TABLE 2

Effect of KDN21 on the activation of ERK1/2 induced by δ and κ selective agonists HEK293 cells coexpressing δ and κ opioid receptors were grown to approximately 95% confluence and exposed to serum-free media for 6 h before the addition of the indicated opioid agonist and then incubated with 100 nM DPDPE, deltorphin II, U69593, or bremazocine in the presence or absence of KDN21 (100 nM) at 37°C for 10 min. The cells were extensively washed and prepared for MAPK extraction as described under Materials and Methods. Protein (10 μg) was separated via SDS-polyacrylamide gel electrophoresis, followed by immunoblotting using the antibody raised against phosphorylated MAPK. NIH Image software, ver. 1.61, was used to quantify phosphorylated MAPK level. The level of phosphorylated MAPK in the absence of ligand treatment is taken as control and defined as 100%. The data are means ± S.E. of three independent experiments.