Phosducin, β-arrestin and opioid receptor migration

Internalization of G protein-coupled opioid receptors depends on multiple criteria, including the affinity of drugs to their receptors and the state of the receptor-G protein interaction. Most recent studies reveal that cytosolic components like phosducin and arrestin interfere with receptor internalization, that is phosducin impairs receptor phosphorylation and arrestin enhances endocytosis by uncoupling the receptor from its G protein. This study was designed to examine the mutual effect phosducin and arrestin exert on receptor endocytosis. Neuronal NG 108-15 hybrid cells transiently expressing the mu-opioid receptor, which has been fused to green fluorescence protein, were employed to study internalization of the fluorescent mu-opioid receptor construct in living cells by means of confocal laser scanning microscopy. Fluorescent mu-opioid receptors were detected in drug-naive cells both at the cell membrane and at cell surface protrusions, most likely filopodia, microspikes and retraction fibres. The opioid receptors present in the cell membrane internalize upon etorphine (1 nM) exposure, a process clearly blocked in cells overexpressing phosducin. However, coexpression of both phosducin and beta-arrestin 1 reverses this blockade. In contrast to etorphine, morphine fails to internalize mu-receptors expressed in NG 108-15 cells. When arrestin is overexpressed in these cells, morphine gains the ability to induce endocytosis, and this process is left unaffected by phosducin. The findings suggest that endocytosis of activated mu-opioid receptors primarily depends on arrestin-triggered uncoupling of the receptor from its G protein complex. Drug-induced receptor phosphorylation appears of subordinate significance for receptor internalization. (C) 1999 Elsevier Science B.V. All rights reserved.