Voluntary adolescent alcohol exposure does not robustly increase adulthood consumption of alcohol in multiple mouse and rat models.

Adolescence is a period of increased risk taking, including increased alcohol and drug use. Multiple clinical studies report a positive relationship between adolescent alcohol consumption and risk of developing an alcohol use disorder (AUD) in adulthood. However, few preclinical studies have attempted to tease apart the biological contributions of adolescent alcohol exposure, independent of other social, environmental, and stress factors, and studies that have been conducted show mixed results. Here we use several adolescent voluntary consumption of alcohol models, conducted across four labs in three institutes and with two rodent species, to investigate the ramifications of adolescent alcohol consumption on adulthood alcohol consumption in controlled, pre-clinical environments. We consistently demonstrate a lack of robust increases in adulthood alcohol consumption. This work highlights that risks seen in both human datasets and other murine drinking models may be due to unique social and environmental factors, some of which may be unique to humans.

Adolescent intermittent ethanol (AIE) produces lasting, sex-specific changes in rat body fat independent of changes in white blood cell composition.

Early initiation of alcohol use during adolescence, and adolescent binge drinking are risk factors for the development of alcohol use disorder later in life. Adolescence is a time of rapid sex-dependent neural, physiological, and behavioral changes as well as a period of heightened vulnerability to many effects of alcohol. The goal of the present studies was to determine age-related changes in blood (leukocyte populations) and body composition across adolescence and early adulthood, and to investigate whether adolescent intermittent ethanol (AIE) exposure would alter the trajectory of adolescent development on these broad physiological parameters. We observed significant ontogenetic changes in leukocyte populations that were mirrored by an age-related increase in cytokine expression among mixed populations of circulating leukocytes. Despite these developmental changes, AIE did not significantly alter overall leukocyte numbers or cytokine gene expression. However, AIE led to sex-specific changes in body fat mass and fat percentage, with AIE-exposed male rats showing significantly decreased fat levels and female rats showing significantly increased fat levels relative to controls. These changes suggest that while AIE may not alter overall leukocyte levels, more complex phenotypic changes in leukocyte populations could underlie previously reported differences in cytokine expression. Coupled with long-term shifts in adipocyte levels, this could have long-lasting effects on innate immunity and the capacity of individuals to respond to later immunological and physiological threats.

Intermittent Exposure to a Single Bottle of Ethanol Modulates Stress Sensitivity: Impact of Age at Exposure Initiation.

Alcohol use during adolescence is a serious public health problem, with binge drinking and high-intensity drinking being particularly harmful to the developing adolescent brain. To investigate the adverse consequences of binge drinking and high-intensity adolescent drinking, adolescent rodents were intermittently exposed to ethanol through intragastric gavage, intraperitoneal injection, or vapor inhalation. These models revealed the long-lasting behavioral and neural consequences of adolescent intermittent ethanol (AIE) exposure. The present study was designed to characterize a different AIE model, namely, intermittent exposure to a single bottle of 10% ethanol as the only source of fluids on a 2 days on/2 days off (water days) schedule, and to determine whether this AIE exposure model would produce changes in hormonal and neuroimmune responsiveness to challenges of differing modalities. Assessments of ethanol intake as well as blood and brain ethanol concentrations (BECs and BrECs, respectively) in adult male and female rats (Experiment 1) revealed that BECs and BrECs peaked following access to ethanol for a 2 h period when assessed 1 h into the dark cycle. Experiment 2 revealed age differences in ethanol intake, BECs, and BrECs following a 2 h access to ethanol (1 h into the dark cycle), with adolescents ingesting more ethanol and reaching higher BECs as well as BrECs than adults. In Experiment 3, intermittent exposure to a single bottle of 10% ethanol for 10 cycles of 2 days on/2 days off was initiated either in early or late adolescence, followed by an acute systemic immune challenge with lipopolysaccharide (LPS) in adulthood. LPS increased corticosterone and progesterone levels regardless of sex and prior ethanol history, whereas an LPS-induced increase in cytokine gene expression in the hippocampus was evident only in ethanol-exposed males and females, with females who underwent early exposure to ethanol being more affected than their later-exposed counterparts. In Experiment 4, intermittent ethanol exposure in females was initiated either in adolescence or adulthood and lasted for 12 ethanol exposure cycles. Then, behavioral (freezing behavior), hormonal (corticosterone and progesterone levels), and neuroimmune (cytokine gene expression in the PVN, amygdala, and hippocampus) responses to novel environments (mild stressors) and shock (intense stressors) were assessed. More pronounced behavioral and hormonal changes, as well as changes in cytokine gene expression, were evident in the shock condition than following placement in the novel environment, with prior history of ethanol exposure not playing a substantial role. Interleukin (IL)-1ß gene expression was enhanced by shock in the PVN, whereas shock-induced increases in IL-6 gene expression were evident in the hippocampus. Together, these findings demonstrate that our intermittent adolescent exposure model enhances responsiveness to immune but not stress challenges, with females being more vulnerable to this AIE effect than males.

Sex-specific effects of ethanol consumption in older Fischer 344 rats on microglial dynamics and Aß(1-42) accumulation.

Chronic alcohol consumption, Alzheimer's disease (AD), and vascular dementia are all associated with cognitive decline later in life, raising questions about whether their underlying neuropathology may share some common features. Indeed, recent evidence suggests that ethanol exposure during adolescence or intermittent drinking in young adulthood increased neuropathological markers of AD, including both tau phosphorylation and beta-amyloid (Aß) accumulation. The goal of the present study was to determine whether alcohol consumption later in life, a time when microglia and other neuroimmune processes tend to become overactive, would influence microglial clearance of Aß(1-42), focusing specifically on microglia in close proximity to the neurovasculature. To do this, male and female Fischer 344 rats were exposed to a combination of voluntary and involuntary ethanol consumption from ∼10 months of age through ∼14 months of age. Immunofluorescence revealed profound sex differences in microglial co-localization, with Aß(1-42) showing that aged female rats with a history of ethanol consumption had a higher number of iba1+ cells and marginally reduced expression of Aß(1-42), suggesting greater phagocytic activity of Aß(1-42) among females after chronic ethanol consumption later in life. Interestingly, these effects were most prominent in Iba1+ cells near neurovasculature that was stained with tomato lectin. In contrast, no significant effects of ethanol consumption were observed on any markers in males. These findings are among the first reports of a sex-specific increase in microglia-mediated phagocytosis of Aß(1-42) by perivascular microglia in aged, ethanol-consuming rats, and may have important implications for understanding mechanisms of cognitive decline associated with chronic drinking.

Neuroendocrine and neuroimmune responses in male and female rats: evidence for functional immaturity of the neuroimmune system during early adolescence.

Adolescence is a developmental period characterized by rapid behavioral and physiological changes, including enhanced vulnerability to stress. Recent studies using rodent models of adolescence have demonstrated age differences in neuroendocrine responses and blunted neuroimmune responding to pharmacological challenges. The present study was designed to test whether this neuroimmune insensitivity would generalize to a non-pharmacological stress challenge. Male and female adolescent (P29-33) and adult (P70-80) Sprague Dawley rats were exposed to intermittent footshock for one-, two-, or two-hours + recovery. Plasma corticosterone and progesterone levels as well as gene expression of several cytokines and c-Fos gene expression in the paraventricular nucleus of the hypothalamus (PVN), the medial amygdala (MeA), and the ventral hippocampus (vHPC) were analyzed. The results of the present study demonstrated differences in response to footshock, with these differences dependent on age, sex, and brain region of interest. Adult males and females demonstrated time-dependent increases in IL-1ß and IL-1R2 in the PVN, with these changes not evident in adolescent males and substantially blunted in adolescent females. TNFα expression was decreased in all regions of interest, with adults demonstrating more suppression relative to adolescents and age differences more apparent in males than in females. IL-6 expression was affected by footshock predominantly in the vHPC of adolescent and adult males and females, with females demonstrating prolonged elevation of IL-6 gene expression. In summary, central cytokine responses to acute stressor exposure are blunted in adolescent rats, with the most pronounced immaturity evident for the brain IL-1 signaling system.

Male, but not female, Sprague Dawley rats display enhanced fear learning following acute ethanol withdrawal (hangover).

The present studies investigated the effects of withdrawal from a single binge-like dose of ethanol (hangover) on fear conditioning in male and female Sprague Dawley rats. In Experiment 1, males and females were given 0 or 3.5 g/kg ethanol intraperitoneally (i.p.) and then conditioned to contextual fear 24 h post injection. Withdrawal from acute ethanol enhanced expression of the conditioned freezing response in males, but not in females. Experiment 2 demonstrated that in males, withdrawal from acute ethanol administered 24 h prior to conditioning enhanced contextual fear conditioning, but not auditory-cued fear conditioning. In Experiment 3, male and female rats were given 3.5 g/kg ethanol, and blood ethanol concentrations (BECs) were assessed at various time points for determination of ethanol clearance. Female rats cleared ethanol at a higher rate than males, with 10 h required for females and 14 for males to eliminate ethanol from their systems. Because females cleared ethanol faster than males, in Experiment 4, females were conditioned 18 h after ethanol administration to keep the interval between ethanol clearance and fear conditioning similar to that of males. Withdrawal from acute ethanol given 18 h prior to conditioning did not affect both contextual and auditory-cued fear conditioning in females. In summary, these results highlight sex differences in the impact of withdrawal from acute ethanol (hangover) on fear learning; suggesting that males are more sensitive to hangover-associated enhancement of negative affect than females.