Real-Time G-Protein-Coupled Receptor Imaging to Understand and Quantify Receptor Dynamics
Figure 7
Study of D2LR internalization. Receptor internalization was studied for 180 min on HEK-293 cells expressing D2LR-YFP or with D2LR-YFP and GRK2. A stack comprising the central part of the cells was used for analysis. Fluorescence intensity was measured across a line crossing the cell in (a), (d), and (g) and was computed and plotted in, respectively (b), (e), and (h). For every plot the minimum of fluorescence intensity (blue circle) and the two maxima at both sides of the minimum (red and green circles) were detected. A ratio was established between the two maxima and the minimum, and the ratio evolution over time was plotted for each cell (c, f, and i). Normalized ratio of all analyzed cells is represented in (j). Using this approach, it was possible to detect statistical significant differences (One-way ANOVA) between 100 nM quinpirole-treated cells expressing D2LR-YFP and GRK2 and untreated or quinpirole-treated cells 97 minutes after treatment addition. In the absence of GRK2 coexpression there is not any detectable D2LR-YFP internalization.
