Aim To observe whether protein phosphorylation and dephosporylation in nuclei play roles in the development of myocardial hypertrophy and distribution of protein kinases and phosphatases in cell fractions were determined. Methods The model of hypertensive rat was established by abdominal aortic constriction. Velocity and isopyknic gradient centrifugation was employed to fractionate rat myocardium to membrane, cytosole and nuclei. Enzymatic methods were employed to determine kinases and phosphatases. Results Compared with control group, the activity of mitogen activated protein kinase (MAPK) increased by 82.03% (p<0.05) in nuclear, changed without significance in membranous fraction, whereas declined by 53.69% (p<0.01) in cytosolic fraction; the activity of protein kinase A (PKA) in different fractions were close to those of control; the activity of PPase 2A increased by 44.95% (p<0.05) and 36.75% (p<0.05) respectively in nuclear and membranous fractions, changed without significance in cytosolic fraction; the activity of PPase 2B increase by 43.57% (p<0.05) in nuclear and changed without significance in membranous and cytosolic fractions; the activity of PPase 2C in different fractions were close to those of control. Conclusions Nuclear MAPK, PPase 2A and PPase 2B might be involved in developing overload induced cardiac hypertrophy.