Adolescent intermittent ethanol exposure alters adult exploratory and affective behaviors, and cerebellar Grin2b expression in C57BL/6J mice.

Binge drinking is one of the most common patterns (more than 90%) of alcohol consumption by young people. During adolescence, the brain undergoes maturational changes that influence behavioral control and affective behaviors, such as cerebellar brain volume and function in adulthood. We investigated long-term impacts of adolescent binge ethanol exposure on affective and exploratory behaviors and cerebellar gene expression in adult male and female mice. Further, the cerebellum is increasingly recognized as a brain region integrating a multitude of behaviors that span from the traditional primary sensory-motor to affective functions, such as anxiety and stress reactivity. Therefore, we investigated the persistent effects of adolescent intermittent ethanol (AIE) on exploratory and affective behaviors and began to elucidate the role of the cerebellum in these behaviors through excitatory signaling gene expression. We exposed C57BL/6J mice to AIE or air (control) vapor inhalation from postnatal day 28-42. After prolonged abstinence (>34 days), in young adulthood (PND 77+) we assessed behavior in the open field, light/dark, tail suspension, and forced swim stress tests to determine changes in affective behaviors including anxiety-like, depressive-like, and stress reactivity behavior. Excitatory signaling gene mRNA levels of fragile X messenger ribonucleoprotein (FMR1), glutamate receptors (Grin2a, Grin2b and Grm5) and excitatory synaptic markers (PSD-95 and Eaat1) were measured in the cerebellum of adult control and AIE-exposed mice. AIE-exposed mice showed decreased exploratory behaviors in the open field test (OFT) where both sexes show reduced ambulation, however only females exhibited a reduction in rearing. Additionally, in the OFT, AIE-exposed females also exhibited increased anxiety-like behavior (entries to center zone). In the forced swim stress test, AIE-exposed male mice, but not females, spent less time immobile compared to their same-sex controls, indicative of sex-specific changes in stress reactivity. Male and female AIE-exposed mice showed increased Grin2b (Glutamate Ionotropic Receptor NMDA Type Subunit 2B) mRNA levels in the cerebellum compared to their same-sex controls. Together, these data show that adolescent binge-like ethanol exposure altered both exploratory and affective behaviors in a sex-specific manner and modified cerebellar Grin2b expression in adult mice. This indicates the cerebellum may serve as an important brain region that is susceptible to long-term molecular changes after AIE.

Adolescent Intermittent Ethanol Exposure Alters Adult Exploratory and Affective Behaviors, and Cerebellar Grin2B Expression in C57BL/6J Mice.

Binge drinking is one of the most common patterns (more than 90%) of alcohol consumption by young people. During adolescence, the brain undergoes maturational changes that influence behavioral control and affective behaviors, such as cerebellar brain volume and function in adulthood. We investigated long-term impacts of adolescent binge ethanol exposure on affective and exploratory behaviors and cerebellar gene expression in adult male and female mice. Further, the cerebellum is increasingly recognized as a brain region integrating a multitude of behaviors that span from the traditional primary sensory-motor to affective functions, such as anxiety and stress reactivity. Therefore, we investigated the persistent effects of adolescent intermittent ethanol (AIE) on exploratory and affective behaviors and began to elucidate the role of the cerebellum in these behaviors through excitatory signaling gene expression. We exposed C57BL/6J mice to AIE or air (control) vapor inhalation from postnatal day 28-42. After prolonged abstinence (>34 days), in young adulthood (PND 77+) we assessed behavior in the open field, light/dark, tail suspension, and forced swim stress tests to determine changes in affective behaviors including anxiety-like, depressive-like, and stress reactivity behavior. Excitatory signaling gene mRNA levels of fragile X messenger ribonucleoprotein ( FMR1) , glutamate receptors ( Grin2a , Grin2B and Grm5 ) and excitatory synaptic markers (PSD-95 and Eaat1) were measured in the cerebellum of adult control and AIE-exposed mice. AIE-exposed mice showed decreased exploratory behaviors in the open field test (OFT) where both sexes show reduced ambulation, however only females exhibited a reduction in rearing. Additionally, in the OFT, AIE-exposed females also exhibited increased anxiety-like behavior (entries to center zone). In the forced swim stress test, AIE-exposed male mice, but not females, spent less time immobile compared to their same-sex controls, indicative of sex-specific changes in stress reactivity. Male and female AIE-exposed mice showed increased Grin2B (Glutamate Ionotropic Receptor NMDA Type Subunit 2B) mRNA levels in the cerebellum compared to their same-sex controls. Together, these data show that adolescent binge-like ethanol exposure altered both exploratory and affective behaviors in a sex-specific manner and modified cerebellar Grin2B expression in adult mice. This indicates the cerebellum may serve as an important brain region that is susceptible to long-term molecular changes after AIE. Highlights: Adolescent intermittent ethanol (AIE) exposure decreased exploratory behavior in adult male and female mice.In females, but not males, AIE increased anxiety-like behavior.In males, but not females, AIE reduced stress reactivity in adulthood.These findings indicate sex differences in the enduring effects of AIE on exploratory and affective behaviors. Cerebellar Grin2B mRNA levels were increased in adulthood in both male and female AIE-exposed mice. These findings add to the small, but growing literature on behavioral AIE effects in mice, and establish cerebellar excitatory synaptic gene expression as an enduring effect of adolescent ethanol exposure.

Targeted epigenomic editing ameliorates adult anxiety and excessive drinking after adolescent alcohol exposure.

Adolescent binge drinking is a major risk factor for psychiatric disorders later in life including alcohol use disorder. Adolescent alcohol exposure induces epigenetic reprogramming at the enhancer region of the activity-regulated cytoskeleton-associated protein (Arc) immediate-early gene, known as synaptic activity response element (SARE), and decreases Arc expression in the amygdala of both rodents and humans. The causal role of amygdalar epigenomic regulation at Arc SARE in adult anxiety and drinking after adolescent alcohol exposure is unknown. Here, we show that dCas9-P300 increases histone acetylation at the Arc SARE and normalizes deficits in Arc expression, leading to attenuation of adult anxiety and excessive alcohol drinking in a rat model of adolescent alcohol exposure. Conversely, dCas9-KRAB increases repressive histone methylation at the Arc SARE, decreases Arc expression, and produces anxiety and alcohol drinking in control rats. These results demonstrate that epigenomic editing in the amygdala can ameliorate adult psychopathology after adolescent alcohol exposure.

Persistence of cerebellar ataxia during chronic ethanol exposure is associated with epigenetic up-regulation of Fmr1 gene expression in rat cerebellum.

BACKGROUND: Alcohol intoxication produces ataxia by affecting the cerebellum, which coordinates movements. Fragile X mental retardation (FMR) protein is a complex regulator of RNA and synaptic plasticity implicated in fragile X-associated tremor/ataxia syndrome, which features ataxia and increased Fmr1 mRNA expression resulting from epigenetic dysregulation of FMRP. We recently demonstrated that acute ethanol-induced ataxia is associated with increased cerebellar Fmr1 gene expression via histone modifications in rats, but it is unknown whether similar behavioral and molecular changes occur following chronic ethanol exposure. Here, we investigated the effects of chronic ethanol exposure on ataxia and epigenetically regulated changes in Fmr1 expression in the cerebellum. METHODS: Male adult Sprague-Dawley rats were trained on the accelerating rotarod and then fed with chronic ethanol or a control Lieber-DeCarli diet while undergoing periodic behavioral testing for ataxia during ethanol exposure and withdrawal. Cerebellar tissues were analyzed for expression of the Fmr1 gene and its targets using a real-time quantitative polymerase chain reaction assay. The epigenetic regulation of Fmr1 was also investigated using a chromatin immunoprecipitation assay. RESULTS: Ataxic behavior measured by the accelerating rotarod behavioral test developed during chronic ethanol treatment and persisted at both the 8-h and 24-h withdrawal time points compared to control diet-fed rats. In addition, chronic ethanol treatment resulted in up-regulated expression of Fmr1 mRNA and increased activating epigenetic marks H3K27 acetylation and H3K4 trimethylation at 2 sites within the Fmr1 promoter. Finally, measurement of the expression of relevant FMRP mRNA targets in the cerebellum showed that chronic ethanol up-regulated cAMP response element binding (CREB) Creb1, Psd95, Grm5, and Grin2b mRNA expression without altering Grin2a, Eaa1, or histone acetyltransferases CREB binding protein (Cbp) or p300 mRNA transcripts. CONCLUSIONS: These results suggest that epigenetic regulation of Fmr1 and subsequent FMRP regulation of target mRNA transcripts constitute neuroadaptations in the cerebellum that may underlie the persistence of ataxic behavior during chronic ethanol exposure and withdrawal.

Epigenetic mechanisms underlying pathobiology of alcohol use disorder.

PURPOSE OF REVIEW: Chronic alcohol use is a worldwide problem with multifaceted consequences including multiplying medical costs and sequelae, societal effects like drunk driving and assault, and lost economic productivity. These large-scale outcomes are driven by the consumption of ethanol, a small permeable molecule that has myriad effects in the human body, particularly in the liver and brain. In this review, we have summarized effects of acute and chronic alcohol consumption on epigenetic mechanisms that may drive pathobiology of Alcohol Use Disorder (AUD) while identifying areas of need for future research. RECENT FINDINGS: Epigenetics has emerged as an interesting field of biology at the intersection of genetics and the environment, and ethanol in particular has been identified as a potent modulator of the epigenome with various effects on DNA methylation, histone modifications, and non-coding RNAs. These changes alter chromatin dynamics and regulate gene expression that contribute to behavioral and physiological changes leading to the development of AUD psychopathology and cancer pathology. SUMMARY: Evidence and discussion presented here from preclinical results and available translational studies have increased our knowledge of the epigenetic effects of alcohol consumption. These studies have identified targets that can be used to develop better therapies to reduce chronic alcohol abuse and mitigate its societal burden and pathophysiology.