Effect of Ca²⁺ on the magnitude of the rehydration response. Experimental conditions were the same as in Figure 3A except that the total Ca²⁺ concentrations in the cell suspension were those indicated in panels B to D. Ionophore addition to 5% Hct RBC suspensions when the [Ca²⁺]_s was higher than 100 μM was followed by rapid and extensive RBC lysis, unrelated to slow Pcat-mediated hydration, thus precluding investigation of Pcat at higher Ca²⁺ levels. In panel A the medium contained 50 μM Na-EGTA and no added calcium in order to chelate contaminant [Ca²⁺]_o (less than 10 μM) and ensure [Ca²⁺]_i levels below Gardos channel activation.