Figure 2
Agonist-induced internalization of μ-OR in genetically modified cell lines. Internalization of μ-OR in response to a range of concentrations of (a) DAMGO, (b), fentanyl (c) loperamide, or (d) morphine in parental HEK293A (WT) cells or HEK293A cell lines with deletion of GRK2 (ΔGRK2), GRK3 (ΔGRK3), GRK2 and -3 (ΔGRK2/3) or β-arrestin1 and -2 (Δβ-Arr1/2). Data represent the mean ± SEM of the area under the curve of 90 min real-time internalization experiments after subtraction of the buffer response in parental cells from 3–5 independent experiments carried out in duplicate. Error bars not shown lie within the dimension of the symbol. (e) Emax of agonist responses in parental cells. Statistical comparison by one-way ANOVA with Dunnett’s multiple comparisons test. (f) Basal μ-OR internalization determined by taking the area under the curve of the buffer responses. (g) μ-OR cell surface expression determined by measuring donor signals in absence of acceptor. (e–g) Data represent the mean of individual experiments (circles) as well as the mean ± SEM (columns) from 4–5 (e), 3–4 (f) or 6–9 (g) independent experiments with 2 (e), 4–8 (f) or 32 (g) replicates per experiment. (f-g) Values in knockout cells were compared to the parental cells by one-way ANOVA with Dunnett’s multiple comparisons test. *P = 0.01–0.05, **P = 0.001–0.01, ***P < 0.001. a.u., arbitrary units.
