Angiotensin II increases cytosolic Ca²⁺ levels in human neutrophils. Fura-2–loaded neutrophils (10⁷ cells/mL) were incubated with 100 nM fMLP (A) or with 100 nM Ang II (B-E) under the indicated conditions for 4 minutes, and changes in intracellular Ca²⁺ levels were fluorometrically recorded. The arrow in each panel indicates the time of fMLP, Ang II, or Ptx addition. (A-B) Cell suspension was incubated in the presence of external 2 mM CaCl₂, and either fMLP (A) or Ang II (B) was added. (C) Cells were treated with 0.1 mM EGTA for 5 minutes in the absence of external Ca²⁺ before the addition of Ang II. (D) Intracellular Ca²⁺ stores were first depleted with 1 μM thapsigargin and 0.1 mM EGTA for 5 minutes, followed by washing. Then the cells were treated simultaneously with Ang II and 2 mM CaCl₂. (E) Cells were pretreated with Ptx before the addition of Ang II. Each graph is representative of a set of 3 experiments that yielded similar results.