Pregnenolone blocks cannabinoid-induced acute psychotic-like states in mice

Abstract

Cannabis-induced acute psychotic-like states (CIAPS) represent a growing health issue, but their underlying neurobiological mechanisms are poorly understood. The use of antipsychotics and benzodiazepines against CIAPS is limited by side effects and/or by their ability to tackle only certain aspects of psychosis. Thus, safer wide-spectrum treatments are currently needed. Although the blockade of cannabinoid type-1 receptor (CB1) had been suggested as a therapeutical means against CIAPS, the use of orthosteric CB1 receptor full antagonists is strongly limited by undesired side effects and low efficacy. The neurosteroid pregnenolone has been recently shown to act as a potent endogenous allosteric signal-specific inhibitor of CB1 receptors. Thus, we tested in mice the potential therapeutic use of pregnenolone against acute psychotic-like effects of Δ⁹-tetrahydrocannabinol (THC), the main psychoactive component of cannabis. We found that pregnenolone blocks a wide spectrum of THC-induced endophenotypes typically associated with psychotic-like states, including impairments in cognitive functions, somatosensory gating and social interaction. In order to capture THC-induced positive psychotic-like symptoms (e.g. perceptual delusions), we adapted a behavioral paradigm based on associations between different sensory modalities and selective devaluation, allowing the measurement of mental sensory representations in mice. Acting at hippocampal CB1 receptors, THC impaired the correct processing of mental sensory representations (reality testing) in an antipsychotic- and pregnenolone-sensitive manner. Overall, this work reveals that signal-specific inhibitors mimicking pregnenolone effects can be considered as promising new therapeutic tools to treat CIAPS.

Figure 1. Pregnenolone blocks the cognitive impairment induced by THC.

(A) Dose-response of THC effects in the spontaneous alternation task [1-Way ANOVA, F(3,32)=3.8, p<0.001, n=6-12]. (B) Effect of pregnenolone (6 mg/kg, s.c.) 10 min before or 30 min after THC (3 mg/kg, i.p.) administration on the impairment of spontaneous alternation [before: 2-Way ANOVA, interaction; F(1,58)=14.83, p<0.001; after: 2-Way ANOVA, interaction; F(1,62)=6.871, p<0.05, n=10-20]. (C-D) Effect of pregnenolone (6 mg/kg, s.c.) on the cognitive impairment induced by THC (5 mg/kg, i.p.) in the delayed matching-to-place version of the Morris water maze. (C) Saving ratio for the latency [2-Way ANOVA, interaction, F(1,64)=6.48, p<0.05, n=15-20]. (D) Saving ratio for the path [2-Way ANOVA, interaction, F(1,64)=5.09, p<0.05, n=15-20]. **, p<0.01; ***, p<0.001 for the effect of THC. ^#, p<0.05; ^##, p<0.01 for the effect of pregnenolone in THC injected mice.

Figure 2. Pregnenolone blocks the effects of THC on PPI and social interaction.

(A) Dose response of THC effects in the PPI test [1-Way ANOVA, pre-pulse 73 db (PP73), F(3,37)=0.47, p<0.01; PP76, F(3,37)=0.14, p<0.05; PP82, F(3,37)=0.11, p<0.05; n=7-13]. (B) Effect of pregnenolone (6 mg/kg, s.c.) on the reduction of PPI induced by THC (10 mg/kg, i.p.) [2-Way ANOVA, interaction; PP73 F(1,35)=11.93, p<0.01; PP76 F(1,35)=6.31, p<0.05; PP82 F(1,35)=7.63, p<0.01; n=8-12]. (C) Dose response analysis of THC effects in the social interaction task [1-Way ANOVA, F(3,33)=11.23, p<0.01, n=10-12]. (D) Effect of pregnenolone on the reduction of social interaction induced by THC (3 mg/kg, i.p.) [2-Way ANOVA, interaction, F(1,47)=4.39, p<0.05, n=10-15]. *, p<0.05; ** p<0.01; ***, p<0.001 for the effect of THC. ^#, p<0.05, ^##, p<0.01 for the effect of pregnenolone in THC injected mice.

Figure 3. Pregnenolone blocks the hyperlocomotor effect induced by THC.

(A) Dose response analysis of the effects of THC on spontaneous locomotion [1-Way ANOVA, F(3,28)=0.7, p<0.001, n=8]. (B) Effect of pregnenolone (6 mg/kg, s.c.) on the hyperlocomotion induced by 0.3 mg/kg (i.p.) [2-Way ANOVA, interaction, F(1,33)=6.8, p<0.05, n=10-12]. *, p<0.05; **, p<0.01; ***, p<0.001 as compared to vehicle control. ^#, p<0.05 as compared to THC alone group.

Figure 4. Validation of the mediated aversion “reality testing” paradigm in mice.

(A) Schematic representation of the behavioral protocol. For further details see Methods and Supplementary Figure 4. (B-C) Effect of the number of odor-taste pairings during preconditioning phase on odor-mediated taste aversion: (B) Consumption of tastes (i.e. mediated CS) [2-Way ANOVA with repeated measures, interaction, F(2,27)=3.98, p<0.05; n=10]. Solid bars indicate consumption of taste paired with devaluated odor (mCS+), and dashed bars indicate consumption of taste paired with undevaluated odor (mCS-, see methods). (C) Aversive index [1-Way ANOVA, F(2,27)=0.3, p<0.05; n=10-12]. (D-E) Effects of psychotogenic treatments on mediated aversion “reality testing” paradigm (mice receiving 6 odor/taste pairings during pre-conditioning): (D) Aversion index in adult animals subchronically treated with MK801 during adolescence (post-natal days 21-28, 1 mg/kg, i.p.) (t-test, p<0.05, n=10-11). (E) Effect of acute pre-test administration of amphetamine (2.5 mg/kg, i.p.) [2-Way ANOVA, interaction, F(1,56)=5.9, p<0.05; n=10-20]. *, p<0.05; **, p<0.01 as compared to vehicle control, ***, p<0.001 as compared to mCS+; @, p<0.05 as compared to 1- and 6-pairings groups; ^#, p<0.05 as compared to amphetamine group.

Figure 5. Pregnenolone blocks the THC effects on “reality testing” in mice.

(A-C) Effect of pre-test THC administration (1 mg/kg, i.p.) on mediated aversion in mice trained with 6 odor-taste pairings. (A) Risperidone (0.3 mg/kg, i.p.) effect on the THC induced “reality testing” impairment [2-Way ANOVA, interaction, F(1,53)=5.9, p<0.05; n=11-19]. (B) Effect of the orthosteric CB1 receptor antagonist rimonabant (1 mg/kg, i.p.) [2-Way ANOVA, interaction, F(1,39)=7.1, p<0.05; n=10-11]. (C) Effect of the signal-specific allosteric inhibitor of CB1 receptors, pregnenolone (6 mg/kg, s.c.) before [2-Way ANOVA, interaction, F(1,46)=9.172, p<0.01; n=14-18] and after THC administration. (D) Effect of hippocampal injections of rimonabant [2-Way ANOVA, interaction, F(1,58)=12.04, p<0.001; n=11-21] and pregnenolone [2-Way ANOVA, interaction, F(1,55)=4.252, p<0.05; n=14-18] (3 μg/μl) before THC administration.*, p<0.05, **, p<0.01, ***, p<0.001 for the effect of THC; ^#, p<0.05, ^##, p<0.01, ^###, p<0.01 for the effect of pregnenolone in THC injected mice.