Passive chloride permeability charge coupled to H+-ATPase of avian osteoclast ruffled membrane

H. C. Blair, S. L. Teitelbaum, H. L. Tan, C. M. Koziol, P. H. Schlesinger

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Abstract

We prepared proton-transporting membrane vesicles from the avian osteoclast's ruffled membrane, a specialized region of the cell surface that acidifies the bone resorption space. We demonstrated a unique conductive Cl- permeability that is charge coupled to the vesicle H+-ATPase and is required for acidification. Ion replacement indicated an anion selectivity of Br- ~ Cl- > SO42- > NO3- ~ SCN- in supporting acidification. The anion channel blocker 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (10 μM) was a competitive inhibitor of acidification and raised the Michaelis constant for ATP of the proton pump ~11-fold in 120 mM KCl. Inhibition was reversed by valinomycin, which provides an alternate path for charge neutralization. The Cl- dependence of acidification was nonlinear and yielded a Hill coefficient of 3-4, showing that it is distinct from a linear Cl- dependence reported for acidification of renal cortical endosomes. The K+ ionophore valinomycin augmented H+ transport in K2SO4, and not in KCl. Dependence of Cl- transport on membrane potential was confirmed by direct measurement of 36Cl- transport. We uncoupled charge transport from proton transport with a large excess of ammonia, which had no effect on 36Cl- accumulation in vesicles, and by measuring 36Cl- accumulation in response to a membrane diffusion potential, produced with a [K+] gradient and valinomycin in the absence of ATP. These experiments demonstrate that the electrogenic proton pump of the osteoclast ruffled membrane is charge coupled to a passive Cl- permeability in the same membrane.

Original languageEnglish
Pages (from-to)C1315-C1324
JournalAmerican Journal of Physiology - Cell Physiology
Volume260
Issue number6 29/6
DOIs
StatePublished - Jan 1 1991

Keywords

  • Anion transport
  • Chloride channel
  • Proton pump

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