1. Mechanosensitive ion channels in articular nociceptors drive mechanical allodynia in osteoarthritis 
B.H. He, M. Christin, S. Mouchbahani-Constance, A. Davidova, R. Sharif-Naeini 
Osteoarthritis and Cartilage 
Volume 25, Issue 12, Pages (December 2017)
DOI: /j.joca
Copyright © 2017 Osteoarthritis Research Society International Terms and Conditions

2. Fig. 1
Correlation between joint pathology and pain intensity in the MIA model. A. Representative histological sections from saline- (Control), and 250 μg MIA-injected mice (1 month post injection). Upper panels show hematoxylin and eosin stain. Lower panels show toluidine blue stains for the articular cartilage and other glycosaminoglycan containing structures. Control knees have a normal structure of cartilage (C) and meniscus (M). In OA knees, there is a thickening of the meniscus and a hypertrophic zone developing in the subchondral bone (arrow). B. Mean (±SEM) histopathology scores of ipsilateral knees injected with different doses of MIA, the medial femoral condyle (MFC), the lateral femoral condyle (LFC), the medial tibial plateau (MTP) or the lateral tibial plateau (LTP). * = P < 0.05; Kruskal–Wallis One Way ANOVA on ranks. N = 5–10 per group. C. Mean (±SEM) pain scores assessed using the knee-bend test. There is a significant difference in pain score for all MIA scores, except 25 μg, compared to control by day 5 (Two-way ANOVA with Tukey, *, **, *** = P < 0.05; P < 0.01; P < 0.001, respectively). N = 5–7 for all groups. D. Relation between the extent of cartilage damage and primary mechanical allodynia, assessed at all four sites of observation. Spearman correlation (r) and significance (p) values are given on the panels.
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2017 Osteoarthritis Research Society International Terms and Conditions

3. Fig. 2
MSICs have a reduced activation threshold in OA nociceptors. A. Representative cell-attached recordings of OA nociceptors stimulated with increasing levels of negative pressure through the recording electrode. Inset: visual identification of an articular nociceptor (black arrow). Image also displays a red neuron (red arrow) which is an articular afferent but non nociceptor, and a nociceptor (red arrowhead) which does not innervate the knee. B. Mean (±SEM) activation threshold of the MSICs in articular nociceptors. Only OA nociceptors innervating the ipsilateral knee display significantly reduced activation threshold. * = P < 0.05 (Kruskal–Wallis One Way ANOVA on ranks with Dunn's Multiple Comparison test). N = 52–83 (see on panel). C. Mean (±SEM) mechanically-activated currents in articular nociceptors of saline- or MIA-injected mice (3 weeks post-injection). Mechanically evoked current is significantly higher than control in both MIA groups as early as −10 mm Hg (Two-way ANOVA with Bonferroni post-hoc test, compared to 0 μg group). N = 82–122 (see on panel). ** = P < 0.01, compared to 0 μg group. D. Mean (±SEM) frequency of active patches and (E) Mean (±SEM) number of channels per patch. No statistical difference, P > 0.05 (One Way ANOVA on Ranks).
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2017 Osteoarthritis Research Society International Terms and Conditions

4. Fig. 3
Single channel analysis suggests that the same MSIC is present on articular nociceptors of both in the naïve and OA mice. A. Representative traces of mechanically-evoked single channel openings at different voltages. B. Mean (±SEM) current amplitudes of MSICs from both naïve and OA articular nociceptors. Data were fit with linear regressions and show a lack of significant difference (P > 0.05, ANCOVA). N = 8–14 for all groups. C. Mean (±SEM) single channel conductances obtained from the data in B. D. Mean (±SEM) reversal potential of the mechanically-evoked currents, extrapolated from the data in panel B. (P > 0.05, Kruskal–Wallis One Way ANOVA on Ranks for C and D).
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2017 Osteoarthritis Research Society International Terms and Conditions

5. Fig. 4
MSICs in articular nociceptors are GsMTx4-sensitive and contribute to mechanical allodynia in OA mice. A. Representative traces of mechanically-evoked currents from an outside-out recording. A minimal positive pressure (P) is applied to open MSICs. B. Mean (±SEM) mechanically-evoked current (normalized to control (Ctrl)) is reversibly reduced by GsMTx4 (One-way ANOVA, with Dunn's multiple comparisons, n = 17, 12, 7 for Control, GsMTx4 and Wash respectively). *** = P < 0.001, compared to Ctrl group. C. Mean (±SEM) channel open time is significantly and reversibly reduced by GsMTx4 superfusion (Kruskal–Wallis with Dunn's multiple comparisons, n = 161, 85, 100 for Control, GsMTx4 and Wash respectively). D. Knee injection of 5% lidocaine (Lido) significantly reduces the pain score of OA mice injected with 250 μg MIA compared to pre-injection pain score (Paired t test, P = 0.0371; n = 8). Knee injection of 10 μM (Gs-10) and 50 μM GsMTx4 (Gs-50) significantly reduce the pain score (Paired t test, P =  and P =  respectively, n = 10 for both). The injection of 50 μM Gadolinium (GdCl3) also significantly reduces the pain score of OA mice (Paired t test, P = 0.0026, n = 7). The injection of 5 μM GsMTx4 (Gs-5), however, did not affect the pain score (Paired t test, P = 0.5237, n = 9). E. Representative spinal cord sections demonstrating the induction of Fos in neurons of the superficial dorsal horn after joint movement in OA mice. Joint movement does not induce Fos in naïve mice. The same manipulation in OA mice (250 μg MIA, 21 days) produces a significant Fos induction compared to the saline group (Mann Whitney, P = 0.0019). Knee injection of GsMTx4 (50 μM) 20 min prior to joint movement significantly reduces the number of Fos-positive neurons (Mann Whitney, P =  vs 250 μg MIA). F. Mean (±SEM) number of Fos-positive neurons per section after joint movement in naïve or OA (100 and 250 μg MIA) mice, pre-treated with saline or GsMTx4 (50 μM). No significant changes in Fos induction are observed in deeper layers of the dorsal horn. N = 8 mice for all groups except 100 μg MIA where N = 15.
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2017 Osteoarthritis Research Society International Terms and Conditions

6. Fig. S1
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2017 Osteoarthritis Research Society International Terms and Conditions

7. Fig. S2
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2017 Osteoarthritis Research Society International Terms and Conditions

8. Fig. S3
Osteoarthritis and Cartilage  , DOI: ( /j.joca )
Copyright © 2017 Osteoarthritis Research Society International Terms and Conditions