Selective pharmacological inhibition of endocannabinoid degradation: effects on physical and psychological stress-induced HPA axis activation in rats

Gemma Ford, Ryan Butler, Mary Reynolds, David Finn. National Uinversity of Ireland, Galway, Galway, Ireland.

Evidence suggests a role for endocannabinoid signalling in pain, anxiety and hypothalamo-pituitary-adrenal (HPA) axis function. Plasma adrenocorticotrophic hormone (ACTH) and corticosterone (CORT) are elevated in rats after cannabinoid administration (Manzanares et al., 1999; Roche et al., 2006). We investigated the effects of pharmacological inhibition of endocannabinoid degradation on stress-induced HPA axis activity. The effects of two stressors, one physical (formalin injection) and one psychological (conditioned fear), alone or combined, on ACTH and CORT responses, were investigated.

Male Lister-Hooded rats (250-330 g, n=7-10 per group) were handled for 3 days before conditioning. The fear-conditioning paradigm was footshock paired with context (10 x 1 s footshocks, 0.4 mA or no footshock controls). Intra-plantar formalin (50μl 2.5% formalin or saline into the right hindpaw) was administered 30 mins prior to a 15 min re-exposure to the aversive context, 24 hours post-footshock. Thirty minutes prior to formalin, animals received an intra-peritoneal injection of either the fatty acid amide hydrolase (FAAH) inhibitor, URB597 (0.3 mg kg-1), the putative monoacyglycerol lipase (MGL) inhibitor, URB754 (13 mg kg-1), or vehicle (ethanol:cremophor:saline). Trunk blood was collected 45 min post-formalin. Plasma corticosterone and ACTH were measured using EIA and ELISA kits (IDS Ltd, UK). Data were analysed by ANOVA and Student-Newman-Keuls or Fisher’s post-hoc tests and expressed as mean ± SEM.

Intra-plantar injection of formalin significantly increased plasma CORT (483 ± 65 ng ml-1) and ACTH (20.7 ± 7.2 pg ml-1) levels in non-fear conditioned animals compared with saline-injected controls (CORT: 286 ± 51 ng ml-1, ACTH: 4.31 ± 1.3 pg ml-1). Fear conditioning significantly reduced formalin-evoked increases in plasma ACTH (3.83 ± 1.2 pg ml-1) but not CORT (527 ± 35 ng ml-1). Plasma CORT levels (537 ± 35 ng ml-1) in non-fear conditioned rats, and both CORT (515 ± 49 ng ml-1) and ACTH levels (19.5 ± 7.69 pg ml-1) in fear-conditioned rats, were significantly increased by prior administration of URB754, compared with saline-injected vehicle controls (Fear conditioned, CORT: 322 ± 39 ng ml-1, ACTH: 4.8 ± 1.7 pg ml-1; non-fear conditioned CORT: 286 ± 51 ng ml-1). Pre-treatment with URB597 did not significantly alter hormone levels.

These data provide evidence for differential modulation of ACTH and CORT with fear conditioning in formalin-treated rats. Further studies are needed to determine the extent to which these differences may relate to the temporal profile of HPA axis activation. The FAAH inhibitor, URB597, had no significant effect on basal or stress-induced HPA axis activity. In light of recent evidence that URB754 does not selectively inhibit MGL, the role of 2-AG in the effects observed remains to be determined.

Manzanares et al., 1999 Brain Res 839, 173-179

Roche et al., 2006 J Neuroimmunology 181, 57-67