Fig. 2. Dynamics of ERK phosphorylation in primary hepatocytes (A), RTgill-W1 cells (B) and RTH-149 hepatoma cells (C) from rainbow trout exposed to hypo-osmolarity, 30 nmol l^–1 EGF or 10 µmol l^–1 CuCl₂, for up to 60 min as indicated. Numbers below the blots are changes in pERK abundance, expressed relative to controls at time zero, denoted as means ± s.e.m. of at least 3 independent experiments. *Statistically significant difference from the control value, as assessed on non-normalized data (P<0.05). c60', controls incubated in standard conditions for 60 min. Activated ERK was determined by western blot analysis using an antibody against the dually phosphorylated form of ERK. The main band detected with this antibody migrated at approximately 44 kDa. In A, the second panel shows an example of a western blot analysis of total ERK abundance using an antibody against the phosphorylated and non-phosphorylated form of ERK (tERK). Similar analyses were made for all cell types under all conditions and indicated that ERK abundance was not altered by the treatments over the time course studied.