Morphine and morphine-like opiates are extensively used in the United States for treatment of pain associated with a wide variety of medical conditions. Although many drugs with strong analgesic properties exist, habituation and tolerance to their analgesic properties develop with prolonged use. For morphine and many related compounds, it is binding to the mu-receptor that mediates both the analgesic activity and the resultant habituation. Recently, it has been shown that some opioid compounds with lower intrinsic activity (i.e., efficacy) at the mu-receptor can nevertheless maintain potent analgesic activity, while possessing much lower abuse potential. Furthermore, low-efficacy mu-agonists have been shown to have potential as treatments for drug abuse. To better understand the relationships among intrinsic activity of a compound, its analgesic activities, and its abuse potential, we have devised the following experiments, using the high affinity, mu-selective antagonist, [3H]CTOP, and the neuroblastoma cell line, SH-SY5Y, which contains mu-receptors that are functionally linked to adenylate cyclase activity in the cells. Efficacies of opioid ligands from several families will be determined based on the definition that relative efficacy equals relative activity at equal receptor occupancy. The opioid activities to be measured will be the ability of compounds to (1) inhibit forskolin-stimulated cAMP accumulation in intact cells and (2) the ability to stimulate GTPase activity. Occupancy will be determined from binding studies in intact cells under conditions identical to those used for measurement of inhibition of cAMP accumulation. General information about receptor-effector coupling for mu- opioid receptors will be gained by treatment of cells with Beta-CNA to inactivate the receptors and pertussis toxin to inactive the G-protein, Gi. The determination of efficacies of selected opiates under these conditions will reveal information about the kinetics of coupling of the receptor to Gi. Efficacies of opiate will also be determined under unusual biological states, such as lowered ATP levels, a condition that would be caused by cerebral ischemia or hypoxia. These studies will give a better understanding of efficacies of addictive and nonaddictive analgesics and possibly provide a simple in vitro screen for low-efficacy opiates with low abuse potential.
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