FIGURE 3.
Quantification of internalization and absence of rapid recycling of CCK2R in HEK cells Flp-InTM CCK2R-293. A, quantification of internalization. Cells were preincubated with CCK (0.1 μm) and different pharmacological agents at 37 °C for the times indicated and, after washing out non-internalized ligand, were incubated with Alexa F 647-CCK at a saturating concentration (1 μm) for quantification of membrane CCK2R. B, recycling. Cells were treated with CCK (0.1 μm) for 1 h, washed several times, and incubated with Alexa F 647-CCK (1 μm) for 45 min or 1, 2, or 3 h after the end of CCK stimulation. Confocal microscopy images are inserted to show the lack of effect of the recycling inhibitor monensin (50 μm) on receptor density at the cell surface. Images are representative of at least three separate experiments. Error bars, S.E.