Predisposition to Alcohol Drinking and Alcohol Consumption Alter Expression of Calcitonin Gene-Related Peptide, Neuropeptide Y, and Microglia in Bed Nucleus of Stria Terminalis in a Subnucleus-Specific Manner.

Ilaria Rossetti,Stefano Morara,Roberta Sau,Luciano Provini,M Paola Castelli,Laura Zambusi,Krzysztof Gonciarz,Paola Maccioni,Giancarlo Colombo

doi:10.3389/fncel.2019.00158

Abstract

Excessive alcohol consumption is often linked to anxiety states and has a major relay center in the anterior part of bed nucleus of stria terminalis (BNST). We analyzed the impact of (i) genetic predisposition to high alcohol preference and consumption, and (ii) alcohol intake on anterior BNST, namely anterolateral (AL), anteromedial (AM), and anteroventral (lateral + medial subdivisions: AVl, AVm) subnuclei. We used two rat lines selectively bred for low- and high-alcohol preference and consumption, named Sardinian alcohol-non preferring (sNP) and -preferring (sP), respectively, the latter showing also inherent anxiety-related behaviors. We analyzed the modulation of calcitonin gene-related peptide (CGRP; exerting anxiogenic effects in BNST), neuropeptide Y (NPY; exerting mainly anxiolytic effects), and microglia activation (neuroinflammation marker, thought to increase anxiety). Calcitonin gene-related peptide-immunofluorescent fibers/terminals did not differ between alcohol-naive sP and sNP rats. Fiber/terminal NPY-immunofluorescent intensity was lower in BNST-AM and BNST-AVm of alcohol-naive sP rats. Activation of microglia (revealed by morphological analysis) was decreased in BNST-AM and increased in BNST-AVm of alcohol-naive sP rats. Prolonged (30 consecutive days), voluntary alcohol intake under the homecage 2-bottle “alcohol vs. water” regimen strongly increased CGRP intensity in BNST of sP rats in a subnucleus-specific manner: in BNST-AL, BNST-AVm, and BNST-AM. CGRP area sum, however, decreased in BNST-AM, without changes in other subnuclei. Alcohol consumption increased NPY expression, in a subnucleus-specific manner, in BNST-AL, BNST-AVl, and BNST-AVm. Alcohol consumption increased many size/shapes parameters in microglial cells, indicative of microglia de-activation. Finally, microglia density was increased in ventral anterior BNST (BNST-AVl, BNST-AVm) by alcohol consumption. In conclusion, genetic predisposition of sP rats to high alcohol intake could be in part mediated by anterior BNST subnuclei showing lower NPY expression and differential microglia activation. Alcohol intake in sP rats produced complex subnucleus-specific changes in BNST, affecting CGRP/NPY expression and microglia and leading to hypothesize that these changes might contribute to the anxiolytic effects of voluntarily consumed alcohol repeatedly observed in sP rats.

Highlights

Excessive alcohol consumption poses a serious threat to human health
An overall analysis of the three groups was carried out: this showed a significant difference in Calcitonin gene-related peptide (CGRP) content among the three groups in bed nucleus of stria terminalis (BNST)-AL (CGRP intensity: F = 27.02, p < 0.0001; CGRP area sum: F = 0.8413, p = 0.4435), BNSTAM (CGRP intensity: F = 54.96, p < 0.0001; CGRP area sum: H = 14.11, p = 0.0009) and BNST-AVm (CGRP intensity: F = 11.84, p = 0.0002; CGRP area sum: H = 9.190, p = 0.0101), but not in BNST-AVl (CGRP intensity: H = 1.071, p = 0.5854; CGRP area sum: H = 1.554, p = 0.4597)
CGRP immunofluorescence area sum showed a decrease in BNST-AM of alcohol-naive Sardinian alcohol-preferring (sP) rats, whereas only a non-significant tendency to decrease or no change was detected in BNST-AL, BNST-AVl, and BNST-AVm of alcohol-naive sP rats (Figure 2)

Summary

Introduction

Excessive alcohol consumption poses a serious threat to human health. This can arise, for example, from increased risk of cancer or cardiovascular diseases (Kunzmann et al, 2018; Wood et al, 2018). Since youth represents a critical period in brain development that is vulnerable to alcohol misuse, binge/heavy-drinking during adolescence and in young adults carries major long-term consequences. Binge and heavy-drinking adolescents and young adults have altered neural structure and activity which could result in neural reorganization and increased risk for developing alcohol use disorder (Cservenka and Brumback, 2017) and can develop longterm cognitive and psychological consequences (Carbia et al, 2018). Alcohol drinking deserves careful analysis and has been the focus of several studies to identify brain areas and mechanisms driving consumption motivations

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