Dual effect of carbonic anhydrase inhibitors on H+ transport by the turtle bladder

1981 ◽  
Vol 240 (5) ◽  
pp. F400-F405 ◽  
Author(s):  
L. H. Norby ◽  
D. Bethencourt ◽  
J. H. Schwartz
Keyword(s):  
Carbonic Anhydrase ◽  
Mucosal Cell ◽  
Measurable Effect ◽  
Electrochemical Gradient ◽  
Dual Effect ◽  
Turtle Bladder ◽  
Noncompetitive Inhibitor ◽  
High Concentrations ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Action Inhibition

Previous studies in isolated turtle bladder have demonstrated that high concentrations of carbonic anhydrase (CA) inhibitors limit H+ transport (JH) by reducing the catalyzed rate of CO2 hydroxylation and also by inhibiting some other step in the acidification process. One possibility is that these inhibitors alter the energy substrate requirement for JH. Recent work has demonstrated that JH may be dependent in part on glucose oxidation via the pentose shunt (PS). The present study was undertaken to determine whether CA inhibitors exert a direct effect on PS metabolism by turtle bladder. Acetazolamide and benzolamide at concentrations of 5 X 10(-4) M significantly reduced the rate of 14CO2 evolution from [1–14C]- but not [6–14C]glucose after JH was abolished by an adverse electrochemical gradient for JH+. These changes are consistent with a reduction in PS metabolism. These same sulfonamides also reduced glucose-6-phosphate dehydrogenase (G-6-PD) activity in mucosal cell homogenates. Acetazolamide decreased the Vmax of G-6-PD but not the Km and, therefore, appears to be a noncompetitive inhibitor of G-6-PD with an estimated Ki of 10(-4) M. The t-butyl analogue of acetazolamide, CL 13850, which is without CA inhibitory activity, had no measurable effect on G-6-PD activity. Accordingly, it is suggested the sulfonamide CA inhibitors may reduce JH by two modes of action, inhibition of CA and inhibition of G-6-PD.

Specialized function of carbonic anhydrase-rich and granular cells of turtle bladder

1982 ◽  
Vol 242 (6) ◽  
pp. F627-F633
Author(s):  
J. H. Schwartz ◽  
D. Bethencourt ◽  
S. Rosen
Keyword(s):  
Carbonic Anhydrase ◽  
Gradient Centrifugation ◽  
Cell Types ◽  
O2 Consumption ◽  
G Cells ◽  
Turtle Bladder ◽  
Specific Transport ◽  
Morphological Criteria ◽  
Transport Inhibitors ◽  
Glucose 6 Phosphate Dehydrogenase

The mucosal surface of the turtle bladder consists of two major cell types, carbonic anhydrase-rich (CA) and granular (G) cells. These cells types may have specific transport functions. To separate and study these cells, they were removed from the bladder by collagenase digestion and then separated into two distant subpopulations by Ficoll discontinuous density-gradient centrifugation. By morphological criteria the lighter subpopulation (band II) consists primarily of CA cells and the denser population (band III) G cells. The distribution of two enzymes important for H+ transport, CA and glucose-6-phosphate dehydrogenase (G-6-PD), and the effect of specific transport inhibitors, acetazolamide and ouabain, on O2 consumption of these cells was examined. The CA activity of band II cells was 4.5 times greater than that of band III cells. The G-6-PD activity of band II cells was 2.2 times greater than that of band III cells. The rate of O2 consumption of band II cells, 4.73 microliters.mg protein-1.h-1, was reduced 40% by 5 X 10(-5) M acetazolamide but not by ouabain. Band III O2 consumption, 3.43 microliters.mg protein-1.h-1, was reduced 30% by 10(-4) M ouabain but not by acetazolamide. On the basis of the distribution of CA and G-6-PD and the effect of acetazolamide and ouabain on O2 consumption, we propose that CA cells are primarily responsible for H+ transport and G cells for Na+ transport.

scholarly journals Hemolytic anemia in hereditary pyrimidine 5'-nucleotidase deficiency: nucleotide inhibition of G6PD and the pentose phosphate shunt

Blood ◽  
1982 ◽  
Vol 60 (5) ◽  
pp. 1212-1218 ◽  
Author(s):  
A Tomoda ◽  
NA Noble ◽  
NA Lachant ◽  
KR Tanaka
Keyword(s):  
Hemolytic Anemia ◽  
Competitive Inhibitor ◽  
Pentose Phosphate ◽  
Similar Degree ◽  
Phosphogluconate Dehydrogenase ◽  
Noncompetitive Inhibitor ◽  
High Concentrations ◽  
Nucleotide Inhibition ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Pentose Phosphate Shunt

We evaluated the erythrocytes of two patients with hereditary pyrimidine 5′-nucleotidase deficiency. Significant findings included an increased reduced glutathione content, increased incubated Heinz body formation, a positive ascorbate cyanide test, and decreased intraerythrocytic pH. The pentose phosphate shunt activity of the patients' red cells as measured by the release of 14CO2 from 14C-1- glucose was decreased compared to high reticulocyte controls. Glucose-6- phosphate dehydrogenase (G6PD) activity in hemolysates from control erythrocytes was inhibited 43% by 5.5 mM cytidine 5′-triphosphate (CTP) and 50% by 5.5 mM in uridine 5′-triphosphate (UTP) at pH 7.1. CTP was a competitive inhibitor for G6P (Ki = 1.7 mM) and a noncompetitive inhibitor for NADP+ (Ki = 7.8 mM). Glutathione peroxidase, glutathione reductase, and 6-phosphogluconate dehydrogenase were not affected by these compounds. Pentose phosphate shunt activity in control red cell hemolysate at pH 7.1 was inhibited to a similar degree by 5.5 mM CTP or UTP. Since the intracellular concentrations of G6P and NADP+ are below their KmS for G6PD, these data suggest that high concentrations of pyrimidine 5′-nucleotides depress pentose phosphate shunt activity in pyrimidin 5′-nucleotidase deficiency. Thus, this impairment of the pentose phosphate pathway appears to contribute to the pathogenesis of hemolysis in pyrimidine 5′-nucleotidase deficiency hemolytic anemia.

scholarly journals Hemolytic anemia in hereditary pyrimidine 5'-nucleotidase deficiency: nucleotide inhibition of G6PD and the pentose phosphate shunt

Blood ◽  
1982 ◽  
Vol 60 (5) ◽  
pp. 1212-1218 ◽  
Author(s):  
A Tomoda ◽  
NA Noble ◽  
NA Lachant ◽  
KR Tanaka
Keyword(s):  
Hemolytic Anemia ◽  
Competitive Inhibitor ◽  
Pentose Phosphate ◽  
Similar Degree ◽  
Phosphogluconate Dehydrogenase ◽  
Noncompetitive Inhibitor ◽  
High Concentrations ◽  
Nucleotide Inhibition ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Pentose Phosphate Shunt

Abstract We evaluated the erythrocytes of two patients with hereditary pyrimidine 5′-nucleotidase deficiency. Significant findings included an increased reduced glutathione content, increased incubated Heinz body formation, a positive ascorbate cyanide test, and decreased intraerythrocytic pH. The pentose phosphate shunt activity of the patients' red cells as measured by the release of 14CO2 from 14C-1- glucose was decreased compared to high reticulocyte controls. Glucose-6- phosphate dehydrogenase (G6PD) activity in hemolysates from control erythrocytes was inhibited 43% by 5.5 mM cytidine 5′-triphosphate (CTP) and 50% by 5.5 mM in uridine 5′-triphosphate (UTP) at pH 7.1. CTP was a competitive inhibitor for G6P (Ki = 1.7 mM) and a noncompetitive inhibitor for NADP+ (Ki = 7.8 mM). Glutathione peroxidase, glutathione reductase, and 6-phosphogluconate dehydrogenase were not affected by these compounds. Pentose phosphate shunt activity in control red cell hemolysate at pH 7.1 was inhibited to a similar degree by 5.5 mM CTP or UTP. Since the intracellular concentrations of G6P and NADP+ are below their KmS for G6PD, these data suggest that high concentrations of pyrimidine 5′-nucleotides depress pentose phosphate shunt activity in pyrimidin 5′-nucleotidase deficiency. Thus, this impairment of the pentose phosphate pathway appears to contribute to the pathogenesis of hemolysis in pyrimidine 5′-nucleotidase deficiency hemolytic anemia.

Dual effect of glycine on horizontal cells of the tiger salamander retina

1991 ◽  
Vol 66 (6) ◽  
pp. 1993-2001 ◽  
Author(s):  
S. Borges ◽  
M. Wilson
Keyword(s):  
Opposite Effect ◽  
Horizontal Cell ◽  
Light Response ◽  
Membrane Resistance ◽  
Horizontal Cells ◽  
Field Size ◽  
Dual Effect ◽  
Light Responses ◽  
Low Concentrations ◽  
High Concentrations

1. The effects of glycine on horizontal cells have been examined by microelectrode recording from superfused retinas isolated from the salamander. 2. Low concentrations of glycine (less than 50 microM) hyperpolarized horizontal cells and increased the magnitude of their light responses. Millimolar concentrations produced the opposite effect of depolarizing these cells and reducing their light response amplitudes. 3. In the presence of Co2+ and Mg2+ at concentrations sufficient to suppress the light response, millimolar glycine still exerted a depolarizing effect on horizontal cells, implying that this effect was largely a direct one on horizontal cell membranes. 4. Although both the rod and the cone contributions to horizontal cell light responses were reduced by millimolar glycine, rod input was reduced more, suggesting that millimolar glycine may also exert a presynaptic effect. 5. Strychnine (10 microns) antagonized the effects of millimolar glycine and, in the absence of exogenously applied glycine, caused horizontal cells to hyperpolarize and their light responses to increase in amplitude. This result implies that, in darkness, glycine is tonically released onto horizontal cells and maintains them in a state of partial depolarization. 6. The low-concentration effect of glycine was accompanied by an increased membrane resistance and receptive field size but no change in the balance of rod and cone input. 7. Low concentrations of glycine were often seen to cause a speeding of light responses, whereas high concentrations sometimes caused a slowing of response kinetics. Response kinetics were found to correlate with horizontal cell dark membrane potential so that, positive to -30 mV, depolarization slowed responses whereas kinetics at more negative values were largely independent of voltage.

Compatible Solutes and Inorganic Ions in the Mangrove Plant Avicennia marina and Their Effects on the Activities of Enzymes

1997 ◽  
Vol 52 (7-8) ◽  
pp. 433-440 ◽  
Author(s):  
Hiroshi Ashihara ◽  
Kyoko Adachi ◽  
Miho Otawa ◽  
Eri Yasumoto ◽  
Yuko Fukushima ◽  
...  
Keyword(s):  
Compatible Solutes ◽  
Inorganic Ions ◽  
Avicennia Marina ◽  
Salt Resistance ◽  
Nmr Studies ◽  
Mangrove Plant ◽  
Young Leaves ◽  
High Concentrations ◽  
Fructose 1,6 Bisphosphate ◽  
Glucose 6 Phosphate Dehydrogenase

Naturally grown two-month-old seedlings of Avicennia marina contain high concentrations of Na+ and Cl-.+ Our NMR studies revealed an accumulation of glycinebetaine, asparagine and stachyose in A. marina. The highest concentration of glycinebetaine was observed in young leaves, while the distribution of stachyose was restricted in stems and roots. A sparagine comprised more than 96% of total free amino acids in roots and 84% in leaves from two-year-old plants. Little or no accumulation of proline or polyols, which are proposed as compatible solutes in other plants, could be detected in A. marina. The activities of phosphofructokinase, pyrophosphate:fructose-6-phosphate 1-phosphotransferase, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase (decarboxylating), phosphoenolpyruvate carboxylase and NAD:malate dehydrogenase from young leaves of A. marina were inhibited by NaCl, while the activity of fructose-1,6-bisphosphate aldolase was activated by 50-200 m M NaCl. There was little or no effect of high concentrations (up to 500 mᴍ ) of glycinebetaine on the activities of any of these enzymes. No significant protection by glycinebetaine was detected against NaCl inhibition of these enzymatic activities. Based on these results, possible mechanisms for the salt-resistance of A. marina cells are discussed.

H+ current response to CO2 and carbonic anhydrase inhibition in turtle bladder

1976 ◽  
Vol 231 (2) ◽  
pp. 565-572 ◽  
Author(s):  
JH Schwartz
Keyword(s):  
Carbonic Anhydrase ◽  
Maximum Rate ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Current Response ◽  
Low Concentrations ◽  
Ph Stat ◽  
High Concentrations ◽  
Co2 Addition

To evaluate the role of CO2 and carbonic anhydrase (CA) in H+ transport (JH) by turtle urinary bladder the effect of CO2 addition, with and without addition of CA inhibitiors, was examined on JH. Since in the presence of exogenous CO2 and HCO3- the pH stat-measured rate of mucosal (M) acidification underestimates JH by the rate of electroneutral HCO3- secretion, the reverse short-circuit current (RSCC) applied across ouabain-treated bladders was used to estimate JH. That the RSCC is a measure of JH was demonstrated by: 1) in the absence of added CO2 and HCO3- the rate of M acidification totally accounted for the RSCC, and 2) increases in RSCC with CO2 addition occurred without changes in Na+ and K+ fluxes or the coupled ration of HCO3- secretion for Cl-absorption. When serosal (S) percent CO2 was progressively progressively increased JH achieved a maximum rate of 64 +/- 3 muA (SE) with 4.5% CO2. At higher S percent CO2 JH did not change, suggesting that factors other than the rate of CO2 hydration were rate limiting. The maximum rate of JH was not decreased by low concentrations of CA inhibitors (acetazolamide, 5 X 10(-5) M), although the percent CO2 at which this maximum rate occurred increased to 8.5%. The increased percent CO2 requirement for the maximum rate of JH with low concentrations of CA inhibitors suggests that these agents alter JH by decreasing the rate of enzymatic CO2 hydration. At high concentrations (acetazolamide, 5 X 10(-4) M) these inhibitors decrease the maximum rate of JH in the presence of CO2, implying that these inhibitors at higher concentrations directly interfere with the H+ transport system.

Mechanism of acid adaptation of a fish living in a pH 3.5 lake

2003 ◽  
Vol 284 (5) ◽  
pp. R1199-R1212 ◽  
Author(s):  
Taku Hirata ◽  
Toyoji Kaneko ◽  
Toshihiro Ono ◽  
Takeru Nakazato ◽  
Norihisa Furukawa ◽  
...  
Keyword(s):  
Carbonic Anhydrase ◽  
Basolateral Membrane ◽  
Single Species ◽  
Carbonic Anhydrase Ii ◽  
Chloride Cells ◽  
Apical Surface ◽  
Acidic Lake ◽  
Apical Side ◽  
High Concentrations

Despite unfavorable conditions, a single species of fish, Osorezan dace, lives in an extremely acidic lake (pH 3.5) in Osorezan, Aomori, Japan. Physiological studies have established that this fish is able to prevent acidification of its plasma and loss of Na+. Here we show that these abilities are mainly attributable to the chloride cells of the gill, which are arranged in a follicular structure and contain high concentrations of Na+-K+-ATPase, carbonic anhydrase II, type 3 Na+/H+ exchanger (NHE3), type 1 Na+-HCO[Formula: see text] cotransporter, and aquaporin-3, all of which are upregulated on acidification. Immunohistochemistry established their chloride cell localization, with NHE3 at the apical surface and the others localized to the basolateral membrane. These results suggest a mechanism by which Osorezan dace adapts to its acidic environment. Most likely, NHE3 on the apical side excretes H+ in exchange for Na+, whereas the electrogenic type 1 Na+-HCO[Formula: see text]cotransporter in the basolateral membrane provides HCO[Formula: see text] for neutralization of plasma using the driving force generated by Na+-K+-ATPase and carbonic anhydrase II. Increased expression of glutamate dehydrogenase was also observed in various tissues of acid-adapted dace, suggesting a significant role of ammonia and bicarbonate generated by glutamine catabolism.

Carbonic anhydrase in turtle bladder mitochondrial-rich luminal and subluminal cells

1991 ◽  
Vol 260 (3) ◽  
pp. F431-F442 ◽  
Author(s):  
C. Fritsche ◽  
J. G. Kleinman ◽  
J. L. Bain ◽  
R. R. Heinen ◽  
D. A. Riley
Keyword(s):  
Epithelial Cells ◽  
Carbonic Anhydrase ◽  
Morphological Feature ◽  
Basal Layer ◽  
Terminal Web ◽  
Turtle Bladder ◽  
Lateral Margins

Bladders from March-April turtles were processed for carbonic anhydrase (CA) cytochemically using the method of D.A. Riley, S. Ellis, and J. Bain (Neuroscience 13: 189, 1984). CA-positive cells comprised 11.1 +/- 0.7% of mucosal epithelial cells. Microplicated (MP) cells comprised 47.2 +/- 1.8% of CA-positive cells and displayed at least two distinct staining patterns: the first was characterized by reaction product that filled the luminal one-third, including the terminal web and microplicae. These cells possessed extensive microplicae, a morphological feature of ongoing H+ secretion. The second was characterized by reaction product distributed throughout cells, excluding the terminal web and microplicae, with greatest intensity in the luminal one-third below the terminal web. These cells possessed flattened microplicae, a morphological feature of diminished H+ secretion. Microvillated (MV) cells comprised 6.0 +/- 1.0% of CA-reactive cells. The basal layer was occupied by 46.8 +/- 1.7% of CA-positive cells, which were termed subluminal (SL) cells. SL cells were mitochondrial rich and did not contact the lumen. Extracellular CA staining was common between the lateral margins of contiguous mitochondrial-rich or non-mitochondrial-rich cells.

Carbonic anhydrase and proton secretion in turtle bladder mitochondrial-rich cells

1991 ◽  
Vol 260 (3) ◽  
pp. F443-F458
Author(s):  
C. Fritsche ◽  
J. G. Kleinman ◽  
J. L. Bain ◽  
R. R. Heinen ◽  
D. A. Riley
Keyword(s):  
Carbonic Anhydrase ◽  
Acid Base ◽  
Proton Secretion ◽  
Turtle Bladder ◽  
Acid Base Status ◽  
Reduced Density

Bladders from actively feeding turtles were processed for carbonic anhydrase (CA) cytochemically. CA-positive cells were identified as microplicated (MP) cells, microvillated (MV) cells, and subluminal (SL) cells. After acute enhancement of H+ secretion with 5% CO2, MP cells displayed extensive microplicae and a reduced density of apical subplasmalemmal vesicles, and they were CA reactive throughout a large part of the cytoplasm including the microplicae. After acute inhibition of H+ secretion with a pH 4.5 mucosal bath, CA staining was excluded from the microplicae and apical subplasmalemmal region of most MP cells, whereas microplicae varied from extensive to reduced, and subapical vesicle density remained elevated. MV cells were characterized by basolateral staining with sparing of the MV and apical subplasmalemmal region in all settings except 1) after 5% CO2 and 2) when MV cells were found in areas in which MP cells were stained to the lumen. These results indicate that CA is active at the site of H+ secretion in MP cells and is correlated with the acute acid-base status of the bladder.

Dual Effect of Platelet Factor Four on the Activities of Factor Xa

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1034-1034
Author(s):  
Martine Marie Fiore ◽  
Ian J Mackie
Keyword(s):  
Factor Xa ◽  
Fold Increase ◽  
Peptide Hydrolysis ◽  
Dual Effect ◽  
Platelet Factor ◽  
Paradoxical Effects ◽  
Heparin Activity ◽  
High Concentrations ◽  
Prothrombin Activation

Abstract Platelet Factor 4 (PF4) is a cationic molecule that binds to heparin with high affinity and neutralises the activity of the latter. Our recent studies indicate that heparin can promote an interaction of fXa with PF4 since neutralization of heparin activity by PF4 was dependent on the concentration of protease. To examine the contribution of PF4 in protease function, fXa activity was determined in chromogenic assays. Upon preincubation with fXa and heparin, PF4 (at a concentration of 100 nM) decreased the kcat of S2765 peptide hydrolysis 4-fold and that of prothrombin activation about 2-fold. These results suggested an effect of PF4 on the primary specificity of the protease. In fact, PF4 exerted a mild effect (30 % decrease) on the Na+ dependence of fXa, consistent with linkage between Na+ and S1. PF4 preincubation with fXa also prevented the binding of the S1 probe p-aminobenzamidine (pAB) while simultaneous addition of PF4 and pAB diminished the contribution of PF4. In the presence of excess fVa (relative to fXa), kinetic parameters measuring fXa amidolytic activity in the presence of PF4 were restored to control values in the absence of PF4. Interestingly, high concentrations of PF4 (> 1 μM) totally restored fXa activity toward peptidyl substrate and strongly enhanced prothrombin activation, indicating a dual effect of PF4 on fXa activities. The inhibitory contribution of PF4 during prothrombin activation was due to a three-fold decreased affinity of fXa for fVa while enhancement of prothrombin activation was accompanied by a three-fold increase in fVa-dependent cofactor activity. Thus, the effects of PF4 possibly involved a region of the heparin/fVabinding exosite that is linked to the S1 and Na+ sites. These findings suggest that PF4 is a probe of fVa-dependent changes occurring in the active site of fXa and provide an explanation for the in vivo paradoxical effects of PF4 reported in the literature.

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