Fig. 1

Chronic NTG injection evoked sustained CM-like phenotypes. A Schematic illustration of chronic NTG injection protocol. Mice of both sexes were i.p. injected with either vehicle (Ctrl) or NTG (0.1 mg/kg). B On day 1, the basal mechanical threshold was the same between the NTG and control group, while the formal developed mechanical hyperalgesia after 2 hrs post-treatment (left, n = 18 per group; p < 0.01 **, and p < 0.001 ***). Sustained basal mechanical hyperalgesia developed after repeated doses of NTG administration (right, n = 18 per group; p < 0.001 ***). C The basal mechanical threshold recovered 6 days after the final NTG injection (n = 4 per group; F(1,24) = 16.2; p < 0.05 *, and p < 0.01 **). D Schematic illustration of periorbital test protocol. The periorbital responses were scored 2 hrs after the i.p. injection of the NTG (left). Sustained mechanical hyperalgesia developed after repeated doses of NTG administration in both sexes of mice (right, n = 9 per group; F(1,112) = 117.8; p < 0.05 *, p < 0.01 **, and p < 0.001 ***). E Sustained mechanical hyperalgesia developed after repeated doses of NTG administration in male mice (n = 5 per group; F(1,4) = 71.6; p < 0.05 *, and p < 0.001 ***). F Sustained mechanical hyperalgesia developed after repeated doses of NTG administration in female mice (n = 4 per group; p < 0.05 *, and p < 0.01 **). G Schematic illustration of acute sumatriptan treatment protocol. Mice of both sexes were i.p. injected with sumatriptan (0.6 mg/kg) 5 min after NTG injection on day 9 (left). Acute sumatriptan injection transiently alleviated chronic NTG induced mechanical hyperalgesia on day 9. The basal response in the NTG group returned to mechanical hyperalgesia on day 10, suggesting that acute sumatriptan treatment only aborted acute pain but did not reverse the chronicity of mechanical hyperalgesia. (right, n = 20 per group; p < 0.001 ***). H Acute sumatriptan injection transiently alleviated chronic NTG induced mechanical hyperalgesia in male mice (n = 10 per group; p < 0.05 *, and p < 0.01 **). I Acute sumatriptan injection transiently alleviated chronic NTG induced mechanical hyperalgesia in female mice (n = 10 per group, p < 0.001 ***). J Schematic illustration of chronic NTG injection protocol. Mice were sacrificed after the NTG post-treatment von Frey test on day 9. K Representative images show that co-labeled CGRP and pERK neurons in the TG after chronic NTG administration were significantly higher than that in the controls (right, n = 4 per group; p = 0.043). Scale bar: 200 μm. l Quantification of the coloalized percentage of CGRP and pERK-positive neurons in the TG after chronic NTG induction (n = 4 per group; p = 0.003). All data shown are mean ± SEM and analyzed by Friedman tests with Dunn’s post hoc test (B, F, G-I) or Bonferroni post hoc test (C-E) or Mann-Whitney U test (K, L). Significance levels set at p < 0.05 *, p < 0.01 **, and p < 0.001 ***