Activation of Spinal α2-Adrenoceptors Using Diluted Bee Venom Stimulation Reduces Cold Allodynia in Neuropathic Pain Rats

Abstract

Cold allodynia is an important distinctive feature of neuropathic pain. The present study examined whether single or repetitive treatment of diluted bee venom (DBV) reduced cold allodynia in sciatic nerve chronic constriction injury (CCI) rats and whether these effects were mediated by spinal adrenergic receptors. Single injection of DBV (0.25 or 2.5 mg/kg) was performed into Zusanli acupoint 2 weeks post CCI, and repetitive DBV (0.25 mg/kg) was injected for 2 weeks beginning on day 15 after CCI surgery. Single treatment of DBV at a low dose (0.25 mg/kg) did not produce any anticold allodynic effect, while a high dose of DBV (2.5 mg/kg) significantly reduced cold allodynia. Moreover, this effect of high-dose DBV was completely blocked by intrathecal pretreatment of idazoxan (α2-adrenoceptor antagonist), but not prazosin (α1-adrenoceptor antagonist) or propranolol (nonselective β-adrenoceptor antagonist). In addition, coadministration of low-dose DBV (0.25 mg/kg) and intrathecal clonidine (α2-adrenoceptor agonist) synergically reduced cold allodynia. On the other hand, repetitive treatments of low-dose DBV showing no motor deficit remarkably suppressed cold allodynia from 7 days after DBV treatment. This effect was also reversed by intrathecal idazoxan injection. These findings demonstrated that single or repetitive stimulation of DBV could alleviate CCI-induced cold allodynia via activation of spinal α2-adrenoceptor.

Figure 1

Effect of single DBV treatment on cold allodynia. A graph illustrating the effect of single treatment with diluted bee venom (DBV) on cold allodynia in neuropathic rats. All the CCI-subjected animal groups (Saline, DBV (2.5 mg), DBV (0.25 mg)) significantly increased paw withdrawal frequency (PWF, %) to innocuous cold stimulus compared with that of the sham surgery animals (Sham, ***P < 0.001). Moreover, the high-dose DBV treatment group (DBV (2.5 mg)) significantly suppressed the CCI-induced PWF from 45 minutes to 90 minutes after DBV treatment compared with the saline-treated group (Saline, ^## P < 0.01 and ^### P < 0.001).

Figure 2

Effect of intrathecal adrenoceptor antagonist injection on the DBV-induced antinociception. The effects of intrathecal adrenoceptor antagonists on the DBV-induced antiallodynic effect to cold stimuli in neuropathic pain animals. The saline pretreatment in DBV-treated rats (Sal + DBV) reduced paw withdrawal frequency (PWF, %) to cold stimulus at 60 and 90 minutes compared with the DBV-treated group (Sal + Sal, **P < 0.01 and ***P < 0.001). The idazoxan-pretreated group (IDX + DBV), but not the PRA- and PRO-treated groups (PRA + DBV and PRO + DBV), significantly reversed PWF from 30 minutes to 90 minutes compared with the saline pretreatment group of DBV-treated rats (Sal + DBV, ^# P < 0.05, ^## P < 0.01 and ^### P < 0.001). All groups were comprised of six animals, respectively. PRA: prazosin, α1-adrenoceptor antagonist, IDX: idazoxzn, α2-adrenoceptor antagonist, PRO: propranolol, nonselective β-adrenoceptor antagonist.

Figure 3

Effect of combined DBV and clonidine treatment on neuropathic pain-induced cold allodynia. A graph demonstrating the analgesic effects of intrathecal clonidine alone or in combination with injection of subcutaneous diluted bee venom (DBV) on CCI-induced cold allodynia. Rats were injected intrathecally with saline (Sal) or clonidine (CLO) 5 minutes after subcutaneous injection of saline or DBV. The intrathecal clonidine-injected rats (Sal + CLO (5 nmol) and Sal + CLO (15 nmol)) dose-dependently suppressed CCI-induced cold allodynic behavior compared with the saline-treated group (Sal + Sal, *P < 0.05 and ***P < 0.001) while the low-dose clonidine group (Sal + CLO (1 nmol)) and DBV injection without clonidine treatment (DBV (0.25 mg) + Sal) had no effect. The combined group of DBV acupuncture and low-dose clonidine (DBV (0.25 mg) + CLO (1 nmol)) significantly reversed CCI-injured cold allodynia compared with the control group (Sal + Sal, *P < 0.05 and ***P < 0.001).

Figure 4

Effect of repetitive DBV treatment on cold allodynia. A graph illustrating the effect of repetitive treatment with diluted bee venom (DBV) from 2 weeks after CCI on cold allodynia in neuropathic rats. The repetitive saline injection group (CCI: R-Sal) significantly increased the paw withdrawal frequency (PWF, %) to 13 days after repetitive saline injection (***P < 0.001 compared with the sham surgery animals, Sham: Naive). The repeated DBV-treated animals (CCI: R-DBV (0.25 mg)) also increased the PWF to day 7 compared with the sham surgery group (**P < 0.01 and ***P < 0.001) and reversed the PWF from 7 days compared with the saline-treated group animals (^### P < 0.001).

Figure 5

Effect of intrathecal α2-adrenoceptor antagonist injection on the repetitive DBV-induced antinociception. The effects of intrathecal α2-adrenoceptor antagonists on repetitive diluted-bee-venom-(DBV-) induced decrease of cold allodynia in CCI animals. Saline and idazoxan were intrathecally injected after repetitive saline and DBV treatment for 2 consecutive weeks. In the repetitive saline injection group, intrathecal idazoxan injection (R-Sal: Idazoxan) did not change the cold allodynic behavior compared with the saline-injected animals (R-Sal: Saline), whereas intrathecal idazoxan injection in the repetitive DBV group (R-DBV (0.25 mg): Idazoxan) significantly reversed the reduced response to cold stimulus compared with the saline-treated rats (R-DBV (0.25 mg): Saline, *P < 0.05).