Abstract

The cysteinyl leukotrienes (cysLTs) LTC₄, LTD₄, and LTE₄ are lipid mediators with physiological and pathophysiological functions. They exert their effects through G protein-coupled receptors (GPCRs), most notably via CysLT₁ and CysLT₂ receptor. The roles of the CysLT₂ receptor are beginning to emerge. Both LTC₄ and LTD₄ are potent agonists for the CysLT₂ receptor; however, LTC₄ is rapidly converted to LTD₄, which is also the main endogenous ligand for the CysLT₁ receptor. A selective and potent agonist at the CysLT₂ receptor would facilitate studies to discern between receptor subtypes. We show here that N-methyl LTC₄ (NMLTC₄), a metabolically stable LTC₄ mimetic, is a potent and selective CysLT₂ receptor agonist. Two expression systems were used to evaluate the functional activity of NMLTC₄ at human and/or mouse CysLT₁ and CysLT₂ receptors. Through the aequorin cell-based assay for calcium-coupled GPCRs, NMLTC₄ was almost equipotent to LTC₄ at CysLT₂ receptors but was the least efficacious at CysLT₁ receptors. In a β-galactosidase–β-arrestin complementation assay, the human (h) CysLT₂ receptor can couple with β-arrestin-2, and NMLTC₄ is slightly more potent for eliciting β-arrestin-2 binding compared with cysLTs. Furthermore, LTE₄ is nearly inactive in this assay compared with its weak partial agonist activity in the aequorin system. In a vascular leakage assay, NMLTC₄ is potent and active in mice overexpressing hCysLT₂ receptor in endothelium, whereas the response is abrogated in CysLT₂ receptor knockout mice. Therefore, NMLTC₄ is a potent subtype selective agonist for the CysLT₂ receptor in vitro and in vivo, and it will be useful to elucidate its biological roles.