Psychological stress increases the level of glucocorticoids in the circulating system. We found that dexamethasome administration in adult mice elevates the expression of COX-2 in the myocardium. With isolated neonatal cardiomyocytes, corticosterone (CT) at physiologically relevant doses (0.01 - 1 µM) induces the expression of COX-2 gene. The induction first appeared at 4 hrs and remained for at least 24 hrs with 1 µM CT treatment. This response is likely cardiomyocyte cell type specific since CT did not induce COX-2 expression in cardiac fibroblasts and glucocorticoids are known to suppress the expression of COX-2 in lymphocytes and several organs. Corticosteroids but not estrogen or progesterone induce COX-2 expression. The glucocorticoid receptor (GR) antagonist, mifepristone (MF), prevented CT from inducing COX-2 gene, suggesting a GR-dependent induction in cardiomyocytes. COX-2 gene promoter deletion and mutation studies indicate a role of CCAAT/enhancer binding protein (C/EBP) in CT induced COX-2 gene expression. Chromatin immunoprecipitation assays reveal that CT caused the binding of both GR and C/EBP to COX-2 promoter, while MF pretreatment blocked such binding. Co-immunoprecipitation experiments demonstrate CT treatment induces the interaction of GR with C/EBP. siRNA against C/EBP prevented CT from activating COX-2 promoter or elevating COX-2 protein. Our data suggest that the interaction between GR and C/EBP contributes to elevated COX-2 gene transcription by CT in cardiomyocytes.