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Rgs6 is required for adult maintenance of dopaminergic neurons in the ventral substantia nigra.

Bifsha P, Yang J, Fisher RA, Drouin J - PLoS Genet. (2014)

Bottom Line: Unilateral cell loss is accompanied by contralateral degenerating neurons that exhibit smaller cell size, altered morphology and reduced dendritic network.The degenerating neurons have low levels of tyrosine hydroxylase (TH) and decreased nuclear Pitx3; accordingly, expression of many Pitx3 target gene products is altered, including Vmat2, Bdnf, Aldh1a1 (Adh2) and Fgf10.Rgs6-dependent protection is thus critical for adult survival and maintenance of the vSNc mDA neurons that are most affected in PD.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Génétique Moléculaire, Institut de Recherches Cliniques de Montréal (IRCM) Montréal, Quebec, Canada; Division of Experimental Medicine, McGill University, Montréal, Quebec, Canada.

ABSTRACT
Parkinson disease (PD) is characterized by the preferential, but poorly understood, vulnerability to degeneration of midbrain dopaminergic (mDA) neurons in the ventral substantia nigra compacta (vSNc). These sensitive mDA neurons express Pitx3, a transcription factor that is critical for their survival during development. We used this dependence to identify, by flow cytometry and expression profiling, the negative regulator of G-protein signaling Rgs6 for its restricted expression in these neurons. In contrast to Pitx3-/- mDA neurons that die during fetal (vSNc) or post-natal (VTA) period, the vSNc mDA neurons of Rgs6-/- mutant mice begin to exhibit unilateral signs of degeneration at around 6 months of age, and by one year cell loss is observed in a fraction of mice. Unilateral cell loss is accompanied by contralateral degenerating neurons that exhibit smaller cell size, altered morphology and reduced dendritic network. The degenerating neurons have low levels of tyrosine hydroxylase (TH) and decreased nuclear Pitx3; accordingly, expression of many Pitx3 target gene products is altered, including Vmat2, Bdnf, Aldh1a1 (Adh2) and Fgf10. These low TH neurons also express markers of increased dopamine signaling, namely increased DAT and phospho-Erk1/2 expression. The late onset degeneration may reflect the protective action of Rgs6 against excessive DA signaling throughout life. Rgs6-dependent protection is thus critical for adult survival and maintenance of the vSNc mDA neurons that are most affected in PD.

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Reduced expression of Pitx3 and its target genes in degenerating neurons.(A) Double immunofluorescence staining against TH (green) and Pitx3 (red) in SNc of control and 1 y-old Rgs6−/− mice that display dysmorphic mDA neurons. Arrowheads indicate Pitx3-negative neurons of dSNc. Scale bar 20 µm. (B) Co-immunofluorescence staining against TH (green), nuclear Dapi (blue) and Vmat2 (red) in SNc of WT and Rgs6−/− mice. Scale bar 20 µm. (C) Co-immunofluorescence staining against TH (green), nuclear Dapi (blue) and Bdnf (red) in SNc and VTA of WT and Rgs6−/− mice. Scale bar 20 µm. (D) Double immunofluorescence staining against TH (green) and Pitx3, Aldh1a1, Fgf10 (red) in control and 1 y-old Rgs6−/− mice that display dysmorphic mDA neurons. Arrowheads indicate unaffected dSNc neurons and arrows point to affected vSNc neurons. Scale bar 100 µm.
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pgen-1004863-g007: Reduced expression of Pitx3 and its target genes in degenerating neurons.(A) Double immunofluorescence staining against TH (green) and Pitx3 (red) in SNc of control and 1 y-old Rgs6−/− mice that display dysmorphic mDA neurons. Arrowheads indicate Pitx3-negative neurons of dSNc. Scale bar 20 µm. (B) Co-immunofluorescence staining against TH (green), nuclear Dapi (blue) and Vmat2 (red) in SNc of WT and Rgs6−/− mice. Scale bar 20 µm. (C) Co-immunofluorescence staining against TH (green), nuclear Dapi (blue) and Bdnf (red) in SNc and VTA of WT and Rgs6−/− mice. Scale bar 20 µm. (D) Double immunofluorescence staining against TH (green) and Pitx3, Aldh1a1, Fgf10 (red) in control and 1 y-old Rgs6−/− mice that display dysmorphic mDA neurons. Arrowheads indicate unaffected dSNc neurons and arrows point to affected vSNc neurons. Scale bar 100 µm.

Mentions: In order to define molecular correlates of Rgs6-dependent degeneration, we assessed Pitx3 expression that was previously shown to have a survival function in SNc [12]. Nuclear Pitx3 was greatly diminished in the dysmorphic mDA neurons while cytoplasmic Pitx3 staining increased, suggesting a shift in sub-cellular localization observed with two different polyclonal Pitx3 antibodies in different cells of the same sections (Fig. 7A). We assessed protein levels of some Pitx3 target genes by immunohistochemistry: these include Bdnf, Aldh1a1 (Adh2), TH, Drd2, DAT, Vmat2 and Fgf10 [19], [23], [40]. The expression of Vmat2 and Bdnf is decreased in THlow neurons (Fig. 7B, C). Expression of Aldh1a1 (Adh2) is also decreased in degenerating vSNc mDA neurons (Fig. 7D): this decrease may in part account for the low TH expression [19]. Our profiling data suggested that Fgf10 expression is repressed by Pitx3 in VTA and indeed, we found de-repression of Fgf10 expression only in THlow degenerating mDA neurons (Fig. 7D). At post-natal day 6 (not shown), we did not observe any change in expression of these genes consistent with a late onset phenotype.


Rgs6 is required for adult maintenance of dopaminergic neurons in the ventral substantia nigra.

Bifsha P, Yang J, Fisher RA, Drouin J - PLoS Genet. (2014)

Reduced expression of Pitx3 and its target genes in degenerating neurons.(A) Double immunofluorescence staining against TH (green) and Pitx3 (red) in SNc of control and 1 y-old Rgs6−/− mice that display dysmorphic mDA neurons. Arrowheads indicate Pitx3-negative neurons of dSNc. Scale bar 20 µm. (B) Co-immunofluorescence staining against TH (green), nuclear Dapi (blue) and Vmat2 (red) in SNc of WT and Rgs6−/− mice. Scale bar 20 µm. (C) Co-immunofluorescence staining against TH (green), nuclear Dapi (blue) and Bdnf (red) in SNc and VTA of WT and Rgs6−/− mice. Scale bar 20 µm. (D) Double immunofluorescence staining against TH (green) and Pitx3, Aldh1a1, Fgf10 (red) in control and 1 y-old Rgs6−/− mice that display dysmorphic mDA neurons. Arrowheads indicate unaffected dSNc neurons and arrows point to affected vSNc neurons. Scale bar 100 µm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263397&req=5

pgen-1004863-g007: Reduced expression of Pitx3 and its target genes in degenerating neurons.(A) Double immunofluorescence staining against TH (green) and Pitx3 (red) in SNc of control and 1 y-old Rgs6−/− mice that display dysmorphic mDA neurons. Arrowheads indicate Pitx3-negative neurons of dSNc. Scale bar 20 µm. (B) Co-immunofluorescence staining against TH (green), nuclear Dapi (blue) and Vmat2 (red) in SNc of WT and Rgs6−/− mice. Scale bar 20 µm. (C) Co-immunofluorescence staining against TH (green), nuclear Dapi (blue) and Bdnf (red) in SNc and VTA of WT and Rgs6−/− mice. Scale bar 20 µm. (D) Double immunofluorescence staining against TH (green) and Pitx3, Aldh1a1, Fgf10 (red) in control and 1 y-old Rgs6−/− mice that display dysmorphic mDA neurons. Arrowheads indicate unaffected dSNc neurons and arrows point to affected vSNc neurons. Scale bar 100 µm.
Mentions: In order to define molecular correlates of Rgs6-dependent degeneration, we assessed Pitx3 expression that was previously shown to have a survival function in SNc [12]. Nuclear Pitx3 was greatly diminished in the dysmorphic mDA neurons while cytoplasmic Pitx3 staining increased, suggesting a shift in sub-cellular localization observed with two different polyclonal Pitx3 antibodies in different cells of the same sections (Fig. 7A). We assessed protein levels of some Pitx3 target genes by immunohistochemistry: these include Bdnf, Aldh1a1 (Adh2), TH, Drd2, DAT, Vmat2 and Fgf10 [19], [23], [40]. The expression of Vmat2 and Bdnf is decreased in THlow neurons (Fig. 7B, C). Expression of Aldh1a1 (Adh2) is also decreased in degenerating vSNc mDA neurons (Fig. 7D): this decrease may in part account for the low TH expression [19]. Our profiling data suggested that Fgf10 expression is repressed by Pitx3 in VTA and indeed, we found de-repression of Fgf10 expression only in THlow degenerating mDA neurons (Fig. 7D). At post-natal day 6 (not shown), we did not observe any change in expression of these genes consistent with a late onset phenotype.

Bottom Line: Unilateral cell loss is accompanied by contralateral degenerating neurons that exhibit smaller cell size, altered morphology and reduced dendritic network.The degenerating neurons have low levels of tyrosine hydroxylase (TH) and decreased nuclear Pitx3; accordingly, expression of many Pitx3 target gene products is altered, including Vmat2, Bdnf, Aldh1a1 (Adh2) and Fgf10.Rgs6-dependent protection is thus critical for adult survival and maintenance of the vSNc mDA neurons that are most affected in PD.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Génétique Moléculaire, Institut de Recherches Cliniques de Montréal (IRCM) Montréal, Quebec, Canada; Division of Experimental Medicine, McGill University, Montréal, Quebec, Canada.

ABSTRACT
Parkinson disease (PD) is characterized by the preferential, but poorly understood, vulnerability to degeneration of midbrain dopaminergic (mDA) neurons in the ventral substantia nigra compacta (vSNc). These sensitive mDA neurons express Pitx3, a transcription factor that is critical for their survival during development. We used this dependence to identify, by flow cytometry and expression profiling, the negative regulator of G-protein signaling Rgs6 for its restricted expression in these neurons. In contrast to Pitx3-/- mDA neurons that die during fetal (vSNc) or post-natal (VTA) period, the vSNc mDA neurons of Rgs6-/- mutant mice begin to exhibit unilateral signs of degeneration at around 6 months of age, and by one year cell loss is observed in a fraction of mice. Unilateral cell loss is accompanied by contralateral degenerating neurons that exhibit smaller cell size, altered morphology and reduced dendritic network. The degenerating neurons have low levels of tyrosine hydroxylase (TH) and decreased nuclear Pitx3; accordingly, expression of many Pitx3 target gene products is altered, including Vmat2, Bdnf, Aldh1a1 (Adh2) and Fgf10. These low TH neurons also express markers of increased dopamine signaling, namely increased DAT and phospho-Erk1/2 expression. The late onset degeneration may reflect the protective action of Rgs6 against excessive DA signaling throughout life. Rgs6-dependent protection is thus critical for adult survival and maintenance of the vSNc mDA neurons that are most affected in PD.

Show MeSH
Related in: MedlinePlus