While the factors which precipitate cardiac arrhythmias are largely unknown, there is no doubt that acute autonomic stimulation is arrhythmogenic. Voltage-gated sodium channels are a critical determinant of normal and abnormal conduction in the heart, and mutant channels with subtle dysfunction can cause life-threatening arrhythmias. At present, the effects of adrenergic stimuli, which activate protein kinase A and C, on the cardiac sodium current remain controversial. The goal of this proposal is to test the hypothesis that protein kinase activatiOn modulates function of human cardiac sodium channels by phosphorylation of the channel alpha-subunit. Electrophysiologic studies will characterize the functional effects of protein kinase A and C stimulation on the major voltage-gated sodium channel in human heart, hHI, with the channel expressed in two different heterologous systems (Xenopus Laevis oocytes and a mammalian cell line). Substantial preliminary data demonstrate that activation of both kinases causes significant effects on hH1 current. Additional studies will be undertaken to define the biochemical basis for kinase effects on hHI. Immunoprecipitation techniques will be used to determine if the channel is directly phosphorylated by kinase under in Vitro and in vivo conditions. To elucidate the functional role of putative phosphorylation sites in the hHI sequence, experiments will be performed using both chimeric human heart-skeletal muscle channels and site-directed mutagenesis, to pinpoint regions of channel sequence and, ultimately, individual amino acids of functional and biochemical importance. Finally, the protein kinase C isoforms present in both the cellular expression systems used and human myocardium will be identified. The effects of human isoforms which are lacking in the cellular systems on hHI will then be tested to understand more fully the relevance of hHI modulation for human heart. The knowledge gained from these studies will improve our understanding of the nature and molecular basis of sodium channel modulation in human heart, and conditions which could conceivably promote or suppress arrhythmias due to changes in channel function.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL055665-02
Application #
2685470
Study Section
Special Emphasis Panel (ZRG4-CVB (03))
Project Start
1997-04-01
Project End
2001-03-31
Budget Start
1998-04-01
Budget End
1999-03-31
Support Year
2
Fiscal Year
1998
Total Cost
Indirect Cost
Name
Vanderbilt University Medical Center
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
004413456
City
Nashville
State
TN
Country
United States
Zip Code
37212
Hallaq, Haifa; Wang, Dao W; Kunic, Jennifer D et al. (2012) Activation of protein kinase C alters the intracellular distribution and mobility of cardiac Na+ channels. Am J Physiol Heart Circ Physiol 302:H782-9
Yang, Zhenjiang; Murray, Katherine T (2011) Ionic mechanisms of pacemaker activity in spontaneously contracting atrial HL-1 cells. J Cardiovasc Pharmacol 57:28-36
Wang, Limin; Gill, Rajan; Pedersen, Theresa L et al. (2009) Triglyceride-rich lipoprotein lipolysis releases neutral and oxidized FFAs that induce endothelial cell inflammation. J Lipid Res 50:204-13
Wang, Limin; Sapuri-Butti, Annapoorna R; Aung, Hnin Hnin et al. (2008) Triglyceride-rich lipoprotein lipolysis increases aggregation of endothelial cell membrane microdomains and produces reactive oxygen species. Am J Physiol Heart Circ Physiol 295:H237-44
Yang, Zhenjiang; Browning, Carrie F; Hallaq, Haifa et al. (2008) Four and a half LIM protein 1: a partner for KCNA5 in human atrium. Cardiovasc Res 78:449-57
Hallaq, Haifa; Yang, Zhenjiang; Viswanathan, Prakash C et al. (2006) Quantitation of protein kinase A-mediated trafficking of cardiac sodium channels in living cells. Cardiovasc Res 72:250-61
Zhou, Jingsong; Shin, Hyeon-Gyu; Yi, Jianxun et al. (2002) Phosphorylation and putative ER retention signals are required for protein kinase A-mediated potentiation of cardiac sodium current. Circ Res 91:540-6
Shin, H G; Murray, K T (2001) Conventional protein kinase C isoforms and cross-activation of protein kinase A regulate cardiac Na+ current. FEBS Lett 495:154-8
Shin, H G; Barnett, J V; Chang, P et al. (2000) Molecular heterogeneity of protein kinase C expression in human ventricle. Cardiovasc Res 48:285-99
Zhou, J; Yi, J; Hu, N et al. (2000) Activation of protein kinase A modulates trafficking of the human cardiac sodium channel in Xenopus oocytes. Circ Res 87:33-8