Phosphorylation of contractile proteins in response to alpha- and beta-adrenergic stimulation in neonatal cardiomyocytes.
alpha- and beta-Adrenergic receptor agonists induce an inotropic response in the adult heart by promoting the phosphorylation of several regulatory proteins, including myosin-binding protein C (MyBP-C), cardiac troponin I (cTnI), and phospholamban (PLB). However, the adrenergic-induced phosphorylation of these proteins has not been characterized in the developing heart. Accordingly, we evaluated MyBP-C, cTnI, and PLB phosphorylation in cultured neonatal rat cardiomyocytes (NRCMs) after alpha- and beta-receptor activation with phenylephrine and isoproterenol. alpha-Receptor stimulation increased, whereas beta-receptor activation reduced MyBP-C phosphorylation. Isoelectric-focusing experiments indicated that the amount of monophosphorylated MyBP-C was sensitive to alpha-adrenergic activation, but diphosphorylated and triphosphorylated MyBP-C levels were largely unaffected. The phosphorylation of cTnI and PLB was consistent with the mechanism observed in adult hearts: alpha- and beta-Receptor stimulation phosphorylated both proteins. For cTnI, the greatest difference associated with beta-receptor activation was observed in the diphosphorylated state, whereas alpha-receptor activation was associated with a marked increase in the tetraphosphorylated protein and absence of the unphosphorylated state. Despite these apparent changes in cTnI and PLB phosphorylation, beta-receptor activation failed to alter calcium transients in NRCMs. Collectively, these findings suggest that, unlike cTnI and PLB, MyBP-C and inotropy are not coupled to beta-adrenergic stimulation in NRCMs. Therefore, cTnI and PLB probably play a more central role in modulating contractile function in NRCMs in response to catecholamines than does MyBP-C, and MyBP-C may have a structural role in stabilizing thick filament assembly rather than influencing cross-bridge formation in developing hearts.
Decker
Robert S
RS
Feinberg Cardiovascular Research Institute, Northwestern University Medical Center, Chicago, Ill 60611, USA.
Rines
Amy K
AK
Nakamura
Sakie
S
Naik
Tejaswitha J
TJ
Wassertsrom
J Andrew
JA
Ardehali
Hossein
H
D000818:Animals; D000831:Animals, Newborn; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D007545:Isoproterenol / Q000494:pharmacology; D009200:Myocardial Contraction; D032383:Myocytes, Cardiac / Q000502:physiology*; D010766:Phosphorylation; D051381:Rats; D011942:Receptors, Adrenergic, alpha / Q000502:physiology*; D011943:Receptors, Adrenergic, beta / Q000502:physiology*; D019210:Troponin I / Q000378:metabolism*; D000099211:Phospholamban; D000099163:Myosin Binding Protein C
D002118:Calcium; D002135:Calcium-Binding Proteins; D002352:Carrier Proteins; D007545:Isoproterenol; D011942:Receptors, Adrenergic, alpha; D011943:Receptors, Adrenergic, beta; D019210:Troponin I; D000099211:Phospholamban; D000099163:Myosin Binding Protein C