In our study, MEAP showed significant anti-nociceptive activities in a dose-dependent manner in the acetic acid-induced writhing tests in mice. In addition, the acetic acid-induced writhing test is commonly used for evaluation of peripheral anti-nociceptive activity of drugs. Acetic acid can indirectly induce the release of endogenous mediators and stimulate the nociceptive neurons that are sensitive to non-steroidal anti-inflammatory drugs. The hot plate test is widely used for evaluating central antinociceptive activities. Results of the hot plate test in our present study indicated that MEAP is not a centrally acting analgesic. Therefore, we presumed that the MEAP exerts peripheral anti-nociceptive activities and performed the formalin test to confirm the hypothesis. The formalin test is a pain model consisting of two different LEE011 phases, which can be separated in time. The early phase is generated peripherally through the activation of nociceptive neurons by the direct action of formalin, and the late phase refers to the inflammatory pain response involving the release of molecules such as prostaglandin, histamine and serotonin. In our present study, the MEAP mainly suppressed pain of the late phase, which suggested that the anti-nociceptive mechanism of MEAP is related to the peripheral anti-nociceptive activity. This result was confirmed by testing MEAP in combination with naloxone. By using the openfield test, a behavior model used to study exploratory and motor activity, we observed no obvious changes of locomotor activity, which provided another piece of solid evidence to support the peripheral anti-nociceptive mechanism of MEAP activity. Cyclooxygenase is the key enzyme in the synthesis of prostaglandins, and COX-2, an inducible enzyme, is responsible for the production of the pro-inflammatory prostaglandins. Many former investigations have shown the important role of COX-2 in the induction of pain and inflammation as well as the analgesic actions of NSAIDs. In addition, COX-2 can induce synthesis of PGE, and the PGE2 released in inflamed tissues sensitizes the terminals of afferent nerve fibers, thereby enhancing nociceptive processing within the spinal cord and brain to evoke hyperalgesia. Thus, one apparently feasible approach against nociception is to suppress the release of COX-2. In our study, MEAP significantly and dose-dependently inhibited expression of COX-2 in the spinal dorsal horns of pain model mice induced by formalin, which indicated that MEAP may be a potent COX-2 inhibitor. Pituitary adenylate cyclase-activating polypeptide is a neuropeptide with multiple roles, including neurotransmitter, neuromodulator and neurotrophic factor. Our recent studies have suggested that PACAP is associated with psychiatric disorders, including schizophrenia.