Alterations in ethanol-induced behaviors and consumption in knock-in mice expressing ethanol-resistant NMDA receptors

Ethanol's action on the brain likely reflects altered function of key ion channels such as glutamatergic N-methyl-D-aspartate receptors (NMDARs). In this study, we determined how expression of a mutant GluN1 subunit (F639A) that reduces ethanol inhibition of NMDARs affects ethanol-induced behaviors in mice. Mice homozygous for the F639A allele died prematurely while heterozygous knock-in mice grew and bred normally. Ethanol (44 mM; ∼0.2 g/dl) significantly inhibited NMDA-mediated EPSCs in wild-type mice but had little effect on responses in knock-in mice. Knock-in mice had normal expression of GluN1 and GluN2B protein across different brain regions and a small reduction in levels of GluN2A in medial prefrontal cortex. Ethanol (0.75-2.0 g/kg; IP) increased locomotor activity in wild-type mice but had no effect on knock-in mice while MK-801 enhanced activity to the same extent in both groups. Ethanol (2.0 g/kg) reduced rotarod performance equally in both groups but knock-in mice recovered faster following a higher dose (2.5 g/kg). In the elevated zero maze, knock-in mice had a blunted anxiolytic response to ethanol (1.25 g/kg) as compared to wild-type animals. No differences were noted between wild-type and knock-in mice for ethanol-induced loss of righting reflex, sleep time, hypothermia or ethanol metabolism. Knock-in mice consumed less ethanol than wild-type mice during daily limited-access sessions but drank more in an intermittent 24 h access paradigm with no change in taste reactivity or conditioned taste aversion. Overall, these data support the hypothesis that NMDA receptors are important in regulating a specific constellation of effects following exposure to ethanol. © 2013 den Hartog et al

AGE Metabolites: A Biomarker Linked to Cancer Disparity?

Abstract Socioeconomic and environmental influences are established factors promoting cancer disparity, but the contribution of biologic factors is not clear. We report a mechanistic link between carbohydrate-derived metabolites and cancer that may provide a biologic consequence of established factors of cancer disparity. Glycation is the nonenzymatic glycosylation of carbohydrates to macromolecules, which produces reactive metabolites called advanced glycation end products (AGE). A sedentary lifestyle and poor diet all promote disease and the AGE accumulation pool in our bodies and also increase cancer risk. We examined AGE metabolites in clinical specimens of African American and European American patients with prostate cancer and found a higher AGE concentration in these specimens among African American patients when compared with European American patients. Elevated AGE levels corresponded with expression of the receptor for AGE (RAGE or AGER). We show that AGE-mediated increases in cancer-associated processes are dependent upon RAGE. Aberrant AGE accumulation may represent a metabolic susceptibility difference that contributes to cancer disparity. Cancer Epidemiol Biomarkers Prev; 23(10); 2186–91. ©2014 AACR.</jats:p

F639A Het mice consume more of a sweetened ethanol solution than wild-type mice in long-access drinking paradigm.

<p>(<i>A</i>), Ethanol intake (mean ±SEM) in wild-type and F639A Het mice with intermittent 24 h access to sweetened ethanol or water (n = 10–11 for each group). Ethanol concentrations were ramped from 3–20% (v/v) and all concentrations also contained 0.2% saccharin (w/v). Symbol (<b>*</b>): indicates main effect of genotype (<b>*</b><i>p</i><0.05, mixed ANOVA). (<i>B</i>), Percent preference for sweetened ethanol solution over water. Symbol (<b>*</b>): indicates main effect of genotype (<b>***</b><i>p</i><0.001, mixed ANOVA). Values are mean ±SEM. (<i>C</i>), Total water intake (mean ±SEM) during ‘off’ drinking days in which mice received 2 bottles containing water.</p

F639A Het mice show altered conditioned taste aversion to a low dose of ethanol as compared to wild-type mice.

<p>Graphs show percent of baseline saccharin solution consumed after repeated pairings with an injection of saline, 1.25/kg, 1.75 g/kg, or 2.5 g/kg of ethanol in (<i>A</i>) wild-type, and (<i>B</i>) F639A Het mice (n = 6–7 for each group). Symbol (<b>*</b>): value significantly different from saline (<b>***</b><i>p</i><0.001, two-way RM ANOVA, Bonferroni's <i>post hoc</i> test). Values are mean ±SEM.</p

Expression of NMDA receptor subunits in wild-type and F639A Het mice (n = 4–5 for each group).

<p>Panels show immunoblot analysis of GluN1 (<i>A</i>), GluN2A (<i>B</i>), and GluN2B (<i>C</i>) from crude membrane fractions prepared from select brain regions. Data are percent of wild-type control (mean ±SEM). Abbreviations: <i>mPFC</i>, medial pre-frontal cortex; <i>DS</i>, dorsal striatum; <i>HC</i>, hippocampus; <i>AMY</i>, amygdala; and <i>NAcc</i>, nucleus accumbens. Symbol (<b>*</b>): value significantly different from control (<b>**</b><i>p</i><0.01, unpaired <i>t</i>-test).</p

Anxiolytic response to ethanol is blunted in F639A Het mice.

<p>(<i>A</i>), Percent of time (mean ±SEM) spent in the open arms of an elevated zero maze following injection of saline or 1.25 g/kg ethanol in F639A Het and wildtype mice (n = 10 for each group). Symbol (<b>*</b>): value significantly different from saline (<b>**</b> p<0.01, two-way ANOVA, Bonferroni's <i>post hoc</i> test). (<i>B</i>), Total distance traveled and (<i>C</i>), total number of arm entries on the elevated zero maze. Symbol (<b>*</b>): value significantly different from saline (<b>*</b><i>p</i><0.05, two-way ANOVA, Bonferroni's <i>post hoc</i> test). Values are mean ±SEM.</p

F639A Het and wild-type mice do not differ in taste reactivity.

<p>Consumption in wild-type and F639A Het mice was measured using a two-bottle choice test with 24 h continuous access to tastants (n = 7 for each group). Left panels show preference ratio for volume of tastant solution consumed over water measured on the 4^th day of access for (<i>A</i>) saccharin, (<i>B</i>) sucrose, and (<i>C</i>) quinine. Right panels show corresponding volumes consumed across days for each tastant. Values are mean ±SEM.</p

F639A Het mice show altered ethanol consumption than wild-type mice in short-access and long-access drinking paradigms.

<p>(<i>A</i>), Ethanol intake (mean ±SEM) in wild-type and F639A Het mice during 2 h limited-access to 15% (v/v) ethanol or water (n = 8 for each group). Symbol (*): indicates main effect of genotype (* <i>p</i><0.05, mixed ANOVA). (<i>B</i>), Ethanol intake (mean ±SEM) in a limited-access DID model in wild-type and F639A Het mice. Mice had access to one bottle containing 20% (v/v) ethanol 3 h into their dark cycle for 2 h and 4 h sessions (n = 11–12 for each group). Dotted lines indicate 4 h sessions. (<i>C</i>), Ethanol intake (mean ±SEM) in wild-type and F639A Het mice during intermittent 24 h access to ethanol or water (n = 10–11 for each group). Ethanol concentrations were ramped from 3, 6, 10% and maintained at 20% (v/v) ethanol. Symbol (*): indicates main effect of genotype (* <i>p</i><0.05, mixed ANOVA). (<i>D</i>), Percent preference for ethanol solution over water-bottle choice in a subset of animals from intermittent access study (n = 6 from each group). Symbol (*): indicates main effect of genotype (* <i>p</i><0.05, mixed ANOVA). Values are mean ±SEM.</p

Hypnotic and hypothermic effects of high doses of ethanol.

<p>Latency to lose righting reflex (LORR; <i>A</i>) and duration of LORR (<i>B</i>) following a 4.0 g/kg injection of ethanol in wild-type and F639A Het mice (n = 17–18 for each group). Values are mean ±SEM. (<i>C</i>), Change in body temperature following a 3.5 g/kg injection of ethanol in wild-type and F639A Het mice (n = 10 for each group). Values are mean ±SEM. (<i>D</i>), Rate of blood ethanol metabolism between wild-type and F639A Het mice. Blood ethanol concentration (mean ±SEM) measured over time following injection with 4.0 g/kg of ethanol (n = 7 for each group).</p