The Effect of Pregabalin on Microglia Differentiation in Rat with Neuropathic pain: A Preliminary Study

Abstract

This study investigated the effects of pregabalin on microglial differentiation in rats with neuropathic pain (NP) induced by sciatic nerve ligation and transection. After confirming NP, the rats were randomly allocated to either a pregabalin or control group. The pregabalin group received intraperitoneal injections of 10 mg/kg pregabalin, while the control group received an equivalent volume of normal saline following surgery. On postoperative day 28, neuronal damage, microglial activity, and microglial differentiation were assessed. The pregabalin group exhibited significantly less neuronal damage compared to the control group, along with a significant decrease in activated microglial expression in both the brain and spinal cord. Pregabalin treatment also significantly altered the microglial phenotype expression, with a decrease in the M1 phenotype percentage and an increase in the M2 phenotype percentage in both the brain (M1 phenotype: 43.52 ± 12.16% and 18.00 ± 8.57% in the control and pregabalin groups, respectively; difference: 27.26 [15.18-42.10], p = 0.002; M2 phenotype: 16.88 ± 6.47% and 39.63 ± 5.82% in the control and pregabalin groups, respectively; difference 22.04 [17.17-32.70], p < 0.001) and the spinal cord ipsilateral to nerve injury (M1 phenotype: 44.35 ± 12.12% and 13.78 ± 5.39% in the control and pregabalin groups, respectively; difference 30.46 [21.73-44.45], p < 0.001; M2 phenotype: 7.64 ± 3.91% and 33.66 ± 7.95% in the control and pregabalin groups, respectively; difference 27.41 [21.21-36.30], p < 0.001). Overall, pregabalin treatment significantly decreased the microglial M1 phenotype while increasing the microglial M2 phenotype in NP rats.

Keywords: Allodynia; Cytokine; Microglia; Neuropathic pain; Pregabalin.

Figure 1

Assessment of mechanical (A) and cold (B) allodynia for neuropathic pain before and after the treatment of normal saline or pregabalin. () Contralateral side for the surgery, Control group, () Contralateral side for the surgery, Pregabalin group, () Ipsilateral side for the surgery, Control group, () Ipsilateral side for the surgery, Pregabalin group. Abbreviations: Control, Control group; Pregabalin, Pregabalin group; D0, before the surgery; D, day after the surgery; before, before the treatment of normal saline or pregabalin; after, 60 minutes after the treatment of normal saline or pregabalin. ^*p < 0.05 compared with D0 in intra-group variance. ^†p < 0.05 compared with Control group.

Figure 2

Neuronal damage in the brain (A) and the L5 segment of the spinal cord (B). Abbreviations: DAPI, 4,6-diamidino-2-phenylindole staining; terminal deoxynucleotidyl transferase (TdT) deoxyuridine triphosphates (dUTP) nick end labeling (TUNEL) staining; Merged, merged image of DAPI staining and TUNEL staining. Control, Control group; Pregabalin, Pregabalin group; Contralateral, contralateral side for surgery; Ipsilateral, Ipsilateral side for the surgery. ^*p < 0.05 compared with Control group. ^†p < 0.05 compared with contralateral side in each group.

Figure 3

Activated microglia in the brain (A) and the L5 segment of the spinal cord (B). Abbreviations: DAPI, 4,6-diamidino-2-phenylindole staining; Iba1, ionized calcium-binding adapter molecule 1 staining; TMEM, rabbit transmembrane protein 119 staining; Merged, merged image of DAPI staining, Iba1 staining and TMEM staining; Control, Control group; Pregabalin, Pregabalin group; Contralateral, contralateral side for surgery; Ipsilateral, Ipsilateral side for the surgery. ^*p < 0.05 compared with Control group. ^†p < 0.05 compared with contralateral side in each group.

Figure 4

Differentiation of microglia in the brain (A) and the L5 segment of the spinal cord (B). Abbreviations: DAPI, 4,6-diamidino-2-phenylindole staining; Iba1, ionized calcium-binding adapter molecule 1 staining; CD, cluster differentiation; Merged, merged image of DAPI staining, Iba1 staining, and CD16 (for M1 phenotype) or CD206 staining (for M2 phenotype); Control, Control group; Pregabalin, Pregabalin group; Contralateral, contralateral side for surgery; Ipsilateral, Ipsilateral side for the surgery. ^*p < 0.05 compared with Control group. ^†p < 0.05 compared with contralateral side in each group.