ß-arrestin-biased Allosteric Modulator of Neurotensin Receptor 1 Reduces Ethanol Drinking and Responses to Ethanol Administration in Rodents.

Alcohol use disorders (AUDs) impose an enormous societal and financial burden, and world-wide, alcohol misuse is the 7th leading cause of premature death1. Despite this, there are currently only 3 FDA approved pharmacological treatments for the treatment of AUDs in the United States. The neurotensin (Nts) system has long been implicated in modulating behaviors associated with alcohol misuse. Recently, a novel compound, SBI-553, that biases the action of Nts receptor 1 (NTSR1) activation, has shown promise in preclinical models of psychostimulant misuse. Here we investigate the efficacy of this compound to alter ethanol-mediated behaviors in a comprehensive battery of experiments assessing ethanol consumption, behavioral responses to ethanol, sensitivity to ethanol, and ethanol metabolism. Additionally, we investigated behavior in avoidance and cognitive assays to monitor potential side effects of SBI-553. We find that SBI-553 reduces binge-like ethanol consumption in mice without altering avoidance behavior or novel object recognition. We also observe sex-dependent differences in physiological responses to sequential ethanol injections in mice. In rats, we show that SBI-553 attenuates sensitivity to the interoceptive effects of ethanol (using a Pavlovian drug discrimination task). Our data suggest that targeting NTSR1 signaling may be promising to attenuate alcohol misuse, and adds to a body of literature that suggests NTSR1 may be a common downstream target involved in the psychoactive effects of multiple reinforcing substances.

Lateral habenula-projecting central amygdala circuits expressing GABA and NPY Y1 receptor modulate binge-like ethanol intake in mice.

The central nucleus of the amygdala (CeA) is a critical brain region in the integration of emotional behaviors and is one of the major output areas of the amygdaloid complex. The CeA is composed of GABAergic interneurons and projection neurons which co-express a range of peptides including neuropeptide Y (NPY). Importantly, GABA and NPY signaling, via the NPY Y1 receptor (Y1R), in the CeA modulate binge-like ethanol intake in rodents and these systems undergo neuroplastic alterations following a history of ethanol consumption. Here we assessed the roles of GABAergic and Y1R+ circuits arising from the CeA and innervating the lateral habenula (LHb), a brain region that modulates the aversive properties of ethanol, in modulating binge-like ethanol intake in mice using "drinking in the dark" (DID) procedures. Using an anterograde cre-inducible reporter virus we established the CeA → LHb circuit in male and female vgat-ires-cre and NPY1r-cre mice. Next, we found that chemogenetic silencing of both the GABAergic or Y1R+ CeA → LHb circuit significantly blunted binge-like intake of a 20% ethanol solution but this same procedure failed to alter the consumption of a 3% sucrose solution. Finally, one, 4-day cycle of DID failed to alter basal or effects of ethanol or NPY on inhibitory transmission in Y1R+ CeA → LHb neurons. The present results suggest that blunting GABAergic tone in LHb-projecting CeA neurons may represent a new approach to preventing the development of AUDs. Drugs that target NPY Y1Rs are potential attractive targets.

Chemogenetic inhibition of corticotropin-releasing factor neurons in the central amygdala alters binge-like ethanol consumption in male mice.

Repetitive bouts of binge drinking can lead to neuroplastic events that alter ethanol's pharmacologic effects and perpetuate excessive consumption. The corticotropin-releasing factor (CRF) system is an example of ethanol-induced neuroadaptations that drive excessive ethanol consumption. Our laboratory has previously shown that CRF antagonist, when infused into the central amygdala (CeA), reduces binge-like ethanol consumption. The present study extends this research by assessing the effects of silencing CRF-producing neurons in CeA on binge-like ethanol drinking stemming from "Drinking in the Dark" (DID) procedures. CRF-ires-Cre mice underwent surgery to infuse Gi/o-coupled Designer Receptors Exclusively Activated by Designer Drugs (DREADD) virus or a control virus into either the CeA or basolateral amygdala (BLA). Gi/o-DREADD-induced CRF-neuronal inhibition in the CeA resulted in a 33% decrease in binge-like ethanol consumption. However, no effect on ethanol consumption was seen after DREADD manipulation in the BLA. Moreover, CeA CRF-neuronal inhibition had no effect on sucrose consumption. The effects of silencing CRF neurons in the CeA on ethanol consumption are not secondary to changes in motor function or anxiety-like behaviors as assessed in the open-field test (OFT). Finally, the DREADD construct's functional ability to inhibit CRF-neuronal activity was demonstrated by reduced ethanol-induced c-Fos following DREADD activation. Together, these data suggest that the CRF neurons in the CeA play an important role in binge ethanol consumption and that inhibition of the CRF-signaling pathway remains a viable target for manipulating binge-like ethanol consumption. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Distinct and Overlapping Patterns of Acute Ethanol-Induced C-Fos Activation in Two Inbred Replicate Lines of Mice Selected for Drinking to High Blood Ethanol Concentrations.

The inbred high drinking in the dark (iHDID1 and iHDID2) strains are two replicate lines bred from the parent HS/Npt (HS) line for achieving binge levels of blood ethanol concentration (≥80 mg/dL BEC) in a four-hour period. In this work, we sought to evaluate differences in baseline and ethanol-induced c-Fos activation between the HS, iHDID1, and iHDID2 genetic lines in brain regions known to process the aversive properties of ethanol. METHODS: Male and female HS, iHDID1, and iHDID2 mice underwent an IP saline 2 3 g/kg ethanol injection. Brain sections were then stained for c-Fos expression in the basolateral/central amygdala (BLA/CeA), bed nucleus of the stria terminals (BNST), A2, locus coeruleus (LC), parabrachial nucleus (PBN), lateral/medial habenula (LHb/MHb), paraventricular nucleus of the thalamus (PVT), periaqueductal gray (PAG), Edinger-Westphal nuclei (EW), and rostromedial tegmental nucleus (RMTg). RESULTS: The iHDID1 and iHDID2 lines showed similar and distinct patterns of regional c-Fos; however, in no region did the two both significantly differ from the HS line together. CONCLUSIONS: These data lend further support to altered baseline or ethanol-induced activation in brain regions associated with processing the aversive properties of ethanol in the iHDID1 and iHDID2 genetic lines.

Assessment of ventral tegmental area-projecting GABAergic neurons from the bed nucleus of the stria terminalis in modulating binge-like ethanol intake.

Corticotropin-releasing factor (CRF) circuitry is a key component in plasticity underlying the transition to ethanol (EtOH) dependence. We have previously shown that chemogenetic silencing of CRF neurons stemming from the dorsolateral bed nucleus of the stria terminalis (dlBNST) and projecting to the ventral tegmental area (VTA) significantly blunts binge-like EtOH consumption. While CRF neurons in the BNST are thought to entail primarily a GABA phenotype, glutamatergic neurons within the BNST also innervate the VTA and influence consummatory behaviors. Here, we combined the well-validated Vgat-ires-Cre transgenic mice with chemogenetic tools to extend our previous findings and corroborate the contribution of the VTA-projecting dlBNST GABAergic circuitry in modulating binge-like EtOH consumption using "drinking-in-the-dark" procedures. Mice were given bilateral injection of Gi-coupled chemogenetic viral vector (or control virus) into the dlBNST and bilateral cannulae into the VTA. On test day, clozapine-N-oxide (CNO; or vehicle) was infused directly into the VTA to silence VTA-projecting dlBNST neurons and subsequent binge-like EtOH consumption was assessed. We then used immunohistochemistry (IHC) to determine the co-expression of CRF and viral vector. Our results showed that relative to vehicle treatment or CNO treatment in mice expressing the control virus, silencing VTA-projecting dlBNST GABAergic neurons by CNO treatment in mice expressing Gi-coupled chemogenetic virus significantly reduced binge-like EtOH intake. This effect was not seen with sucrose consumption. Our IHC results confirm a population of CRF-expressing GABAergic neurons within the dlBNST. This study directly establishes that VTA-projecting GABAergic neurons of the dlBNST modulate binge-like EtOH consumption.