Figure 6

Effect of δ-OR or EGFR down-regulation on ERK phosphorylation by acute exposure to DSLET or EGF. HEK-δ-OR cells were serum-starved overnight prior to exposure to DSLET (10 μM for 24 hours) or EGF (100 ng/ml for 24 hours) to induce receptor down-regulation. After chronic agonist pre-treatment, cell monolayers were washed once with fresh media and exposed, acutely, to either DSLET (1 μM for 5 minutes) or EGF (10 ng/ml for 5 minutes). After acute agonist exposure, cell lysates were prepared and ERK phosphorylation was determined by SDS PAGE, followed by immunoblotting of proteins onto nitrocellulose. PhosphoERK -immunopositive bands were quantified using the MCID system. Band intensities are expressed graphically (hatched bars) as: grain density × band area - [region of detection (ROD)] × pixel. Data points represent the mean ± SEM of three experiments. ^*p ≤ 0.05 compared to untreated samples and ^**p ≤ 0.01 compared to untreated samples as determined by ANOVA and the Tukey post-hoc test.