Ca²⁺-independent phospholipase A₂ is required for agonist-induced Ca²⁺ sensitization of contraction in vascular smooth muscle

Excitatory agonists can induce significant smooth muscle contraction under constant free Ca²⁺ through a mechanism called Ca²⁺ sensitization. Considerable evidence suggests that free arachidonic acid plays an important role in mediating agonist-induced Ca²⁺-sensitization; however, the molecular mechanisms responsible for maintaining and regulating free arachidonic acid level are not completely understood. In the current study, we demonstrated that Ca²⁺-independent phospholipase A₂ (iPLA₂) is expressed in vascular smooth muscle tissues. Inhibition of the endogenous iPLA₂ activity by bromoenol lactone (BEL) decreases basal free arachidonic acid levels and reduces the final free arachidonic acid level after phenylephrine stimulation, without significant effect on the net increase in free arachidonic acid stimulated by phenylephrine. Importantly, BEL treatment diminishes agonist-induced Ca²⁺ sensitization of contraction from 49 ± 3.6 to 12 ± 1.0% (p < 0.01). In contrast, BEL does not affect agonist-induced diacylglycerol production or contraction induced by Ca²⁺, phorbol 12,13-dibutyrate (a protein kinase C activator), or exogenous arachidonic acid. Further, we demonstrate that adenovirus-mediated overexpression of exogenous iPLA₂ in mouse portal vein tissue significantly potentiates serotonin-induced contraction. Our data provide the first evidenee that iPLA₂ is required for maintaining basal free arachidonic acid levels and thus is essential for agonist-induced Ca²⁺-sensitization of contraction in vascular smooth muscle.