Suppression of phospholipase C blocks G(i)-mediated inhibition of adenylyl cyclase activity

The potential effect of inhibition of phospholipase C on the response of G(i)-coupled receptors was investigated in neuroblastoma x glioma hybrid (NG108-15) cells. The phospholipase C specific inhibitor 1-[6-((17β-3- methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl]1H-pyrrole-2,5-dione (U73122), which did not affect basal and forskolin-stimulated adenylyl cyclase activities, time- and dose-dependently blocked δ-opioid receptor-mediated inhibition of adenylyl cyclase activity, the EC₅₀ (0.5 μM) of which was consistent with that for inhibition of bradykinin-dependent phospholipase C activation (EC₅₀ = 1 μM). U73122 treatment also blocked functional responses of m4 muscarinic receptor and α₂-adrenoceptor in NG108-15 cells and three opioid receptors (μ, δ and opioid receptor-like receptor (ORL₁)) in human neuroblastoma SK-N-SH cells. 1-[6-((17β-3-Methoxyestra-1,3,5(10)- trien-17-yl)amino)hexyl]-2,5-pyrrolidinedione (U73343), an inactive analog of U73122, did not show any effect, which suggests that the blockade by U73122 of G(i)-coupled receptor-mediated signaling is probably mediated through inhibition of phospholipase C, although a possible direct modification of G proteins can not be excluded. Furthermore, treatment with U73122 but not U73343 blocked the GTP-induced inhibition of adenylyl cyclase, indicating blockade at the level of G(i) proteins.