Abstract

The effect of lysophosphatidic acid on the phosphorylation and function of α_1b-adrenoceptors transfected into rat-1 fibroblasts was studied. This phospholipid mitogen increased in a concentration-dependent fashion (EC₅₀ ∼50 nM) the phosphorylation of these adrenoceptors. Lysophosphatidic acid-induced α_1b-adrenoceptor phosphorylation was relatively rapid (t_1/2 ∼1 min), intense (2.5-fold), and sustained for at least 60 min. The effect of lysophosphatidic acid was blocked by pretreatment with pertussis toxin. The α_1b-adrenoceptor phosphorylation induced by lysophosphatidic acid was not blocked by genistein, a tyrosine kinase inhibitor, but it was inhibited by inhibitors of protein kinase C (bisindolylmaleimide I, staurosporine, and Ro 31-8220) and phosphoinositide 3-kinase (wortmannin and LY 294002). The ability of norepinephrine to increase cytosol calcium concentration was markedly decreased in cells previously challenged with lysophosphatidic acid. Norepinephrine-induced [³⁵S]GTPγS binding in membrane preparations was used as an index of the functional coupling of the α_1b-adrenoceptors and G proteins. Norepinephrine-stimulated [³⁵S]GTPγS binding was markedly decreased in membranes from cells pretreated with lysophosphatidic acid. This effect of lysophosphatidic acid was blocked by pretreatment with wortmannin or staurosporine. Our data indicate that: 1) activation of lysophosphatidic acid receptors induce phosphorylation of α_1b-adrenoceptors; 2) this effect is mediated through pertussis toxin-sensitive G proteins, phosphatidylinositol 3-kinase, and protein kinase C; and 3) the phosphorylation of α_1b-adrenoceptors induced by the lipid mitogen is associated to adrenoceptor desensitization.