The isolated cardiac myocytes of rats were immediately infected by cosackievirus B3 (CVB3) to investigate the effects of such procedure on the cell cycle, apoptosis and intracellular ionized calcium (Ca 2+ i) of cardiac myocytes. Newborn Balb/c murine cardiac myocytes were cultivated, then infected by CVB3. Intracellular Ca 2+ i was measured by flow cytometer. The calcium in the medium for culturing cardiac myocytes was detected by using atom absorb spectrum test. It was found that CVB3 could markedly inhibit the differentiation and proliferation of the infected cardiac myocytes and induce the apoptosis. The intracellular Ca 2+ i level in the infected group was significantly higher than in the control group (P<0.01). The calcium concentration in the medium for culturing cardiac myocytes in the infected group was significantly lower than in the control group (P<0.05). It was suggested that the apoptosis and intracellular calcium overload of the CVB3-affected cardiac myocytes are likely to play an important role in the pathogenesis of viral myocarditis. The isolated cardiac myocytes of mice were immediately infected by cosackievirus B3 (CVB3) to investigate the effects of such procedures on the cell cycle, apoptosis and intracellular ionized calcium (Ca 2+ i) of cardiac myocytes. Newborn Balb / c murine cardiac myocytes were The calcium in the medium for culturing cardiac myocytes was detected by using atom absorbance spectrum test. It was found that CVB3 could markedly inhibit the differentiation and proliferation of the infected cardiac myocytes and induce the apoptosis. The intracellular Ca 2+ i level in the infected group was significantly higher than in the control group (P <0.01). The calcium concentration in the medium for culturing cardiac myocytes in the infected group was significantly lower than in the control group (P <0.05). It was suggested that the apoptosis and intracellular calcium overload of the CVB3-affected cardiac myo cytes are likely to play an important role in the pathogenesis of viral myocarditis.