Figure 1.

Representative experimental traces from individual SKNMC-H₃ cells showing the Na⁺-dependent pH_i recovery after acute exposure to an NH₄Cl acid pulse. (A) A control cell: The y axis represents the pH_i as determined from the intracellular calibration of the dye in this cell. The cell was initially superfused with Na⁺-Ringer's solution (NaR), then with 20 mM NH₄Cl. Acute exposure to NH₄Cl resulted in acidification of the cytosol to ≈pH_i 6.2 after its removal. In the absence of extracellular Na (0 Na), there was no measurable pH_i recovery. With the reintroduction of extracellular Na⁺ (NaR), pH_i increased with intracellular alkalinization occurring at a rate of 0.094 pH_i units/min (dotted line). The final pH_i, achieved in the cell shown in this trace, was higher than the starting pH_i (6.8 vs. 6.6). This overshoot of the final pH_i relative to the initial resting pH_i was seen in the majority of control cells studied and is shown in the bar graph accompanying A. For the 221 control cells studied, the initial pH_i was 6.69 ± 0.01 and the final pH_i was significantly higher at 6.76 ± 0.01 (P < 0.001). (B) Effect of imetit: The y axis represents pH_i as determined from the intracellular calibration of the dye in this cell. Imetit (100 nM) was present in the superfusate from the addition of NH₄Cl until the end of the protocol as shown. Acute exposure to NH₄Cl resulted in acidification of the cytosol to ≈6.2 after its removal. In the absence of extracellular Na⁺ (0 Na), there was no measurable pH_i recovery. With the reintroduction of extracellular Na⁺ (NaR), pH_i started to increase with intracellular alkalinization occurring at a rate of 0.050 pH_i units/min. The mean rate of Na⁺-dependent pH_i recovery was significantly less than that observed in the control cells (P < 0.0001) (Figs. 2 and 3). The final pH_i achieved in the cell shown in this trace was lower than the starting pH_i (6.7 vs. 6.5). This pH_i undershoot relative to the starting pH_i was seen in the majority of cells exposed to imetit, as shown in the bar graph of B. Unlike control cells, the final pH_i of the 277 imetit-treated cells was significantly lower than the initial pH_i (6.61 ± 0.01 vs. 6.74 ± 0.01; P < 0.0001).