neomycin to PIP2, and recovery from ATP inhibition was abolished by blocking phosphatidylinositol-4-kinase or diacylglycerol kinase. Stimulation by ATP depleted PIP2 from apical membranes, and PIP2 co-immunoprecipitated the beta subunit of ENaC. ENaC was inhibited by ATP stimulation of P2Y2 receptors in Xenopus oocytes. Mutations in the PIP2 binding domain of betaENaC but not gammaENaC reduced ENaC currents without affecting surface expression. Collectively, these data supply evidence for a novel and physiologically relevant regulation of ENaC in epithelial tissues. Although surface expression is controlled by its C terminus, N-terminal binding of betaENaC to PIP2 determines channel activity.

Effects of purinergic stimulation, CFTR and osmotic stress on amiloride-sensitive Na+ transport in epithelia and Xenopus oocytes.

Both stimulation of purinergic receptors by ATP and activation of the cystic fibrosis transmembrane conductance regulator (CFTR) inhibit amiloride-sensitive Na+ transport and activate Cl- secretion. These changes in ion transport may well affect cell volume. We therefore examined whether cell shrinkage or cell swelling do affect amiloride-sensitive Na+ transport in epithelial tissues or Xenopus oocytes and whether osmotic stress interferes with regulation of Na+ transport by ATP or CFTR. Stimulation of purinergic receptors by ATP/UTP or activation of CFTR by IBMX and forskolin inhibited amiloride-sensitive transport in mouse trachea and colon, respectively, by a mechanism that was Cl- dependent. When exposed to a hypertonic but not hypotonic bath solution, amiloride-sensitive Na+ transport was inhibited in mouse trachea and colon, independent of the extracellular Cl- concentration. Both inhibition of Na+ transport by hypertonic bath solution and ATP were additive. When coexpressed in Xenopus oocytes, activation of CFTR by IBMX and forskolin inhibited the epithelial Na+ channel (ENaC) in a Cl- dependent fashion. However, both hypertonic and hypotonic bath solutions showed only minor effects on amiloride-sensitive conductance, independent of the bath Cl- concentration. Moreover, CFTR-induced inhibition of ENaC could be detected in oocytes even after exposure to hypertonic or hypotonic bath solutions. We conclude that amiloride-sensitive Na+ absorption in mouse airways and colon is inhibited by cell shrinkage by a mechanism that does not interfere with purinergic and CFTR-mediated inhibition of ENaC.

Regulation of the mouse Nas1 promoter by vitamin D and thyroid hormone.

The renal sodium-sulfate cotransporter, NaS(i)-1, a protein implicated to control serum sulfate levels, has been shown to be regulated in vivo by 1,25-dihydroxyvitamin D(3) (1,25-(OH)(2)D(3)) and tri-iodothyronine (T(3)). Recently, we cloned the mouse NaS(i)-1 gene ( Nas1) and in the present study identified a 1,25-(OH)(2)D(3)- and T(3)-responsive element located within the Nas1 promoter. Mutational analysis of the Nas1 promoter resulted in identification of a direct repeat 6-type vitamin-D-responsive element (DR6 VDRE) at -525 to -508 and an imperfect inverted repeat 0-type T(3)-responsive element (IR0 T(3)RE) at -436 to -425 which conferred 1,25-(OH)(2)D(3) and T(3) responsiveness, respectively. In summary, we have identified responsive elements that mediate the enhanced transcription of Nas1 by 1,25-(OH)(2)D(3) and T(3), and these mechanisms may provide important clues to the physiological control of sulfate homeostasis.

Physiological roles and regulation of mammalian sulfate transporters.

All cells require inorganic sulfate for normal function. Sulfate is among the most important macronutrients in cells and is the fourth most abundant anion in human plasma (300 microM). Sulfate is the major sulfur source in many organisms, and because it is a hydrophilic anion that cannot passively cross the lipid bilayer of cell membranes, all cells require a mechanism for sulfate influx and efflux to ensure an optimal supply of sulfate in the body. The class of proteins involved in moving sulfate into or out of cells is called sulfate transporters. To date, numerous sulfate transporters have been identified in tissues and cells from many origins. These include the renal sulfate transporters NaSi-1 and sat-1, the ubiquitously expressed diastrophic dysplasia sulfate transporter DTDST, the intestinal sulfate transporter DRA that is linked to congenital chloride diarrhea, and the erythrocyte anion exchanger AE1. These transporters have only been isolated in the last 10-15 years, and their physiological roles and contributions to body sulfate homeostasis are just now beginning to be determined. This review focuses on the structural and functional properties of mammalian sulfate transporters and highlights some of regulatory mechanisms that control their expression in vivo, under normal physiological and pathophysiological states.

Glycosphingolipids modulate renal phosphate transport in potassium deficiency.

BACKGROUND: Potassium (K) deficiency (KD) and/or hypokalemia have been associated with disturbances of phosphate metabolism. The purpose of the present study was to determine the cellular mechanisms that mediate the impairment of renal proximal tubular Na/Pi cotransport in a model of K deficiency in the rat. METHODS: K deficiency in the rat was achieved by feeding rats a K-deficient diet for seven days, which resulted in a marked decrease in serum and tissue K content. RESULTS: K deficiency resulted in a marked increase in urinary Pi excretion and a decrease in the V(max) of brush-border membrane (BBM) Na/Pi cotransport activity (1943 +/- 95 in control vs. 1184 +/- 99 pmol/5 sec/mg BBM protein in K deficiency, P < 0.02). Surprisingly, the decrease in Na/Pi cotransport activity was associated with increases in the abundance of type I (NaPi-1), and type II (NaPi-2) and type III (Glvr-1) Na/Pi protein. The decrease in Na/Pi transport was associated with significant alterations in BBM lipid composition, including increases in sphingomyelin, glucosylceramide, and ganglioside GM3 content and a decrease in BBM lipid fluidity. Inhibition of glucosylceramide synthesis resulted in increases in BBM Na/Pi cotransport activity in control and K-deficient rats. The resultant Na/Pi cotransport activity in K-deficient rats was the same as in control rats (1148 +/- 52 in control + PDMP vs. 1152 +/- 61 pmol/5 sec/mg BBM protein in K deficiency + PDMP). These changes in transport activity occurred independent of further changes in BBM NaPi-2 protein or renal cortical NaPi-2 mRNA abundance. CONCLUSION: K deficiency in the rat causes inhibition of renal Na/Pi cotransport activity by post-translational mechanisms that are mediated in part through alterations in glucosylceramide content and membrane lipid dynamics.

Rapid GLUT-1 mediated glucose transport in erythrocytes from the grey-headed fruit bat (Pteropus poliocephalus).

D-Glucose entry into erythrocytes from adult grey-headed flying fox fruit bats (Pteropus poliocephalus) was rapid and showed saturation at high substrate concentrations. Kinetic parameters were estimated from the concentration dependence of initial rates of zero-trans D-glucose entry at 5.5 degrees C as Michaelis constant (K(m)) 1. 64+/-0.56 mM, and maximal velocity (V(max)) 1162+/-152 micromol.l. cell water(-1).min(-1). D-Glucose entry was inhibited by cytochalasin B; mass law analysis of D-glucose-displaceable cytochalasin B binding gave values of K(d) 37.1+/-5.0 nM and B(max) 361.2+/-9.1 pmol/mg membrane protein. Entry of 2-deoxy-D-glucose, and 3-O-methyl-D-glucose, into P. poliocephalus red cells was rapid, entry of D-fructose was very slow. Glucose transporter polypeptides were identified on immunoblots as a band M(r) 47000-54000 and their identity confirmed by D-glucose-sensitive photolabeling of membranes with [3H]-cytochalasin B. Peptide-N-glycanase F digestion of both human and bat erythrocyte membranes generated GLUT-1-derived bands M(r) 37000. Trypsin digestion of human and fruit bat erythrocyte membranes generated fragmentation patterns consistent with similar GLUT-1 polypeptide structures in both species. Erythrocytes from adult Australian ghost bats (Macroderma gigas), a carnivorous microchiropteran bat, also expressed high levels of GLUT-1.

The mouse Na(+)-sulfate cotransporter gene Nas1. Cloning, tissue distribution, gene structure, chromosomal assignment, and transcriptional regulation by vitamin D.

NaSi-1 is a Na(+)-sulfate cotransporter expressed on the apical membrane of the renal proximal tubule and plays an important role in sulfate reabsorption. To understand the molecular mechanisms that mediate the regulation of NaSi-1, we have isolated and characterized the mouse NaSi-1 cDNA (mNaSi-1), gene (Nas1), and promoter region and determined Nas1 chromosomal localization. The mNaSi-1 cDNA encodes a protein of 594 amino acids with 13 putative transmembrane segments, inducing high affinity Na(+)-dependent transport of sulfate in Xenopus oocytes. Three different mNaSi-1 transcripts derived from alternative polyadenylation and splicing were identified in kidney and intestine. The Nas1 gene is a single copy gene comprising 15 exons spread over 75 kilobase pairs that maps to mouse chromosome 6. Transcription initiation occurs from a single site, 29 base pairs downstream to a TATA box-like sequence. The promoter is AT-rich (61%), contains a number of well characterized cis-acting elements, and can drive basal transcriptional activity in opossum kidney cells but not in COS-1 or NIH3T3 cells. We demonstrated that 1,25-dihydroxyvitamin D(3) stimulated the transcriptional activity of the Nas1 promoter in transiently transfected opossum kidney cells. This study represents the first characterization of the genomic organization of a Na(+)-sulfate cotransporter gene. It also provides the basis for a detailed analysis of Nas1 gene regulation and the tools required for assessing Nas1 role in sulfate homeostasis using targeted gene manipulation in mice.

The human renal sodium sulfate cotransporter (SLC13A1; hNaSi-1) cDNA and gene: organization, chromosomal localization, and functional characterization.

Sulfate plays an essential role during growth, development, bone/cartilage formation, and cellular metabolism. In this study, we have determined the structure of the human Na+-sulfate cotransporter (hNaSi-1) cDNA (Human Genome Nomenclature Committee-approved symbol SLC13A1) and gene (NAS1). hNaSi-1 encodes a protein of 595 amino acids with 13 putative transmembrane domains. hNaSi-1 mRNA expression was exclusive to the human kidney. Expression of hNaSi-1 protein in Xenopus oocytes demonstrated a high-affinity Na+-sulfate cotransporter that was inhibited by selenate, thiosulfate, molybdate, tungstate, citrate, and succinate. Antisense inhibition experiments suggest hNaSi-1 to represent the major Na+-sulfate cotransporter in the human kidney. NAS1 was localized on human chromosome 7, mapped to 7q31-q32, near the sulfate transporter genes, DRA and SUT-1. The NAS1 gene contains 15 exons, spanning over 83 kb in length. Knowledge of the structure, function, and chromosomal localization of hNaSi-1 will permit the screening of NAS1 mutations in humans with disorders in renal sulfate reabsorption and homeostasis.

Ontogeny of renal sulfate transporters: postnatal mRNA and protein expression.

Renal reabsorption of inorganic sulfate changes during growth and development. We have studied the expression of two proximal tubular sulfate transporters, NaS(i)-1 and sat-1, during postnatal maturation. Both NaS(i)-1 and sat-1 mRNA levels increase postnatally, peaking at day 14, with a maximal tenfold increase in NaS(i)-1 and sat-1 mRNA levels compared with day 1. A similar age-dependent increase was observed for sodium-dependent and sodium-independent sulfate uptakes by injection of rat kidney mRNA into Xenopus oocytes. Western blot analysis of renal membranes from rats aged day 1 to day 77 showed no significant changes for NaS(i)-1 protein, whereas sat-1 protein levels paralleled mRNA expression, increasing from day 1 to day 14, followed by a decrease in protein levels thereafter. For NaSi-1 protein expression, translational or posttranslational mechanisms may maintain equal numbers of sulfate proteins on proximal tubular membranes during maturation, whereas sat-1 protein levels are in close agreement with mRNA changes. This is the first study to look at the ontogeny of renal sulfate transporter (NaS(i)-1 and sat-1) expression during postnatal maturation.

Identification of a mammalian brain sulfate transporter.

Sulfate is an essential anion involved in many biosynthetic and pharmacological reactions. Sulfate is an important constituent of myelin membranes in the brain; however, very little is known as to how sulfate enters brain cells. In this study, our aim was to determine whether the mammalian brain possesses a sulfate transporter. Injection of rat brain poly A(+) RNA into Xenopus oocytes led to an induction of Na(+)-independent sulfate transport, which was inhibited by oxalate, probenecid, phenol red, thiosulfate and DIDS. Hybrid depletion using sat-1 antisense oligodeoxyribonucleotides led to a complete inhibition of brain mRNA-induced sulfate transport in Xenopus oocytes, suggesting the presence of a functional sat-1 transcript in the brain. By RT-PCR, sat-1 mRNA was detected throughout the rat brain and in situ hybridisation showed highest sat-1 expression in the hippocampus and cerebellum. This is the first study to identify and characterise a functional mammalian brain sulfate transporter.

Expression of membrane transporters in cane toad Bufo marinus oocytes.

Membrane transport proteins (transporters and ion channels) have been extensively expressed in amphibian oocytes. The aims of this study were to determine whether oocytes from the cane toad Bufo marinus could be used as an alternative expression system to the broadly used Xenopus laevis oocytes. mRNAs encoding plasma membrane transporters NaSi-1 and sat-1 (sulphate transporters), NaDC-1 (dicarboxylate transporter), SGLT-1 (Na(+)/glucose cotransporter) and rBAT and 4F2 hc (amino acid transporters) were injected into B. marinus oocytes. All led to significant induction of their respective transport activities. Uptake rates were comparable with those in X. laevis oocytes, with the exception of rBAT, which was able to induce amino acid uptake only in X. laevis oocytes, suggesting that rBAT may require an endogenous X. laevis oocyte protein that is absent from B. marinus oocytes. Transport kinetics were determined for the NaSi-1 cotransporter in B. marinus oocytes, with identical results to those obtained in X. laevis oocytes. NaSi-1 specificity for the Na(+) cation was determined, and the anions selenate, molybdate, tungstate, oxalate and thiosulphate could all inhibit NaSi-1-induced sulphate transport. This study demonstrates that cane toad oocytes can be used successfully to express plasma membrane proteins, making this a viable heterologous system for the expression of proteins.

Metabolic acidosis regulates rat renal Na-Si cotransport activity.

Recently, we cloned a cDNA (NaSi-1) localized to rat renal proximal tubules and encoding the brush-border membrane (BBM) Na gradient-dependent inorganic sulfate (Si) transport protein (Na-Si cotransporter). The purpose of the present study was to determine the effect of metabolic acidosis (MA) on Na-Si cotransport activity and NaSi-1 protein and mRNA expression. In rats with MA for 24 h (but not 6 or 12 h), there was a significant increase in the fractional excretion of Si, which was associated with a 2.4-fold decrease in BBM Na-Si cotransport activity. The decrease in Na-Si cotransport correlated with a 2.8-fold decrease in BBM NaSi-1 protein abundance and a 2.2-fold decrease in cortical NaSi-1 mRNA abundance. The inhibitory effect of MA on BBM Na-Si cotransport was also sustained in rats with chronic (10 days) MA. In addition, in Xenopus laevis oocytes injected with mRNA from kidney cortex, there was a significant reduction in the induced Na-Si cotransport in rats with MA compared with control rats, suggesting that MA causes a decrease in the abundance of functional mRNA encoding the NaSi-1 cotransporter. These findings indicate that MA reduces Si reabsorption by causing decreases in BBM Na-Si cotransport activity and that decreases in the expression of NaSi-1 protein and mRNA abundance, at least in part, play an important role in the inhibition of Na-Si cotransport activity during MA.