Objective: To investigate the protective effects of soybean isoflavone on hippocampal cholinergic neurons of chronic cerebral ischemia rats. Methods: 30 healthy, adult female SD rats were applied in this study and randomly divided into sham operation group (n=10), model group (n=10) and treatment group (n=10). The rats in model group and treatment group were established chronic cerebral ischemia model by permanently ligating the common carotid arteries on both sides of castrated rats. In sham operation group, the bilateral ovaries and common carotid arteries were only separated without ligature. After the chronic cerebral ischemia rats' model was established, the rats in treatment group were lavaged with soybean isoflavone (9mg/kg/d) for 21 days. SP immunohistochemical staining and MiVnt image analysis system were used to detect and analyze the choline acetyltransferase (ChAT) protein expression in CA1 region of hippocampus of rats in each group. Results: The ChAT protein expression of neurons in CA1 region of hippocampus of rats in sham operation group was obvioulsy higher than model group and treatment group (P˂0.05). The ChAT protein expression of neurons in CA1 region of hippocampus of rats in treatment group was obvioulsy higher than model group (P˂0.05). Conclusions: Soybean isoflavone has protective effects on cholinergic neurons in hippocampus of rats with chronic cerebral ischemia, the mechanisms may be related to soybean isoflavone can up regulate ChAT protein expression in hippocampus.