Hypoxia-induced pulmonary vasoconstriction (HPV) is attributed to an increase in intracellular Ca²⁺ concentration ([Ca²⁺]_i) in pulmonary artery smooth muscle cells (PASMCs). We have reported that phospholipase C-γ1 (PLCγ1) plays a significant role in the hypoxia-induced increase in [Ca²⁺]_i in PASMCs and attendant HPV. In this study, we intended to determine molecular mechanisms for hypoxic Ca²⁺ and contractile responses in PASMCs. Our data reveal that hypoxic vasoconstriction occurs in pulmonary arteries, but not in mesenteric arteries. Hypoxia caused a large increase in [Ca²⁺]_i in PASMCs, which is diminished by the PLC inhibitor U73122 and not by its inactive analog U73433. Hypoxia augments PLCγ1-dependent inositol 1,4,5-trisphosphate (IP₃) generation. Exogenous ROS, hydrogen peroxide (H₂O₂), increases PLCγ1 phosphorylation at tyrosine-783 and IP₃ production. IP₃ receptor-1 (IP₃R1) knock-down remarkably diminishes hypoxia- or H₂O₂-induced increase in [Ca²⁺]_i. Hypoxia or H₂O₂ increases the activity of IP₃Rs, which is significantly reduced in protein kinase C-ε (PKCε) knockout PASMCs. A higher PLCγ1 expression, activity, and basal [Ca²⁺]_i are found in PASMCs, but not in mesenteric artery smooth muscle cells from mice exposed to chronic hypoxia (CH) for 21 days. CH enhances H₂O₂- and ATP-induced increase in [Ca²⁺]_i in PASMCs and PLC-dependent, norepinephrine-evoked pulmonary vasoconstriction. In conclusion, acute hypoxia uniquely causes ROS-dependent PLCγ1 activation, IP₃ production, PKCε activation, IP₃R1 opening, Ca²⁺ release, and contraction in mouse PASMCs; CH enhances PASM PLCγ1 expression, activity, and function, playing an essential role in pulmonary hypertension in mice.