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This Article Full Text Full Text (PDF) All Versions of this Article: 576/2/417    most recent jphysiol.2006.117176v1 Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Sakai, H. Articles by Kuno, M. Search for Related Content PubMed PubMed Citation Articles by Sakai, H. Articles by Kuno, M. Related Collections Cellular

Departments of
¹ Physiology
² Neurology
³ Central Laboratory, Osaka City University Graduate School of Medicine, Abeno-ku, Osaka 545-8585, Japan
⁴ National Institutes of Natural Sciences, Department of Molecular Physiology, Division of Intracellular Metabolism, Okazaki 444-8585, Japan

The vacuolar-type H⁺-ATPase (V-ATPase) in the plasma membrane of a variety of cells serves as an acid-secreting pathway, and its activity is closely related to cellular functions. Massive proton secretion often leads to electrolyte disturbances in the vicinity of the cell and may in turn affect the activity of the V-ATPase. We characterized, for the first time, the proton currents mediated by plasmalemmal V-ATPase in murine osteoclast-like cells and investigated its activity over a wide range of pH gradients across the membrane (pH = extracellular pH – intracellular pH). The V-ATPase currents were identified as outward H⁺ currents and were dependent on ATP and sensitive to the inhibitors bafilomycin A₁ and N,N'-dicyclohexylcarbodiimide. Although H⁺ was transported uphill, the electrochemical gradient for H⁺ affected the current. The currents were increased by elevating pH and depolarization, and were reduced by lowering pH and hyperpolarization. Elevation of extracellular Ca²⁺ (5–40 mM) diminished the currents in a dose-dependent manner and made the voltage dependence more marked. Extracellular Mg²⁺ mimicked the inhibition. With 40 mM Ca²⁺, the currents decreased to < 40% at 0 mV and to < 10% at about –80 mV. Increases in the intracellular Ca²⁺ (0.5–5 µM) did not affect the current. The data suggest that acid secretion through the plasmalemmal V-ATPase is regulated by a combination of the pH gradient, the membrane potential and the extracellular divalent cations. In osteoclasts, the activity-dependent accumulation of acids and Ca²⁺ in the closed extracellular compartment might serve as negative feedback signals for regulating the V-ATPase.

(Received 13 July 2006; accepted after revision 9 August 2006; first published online 10 August 2006)
Corresponding author M. Kuno: Department of Physiology, Osaka City University Graduate School of Medicine, Abeno-ku, Osaka 545-8585, Japan. Email: kunomyk{at}med.osaka-cu.ac.jp

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