Functional expression of mammalian opioid receptors in insect cells and high-throughput screening platforms for receptor ligand mimetics

Lepidopteran cell lines have been engineered to constitutively express high levels of mouse opioid receptors either alone or in combination with human G16 protein. Biochemical and pharmacological studies demonstrate that these lines contain all the mediator G proteins and downstream effectors required for opioid receptor function, including phospholipase C, and that expression of exogenous G16 does not contribute significantly to increased receptor responses upon activation. The activation of the phospholipase C pathway in the transformed cells upon stimulation with known receptor ligands results in easily and quantitatively measurable increases in free intracellular calcium, which can be monitored by automated fluorescent methods, while the addition of specific antagonists blocks the agonist-induced responses. Therefore, the transformed lepidopteran cell lines can be used as sensitive high-throughput screening platforms for fast detection of opioid receptor ligand mimetics (agonists and antagonists) in collections of natural products and synthetic compounds.Received 3 December 2004; received after revision 3 February 2005; accepted 10 February 2005L. Swevers and E. Morou contributed equally to this work.