The BDNF Val68Met polymorphism causes a sex specific alcohol preference over social interaction and also acute tolerance to the anxiolytic effects of alcohol, a phenotype driven by malfunction of BDNF in the ventral hippocampus of male mice

Abstract

Background: The brain-derived neurotrophic factor (BDNF) Valine 66 to Methionine human polymorphism results in impaired activity-dependent BDNF release and has been linked to psychiatric disorders including depression and anxiety. We previously showed that male knock-in mice carrying the mouse Methionine homolog (Met68BDNF) exhibit excessive and compulsive alcohol drinking behaviors as compared to the wild-type Val68BDNF mice.

Objective: Here, we set out to determine the potential mechanism for the heightened and compulsive alcohol drinking phenotypes detected in Met68BDNF mice.

Results: We found that male, but not female Met68BDNF mice exhibit social anxiety-like behaviors. We further show that male Met68BDNF mice exhibit a preference for alcohol over social interaction. In contrast, alcohol place preference without an alternative social reward, is similar in male Met68BDNF and Val68BDNF mice. Since the Met68BDNF mice show social anxiety phenotypes, we tested whether alcohol reliefs anxiety similarly in Met68BDNF and Val68BDNF mice and found that male, but not female Met68BDNF mice are insensitive to the acute anxiolytic action of alcohol. Finally, we show that this acute tolerance to alcohol-dependent anxiolysis can be restored by overexpressing wild-type Val68BDNF in the ventral hippocampus (vHC) of Met68BDNF mice.

Conclusions: Together, our results suggest that excessive alcohol drinking in the Met68BDNF may be attributed, in part, to heighted social anxiety and a lack of alcohol-dependent anxiolysis, a phenotype that is associated with malfunction of BDNF signaling in the vHC of male Met68BDNF mice.

Keywords: Alcohol; BDNF; BDNF Val/Met polymorphism; Ventral hippocampus.

Fig. 1

Male Met68BDNF mice exhibit social anxiety-like behaviors. A Three-chamber sociability and social novelty paradigms. Sociability test (left). The distal chamber contained an empty cage, and the other chamber contained a sex match juvenile C57Bl/6 J mouse (stranger I). Female (green) and male (blue) Val68BDNF, and female (orange) and male (red) Met68BDNF mice were allowed to interact with the stranger I mouse for 15 min, and the time a mouse spent with stranger I mouse was recorded and quantified. Social novelty test (right). Stranger I was relocated to the distal chamber, and a sex-matched novel juvenile C57Bl/6 J mouse (stranger II), was placed in the other chamber. Female (green) and male (blue), Val68BDNF and female (orange) and male (red) Met68BDNF mice were allowed to interact with stranger I or stranger II for 15 min, and the time a mouse spent with stranger I or stranger II was recorded and quantified. B Female and male Val68BDNF and Met68BDNF mice spent about the same amount of time interacting with the stranger I mouse during the sociability test. C Male Met68BDNF mice and male and female Val68BDNF mice spent significantly more time in the chamber containing the stranger I mouse than they do in an empty chamber. D Male Met68BDNF mice spent significantly less time interacting with a stranger II mouse in the social novelty test as compared to male Val68BDNF mice. Female Val68BDNF and Met68BDNF mice spent a similar amount of time interacting with a stranger II mouse. E Male Met68BDNF mice spent significantly more time in a chamber containing the more-familiar stranger I than they do in a chamber containing stranger II. Female Met68BDNF mice spent significantly more time in the chamber containing stranger II than the chamber containing stranger I. F Heatmaps of mean mouse position of male and female Val68BDNF and Met68BDNF mice during the social novelty test shown in (C, E). Stranger I mice are shown on the left of each heatmap, and stranger II are shown on the right side. Dotted lines indicate the approximate position of apparatus walls. All data are represented as mean ± SEM. *** p < 0.001, * p < 0.05; ns, non-significant. Female Val68BDNF: n = 15, all other experimental groups: n = 14

Fig. 2

Male Met68BDNF mice exhibit aberrant social interaction. A Open field social interaction paradigm. Male (blue) and female (green) Val68BDNF and male (red) and female (orange) Met68BDNF mice were placed in an open field apparatus with a novel sex-matched juvenile C57BL/6J mouse for 10 min and body contacts were recorded. B Male Met68BDNF mice spent significantly less time in contact with a novel interaction partner than male Val68BDNF. C There was no significant difference in total body contacts with a novel interaction partner between female Val68BDNF and Met68BDNF mice. ** p < 0.01; ns, non-significant. B Male Val68BDNF: n = 10, male Met68BDNF: n = 9, C female Val68BDNF and Met68BDNF n = 14

Fig. 3

Male Met68BDNF mice demonstrate social aversion and alcohol preference in a social-alcohol place conditioning test. A Outline of the social-alcohol place preference/aversion test. On day 1, mice explored the entire apparatus for 15 min. In the morning of days 2–4 conditioning days, mice received an i.p. injection of saline before a 10-min social interaction period in the social assigned chamber. In the afternoon of days 2–4 mice received an i.p. injection of 2 g/kg of alcohol prior to being placed in the alcohol-paired chamber for 10 min. On day 5, mice were allowed to freely explore the entire apparatus for 15 min and the time spend in the social and alcohol chambers were recorded and quantified. B Male Met68BDNF mice (red) spent significantly less time in the social-paired chamber in the post-test than they did in the pre-test, and significantly more time in the alcohol-paired chamber in the post-test compared to the pre-test. Male Val68BDNF mice (blue) do not exhibit place preference or aversion. C Representative heatmaps of mouse position of Val68BDNF and Met68BDNF cohorts during the social-alcohol CPP/CPA shown in (B). The social chamber is depicted on the left and the alcohol chamber is depicted on the right for each heatmap. The upper portion of the heatmap represents the connecting hallway between chambers. Dotted lines indicate the approximate position of apparatus walls. All data are represented as mean ± SEM; ** p < 0.01, *** p < 0.001. Val68BDNF: n = 12, Met68BDNF: n = 10

Fig. 4

Male Met68BDNF exhibit acute tolerance to the anxiolytic action of alcohol. A Ten minutes following i.p. injection of saline or alcohol (1.25 g/kg), mice were placed in the center of the elevated plus maze and allowed to freely explore the apparatus. The time mice spent in the open arm and in the distal portion of the open arms was measured. B–C Following an i.p. administration of alcohol (hatched bars), female and male mice spent significantly more time in the open arm (B) and the distal portion of the open arm (C) than upon receiving an injection of saline (open bars). Male Met68BDNF mice (red) exploration time in the open arm (B) and the distal portion of the open arm (C) was unaltered following a systemic administration of alcohol (hatched bars) or saline (empty bars). D–E Additional analysis on male mice was performed in which the time in open arms or distal arms after alcohol treatment was normalized to the time in open arm or distal arms of the saline group. F Heatmaps representing mean relative position of animals in each group. Open arms are represented horizontally, and closed arms extend vertically from the center. Dotted lines indicate the approximate position of apparatus walls/ledges. Data are represented as mean ± SEM; * p < 0.05, ** p < 0.01, *** p < 0.001. Male Val68BDNF + saline: n = 15, male Val68BDNF + alcohol: n = 14, male Met68BDNF + saline: n = 13, male Met68BDNF + alcohol: n = 14, female Val68BDNF + saline: n = 6, female Val68BDNF + alcohol: n = 7, female Met68BDNF + saline: n = 7, female Met68BDNF + alcohol: n = 7

Fig. 5

Overexpression of Val68BDNF in the vHC of male Met68BDNF rescues the anxiolytic effect of alcohol. A Confirmation of Val68BDNF overexpression. Image (10 ×) depicts GFP signal in the vHC of a Met68BDNF mouse that received AAV-Val68BDNF in the vHC. B–C Male Met68BDNF that received AAV-GFP (green) in the vHC spent similar time in both the open arm (B) and the distal open arm (C) following i.p. injection of alcohol (1.25 g/kg) (hatched bars) or saline (open bars). Met68BDNF mice that received AAV-Val68BDNF (purple) in the vHC spent significantly more time in the open arm (B) and the distal open arm (C) following a systemic administration of alcohol (hatched bars) than they did following saline administration (open bars). D Heatmaps of group mean position on the elevated plus maze. Open arms are horizontal, and closed arms extend vertically from the center. Dotted lines indicate the approximate position of apparatus walls/ledges. Data represented as mean ± SEM; *** p < 0.001. Male Met68BDNF + AAV-GFP: n = 11, Male Met68BDNF + AAV-Val68BDNF: n = 10