MECHANISMS OF OPIATE/OPIOID PEPTIDE ACTION

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The long-range goal of these studies is to understand the mechanisms by which opiates and opioid peptides alter normal neuronal function by analyzing receptor-mediated events in cultured cells of nervous system origin. We propose to use neurotumor cells and sensory neurons that possess Mu, Kappa and Delta receptors to study both the acute and chronic effects of opiates and opioid peptides so that eventually we can devise a rational therapy for narcotic addiction. Acute effects of opiates (0-30min) will be studied in a neuroblastoma x Chinese hamster neuron hybrid cell line (NCB-20) which expresses both delta (enkephalin) and sigma-like (benzomorphan) opiate receptors and voltage-sensitive calcium channels (VSCC), and in rat pheochromocytoma (PC12) cells which express VSCC, release neurotransmitters such as norepinephrine (NE), and which can be induced with NGF to express enkephalin receptors. Particular emphasis will be placed on the role of opiates in regulating calcium fluxes internal calcium mobilization, cyclic AMP-dependent protein phosphorylation, calcium-dependent protein phosphorylation, polyphosphoinositide turnover, inositol triphosphate release and modulation of K+ flux. We will also use dorsal root ganglion cells in which substance P release, evoked by cell depolarization, is blocked by opiate agonists. It is our primary hypothesis that an opiate-induced reduction of cellular cyclic AMP levels may result in a reduced phosphorylation of a dihydropyridine-sensitive calcium channel in these cells. This would lead to a reduction in evoked influx of calcium ions and subsequent inhibition of transmitter release.