Cl- and regulation of pH by MDCK-C11 cells
The interaction between H+ extrusion via H+-ATPase and Cl- conductance was studied in the C11 clone of MDCK cells, akin to the intercalated cells of the collecting duct. Cell pH (pHi) was measured by fluorescence microscopy using the fluorescein-derived probe BCECF-AM. Control recovery rate measured...
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| Format: | Article |
| Language: | English |
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Associação Brasileira de Divulgação Científica
2007-05-01
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| Series: | Brazilian Journal of Medical and Biological Research |
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| Online Access: | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2007000500012 |
| _version_ | 1819263368113946624 |
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| author | A.L. Tararthuch R. Fernandez G. Malnic |
| author_facet | A.L. Tararthuch R. Fernandez G. Malnic |
| author_sort | A.L. Tararthuch |
| collection | DOAJ |
| description | The interaction between H+ extrusion via H+-ATPase and Cl- conductance was studied in the C11 clone of MDCK cells, akin to the intercalated cells of the collecting duct. Cell pH (pHi) was measured by fluorescence microscopy using the fluorescein-derived probe BCECF-AM. Control recovery rate measured after a 20 mM NH4Cl acid pulse was 0.136 ± 0.008 pH units/min (dpHi/dt) in Na+ Ringer and 0.032 ± 0.003 in the absence of Na+ (0 Na+). With 0 Na+ plus the Cl- channel inhibitor NPPB (10 µM), recovery was reduced to 0.014 ± 0.001 dpHi/dt. 8-Br-cAMP, known to activate CFTR Cl- channels, increased dpHi/dt in 0 Na+ to 0.061 ± 0.009 and also in the presence of 46 nM concanamycin and 50 µM Schering 28080. Since it is thought that the Cl- dependence of H+-ATPase might be due to its electrogenic nature and the establishment of a +PD (potential difference) across the cell membrane, the effect of 10 µM valinomycin at high (100 mM) K+ was tested in our cells. In Na+ Ringer, dpHi/dt was increased, but no effect was detected in 0 Na+ Ringer in the presence of NPPB, indicating that in intact C11 cells the effect of blocking Cl- channels on dpHi/dt was not due to an adverse electrical gradient. The effect of 100 µM ATP was studied in 0 Na+ Ringer solution; this treatment caused a significant inhibition of dpHi/dt, reversed by 50 µM Bapta. We have shown that H+-ATPase present in MDCK C11 cells depends on Cl- ions and their channels, being regulated by cAMP and ATP, but not by the electrical gradient established by electrogenic H+ transport. |
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| format | Article |
| id | doaj.art-3d08e4f5d5fb490cb0179184e378e6d8 |
| institution | Directory Open Access Journal |
| issn | 0100-879X 1414-431X |
| language | English |
| last_indexed | 2024-12-23T20:12:28Z |
| publishDate | 2007-05-01 |
| publisher | Associação Brasileira de Divulgação Científica |
| record_format | Article |
| series | Brazilian Journal of Medical and Biological Research |
| spelling | doaj.art-3d08e4f5d5fb490cb0179184e378e6d82022-12-21T17:32:47ZengAssociação Brasileira de Divulgação CientíficaBrazilian Journal of Medical and Biological Research0100-879X1414-431X2007-05-0140568769610.1590/S0100-879X2007000500012Cl- and regulation of pH by MDCK-C11 cellsA.L. TararthuchR. FernandezG. MalnicThe interaction between H+ extrusion via H+-ATPase and Cl- conductance was studied in the C11 clone of MDCK cells, akin to the intercalated cells of the collecting duct. Cell pH (pHi) was measured by fluorescence microscopy using the fluorescein-derived probe BCECF-AM. Control recovery rate measured after a 20 mM NH4Cl acid pulse was 0.136 ± 0.008 pH units/min (dpHi/dt) in Na+ Ringer and 0.032 ± 0.003 in the absence of Na+ (0 Na+). With 0 Na+ plus the Cl- channel inhibitor NPPB (10 µM), recovery was reduced to 0.014 ± 0.001 dpHi/dt. 8-Br-cAMP, known to activate CFTR Cl- channels, increased dpHi/dt in 0 Na+ to 0.061 ± 0.009 and also in the presence of 46 nM concanamycin and 50 µM Schering 28080. Since it is thought that the Cl- dependence of H+-ATPase might be due to its electrogenic nature and the establishment of a +PD (potential difference) across the cell membrane, the effect of 10 µM valinomycin at high (100 mM) K+ was tested in our cells. In Na+ Ringer, dpHi/dt was increased, but no effect was detected in 0 Na+ Ringer in the presence of NPPB, indicating that in intact C11 cells the effect of blocking Cl- channels on dpHi/dt was not due to an adverse electrical gradient. The effect of 100 µM ATP was studied in 0 Na+ Ringer solution; this treatment caused a significant inhibition of dpHi/dt, reversed by 50 µM Bapta. We have shown that H+-ATPase present in MDCK C11 cells depends on Cl- ions and their channels, being regulated by cAMP and ATP, but not by the electrical gradient established by electrogenic H+ transport.http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2007000500012Cell pHcAMPATPH+-ATPaseValinomycinCl- |
| spellingShingle | A.L. Tararthuch R. Fernandez G. Malnic Cl- and regulation of pH by MDCK-C11 cells Brazilian Journal of Medical and Biological Research Cell pH cAMP ATP H+-ATPase Valinomycin Cl- |
| title | Cl- and regulation of pH by MDCK-C11 cells |
| title_full | Cl- and regulation of pH by MDCK-C11 cells |
| title_fullStr | Cl- and regulation of pH by MDCK-C11 cells |
| title_full_unstemmed | Cl- and regulation of pH by MDCK-C11 cells |
| title_short | Cl- and regulation of pH by MDCK-C11 cells |
| title_sort | cl and regulation of ph by mdck c11 cells |
| topic | Cell pH cAMP ATP H+-ATPase Valinomycin Cl- |
| url | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2007000500012 |
| work_keys_str_mv | AT altararthuch clandregulationofphbymdckc11cells AT rfernandez clandregulationofphbymdckc11cells AT gmalnic clandregulationofphbymdckc11cells |