Effect of reactive oxygen species on NH4+ permeation in Xenopus laevis oocytes.
To investigate the effects of reactive oxygen species (ROS) on NH4+ 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 NH4Cl-containing solution. Under control conditions, NH4Cl 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) (DeltapH(i)) of 0.59 +/- 0.06 pH units in 12 min]; the initial rate of cell acidification (dpH(i)/dt) was 0.06 +/- 0.01 pH units/min. Incubation of the oocytes in the presence of H2O2 or beta-amyloid protein had no marked effect on the NH4Cl-induced DeltapH(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 DeltapH(i) and dpH(i)/dt. These increases in DeltapH(i) and dpH(i)/dt were prevented by the ROS scavengers histidine and desferrioxamine, suggesting involvement of the reactive species (1)DeltagO2 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 NH4+ influx into the oocyte via activation of a DPC-sensitive nonselective cation conductance pathway.
Cougnon
Marc
M
Institut National de la Santé et de la Recherche Médicale Unité 467, Université Paris V, Faculté de Médecine Necker-Enfants Malades, 75730 Paris Cedex 15, France.
Benammou
Samia
S
Brouillard
Franck
F
Hulin
Philippe
P
Planelles
Gabrielle
G
D000643:Ammonium Chloride / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D002412:Cations / Q000378:metabolism; D016166:Free Radical Scavengers / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007424:Intracellular Fluid / Q000378:metabolism; D017736:Ion-Selective Electrodes; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008839:Microelectrodes; D009865:Oocytes / Q000378:metabolism*; D016877:Oxidants / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D010539:Permeability / Q000187:drug effects; D010777:Photochemistry; D017382:Reactive Oxygen Species / Q000378:metabolism / Q000494:pharmacology*; D012395:Rose Bengal / Q000494:pharmacology; D019820:Xanthine / Q000378:metabolism / Q000494:pharmacology; D014969:Xanthine Oxidase / Q000378:metabolism / Q000494:pharmacology; D014982:Xenopus laevis; D020122:tert-Butylhydroperoxide / Q000494:pharmacology
D002412:Cations; D016166:Free Radical Scavengers; D016877:Oxidants; D017382:Reactive Oxygen Species; D000643:Ammonium Chloride; D019820:Xanthine; D012395:Rose Bengal; D020122:tert-Butylhydroperoxide; D006861:Hydrogen Peroxide; D014969:Xanthine Oxidase