Abstract Background This study aimed to investigate dynamic urinary proteome changes of electroacupuncture (EP) on cerebral ischemia‑reperfusion (CI/R) injured rats and to explore the therapeutic biological mechanisms of EP. Methods First, changed urinary proteins were found in EP stimulation in healthy rats. Then, we used a CI/R injury rat model induced by Pulsinelli's four‐vessel occlusion (4‐VO) method to explore the function of EP on urinary proteome in CI/R injury. Urine samples were collected for proteome analysis by liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) and bioinformatics analysis. Results In total, 384 proteins were identified, among which 47 proteins (23 upregulated, 24 downregulated) were differentially expressed with 0.6‐log FC and p