The long term goal of this proposed research has been and remains to determine how insulin and diabetes affect regulation of glycogen metabolism of cardiac muscle. The regulation of cardiac glycogen metabolism will be studied at the molecular level using isolated cardiomyocytes and primary cultures of adult cardiomyocytes. Studies will utilize hearts from normal, alloxan diabetic and BB/W spontaneously diabetic rats. The hypotheses to be tested are that 1) Cardiac glycogen synthase is acutely regulated by insulin and epinephrine through changes in the phosphorylation state of specific serine residues. 2) The expression of glycogen synthase and phosphorylase is chronically controlled by hormones. 3) Phosphoprotein phosphatase-1 (PP-1) is the physiologically relevant glycogen synthase phosphatase in heart. 4) The form of PP-1 active against glycogen synthase is chronically controlled by insulin or a hormonal axis modulated by the diabetic state. and that 5) Utilization of the diabetic state as a manipulative tool and restoration therapy in a chemically defined serum-free medium will facilitate a better understanding of the mechanism of hormone action in cardiac tissue.
Specific aims are to 1) Identify amino acid sequences of specific rat heart synthase peptides associated with insulin and epinephrine regulation through phosphorylation/dephosphorylation. 2) Use the antibodies which we have developed in concert with isolated cardiomyocytes and primary cultures of cardiomyocytes to study the hormonal control of turnover of synthase and phosphorylase. 3) Using antibodies to PP-1, determine if PP-1 is a likely candidate for the physiologically active form of synthase phosphatase. 4) Restore the diabetes-related decrease in synthase phosphatase activity using hormonal intervention in the primary culture system. and 5) Study the mechanism of the chronic regulation (turnover) of synthase phosphatase by hormones. All of these specific aims focus around gaining a better understanding of the mechanisms of chronic and acute regulation of rat heart glycogen metabolism as affected by diabetes and hormonal intervention. Results of these studies should increase our knowledge in this area in heart, and hopefully, eventually lead to an improved prognosis for our large diabetic population.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37HL020476-19
Application #
2215351
Study Section
Special Emphasis Panel (NSS)
Project Start
1976-09-01
Project End
1997-03-31
Budget Start
1995-04-01
Budget End
1996-03-31
Support Year
19
Fiscal Year
1995
Total Cost
Indirect Cost
Name
University of Massachusetts Medical School Worcester
Department
Biochemistry
Type
Schools of Medicine
DUNS #
660735098
City
Worcester
State
MA
Country
United States
Zip Code
01655
De Luca, J P; Garnache, A K; Rulfs, J et al. (1999) Wortmannin inhibits insulin-stimulated activation of protein phosphatase 1 in rat cardiomyocytes. Am J Physiol 276:H1520-6
Buczek-Thomas, J A; Miller Jr, T B (1995) Identification of the molecular basis for phosphorylase hypersensitivity in cultured diabetic cardiomyocytes. Mol Cell Biochem 145:131-9
Buczek-Thomas, J A; Miller Jr, T B (1995) Cyclic GMP accumulation in normal and diabetic primary culture adult rat ventricular cardiomyocytes: a minor role for nitric oxide in phosphorylase activation. Cell Signal 7:591-8
Miller, C; Rulfs, J; Jaspers, S R et al. (1994) Transformation of adult ventricular myocytes with the temperature sensitive A58 (tsA58) mutant of the SV40 large T antigen. Mol Cell Biochem 136:29-34
Jaspers, S R; Garnache, A K; Miller Jr, T B (1993) Factors affecting the activation of glycogen synthase in primary culture cardiomyocytes. J Mol Cell Cardiol 25:1171-8
Buczek-Thomas, J A; Jaspers, S R; Miller Jr, T B (1992) Post-receptor defect accounts for phosphorylase hypersensitivity in cultured diabetic cardiomyocytes. Mol Cell Biochem 117:63-70
Buczek-Thomas, J A; Jaspers, S R; Miller Jr, T B (1992) Adrenergic activation of glycogen phosphorylase in primary culture diabetic cardiomyocytes. Am J Physiol 262:H649-53
Jaspers, S R; Miller Jr, T B (1991) Purification and the immunological characterization of rat protein phosphatase 2A: enzyme levels in diabetic liver and heart. Mol Cell Biochem 101:167-74
Klarlund, J K; Jaspers, S R; Khalaf, N et al. (1991) An insulin-stimulated kemptide kinase purified from rat liver is deactivated by phosphatase 2A. J Biol Chem 266:4052-5
Jaspers, S R; Rulfs, J; Johnson, G L et al. (1989) Amino-terminal sequence analysis of rat heart and muscle glycogen synthase: homology to the rabbit enzyme and the implications for hormonal control. Arch Biochem Biophys 268:630-6

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