Phosducin, beta-arrestin and opioid receptor migration

Internalization of G protein-coupled opioid receptors depends on multiple c riteria, including the affinity of drugs to their receptors and the state o f the receptor-G protein interaction. Most recent studies reveal that cytos olic components like phosducin and arrestin interfere with receptor interna lization, that is phosducin impairs receptor phosphorylation and arrestin e nhances endocytosis by uncoupling the receptor from its G protein. This stu dy was designed to examine the mutual effect phosducin and arrestin exert o n receptor endocytosis. Neuronal NG 108-15 hybrid cells transiently express ing the mu-opioid receptor, which has been fused to green fluorescence prot ein, were employed to study internalization of the fluorescent mu-opioid re ceptor construct in living cells by means of confocal laser scanning micros copy. Fluorescent mu-opioid receptors were detected in drug-naive cells bot h at the cell membrane and at cell surface protrusions, most likely filopod ia, 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 p hosducin and beta-arrestin 1 reverses this blockade. In contrast to etorphi ne, 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. T he findings suggest that endocytosis of activated mu-opioid receptors prima rily depends on arrestin-triggered uncoupling of the receptor from its G pr otein complex. Drug-induced receptor phosphorylation appears of subordinate significance for receptor internalization. (C) 1999 Elsevier Science B.V. All rights reserved.