Supplementary Figure 3: Behavioral tests in VT3^Lbx1-ablated mice versus control littermates

(a) The Rota-Rod assay. No significant difference in falling latencies was detected between VT3^Lbx1-ablated (“VT3^Lbx1 Abl”; purple circles) and control (blue circles) mice (n = 10 in each group; two-tailed Student's unpaired t test, P = 0.4126, t₍₁₈₎ = 0.8388). (b,c) Touch sensation. No significant difference was detected in response numbers per three trials of brushing (b, n = 11, Control; n = 11, VT3^Lbx1 Abl; two-tailed Student's unpaired t test, P = 0.2500, t₍₂₀₎ = 1.185). The quick hindpaw lifting and/or walking away in response to brushing was scored as 1 for this assay, but would be scored as 0 for the dynamic allodynia assay described in Fig. 2. No significant difference in latencies of removing sticky tape was detected (c, n = 9, Control; n = 10, VT3^Lbx1 Abl; two-tailed Student's unpaired t test, P = 0.8383, t₍₁₇₎ = 0.2072). (d) No significant difference was detected in the Hargreaves heat test (n = 11 in each group; two-tailed Student's unpaired t test, P = 0.6379, t₍₂₀₎ = 0.4779). (e) No significant difference was detected in hot plate assays (n = 10 in each group; two-tailed Student's unpaired t test for flinch and lick latency at each temperature: P = 0.3770, t₍₁₈₎ = 0.9059; P = 0.4705, t₍₁₈₎ = 0.7372); P = 0.5331, t₍₁₈₎ = 0.6355; P = 0.1897, t₍₁₈₎ = 1.363; P = 0.1201, t₍₁₈₎ = 1.632; P = 0.7778, t₍₁₈₎ = 0.2864. (f) Cold plate assays. No significant difference in latencies of forepaw flinching or hindpaw licking was detected (n = 10 in each group; two-tailed Student's unpaired t test, P = 0.636, t₍₁₈₎ = 0.4815; P = 0.4368, t₍₁₈₎ = 0.7953). (g) No significant difference in behavioral responses to acetone-evoked cooling was detected (n = 10 in each group; two-tailed Student's unpaired t test, P = 0.7621, t₍₁₈₎ = 0.3073). (h) Itch test by nape injection of compound 48/80 (100 μg). No significant difference in evoked scratching bouts was detected (n = 9, Control; n = 10, VT3^Lbx1 Abl; two-tailed Student's unpaired t test, P = 0.2335, t₍₁₇₎ = 1.235). (i) Itch test by nape injection of chloroquine (200 μg). No significant difference in evoked scratching bouts was detected (n = 7, Control; n = 8, VT3^Lbx1 Abl; two-tailed Student's unpaired t test, P = 0.3681, t₍₁₃₎ = 0.9324). (j) Cold allodynia assays. No significant difference in cold allodynia at day 7 after SNI was detected (n = 5, Control; n = 5, VT3^Lbx1 Abl; two-tailed Student's unpaired t test, P = 0.9329, t₍₈₎ = 0.8687). Student’s unpaired t test. Data are represented as mean ± SEM.