Electron currents, H₂O₂ production, and membrane potential of VSOP/Hv1^−/− neutrophils. (A and B) Electron current recorded at −60 mV in a WT blood neutrophil in the perforated patch configuration. Currents were evoked by 100 nM PMA and blocked by 1 µM DPI. (B) Mean amplitude of the PMA-activated electron currents. Data are means ± SEM of 6 WT and 11 VSOP/Hv1^−/− neutrophils from five WT and nine VSOP/Hv1^−/− mice that were tested in 17 independent experiments. ns, not significant at P < 0.05 by an unpaired Student's t test. (C) Time-dependent H₂O₂ production in WT and VSOP/Hv1^−/− blood neutrophils activated with PMA in the absence or presence of 1 mM of the proton channel blocker Zn²⁺ or 40 µg/ml of the proton-permeable channel gramicidin. Data are from two representative experiments that were independently performed more than three times. (D) Mean H₂O₂ production from WT and VSOP/Hv1^−/− neutrophils. Data are means ± SD of three to five separate experiments done in triplicate from six WT and five VSOP/Hv1^−/− mice. ***, P < 0.0001; *, P < 0.05, unpaired Student's t test. (E and F) Membrane potential changes measured with DiBAC4(3) during sequential addition of 1 µM PMA and 100 µM Zn²⁺ to blood neutrophils. (E) Mean responses of eight WT and nine VSOP/Hv1^−/− neutrophils from four independent experiments. PMA evoked a larger depolarization in VSOP/Hv1^−/− cells. Arrow indicates the direction of depolarization. (F) Mean change in DiBAC4(3) fluorescence evoked by PMA and PMA+Zn²⁺. Data are means ± SEM of eight WT and nine VSOP/Hv1^−/− blood neutrophils from four independent experiments using five WT and four VSOP/Hv1^−/− mice. *, P < 0.05, unpaired Student's t test.