Effects of hydroxyurea on intracellular calcium concentration in HUVECs. (A) The effect of hydroxyurea on [Ca2⁺]_i was independent of the time of its application (left panel vs right panel) and was not mimicked by the addition of medium 199 (right panel). Traces shown are representative from at least 3 experiments, each performed on 15 to 30 single cells during incubation with hydroxyurea. Gray areas indicate the duration of hydroxyurea treatment. Notice a variety in times of initiation of [Ca2⁺]_i response after the addition of hydroxyurea. (B) Dependence of hydroxyurea-induced calcium response on calcium mobilization from intracellular stores. Comparison of [Ca2⁺]_i signals induced by hydroxyurea (bottom trace) and endothelin-1, the native calcium-mobilizing agonist for these cells (upper panel). (C) Ionomycin-induced [Ca2⁺]_i signals in controls (upper trace) and hydroxyurea-treated cells (bottom trace). (D) The lack of effects of hydroxyurea on [Ca2⁺]_i response in cells with blocked endoplasmic reticulum calcium pump with thapsigargin. To avoid the UV damage of cells, the ratio of light intensities during thapsigargin treatment was followed at a rate of 1 point per 20 seconds.