Figure 4
β-arrestin2 recruitment of μ-OR in genetically modified cell lines. Recruitment of β-arrestin2-RlucII to the cell membrane monitored by ebBRET in response to 6 min stimulation of μ-OR by 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), or GRK2 and -3 (ΔGRK2/3). Data represent the mean ± SEM of the BRET2 ratio after subtraction of the buffer response in parental cells from 3–4 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. **P = 0.001–0.01, ***P < 0.001. (f) Basal β-arrestin2 recruitment determined as the buffer response. (g) μ-OR surface expression measured by ELISA and normalized to expression in parental cells. (e–g) Data represent the mean of individual experiments (circles) and the mean ± SEM (columns) from 3 (e,f) or 3–4 (g) independent experiments with 2 (e), 6–8 (f) or 3 (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 and no significant differences were found (P > 0.05).
