| Intracellular pH in the OK cell. I. Identification of H+ conductance and observations on buffering capacity. | |
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MedLine Citation:
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PMID: 1662906 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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The regulation of intracellular pH (pHi) in the opossum kidney (OK) cell line was studied in vitro using the pH-sensitive excitation ratio of 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. Recovery from an NH4Cl acid load disclosed a Na-dependent component blocked by amiloride and a smaller Na-independent component. The Na-independent recovery rate was proportional to the H+ gradient from cell to buffer and was zero in the absence of an electrochemical gradient. The Na-independent recovery was not affected by N-ethylmaleimide, dicyclohexylcarbodiimide, HCO3, phloretin, or ZnCl2 but was accelerated in depolarized cells and by membrane-fluidizing drugs and was inhibited by glutaraldehyde. The apparent cellular buffering capacity changed in proportion to this H+ conductance. Consistent with an electrogenic H+ leak, steady-state cell pH alkalinized with depolarization and acidified with hyperpolarization. Removal of buffer Na+ produced a profound acidification, as did amiloride. In 0-Na+ buffers, extremely large cell-to-buffer H+ gradients were present and proportional to buffer pH. 4-Acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid and 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid had no effect on steady-state pHi. Measurements of intracellular buffering capacity were derived from the change of cell pH induced by withdrawing NH4Cl. This buffering capacity was increased threefold in Na-free buffers, whereas the value measured by direct titration of cell lysate was the same or less than that of control cells. The NH4Cl-derived buffering capacity varied in direct proportion to the magnitude of the H+ leak. Drugs that changed H+ permeability produced the apparent changes of the measured buffering capacity within a few minutes. We conclude that, in HCO3-free buffer, the OK cell uses two membrane acid-base transport pathways: a Na-H antiporter active at physiological pH and a substantial passive H+ conductance. The results also reveal that the NH4Cl-derived buffering capacity is subject to artifacts, possibly due to a finite leak of ionic NH4+. |
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Authors:
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M Graber; J DiPaola; F L Hsiang; C Barry; E Pastoriza |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S. |
Journal Detail:
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Title: The American journal of physiology Volume: 261 ISSN: 0002-9513 ISO Abbreviation: Am. J. Physiol. Publication Date: 1991 Dec |
Date Detail:
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Created Date: 1992-02-14 Completed Date: 1992-02-14 Revised Date: 2006-11-15 |
Medline Journal Info:
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Nlm Unique ID: 0370511 Medline TA: Am J Physiol Country: UNITED STATES |
Other Details:
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Languages: eng Pagination: C1143-53 Citation Subset: IM |
Affiliation:
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Veterans Affairs Medical Center, Northport 11768. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Amiloride
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pharmacology Animals Buffers Carrier Proteins / metabolism* Cell Line Cell Membrane Permeability / drug effects Dimethyl Sulfoxide / pharmacology Ethylmaleimide / pharmacology Fluoresceins Hydrogen-Ion Concentration Kidney Cortex / enzymology, metabolism* Membrane Potentials Opossums Proton-Translocating ATPases / antagonists & inhibitors, metabolism* Protons Sodium / metabolism Sodium-Hydrogen Antiporter |
| Chemical | |
Reg. No./Substance:
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0/Buffers; 0/Carrier Proteins; 0/Fluoresceins; 0/Protons; 0/Sodium-Hydrogen Antiporter; 128-53-0/Ethylmaleimide; 2609-46-3/Amiloride; 67-68-5/Dimethyl Sulfoxide; 7440-23-5/Sodium; 85138-49-4/2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein; EC 3.6.3.14/Proton-Translocating ATPases |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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