Timeframe Analysis of Novel Synthetic Cannabinoids Effects: A Study on Behavioral Response and Endogenous Cannabinoids Disruption

Abstract

This study investigates the impact of SCs consumption by assessing the effects of three novel synthetic cannabinoids (SCs); MDMB-CHMINACA, 5F-ADB-PINACA, and APICA post-drug treatment. SCs are known for their rapid onset (<1 min) and prolonged duration (≥5 h). Therefore, this research aimed to assess behavioral responses and their correlation with endocannabinoids (ECs) accumulation in the hippocampus, and EC's metabolic enzymes alteration at different timeframes (1-3-5-h) following drug administration. Different extents of locomotive disruption and sustained anxiety-like symptoms were observed throughout all-encompassing timeframes of drug administration. Notably, MDMB-CHMINACA induced significant memory impairment at 1 and 3 h. Elevated levels of anandamide (AEA) and 2-arachidonoyl glycerol (2-AG) were detected 1 h post-MDMB-CHMINACA and 5F-ADB-PINACA administration. Reduced mRNA expression levels of fatty acid amide hydrolase (FAAH), monoacylglycerol lipase (MAGL) (AEA and 2-AG degrading enzymes, respectively), and brain-derived neurotrophic factor (BDNF) occurred at 1 h, with FAAH levels remaining reduced at 3 h. These findings suggest a connection between increased EC content and decreased BDNF expression following SC exposure. Cognitive disruption, particularly motor coordination decline and progressive loss manifested in a time-dependent manner across all the analyzed SCs. Our study highlights the importance of adopting a temporal framework when assessing the effects of SCs.

Keywords: 5F-ADB-PINACA; APICA; JWH-018; MDMB-CHMINACA; behavior; indazole/indole–carboxamide; locomotive; memory.

Figure 1

Schematic representation of the chemical structures and substituents of indole/indazole derivatives synthetic cannabinoids; MDMB-CHMINACA, 5F-ADB-PINACA, APICA. The ease of insertion of an additional N atom into the heterocyclic system of the classic synthetic cannabinoid JWH-018 and the archetypal aminoalkylindole-based synthetic cannabinoid MDMB-CHMICA structures along with the presence of a carboxamide group, diverse substituents on both the N-1 atom of the indazole ring, and the carboxamide N atom promote the synthesis of many indazole-3-carboxamide based SCs. They usually comprise an indazole-carboxamides or N-1-adamantyl indole-carboxamide at the 3-position. Adapted from [29].

Figure 2

Schematic diagram of behavioral studies. (A) Outline of the exploration trial and anxiety assessment configuration. (B) Layout of the recognition trial arena tests. Adapted from [39].

Figure 3

Effects of 1 mg/kg systemic administration of MDMB-CHMINACA, APICA, and 5F-ADB-PINACA on the (A) total distance traveled (cm) and (B) velocity (cm/s) during the open field test in mice. Drugs were administered 1 h, 3 h, and 5 h prior to the test. All drug-treated groups were compared with the respective vehicle-treated group (control). Each bar represents the mean ± SEM of 10 mice for each treatment. Statistical analysis was performed by two-way ANOVA followed by Tukey’s post hoc for multiple comparison among data set. * p < 0.05, ** p < 0.01, and *** p < 0.001, indicate a significant difference compared to controls, and + p < 0.05, and +++ p < 0.001 versus 1 h vehicle treated group.

Figure 4

Effect of 1 mg/kg systemic administration of MDMB-CHMINACA, APICA, and 5F-ADB-PINACA on the anxiety-like behavior of mice. Anxiety-like behavior was measured as the amount of time (s) mice spent in the INNER area vs. the OUTER area of the arena during the OFT. Longer periods in the OUTER area reflect anxiety-like behavior. All drug-treated groups’ INNER fragments were compared to the INNER fragment of the vehicle-treated groups (control). Each bar represents the mean ± SEM of 10 mice for each treatment. Statistical analysis was performed by two-way ANOVA followed by Bonferroni’s post hoc for multiple comparison among data set * p < 0.05, ** p < 0.01, and *** p < 0.001, indicate significant differences compared to control. ns, not significant.

Figure 5

Effects of 1 mg/kg systemic administration of MDMB-CHMINACA, APICA, and 5F-ADB-PINACA on the recognition index for the novel object recognition test in mice. Trial is based on the index (%) of exploration and consequent preferences of mice toward a novel object compared to a previously familiarized object. Values closer to one indicate no memory and learning impairment and values below 0.5 suggest strong memory impairment. All drug treatments were administered intraperitoneally. Each bar represents the mean ± SEM of 10 mice for each treatment. Statistical analysis was performed by two-way ANOVA followed by Tukey’s post hoc for multiple comparison among data set. ** p < 0.01 and *** p < 0.001 indicate significant differences compared to control (vehicle).

Figure 6

Effect of MDMB-CHMINACA, APICA, and 5F-ADB-PINACA on the mRNA expression levels of brain-derived neurotrophic factor (BDNF). Mice were administered 1 mg/kg of either MDMB-CHMINACA, APICA, 5F-DB-PINACA, or vehicle (control) at 1 h, 3 h, and 5 h, as shown. Their hippocampi were collected after treatment. The indicated/relative levels of mRNA were analyzed via quantitative reverse transcription real-time polymerase chain reaction (qRT-PCR) and normalized to those of β-actin. The bars represent the mean ± SEM of 5 mice. Statistical analysis was performed by two-way ANOVA followed by Tukey’s post hoc for multiple comparison among data set. ** p < 0.01 indicates significant difference compared to control.

Figure 7

Quantification of the endogenous cannabinoids (A) AEA and (B) 2-AG content in hippocampal tissue of male C57BL/6J mice 1, 3, and 5 h after synthetic cannabinoid treatment. Each mouse was administered a 1 mg/kg dose of MDMB-CHMINACA, APICA, 5F-ADB-PINACA, or vehicle (control) as shown. Subsequently, endocannabinoid content was quantified via UPLC-TOF/MS. Each drug treatment was compared to its control (vehicle). Each bar represents the mean ± SEM of 5 mice for each treatment. Statistical analysis was performed by two-way ANOVA followed by Tukey’s post hoc for multiple comparison among data set. * p < 0.05, ** p < 0.01, and *** p < 0.001, indicate significant differences compared to control.

Figure 8

Effect of MDMB-CHMINACA, APICA, and 5F-ADB-PINACA on the mRNA expression of (A) AEA-degrading enzyme (FAAH) and (B) 2-AG degrading enzyme (MAGL). Mice were administered 1 mg/kg of MDMB-CHMINACA, APICA, 5F-DB-PINACA, or vehicle (control) at 1, 3, and 5 h, as shown. Their hippocampi were collected after treatment. The indicated/relative levels of mRNA were analyzed via quantitative reverse transcription real-time polymerase chain reaction (qRT-PCR) and normalized to those of β-actin. The bars represent the mean ± SEM of 5 mice. Statistical analysis was performed by two-way ANOVA followed by Tukey’s post hoc for multiple comparison among data set. * p < 0.05 and ** p < 0.01, indicate significant differences compared to control.