Domen, M. | bioRxiv Science

bioRxiv2026

Prenatal alcohol and cannabinoid co-exposure increases ethanol-seeking in adult offspring and produces sex-dependent medial prefrontal cannabinoid receptor 1 expression.

Background: Alcohol and cannabinoids are frequently used during pregnancy, however their combined effects on offspring outcomes are currently under-investigated. In this study, co-exposure was compared to single-drug and drug-free exposures for influences on adult alcohol-seeking and cannabinoid system proteomics. Methods: Pregnant C57BL/6J mice received ethanol vapor (ALC), CP-55,940 (0.75 mg/kg; CB), both (ALC+CB), or neither on gestational days 12-15. Beginning on Postnatal Day 200, male and female offspring underwent home-cage ethanol drinking (N=108) and operant self-administration, progressive-ratio, extinction, and reinstatement testing (N=96). CNR1 and CNRIP1 protein abundance was quantified in the medial prefrontal cortex (mPFC) and dorsomedial striatum (N=57) and compared to within-subject drinking behaviors. Results: Prenatal CB exposure increased cumulative home-cage ethanol intake, primarily in males. At 40% ethanol under progressive-ratio schedules, ALC+CB males consumed more than all other male groups, whereas all exposed female groups consumed more than controls. ALC+CB males also showed greater extinction responding than all other male groups and greater reinstatement intake relative to baseline than CON and ALC males. ALC+CB increased mPFC CNR1 in males relative to all other groups, whereas CB and ALC+CB reduced mPFC CNR1 in females. Higher mPFC CNR1 was associated with fixed-ratio and 40% progressive-ratio intake, extinction, and reinstatement exclusively in males. Conclusions: Prenatal co-exposure produced a persistent male phenotype of enhanced ethanol seeking that was distinct from either exposure alone and accompanied by elevated mPFC CNR1. This behavioral and molecular convergence justifies future investigations manipulating corticostriatal cannabinoid signaling to attenuate exposure-induced ethanol consumption in late adulthood.

developmental biology↗

bioRxiv2025

Early Life Outcomes of Prenatal Exposure to Alcohol and Synthetic Cannabinoids in Mice

This study explores the effects of prenatal co-exposure to alcohol and synthetic cannabinoids on offspring viability, physical development, and neurobehavioral outcomes in young adulthood. The aim is to identify distinct outcomes of co-exposure compared to single-drug exposures and to examine potential sex-specific vulnerabilities in motor coordination and exploratory behaviors. Pregnant C57Bl/6J mice were assigned to one of four treatment groups: Control, Alcohol-exposed, Cannabinoid-exposed, or Alcohol+Cannabinoid-exposed, with drug administration occurring between Gestational Days 12-15. Offspring were first evaluated at birth for survival, physical malformations, and developmental delays. Subsequently, young adult offspring were assessed for motor coordination using rotarod tests and exploratory behavior using open field tests. Our results indicate that alcohol and cannabinoid co-exposure significantly reduced offspring survival and litter sizes compared to controls. Non-viable offspring displayed craniofacial abnormalities, limb malformations, and developmental delays. Behavioral assessments in young adulthood demonstrated that all forms of prenatal drug exposure impaired motor coordination in males, while alcohol and cannabinoid exposures independently produced impairments in females. In the open field test, co-exposed male offspring exhibited reduced center exploration, indicative of anxiety-like behavior. Co-exposed offspring, regardless of sex, demonstrated hyperactivity, characterized by increased speed and distance traveled. Together, these findings underscore the heightened risks associated with prenatal polysubstance exposure, which exacerbates offspring mortality and induces sex-specific neurobehavioral deficits. This study highlights the distinct outcomes associated with prenatal co-exposure, and the need for future research to investigate underlying mechanisms driving these developmental disruptions and sex-specific susceptibilities. Structured AbstractO_ST_ABSPurposeC_ST_ABSThis study investigates the effects of prenatal co-exposure to alcohol and synthetic cannabinoids on offspring viability, physical development, and neurobehavioral outcomes in young adulthood. The goal of this investigation is to determine whether prenatal co-exposure produces distinct outcomes from single-drug exposures, including sex-specific vulnerabilities in motor coordination and exploratory behaviors. MethodsPregnant C57Bl/6J mice were randomly assigned to one of four treatment groups: drug-free controls, alcohol (ALC)-exposed, cannabinoid (CP-55,940, CB)-exposed or ALC+CB-exposed, with drug exposure occurring between Gestational Days 12-15. Offspring viability, physical malformations, and developmental delays were first assessed at birth. Then, behavioral evaluations, including rotarod and open field tests, were conducted on young adult offspring (Postnatal Days 100-120). ResultsALC+CB exposure significantly decreased litter survival (p = 0.006) and offspring viability compared to controls. Non-viable offspring exhibited craniofacial abnormalities, limb malformations, and developmental delays. Assessments of rotarod performance revealed that all exposures reduced motor coordination in males compared to controls (p < 0.05), while ALC and CB exposures alone produced this outcome in females. Open field tests indicated that ALC+CB exposure reduced time in the center of the arena in male offspring exclusively, while this same exposure increased hyperactivity compared to single-drug and control groups, independent of sex (p < 0.05). ConclusionsPrenatal co-exposure to alcohol and synthetic cannabinoids exacerbates offspring mortality and induces sex-specific deficits in neurobehavioral motor outcomes. These findings highlight the distinct risks of polysubstance exposure during pregnancy and underscore the need for targeted interventions to mitigate the effects of prenatal polysubstance exposure on offspring health outcomes. HighlightsO_LIAlcohol and cannabinoid co-exposure significantly reduces offspring survival. C_LIO_LIReduced survival is associated with offspring craniofacial and limb malformations. C_LIO_LIPrenatal exposures produce sex-specific impairments in motor coordination. C_LIO_LICo-exposed offspring display hyperactivity in young adulthood. C_LI

animal behavior and cognition↗