The role and mechanism of calcineurin injecting into anterior cingulate cortex in alleviating pain in rats following experimental tooth movement

Abstract

Objective To explore the analgesic effect and mechanism of injection of calcineurin in anterior cingulate cortex (ACC) on experimental tooth movement in rats. Methods According to random number table, 60 Sprague-Dawley rats (200-250 g) were randomly divided into four groups: sham group (10) , force group (10) , 0.9% sodium chloride solution group (20) and calcineurin (CaN) solution group (20) . The four groups were treated in different modes: tooth movement modeling but with no force, modeling and applying force, applying force and injecting saline, applying force and injecting CaN. Mouth wiping behaviors and rat grimace scale (RGS) were recorded at the time before brain cannulation surgery in ACC, before modeling, 2 d after modeling and 30 min, 2 h, 4 h, 24 h, 3 d, 5 d and 7 d after injection to assess pain perception in rats. The phosphorylation of α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor subunit GluA1 and the expression of phosphorylated protein kinase A (pPKA) and phosphorylated protein kinase Cγ (pPKCγ) were detected with Western blot. Results At 2 d after tooth movement, the average mouth wiping time of the sham-operated group and the force group were (43 ± 9) s and (158 ± 21) s, respectively. Besides, the RGS values of these two groups were 19 ± 3 and 53 ± 6, respectively. Compared with the sham-operated group, pain behaviors (tMouthwiping= 15.876, PMouth wiping<0.001; tRGS= 16.250, PRGS<0.001) and the expressions of pGluA1-831 and pPKCγ in ACC were both increased in the force group (t831= 3.060, P831= 0.013; tPKCγ= 2.831, PPKCγ= 0.011) . At 2 h, 4 h and 24 h after injection into the ACC area, mean mouth wiping time of CaN solution group was (64 ± 26) , (67 ± 18) , and (93 ± 21) s, respectively, which was significantly less than that in the 0.9% sodium chloride solution group, with average mouth wiping time of (147 ± 31) , (136 ± 29) , (119 ± 30) s, respectively. Meanwhile, at 30 min, 2 h, 4 h, 24 h and 3 d after administration, the RGS values of the CaN solution group were also dramatically lower than those in the 0.9% sodium chloride solution group. In addition, the protein levels of pGluA1-831 and pPKCγ in ACC significantly decreased at 2 h after the injection of calcineurin (t831= 7.916, P831<0.001; tPKCγ= 3.230, PPKCγ= 0.005) . Conclusions Pain in rats following orthodontic tooth movement is associated with phosphorylation of AMPA receptor subunit GluA1 at Ser831, which is probably mediated by protein kinase C gamma. Calcineurin alleviates pain of orthodontic tooth movement by dephosphorylation of GluA1. Key words: Calcineurin; Receptors, α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid; Toothache; Gyrus cinguli; Tooth movement