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Effect of reactive oxygen species on NH permeation in Xenopus laevis oocytes

Effect of reactive oxygen species on NH permeation in Xenopus laevis oocytes Abstract To investigate the effects of reactive oxygen species (ROS) on NH permeation in Xenopus laevis oocytes, we used intracellular double-barreled microelectrodes to monitor the changes in membrane potential ( V m ) and intracellular pH (pH i ) induced by a 20 mM NH 4 Cl-containing solution. Under control conditions, NH 4 Cl exposure induced a large membrane depolarization (to V m = 4.0 ± 1.5 mV; n = 21) and intracellular acidification reaching a change in pH i (ΔpH i ) of 0.59 ± 0.06 pH units in 12 min; the initial rate of cell acidification (dpH i /d t ) was 0.06 ± 0.01 pH units/min. Incubation of the oocytes in the presence of H 2 O 2 or β-amyloid protein had no marked effect on the NH 4 Cl-induced ΔpH i . By contrast, in the presence of photoactivated rose bengal (RB), tert -butyl-hydroxyperoxide ( t -BHP), or xanthine/xanthine oxidase (X/XO), the same experimental maneuver induced significantly greater ΔpH i and dpH i /d t . These increases in ΔpH i and dpH i /d t were prevented by the ROS scavengers histidine and desferrioxamine, suggesting involvement of the reactive species 1 ΔgO 2 and ·OH. Using the voltage-clamp technique to identify the mechanism underlying the ROS-measured effects, we found that RB induced a large increase in the oocyte membrane conductance ( G m ). This RB-induced G m increase was prevented by 1 mM diphenylamine-2-carboxylate (DPC) and by a low Na + concentration in the bath. We conclude that RB, t -BHP, and X/XO enhance NH influx into the oocyte via activation of a DPC-sensitive nonselective cation conductance pathway. ammonium ions nonselective cationic conductance Footnotes Address for reprint requests and other correspondence: G. Planelles, Inserm U 467, Université Paris V, Faculté de Médecine Necker-Enfants Malades, 156 rue de Vaugirard, 75730 Paris Cedex 15, France (E-mail: planelle@necker.fr ). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “ advertisement ” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. First published February 6, 2002;10.1152/ajpcell.00410.2001 Copyright © 2002 the American Physiological Society http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png AJP - Cell Physiology The American Physiological Society

Effect of reactive oxygen species on NH permeation in Xenopus laevis oocytes

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Publisher
The American Physiological Society
Copyright
Copyright © 2010 the American Physiological Society
ISSN
0363-6143
eISSN
1522-1563
DOI
10.1152/ajpcell.00410.2001
pmid
11997259
Publisher site
See Article on Publisher Site

Abstract

Abstract To investigate the effects of reactive oxygen species (ROS) on NH permeation in Xenopus laevis oocytes, we used intracellular double-barreled microelectrodes to monitor the changes in membrane potential ( V m ) and intracellular pH (pH i ) induced by a 20 mM NH 4 Cl-containing solution. Under control conditions, NH 4 Cl exposure induced a large membrane depolarization (to V m = 4.0 ± 1.5 mV; n = 21) and intracellular acidification reaching a change in pH i (ΔpH i ) of 0.59 ± 0.06 pH units in 12 min; the initial rate of cell acidification (dpH i /d t ) was 0.06 ± 0.01 pH units/min. Incubation of the oocytes in the presence of H 2 O 2 or β-amyloid protein had no marked effect on the NH 4 Cl-induced ΔpH i . By contrast, in the presence of photoactivated rose bengal (RB), tert -butyl-hydroxyperoxide ( t -BHP), or xanthine/xanthine oxidase (X/XO), the same experimental maneuver induced significantly greater ΔpH i and dpH i /d t . These increases in ΔpH i and dpH i /d t were prevented by the ROS scavengers histidine and desferrioxamine, suggesting involvement of the reactive species 1 ΔgO 2 and ·OH. Using the voltage-clamp technique to identify the mechanism underlying the ROS-measured effects, we found that RB induced a large increase in the oocyte membrane conductance ( G m ). This RB-induced G m increase was prevented by 1 mM diphenylamine-2-carboxylate (DPC) and by a low Na + concentration in the bath. We conclude that RB, t -BHP, and X/XO enhance NH influx into the oocyte via activation of a DPC-sensitive nonselective cation conductance pathway. ammonium ions nonselective cationic conductance Footnotes Address for reprint requests and other correspondence: G. Planelles, Inserm U 467, Université Paris V, Faculté de Médecine Necker-Enfants Malades, 156 rue de Vaugirard, 75730 Paris Cedex 15, France (E-mail: planelle@necker.fr ). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “ advertisement ” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. First published February 6, 2002;10.1152/ajpcell.00410.2001 Copyright © 2002 the American Physiological Society

Journal

AJP - Cell Physiology – The American Physiological Society

Published: Jun 1, 2002

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