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Targeted deletion of Vglut2 expression in the embryonal telencephalon promotes an anxiolytic phenotype of the adult mouse.

Nordenankar K, Bergfors A, Wallén-Mackenzie Å - Ups. J. Med. Sci. (2015)

Bottom Line: Anxiety is a natural emotion experienced by all individuals.Anxiolysis, the reduction of anxiety, is mediated via several large groups of therapeutical compounds, but the relief is often only temporary, and increased knowledge of the neurobiology underlying anxiety is needed in order to improve future therapies.Our results suggest that both embryonal and adolescent forebrain expression of Vglut2 normally contributes to balancing the level of anxiety.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Unit of Functional Neurobiology and Unit of Developmental Genetics, Uppsala University , Box 593, S-75214 Uppsala , Sweden.

ABSTRACT

Background: Anxiety is a natural emotion experienced by all individuals. However, when anxiety becomes excessive, it contributes to the substantial group of anxiety disorders that affect one in three people and thus are among the most common psychiatric disorders. Anxiolysis, the reduction of anxiety, is mediated via several large groups of therapeutical compounds, but the relief is often only temporary, and increased knowledge of the neurobiology underlying anxiety is needed in order to improve future therapies.

Aim: We previously demonstrated that mice lacking forebrain expression of the Vesicular glutamate transporter 2 (Vglut2) from adolescence showed a strong anxiolytic behaviour as adults. In the current study, we wished to analyse if removal of Vglut2 expression already from mid-gestation of the mouse embryo would give rise to similar anxiolysis in the adult mouse.

Methods: We produced transgenic mice lacking Vglut2 from mid-gestation and analysed their affective behaviour, including anxiety, when they had reached adulthood.

Results: The transgenic mice lacking Vglut2 expression from mid-gestation showed certain signs of anxiolytic behaviour, but this phenotype was not as prominent as when Vglut2 was removed during adolescence.

Conclusion: Our results suggest that both embryonal and adolescent forebrain expression of Vglut2 normally contributes to balancing the level of anxiety. As the neurobiological basis for anxiety is similar across species, our results in mice may help improve the current understanding of the neurocircuitry of anxiety, and hence anxiolysis, also in humans.

No MeSH data available.


Related in: MedlinePlus

Specific deletion of Vglut2 in selected forebrain target areas. Floating in situ hybridization on coronal brain (70 μm) sections from control mice and Vglut2f/f;Emx1-Cre cKO mice (A–H) using a DIG-labelled Vglut2 probe. Close-ups as indicated in I–R, which demonstrate that cells expressing Vglut2 mRNA was absent in the mitral cell (Mi) layer and GI layer in the olfactory bulb (I–J). Vglut2 mRNA was also absent in the RSG of the medial cortex in the Vglut2f/f;Emx1-Cre mice (K, L). There is a loss of Vglut2 mRNA in the BMA, in the ACo the mRNA is partially deleted, and the Me amygdala and MePV are unaltered in the Vglut2f/f;Emx1-Cre mice (M–P). Vglut2 mRNA positive cells were present in the Sub in control mice but not in Vglut2f/f;Emx1-Cre cKO mice (Q, R). ACo = anterior cortical amygdaloid area; BL = basolateral amygdaloid nucleus; BM = basomedial amygdaloid nucleus anterior part; cKO = conditional knock-out; DIG = digoxigenin; GI = periglomerular layer; Me = medial amygdaloid nucleus; MePV = posteriorventral medial amygdaloid nucleus; Mi = mitral cell layer; RSG = retrosplenial group; Sub = subiculum; Bregma interval (dorsal, ventral) is shown in lower right corner.
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Figure 1: Specific deletion of Vglut2 in selected forebrain target areas. Floating in situ hybridization on coronal brain (70 μm) sections from control mice and Vglut2f/f;Emx1-Cre cKO mice (A–H) using a DIG-labelled Vglut2 probe. Close-ups as indicated in I–R, which demonstrate that cells expressing Vglut2 mRNA was absent in the mitral cell (Mi) layer and GI layer in the olfactory bulb (I–J). Vglut2 mRNA was also absent in the RSG of the medial cortex in the Vglut2f/f;Emx1-Cre mice (K, L). There is a loss of Vglut2 mRNA in the BMA, in the ACo the mRNA is partially deleted, and the Me amygdala and MePV are unaltered in the Vglut2f/f;Emx1-Cre mice (M–P). Vglut2 mRNA positive cells were present in the Sub in control mice but not in Vglut2f/f;Emx1-Cre cKO mice (Q, R). ACo = anterior cortical amygdaloid area; BL = basolateral amygdaloid nucleus; BM = basomedial amygdaloid nucleus anterior part; cKO = conditional knock-out; DIG = digoxigenin; GI = periglomerular layer; Me = medial amygdaloid nucleus; MePV = posteriorventral medial amygdaloid nucleus; Mi = mitral cell layer; RSG = retrosplenial group; Sub = subiculum; Bregma interval (dorsal, ventral) is shown in lower right corner.

Mentions: The Emx1-driven Cre activity in the Emx1IRES-Cre knock-in mouse line was investigated previously by reporter-gene analysis of the R26R strain and shown to be dorsally located in cortical subdivisions of the telencephalon from embryonic day (E) 10.5 and to stay regionally distributed also in the adult telencephalic area (31). Areas characterized by Emx1-driven Cre activity in the adult included the whole neocortex and the entire hippocampal formation as defined by the subiculum, hippocampus proper, and the dentate gyrus. The olfactory bulb and several, but not all, subpopulations of the amygdala were also characterized by Emx1-driven Cre activity. Cre-active areas included the lateral (L), centrolateral (CL), basolateral (BL), and basomedial (BM) amygdala, while leaving untouched the adjacently located medial (Me) and central (C) amygdaloid nuclei as well as the bed nucleus of stria terminalis (Bst) (the two last-mentioned a part of the so-called extended amygdala). Somewhat weaker activity was detected in the anterior cortical (ACo) amygdala (31). Guided by this previous report, brain sections from Vglut2f/f;Emx1-Cre(tg/wt) cKO mice and littermate Vglut2f/f;Emx1-Cre(wt/wt) control mice, produced as described in ‘Materials and methods’, were analysed by in situ hybridization (ISH) at the adult stage in order to ascertain the targeted deletion of Vglut2. As we previously reported, expression of Vglut2 in the adult telencephalon was found in the retrosplenial group (RSG) and layers III and V/VI (Figure 1E,Q) (13). Expression was also found in the mitral and deep periglomerular cells of the olfactory bulb (Figure 1A, I), much resembling the expression previously reported in the rat olfactory system during embryonal development (14). In accordance with the reported Cre-activity of the Emx1IRES-Cre mouse line (31), Vglut2 mRNA expression was found deleted in all of these telencephalic areas in the cKO brains (Figure 1R, E, B and J). Further, as described before, Vglut2 expression in controls was evident in the subiculum, but not in the rest of the hippocampal formation (Figure 1Q), and in several subnuclei of the amygdala complex, including the ACo, the BM, and anterior and posteroventral Me nuclei (Figure 1M, O). In the cKO mice, the subicular and BM Vglut2 expression was found absent, again in accordance with the reported Cre activity (Figure 1R, N) (31). However, Vglut2 expression in the ACo appeared reduced only (Figure 1N), in line with the limited Cre activity described in this area (31), while the expression in the Me appeared normal, fitting the apparent lack of Cre activity in these areas (Figure 1N, P) (31). In addition, we addressed Vglut2 expression in all of the remaining brain and found it normal compared to control mice (Figure 1G, H; Table I) (Supplementary Table S1, available online). Thus, in the adult Vglut2f/f;Emx1-Cre(tg/wt) cKO mice, the targeted deletion of Vglut2 is specific to the olfactory bulb, cortical subregions, the subiculum, and the BM and ACo areas of the amygdala, regions that all express the Emx1IRES-Cre bicistronic construct.


Targeted deletion of Vglut2 expression in the embryonal telencephalon promotes an anxiolytic phenotype of the adult mouse.

Nordenankar K, Bergfors A, Wallén-Mackenzie Å - Ups. J. Med. Sci. (2015)

Specific deletion of Vglut2 in selected forebrain target areas. Floating in situ hybridization on coronal brain (70 μm) sections from control mice and Vglut2f/f;Emx1-Cre cKO mice (A–H) using a DIG-labelled Vglut2 probe. Close-ups as indicated in I–R, which demonstrate that cells expressing Vglut2 mRNA was absent in the mitral cell (Mi) layer and GI layer in the olfactory bulb (I–J). Vglut2 mRNA was also absent in the RSG of the medial cortex in the Vglut2f/f;Emx1-Cre mice (K, L). There is a loss of Vglut2 mRNA in the BMA, in the ACo the mRNA is partially deleted, and the Me amygdala and MePV are unaltered in the Vglut2f/f;Emx1-Cre mice (M–P). Vglut2 mRNA positive cells were present in the Sub in control mice but not in Vglut2f/f;Emx1-Cre cKO mice (Q, R). ACo = anterior cortical amygdaloid area; BL = basolateral amygdaloid nucleus; BM = basomedial amygdaloid nucleus anterior part; cKO = conditional knock-out; DIG = digoxigenin; GI = periglomerular layer; Me = medial amygdaloid nucleus; MePV = posteriorventral medial amygdaloid nucleus; Mi = mitral cell layer; RSG = retrosplenial group; Sub = subiculum; Bregma interval (dorsal, ventral) is shown in lower right corner.
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Figure 1: Specific deletion of Vglut2 in selected forebrain target areas. Floating in situ hybridization on coronal brain (70 μm) sections from control mice and Vglut2f/f;Emx1-Cre cKO mice (A–H) using a DIG-labelled Vglut2 probe. Close-ups as indicated in I–R, which demonstrate that cells expressing Vglut2 mRNA was absent in the mitral cell (Mi) layer and GI layer in the olfactory bulb (I–J). Vglut2 mRNA was also absent in the RSG of the medial cortex in the Vglut2f/f;Emx1-Cre mice (K, L). There is a loss of Vglut2 mRNA in the BMA, in the ACo the mRNA is partially deleted, and the Me amygdala and MePV are unaltered in the Vglut2f/f;Emx1-Cre mice (M–P). Vglut2 mRNA positive cells were present in the Sub in control mice but not in Vglut2f/f;Emx1-Cre cKO mice (Q, R). ACo = anterior cortical amygdaloid area; BL = basolateral amygdaloid nucleus; BM = basomedial amygdaloid nucleus anterior part; cKO = conditional knock-out; DIG = digoxigenin; GI = periglomerular layer; Me = medial amygdaloid nucleus; MePV = posteriorventral medial amygdaloid nucleus; Mi = mitral cell layer; RSG = retrosplenial group; Sub = subiculum; Bregma interval (dorsal, ventral) is shown in lower right corner.
Mentions: The Emx1-driven Cre activity in the Emx1IRES-Cre knock-in mouse line was investigated previously by reporter-gene analysis of the R26R strain and shown to be dorsally located in cortical subdivisions of the telencephalon from embryonic day (E) 10.5 and to stay regionally distributed also in the adult telencephalic area (31). Areas characterized by Emx1-driven Cre activity in the adult included the whole neocortex and the entire hippocampal formation as defined by the subiculum, hippocampus proper, and the dentate gyrus. The olfactory bulb and several, but not all, subpopulations of the amygdala were also characterized by Emx1-driven Cre activity. Cre-active areas included the lateral (L), centrolateral (CL), basolateral (BL), and basomedial (BM) amygdala, while leaving untouched the adjacently located medial (Me) and central (C) amygdaloid nuclei as well as the bed nucleus of stria terminalis (Bst) (the two last-mentioned a part of the so-called extended amygdala). Somewhat weaker activity was detected in the anterior cortical (ACo) amygdala (31). Guided by this previous report, brain sections from Vglut2f/f;Emx1-Cre(tg/wt) cKO mice and littermate Vglut2f/f;Emx1-Cre(wt/wt) control mice, produced as described in ‘Materials and methods’, were analysed by in situ hybridization (ISH) at the adult stage in order to ascertain the targeted deletion of Vglut2. As we previously reported, expression of Vglut2 in the adult telencephalon was found in the retrosplenial group (RSG) and layers III and V/VI (Figure 1E,Q) (13). Expression was also found in the mitral and deep periglomerular cells of the olfactory bulb (Figure 1A, I), much resembling the expression previously reported in the rat olfactory system during embryonal development (14). In accordance with the reported Cre-activity of the Emx1IRES-Cre mouse line (31), Vglut2 mRNA expression was found deleted in all of these telencephalic areas in the cKO brains (Figure 1R, E, B and J). Further, as described before, Vglut2 expression in controls was evident in the subiculum, but not in the rest of the hippocampal formation (Figure 1Q), and in several subnuclei of the amygdala complex, including the ACo, the BM, and anterior and posteroventral Me nuclei (Figure 1M, O). In the cKO mice, the subicular and BM Vglut2 expression was found absent, again in accordance with the reported Cre activity (Figure 1R, N) (31). However, Vglut2 expression in the ACo appeared reduced only (Figure 1N), in line with the limited Cre activity described in this area (31), while the expression in the Me appeared normal, fitting the apparent lack of Cre activity in these areas (Figure 1N, P) (31). In addition, we addressed Vglut2 expression in all of the remaining brain and found it normal compared to control mice (Figure 1G, H; Table I) (Supplementary Table S1, available online). Thus, in the adult Vglut2f/f;Emx1-Cre(tg/wt) cKO mice, the targeted deletion of Vglut2 is specific to the olfactory bulb, cortical subregions, the subiculum, and the BM and ACo areas of the amygdala, regions that all express the Emx1IRES-Cre bicistronic construct.

Bottom Line: Anxiety is a natural emotion experienced by all individuals.Anxiolysis, the reduction of anxiety, is mediated via several large groups of therapeutical compounds, but the relief is often only temporary, and increased knowledge of the neurobiology underlying anxiety is needed in order to improve future therapies.Our results suggest that both embryonal and adolescent forebrain expression of Vglut2 normally contributes to balancing the level of anxiety.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Unit of Functional Neurobiology and Unit of Developmental Genetics, Uppsala University , Box 593, S-75214 Uppsala , Sweden.

ABSTRACT

Background: Anxiety is a natural emotion experienced by all individuals. However, when anxiety becomes excessive, it contributes to the substantial group of anxiety disorders that affect one in three people and thus are among the most common psychiatric disorders. Anxiolysis, the reduction of anxiety, is mediated via several large groups of therapeutical compounds, but the relief is often only temporary, and increased knowledge of the neurobiology underlying anxiety is needed in order to improve future therapies.

Aim: We previously demonstrated that mice lacking forebrain expression of the Vesicular glutamate transporter 2 (Vglut2) from adolescence showed a strong anxiolytic behaviour as adults. In the current study, we wished to analyse if removal of Vglut2 expression already from mid-gestation of the mouse embryo would give rise to similar anxiolysis in the adult mouse.

Methods: We produced transgenic mice lacking Vglut2 from mid-gestation and analysed their affective behaviour, including anxiety, when they had reached adulthood.

Results: The transgenic mice lacking Vglut2 expression from mid-gestation showed certain signs of anxiolytic behaviour, but this phenotype was not as prominent as when Vglut2 was removed during adolescence.

Conclusion: Our results suggest that both embryonal and adolescent forebrain expression of Vglut2 normally contributes to balancing the level of anxiety. As the neurobiological basis for anxiety is similar across species, our results in mice may help improve the current understanding of the neurocircuitry of anxiety, and hence anxiolysis, also in humans.

No MeSH data available.


Related in: MedlinePlus