Peripheral and central sites of action for the non-selective cannabinoid agonist WIN 55,212-2 in a rat model of post-operative pain

Abstract

Background and purpose: Activation of cannabinoid (CB) receptors decreases nociceptive transmission in inflammatory or neuropathic pain states. However, the effects of CB receptor agonists in post-operative pain remain to be investigated. Here, we characterized the anti-allodynic effects of WIN 55,212-2 (WIN) in a rat model of post-operative pain.

Experimental approach: WIN 55,212-2 was characterized in radioligand binding and in vitro functional assays at rat and human CB(1) and CB(2) receptors. Analgesic activity and site(s) of action of WIN were assessed in the skin incision-induced post-operative pain model in rats; receptor specificity was investigated using selective CB(1) and CB(2) receptor antagonists.

Key results: WIN 55,212-2 exhibited non-selective affinity and agonist efficacy at human and rat CB(1) versus CB(2) receptors. Systemic administration of WIN decreased injury-induced mechanical allodynia and these effects were reversed by pretreatment with a CB(1) receptor antagonist, but not with a CB(2) receptor antagonist, given by systemic, intrathecal and supraspinal routes. In addition, peripheral administration of both CB(1) and CB(2) antagonists blocked systemic WIN-induced analgesic activity.

Conclusions and implications: Both CB(1) and CB(2) receptors were involved in the peripheral anti-allodynic effect of systemic WIN in a pre-clinical model of post-operative pain. In contrast, the centrally mediated anti-allodynic activity of systemic WIN is mostly due to the activation of CB(1) but not CB(2) receptors at both the spinal cord and brain levels. However, the increased potency of WIN following i.c.v. administration suggests that its main site of action is at CB(1) receptors in the brain.

Figure 1

Effects of systemic WIN 55,212-2 (WIN) on skin incision-induced post-operative pain demonstrated as the increase of paw withdrawal threshold. (A) WIN (1, 3 and 10 µmol·kg⁻¹) was injected i.p. in rats 90 min after surgery. (B) Reversal of WIN-induced analgesic efficacy by pretreatment with CB receptor antagonists, selective CB₁ receptor antagonist SR141716A (SR1, 30 µmol·kg ⁻¹, i.p.) or selective CB₂ receptor antagonist SR144528 (SR2, 10 µmol·kg⁻¹, i.p.) was injected 15 min before the administration of WIN (5 µmol·kg⁻¹, i.p.). Mechanical allodynia was assessed 30 min following WIN treatment. Data are mean ± SEM. *P < 0.05, **P < 0.01 versus vehicle (Veh) treated group, ##P < 0.01, SR1 i.p./WIN i.p. versus Veh i.p./WIN i.p. group, n = 6 per group. Dashed line: paw withdrawal threshold of non-injured paw. CB, cannabinoid.

Figure 2

Effects of peripheral injection of WIN 55,212-2 (WIN) on skin incision-induced post-operative pain demonstrated as the increase of paw withdrawal threshold. (A) WIN (30, 100, 300 nmol per paw) was intraplantarly (i.pl.) injected in rats 90 min after surgery, WIN (100, 300 nmol per paw) was also injected i.pl. to the contralateral (Con) non-injured paw in a separate group of animals. (B) Reversal of WIN-induced anti-allodynic effects by peripheral pretreatment with CB antagonists, selective CB₁ receptor antagonist SR141716A (SR1, 150 nmol per paw) or selective CB₂ receptor antagonist SR144528 (SR2, 150 nmol per paw) was injected i.pl. 15 min before the administration of WIN (5 µmol·kg⁻¹, i.p.). Mechanical allodynia was assessed 30 min after WIN treatment. Data are mean ± SEM. *P < 0.05, **P < 0.01 versus vehicle (Veh) treated group, ##P < 0.01, SR1 i.pl./WIN i.p. or SR2 i.pl./WIN i.p. versus Veh i.p./WIN i.p. group, n = 6 per group. Dashed line: paw withdrawal threshold of non-injured paw. CB, cannabinoid.

Figure 3

Effects of spinal application of WIN 55,212-2 (WIN) on skin incision-induced post-operative pain demonstrated as the increase of paw withdrawal threshold. (A) WIN (30, 100, 300 nmol per rat) was intrathecally (i.t.) applied in rats 90 min after surgery. (B) Reversal of WIN-induced analgesic efficacy by spinal pretreatment with CB antagonists, selective CB₁ receptor antagonist SR141716A (SR1, 150 nmol per rat) or selective CB₂ receptor antagonist SR144528 (SR2, 150 nmol per rat) was intrathecally delivered 15 min before the administration of WIN (5 µmol·kg⁻¹, i.p.). Mechanical allodynia was assessed 30 min after WIN treatment. Data are mean ± SEM. *P < 0.05, **P < 0.01 versus vehicle (Veh) treated group, ##P < 0.01, SR1 i.t./WIN i.p. versus Veh i.t./WIN i.p. group, n = 6 per group. Dashed line: paw withdrawal threshold of non-injured paw. CB, cannabinoid.

Figure 4

Effects of supraspinal infusion of WIN 55,212-2 (WIN) on skin incision-induced post-operative pain demonstrated as the increase of paw withdrawal threshold. (A) WIN (3,10, 30, 100 nmol per rat) was intracerebroventricularly (i.c.v.) infused in rats 90 min after surgery. (B) Reversal of WIN-induced analgesic efficacy by supraspinaly pretreatment with CB antagonists, selective CB₁ receptor antagonist SR141716A (SR1, 150 nmol per rat) or selective CB₂ receptor antagonist SR144528 (SR2, 150 nmol per rat) was i.c.v. infused 15 min before the administration of WIN (5 µmol·kg⁻¹, i.p.). Mechanical allodynia was assessed 30 min after WIN treatment. Data are mean ± SEM. *P < 0.05, **P < 0.01 versus vehicle (Veh) treated group, ## P < 0.01, SR1 i.c.v./WIN i.p. versus Veh i.c.v./WIN i.p. group, n = 6 per group. Dashed line: paw withdrawal threshold of non-injured paw. CB, cannabinoid.

Figure 5

Effects of systemic WIN 55,212-2 (WIN) on spontaneous horizontal exploratory behaviours and rotarod performance. (A) WIN (1, 3 and 10 µmol·kg⁻¹) was i.p. administered in naïve rats 30 min before the assessment of horizontal exploratory activity. (B) WIN (1, 3 and 10 µmol·kg⁻¹) was i.p. administered in naïve rats 30 min before the recording of fall latency from rotarod performance. Data are mean ± SEM. *P < 0.05, **P < 0.01 versus vehicle (Veh) treated group, n = 6–8 per group.