Susan M. Brasser

T1r3 taste receptor involvement in gustatory neural responses to ethanol and oral ethanol preference

Elevated alcohol consumption is associated with enhanced preference for sweet substances across species and may be mediated by oral alcohol-induced activation of neurobiological substrates for sweet taste. Here, we directly examined the contribution of the T1r3 receptor protein, important for sweet taste detection in mammals, to ethanol intake and preference and the neural processing of ethanol taste by measuring behavioral and central neurophysiological responses to oral alcohol in T1r3 receptor-deficient mice and their C57BL/6J background strain. T1r3 knockout and wild-type mice were tested in behavioral preference assays for long-term voluntary intake of a broad concentration range of ethanol, sucrose, and quinine. For neurophysiological experiments, separate groups of mice of each genotype were anesthetized, and taste responses to ethanol and stimuli of different taste qualities were electrophysiologically recorded from gustatory neurons in the nucleus of the solitary tract. Mice lacking the T1r3 receptor were behaviorally indifferent to alcohol (i.e., ∼50% preference values) at concentrations typically preferred by wild-type mice (5-15%). Central neural taste responses to ethanol in T1r3-deficient mice were significantly lower compared with C57BL/6J controls, a strain for which oral ethanol stimulation produced a concentration-dependent activation of sweet-responsive NTS gustatory neurons. An attenuated difference in ethanol preference between knockouts and controls at concentrations >15% indicated that other sensory and/or postingestive effects of ethanol compete with sweet taste input at high concentrations. As expected, T1r3 knockouts exhibited strongly suppressed behavioral and neural taste responses to sweeteners but did not differ from wild-type mice in responses to prototypic salt, acid, or bitter stimuli. These data implicate the T1r3 receptor in the sensory detection and transduction of ethanol taste.

Chemosensory responsiveness to ethanol and its individual sensory components in alcohol-preferring, alcohol-nonpreferring and genetically heterogeneous rats

Alcohol activates orosensory circuits that project to motivationally relevant limbic forebrain areas that control appetite, feeding and drinking. To date, limited data exists regarding the contribution of chemosensory‐derived ethanol reinforcement to ethanol preference and consumption. Measures of taste reactivity to intra‐orally infused ethanol have not found differences in initial orofacial responses to alcohol between alcohol‐preferring (P) and alcohol‐non‐preferring (NP) genetically selected rat lines. Yet, in voluntary intake tests, P rats prefer highly concentrated ethanol upon initial exposure, suggesting an early sensory‐mediated attraction. Here, we directly compared self‐initiated chemosensory responding for alcohol and prototypic sweet, bitter and oral trigeminal stimuli among selectively bred P, NP and non‐selected Wistar (WI) outbred lines to determine whether differential sensory responsiveness to ethanol and its putative sensory components are phenotypically associated with genetically influenced alcohol preference. Rats were tested for immediate short‐term lick responses to alcohol (3–40%), sucrose (0.01–1 M), quinine (0.01–3 mM) and capsaicin (0.003–1 mM) in a brief‐access assay designed to index orosensory‐guided behavior. P rats exhibited elevated short‐term lick responses to both alcohol and sucrose relative to NP and WI lines across a broad range of concentrations of each stimulus and in the absence of blood alcohol levels that would produce significant post‐absorptive effects. There was no consistent relationship between genetically mediated alcohol preference and orosensory avoidance of quinine or capsaicin. These data indicate that enhanced initial chemosensory attraction to ethanol and sweet stimuli are phenotypes associated with genetic alcohol preference and are considered within the framework of downstream activation of oral appetitive reward circuits.

Chronic Voluntary Ethanol Consumption Induces Favorable Ceramide Profiles in Selectively Bred Alcohol-Preferring (P) Rats.

Heavy alcohol consumption has detrimental neurologic effects, inducing widespread neuronal loss in both fetuses and adults. One proposed mechanism of ethanol-induced cell loss with sufficient exposure is an elevation in concentrations of bioactive lipids that mediate apoptosis, including the membrane sphingolipid metabolites ceramide and sphingosine. While these naturally-occurring lipids serve as important modulators of normal neuronal development, elevated levels resulting from various extracellular insults have been implicated in pathological apoptosis of neurons and oligodendrocytes in several neuroinflammatory and neurodegenerative disorders. Prior work has shown that acute administration of ethanol to developing mice increases levels of ceramide in multiple brain regions, hypothesized to be a mediator of fetal alcohol-induced neuronal loss. Elevated ceramide levels have also been implicated in ethanol-mediated neurodegeneration in adult animals and humans. Here, we determined the effect of chronic voluntary ethanol consumption on lipid profiles in brain and peripheral tissues from adult alcohol-preferring (P) rats to further examine alterations in lipid composition as a potential contributor to ethanol-induced cellular damage. P rats were exposed for 13 weeks to a 20% ethanol intermittent-access drinking paradigm (45 ethanol sessions total) or were given access only to water (control). Following the final session, tissues were collected for subsequent chromatographic analysis of lipid content and enzymatic gene expression. Contrary to expectations, ethanol-exposed rats displayed substantial reductions in concentrations of ceramides in forebrain and heart relative to non-exposed controls, and modest but significant decreases in liver cholesterol. qRT-PCR analysis showed a reduction in the expression of sphingolipid delta(4)-desaturase (Degs2), an enzyme involved in de novo ceramide synthesis. These findings indicate that ethanol intake levels achieved by alcohol-preferring P rats as a result of chronic voluntary exposure may have favorable vs. detrimental effects on lipid profiles in this genetic line, consistent with data supporting beneficial cardioprotective and neuroprotective effects of moderate ethanol consumption.

Differential neural representation of oral ethanol by central taste-sensitive neurons in ethanol-preferring and genetically heterogeneous rats

In randomly bred rats, orally applied ethanol stimulates neural substrates for appetitive sweet taste. To study associations between ethanols oral sensory characteristics and genetically mediated ethanol preference, we made electrophysiological recordings of oral responses (spike density) by taste-sensitive nucleus tractus solitarii neurons in anesthetized selectively bred ethanol-preferring (P) rats and their genetically heterogeneous Wistar (W) control strain. Stimuli (25 total) included ethanol [3%, 5%, 10%, 15%, 25%, and 40% (vol/vol)], a sucrose series (0.01, 0.03, 0.1, 0.3, 0.5, and 1 M), and other sweet, salt, acidic, and bitter stimuli; 50 P and 39 W neurons were sampled. k-means clustering applied to the sucrose response series identified cells showing high (S(1)) or relatively low (S(0)) sensitivity to sucrose. A three-way factorial analysis revealed that activity to ethanol was influenced by a neurons sensitivity to sucrose, ethanol concentration, and rat line (P = 0.01). Ethanol produced concentration-dependent responses in S(1) neurons that were larger than those in S(0) cells. Although responses to ethanol by S(1) cells did not differ between lines, neuronal firing rates to ethanol in S(0) cells increased across concentration only in P rats. Correlation and multivariate analyses revealed that ethanol evoked responses in W neurons that were strongly and selectively associated with activity to sweet stimuli, whereas responses to ethanol by P neurons were not easily associated with activity to representative sweet, sodium salt, acidic, or bitter stimuli. These findings show differential central neural representation of oral ethanol between genetically heterogeneous rats and P rats genetically selected to prefer alcohol.

Alcohol sensory processing and its relevance for ingestion.

Alcohol possesses complex sensory attributes that are first detected by the body via sensory receptors and afferent fibers that promptly transmit signals to brain areas involved in mediating ingestive motivation, reinforcement, and addictive behavior. Given that the chemosensory cues accompanying alcohol consumption are among the most intimate, consistent, and immediate predictors of alcohols postabsorptive effects, with experience these stimuli also gain powerful associative incentive value to elicit craving and related physiologic changes, maintenance of ongoing alcohol use, and reinstatement of drug seeking after periods of abstinence. Despite the above, preclinical research has traditionally dichotomized alcohols taste and postingestive influences as independent regulators of motivation to drink. The present review summarizes current evidence regarding alcohols ability to directly activate peripheral and central oral chemosensory circuits, relevance for intake of the drug, and provides a framework for moving beyond a dissociation between the sensory and postabsorptive effects of alcohol to understand their neurobiological integration and significance for alcohol addiction.

Effects of moderate alcohol consumption on gene expression related to colonic inflammation and antioxidant enzymes in rats

Excessive alcohol consumption is a risk factor associated with colorectal cancer; however, some studies have reported that moderate alcohol consumption may not contribute additional risk for developing colorectal cancer while others suggest that moderate alcohol consumption provides a protective effect that reduces colorectal cancer risk. The purpose of this study was to determine the effects of moderate voluntary alcohol (20% ethanol) intake on alternate days for 3 months in outbred Wistar rats on risk factors associated with colorectal cancer development. Colonic gene expression of cyclooxygenase-2, RelA, 8-oxoguanine DNA glycosylase 1, superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, glutathione-S-transferase M1, and aldehyde dehydrogenase 2 were determined. Blood alcohol content, liver function enzyme activities, and 8-oxo-deoxyguanosine DNA adducts were also assessed. Alcohol-treated rats were found to have significantly lower 8-oxo-deoxyguanosine levels in blood, a marker of DNA damage. Alanine aminotransferase and lactate dehydrogenase were both significantly lower in the alcohol group. Moderate alcohol significantly decreased cyclooxygenase-2 gene expression, an inflammatory marker associated with colorectal cancer risk. The alcohol group had significantly increased glutathione-S-transferase M1 expression, an antioxidant enzyme that helps detoxify carcinogens, such as acetaldehyde, and significantly increased aldehyde dehydrogenase 2 expression, which allows for greater acetaldehyde clearance. Increased expression of glutathione-S-transferase M1 and aldehyde dehydrogenase 2 likely contributed to reduce mucosal damage that is caused by acetaldehyde accumulation. These results indicate that moderate alcohol may reduce the risk for colorectal cancer development, which was evidenced by reduced inflammation activity and lower DNA damage after alcohol exposure.

Effects of moderate, voluntary ethanol consumption on the rat and human gut microbiome: Effects of ethanol consumption

Many alcohol‐induced health complications are directly attributable to the toxicity of alcohol or its metabolites, but another potential health impact of alcohol may be on the microbial communities of the human gut. Clear distinctions between healthy and diseased‐state gut microbiota have been observed in subjects with metabolic diseases, and recent studies suggest that chronic alcoholism is linked to gut microbiome dysbiosis. Here, we investigated the effects of moderate levels of alcohol consumption on the gut microbiome in both rats and humans. The gut microbiota of rats voluntarily consuming a 20 percent ethanol solution, on alternate days, were compared with a non‐exposed control group to identify differential taxonomic and functional profiles. Gut microbial diversity profiles were determined using culture‐independent amplification, next‐generation sequencing and bioinformatic analysis of bacterial 16S ribosomal RNA gene sequence libraries. Our results showed that, compared with controls, ethanol‐consuming rats experienced a significant decline in the biodiversity of their gut microbiomes, a state generally associated with dysbiosis. We also observed significant shifts in the overall diversity of the gut microbial communities and a dramatic change in the relative abundance of particular microbes, such as the Lactobacilli. We also compared our results to human fecal microbiome data collected as part of the citizen science American Gut Project. In contrast to the rat data, human drinkers had significantly higher gut microbial biodiversity than non‐drinkers. However, we also observed that microbes that differed among the human subjects displayed similar trends in the rat model, including bacteria implicated in metabolic disease.