Endocytosis machinery is required for beta1-adrenergic receptor-induced hypertrophy in neonatal rat cardiac myocytes.
OBJECTIVE
Cardiac hypertrophy by activation of the beta-adrenergic receptor (beta AR) is mediated more efficiently by the beta1-AR than by the beta2-AR. We investigated the signalling mechanism by which the beta1-AR mediates cardiac hypertrophy.


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 microM), a non-selective beta-AR agonist, caused selective downregulation of the beta1-AR (control beta1 vs. beta2: 35 vs. 65%, Bmax 78 +/- 4 fmol/mg; 4 h, 10 vs. 90%, 61 +/- 5 fmol/mg). Concanavalin A (Con A, 0.5 microg/mL), an inhibitor of endocytosis, prevented downregulation of beta1-ARs by ISO treatment (4 h, 35 vs. 65%, 73 +/- 8 fmol/mg), suggesting that beta1-ARs selectively undergo endocytosis. Interference with beta1-AR endocytosis by Con A, carboxyl terminal peptide of beta-AR kinase-1, dominant negative (DN) beta-arrestin-1, or DN dynamin inhibited beta-adrenergic hypertrophy, suggesting that the endocytosis machinery plays a key role in mediating beta-adrenergic hypertrophy. Activation of Akt by the beta1-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.


CONCLUSIONS
Activation of the endocytosis machinery is required for activation of Akt, which, in turn, critically mediates beta1-AR-induced cardiac hypertrophy.
Morisco
Carmine
C
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.
Marrone
Chiara
C
Galeotti
Jonathan
J
Shao
Dan
D
Vatner
Dorothy E
DE
Vatner
Stephen F
SF
Sadoshima
Junichi
J
D058665:Adrenergic beta-1 Receptor Agonists; D000318:Adrenergic beta-Agonists / Q000633:toxicity*; D000818:Animals; D000831:Animals, Newborn; D019390:Arrestins / Q000235:genetics / Q000378:metabolism; D009320:Atrial Natriuretic Factor / Q000235:genetics / Q000378:metabolism; D006332:Cardiomegaly / Q000139:chemically induced / Q000378:metabolism* / Q000473:pathology; D048429:Cell Size; D002478:Cells, Cultured; D003208:Concanavalin A / Q000494:pharmacology; D034281:Dynamins / Q000235:genetics / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects*; D004789:Enzyme Activation; D044387:GTP-Binding Protein beta Subunits / Q000378:metabolism; D044388:GTP-Binding Protein gamma Subunits / Q000378:metabolism; D007545:Isoproterenol / Q000633:toxicity*; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D018342:Receptors, Adrenergic, beta-1 / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D013997:Time Factors; D014158:Transcription, Genetic / Q000187:drug effects; D000071559:beta-Arrestin 1; D000071557:beta-Arrestins; D019061:src-Family Kinases / Q000378:metabolism
D058665:Adrenergic beta-1 Receptor Agonists; D000318:Adrenergic beta-Agonists; C000605256:Arrb1 protein, rat; D019390:Arrestins; C118981:G-protein Beta gamma; D044387:GTP-Binding Protein beta Subunits; D044388:GTP-Binding Protein gamma Subunits; D018342:Receptors, Adrenergic, beta-1; D000071559:beta-Arrestin 1; D000071557:beta-Arrestins; D003208:Concanavalin A; D009320:Atrial Natriuretic Factor; D019061:src-Family Kinases; D051057:Proto-Oncogene Proteins c-akt; D034281:Dynamins; D007545:Isoproterenol