Figure 7. KMO is upregulated in the spinal cord after SNI and mediates neuropathic pain.

(A) Mechanical nociceptive threshold was evaluated before and up to 24 hours after intrathecal injection of equimolar doses (8 nmol) of kynurenine (Kyn), 3-hydroxykynurenine (3-Hk), 3-hydroxyanthranilic acid (3-Haa), or vehicle (saline) in WT mice (n = 5–6). This panel shows representative curves obtained in the dose-response analysis of mechanical allodynia caused by equimolar doses of Kyn, 3-Hk, 3-Haa in WT mice (see Supplemental Figure 8). (B) Time course of 3-Hk levels in the ipsilateral dorsal horn of the spinal cord of mice after sham (14 days) and SNI surgeries (n = 4–5 per time point). (C) Time course of Kmo mRNA expression in the ipsilateral dorsal horn of the spinal cord spinal cord after sham (14 days) and SNI surgeries. (n = 5–8). (D) Western blotting analysis of KMO expression in the ipsilateral dorsal horn of the spinal cord after sham (14 days) or SNI surgery (10 and 14 days) (n = 4–5). (E) Representative image of KMO expression analyzed by immunofluorescence in the dorsal horn of the spinal cord after sham or SNI induction (14 days). Scale bar: 50 μm. (F) Mechanical nociceptive threshold was determined before and 14 days after SNI. Mice were then treated intrathecally (i.t.) with Ro 61-8048 (KMO inhibitor; 3–300 nmol) or vehicle and mechanical allodynia was measured up to 7 hours after treatments (n = 5). (G) Mechanical nociceptive threshold was determined before and 10 days after SNI followed by intrathecal treatment with shRNA against KMO, shRNA scramble, or vehicle (indicated arrows) and mechanical allodynia was measured up to 15 days after SNI (n = 6). Data are expressed as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001 versus sham or saline injected; ^##P < 0.01, ^###P < 0.001 versus mice treated with Ro 61-8048 or scramble shRNA by 2-way ANOVA with Bonferroni’s post hoc test (A, F, and G) or 1-way ANOVA with Bonferroni’s post hoc test (B–D).