Sensitivity and Resilience to Predator Stress-Enhanced Ethanol Drinking Is Associated With Sex-Dependent Differences in Stress-Regulating Systems

Abstract

Stress can increase ethanol drinking, and evidence confirms an association between post-traumatic stress disorder (PTSD) and the development of alcohol use disorder (AUD). Exposure to predator odor is considered a traumatic stressor, and predator stress (PS) has been used extensively as an animal model of PTSD. Our prior work determined that repeated exposure to intermittent PS significantly increased anxiety-related behavior, corticosterone levels, and neuronal activation in the hippocampus and prefrontal cortex in naïve male and female C57BL/6J mice. Intermittent PS exposure also increased subsequent ethanol drinking in a subgroup of animals, with heterogeneity of responses as seen with comorbid PTSD and AUD. The present studies built upon this prior work and began to characterize “sensitivity” and “resilience” to PS-enhanced drinking. Ethanol drinking was measured during baseline, intermittent PS exposure, and post-stress; mice were euthanized after 24-h abstinence. Calculation of median and interquartile ranges identified “sensitive” (>20% increase in drinking over baseline) and “resilient” (no change or decrease in drinking from baseline) subgroups. Intermittent PS significantly increased subsequent ethanol intake in 24% of male (↑60%) and in 20% of female (↑71%) C57BL/6J mice in the “sensitive” subgroup. Plasma corticosterone levels were increased significantly after PS in both sexes, but levels were lower in the “sensitive” vs. “resilient” subgroups. In representative mice from “sensitive” and “resilient” subgroups, prefrontal cortex and hippocampus were analyzed by Western Blotting for levels of corticotropin releasing factor (CRF) receptor 1, CRF receptor 2, CRF binding protein, and glucocorticoid receptor, vs. separate naïve age-matched mice. In prefrontal cortex, CRF receptor 1, CRF receptor 2, CRF binding protein, and glucocorticoid receptor levels were significantly higher in “sensitive” vs. naïve and “resilient” mice only in females. In hippocampus, CRF receptor 1, CRF receptor 2 and glucocorticoid receptor levels were significantly lower in “resilient” vs. naïve and “sensitive” mice across both sexes. These results indicate that sex strongly influences the effects of ethanol drinking and stress on proteins regulating stress and anxiety responses. They further suggest that targeting the CRF system and glucocorticoid receptors in AUD needs to consider the comorbidity of PTSD with AUD and sex of treated individuals.

Predator odor stress reactivity, alcohol drinking and the endocannabinoid system

“…In mice, repeated exposure to dirty rat bedding can also be used to subgroup rats into “sensitive” and “resilient” phenotypes based on percent change in ethanol drinking before and after repeated stressor exposure ( Alavi et al, 2022 ). Mice that show approximately 20% increase in home cage alcohol drinking after the stressor exposure compared to baseline drinking are defined as “sensitive”, while “resilient” mice show decreased or no change in drinking after the stressor ( Alavi et al, 2022 ). This method to examine individual differences focuses on drinking behavior rather than behavioral coping strategies during the stressor such as avoidance or immobility.…”

“…Exposure to a predator odor stressor has been shown to be a valid and reliable animal model to examine symptom profiles of PTSD. For example, predator odor models can induce PTSD phenotypes including hyperarousal ( Blount et al, 2023 ; Schwendt et al, 2018 ), avoidance behavior ( Edwards et al, 2013 ; Weera et al, 2020 ; Albrechet-Souza et al, 2020 ; Albrechet-Souza and Gilpin, 2019 ; Brodnik et al, 2017 , 2020 ), contextual stress memory ( Ornelas et al, 2021a , 2021b ; Tyler et al, 2020 ) and increases in plasma corticosterone levels ( Ornelas et al, 2021a ; Alavi et al, 2022 ; Finn et al, 2018 ; Cozzoli et al, 2014 ), indicative of changes in hypothalamic-pituitary-adrenal (HPA) axis activity. The HPA axis is the primary neuroendocrine system that regulates physiological and behavioral responses to stress.…”

“…Another predator odor stressor that has been extensively used in mice, is exposure to dirty rat bedding ( Alavi et al, 2022 ; Finn et al, 2018 ; Cozzoli et al, 2014 ; Devaud et al, 2020 ; Manjoch et al, 2016 ; Clark et al, 2023 ; Nipper et al, 2024 ). This model has been used to study sex differences in response to stress, increases in ethanol intake after stressor exposure and neurochemical changes induced by predator odor stress.…”

“…In the hippocampus, both male and female mice showed increased corticotropin releasing factor receptor 1 after predator odor exposure. Finally, hippocampal GR expression was increased in female mice, but not males, after a history of binge ethanol and exposure to predator odor exposure ( Alavi et al, 2022 ). This stressor model and a history of binge drinking has also showed significant sex differences in proteins associated in steroidogenesis and synaptic plasticity levels in the PFC and hippocampus, key regions associated with PTSD ( Devaud et al, 2020 ).…”

Predator Odor Stressor, 2,3,5-trimethyl-3-thiazoline (TMT): Assessment of stress reactive behaviors during an animal model of traumatic stress in rats

“…Predator odor stressor exposure can also be used to investigate the complex interaction between increased alcohol consumption and stress, which may help in understanding the neurobiology associated with comorbid PTSD and alcohol use disorder (Ornelas, Tyler, et al, 2021; Weera et al, 2020). For example, several studies have shown increased alcohol drinking (Alavi et al, 2022; Albrechet-Souza, Schratz, & Gilpin, 2020; Makhijani, Franklin, Van Voorhies, Fortino, & Besheer, 2021) or alcohol relapse-like behavior (Becker, Lopez, King, & Griffin, 2023; King & Becker, 2019) following exposure to varying predator odor stressors, including TMT. Importantly, not all individuals who experience a traumatic event develop PTSD.…”