Fig. 4. Time course of the effect of thermal stress and synergistic effect of Cu²⁺ combined with hyperthermia (30°C) upon p38-MAPK phosphorylation in M. galloprovincialis mantle tissue. Phosphorylated p38-MAPK was detected in extracts (100 µg of protein) from control animals maintained at 15°C (Con) or animals maintained at 4°C (A, top panel) or 30°C (A, bottom panel) for the indicated times. The molecular mass markers (kDa) are shown to the right of the panel. (C) Phosphorylated p38-MAPK (top panel) or total p38-MAPK (bottom panel) levels were detected in extracts (100 µg of protein) from control animals maintained at 15°C and animals maintained at 30°C for 30 min, either in the absence or presence of 1 µmol l^–1 CuCl₂. Western blots shown are representative of four independent experiments. (B,D) Densitometric analysis of phospho-p38-MAPK bands by laser scanning. Results are means ± S.E.M. for four independent experiments performed with similar results. P<0.001, *P<0.05, **P<0.01 versus control value.