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Research Article Free access | 10.1172/JCI119584
Division of Nephrology, Department of Internal Medicine, University of Michigan and Veteran's Affairs Medical Center, Ann Arbor, Michigan 48109, USA. wnberg@umich.edu
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Division of Nephrology, Department of Internal Medicine, University of Michigan and Veteran's Affairs Medical Center, Ann Arbor, Michigan 48109, USA. wnberg@umich.edu
Find articles by Davis, J. in: JCI | PubMed | Google Scholar
Division of Nephrology, Department of Internal Medicine, University of Michigan and Veteran's Affairs Medical Center, Ann Arbor, Michigan 48109, USA. wnberg@umich.edu
Find articles by Venkatachalam, M. in: JCI | PubMed | Google Scholar
Published August 1, 1997 - More info
Previous studies have shown that cytosolic-free Ca2+ (Caf) increases to at least low micromolar concentrations during ATP depletion of isolated kidney proximal tubules. However, peak levels could not be determined precisely with the Ca2+-sensitive fluorophore, fura-2, because of its high affinity for Ca2+. Now, we have used two low affinity Ca2+ fluorophores, mag-fura-2 (furaptra) and fura-2FF, to quantitate the full magnitude of Caf increase. Between 30 and 60 min after treatment with antimycin to deplete ATP in the presence of glycine to prevent lytic plasma membrane damage, Caf measured with mag-fura-2 exceeded 10 microM in 91% of tubules studied and 68% had increases to greater than 100 microM. Caf increases of similar magnitude that were dependent on influx of medium Ca2+ were also seen using the new low Ca2+ affinity, Mg2+-insensitive, fluorophore fura-2FF in tubules depleted of ATP by hypoxia, and these increases were reversed by reoxygenation. Total cell Ca2+ levels in antimycin-treated or hypoxic tubules did not change, suggesting that mitochondria were not buffering the increased Caf during ATP depletion. Considered in the context of the high degree of structural preservation of glycine-treated tubule cells during ATP depletion and the commonly assumed Ca2+ requirements for phospholipid hydrolysis, actin disassembly, and Ca2+-mediated structural damage, the remarkable elevations of Caf demonstrated here suggest an unexpected resistance to the deleterious effects of increased Caf during energy deprivation in the presence of glycine.