Traditional Chinese Veterinary Medicine (TCVM) including acupuncture has been used to treat disease and relieve pain in horses for more than 2,000 years. A review of the current knowledge of the mechanisms of acupuncture stimulation is presented. An emphasis is placed on electro-acupuncture, which can release neurotransmitters such as 5-hydroxytryptamine (serotonin) and endogenous opioids including β-endorphin, to relieve pain. Acupuncture has been shown to be a viable integrative treatment for equine back pain, foot pain, cervical stiffness, laryngeal hemiplegia and infertility. Acupoint sensitivity on palpation may be useful for the assessment of lameness along with conventional diagnostics in horses. Future well-designed studies are needed in order to strengthen the recommendation for acupuncture in the diagnosis and treatment of clinical conditions in horses.