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Childhood stress and trauma are associated with substance use disorders in adulthood, but the neurological changes that confer increased vulnerability are largely unknown. In this study, maternal separation [MS] stress, restricted to the pre-weaning period, was used as a model to study mechanisms of protracted effects of childhood stress/traumatic experiences on binge drinking and impulsivity. Using an operant self-administration model of binge drinking and a delay discounting assay to measure impulsive-like behavior, we report that early life stress due to MS facilitated acquisition of binge drinking and impulsivity during adulthood in rats. Previous studies have shown heightened levels of corticotropin releasing factor (CRF) after MS, and here, we add that MS increased expression levels of GABAA α2 subunit in central stress circuits. To investigate the precise role of these circuits in regulating impulsivity and binge drinking, the CRF1 receptor antagonist antalarmin and the novel GABAA α2 subunit ligand 3-PBC were infused into the central amygdala [CeA] and medial prefrontal cortex [mPFC]. Antalarmin and 3-PBC at each site markedly reduced impulsivity and produced profound reductions on binge-motivated alcohol drinking, without altering responding for sucrose. Furthermore, whole-cell patch-clamp studies showed that low concentrations of 3-PBC directly reversed the effect of relatively high concentrations of ethanol on α2β3γ2 GABAA receptors, by a benzodiazepine site-independent mechanism. Together, our data provide strong evidence that maternal separation, i.e., early life stress, is a risk factor for binge drinking, and is linked to impulsivity, another key risk factor for excessive alcohol drinking. We further show that pharmacological manipulation of CRF and GABA receptor signaling is effective to reverse binge drinking and impulsive-like behavior in MS rats. These results provide novel insights into the role of the brain stress systems in the development of impulsivity and excessive alcohol consumption.

The enteric nervous system (ENS) orchestrates a broad range of important gastrointestinal functions such as intestinal motility and gastric secretion. The ENS can be affected by environmental factors, diet and disease. Changes due to these alterations are often hard to evaluate in detail when whole gut samples are used. Analyses based on pure ENS tissue can more effectively reflect the ongoing changes during pathological processes. Here, we present an optimized approach for the isolation of pure myenteric plexus (MP) from adult mouse and human. To do so, muscle tissue was individually digested with a purified collagenase. After incubation and a gentle mechanical disruption step, MP networks could be collected with anatomical integrity. These tissues could be stored and used either for immediate genomic, proteomic or in vitro approaches, and enteric neurospheres could be generated and differentiated. In a pilot experiment, the influence of bacterial lipopolysaccharide on human MP was analyzed using 2-dimensional gel electrophoresis. The method also allows investigation of factors that are secreted by myenteric tissue in vitro. The isolation of pure MP in large amounts allows new analytical approaches that can provide a new perspective in evaluating changes of the ENS in experimental models, human disease and aging.

The behavioral and functional significance of the extrasynaptic inhibitory GABAA receptors in the brain is still poorly known. We used a transgenic mouse line expressing the GABAA receptor α6 subunit gene in the forebrain under the Thy‐1.2 promoter (Thy1α6) mice ectopically expressing α6 subunits especially in the hippocampus to study how extrasynaptically enriched αβ(γ2)‐type receptors alter animal behavior and receptor responses. In these mice extrasynaptic α6β receptors make up about 10% of the hippocampal GABAA receptors resulting in imbalance between synaptic and extrasynaptic inhibition. The synthetic GABA‐site competitive agonist gaboxadol (4,5,6,7‐tetrahydroisoxazolo[5,4‐c]pyridin‐3‐ol; 3 mg/kg) induced remarkable anxiolytic‐like response in the light : dark exploration and elevated plus‐maze tests in Thy1α6 mice, while being almost inactive in wild‐type mice. The transgenic mice also lost quicker and for longer time their righting reflex after 25 mg/kg gaboxadol than wild‐type mice. In hippocampal sections of Thy1α6 mice, the α6β receptors could be visualized autoradiographically by interactions between gaboxadol and GABA via [35S]TBPS binding to the GABAA receptor ionophore. Gaboxadol inhibition of the binding could be partially prevented by GABA. Electrophysiology of recombinant GABAA receptors revealed that GABA was a partial agonist at α6β3 and α6β3δ receptors, but a full agonist at α6β3γ2 receptors when compared with gaboxadol. The results suggest strong behavioral effects via selective pharmacological activation of enriched extrasynaptic αβ GABAA receptors, and the mouse model represents an example of the functional consequences of altered balance between extrasynaptic and synaptic inhibition.

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