Objective The manifestations of aging include cognitive deficits and muscular dysfunction, which are closely linked to impairment of mitochondrial biogenesis. Berberine, an isoquinoline alkaloid, presents multiple anti-diabetic pharmacological effects. Evidence has indicated that insulin resistance and cognitive impairment share the same pathogenesis, and berberine could reverse glucose metabolism abnormalities and muscle mitochondrial dysfunction induced by a high-fat diet. This study was used to investigate whether berberine could be used as an anti-aging drug to prevent cognitive deficits and muscular dysfunction in natural aging. Methods Biochemical indicators and an intraperitoneal glucose tolerance test were tested in 5-monthold rats (5 mo group), 24-month-old rats (24 mo group) and 24-month-old rats that had undergone 6 months of berberine treatment (BBR group). A Morris water maze test was conducted to assess the cognitive ability of the rats. Insulin resistance in whole-body was evaluated by intraperitoneal glucose tolerance test (IPGTT). The morphology of the skeletal muscle tissue was observed by hematoxylin-eosin (HE) staining. The levels of total cholesterol, triglyceride, ATP and reactive oxygen species (ROS) were assessed with corresponding reagent kits. The protein expressions of GLUT4, AMPK, SIRT1 and PGC-1α in skeletal muscle were examined by Western blot. Results The results showed that administration of berberine for 6 months significantly improved cognitive deficits and insulin resistance in naturally aging rats (p<0.01). Furthermore, berberine treatment helped normalize the disordered alignment and the decreased number of muscle fibers (p<0.01) in the skeletal muscle of 24 mo rats. Berberine decreased the levels of ROS in both the serum and the skeletal muscle of 24 mo rats (p<0.01). Berberine increased the protein expression of p-AMPK, SIRT1 and PGC-1α and increased the production of ATP in the skeletal muscle of aging rats (p<0.01). Conclusions Berberine markedly ameliorates aging-related reductions in cognitive ability and muscular function, and the activation of the AMPK/SIRT1/PGC-1α pathway in skeletal muscle may be the underlying protective mechanism of berberine on muscular function.