Regulation of intracellular pH by a neuronal homolog of the erythrocyte anion exchanger

We have isolated AE3, a novel gene expressed primarily in brain neurons and in heart. The predicted AE3 polypeptide shares a high degree of identity with the anion exchange and cytoskeletal binding domains of the erythrocyte band 3 protein. Expression of AE3 cDNA in COS cells leads to chronic cytoplasmic acidification and to chloride- and bicarbonate-dependent changes in intracellular pH, confirming that this gene product is an anion exchanger. Characterization of an AE3 mutant lacking the NH2-terminal 645 amino acids demonstrates that the COOH-terminal half of the polypeptide is both necessary and sufficient for correct insertion into the plasma membrane and for anion exchange activity. The NH2-terminal domain may play a role in regulating the activity of the exchanger and may be involved in the structural organization of the cytoskeleton in neurons.     

Synthesis,characterization, and electrocatalytic behaviour of hydrothermally grown nanostructured La2O3 and La2O3/K-complex

Rare earth metals play a conspicuous role in magnetic resonance imaging (MRI) for detecting cancerous cells. The alkali metal potassium is a neurotransmitter in the sodium–potassium pump in biomedical sciences. This unique property of rare earth metals and potassium drew our attention to carry forward this study. Therefore, in this work, previously synthesized potassium (K) complexes formed by the reflux of 4-N,N-dimethylaminobenzoic acid (DBA) and potassium hydroxide in methanol, and named [(μ2–4-N,N-dimethylaminobenzoate-κO)(μ2–4-N,N-dimethylaminobenzoic acid-κO)(4-N,N-dimethylaminobenzoic acid-κO) potassium(I) coordination polymer)] were treated hydrothermally with La₂O₃ nanomaterials to obtain a nanohybrid La₂O₃/K-complex. After that, the K-complex was analyzed using single-crystal X-ray diffraction and ¹H and ¹³C NMR spectroscopy. In addition, the structural and morphological properties of the as-prepared nanostructured La₂O₃/K-complex were also characterized, which involved an investigation using X-ray diffraction (XRD)spectroscopy, Fourier transform infrared (FTIR) spectroscopy, atomic force spectroscopy (AFM), transmission electron microscopy (TEM), and energy dispersive X-ray (EDX) analysis. After this, the electrochemical redox behaviour of the synthesized nanohybrid material was studied using cyclic voltammetry (CV) and differential pulse voltammetry (DPV). Therefore, the results from these studies revealed that the as-prepared material was a La₂O₃/K-complex that has a promising future role in sensing various analytes, as it showed effective electrocatalytic behaviour.     

Functional differences among nonerythroid anion exchangers expressed in a transfected human cell line

A new transient expression system has been developed to investigate the function of anion exchangers in vivo. Human 293 cells were cotransfected with AE2 or AE3 cDNA together with a plasmid encoding a cell surface marker protein. Staining of the cells with antibody directed against a cell surface epitope present in the marker protein permitted the detection of cells expressing functional anion exchangers. Intracellular pH (pHi) recording in individual transfectants loaded with the fluorescent pHi indicator, 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein, was used to determine the flux of HCO3- as a measure of Cl-/HCO3- exchange activity. Cells expressing either anion exchanger displayed significantly enhanced Cl-/HCO3- exchange activity compared with controls expressing only the marker. Transfection with either anion exchanger or with control plasmid resulted in altered intrinsic buffering capacity profiles compared with untransfected controls. Expression of either AE2 or AE3 did not result in changes in resting pHi. The activities of both AE2 and AE3 were stimulated at alkaline pHi, suggesting that an internal protonation site in AE2 and AE3 may regulate their activities. Both exchangers were inhibited reversibly and irreversibly by the anion 4,4'-diisothiocyanostilbene-2,2'-disulfonate with IC50 values of 142 and 0.43 microM for AE2 and AE3, respectively. These data indicate that structural differences in these highly conserved anion exchangers give rise to differences in affinities at the external anion binding site.     

Inhibitory effect of iron on the uptake of lead by erythrocytes.

It is well known that more than 90% of the lead found in blood is associated with the erythrocytes. The present $\text{in vitro}$ experiments show that the uptake of lead-203 by rabbit erythrocytes is inhibited by the presence of non-radioactive lead or iron or by reduction of the incubation temperature. The inhibitory effect of iron on radioactive lead uptake by erythrocytes is also demonstrable $\text{in vivo}$.When lead-203 is incorporated into erythrocytes $\text{in vitro}$, about 10% of the radioactivity is attached to the membrane and the remainder is found in the cytoplasm associated with hemoglobin and an unidentified low molecular weight intracellular component. In the presence of 25 μg/ml of added iron (Fe⁺⁺⁺) the uptake of radioactive lead by erythrocytes is reduced to 21.7±5.1% and membrane binding accounts for approximately 5% of this total. Chromatographic analyses of hemolysates show that the reduction in cytoplasmic labeling is directly related to decreased lead binding to the low molecular weight component, since hemoglobin binding remains unchanged.This work suggests that in addition to the interaction between iron and lead which occurs during the biosynthesis of heme, these metals may directly compete for specific erythrocyte binding sites.     

Two silicone nontoxic fouling release coatings: Hydrosilation cured PDMS and CaCO3 filled,ethoxysiloxane cured RTV11

Two silicone coatings have been evaluated for barnacle adhesion. One coating is an unfilled hydrosilation cured polydimethylsiloxane (PDMS) network, while the other is a room temperature vulcanized (RTV), filled, ethoxysiloxane cured PDMS elastomer, RTV11^?. The adhesion strength of one species of barnacle, Balanus eburneus, to the hydrosilation coatings is in the range of 0.37–0.60 kg cm^‐2 while the corresponding range for RTV11 is 0.64–0.90 kg cm^‐2. The easier release of B. eburneus from the hydrosilation cured network compared to RTV11 is discussed in relationship to differences in bulk and surface properties. Preliminary results suggest bulk modulus may be the most important parameter in determining barnacle adhesion strength. In light or mechanical property analysis, a re‐evaluation of surface properties and chemical stability is presented.     

Biological effects of power frequency magnetic fields: Neurochemical and toxicological changes in developing chick embryos

BACKGROUND: There are several reports that indicate a linkage between exposure to power frequency (50 - 60 Hz) magnetic fields with abnormalities in the early embryonic development of the chicken. The present study was designed to understand whether power frequency electromagnetic fields could act as an environmental insult and invoke any neurochemical or toxicological changes in developing chick embryo model. METHODS: Fertilized chicken eggs were subjected to continuous exposure to magnetic fields (50 Hz) of varying intensities (5, 50 or 100 microT) for a period of up to 15 days. The embryos were taken out of the eggs on day 5, day 10 and day 15. Neurochemical (norepinephrine and 5-hydroxytryptamine) and amino acid (tyrosine, glutamine and tryptophan) contents were measured, along with an assay of the enzyme glutamine synthetase in the brain. Preliminary toxicological investigations were carried out based on aminotransferases (AST and ALT) and lactate dehydrogenase activities in the whole embryo as well as in the liver. RESULTS: The study revealed that there was a significant increase (p < 0.01 and p < 0.001) in the level of norepinephrine accompanied by a significant decrease (p < 0.01 and p < 0.001) in the tyrosine content in the brain on day 15 following exposure to 5, 50 and 100 microT magnetic fields. There was a significant increase (p < 0.001) in glutamine synthetase activity resulting in the significantly enhanced (p < 0.001) level of glutamine in the brain on day 15 (for 100 microT only). The possible mechanisms for these alterations are discussed. Further, magnetic fields had no effect on the levels of tryptophan and 5-hydroxytryptamine in the brain. Similarly, there was no effect on the activity of either aminotransferases or lactate dehydrogenase in the whole embryo or liver due to magnetic field exposure. CONCLUSIONS: Based on these studies we conclude that magnetic field-induced changes in norepinephrine levels might help explain alterations in the circadian rhythm, observed during magnetic field stress. Also, the enhanced level of glutamine can act as a contributing factor for developmental abnormalities.     

Cysteine residues in the nucleotide binding domains regulate the conductance state of CFTR channels

Gating of cystic fibrosis transmembrane conductance regulator (CFTR) channels requires intermolecular or interdomain interactions, but the exact nature and physiological significance of those interactions remains uncertain. Subconductance states of the channel may result from alterations in interactions among domains, and studying mutant channels enriched for a single conductance type may elucidate those interactions. Analysis of CFTR channels in inside-out patches revealed that mutation of cysteine residues in NBD1 and NBD2 affects the frequency of channel opening to the full-size versus a 3-pS subconductance. Mutating cysteines in NBD1 resulted in channels that open almost exclusively to the 3-pS subconductance, while mutations of cysteines in NBD2 decreased the frequency of subconductance openings. Wild-type channels open to both size conductances and make fast transitions between them within a single open burst. Full-size and subconductance openings of both mutant and wild-type channels are similarly activated by ATP and phosphorylation. However, the different size conductances open very differently in the presence of a nonhydrolyzable ATP analog, with subconductance openings significantly shortened by ATPgammaS, while full-size channels are locked open. In wild-type channels, reducing conditions increase the frequency and decrease the open time of subconductance channels, while oxidizing conditions decrease the frequency of subconductance openings. In contrast, in the cysteine mutants studied, altering redox potential has little effect on gating of the subconductance.