Investigating the Roles of Sex Chromosome Complement and Gonadal Status in a Binge Model of Ethanol Consumption
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Research Team
Research Team
Jordan Krenz
Jordan Krenz
Undergraduate Research Assistant
Psychology, Neuroscience Co-Major '27
Emalie Werkowski
Emalie Werkowski
Undergraduate Research Assistant
Psychology, Neuroscience Minor '27
Lindsey O'Lalde
Lindsey O'Lalde
Undergraduate Research Assistant
Psychology, Neuroscience Minor '26
Dr. Anna Radke
Dr. Anna Radke
Principal Investigator, RAD Lab
Department of Psychology
Abstract/Introduction
Abstract/Introduction
Sex differences in ethanol (EtOH) consumption are well established in rodent models, where females typically drink more than males, paralleling the narrowing gender gap in human alcohol use disorder (AUD) prevalence in recent decades. The Drinking in the Dark (DID) paradigm provides a reliable assay for binge-like drinking and escalation of intake in mice. Biological contributors to these sex differences include both gonadal hormones and sex chromosome complement. The Four Core Genotypes (FCG) model dissociates chromosomal sex (XX vs. XY) from gonadal phenotype (Sry+ vs. Sry–), enabling direct tests of how each factor influences motivated behaviors. Previous work demonstrates that removal of gonadal hormones alters binge-like and aversion-resistant drinking across both C57BL/6J and FCG mouse lines, highlighting a regulatory role for testosterone in EtOH intake. Using a single-bottle DID paradigm, the present study investigates how sex chromosome complement and gonadal status interact to shape binge-like EtOH consumption.
Research Question
Research Question
Does the sex chromosome complement, gonadal status, or their interaction drive differences in binge-like ethanol consumption in a single bottle DID paradigm?
Methods
Methods
64 adult gonadal male (Sry+) and female (Sry-) XX and XY FCG mice were selected for this experiment. Mice were generated from breeding pairs obtained from Jackson Laboratory, Bar Harbor, ME and have since been bred in-house. Male (XY/Sry+) FCG mice were paired with C57BL/6J females in harems for breeding. FCG mice (16 per genotype) completed the one-week DID paradigm in two separate cohorts consisting of 32 mice.
During the dark cycle, mice were given access to a single bottle containing 20% ethanol. Ethanol access was limited to 2 hours per day for 4 consecutive days (Monday–Thursday), followed by a 4-hour access period on the fifth day (Friday). Outside of the DID testing periods, mice had continuous access to a single bottle of reverse osmosis (RO) water placed on the home cage.
Consumption is expressed as grams of ethanol per kilogram of body weight (g/kg) with body weights measured every weekday. Any spillage was accounted for with empty “dummy” cages (no mouse). Data was analyzed using GraphPad Prism statistical software. Analysis was conducted via two-way repeated measures ANOVA.
Results/Findings and Discussion
Results/Findings and Discussion
EtOH consumption increased across 2-hour sessions, with sex hormones influencing consumption levels.
Analysis of EtOH consumption across sessions revealed a non-discriminative difference in consumption. A mixed effects analysis was run to evaluate the effects of session, chromosome complement, and Sry status across the first four sessions, with 2-hour access. There was a significant main effect of the session (F(2.840, 169.5) = 5.856, p = 0.0010, ε = 0.9468), as well as a main effect of Sry gene (F(1, 60) = 14.44, p = 0.0003), indicating a consistency in the trend of the absence of the gene in the promotion of ethanol consumption that has been established. There were no significant main effects for chromosome influence (F(1, 60) = 0.7190, p = 0.3999) on EtOH consumption. There were no significant interactions between session and sex chromosome (F(2.840, 169.5) = 0.443, p = 0.7113, ε = 0.9468), session and Sry influence (F(2.840, 169.5) = 2.128, p = 0.1020, ε = 0.9468), sex chromosome and Sry (F(1, 60) = 0.001464, p = 0.9696), or session, sex chromosome, and Sry (F(2.840, 169.5) = 0.8339, p = 0.4716, ε = 0.9468).
EtOH consumption increased over a 4-hour session, with sex hormones influencing consumption levels.
Analysis of EtOH consumption over the fifth session, lasting four hours, revealed a similar trend of increased consumption between the previous 2-hour sessions and the 4-hour session. A mixed effects analysis was once again run, demonstrating that the Sry- mice had again consumed more EtOH than Sry+ mice, reflecting the significant main effect of the absence of the gene on consumption (F(1, 60) = 18.03, p < 0.0001). Additionally, we saw a significant increase in EtOH consumption over the fifth session of the week, aligning with the overarching results of increased consumption between sessions 1 and 4, with a notable increase in session 5 as well (F(3.650, 217.2) = 40.45, p < 0.0001, ε = 0.9125). There were no significant findings by sex chromosome complements across the week (F(1, 60) = 0.2676, p = 0.6068), again reflecting a lack of influence in sex chromosome type on the consumption levels. Additionally, there were no significant interactions between session and sex chromosome (F(3.650, 217.2) = 0.6001, p = 0.6479, ε = 0.9125), session and Sry type (F(3.650, 217.2) = 1.811, p = 0.1339, ε = 0.9125), sex chromosome and Sry type (F(1, 60) = 0.08829, p = 0.7674), or between session, sex chromosome, and Sry (F(3.650, 217.2) = 0.8597, p = 0.4807, ε = 0.9125).
Conclusions and Future Study
Conclusions and Future Study
Consumption levels displayed a significant increase between sessions 1 and 4, aligning with established findings in the binge-like behavior of mice in EtOH drinking.
These results suggest that gonadal female (Sry-) FCG mice display greater EtOH consumption levels, despite a lack of influence of sex chromosome complement.
Results saw no influence made by sex chromosome complement (XX or XY) on either consumption levels across sessions or in interaction with gonadal sex (Sry- or Sry+).
These results display consumption levels in binge-like behaviors influenced across multiple sessions and by gonadal sex (Sry), however there were no observed influences made by sex chromosome type or interactions between each.
Future directions include a greater understanding of how sex chromosome complements influence gonadal sex in binge-drinking behaviors in mice, with a search for significant influences of one on the other.
Future research may also question sex chromosome complements and their influence on levels of binge-drinking behaviors over multiple sessions. Both
Future directions may include a longer timeline to include a greater number of sessions for each length of exposure to EtOH in order to see a greater interaction between sex chromosome complements and gonadal sex on binge-like behaviors.
The following is an image of poster presented at the 2026 Undergraduate Research Forum.
Acknowledgements
Acknowledgements
This research is support by the National Institute on Alcohol Abuse and Alcoholism.
References
References
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NACE Career Readiness Competencies
NACE Career Readiness Competencies
Critical Thinking – Analyzed data from the Four Core Genotypes mouse model to disentangle chromosomal vs. hormonal contributions to observed sex differences.
Teamwork – Collaborated with lab members on animal handling, data collection, and maintaining consistent experimental conditions across groups.
Communication – Presented research findings clearly in posters and discussions, translating complex sex chromosome mechanisms into accessible scientific insights.
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