Oxidative stress decreases pH_i and Na⁺/H⁺ exchange and increases excitability of solitary complex neurons from rat brain slices

Putative chemoreceptors in the solitary complex (SC) are sensitive to hypercapnia and oxidative stress. We tested the hypothesis that oxidative stress stimulates SC neurons by a mechanism independent of intracellular pH (pHi). pHi was measured by using ratiometric fluorescence imaging microscopy, utilizing either the pH-sensitive fluorescent dye BCECF or, during whole cell recordings, pyranine in SC neurons in brain stem slices from rat pups. Oxidative stress decreased pHi in 270 of 436 (62%) SC neurons tested. Chloramine-T (CT), N -chlorosuccinimide (NCS), dihydroxyfumaric acid, and H2O2 decreased pHi by 0.19 ± 0.007, 0.20 ± 0.015, 0.15 ± 0.013, and 0.08 ± 0.002 pH unit, respectively. Hypercapnia decreased pHi by 0.26 ± 0.006 pH unit ( n = 95). The combination of hypercapnia and CT or NCS had an additive effect on pHi, causing a 0.42 ± 0.03 ( n = 21) pH unit acidification. CT slowed pHi recovery mediated by Na+/H+ exchange (NHE) from NH4Cl-induced acidification by 53% ( n = 20) in \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{CO}\_{2}{/}\mathrm{HCO}\_{3}^{-}\) \end{document}-buffered medium and by 58% ( n = 10) in HEPES-buffered medium. CT increased firing rate in 14 of 16 SC neurons, and there was no difference in the firing rate response to CT with or without a corresponding change in pHi. These results indicate that oxidative stress 1 ) decreases pHi in some SC neurons, 2 ) together with hypercapnia has an additive effect on pHi, 3 ) partially inhibits NHE, and 4 ) directly affects excitability of CO2/H+-chemosensitive SC neurons independently of pHi changes. These findings suggest that oxidative stress acidifies SC neurons in part by inhibiting NHE, and this acidification may contribute ultimately to respiratory control dysfunction.