(A) Cardiomyocytes were unstimulated (Con), exposed to 100 μM PE, 50 μM isoprenaline (ISO; 5 min) or to 20 μM propranolol (Prop; 15 min), or exposed to propranolol (10 min) before addition of PE (Prop/PE) or isoprenaline (Prop/ISO) (5 min). Samples were immunoblotted for phospho-MKK1/2 (P-MKK1/2), total MKK1/2 (T-MKK1/2), phospho-ERK1/2 (P-ERKs) and total ERK1/2 (T-ERKs). The experiment was repeated with similar results. (B) Immunoblots of GAPDH or CREB in cytosolic (C) and NPE (N) fractions from cardiomyocytes. (C and E) Cardiomyocytes were unstimulated (Con), or exposed to 100 nM ET-1, 100 μM PE or 50 nM A61603 for the times indicated (C) or for 5 min (E). Fractions were immunoblotted for phospho-ERK1/2 or total ERK1/2. Blots are representative of at least three experiments with different cardiomyocyte preparations. (D) Densitometric analysis of the blots in (C). The graphs on the left-hand side show data normalized to maximum values. The graphs on the right-hand show the relative percentage in each fraction. Results are means±S.E.M. for three independent myocyte preparations.