Activation of NADPH oxidase-related proton and electron currents in human eosinophils by arachidonic acid.
1. Effects of arachidonic acid (AA) on proton and electron currents in human eosinophils were studied using the permeabilized-patch voltage-clamp technique, using an applied NH4+ gradient to control pH(i). 2. Superoxide anion (O2-) release was assessed by cytochrome c reduction in human eosinophils. Significant O2- release was stimulated by 5-10 microM AA. 3. AA activated diphenylene iodinium (DPI)-inhibitable inward current reflecting electron efflux through NADPH oxidase. These electron currents (I(e)) were elicited in human eosinophils at AA concentrations (3-10 microM) similar to those that induced O2- release. 4. The voltage-gated proton conductance (g(H)) in eosinophils stimulated with AA was profoundly enhanced: H+ current amplitude (I(H)) increased 4.6 times, activation was 4 times faster, and the H+ conductance-voltage (g(H)-V) relationship was shifted to substantially more negative voltages. The electrophysiological effects of AA resembled those reported for PMA, except that AA did not consistently slow tau(tail) (deactivation of H+ currents). 5. The stimulation of both proton and electron currents by AA was reversible upon washout. Repeated exposure elicited repeated responses. The activation of H+ currents by AA was dissociable from its activation of NADPH oxidase; H+ currents were enhanced at low concentrations of AA that did not elicit detectable I(e) or when NADPH oxidase was inhibited by DPI. 6. Most of the effects of AA on H+ currents qualitatively resemble those reported in whole-cell studies, reflecting a more direct action than PMA. The results are compatible with AA being an immediate activator of both NADPH oxidase and proton channels in human eosinophils.
Cherny
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Department of Molecular Biophysics and Physiology, Rush Presbyterian St Luke's Medical Center, Chicago, IL 60612, USA. tdecours@rush.edu
Henderson
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LM
Xu
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Thomas
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DeCoursey
T E
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D016718:Arachidonic Acid / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004583:Electrons; D004804:Eosinophils / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D015640:Ion Channel Gating / Q000187:drug effects*; D007473:Ion Channels / Q000819:agonists*; D019255:NADPH Oxidases / Q000502:physiology*; D010101:Oxygen Consumption / Q000187:drug effects; D011522:Protons; D013268:Stimulation, Chemical
D007473:Ion Channels; D011522:Protons; D016718:Arachidonic Acid; D019255:NADPH Oxidases