Evidence for the requirement of OR internalization in the opioid activation of ERK in HEK-293 cells. Cells cotransfected with δ-, μ-, or κ-OR, DWT, or mutant K44A were treated with 1 μm DPDPE (A), 0.1 μm DADLE (B), 1 μmDAMGE (C), or 1 μm U69,593 (D) for different time intervals.Top, Representative autoradiograms showing the phosphorylated MBP bands. Bottom, Quantification of ERK activity. n = 2–6; *p < 0.05.