Evaluation of the participation of the endocannabinoid system in the extinction of conditioned fear memory of mice

Author(s)

Larissa Fernanda Matias Werworn

• Document type: Master's Dissertation
• Press: Ribeirão Preto.
• Institution: Universidade de São Paulo (USP). Faculdade de Ciências Farmacêuticas de Ribeirão Preto (PCARP/BC)
• Defense date: 2022-08-25

Examining board members

Sabrina Francesca de Souza Lisboa; Amanda Ribeiro de Oliveira; Cristina Aparecida Jark Stern

• Advisor: Sabrina Francesca de Souza Lisboa

Abstract

Posttraumatic stress disorder (PTSD), a debilitating psychiatric disorder, can develop after exposure to traumatic events. The approved treatments are little effective, have long latency period to start the action, and numerous adverse effects. Therefore, studies that investigate mechanisms involved in PTSD are importante to provide new therapeutic perspectives more effectively. The endocanabinoid (eCB) system is among the biological systems that can possibly be involved in the PTSD pathophysiology. Initially, the objective was to test the hypothesis that a repeated social defeat stress (RSDS)-induced dysfunction in the eCB signaling would be involved in PTSD-related behavioral changes. For this purpose, we performed experiments aiming at standardizing RSDS and contextual fear conditioning (CFC) protocols, using different behavioral tests after stress (Study 1). To perform the RSDS, a CD-1 mouse was introduced into the homecage of the C57Bl/6 mice, in order to disturb the social hierarchy and attack the residente animals (6 consecutive days, 2 hours a day). In one of the experiments, AA-5HT (TRPV1 antagonist and FAAH inhibitor; 0.1, 0.3 and 1.0 mg/kg, i.p.) was administered 1 hour before each stress session. After 1 week, the CFC protocol was performed, in which the mice were placed in a conditioning chamber for 7 minutes and 30 seconds, where They received 3 random electric footshocks (0.75 mA, 2s each). After 24 hours, They were re-exposed to the chamber, without footshocks, for 20 minutes (session of evocation and acquisition of extinction; day 2). After another 24 hours, They were re-exposed again to the chamber (10 minutes) to test the evocation of extinction memory (day 3). RSDS protocol did not induce consistente anxiogenic effect or CFC memory extinction déficits, as expected and previously reported. In the experimente with AA-5HT, althought RSDS also did not induce changes in the CFC, the effects of the drug are inconclusive due to the low number of animals used. Due to the lack of efficacy of RSDS and the fact that the MCC protocol produced consistent results, the objective of this work was changed to: test the hypothesis that CB2 receptors would modulate CFC memory extinction. For this purpose, in the same CFC protocol explained before, a CB2 receptor agonist (JWH133) and a CB2 antagonist/inverse agonist (AM630), both at doses of 0.5, 1.0, 2.0 or 4.0 mg/kg, i.p., were administered 30 minutes prior to box re-exposure on Day 2. JWH133 (2.0 mg/kg) facilitated the acquisition of extinction memory without, however, interfering with the evocation of this memory, at least in short term. AM630 (2.0 mg/kg) also had a slight possible effect facilitating extinction (Day 2). One possibility could be the action of eCBs on CB1, so an ineffective dose of AM251 was administered 30 minutes before AM630. The expected effect was not observed. Considering the importance of eCBs in more aversive situations, an experiment was carried out with a protocol involving more intense shocks (3x1.5 mA, 2s) with JWH133 (2.0 and 4.0 mg/kg). JWH133 (2.0 mg/kg) had a more robust facilitating effect on the acquisition of extinction memory, without interfering with the evocation of this memory. The highest dose (4.0 mg/kg) did not interfere with the acquisition of extinction memory, but impaired the evocation of this memory on the following day, evidencing a dual response. These results suggest that CB2 receptors may be involved in the CFC memory extinction process, but in a complex way, probably involving balance of action of eCBs in different receptors and in receptors with different cellular locations. Therefore, more studies are needed to elucidate these questions. (AU)

• FAPESP's process: 19/19226-5 - Evaluation of the involvement of endovanilloid and endocannabinoid systems in behavioral and plastic consequences after trauma exposure in mice
• Grantee: Larissa Fernanda Matias Werworn
• Support Opportunities: Scholarships in Brazil - Master