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

1 Department of Anatomy and Physiology, Environmental and Hyperbaric Cell Biology Facility, and 2 Department of Community Health, Wright State University School of Medicine, Dayton, Ohio 45435 Submitted 28 July 2003 ; accepted in final form 5 December 2003 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 (pH i ). pH i 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 pH i in 270 of 436 (62%) SC neurons tested. Chloramine-T (CT), N -chlorosuccinimide (NCS), dihydroxyfumaric acid, and H 2 O 2 decreased pH i by 0.19 ± 0.007, 0.20 ± 0.015, 0.15 ± 0.013, and 0.08 ± 0.002 pH unit, respectively. Hypercapnia decreased pH i by 0.26 ± 0.006 pH unit ( n = 95). The combination of hypercapnia and CT or NCS had an additive effect on pH i , causing a 0.42 ± 0.03 ( n = 21) pH unit acidification. CT slowed pH i recovery mediated by Na + /H + exchange (NHE) from NH 4 Cl-induced acidification by 53% ( n = 20) in -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 pH i . These results indicate that oxidative stress 1 ) decreases pH i in some SC neurons, 2 ) together with hypercapnia has an additive effect on pH i , 3 ) partially inhibits NHE, and 4 ) directly affects excitability of CO 2 /H + -chemosensitive SC neurons independently of pH i 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. hyperoxic hyperventilation; O 2 toxicity; pH regulation; brain stem; reactive oxygen species Address for reprint requests and other correspondence: J. B. Dean, Dept. of Anatomy and Physiology, Rm. 235C Bio. Sci Bldg., 3640 Col. Glenn Hwy., Wright State Univ., Dayton, OH 45435 (E-mail: jay.dean{at}wright.edu ).