Involvement of cannabinoid receptors in the rostral ventromedial medulla in a rat model of spinal nerve ligation

A Zaparolli, NC Coimbra, SH Ferreira. School of Medicine of Ribeirao Preto, Ribeirao Preto, Sao Paulo, Brazil

Cannabinoid agonists produce antinociception in several pain models. Studies have demonstrated the efficacy of cannabinoids and modulators of the endocannabinoid system in suppressing neuropathic pain in animal models. Cannabinoids suppress hyperalgesia and allodynia, induced by diverse neuropathic pain states through CB1 and CB2-specific mechanisms. The rostral ventromedial medulla (RVM) is thought to play a crucial role in the antinociceptive actions of cannabinoids (1). However, the involvement of cannabinoids within CNS regions which mediate antinociception is unknown in neuropathic pain models. The aim of the present study was to investigate the involvement of cannabinoid receptors into the RVM in the L5 spinal nerve ligation (SNL) model in rats.

Male Wistar rats (n = 5-14/group), weighing 180–200 g, were used. The protocol (206/2011) was in accordance with the ethical rules for animal experimentation of the University of São Paulo at Ribeirão Preto, which agrees with the Animal Research Ethics adopted by the Brazilian College of Animal Experimentation (COBEA).

The animals were anaesthetized with intraperitoneal (IP) injection of 0.1mL of 10% ketamine (Agener; at 25 mg/kg) and 0.2 mL of 4% xylazine (Dopaser; at 10 mg/kg) and submitted to a stereotaxic surgery for the introduction of guide-cannula into the nucleus raphe magnus, according to the following coordinates: AP: -10.8mm; ML: 0.0mm; DV: 9.5mm. Then, ligation of L5 spinal nerve was carried out in the same group (2). Withdrawal threshold to von Frey filament application to the right hind paw was determined before and after microinjection of ACEA (0.01-1µg/0.2µl), AM1241 (0.1-3µg/0.2µl), URB597 (0.5-4ηmol/0.2µl), URB597 (4ηmol/0.2µl) + AM251 (2.5ηmol/0.2µl) or vehicle into the RVM seven days after the post-operative recovery. Data were submitted to one-way analysis of variance (ANOVA) followed by the post-hoc Bonferroni’s test.

Microinjection of FAAH inhibitor URB597 into the RVM decreased the mechanical hypernociception in the L5 spinal nerve ligation model. Cannabinoid-1 receptor (CB1) antagonist AM251 altered the effect of URB597 microinjected into RVM. The cannabinoid CB1 receptor agonist ACEA into the RVM attenuated the hypernociception in the L5 SNL model. However, cannabinoid CB2 receptor agonist AM1241 did not alter the mechanical hypernociception in the L5 SNL model (Table 1).

Table 1 Effect of FAAH inhibitor, CB1 and CB2 agonists into the RVM in the L5 SNL model

* p<0.05 vs. Naïve, #p<0.05 vs. L5 SNL or vs. Vehicle

EF50 is defined as the von Frey filament force (g) that produces a 50% response frequency

Our data suggest that cannabinoids can reduce nociceptive responses, in part, by modulating descending pathways of pain by acting at CB1 receptors in a rat model of spinal nerve ligation.

(1) Vaughan CW et al, BrJ Pharmacol 127: 935-940, 1999

(2) Mitrirattanakul S et al, Pain 126:102-114, 2006