Tropisetron attenuates neuroinflammation and chronic neuropathic pain via α7nAChR activation in the spinal cord in rats

Abstract

Objectives: Tropisetron is an alpha 7 nicotinic acetylcholine receptor (α7nAChR) agonist and is a commonly used antiemetic clinically. α7nAChRs activation modulating nociception transmissions and cholinergic anti-inflammation may decrease neuropathic pain. This study was set to investigate the effects of tropisetron on neuropathic pain and neuroinflammation as well as the underlying mechanisms in rats.

Methods: Neuropathic pain behavior was assessed in rats using the paw mechanical withdrawal threshold and paw thermal withdrawal latency before and after the establishment of a spared nerve injury (SNI) pain model in rats treated with tropisetron treatment in the presence or absence of the α7nAChR antagonist methyllycaconitine (MLA) through intrathecal injection. Their spinal cords were then harvested for inflammatory cytokines, the α7nAChR, p38 mitogen-activated protein kinase (p-p38MAPK) and cAMP-response element binding protein (CREB) measurement.

Results: Tropisetron effectively alleviated mechanical allodynia and thermal hyperalgesia; decreased IL-6, IL-1ß and TNF-a; and down-regulated the phosphorylation of p38MAPK and CREB. Pre-treatment with MLA abolished these effects of tropisetron.

Conclusion: Our data indicate that tropisetron alleviates neuropathic pain may through inhibition of the p38MAPK-CREB pathway via α7nAChR activation. Thus, tropisetron may be a potential new therapeutic strategy for chronic neuropathic pain.

Keywords: Chronic neuropathic pain; Neuroinflammation; Pain sensitization; Tropisetron; α7nAChR.

Figure 1

Intrathecal injection of tropisetron attenuates SNI-induced chronic neuropathic pain in rats. (A) Tropisetron intrathecally increased the ipsilateral PWMT in SNI rats. (B) The dose–response curve of tropisetron administration in SNI rats. (C, D) The change of PMWT and PTWL in each group at indicated time points. The ED95 of tropisetron (17.47 μg) was used in subsequent experiments. All data are presented as means ± SD (n = 6 per group). *P < .05 and ***P < .001 vs. SNI.

Figure 2

Intrathecal injection of tropisetron reduced SNI-induced inflammatory cytokine release in the rat spinal cord. (A–C) The levels of IL-6, IL-1β, and TNF-α in the spinal cord from each group 1 h after injection, as detected by ELISA. All data are presented as means ± SD (n = 6 per group). ***P < .001 vs. Sham; ##P < .01and ###P < .001 vs. SNI.

Figure 3

α7nAChR expression in the rat spinal cord in each treatment group 1 h after injection, as measured by immunofluorescence and WB. (A) Representative images of spinal dorsal horn regions co-stained with α7nAChR (red) and DAPI (blue). Original magnification: 400×. Scale bar: 25 μm. (B) Quantitation of α7nAChR-positive cells (white arrow in A). (C) Expression of representative images of western blot, and (D) densitometric analysis of α7nAChR in the spinal cord. Data are presented as means ± SD (n = 4 per group). ***P < .001 vs. SNI. ns: not significant.

Figure 4

Effect of the α7nAChR antagonist MLA on the analgesic effect of tropisetron. (A) MLA abolished the effect of tropisetron on the PMWT of SNI rats. (B, C) The change in the PMWT and PTWL after administration of MLA in each group. The dose of MLA (10 μg) was used in subsequent experiments. All data are presented as means ± SD (n = 6 per group). **P < .01 and ***P < .001 vs. SNI + N.S. + tropisetron; #P < .05, ##P < .01and ###P < .001 vs. SNI + MLA + N.S.

Figure 5

Effect of the α7nAChR antagonist MLA on the anti-inflammatory action of tropisetron. (A–C) The levels of IL-6, IL-1β and TNF-α in the spinal cord in each group, as detected by ELISA. All data are presented as means ± SD, n = 6 per group. ns: not significant; *P < .05 and **P < .01 vs. SNI + N.S. + tropisetron; #P < .05 and ##P < .01 vs. SNI + MLA + tropisetron.

Figure 6

Effects of tropisetron and MLA on the expression of α7nAChR, p-p38, and p-CREB in spinal cord tissues. (A) Representative images of western blot, and (B, C) densitometric analyses of phosphorylation/total p38 and phosphorylation/total CREB in the spinal cord as detected by western blotting. Data are presented as means ± SD (n = 4 per group). ***P < .001 vs. Sham; ###P < .001 vs. SNI + N.S.; +P < .05 and ++P < .01 vs. SNI + N.S. + tropisetron.