ACTIVATION of phospholipase C (PLC) in response to stimulation of δ-opioid, m4 muscarinic and α2 adrenergic receptors was observed in NG108-15 cells. Treatment with PLC specific inhibitors, U73122 and ET-18-OCH3, blocked δ-opioid receptor-mediated activation of G proteins with no effect on opioid binding to the receptors. U73122 treatment also suppressed functional responses of m4 muscarinic and α2 adrenergic receptors in NG108-15 cells. Furthermore, the G protein activation mediated by μ- and δ-opioid receptors and opioid receptor-like receptor (ORL1) were abolished by U73122 in SK-N-SH neuroblastoma cells. Inhibition of adenylyl cyclase induced by high concentrations of GTP was blocked by U73122, suggesting that blockade is at the level of G proteins. Our results thus indicated that inhibition of PLC leads to blockade of Gi protein activation mediated by opioid receptors or other Gi-coupled receptors.