Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Chlorides”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 919 records · Page 51Linked to original sources
PubMedDate not supplied

Comparison of the Volhard and potentiometric methods for the determination of chloride in meat products: collaborative study.

A collaborative study of the determination of chloride in meat products was conducted by the International Organization for Standardization (ISO) to compare the ISO 1841 method (Volhard titration) with the FAO/WHO Codex Alimentarius Committee method (potentiometric titration). Five canned luncheon meat products containing 0.25-2.0% sodium chloride at 4 different spiking levels were analyzed by 11 laboratories. The data were analyzed by ISO statistics (ISO 5725) and by AOAC statistics (Youden-Steiner), the major differences being in the rejection of outliers and in the statement of precision parameters. Good agreement was found between the mean chloride contents of the products as determined by both methods and with the added amounts, although statistically significantly higher sodium chloride recoveries were obtained with the potentiometric method. The within-laboratory variability (repeatability) is greater for the Volhard method, especially for chloride levels below 1.0%. Therefore it is proposed to set the lowest level of determination for the Volhard method at about 1.0% sodium chloride. The among-laboratories variability (reproducibility) of the potentiometric method was comparable with the results from the collaborative studies for chloride in cheese, giving acceptable values for relative standard deviations of 1.5-3.0% for meat products with 0.3-2.0% added sodium chloride. It is recommended that further work be conducted to reduce or eliminate the systematic error present with the potentiometric method as applied to meat and meat products.

Animals↗
PubMed1995

Volume-sensitive chloride channels associated with human cervical carcinogenesis.

Previous controversy has risen from the purported equivalence of the volume-sensitive chloride channels with P-glycoprotein. The aim of this study was to investigate the association between expression of volume-sensitive Cl- channels and the process of malignant transformation of cervical epithelial cells. We studied the activations of volume-sensitive and cAMP-mediated chloride currents in various human cervical squamous cells that were representative of different stages of cervical carcinogenesis, i.e., normal cervical epithelium, low-grade cervical intraepithelial neoplasia, carcinoma in situ, and invasive carcinoma using the whole-cell patch clamp technique. The volume-sensitive chloride channels, however, were significantly activated only in the four cervical cancer cell lines, primary culture cells of carcinoma in situ, and invasive cancer of the cervix. The expression of volume-sensitive chloride currents was independent of the state of human papillomavirus positivity. When these cells were exposed to hypotonic shock, the cells swelled, and outward rectified chloride currents were observed. These effects were readily reversed by returning the cells to isotonic medium. In addition, 4,4'-diisothiocyanatostilbene-2,2-disulfonic acid, 1,9-dideoxyforskolin, and verapamil reversibly abolished the volume-sensitive Cl- currents. In contrast, none of the cells from normal cervices and human papillomavirus-immortalized cell lines, the in vitro equivalent of low-grade cervical intraepithelial neoplasia, developed substantial chloride currents on exposure to hypotonicity. cAMP-mediated chloride currents were ubiquitously activated in all cervical squamous cells, regardless of the stages of carcinogenesis. This is the first report suggesting an in vivo association between the development of volume-sensitive chloride currents and human carcinogenesis.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1963

FINE STRUCTURE OF CHLORIDE CELLS FROM THREE SPECIES OF FUNDULUS.

A MORPHOLOGICAL BASIS FOR OSMOREGULATION IN THE TELEOSTS WAS STUDIED BY COMPARING THE FINE STRUCTURE OF CHLORIDE CELLS FOUND IN EPITHELIA OF THE GILLS OF THREE SPECIES OF FISH: Fundulus heteroclitus which can survive in a wide range of salinities, and F. similis and F. chrysotus which are usually restricted to salt water and fresh water environments, respectively. Gills were removed from F. heteroclitus which had been laboratory adapted to either sea water or pond water. For a comparison, gills were also removed from the marine F. similis and the fresh water F. chrysotus which had been adapted to their natural environments. Gill-filaments were fixed in Millonig's phosphate buffered (pH 7.4), 1 per cent osmium tetroxide and were embedded in Epon. Thin sections of filaments were stained with lead hydroxide. The cytoplasm of chloride cells of all three species of Fundulus is heavily populated with mitochondria and is filled with tubules of the agranular endoplasmic reticulum (ER). An orderly secretory cycle was indicated for chloride cells of salt water adapted F. heteroclitus and the marine F. similis. An amorphous material is observed in the agranular ER. Its density increases towards the apical end of the cell. In the apical cytoplasm, tubules of the agranular ER appear to converge and to discharge the amorphous material into an apical cavity. Except for the actual opening of the apical cavity, the distal end of salt water adapted chloride cells is characteristically shielded from the hypertonic environment by thin cytoplasmic flanges projecting from the neighboring epithelial cells. Chloride cells of the fresh water F. chrysotus resemble chloride cells of pond water adapted F. heteroclitus, in that these cells do not have apical cavities with the functional appearance of those in the sea water adapted forms. The distal end of fresh water adapted chloride cells is typically exposed to the free surface of the gill-filament. The possible function of the cell type is discussed.

Adaptation, Biological↗
PubMedDate not supplied

In vivo and in vitro studies on the genotoxicity of cadmium chloride in mice.

The genotoxic effect of cadmium chloride was evaluated in chromosomes of experimental mice using in vivo and in vitro studies. In vivo the induction of micronuclei, sister chromatid exchange in mouse bone marrow and chromosomal aberrations in both somatic and germ cells was investigated. Doses 1.9, 5.7 and 7.6 mg kg(-1) body wt. (single i.p. treatment) induced a significant and dose-dependent increase in the percentage of polychromatic erythrocytes with micronuclei. Such a percentage reached 2.1% with the highest tested dose, compared with 0.57% for the control (non-treated) and 2.2% for mitomycin c as the positive control. The dose of 1.9 mg kg(-1) body wt. had no significant effect with respect to sister chromatid exchange (SCE) but the doses of 5.7 and 7.6 mg kg(-1)body wt. increased the frequency of SCEs significantly. The frequency of SCE reached 7.35 +/- 0.26 per cell after treatment with the highest tested dose, which is a less than twofold increase compared with the control frequency of 4.6 +/- 0.42 per cell. However mitomycin c induced a much higher effect (12.1 +/- 0.73). Cadmium chloride also induced a significant increase in the percentage of chromosomal aberrations in mouse bone marrow at the doses of 5.7 and 9.5 mg kg(-1) body wt. (single i.p. treatment). The effect is a function of cadmium chloride concentration. Moreover, cadmium chloride induced its maximum effect concerning the induction of chromosomal aberrations in mouse bone marrow cells 24 h after treatment, compared with 12 and 48 h. In germ cells, chromosomal aberrations were observed in mouse spermatocytes 12 days post-treatment with the dose of 5.7 mg kg(-1) body wt. Moreover, a pronounced reduction in the number of spermatocytes was observed after administration of cadmium chloride (0.9, 1.9 and 5.7 mg kg(-1) body wt.) In in vitro studies, the three tested concentrations of 10, 15 and 20 microgram ml(-1) cadmium chloride induced a statistically significant increase in the frequency of SCEs in cultured mouse spleen cells. The concentrations of 15 and 20 microgram ml(-1) also induced chromosomal aberrations in mouse spleen culture. The ability of vitamin C (l-ascorbic acid) to minimize the incidence of chromosomal aberrations induced by cadmium chloride in cultured mouse spleen cells was investigated. Vitamin C at the concentrations of 3 and 6 microgram ml(-1) significantly minimized the percentage of aberrant cells induced by cadmium chloride.

Animals↗
PubMed1987

Two types of chloride cells in the gill epithelium of a freshwater-adapted euryhaline fish: Lebistes reticulatus; their modifications during adaptation to saltwater.

Two types of chloride cells were identified in the gill epithelium of freshwater-adapted guppies. One type, referred to as an "alpha-chloride cell," was a pale, elongated cell located at the base of the secondary lamella in close contact with the arterioarterial pillar capillaries. In its cytoplasm, membranous tubules in continuity with its basolateral plasma membrane formed an extended tridimensional network. The vesiculotubular system (Pisam: Anat. Rec. 200:401-414, 1981) consisted of a few tubules and vesicles located next to the apical plasma membrane. A second type, referred to as a "beta-chloride cell," was a darker, ovoid cell located in the interlamellar region of the primary epithelium facing the central venous sinus. Membranous tubules in continuity with the basolateral plasma membrane were unevenly distributed in the cytoplasm. A prominent vesiculotubular system composed of numerous vesicles and tubules was found between the Golgi apparatus and the apical surface. During seawater adaptation, the alpha-chloride cells increased in size and progressively transformed into characteristic "seawater alpha-chloride cells" with a well-developed, regular, tight tubular network and numerous vesicles and tubules of the vesiculotubular system accumulated below the apical pit. The beta-chloride cells underwent a progressive degeneration and disappeared. Thus, in freshwater-adapted guppies, there are two types of chloride cells, alpha and beta, respectively, related to the arterial and the venous vessels, whereas in seawater-adapted fishes, a single type of cell, the alpha-chloride cell, was related to both the arterial and venous channels.

Adaptation, Physiological↗
PubMed1979

Comparison of chloride transport in mouse erythrocytes and Friend virus-transformed erythroleukemic cells.

Friend erythroleukemic cells, which grow continuously in tissue culture, resemble in many respects early precursors of mouse erythrocytes. To determine whether or not the membranes of these cells exhibit the rapid and selective exchange of chloride, a specialized feature of the mature erythrocyte membrane, anion fluxes were compared in Friend cells and mouse erythrocytes. The chloride flux in Friend cells at 37 degrees C was about 800-fold lower than in mouse erythrocytes (extrapolated from data at lower temperatures). This difference could not be accounted for by the somewhat lower chloride concentration in Friend cells relative to erythrocytes. Comparison of chloride and sulfate fluxes revealed that the Friend cells had over a 1,000-fold lower selectivity for chloride versus sulphate than did the mouse red cells. The temperature dependence of chloride fluxes in Friend cells corresponded to an Arrhenius activation energy of 17.9 kcal/mol, in contrast to over 30 kcal/mol for mature red cells. The chloride flux in Friend cells was also 10-fold less sensitive to the inhibitor, furosemide, than was the flux in mature red cells. The selective chloride exchange system of the mature erythrocyte therefore does not seem to be functional at the stage represented by the Friend cell, and must appear at some later stage of erythroid maturation.

Animals↗
PubMed1989

Beta-adrenergic control of cell volume and chloride transport in an established rat submandibular cell line.

Rat submandibular cells treated with methylcholanthrene are able to be propagated in continuous culture while retaining beta-adrenergic responsiveness. A specific clone, RSMT-A5, has been isolated and studied in detail. RSMT-A5 cells possess beta-adrenergic receptors (BARS) as judged by [3H]-dihydroalprenolol ([3H]-DHA) binding studies. [3H]-DHA binds to RSMT-A5 membranes in a specific and saturable manner with respect to time and [3H]-DHA concentration. Specific binding is saturable within three min of incubation, and a Scatchard analysis reveals a single class of high affinity binding sites with an equilibrium dissociation constant of 0.62 +/- 0.03 nM and a receptor density of 101 +/- 4 fmole/mg protein. Antagonist competition studies indicate that the BARs are primarily of the beta 2-subtype. The BARs are functional since isoproterenol stimulation results in an increased intracellular cAMP content, marked morphological change, and decreased cell volume and chloride content. These same responses can be evoked by treating RSMT-A5 cells with 8-bromo-cAMP. Ion transport inhibitors such as bumetanide (an inhibitor of Na/K/Cl cotransport), SITS and DIDS (inhibitors of chloride-bicarbonate exchange), amiloride (an inhibitor of Na-H exchange), ouabain (an inhibitor of Na/K-ATPase), and dipyridamole and 9-anthracene carboxylic acid (chloride channel blockers) fail to inhibit the isoproterenol-stimulated change in chloride content. The effects of either isoproterenol or 8-bromo-cAMP on both chloride content and cell volume can be inhibited by the chloride channel blocker N-phenylanthranilic acid, however. Taken together, our results indicate that RSMT-A5 cells possess a beta-adrenergic receptor system which controls intracellular volume and chloride content by modulating transport processes that are 1) cAMP-responsive and 2) inhibitable by the putative chloride channel blocker N-phenylanthranilic acid.

Adrenergic beta-Agonists↗
PubMed1995

Effects of mutations in cAMP-dependent protein kinase on chloride efflux in Caco-2 human colonic carcinoma cells.

In order to evaluate the importance of cAMP and cAMP-dependent protein kinase (cAMPdPK) in the regulation of chloride efflux via the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel, Caco-2, human colonic carcinoma cells were transfected with an expression vector encoding a mutant form of regulatory subunit of cAMPdPK under control of the mouse metallothionein 1 promoter. Four stable transformants were isolated that expressed the mutant subunit in a Zn(2+)-inducible manner and exhibited Zn(2+)-inducible inhibition of cAMPdPK activity. The parental and transformed Caco-2 cells were examined for their abilities to regulate chloride efflux in response to various secretagogues using a radioactive iodide-efflux assay. In the transformants, induction of the protein kinase mutation with ZnSO4 markedly decreased chloride efflux in response to forskolin, the 8-(4-chlorophenylthio) analog of cAMP, vasoactive intestinal polypeptide, prostaglandin E2 and isoproterenol, whereas Zn(2+)-treated parental cells remained responsive to these secretagogues. Treatment with carbachol, calcium ionophores or phorbol ester did not acutely affect chloride efflux. Together, these studies indicate that cAMP and cAMPdPK are essential components of secretagogue-regulated chloride channel activity in the Caco-2 cell line. In whole cell patch clamp recordings, induction of the cAMPdPK mutation inhibited anionic conductances indicative of the CFTR chloride channel, whereas purified catalytic subunit of cAMPdPK, added intracellularly, reversed the inhibition. These latter results demonstrate that the CFTR chloride channels in the protein kinase-defective transformants are normal and that the protein kinase mutation specifically affects their regulation, presumably by direct phosphorylation.

Carbachol↗
PubMed1977

Killifish opercular skin: a flat epithelium with a high density of chloride cells.

In teleosts the head region, particularly the gills, plays the key role in osmoregulatory NaCl transport, presumably by mechanisms located in the chloride cell. As interest has focused on specific mechanisms of chloride cell function, two classical preparations, the intact fish and the isolated, perfused gill, have continued to serve as the only available model systems. However, both of these preparations have severe limitations, e.g., as they are not flat sheets, they cannot be studied readily under the ideal thermodynamic conditions achieved with the short-circuit current technique. The present sutdy describes the histology and ultrastructure of a particular area of skin in the head region of both killifish (Fundulus heteroclitus) and sea raven (Hemitripterus americanus). This skin lies on the inside of the operculum and possesses a flat epithelium containing chloride cells. In the present study the identity of the chloride cell in this epithelium was definitively established with the electron microscope. Although the opercular epithelium from the marine sea raven contains few chloride cells, that from the euryhaline killifish adapted to pond water, 100%-, and 200% artificial seawater is predominately chloride cells. Significantly, the teleost gill has never been reported to contain more than 10% chloride cells. Thus the opercular skin of the killifish can serve as a model to study the adaptive role of chloride cells in euryhaline teleosts. A separate electrophysiological study has deminstrated that the short-circuit current technique can be applied to this skin.

Animals↗
PubMed1979

Ultrastructure of chloride cells in young adults of the anadromous sea lamprey, Petromyzon marinus L., in fresh water and during adaptation to sea water.

The chloride cells in the interlamellar areas of the gills of young adult, anadromous sea lampreys, Petromyzon marinus L., captured in fresh water undergo structural modification during the adaptation of these animals to sea water. In fresh water the chloride cells are partially overlapped by mucus-secreting superficial cells and contain an extensive reticulum of cytoplasmic tubules, which are confluent with both lateral and basal plasma membranes, numerous mitochondria, a Golgi complex of moderate size, and numerous apical vesicles. Adaptation to sea water results in a retraction of the superficial cells, exposing the entire apical surface of the chloride cells, and a proliferation of both cytoplasmic tubules and mitochondria. Extensive enlargement of the Golgi complex in the chloride cells of these animals suggests the involvement of this organelle in the proliferation of cytoplasmic tubules. The extracellular tracer, ruthenium red, enters the tubules from the lateral or basal intercellular spaces in both freshwater- and seawater-adapted animals but never enters either tubules or vesicles from the apical surfaces, indicating that these are not confluent. The presence of dividing basal cells and newly-forming chloride cells, combined with evidence of degeneration of chloride cells, suggests that there is a turnover of this cell type. Both superficial and basal cells are phagocytic and involved in heterophagy of degenerating chloride cells. This phenomenon occurs in both fresh water and sea water indicating that the chloride cells may be functional in both environments.

Adaptation, Physiological↗
PubMed1999

Lithium chloride-induced cardiovascular changes in rabbits are mediated by adenosine triphosphate-sensitive potassium channels.

The effect of lithium chloride on the mean arterial blood pressure, heart rate and heart contractility was evaluated in rabbits. The intravenous administration of 50 mg kg-1 lithium chloride as a bolus injection into rabbits produced a progressive decrease in the mean arterial blood pressure, heart rate and heart contractility during the 2-h period of investigation. Pretreatment of animals with 5 mg kg-1 glibenclamide, a potent inhibitor of adenosine triphosphate-sensitive potassium channels, markedly inhibited the cardiovascular changes induced by lithium chloride. Doubling the dose of glibenclamide nearly abolished these effects of lithium chloride. Similarly, lithium chloride produced a concentration- dependent relaxation of noradrenaline-induced contractions in isolated aortic strips of rabbits. This relaxant effect of lithium chloride was inhibited by pretreatment of the aortic strips with glibenclamide. Doubling the concentration of glibenclamide in the bathing fluid nearly abolished the effect of lithium. Diazoxide and verapamil potentiated the relaxant effect of lithium chloride on the isolated noradrenaline-contracted aortic strips. Pretreatment with glibenclamide markedly reversed the effect of diazoxide but not that of verapamil. The intravenous administration of lithium was also found to be capable of increasing the plasma potassium level and of decreasing the intracellular levels of adenosine triphosphate in cardiac and vascular tissues in a time-dependent manner. The plasma sodium and calcium levels were not changed. These results provide evidence that the hypotensive and cardiac depressant effects of lithium chloride are mediated by activation of adenosine triphosphate-sensitive potassium channels.

Adenosine Triphosphate↗
PubMed1983

Covalent protein binding of vinyl chloride metabolites during co-incubation of freshly isolated hepatocytes and hepatic sinusoidal cells of rats.

Proteins of isolated rat hepatic sinusoidal cells incubated with 14C-vinyl chloride, rat liver microsomes and an NADPH-regenerating system were alkylated by vinyl chloride metabolites formed by microsomes. This suggests that reactive vinyl chloride metabolites can penetrate sinusoidal cells. Protein alkylation in isolated hepatic sinusoidal cells was higher when these were co-incubated with isolated hepatocytes, indicating that reactive vinyl chloride metabolites formed by hepatocytes are stable enough to diffuse out of hepatocytes into sinusoidal cells. Glutathione added to the incubation medium inhibited the covalent protein binding of vinyl chloride metabolites in sinusoidal cells as well as in hepatocytes incubated separately and deleted the increased protein binding in sinusoidal cells co-incubated with hepatocytes. The data indicate that glutathione present in the incubation medium traps reactive vinyl chloride metabolites formed by hepatocytes which otherwise would react with cell constituents of sinusoidal cells. If similar conditions exist in vivo, the alkylation of DNA of liver endothelial cells by vinyl chloride metabolites formed in hepatocytes is possible. This would explain the induction of hemangioendotheliomas of the liver by vinyl chloride.

Alkylation↗
PubMed1989

Extracellular sodium and chloride depletion enhances nonexocytotic noradrenaline release induced by energy deficiency in rat heart.

The effect of either extracellular sodium or extracellular chloride reduction on the release of endogenous noradrenaline and its deaminated metabolite dihydroxyphenylglycol (DOPEG) has been studied in the isolated perfused rat heart under conditions of ischaemia and cyanide intoxication. The overflow of noradrenaline and DOPEG was determined by high pressure liquid chromatography. The efflux of DOPEG, the predominant neuronal noradrenaline adrenaline metabolite, served as indicator of the free axoplasmic plasmic amine concentration. A calcium-free perfusion buffer was used to avoid exocytotic noradrenaline release. Sodium and chloride in the perfusion buffer were replaced by lithium and isethionate, respectively. (1) Reduction of extracellular sodium or chloride increased noradrenaline overflow in ischaemia. The release was suppressed by the uptake1 blocker cocaine indicating carrier-mediated outward transport of noradrenaline. (2) In cyanide intoxication sodium or chloride reduction accelerated the onset of DOPEG efflux reflecting increased axoplasmic noradrenaline concentrations. This was accompanied by increased noradrenaline release. The ratio of noradrenaline/DOPEG overflow was increased by reduced sodium or chloride, indicating facilitation of carrier-mediated noradrenaline net outward transport. (3) In the presence of unaltered energy metabolism overflow of both, noradrenaline and DOPEG, was not enhanced by sodium or chloride reduction. The results demonstrate that reduction of extracellular sodium or chloride has two effects on noradrenaline release from the sympathetic neuron with reduced energy supply. First, reduced sodium or chloride induces increased axoplasmic noradrenaline concentrations by interference with vesicular storage function. Second, both interventions enhance carrier-mediated noradrenaline release.

Animals↗
PubMed1975

Chloride cells of larval Notonecta glauca and Naucoris cimicoides (Hemiptera, Hydrocorisae). Fine structure and cell counts at different salinities.

The chloride cells of the larval waterbugs, Notonecta glauca L. and Naucoris cimicoides L., ultrastructurally resemble the caviform chloride cells of other aquatic insects as well as those of teleost fish. The predominant features are abundant mitochondria, basolateral plasma membrane infoldings and an apical cavity possessing numerous microvilli. After histochemical precipitation of chloride, dense deposits of silver chloride are present in the apical region. The cuticular area overlying the chloride cells is extremely reduced in thickness and differentiated into a plug-like structure. The 1st instars of Notonecta glauca raised from the eggs at various hypo-osmotic salt concentrations have identical numbers of thoracal chloride cells regardless of the external salinity. The number of cells progressively increases with each moult. However, in corresponding larval stages, there is a significant decrease in the number of thoracal chloride cells relative to increases in the external salinity. These results are consistent with the presumptive involvement of the chloride cells in osmotic hyperregulation.

Animals↗
PubMed1995

Apical membrane sodium and chloride entry during osmotic swelling of renal (A6) epithelial cells.

To assess the role of chloride in cell volume and sodium transport regulation, we measured cell height changes (CH), transepithelial chloride and sodium fluxes, and intracellular chloride content during challenge with hyposmotic solutions under open circuit (OC) conditions. CH maximally increased following hyposmotic challenge within approximately 5 minutes. The change in CH was smaller under short circuit (SC) conditions or following replacement of chloride in the mucosal solution by gluconate or cyclamate (Cl(-)-freem). When corrected for the osmotically inactive cell volume (30 +/- 2%), delta CH for controls (OC) were greater than predicted for an ideal osmometer. In contrast, delta CH for Cl(-)-freem or SC conditions were similar to that predicted for an ideal osmometer. Na+ and Cl- mucosa-to-serosa fluxes increased following hyposmotic challenge. Chloride fluxes increased maximally within 5 min, then decreased. In contrast, the Na+ flux increased slowly and reached a steady state after approximately 25 min. Under isosmotic conditions, exposure to Cl(-)-freem solutions led to decreases in the transepithelial conductance, Na+ flux, and CH. Chloride permeabilities in the apical and basolateral membranes were detected using the fluorescent intracellular chloride indicator MQAE. The results indicate that during osmotic swelling, the entry of both sodium and chloride is increased. The time courses of these increases differ, suggesting distinct mechanisms for the osmotic regulation of these apical membrane transport processes.

Animals↗
PubMed1981

Sodium-chloride transport in the thick ascending limb of Henle's loop. Oxygen consumption studies in isolated cells.

Isolated cells were prepared from the medullary thick ascending limb of Henle's loop (TALH) and the response of oxygen consumption was correlated with the active chloride transport system found in these cells. Oxygen consumption was 31.6 microliters O2/mg protein . h and inhibited 50% by the absence of either sodium or chloride in the incubation medium. The absence of both sodium and chloride produced no further inhibition of oxygen consumption. Ouabain (10(-4) M) inhibited oxygen consumption by 50% and the inhibitory effect depended on the presence of both sodium and chloride in the incubation medium. Further, furosemide inhibited oxygen consumption by a maximum of 50% at 10(-3) M and also had no inhibitory effect if either sodium or chloride were absent. Furosemide had no effect on the Na, K-ATPase activity or ATP levels of the TALH cells. Thus, the data suggest or ATP levels of the TALH cells. Thus, the data suggest that 50% of the oxygen consumption of the TALH cells is related to the movement of sodium and chloride into the cell and that the ions may be transported in a coupled manner. In addition the effect of various diuretics on oxygen consumption in the isolated TALH cells was tested. The diuretics could be grouped in three categories: (1) highly effective in inhibiting chloride-dependent oxygen consumption with an apparent inhibitory constant (Ki) of around 10(-6) M, including the diuretics furosemide, bumetanide, ethacrynic acid-cysteine and piretanide, (2) diuretics which were less effective in inhibiting oxygen consumption with an apparent Ki of around 10(-4) M, HOE 740 and ethacrynic acid, and (3) diuretics which were ineffective in inhibiting chloride-dependent oxygen consumption, amiloride and hydrochlorothiazide.

Adenosine Triphosphate↗
PubMed1993

Random chemical modification of the oxygen-linked chloride-binding sites of hemoglobin: those in the central dyad axis may influence the transition between deoxy- and oxy-hemoglobin.

The features of random chemical modification are defined with reference to acetylation of bovine hemoglobin, which has been performed in a random manner so that all of the amino groups that participate in functional chloride binding (i.e., those that are oxygen-linked) could be identified. Random chemical modification, which has objectives different from those of both specific (selective) and extensive chemical modification, has been achieved for bovine hemoglobin with the mild reagent, 14C-methyl acetate phosphate; retention of function was demonstrated by a Hill coefficient of n = 2.2 for the modified hemoglobin. After removal of unmodified Hb chains, the mixture of randomly modified acetylated alpha or beta chains was subjected to tandem treatment with trypsin and chymotrypsin. Peptides were purified by HPLC and identified by amino acid analysis. The amount of radioactivity in the acetylated amino group of a purified peptide was taken as an estimate of the degree of chloride binding. For bovine Hb, two amino groups of the alpha-chain (Val-1 and Lys-99) and three amino groups of the beta-chain (Met-1, Lys-81, and Lys-103) were shown to be oxygen-linked (i.e., to have incorporated significantly more radioactivity in the deoxy conformation compared to the same site in the oxy conformation). Three of these sites were already known chloride-binding sites [i.e., Val-1(alpha), the N-terminus of the alpha-chain, and two sites between the 2 beta-chains of bovine hemoglobin, Met-1(beta) and Lys-81(beta); these findings support the conclusions of the random modification approach. Two other chloride-binding sites, Lys-99(alpha) and Lys-103(beta), align the sides of the central dyad axis connecting the two well-known major chloride-binding sites of bovine Hb. The interrelationship of these five chloride-binding sites was assessed by improved molecular graphics. When viewed through the central dyad axis, the functional chloride-binding sites in the central cavity appear to be symmetrically related and to connect the two major chloride-binding sites. Modifiers or mutants that are directed at these regions in the central dyad axis may favor the deoxy conformation to provide a lower oxygen affinity by preventing the constriction of the central cavity that normally occurs upon oxygenation.

Acetylation↗
PubMed1982

Effects of inhibition of chloride transport on intracellular sodium activity in distal amphibian nephron.

Previous experiments had demonstrated that cell chloride activities in early distal tubule cells of Amphiuma are above equilibrium distribution. Chloride activities fell sharply towards electrochemical equilibrium following perfusion of the tubular lumen with furosemide or with sodium-free solutions. These results suggested a furosemide-sensitive sodium chloride cotransport system in the luminal cell membrane. The present experiments were carried out to evaluate directly the electrochemical driving forces acting on sodium ions under similar experimental conditions. Intracellular sodium activity measurements were performed in the doubly-perfused kidney of Amphiuma by means of single-barreled liquid ion-exchange microelectrodes. Basolateral cell membrane potential and resistance ratio measurements of tubular cell membranes were also carried out under control conditions and after inhibition of chloride transport by luminal application of furosemide (5 . 10(-5) mol/l) or by omission of chloride. Control conditions were characterized by a steep downhill electrochemical gradient for sodium ions from lumen to cell. Inhibition of chloride transport led to a sharp decrease of intracellular sodium activity and to hyperpolarization of the peritubular membrane potential while the resistance ratio of the tubular cell membranes did not change significantly. These results demonstrate the presence of low cellular sodium activities in early distal tubule cells. The sharp decline of cell sodium after furosemide and after luminal chloride removal is consistent with inhibition of a sodium chloride cotransport system and continued peritubular sodium extrusion. The latter can increase the electrochemical gradient of sodium ions beyond that observed under control conditions.

Animals↗