Type 2 diabetes mellitus is characterized by islet β-cell dysfunction and its incidence increases with age. However, the mechanisms underlying the effect of aging on islet β-cell function are not fully understood. We characterized β-cell function in 4-month-old (young), 14-month-old (adult), and 24-month-old (old) male Wistar rats, and found that islet β-cell function decreased gradually with age. Old rats displayed oral glucose intolerance and exhibited a decrease in glucose-stimulated insulin release (GSIR) and palmitic acid-stimulated insulin release (PSIR). Furthermore, total superoxide dismutase (T-SOD), CuZn superoxide dismutase (CuZn-SOD), and glutathione peroxidase (GSH-Px) activity decreased, whereas serum malondialdehyde (MDA) levels increased in the older rats. Moreover, we detected a significant reduction in β-cell proliferation and an increase in the frequency of apoptotic β-cells in the islets of rats in the old group. Finally, Anxa1 expression in the islets of old rats was significantly upregulated. These data provide new insights into the development of age-related β-cell dysfunction in rats.