Figure Legends Figure 1. AITC and capsaicin-evoked nocifensive behaviour and mechanical allodynia(A) Acute nocifensive behaviour and HPMA after intraplantar (i.pl., 20 µl), AITC (200 nmol) or vehicle (veh) in rats pre-treated (0.5 h) with intraperitoneal (i.p.) or local (i.pl.) A967079 (A96, 100 mg kg-1 and 300 nmol, respectively) or veh. (B) Acute nocifensive behaviour and HPMA after capsaicin (CPS, 10 nmol, i.pl.) or veh in rats pre-treated (0.5 h) with capsazepine (CPZ, 4 mg kg-1, i.p.) or veh. (C) Acute nocifensive behaviour and HPMA after CPS (10 nmol, i.pl.) or veh in rats pre-treated (0.5 h) with A96 (100 mg kg-1, i.p.) and A96 (300 nmol, i.pl.) or veh. (D) HPMA after CPS (10 nmol, i.pl.), AITC (200 nmol, i.pl.) or veh in rats post-treated (0.5 h) with CPZ (4 mg kg-1, i.p.), A96 (100 mg kg-1, i.p.) or veh. Mean±SEM, n=6 rats per group. Dash (-) is a combination of vehicles. Arrows indicates time of administration. ***P<0.001 vs. Veh;###P<0.001 vs. AITC/Veh A96, CPS/Veh CPZ, CPS/Veh A96. 1-way and 2-way ANOVA, Bonferroni correction.Figure 2. SP and CGRP-evoked mechanical allodynia(A) Acute nocifensive behaviour and HPMA after intraplantar (i.pl., 20 µl), SP (3.5 nmol) or vehicle (veh) in rats pre-treated (0.5 h) with local (i.pl.) L733,060 (20 nmol) or veh. (B) Acute nocifensive behaviour and HPMA after CGRP (1.5 nmol, i.pl.) or veh in rats pre-treated (0.5 h) with olcegepant (1 nmol) or veh. Acute nocifensive behaviour and HPMA after AITC (C) (200 nmol, i.pl.), capsaicin (D) (CPS, 10 noml, i.pl.) or veh in rats pre-treated (0.5 h) with olcegepant (1 nmol) or veh. Acute nocifensive behaviour and HPMA after AITC (E) (200 nmol, i.pl.), capsaicin (F) (CPS, 10 noml, i.pl.) or veh in rats pre-treated (0.5 h) with L733,060 (20 nmol) or veh. HPMA after CGRP (1.5 nmol, i.pl.) or veh in rats pre-treated (G) (0.5 h) or post-treated (H) (0.5 h) with A967079 (A96, 300 nmol, i.pl.) or veh. HPMA after CGRP (1.5 nmol, i.pl.) or veh in rats pre-treated (0.5 h) with capsazepine (I) (CPZ, 4 mg kg-1, intraperitoneal, i.p.) or veh, or (K) in rats post-treated (0.5 h) with olcegepant (1 nmol) or veh. Mean±SEM, n=6 rats per group. Dash (-) is a combination of vehicles. Arrows indicates time of administration. ***P<0.001 vs. Veh;#P<0.05,###P<0.001 vs. SP/Veh L733,060, CGRP/Veh olcegepant, CPS/Veh olcegepant, CGRP/Veh A96. 1-way and 2-way ANOVA, Bonferroni correction.Figure 3. CGRP, AITC and capsaicin induce mechanical allodyniavia NO production, and ROS production. HPMA after intraplantar (i.pl., 20 µl) CGRP (1.5 nmol) or vehicle (veh) in rats pre-treated (0.5 h) or post-treated (0.5 h) with local (i.pl.) (A) SQ22536 (25 nmol), (B) L-NAME (1 µmol), (C) PBN (670 nmol) or veh. Acute nocifensive behaviour and HPMA after (D) capsaicin (CPS, 10 nmol, i.pl.), (E) AITC (200 nmol, i.pl.) or veh in rats pre-treated (0.5 h) or post-treated (0.5 h) with local (i.pl.) PBN (670 nmol) or veh. Acute nocifensive behaviour and HPMA after AITC (200 nmol, i.pl.) or veh in rats treated (once a day for 4 consecutive days) with (F) intratechal (i.th, 10 µl) or (G) perineural (p.n., 10 µl) TRPA1 antisense (AS) or mismatch (MM) oligonucleotide (ODN) (10 nmol). Acute nocifensive behaviour and HPMA after (H) CPS (10 nmol, i.pl.), (I) CGRP (1.5 nmol, i.pl.) or veh in rats treated (once a day for 4 consecutive days) TRPA1 AS/MM ODN (10 nmol). Mean±SEM, n=6 rats per group. Arrows indicates time of administration. ***P<0.001 vs. Veh;###P<0.001 vs. CGRP/Veh SQ22536, L-NAME, PBN, CPS-AITC/Veh PBN, AITC-CPS-CGRP/MM. 1-way and 2-way ANOVA, Bonferroni correction.Figure 4. TRPA1 expression and function in primary rat Schwann cells. (A) Representative images of TRPA1 mRNA expression in rat dorsal root ganglia (DRG) and sciatic nerve (Scale bar: 20 µm, inset 10 µm) (n=3 subjects). DapB, negative control. (B) Representative real-time PCR plot and cumulative data for S100 and Trpa1 mRNA in rat Schwann cells (n=3 independent experiments). (C and D) Typical traces and cumulative data of Ca2+ response (F340/F380) in primary rat Schwann cells stimulated with AITC (1 mM), CGRP (10 μM) or vehicle (veh) in presence of A967079 (A96, 50 μM), olcegepant (100 nM) or veh (n=3 independent experiments). Mean±SEM. Dash (-) is a combination of vehicles. ***P<0.001 vs. Veh; ###P<0.001 vs. AITC CGRP. 1-way ANOVA, Bonferroni correction.Figure 5. TRPA1 activation mediates HPMA in partial sciatic nerve ligation (pSNL), but not in chronic constriction injury (CCI) model. HPMA in rats 15 days after (A) CCI, (B) pSNL or sham procedure treated with intraperitoneal (i.p.) AMG0902 (AMG, 30 and 100 mg kg-1), A967079 (A96, 10, 30 and 100 mg kg-1) or vehicle (veh). HPMA in rats 15 days after (C) CCI, (D) pSNL or sham procedure treated with capsazepine (CPZ, 4 mg kg-1, i.p.) or veh. HPMA in rats 15 days after (E) CCI, (E) pSNL or sham procedure treated (once a day for 4 consecutive days starting from day 10 to day14 after surgery) with perineural (p.n., 10 µl) TRPA1 antisense (AS) or mismatch (MM) oligonucleotide (ODN) (10 nmol). HPMA in rats 15 days after (G) CCI, (H) pSNL or sham procedure treated with intraplantar (i.pl.) olcegepant (1 nmol) or veh. HPMA in rats 15 days after (I) CCI, (J) pSNL or sham procedure treated with PBN (670 nmol, i.pl.) or veh. Mean±SEM, n=6 rats per group. Arrows indicates time of administration. ***P<0.001 vs. Sham/Veh;###P<0.001 vs. pSNL-CCI/veh AMG/A96, pSNL/MM, pSNL-CCI/veh PBN. 2-way ANOVA, Bonferroni correction.