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As a major amino acid, glycine has multiple functions in metabolism, growth, immunity, cytoprotection, and survival. The aim of this study was to determine the effects of glycine on pathologic cardiac hypertrophy and the mechanism underlying it. Pre-treatment with glycine significantly attenuated murine cardiac hypertrophy induced by transverse aortic constriction or by administration of angiotensin II (Ang II). This action was associated with a suppressive extracellular signal-regulated kinase 1/2 phosphorylation in myocardium. The cardioprotective effect of glycine disappeared when endogenous glycine receptor α2 was knocked down by mRNA interference in rats. Co-culture experiments revealed that glycine could also antagonize Ang II stimulated release of transforming growth factor β and endothelin-1 by cardiomyocytes, which prevented an over-production of collagens in rat fibroblasts. These results, for the first time, demonstrate that glycine may be a novel cardioprotector against pressure overload induced cardiac hypertrophy. Thus, glycine would be useful in the prevention of cardiac hypertrophy and heart failure.