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This Article Full Text Full Text (PDF) All Versions of this Article: 290/3/H948    most recent 00868.2005v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Web of Science (2) Citing Articles via Google Scholar Google Scholar Articles by Hecquet, C. Articles by Erdös, E. G. Search for Related Content PubMed PubMed Citation Articles by Hecquet, C. Articles by Erdös, E. G.

Positive cooperativity between the thrombin and bradykinin B₂ receptors enhances arachidonic acid release

Departments of ¹Pharmacology and ²Anesthesiology, University of Illinois College of Medicine at Chicago, Chicago, Illinois

Submitted 12 August 2005 ; accepted in final form 20 September 2005

Bradykinin (BK) or kallikreins activate B₂ receptors (R) that couple G_i and G_q proteins to release arachidonic acid (AA) and elevate intracellular Ca²⁺ concentration ([Ca²⁺]_i). Thrombin cleaves the protease-activated-receptor-1 (PAR1) that couples G_i, G_q, and G_12/13 proteins. In Chinese hamster ovary cells stably transfected with human B₂R, thrombin liberated little AA, but it significantly potentiated AA release by B₂R agonists. We explored mechanisms of cooperativity between constitutively expressed PAR1 and B₂R. We also examined human endothelial cells expressing both Rs constitutively. The PAR1 agonist hexapeptide (TRAP) was as effective as thrombin. Inhibitors of components of G_i, G_q, and G_12/13 signaling pathways, and a protein kinase C (PKC)- inhibitor, Gö-6976, blocked potentiation, while phorbol, an activator, enhanced it. Several inhibitors, including a RhoA kinase inhibitor, a [Ca²⁺]_i antagonist, and an inositol-(1,3,4)-trisphosphate R antagonist, reduced mobilization of [Ca²⁺]_i by thrombin and blocked potentiation of AA release by B₂R agonists. Because either a nonselective inhibitor (isotetrandrine) of phospholipase A₂ (PLA₂) or a Ca²⁺-dependent PLA₂ inhibitor abolished potentiation of AA release by thrombin, while a Ca²⁺-independent PLA₂ inhibitor did not, we concluded that the mechanism involves Ca²⁺-dependent PLA₂ activation. Both thrombin and TRAP modified activation and phosphorylation of the B₂R induced by BK. In lower concentrations they enhanced it, while higher concentrations inhibited phosphorylation and diminished B₂R activation. Protection of the NH₂-terminal Ser¹-Phe² bond of TRAP by an aminopeptidase inhibitor made this peptide much more active than the unprotected agonist. Thus PAR1 activation enhances AA release by B₂R agonists through signal transduction pathway.

protease-activated-receptor-1; thrombin receptor activator peptide; protein kinases; potentiation; calcium; kallikrein