Fig. 3. ATP dose-response curve. (A) Relationship between total increase in capacitance C_m (measured as the difference between the final plateau value and the initial capacitance) and time in ATP-depleting medium. The time was counted until the time of patch rupture. Open squares and filled circles represent mean values using 300 µM Ca²⁺ (n=3-13) and 10 µM Ca²⁺ (n=5-10) in the pipette, respectively. (B) The same data sets as in A are shown but plotted as a function of intracellular ATP, as calculated using the fitting function from Fig. 1B, at the time of patch rupture. Solid lines in B are logistic functions fitted to the data. For 10 µM Ca²⁺ K_m=328 µM and the cooperativity factor P=2.0, for 300 µM Ca²⁺ K_m=95 µM and P=3.7. (C) Reversibility of ATP depletion. Cells were ATP-depleted for 39±6.5 minutes or 65±8 minutes to lower intracellular ATP to the critical level for secondary/tertiary granules and primary granules, respectively. These cells were then stimulated with 10 µM Ca²⁺ or 300 µM Ca²⁺, respectively, including 1 mM ATP in the patch pipette (reperfusion). Mean capacitance increase from control cells (no ATP depletion) or cells reperfused with ATP after depletion, n=4-8.