Aim To explore the effect of Calpain 1 on astragaloside Ⅳ （ASⅣ） inhibiting myocardial apoptosis induced by isoproterenol （ISO）. Methods 48 SD rats were randomly divided into 6 groups, 8 rats in each group: control group, ISO group, ISO＋propranolol 40 mg/（kg·d） group, ISO＋ASⅣ 20 mg/（kg·d） group, ISO＋ASⅣ 40 mg/（kg·d） group and ISO＋ASⅣ 80 mg/（kg·d） group. Administered groups received continually intragastric administration for 2 weeks, and ISO was intraperitoneal injected for 2 weeks in the day after that. 2 weeks later, TUNEL and electron microscopy experiments were performed to detect the myocardial apoptosis and mitochondrial changes respectively. Western blot was used to semi-quantify the expression of PARP1, mitochondrial Calpain 1 and apoptosis inducing factor （AIF）. Results Compared to control group, ISO group had a higher apoptosis percentage and mitochondrial swelling, cristae breakage were observed. In addition, the expression of poly ADP ribose polymerase 1 （PARP1）, mitochondrial Calpain 1 increased while mitochondrial AIF decreased. Compared to ISO group, ASⅣ group with different dose could decrease the apoptosis percentage and mitochondrial damage. Western blot results revealed that ASⅣ could decrease PARP1, mitochondrial Calpain 1 expression and increase mitochondrial AIF. Conclusions ASⅣ can protect myocardial apoptosis induced by ISO. The possible mechanism can be related to the inhibition of mitochondrial Calpain 1 expression, then reduce the release of mitochondria AIF into the cytosol and translocation to the nuclei.