INVOLVEMENT OF A PERTUSSIS TOXIN-SENSITIVE G-PROTEIN-COUPLED PHOSPHOLIPASE-A2 IN AGONIST-STIMULATED ARACHIDONIC-ACID RELEASE IN MEMBRANES ISOLATED FROM BOVINE IRIS SPHINCTER SMOOTH-MUSCLE

We have shown that in bovine iris sphincter membranes G proteins are i nvolved in coupling muscarinic-, PGF2alpha-, endothelin- and platelet- activating factor receptors to the activation of phospholipase A2 and the release of arachidonic acid. GTPgammaS and GTPgammaS plus carbacho l stimulated arachidonic acid release in the membranes in a dose- and time-dependent manner. Nucleotide stimulation was specific to GTPgamma S, since GDP, GDPbetaS and ATP had no effect. The stimulatory effect o f GTPgammaS plus carbachol was blocked by atropine and it required the presence of physiological concentrations of Ca2-. AIF4-, which bypass es the receptor and directly activates the G protein, induced arachido nic acid liberation in the intact ins sphincter, but was ineffective i n the membranes. Addition of GTPgammaS plus carbachol to sphincter mus cle membranes prelabeled with [H-3]inositol or H-3-arachidonic acid re sulted in the formation of lysophosphatidylinositol and the liberation of arachidonic acid, thus suggesting the involvement of phospholipase A2. In vitro treatment of the iris membranes with pertussis toxic inh ibited arachidonic acid release by the agonists. This is in contrast t o the pertussis toxin-insensitive G protein that activates phospholipa se C in this tissue (22). These data demonstrate that in the iris sphi ncter a G protein is involved in the step between receptor activation and the activation of phospholipase A2, and that arachidonic acid rele ase in this tissue is mediated by a pertussis-toxin-sensitive G protei n-coupled phospholipase A2. Thus, GTP can regulate arachidonic acid re lease and its subsequent conversion into eicosanoids by stimulating it s formation.