Figure 1.

Concentration-response relationships of agonists at WT D₂R and S193^5.42A. Concentration–response curves for GIRK activation elicited by application of DA, (S)-5-OH-DPAT, (R)-5-OH-DPAT, and p-tyramine (structures shown in panel (A)) in oocytes co-expressing GIRK1/4 subunits and RGS4 with (B) WT D₂R and (C) D₂R S193^5.42A. (D) Agonist pEC₅₀s at WT and S193^5.42A D₂R: pEC₅₀ for DA (WT: pEC₅₀ = 7.70 ± 0.07, n = 5; S193^5.42A: pEC₅₀ = 5.03 ± 0.04, n = 8), (S)-5-OH-DPAT (WT: pEC₅₀ = 8.28 ± 0.08, n = 11; S193^5.42A: pEC₅₀ = 6.56 ± 0.05, n = 12), (R)-5-OH-DPAT (WT: pEC₅₀ = 6.85 ± 0.16, n = 11; S193^5.42A: pEC₅₀ = 7.07 ± 0.08, n = 14), and p-tyramine (WT: pEC₅₀ = 4.00 ± 0.21, n = 6; S193^5.42A: pEC₅₀ = 4.32 ± 0.12, n = 6–12). Comparison of pEC₅₀s using two-way ANOVA yielded significant main effects of agonist (F_{(3, 65)} = 281.8) and of the S193^5.42A mutation (F_{(1, 65)} = 139.2), as well as a significant interaction between these two factors (F_{(3, 65)} = 79.98, p < 0.001 for each main effect). Sidak’s multiple comparisons test further revealed that the pEC₅₀s of DA and (S)-5-OH-DPAT, but not p-tyramine and (R)-5-OH-DPAT, differed significantly between WT and mutant D₂R, as indicated by asterisks; ***, p < 0.001. (E) Relative efficacies at WT D₂R and S193^5.42A for DA (WT: 1.00 ± 0.02; S193^5.42A: 1.04 ± 0.04), (S)-5-OH-DPAT (WT: 0.52 ± 0.01; S193^5.42A: 0.64 ± 0.01), (R)-5-OH-DPAT (WT: 0.11 ± 0.01; S193^5.42A: 0.63 ± 0.02), and p-tyramine (WT: 0.09 ± 0.01; S193^5.42A: 0.37 ± 0.01). WT and S193^5.42A responses were normalized to the response evoked by 1 µM and 300 µM DA, respectively. The efficacy values were obtained from the fitted parameter Top, from the corresponding concentration-response curves (see Materials and Methods). The number of oocytes used for each condition is indicated on the bars in (D,E) and corresponds to the number recorded to generate the data points plotted in (B,C). Experiments were performed using a buffer perfusion rate of 1 mL/min. Data are presented as mean ± SEM.