title: Myocyte contractile activity modulates norepinephrine cytotoxicity and survival effects of neuregulin-1beta.
abstract: The purpose of this study is to test the hypothesis that mechanical and electrical activity in adult rat ventricular myocytes (ARVM) alters responses to proapoptotic and prosurvival ligands. The effects of electrical stimulation on myocyte survival, stress signaling, response to beta-adrenergic receptor (beta-AR)-stimulated apoptosis, and neuregulin-1beta (NRG) were examined. Electrical stimulation (6.6 V/cm; 0, 2, and 5 Hz; 2-ms duration; alternating polarity) of ARVM resulted in more than 70% capture. Although ARVM paced for 48 h showed higher mitochondrial uptake of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (P < 0.05, 0 vs. 2 and 5 Hz), electrical stimulation had little effect on cell survival assessed by trypan blue uptake, CPK release, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining. Electrical stimulation for 24 h did not induce stress response (heat shock protein 70, 90) nor stress kinase (Erk, JNK, p38) activation. NRG stimulation of Erk and Akt was similar between paced and quiescent cells. Pacing sensitized myocytes to beta-AR-stimulated JNK phosphorylation and cell death with 0.1 microM norepinephrine (NE) in paced myocytes causing equivalent cytotoxicity to 10 microM NE in quiescent cells. NRG suppressed beta-AR-induced apoptosis through a phosphatidylinositol-3-kinase-dependent pathway in both paced and quiescent cells, although it is overwhelmed by high-NE concentration in paced cells. Thus myocyte contractility modulates both NE cytotoxicity as well as the cytoprotective effect of NRG. These results demonstrate the feasibility and importance of using electrically paced cardiomyocytes in primary culture when examining the signaling pathways of cell survival.
authors.affiliation: Whitaker Cardiovascular Institute and Cardiovascular Divisions, Department of Medicine, Boston University Medical Center, Boston, MA 02118, USA.
authors.forename: Yukio
authors.initials: Y
authors.lastname: Kuramochi
authors.forename: Chee Chew
authors.initials: CC
authors.lastname: Lim
authors.forename: Xinxin
authors.initials: X
authors.lastname: Guo
authors.forename: Wilson S
authors.initials: WS
authors.lastname: Colucci
authors.forename: Ronglih
authors.initials: R
authors.lastname: Liao
authors.forename: Douglas B
authors.initials: DB
authors.lastname: Sawyer
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D004558:Electric Stimulation; D022062:Electrophysiologic Techniques, Cardiac; D004789:Enzyme Activation / Q000502:physiology; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D006352:Heart Ventricles; D048031:JNK Mitogen-Activated Protein Kinases; D008929:Mitochondria, Heart / Q000502:physiology; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009200:Myocardial Contraction / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000502:physiology*; D020890:Neuregulin-1 / Q000494:pharmacology*; D009638:Norepinephrine / Q000494:pharmacology*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D020033:Protein Isoforms / Q000494:pharmacology; D051381:Rats; D013997:Time Factors
chemical_list: D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D020890:Neuregulin-1; D020033:Protein Isoforms; D019869:Phosphatidylinositol 3-Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D009638:Norepinephrine