Endocytosis machinery is required for β1-adrenergic receptor-induced hypertrophy in neonatal rat cardiac myocytes

Aims Cardiac hypertrophy by activation of the P-adrenergic receptor (PAR) is mediated more efficiently by the beta(1)-AR than by the beta(2)-AR. We investigated the signalling mechanism by which the beta(1)-AR mediates cardiac hypertrophy. Methods and results Experiments were performed in cultured neonatal rat cardiomyocytes. Hypertrophy was determined by the protein/DNA content and atrial. natriuretic factor transcription. Phosphorytation of Akt and Src was assessed by immunoblotting. Isoproterenot (ISO, 10 mu M), a non-selective beta-AR agonist, caused selective downregulation of the beta(1)-AR (control beta(1) vs. beta(2): 35 vs. 65%, B-max 78 +/- 4 fmol/mg; 4 h, 10 vs. 90%, 61 +/- 5 fmol/mg). Concanavalin A (Con A, 0.5 mu g/mL), an inhibitor of endocytosis, prevented downregulation of beta(1)-ARs by ISO treatment (4 h, 35 vs. 65%, 73 +/- 8 fmol/mg), suggesting that beta(1)-ARs selectively undergo endocytosis. Interference with beta(1)-AR endocytosis by Con A, carboxyl terminal peptide of beta-AR kinase-1, dominant negative (DN) beta-arrestin-1, or DN dynamin inhibited p-adrenergic hypertrophy, suggesting that the endocytosis machinery plays a key role in mediating beta-adrenergic hypertrophy. Activation of Akt by the beta(1)-AR was blocked by inhibition of the endocytosis machinery, suggesting that endocytosis mediates activation of Akt. Akt plays a critical role in beta-adrenergic hypertrophy, since DN Akt blocked ISO-induced hypertrophy. beta-Adrenergic activation of Akt is mediated by Src, which associates with the endocytosis machinery and is necessary and sufficient to mediate beta-adrenergic hypertrophy. Conclusion Activation of the endocytosis machinery is required for activation of Akt, which, in turn, critically mediates beta(1)-AR-induced cardiac hypertrophy.