The dynamic changes of ACh activity in the spinal dorsal horn of rats were recorded by ACh ion selective microelectrodes. The influence of applying capsaicin to the spinal surface, administering anti-SP surum to the interior of dorsal horn, cutting off dorsal roots and electroacupuncturing at "points" on the ACh activity were studied. The results showed that electroacupuncture at "points" and capsaicin could facilitate ACh release of the spinal dorsal horn, it was fully inhibited by cutting off dorsal roots or obviously inhibited by administering anti-SP surum. The above results indicated that ACh might participate in the primary afferent of painsensation and acupuncture sensation as a neurotransmitter and SP might adjust the release of ACh in spinal dorsal horn.