FIGURE 14.

Effect of the NHE antagonist HOE694 on pH recovery from an intracellular acid load in mouse primary hippocampal neurons. Primary cultures of mouse hippocampal neurons (14–21 days in vitro) were grown on 12-mm coverslips and loaded for 15 min with the pH-sensitive fluorescent dye BCECF-AM (50 μm). Coverslips were placed into a Live Cell Instruments magnetic chamber and incubated with HEPES-buffered ACSF (contains 126 mm NaCl). The cells were examined at 37 °C using a Zeiss LSM710 confocal microscope. A, ACSF was perfused onto the cells at ∼2 ml/min for 1 min prior to acid loading with NH₄Cl. Following NH₄Cl treatment, the cells were allowed to recover in Na⁺-rich ACSF medium containing increasing concentrations (10–1000 μm) of the NHE-specific antagonist HOE694 as indicated, or in some cases ACSF medium where the NaCl was substituted with molar equivalents of choline chloride (i.e. 0 mm NaCl). The rate of pH_i recovery following acidification was monitored as a function of time. B, steady-state pH_i of cultured hippocampal neurons was monitored as a function of time following addition of diluent (control) or increasing concentrations of HOE694. The data represent the mean ± S.E. of three independent experiments, with ∼10 cells per experiment.