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.
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