Cardiovascular Research Advance Access originally published online on January 14, 2008
Cardiovascular Research 2008 78(1):36-44; doi:10.1093/cvr/cvn008
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Endocytosis machinery is required for β1-adrenergic receptor-induced hypertrophy in neonatal rat cardiac myocytes
1 Department of Cell Biology and Molecular Medicine, Cardiovascular Research Institute, UMDNJ, New Jersey Medical School, 185 South Orange Avenue, MSB G-609, Newark, NJ 07103, USA
2 Department of Internal Medicine, Federico II University, Naples, Italy
*Corresponding author. Tel: + 1 973 972 8619; fax: + 1 973 972 7489. E-mail address: sadoshju{at}umdnj.edu
Aims: Cardiac hypertrophy by activation of the β-adrenergic receptor (βAR) is mediated more efficiently by the β1-AR than by the β2-AR. We investigated the signalling mechanism by which the β1-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. Phosphorylation of Akt and Src was assessed by immunoblotting. Isoproterenol (ISO, 10 µM), a non-selective β-AR agonist, caused selective downregulation of the β1-AR (control β1 vs. β2: 35 vs. 65%, Bmax 78 ± 4 fmol/mg; 4 h, 10 vs. 90%, 61 ± 5 fmol/mg). Concanavalin A (Con A, 0.5 µg/mL), an inhibitor of endocytosis, prevented downregulation of β1-ARs by ISO treatment (4 h, 35 vs. 65%, 73 ± 8 fmol/mg), suggesting that β1-ARs selectively undergo endocytosis. Interference with β1-AR endocytosis by Con A, carboxyl terminal peptide of β-AR kinase-1, dominant negative (DN) β-arrestin-1, or DN dynamin inhibited β-adrenergic hypertrophy, suggesting that the endocytosis machinery plays a key role in mediating β-adrenergic hypertrophy. Activation of Akt by the β1-AR was blocked by inhibition of the endocytosis machinery, suggesting that endocytosis mediates activation of Akt. Akt plays a critical role in β-adrenergic hypertrophy, since DN Akt blocked ISO-induced hypertrophy. β-Adrenergic activation of Akt is mediated by Src, which associates with the endocytosis machinery and is necessary and sufficient to mediate β-adrenergic hypertrophy.
Conclusion: Activation of the endocytosis machinery is required for activation of Akt, which, in turn, critically mediates β1-AR-induced cardiac hypertrophy.
KEYWORDS Myocytes; Cell culture; Protein kinases; Protein phosphorylation
Time for primary review: 31 days
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