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  • 1
    ISSN: 1432-2013
    Keywords: BCECF Cell pH Corneal endothelial cells Na+-HCO3– cotransporter RT-PCR
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract. Although bicarbonate transport in corneal endothelium has been suggested to be coupled to Na+, the underlying molecular mechanism has not been clarified. In the present study we investigated whether a recently cloned Na+-HCO3 – cotransporter (NBC-1) is responsible for this process, and, if so, whether the endothelium expresses a separate isoform or one of the other two isoforms that have recently been identified (kNBC-1 from kidney and pNBC-1 from pancreas). Using primers designed for specific and common regions we demonstrated by reverse transcriptase polymerase chain reaction (RT-PCR) that both kNBC-1 and pNBC-1 are expressed in cultured human corneal endothelial cells. In addition functional studies with a pH-sensitive fluorescence probe were performed. In the presence of HCO3 –/CO2 a pH regulatory process was demonstrated which depends on the presence of Na+ and membrane potential, but is independent of Cl– and is inhibited by the disulfonic stilbene DIDS. These results support the presence of NBC-1 as the major bicarbonate transport system in corneal endothelium.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 422 (1992), S. 60-65 
    ISSN: 1432-2013
    Keywords: Renal proximal tubule ; S2 segment ; Rheogenic Na+-HCO 3 − cotransport ; Cl−/HCO 3 − exchange ; Carbonic anhydrase inhibitor
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The influence of the carbonic anhydrase inhibitor acetazolamide (ACZ) was investigated on HCO 3 − transport mechanisms in the basolateral cell membrane of rabbit renal proximal tubule. Experiments were performed on isolated S2 segments using double-barrelled microelectrodes to measure cell membrane potential (V b) and cell pH (pHi) during step changes in bath perfusate ion concentrations. Peritubular application of ACZ (1 mmol/l) reduced the initial V b response to 10∶1 reduction of bath HCO 3 − concentration only slightly, from +53.8±4.2 mV to+49.1±0.3 mV (n=5), but caused an intermittent overshooting repolarization in the secondary V b response. In conjunction with these effects it left the initial pHi response virtually unchanged but induced a secondary slow acidification. These observation indicate that — under the present experimental conditions — ACZ does not block the Na+-HCO 3 − cotransporter but acts via inhibition of cytosolic carbonic anhydrase. This was confirmed by studying the effect of elevated intracellular HCO 3 − concentrations under reduced flux conditions and by comparing the concentration dependence of the V b response with the inhibition kinetics of cytosolic carbonic anhydrase. In contrast, peritubular ACZ inhibited Na+-independent Cl−/HCO 3 − exchange in the basolateral cell membrane of S2 segments directly in a similar way to that described in the preceding publication for S3 segments.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 422 (1992), S. 55-59 
    ISSN: 1432-2013
    Keywords: Renal proximal tubule ; S3 segment ; Cl−/HCO 3 − exchange ; Carbonic anhydrase inhibitors
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Cell pH (pHi) and cell membrane potential (V b) were measured in isolated S3 segments of rabbit renal proximal tubule with double-barrelled microelectrodes to search for a possible effect of the carbonic anhydrase inhibitor, acetazolamide (ACZ), on Cl−/HCO 3 − exchange in the basolateral cell membrane. ACZ was found to retard and reduce the pHi response to bath Cl− removal reversibly with half-maximal inhibition at 0.42 mmol/l and a rather flat concentration dependence (Hill coefficient ≈ 0.36). To determine whether the retardation resulted from inhibition of cytoplasmic carbonic anhydrase, which might have delayed the attainment of HCO 3 − /CO2 equilibrium, we have measured the response of pHi to step changes in PCO2 in the presence and absence of ACZ. ACZ greatly retarded the pHi response to CO2 steps; however, the concentration dependence differed (half-maximal inhibition at 18 μmol/l) and even at maximal ACZ concentrations the response to CO2 steps was more than twice as fast as the response to Cl− replacement. Since, in addition, the ACZ inhibition of Cl−/HCO 3 − exchange could not be overcome by increasing PCO2 we conclude that the ACZ effect on Cl−/HCO 3 − exchange in rabbit proximal tubule S3 segments does not result from inhibition of cytosolic or membrane-bound carbonic anhydrase, but from a direct interaction with the exchanger molecule.
    Type of Medium: Electronic Resource
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