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Unveiling the alcohol-dependent alterations in local dendritic translation of mRNAs in the prefrontal cortex during adolescence

Periodic Reporting for period 1 - ALCO-ADO (Unveiling the alcohol-dependent alterations in local dendritic translation of mRNAs in the prefrontal cortex during adolescence)

Reporting period: 2019-04-01 to 2021-03-31

Alcohol use disorder (AUD) is a devastating relapsing disease which represents the fourth leading cause of preventable death worldwide. AUD has been considered as a pathological condition that develops in adulthood, but emerging evidence suggest that the roots of alcohol addiction begin to grow earlier in life, precisely during adolescence. Indeed, studies have suggested that heavy alcohol intake perturbs the maturation of the adolescent brain, inducing neuroadaptations that in turn increase the risk for developing AUD and comorbid psychopathology later in life. In addition, the historic gender gap in AUD, characterized by a greater prevalence in men, has been narrowing over the past years, highlighting the importance of understanding sex differences in the biology and treatments of AUD. However, the current literature on sex differences in the etiology of AUD is still very limited.
The long-term objectives of this research are (i) to unveil how alcohol consumption during adolescence alters brain maturation and leads to long-lasting behavioral impairments, (ii) to reveal how alcohol differentially impacts male and female brain maturation; (iii) to identify new mechanisms and targets of alcohol’s actions in order to develop new therapies for treating AUD.

Conclusions of the action
We showed that voluntary alcohol binge-drinking during adolescence induces delayed appearance of behavioral defects in both sexes (Van Hees et al, under review, https://doi.org/10.1101/2020.08.11.245878). Moreover, our preliminary results suggest that AAE may modulate the activity of local translation regulators and interferes with PFC maturation.
WP1: Uncovering the structural, physiological and behavioral defects induced by AAE in the PFC
Van Hees et al, https://doi.org/10.1101/2020.08.11.245878. Our findings are of great importance regarding the major public health issue that is adolescent binge-drinking, and could help refining the prevention strategies against harmful alcohol use in youth.
Regarding the functional studies, our preliminary results suggest that AAE significantly increases the frequency of sEPSCs in layer V neurons of the prelimbic cortex (Figure 1). Morphological studies are currently being performed. Active and passive properties of PFC neurons as well as functional synaptic transmission will also be assessed.
WP2: Deciphering whether AAE modulates the activity of translation regulators in PFC neurons
Our preliminary results suggest that AAE induces the long-lasting activation of mTORC1 in the PFC of male and female mice (Figure 2), which may be restricted to Ctip2+ layer V neurons (Figure 3). We will test whether modulation of mTORC1 activity reverses the behavioral deficits by using the FLEX constructs, which have been validated.
WP3: Unveiling the alcohol-induced alterations in local dendritic translation occurring in specific PFC neuronal populations
Ribosome-associated mRNAs were purified from projection neurons (Ribotag x VGlut-Cre mice). The specificity of HA-expression was validated (Figure 4). Samples are currently being processed by Diagenode sprl (Liège, Belgium). The results will show the alcohol-induced alterations in local dendritic translation in projection neurons of the PFC. Candidate synaptic mRNAs will be validated by quantitative RT-PCR after Ribotag immunoprecipitation/translating mRNA extraction, as well as after polysomal RNA extraction from purified synaptic fractions. In addition, the involvement of mTORC1 in the translational control of specific mRNA will be assessed.

Dissemination
- Scientific publication: https://doi.org/10.1101/2020.08.11.245878
- Oral presentation: European Society for Biomedical Research on Alcoholism (ESBRA) meeting 2019, Lille, France.
- Poster presentation: BSCDB Meeting 2019, Liege, Belgium.
- Seminars in schools, in collaboration with Rejouisciences and ULiège. (Will be resumed in September 2021 according to the sanitary situation.
- Participation in “Binôme – Dislocation Cervicale”, original theater play inspired from scientific research..(https://www.news.uliege.be/cms/c_11061658/fr/binome-dislocation-cervicale-avignon; https://www.liegecreative.be/evenements/binome-le-poete-et-le-savant; https://vimeo.com/359769867).
- Television/radio Interviews:
o http://www.vivreici.be/videos/detail_sophie-laguesse-n-abimez-pas-votre-cerveau-pour-quelques-verres-d-alcool-en-trop?videoId=1138436)

o https://www.rtbf.be/auvio/detail_tendances-premiere-le-dossier?id=2602931)

- Public Conferences : Doc’s Café, Lion’s Club Liège Airport February 2020, Lion’s club Spa December 2019, Maria Goretti October 2019, Fondation Leon Fredericq academic meeting November 2019.
Alcohol is the most consumed drug among adolescents, with 40% of them reporting regularly experiencing binge-drinking episodes, consuming more than 5 drinks in two hours and reaching blood alcohol concentrations above 0.08g/dl. This pattern of alcohol consumption is particularly harmful as it may interfere with the ongoing maturation of frontal brain circuits, and clinical studies have shown that AAE significantly increases the risk of developing psychiatric and behavioral disorders later in life, including ddiction. However, the precise cellular mechanisms underlying behavioral deficits, the molecular mechanisms underlying alcohol-induced defects in PFC maturation, and possible gender differences are still poorly understood.

We developed a mouse model of voluntary alcohol binge-drinking during adolescence, and reported that AAE induces the progressive development of behavioral impairments related to PFC malfunction. Although differences were noted between males and females, AAE similarly impacted male and female behaviors. Importantly, no major behavioral impairments was observed short-term after AAE, suggesting that AAE induces a progressive development of behavioral defects, which only emerge in adulthood. Our findings are of great importance regarding the major public health issue that is adolescent binge-drinking, and could help refining the prevention strategies against harmful alcohol use in youth. All data are available in a Mendeley dataset DOI: 10.17632/gtnmrbtmt4.1.

It has been shown in adult mice that excessive alcohol consumption modifies synaptic protein composition in brain regions associated with the mesocorticolimbic pathway, promoting the development and maintenance of alcohol addiction. The mammalian target of rapamycin complex 1 (mTORC1) and the eukaryotic initiation factor 2α (eIF2α) are master regulators of local translation. However, the alcohol- dependent modulation of mTORC1 and eIF2α activity in the maturating PFC and the consequences of alcohol-induced defects in local translation have remained unknown.

Our preliminary results suggest that AAE activates mTORC1 but not eIF2a activity specifically in the prelimbic and infralimbic regions of the PFC. Furthermore, we observed that the AAE-dependent increased mTORC1 activity is mostly present in the Ctip2+ layer V neurons. We are investigating whether modulation of mTORC1 activity in the PFC projection neurons rescues the AAE-induced behavioral defects.

We performed a Ribotag-RNA-seq experiment to specifically analyze the “translatome” of PFC projection neurons in AAE and water-exposed P43 animals. We are currently performing the bioinformatics analysis in order to identify synaptic RNA candidates whose translation is modulated by AAE in the PFC.
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