Prenatal cannabis exposure (PCE) has been associated with altered prefrontal cortex (PFC) activity and connectivity in adulthood, potentially increasing the risk of psychopathology later in life. This risk is thought to involve a complex interplay between the endocannabinoid and dopaminergic systems. We investigated the transcriptional regulation of genes associated with these systems in an animal model of PCE during adolescence, focusing on DNA methylation and specific microRNAs (miRNAs). Our study revealed increased mRNA levels of dopamine D1 and D2 receptors (Drd1 and Drd2) in the PFC, with a notable effect on Drd2 in male offspring. Notably, we observed a consistent reduction in Drd2 DNA methylation levels in PCE male rats. Both Drd1 and Drd2 expressions were regulated by selective miRNAs. Accordingly, we found changes in the excitability of PFC pyramidal neurons in male adolescent PCE offspring, along with alterations in the Netrin-1/DCC guidance cue system. Our findings highlight PCE-induced modifications of the PFC dopaminergic system while maintaining stable gene expression of the endocannabinoid system in male offspring. Changes in this complex interaction during sensitive developmental periods like adolescence might lead to sex-dependent divergent behavioral outcomes induced by PCE.

Background/Objectives: High-altitude exposure represents a complex psychophysiological stressor involving hypoxia, physical effort, sleep disruption and psychological strain. The endocannabinoid system (ECS) plays a key role in stress regulation, yet its epigenetic modulation under extreme environmental conditions remains poorly characterized. This pilot and exploratory study investigated DNA methylation and descriptive microRNA (miRNA) expression patterns of CNR1 and FAAH genes, and their associations with mood and anxiety outcomes, in trekkers exposed to Himalayan high altitude. Methods: Twenty-one healthy lowlanders completed a longitudinal expedition from 2860 m to 5050 m. Psychometric measures (SVARAD, BDI, SAS, SHAPS) and saliva samples were collected at baseline (T0) and at high altitude (T1). DNA methylation of CNR1 and FAAH regulatory regions was quantified by pyrosequencing. Exosomal miRNAs targeting these genes were profiled using qRT-PCR, on pooled samples; results are presented descriptively. Results: DNA methylation analysis revealed heterogeneous, sex-specific epigenetic patterns following high-altitude exposure. A significant increase in CNR1 promoter methylation at CpG4 was observed in males at T1, whereas methylation remained largely stable in females. Descriptive miRNA expression data showed bidirectional differences between groups, consistent with context-dependent stress regulation. Convergent directional patterns between miR-23b-3p expression and CNR1 methylation in males were observed. However, given the descriptive nature of the miRNA data, this observation is purely exploratory and requires replication before any mechanistic conclusions can be drawn. Psychometrically, participants showed a mild mood decline without overt clinical symptoms. Sex-specific differences in the relationship between CNR1 methylation and psychometric outcomes were observed and warrant further investigation in adequately powered cohorts. Conclusions: These preliminary findings suggest that CNR1 epigenetic regulation warrants further investigation as a potential indicator of stress adaptation and psychological responses and underscore the need to consider sex differences when evaluating resilience and vulnerability to extreme environments.