The Cortical Neuroimmune Regulator TANK Affects Emotional Processing and Enhances Alcohol Drinking: A Translational Study

Abstract

Alcohol abuse is a major public health problem worldwide. Understanding the molecular mechanisms that control regular drinking may help to reduce hazards of alcohol consumption. While immunological mechanisms have been related to alcohol drinking, most studies reported changes in immune function that are secondary to alcohol use. In this report, we analyse how the gene "TRAF family member-associated NF-κB activator" (TANK) affects alcohol drinking behavior. Based on our recent discovery in a large GWAS dataset that suggested an association of TANK, SNP rs197273, with alcohol drinking, we report that SNP rs197273 in TANK is associated both with gene expression (P = 1.16 × 10-19) and regional methylation (P = 5.90 × 10-25). A tank knock out mouse model suggests a role of TANK in alcohol drinking, anxiety-related behavior, as well as alcohol exposure induced activation of insular cortex NF-κB. Functional and structural neuroimaging studies among up to 1896 adolescents reveal that TANK is involved in the control of brain activity in areas of aversive interoceptive processing, including the insular cortex, but not in areas related to reinforcement, reward processing or impulsiveness. Our findings suggest that the cortical neuroimmune regulator TANK is associated with enhanced aversive emotional processing that better protects from the establishment of alcohol drinking behavior.

Keywords: NF-κB; TANK; alcohol; anxiety; drinking; insular cortex.

Figure 1.

Schematic diagrams of the TANK gene structure. (a) Possible mechanisms linking the TANK single-nucleotide polymorphism rs197273 to TANK gene expression through DNA methylation in the (b) Framingham and (c) in the IMAGEN study.

Figure 2.

TANK is required to establish normal alcohol drinking in mice. tank KO (n = 11) and wild-type (WT; n = 12) mice were tested in a free-choice 2-bottle drinking paradigm for their alcohol consumption. (a) Amount of alcohol consumed at different concentrations of the drinking fluid. (b) Preference of alcohol versus water. (c) Total fluid consumption during testing (pre-planned Bonferroni-corrected LSD test; *P < 0.05, **P < 0.01; vs. WT). (d) Consumption of 16 vol.% alcohol after 2 weeks of drinking and alcohol deprivation effect (ADE) in tank KO (n = 11) and WT (n = 12) mice. After continuous drinking, animals were withdrawn from alcohol for 2 times 3 weeks (dotted lines) and reinstated (R) for 4 days, respectively, (e) alcohol preference and (f) total fluid consumption during ADE (pre-planned Bonferroni-corrected LSD test; *P < 0.05, ^**P < 0.01; ***P < 0.001 vs. baseline (Bl)).

Figure 3.

TANK controls preference for sweet taste in a 2-bottle free-choice paradigm in tank KO (n = 11) and wild-type (WT; n = 12) mice. (a) Sucrose (sweet) preference and quinine (bitter) avoidance test in a free-choice 2-bottle drinking paradigm indicates no difference between tank KO (n = 11) and WT mice (n = 12) in the avoidance of bitter tasting quinine, but a reduced preference for sucrose (pre-planned Bonferroni-corrected LSD test; ^##P < 0.01). (b) A saccharine preference test in a free-choice 2-bottle drinking paradigm indicates a reduced preference for sweet tasting, but caloric neutral saccharin in naïve tank KO mice (n = 12) versus WT (n = 12) (pre-planned Bonferroni-corrected LSD test; ^#P < 0.05). (c) TANK has no effect on alcohol bioavailability in mice. Blood alcohol concentration in tank KO (n = 10) and WT mice (n = 10) after alcohol injection (3.5 g/kg, i.p.). Over the 3-h tested, there was no difference in alcohol bioavailability between genotypes (P > 0.05).

Figure 4.

Association of whole-brain activity during social emotional processing with the tank gene. Brain regions associated with (a/c) TANK rs197273 and (b/d) TANK gene score during angry faces task. Red and blue colors represent a positive association and a negative association, respectively.

Figure 5.

TANK controls anxiety-related behavior in mice. Tank KO mice display more anxiety-related behavior in a novel environment than wild-type (WT) mice. Enhanced anxiety in the elevated plus maze test (EPM) by tank KO (n = 12) vs. WT (n = 11) mice shown by (a) reduced time spent and (b) less distance moved on open arms (P < 0.05) and (c) less relative time in open arms. (d) No effect on depression-related behavior in the novelty-suppressed feeding test in tank KO mice. (e) In the open field (OF) test, tank KO mice (n = 12) spend less time in the anxiogenic center of the maze than WT mice (n = 11). (f) The number of OF center entries is reduced in tank KO mice. Locomotor activity of tank KO mice is reduced in (g) the center of the maze, but also (h) when total locomotion is considered (pre-planned Bonferroni-corrected LSD test; ^#P < 0.05; ^$P < 0.001 vs. WT).

Figure 6.

TANK controls neuroimmune activation after alcohol and/or lipopolysaccharide (LPS) in the insular cortex of mice. (a) Images of p-NFκB-p65+immunoreactive cells (+IR) of the anterior insular cortex of tank KO and WT mice after alcohol (5 g/kg, i.g., 10 days) and LPS (3 mg/kg, i.p.) treatment (scale bar = 30 μm). (b) Quantification of p-NFκB-p65+IR cells. BioQuant image analysis shows that cell number of p-NFκB-p65+IR was significantly increased after 10 days of alcohol (i.g.) or LPS alone. Pretreatment of alcohol significantly enhances LPS-induced p-NFκB-p65 immunoreactivity in both WT and tank KO mice (ANOVA: P < 0.01). Anterior insular cortex of tank KO mice shows significantly decreased p-NFκB-p65+IR cells after alcohol and/or LPS compared with WT mice (t-test; *P < 0.05, **P < 0.01 vs. control treatment; ^§§P < 0.01 vs. WT mice with same treatment).