Botulinum toxin (BT-A) is the most potent known toxin and at the same time a pharmacologically useful protein that has a wide clinical application. Animal studies and clinical studies have shown long-term antinociceptive activity of BT-A in some chronic pain states, but the mechanism of action has not yet been clarified. For a long time, the analgesic effect was considered to be at the peripheral level by inhibition of the exocytosis of the neurotransmitters involved in nociception. However, a growing number of evidence suggests that after peripheral administration BT-A enters the neuron and through axonal transport comes to the central nerve endings in the dorsal horn of the spinal cord, where he inhibits the exocytosis of the neurotransmitters involved in transferring the painful information to the next neuron in the pain pathway, but it is still necessary to investigate whether this happens segmentally in spinal cord or the antinociceptive mechanism also takes place on supraspinal level. The purpose of this study is to investigate the antinociceptive effect of BT-A after its application in brain chambers or in cisterna magna on two pain models. One of them is formalin-induced pain model that takes place on area innervated by ischiadic nerve, while the other is orofacial formalin-induced pain model conducted on area inervated by trigeminal nerve. This study was performed on male Wistar rats that were 3-4 months old, at Department of Pharmacology of the Faculty of Medicine (as part of current approved projects). In the formalin test on the rat hind-paw BT-A (1 i.j./kg) was administered i.c.v. and i.c. In orofacial test BT-A was applied i.c. The supraspinally applied BT-A neither reduced pain in the first (acute) phase, nor in the second (inflammatory) phase of the formalin test performed on the rats hind-paw, while BT-A applied i.c. reduced pain in the second phase of the orofacial formalin test. The obtained results show that BT-A, when administered supraspinally, does not have analgesic effect on peripheral pain although it travels to the nociceptive regions of the brain which is proved by presence of cleaved substrate, SNAP-25. Taking into account the results of previous researches the results of this study are additional evidence that the antinociceptive activity of BT-A takes place segmentally at the spinal cord level.