Involvement of spinal serotonin receptors in electroacupuncture anti-hyperalgesia in an inflammatory pain rat model

Abstract

We previously showed that electroacupuncture (EA) activates medulla-spinal serotonin-containing neurons. The present study investigated the effects of intrathecal 5,7-dihydroxytryptamine creatinine sulfate, a selective neurotoxin for serotonergic terminals, the 5-hydroxytryptamine 1A receptor (5-HT1AR) antagonist NAN-190 hydrobromide and the 5-HT2C receptor (5-HT2CR) antagonist SB-242,084 on EA anti-hyperalgesia. EA was given twice at acupoint GB30 after complete Freund's adjuvant (CFA) injection into hind paw. CFA-induced hyperalgesia was measured by assessing hind paw withdrawal latency (PWL) to a noxious thermal stimulus 30 min post-EA. Serotonin depletion and the 5-HT1AR antagonist blocked EA anti-hyperalgesia; the 5-HT2CR antagonist did not. Immunohistochemical staining showed that spinal 5-HT1AR was expressed and that 5-HT2CR was absent in naive and CFA-injected animals 2.5 h post-CFA. These results show a correlation between EA anti-hyperalgesia and receptor expression. Collectively, the data show that EA activates supraspinal serotonin neurons to release 5-HT, which acts on spinal 5-HT1AR to inhibit hyperalgesia.

Fig. 1

Time course of experiments 1–3.

Fig. 2

PWLs before and 9 days after 5,7-DHT (n=16) treatment. There were no significant changes between left and right, or before and after 5,7-DHT.

Fig. 3

Effect of 5,7-DHT treatment on EA-produced anti-hyperalgesia in rats with CFA-induced inflammation (n=8 per group). CFA induced a significant (P<0.05) decrease of ipsilateral PWL in both saline- and 5,7-DHT-treated rats 2.5 hr post-CFA compared to baseline. EA significantly increased PWL in saline- but not 5,7-DHT-treated rats compared to sham EA. #P<0.05 vs saline + sham EA; *P<0.05 vs 5,7-DHT + EA.

Fig. 4

Effect of pretreatment with the 5-HT receptor 2C antagonist SB-242,084 on EA-produced anti-hyperalgesia in rats with CFA-induced inflammation (n=8 per group). CFA induced a significant (P<0.05) decrease of ipsilateral PWL in both saline- and SB-242,084-treated rats 2.5 hr post-CFA compared to baseline. Compared to sham, EA significantly increased PWL at 2.5 hr after either saline or SB-242,084 treatment. #P<0.05 vs saline + sham EA; *P<0.05 vs SB-242,084 + sham EA.

Fig. 5

Effect of pretreatment with 5-HT receptor 1A antagonist NAN-190 hydrobromide on EA-produced anti-hyperalgesia in rats with CFA-induced inflammation (n=7 per group). CFA induced significant (P<0.05) decreases of ipsilateral PWL in both saline-and NAN-190 hydrobromide-treated rats 2.5 post-CFA compared to baseline. EA significantly increased PWLs 2.5 hr in vehicle- but not NAN-190 hydrobromide-treated rats compared to sham EA. #P<0.05 vs vehicle + sham EA; *P<0.05 vs NAN-190 hydrobromide + EA.

Fig. 6

Immunostaining data shows that 5-HT1A receptor expression was similar in naïve animals (A, 16 ± 3) and 2.5 hr post-CFA injection (B, 15± 3). The inset in A is higher magnification of arrow-pointed neurons. Scale is 100 μm.

Fig. 7

Immunostaining data shows that 5-HT2C receptors were absent in naive animals (A) and 2.5 hr (B) post-CFA injection. Scale is 100 μm.