Figure 3.

NO delivered from donors produces a concentration-dependent biphasic effect on NHE1 activity. (A) Wild-type rat myocytes. Hepes-buffered superfusates, 37°C. The short half-life NO donor, sodium nitroprusside (SNP), was added to solutions 4–6 min before activating NHE1. With 1 µM SNP, NO is expected to peak at 600 nM during pH_i recovery (control: N = 35 cells from five animals; SNP: N = 31 cells from five animals). (B) Experiments repeated on myocytes that had been pre-treated with 6 µM ODQ (1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one), an inhibitor of guanylyl cyclase (GC; ODQ: N = 24 cells from four animals; ODQ+SNP: N = 26 cells from four animals). (C) CO₂/${\text{HCO}}_3^{-}$-buffered superfusates, 37°C; 30 µM dimethylamiloride (DMA) included to block NHE isoforms. pH_i recovery mediated by ${\text{HCO}}_3^{-}$ dependent transporters is unaffected by NO (control: N = 15 cells from three animals; SNP: N = 15 cells from three animals). (D) Wild-type rat myocytes. Hepes-buffered superfusates, 37°C. The long half-life NO donor NOC12 was added to solutions 10 min before activating NHE1. At a concentration of 5 µM, NOC12 is expected to maintain NO above 300 nM for the duration of pH_i recovery. (E) NHE1 fluxes. Control (N = 21 from five animals), effect of NOC12 (N = 18 from five animals), and effect of NOC12 on ODQ-pretreated cells (N = 26 from five animals). Significant difference between control and NOC12 (P < 0.0001), but not between control and NOC12+ODQ (P = 0.06). (F) Lower concentrations of NO (<30 nM) were produced by mixing donor NOC12 (300 µM) with scavenger CPTIO (100 µM). (G) NHE1 fluxes. Control (N = 21 from five animals), effect of NOC12/CPTIO (N = 19 from five animals), and effect of NOC12/CPTIO on ODQ-pretreated cells (N = 10 from four animals). Significant difference between control and NOC12 (P < 0.0001), but not between control and NOC12/CPTIO+ODQ (P = 0.08).