Time-course changes in tissue levels of transcripts encoding ATF3 (A), OX-42 (B) or GFAP (C) in dorsal root ganglia and spinal cord at various times after spinal cord transection.

<p>Real-time RT-qPCR determinations were made in T6–T8 and T9–T11 dorsal root ganglia, T6–T8 and T9–T11 spinal cord segments and the cervical and lumbar enlargements at various times (in days, D, abscissa) after spinal cord transection at T8–T9 level. Data are expressed as the ratio of specific mRNA over GaPDH mRNA [R.Q.(A.U.)]. Each bar is the mean + S.E.M. of n independent determinations (D2, D4, D9, D15, D21: n = 6; D60: n = 12). Sham values at every postoperative time are pooled under “C” (control) on abscissa. * P<0.05, * P<0.01, *** P<0.001 compared to respective values in sham-operated rats (C).Two-way ANOVA followed by Bonferroni test.</p

Short- and long-term changes in levels of transcripts encoding P2×4 (A), P2×7 (B) and TLR4 (C) in dorsal root ganglia and spinal tissues in spinal cord-transected rats.

<p>Real-time RT-qPCR determinations were made in T6–T8 and T9–T11 dorsal root ganglia and T6–T8 and T9–T11 spinal segments at day 2 or 60 (abscissa) after spinal cord transection at T8–T9 level. Data are expressed as the ratio of specific mRNA over GaPDH mRNA [R.Q.(A.U.)]. Each bar is the mean + S.E.M. of n independent determinations (D2: n = 6; D60: n = 12). Sham values at every postoperative time are pooled under “C” (control) on abscissa. ** P<0.01, *** P<0.001 compared to respective levels in sham-operated rats (C). Two-way ANOVA followed by Bonferroni test.</p

Time-course changes in nocifensive reactions to von Frey filaments application in the « at-level » allodynic territory rostral to the lesion in spinal cord-transected rats.

<p>Pressure threshold values to trigger biting (of the filament), shaking or escape were determined using the “up-down” method with a graded series of von Frey filaments applied onto the allodynic at-level area on the back at various times (in days) after surgery (0 on abscissa). Each bar is the mean + S.E.M. of independent determinations in 8 rats. *** P<0.001 compared to control (intact) rats (C on abscissa). One-way ANOVA for repeated measures followed by Dunnett's test.</p

Short- and long-term changes in levels of transcripts encoding IL-6 (A), IL-1β (B), TNF-α (C) and IL-10 in dorsal root ganglia and spinal tissues in spinal cord-transected rats.

<p>Real-time RT-qPCR determinations were made in T6–T8 and T9–T11 dorsal root ganglia and T6–T8 and T9–T11 spinal segments at day (D) 2, 15 or 60 (abscissa) after spinal cord transection at T8–T9 level. Data are expressed as the ratio of specific mRNA over GaPDH mRNA [R.Q.(A.U.)]. Each bar is the mean + S.E.M. of n independent determinations (D2, D15: n = 6; D60: n = 12). Sham values at every postoperative time are pooled under “C” (control) on abscissa. * P<0.05, * P<0.01, *** P<0.001 compared to respective values in sham-operated rats (C). Two-way ANOVA followed by Bonferroni test.</p

Hyper-responsiveness to thermal stimulation in spinal cord-transected rats.

<p><b>A</b>– Latency (in sec) to hindpaw withdrawal was determined after paw immersion into a bath of hot (46°C) or cold (10°C) water, two weeks after the surgery. Each bar is the mean + S.E.M. of independent determinations in 9 SCT rats and 5 sham-operated rats. ** P<0.01, *** P<0.001 compared to respective values in sham-operated rats. Student's t test. <b>B</b> – Behavioral responses to the acetone drop test applied at the surgical scar two weeks after surgery. The number of shakes and the time (in sec) spent in escape attempts and licking of the back were measured for one minute after acetone drops application. Each bar is the mean + S.E.M. of independent determinations in 8 SCT rats and 7 sham-operated rats. * P<0.05, ** P<0.01 compared to respective values in sham-operated rats. Student's t test.</p

Increased expression of ATF3, OX-42 and GFAP mRNAs in the dorsal and ventral halves of spinal cord segments just above (T6–T8) and below (T9–T11) the surgery level in spinal cord-transected rats.

<p>Real time RT-qPCR determinations were made at day 17 after surgery. Data are expressed as the ratio of specific mRNA over GaPDH mRNA [R.Q.(A.U.)]. Each bar is the mean + S.E.M. of 10 independent determinations in both SCT (black bars) and sham-operated (empty bars) rats. *** P<0.001 compared to respective values in sham-operated rats. Two-way ANOVA followed by Bonferroni test.</p

Anti-allodynic effects of acute administration of morphine (A), tapentadol (B), ketamine (C) or baclofen (D) in spinal cord-transected rats.

<p>Acute administration of morphine (1, 3 or 10 mg/kg s.c.), tapentadol (10 or 20 mg/kg i.p.), ketamine (50 mg/kg i.p.), baclofen (10 mg/kg i.p.) or their respective vehicle was performed (0 on abscissa, arrow) in rats whose spinal cord had been transected at T8–T9 level one month before. Pressure threshold values to trigger nocifensive biting were determined using von Frey filaments applied within the at-level allodynic territory at various times after treatment. Each point is the mean + S.E.M. of independent determinations in n rats. C on abscissa: Control (naive) rats (prior to surgery). P<0.05, ** P<0.01, *** P<0.001 compared to respective values in vehicle-treated rats. One-way ANOVA for repeated measures followed by Dunnett's test.</p

Body territories with increased mechanical sensitivity in spinal cord-transected rats.

<p>Pressure threshold values to trigger nocifensive responses were determined using a graded series of von Frey filaments applied throughout the body. Comparison with sham-operated rats (C) showed that pressure threshold values differed in SCT rats only in a limited territory (6 cm²) bordering rostrally the spinal cord section (at T8–T9, horizontal bar with arrow heads) and in hindpaws (black areas tested), where reactions were obtained for pressure values significantly less than in controls. Time course (day 2 to day 60) changes in spinal cord transected rats showed that supersensitivity (allodynia) in the at-level area just rostral to the lesion was already detected at day 2 (D2) post-surgery, then extended and increased up to a plateau reached at D14 post-surgery. At hindpaw level, supersensitivity developed much later (from D21 post-surgery). Data were obtained in 8–14 rats at each time.</p

Time-course changes in the pressure threshold value to trigger hindpaw withdrawal in spinal cord-transected rats.

<p>Pressure threshold values were determined using a graded series of von Frey filaments applied onto hindpaw. Each point is the mean + S.E.M. of independent determinations in 6 rats. « Cut-off SCT » corresponded to the maximal pressure tested in spinal cord-transected rats; even at this high pressure level (100 g), no response of hindpaws was evoked for the first 9 days post-surgery. The « Threshold sham » corresponded to the minimal pressure (60 g) to which sham-operated animals start to respond by hindpaw withdrawal. ** P<0.01, *** P<0.001, significantly different from 100 g « cut-off SCT » value. One-way ANOVA for repeated measures followed by Dunnett's test.</p

Effect of sex and HD mutation on saccharin-preference test and total fluid intake.

<p>(A) We found a significant interaction between sex and genotype (F_(1,44) = 5.84, p<0.05) on saccharin preference (expressed as % of total fluid intake). Indeed compared to WT animals, only HD female mice exhibited reduced saccharin preference. (B) Interestingly looking at total fluid intake (expressed in mL), we revealed an overall effect of genotype (F_(1,44) = 12.3, p<0.01) but no significant effect of sex or interactions. Values represent means (± SEM) of n = 8–14 mice per group. WT vs. HD: (*) p<0.05, (**) p<0.01.</p