2.4 Neuropathic pain model
The spared nerve injury (SNI) model of neuropathic pain was used. This partial denervation model involves ligation and excision of two of the three distal branches of the sciatic nerve. Traditional SNI involves ligating the common peroneal and tibial nerves and sparing the sural nerve (Decosterd & Woolf, 2000). We used a variation in which the tibial nerve was spared (SNIt) and the common peroneal and sural nerves were ligated and cut. Baseline responses to mechanical stimuli were determined before surgery. Then mice were deeply anesthetized with isoflurane 1.5-4.0% (Abbott laboratories, North Chicago, IL), and toe pinch was used to verify that mice were under the surgical plane of anesthesia. Under aseptic conditions, the sciatic nerve was exposed at mid-thigh level through the biceps femoris muscle. The common peroneal and sural nerves were tightly ligated with 6.0 silk and axotomized distal to the ligation. Care was taken to avoid any contact with or stretching of the intact tibial nerve. Muscle and skin were closed in two layers with 4.0 silk and staples, respectively. In sham surgeries, the branches of the sciatic nerve were exposed but not ligated. The animals were returned to their cages after surgery and monitored during recovery. SNItproduces early (<24 hr) and prolonged (>12 weeks) mechanical hypersensitivity and neuropathic pain predominantly in the area innervated by the tibial nerve (Tallarida, 2006). Behavioral experiments were initiated 2 days after surgery.