Abstract
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.