Objectives: To further explore the role of PKA signaling pathway in UII induced cardiomyocyte hypertrophy. Methods: Isolate and culture the neonatal rat myocardial cells. The primary myocardial cells were randomly divided into 4 groups: (1) the normal control group; (2) the UII 10–8 mol/L group; (3) KT-5720 + UII 10–8 mol/L group; (4) KT-5720 group. Cell planimetric area was viewed using an inverted fluorescent microscope. BCA method was used to detect total intracellular protein in myocardial cells and Hoechst 33258 staining to detect total intracellular DNA. Then the protein/DNA ratio was calculated. Fura-3AM fluorescence staining was used to detect myocardial cell cytoplasm Ca2 + concentration. Intracellular p-PKA, PKA, p-PLN, PLN, SERCA2a protein content was measured by Western blot assay. Results: (1) UII induced myocardial hypertrophy in the concentration of 10–8 mol/L. Pretreatment with KT-5720 could inhibit the hypertrophy of large extent. (2) UII could lead to sharp increase in intracellular Ca2 + concentration; KT-5720 pretreatment also attenuated this increase degree. (3) KT-5720 pretreatment significantly increased the expression of p-PKA, p-PLN and SERCA2a protein which were induced decreasing by UII. Conclusion: PKA signaling pathway plays an important role in UII induced hypertrophy of myocardial cells.