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Increased hippocampal excitability and impaired spatial memory function in mice lacking VGLUT2 selectively in neurons defined by tyrosine hydroxylase promoter activity.

Nordenankar K, Smith-Anttila CJ, Schweizer N, Viereckel T, Birgner C, Mejia-Toiber J, Morales M, Leao RN, Wallén-Mackenzie Å - Brain Struct Funct (2014)

Bottom Line: The transient phase is, however, not specific to catecholaminergic neurons, a feature taken to advantage here as it enabled Vglut2 gene targeting within all three A10 populations expressing this gene, thus creating a new conditional knockout.Electrophysiological analyses revealed a profound alteration of oscillatory activity in the CA3 region of the hippocampus.In addition to identifying a novel role for Vglut2 in hippocampus function, this study points to the need for improved genetic tools for targeting of the diversity of subpopulations of the A10 area.

View Article: PubMed Central - PubMed

Affiliation: Unit of Functional Neurobiology and Unit of Developmental Genetics, Biomedical Center, Department of Neuroscience, Uppsala University, Box 593, S-751 24, Uppsala, Sweden.

ABSTRACT
Three populations of neurons expressing the vesicular glutamate transporter 2 (Vglut2) were recently described in the A10 area of the mouse midbrain, of which two populations were shown to express the gene encoding, the rate-limiting enzyme for catecholamine synthesis, tyrosine hydroxylase (TH).One of these populations ("TH-Vglut2 Class1") also expressed the dopamine transporter (DAT) gene while one did not ("TH-Vglut2 Class2"), and the remaining population did not express TH at all ("Vglut2-only"). TH is known to be expressed by a promoter which shows two phases of activation, a transient one early during embryonal development, and a later one which gives rise to stable endogenous expression of the TH gene. The transient phase is, however, not specific to catecholaminergic neurons, a feature taken to advantage here as it enabled Vglut2 gene targeting within all three A10 populations expressing this gene, thus creating a new conditional knockout. These knockout mice showed impairment in spatial memory function. Electrophysiological analyses revealed a profound alteration of oscillatory activity in the CA3 region of the hippocampus. In addition to identifying a novel role for Vglut2 in hippocampus function, this study points to the need for improved genetic tools for targeting of the diversity of subpopulations of the A10 area.

No MeSH data available.


Related in: MedlinePlus

Immunohistochemical and in situ hybridization analysis on sagittal sections of the embryonal mouse brain. Immunofluorescent analysis of VGLUT2 (red) with cell nuclei labelled with DAPI (blue) at embryonic (E) day 12. Synaptic colocalization of VGLUT2 (red) and Synapsin (green) (a); Nestin-expressing neuronal progenitors (green) interspersed with VGLUT2 (red) (b); localization of VGLUT2 (red) within and parallel to fibrous structures immunoreactive for neuronal marker alpha-Internexin (a-Internexin; green) (c left and right); localization of vesicular VGLUT2 (red dots) within neuronal fibre bundles labelled by β-III-Tubulin (Tuj1; green) (d); overview (E15) of ventral midbrain (VM) to visualize the area above mesencephalic flexure (MF) and TH-positive (green) neurons with projections in median forebrain bundle (MFB) to striatum (e), dotted square marks the area of sections shown in f, g; VGLUT2-positive fibres (note, green) in the VM above the MF, no colocalization with the marker for inhibitory neurons, VIAAT (red) (f); VGLUT2-positive fibres (red) in VM area of TH-positive neurons (green) (g); (h) in situ hybridization combined with immunohistochemistry of Vglut2 mRNA (black) and TH protein (green) in the VM.Cells positive for both Vglut2 (black) and TH (green) marked with single asterisk cells positive for Vglut2 but not TH marked with hash
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Fig1: Immunohistochemical and in situ hybridization analysis on sagittal sections of the embryonal mouse brain. Immunofluorescent analysis of VGLUT2 (red) with cell nuclei labelled with DAPI (blue) at embryonic (E) day 12. Synaptic colocalization of VGLUT2 (red) and Synapsin (green) (a); Nestin-expressing neuronal progenitors (green) interspersed with VGLUT2 (red) (b); localization of VGLUT2 (red) within and parallel to fibrous structures immunoreactive for neuronal marker alpha-Internexin (a-Internexin; green) (c left and right); localization of vesicular VGLUT2 (red dots) within neuronal fibre bundles labelled by β-III-Tubulin (Tuj1; green) (d); overview (E15) of ventral midbrain (VM) to visualize the area above mesencephalic flexure (MF) and TH-positive (green) neurons with projections in median forebrain bundle (MFB) to striatum (e), dotted square marks the area of sections shown in f, g; VGLUT2-positive fibres (note, green) in the VM above the MF, no colocalization with the marker for inhibitory neurons, VIAAT (red) (f); VGLUT2-positive fibres (red) in VM area of TH-positive neurons (green) (g); (h) in situ hybridization combined with immunohistochemistry of Vglut2 mRNA (black) and TH protein (green) in the VM.Cells positive for both Vglut2 (black) and TH (green) marked with single asterisk cells positive for Vglut2 but not TH marked with hash

Mentions: We have previously shown that Vglut2 is prominently expressed in the subventricular zone in several cortical, subcortical and brainstem structures, including in TH-expressing neurons in the ventral midbrain, of the mouse already at midgestation (Birgner et al. 2010). To analyse if Vglut2 gene expression at these early developmental stages results in protein products, we analysed localization of the VGLUT2 protein in combination with a range of various molecular markers by immunohistochemistry (IHC). Ample VGLUT2 protein was found in the di-, mes- and metencephalon of the developing mouse brain already at E12, detected in the distinct, dotted appearance of presynaptic structures where it colocalised with the presynaptic marker Synapsin (Fig. 1a, representative example). VGLUT2 protein did not colocalize with Nestin, a marker for dividing neural progenitor cells (Lendahl et al. 1990) (Fig. 1b), but was detected in elongated structures resembling fibre bundles, where it showed some, but not complete colocalization with alpha-Internexin (Fig. 1c), a class IV intermediate filament protein commonly used as a marker for subclasses of neurons and the general neuronal fibre marker beta-III-Tubulin (TUJ1) (Fig. 1d). These fibre structures are usually not observed when analysing VGLUT2 protein immunoreactivity in the adult brain, where it is exclusively presynaptic and vesicular, suggesting that VGLUT2 is present in the embryo already during the establishment of the developing axonal structures. When analysing the ventral midbrain (Fig. 1e for localization of area), VGLUT2 was found not to colocalize with VIAAT, a molecular marker for inhibitory neurons (Fig. 1f). TH-expressing neurons were found intermingled with VGLUT2-positive fibres, suggesting ample glutamatergic afferent input into this area (Fig. 1g). With the reports of a “Glu-only” neuronal population in the adult VTA (Hnasko et al. 2012; Kawano et al. 2006; Li et al. 2013; Yamaguchi et al. 2007), we wished to assess Vglut2 expression in the ventral mesencephalic flexure where VTA and SNc neurons form (Wang et al. 1995; Ye et al. 1998). Vglut2 in situ hybridization analysis at E12 combined with IHC for TH showed a mixture of cells expressing both Vglut2 and TH (“TH–Vglut2”), cells expressing Vglut2 but not TH (“Vglut2-only”), and cells expressing TH but not Vglut2 (“TH-only”) at all three embryonal stages examined (Fig. 1h for representative example, and Fig. 8 for illustration of these populations). Together, these analyses show that the Vglut2 gene is expressed from midgestation and onwards in the developing mouse brain, including the developing VTA area where it is found in both TH-expressing and non-TH-expressing cells.Fig. 1


Increased hippocampal excitability and impaired spatial memory function in mice lacking VGLUT2 selectively in neurons defined by tyrosine hydroxylase promoter activity.

Nordenankar K, Smith-Anttila CJ, Schweizer N, Viereckel T, Birgner C, Mejia-Toiber J, Morales M, Leao RN, Wallén-Mackenzie Å - Brain Struct Funct (2014)

Immunohistochemical and in situ hybridization analysis on sagittal sections of the embryonal mouse brain. Immunofluorescent analysis of VGLUT2 (red) with cell nuclei labelled with DAPI (blue) at embryonic (E) day 12. Synaptic colocalization of VGLUT2 (red) and Synapsin (green) (a); Nestin-expressing neuronal progenitors (green) interspersed with VGLUT2 (red) (b); localization of VGLUT2 (red) within and parallel to fibrous structures immunoreactive for neuronal marker alpha-Internexin (a-Internexin; green) (c left and right); localization of vesicular VGLUT2 (red dots) within neuronal fibre bundles labelled by β-III-Tubulin (Tuj1; green) (d); overview (E15) of ventral midbrain (VM) to visualize the area above mesencephalic flexure (MF) and TH-positive (green) neurons with projections in median forebrain bundle (MFB) to striatum (e), dotted square marks the area of sections shown in f, g; VGLUT2-positive fibres (note, green) in the VM above the MF, no colocalization with the marker for inhibitory neurons, VIAAT (red) (f); VGLUT2-positive fibres (red) in VM area of TH-positive neurons (green) (g); (h) in situ hybridization combined with immunohistochemistry of Vglut2 mRNA (black) and TH protein (green) in the VM.Cells positive for both Vglut2 (black) and TH (green) marked with single asterisk cells positive for Vglut2 but not TH marked with hash
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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Fig1: Immunohistochemical and in situ hybridization analysis on sagittal sections of the embryonal mouse brain. Immunofluorescent analysis of VGLUT2 (red) with cell nuclei labelled with DAPI (blue) at embryonic (E) day 12. Synaptic colocalization of VGLUT2 (red) and Synapsin (green) (a); Nestin-expressing neuronal progenitors (green) interspersed with VGLUT2 (red) (b); localization of VGLUT2 (red) within and parallel to fibrous structures immunoreactive for neuronal marker alpha-Internexin (a-Internexin; green) (c left and right); localization of vesicular VGLUT2 (red dots) within neuronal fibre bundles labelled by β-III-Tubulin (Tuj1; green) (d); overview (E15) of ventral midbrain (VM) to visualize the area above mesencephalic flexure (MF) and TH-positive (green) neurons with projections in median forebrain bundle (MFB) to striatum (e), dotted square marks the area of sections shown in f, g; VGLUT2-positive fibres (note, green) in the VM above the MF, no colocalization with the marker for inhibitory neurons, VIAAT (red) (f); VGLUT2-positive fibres (red) in VM area of TH-positive neurons (green) (g); (h) in situ hybridization combined with immunohistochemistry of Vglut2 mRNA (black) and TH protein (green) in the VM.Cells positive for both Vglut2 (black) and TH (green) marked with single asterisk cells positive for Vglut2 but not TH marked with hash
Mentions: We have previously shown that Vglut2 is prominently expressed in the subventricular zone in several cortical, subcortical and brainstem structures, including in TH-expressing neurons in the ventral midbrain, of the mouse already at midgestation (Birgner et al. 2010). To analyse if Vglut2 gene expression at these early developmental stages results in protein products, we analysed localization of the VGLUT2 protein in combination with a range of various molecular markers by immunohistochemistry (IHC). Ample VGLUT2 protein was found in the di-, mes- and metencephalon of the developing mouse brain already at E12, detected in the distinct, dotted appearance of presynaptic structures where it colocalised with the presynaptic marker Synapsin (Fig. 1a, representative example). VGLUT2 protein did not colocalize with Nestin, a marker for dividing neural progenitor cells (Lendahl et al. 1990) (Fig. 1b), but was detected in elongated structures resembling fibre bundles, where it showed some, but not complete colocalization with alpha-Internexin (Fig. 1c), a class IV intermediate filament protein commonly used as a marker for subclasses of neurons and the general neuronal fibre marker beta-III-Tubulin (TUJ1) (Fig. 1d). These fibre structures are usually not observed when analysing VGLUT2 protein immunoreactivity in the adult brain, where it is exclusively presynaptic and vesicular, suggesting that VGLUT2 is present in the embryo already during the establishment of the developing axonal structures. When analysing the ventral midbrain (Fig. 1e for localization of area), VGLUT2 was found not to colocalize with VIAAT, a molecular marker for inhibitory neurons (Fig. 1f). TH-expressing neurons were found intermingled with VGLUT2-positive fibres, suggesting ample glutamatergic afferent input into this area (Fig. 1g). With the reports of a “Glu-only” neuronal population in the adult VTA (Hnasko et al. 2012; Kawano et al. 2006; Li et al. 2013; Yamaguchi et al. 2007), we wished to assess Vglut2 expression in the ventral mesencephalic flexure where VTA and SNc neurons form (Wang et al. 1995; Ye et al. 1998). Vglut2 in situ hybridization analysis at E12 combined with IHC for TH showed a mixture of cells expressing both Vglut2 and TH (“TH–Vglut2”), cells expressing Vglut2 but not TH (“Vglut2-only”), and cells expressing TH but not Vglut2 (“TH-only”) at all three embryonal stages examined (Fig. 1h for representative example, and Fig. 8 for illustration of these populations). Together, these analyses show that the Vglut2 gene is expressed from midgestation and onwards in the developing mouse brain, including the developing VTA area where it is found in both TH-expressing and non-TH-expressing cells.Fig. 1

Bottom Line: The transient phase is, however, not specific to catecholaminergic neurons, a feature taken to advantage here as it enabled Vglut2 gene targeting within all three A10 populations expressing this gene, thus creating a new conditional knockout.Electrophysiological analyses revealed a profound alteration of oscillatory activity in the CA3 region of the hippocampus.In addition to identifying a novel role for Vglut2 in hippocampus function, this study points to the need for improved genetic tools for targeting of the diversity of subpopulations of the A10 area.

View Article: PubMed Central - PubMed

Affiliation: Unit of Functional Neurobiology and Unit of Developmental Genetics, Biomedical Center, Department of Neuroscience, Uppsala University, Box 593, S-751 24, Uppsala, Sweden.

ABSTRACT
Three populations of neurons expressing the vesicular glutamate transporter 2 (Vglut2) were recently described in the A10 area of the mouse midbrain, of which two populations were shown to express the gene encoding, the rate-limiting enzyme for catecholamine synthesis, tyrosine hydroxylase (TH).One of these populations ("TH-Vglut2 Class1") also expressed the dopamine transporter (DAT) gene while one did not ("TH-Vglut2 Class2"), and the remaining population did not express TH at all ("Vglut2-only"). TH is known to be expressed by a promoter which shows two phases of activation, a transient one early during embryonal development, and a later one which gives rise to stable endogenous expression of the TH gene. The transient phase is, however, not specific to catecholaminergic neurons, a feature taken to advantage here as it enabled Vglut2 gene targeting within all three A10 populations expressing this gene, thus creating a new conditional knockout. These knockout mice showed impairment in spatial memory function. Electrophysiological analyses revealed a profound alteration of oscillatory activity in the CA3 region of the hippocampus. In addition to identifying a novel role for Vglut2 in hippocampus function, this study points to the need for improved genetic tools for targeting of the diversity of subpopulations of the A10 area.

No MeSH data available.


Related in: MedlinePlus