The present invention pertains to fields of biochemistry and molecular biology, relates to an ±O-superfamily conotoxin peptide, pharmaceutical composition thereof, preparation method and use thereof. The present invention further relates to a propeptide of the conotoxin peptide, nucleic acid construct thereof, expression vector and transformed cell thereof, and fusion protein thereof. The present invention further relates to a method for blocking acetylcholine receptors as well as a use of the conotoxin peptide in the manufacture of a medicament. The new ±O-superfamily conotoxin peptide of the present invention is capable of specifically blocking acetylcholine receptor (nAChRs) (e.g., ±9±10 nAChR), and NMDA receptor (e.g., NR2C NMDAR), and has activity for treatment of neuralgia, addiction, and activity for treatment of chemotherapy of cancers, breast cancer, lung cancer, wound healing, epilepsia, ischemia, and thus is promising in the manufacture of analgesic, a medicament for treatment of addiction, and a tool drug for neuroscience.