Figure 1.
Behavioural and articular histological assessment of MIA animals.
A – withdrawals in response to 1 g, 6 g or 8 g von frey hair or acetone applied to the plantar hindpaw 3, 7 or 14 days after intra-articular saline, 2 mg or 1 mg MIA injection (Kruskal Wallis test of: timecourse, P<0.05 +, P<0.01 ++, P<0.001 +++ vs preinjection baseline; inter-group comparison P<0.05 *, P<0.01 ** n = 12 animals/group). B – incapacitance testing of 2 mg or 1 mg MIA treated animals at 3, 7 and 14d after treatment (2-way ANOVA with Bonferroni's post test, n = 12 animals/group. P>0.05 at all time points). C – 2 mg MIA treated rat knee sagittal sections from day 14, stained with toluidine blue to visualise cartilage proteoglycan content. Fem = femoral condyl, Tib = tibial condyl. Ant = anterior aspect of knee, Post = posterior aspect. D – 1 mg MIA rat knee sections from day 14. E – saline injected contralateral control knee from day 14.
Figure 2.
Cell size distribution for BIIITubulin (green bars, left y axis) and ATF-3 (red bars, right y axis) expressing profiles in DRG L3, L4 and L5 in sham or 2 mg MIA animals 3, 7 and 14 days after injection.
n = 9 sham, 7 at day 3, 12 at day 7, 7 at day 14.
Figure 3.
Approximation of total DRG ATF-3 expression.
A, B, C - DRG profiles show nuclear expression of ATF-3 7 days after 2 mg (A) or 1 mg (B) MIA, but not 14 days after saline sham injection (C). D - Estimated fraction of DRG profiles expressing ATF-3 in 2 mg MIA-treated animals (One-way ANOVA with Dunnett's multiple comparison test, n = 9×14d sham, 7×3d, 12×7d MIA, 7×14d MIA, P<0.01 **, P<0.05 *). E - Comparison of ATF-3 expression in 2 mg vs 1 mg MIA injected groups at 7 day timepoint (1 way ANOVA with Bonferroni's multiple comparison test, n = 9×7d sham, 12×2 mg MIA, 12×1 mg MIA, P<0.01 ** P<0.001 ***).
Figure 4.
Quantification of intrapepidermal nerve fibre density in plantar hindpaw following MIA treatment.
A, B - sections of naïve (A) and MIA (B) plantar hindpaw skin containing PGP9.5 immunoreactive intraepidermal nerve fibres (arrows). C – quantification of IENFs at the 7 and 14 day timepoints (paired t-test of ipsi vs contralateral density, n = 8×7d, 4×14d, P<0.05). IENF density reduction is not seen in 1 mg MIA animals.
Figure 5.
Quantification of microglial activation in spinal cord following MIA treatment.
A, B - expression of the microglial marker iba1 in L4 spinal cord in 2 mg (A) and 1 mg (B) MIA groups. C, D – Quantification of microglia with effector morphology in dorsal (C) and ventral (D) horn (1-way ANOVA, n = 4×7d sham, 11×2 mg MIA, P<0.05 *). E – Comparison of dorsal horn microgliosis in 2 mg vs 1 mg MIA groups at the 7d timepoint (1-way ANOVA, n = 4×7d sham, 11×2 mg MIA, 8×1 mg MIA, P<0.05 *).
Figure 6.
Effect of pregabalin treatment (10 mg/kg s.c.) on evoked responses of deep dorsal horn WDR neurones.
A - Electrically evoked responses in the Aß, A∂ and C fibre range, as well as PD (post-discharge/repetitive firing) expressed as % of predose baseline in 2 mg MIA, 1 mg MIA or sham injected animals. B - Mechanically evoked responses expressed as % of predose baseline in 2 mg MIA, 1 mg MIA or sham injected animals. C - Thermally evoked responses expressed as % of predose baseline in 2 mg MIA, 1 mg MIA or sham injected animals. Note the significantly greater effectiveness of pregabalin in the 2 mg treated group (normalized post-drug responses compared using Kruskal Wallis test with Dunn's posttest. n = 14d sham×9, 1 mg MIA×8, 2 mg MIA×9. P<0.05 *, P<0.01, **). The sham and 2 mg data is included for comparison, but has previously been published in a different form [19].