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The A9 dopamine neuron component in grafts of ventral mesencephalon is an important determinant for recovery of motor function in a rat model of Parkinson's disease.

Grealish S, Jönsson ME, Li M, Kirik D, Björklund A, Thompson LH - Brain (2010)

Bottom Line: Here, we report results from a series of grafting experiments where the anatomical and functional properties of grafts either selectively lacking in A9 neurons, or with a typical A9/A10 composition were compared.The findings highlight dopamine neuronal subtype composition as a potentially important parameter to monitor in order to understand the variable nature of functional outcome better in transplantation studies.Furthermore, the results have interesting implications for current efforts in this field to generate well-characterized and standardized preparations of transplantable dopamine neuronal progenitors from stem cells.

View Article: PubMed Central - PubMed

Affiliation: Wallenberg Neuroscience Centre, Lund University, Lund, Sweden.

ABSTRACT
Grafts of foetal ventral mesencephalon, used in cell replacement therapy for Parkinson's disease, are known to contain a mix of dopamine neuronal subtypes including the A9 neurons of the substantia nigra and the A10 neurons of the ventral tegmental area. However, the relative importance of these subtypes for functional repair of the brain affected by Parkinson's disease has not been studied thoroughly. Here, we report results from a series of grafting experiments where the anatomical and functional properties of grafts either selectively lacking in A9 neurons, or with a typical A9/A10 composition were compared. The results show that the A9 component of intrastriatal grafts is of critical importance for recovery in tests on motor performance, in a rodent model of Parkinson's disease. Analysis at the histological level indicates that this is likely to be due to the unique ability of A9 neurons to innervate and functionally activate their target structure, the dorsolateral region of the host striatum. The findings highlight dopamine neuronal subtype composition as a potentially important parameter to monitor in order to understand the variable nature of functional outcome better in transplantation studies. Furthermore, the results have interesting implications for current efforts in this field to generate well-characterized and standardized preparations of transplantable dopamine neuronal progenitors from stem cells.

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Connectivity of Pitx3WT/GFP and Pitx3GFP/GFP grafts. Immunohistochemistry for TH in representative coronal sections from the ungrafted lesion control (A), Pitx3WT/GFP graft (B), and Pitx3GFP/GFP graft (C) groups, 12 weeks after transplantation. Two sections through the forebrain (∼bregma + 3.0 mm, left section and −2.0 mm, middle section) and one section through the midbrain are shown. The left-hand side of the lesion control brain shows the intact nigrostriatal projection system, while the right-hand side illustrates the substantial loss of TH+ neurons from the substantia nigra and reduction of TH+ terminal staining in the dorsolateral striatum resulting from the terminal 6-hydroxydopamine lesion (A). Dashed lines in (A) identify the dorsolateral (a) and ventromedial (b) quadrants of the striatum used to quantify TH+ fibre density in the three groups. Boxed areas are enlarged and illustrate the level of TH+ terminal staining in the dorsolateral striatum from each group. Part of the grafts, containing intensely TH+ cell bodies, can be seen on the left side of the boxed panels from the two grafted groups (B, C). Darkfield photographs illustrate immunohistochemical detection of GFP in the dorsolateral striatum in Pitx3WT/GFP graft (D) and Pitx3GFP/GFP graft (E) groups. The graft core (G) can be seen in the lower left part of these panels. The average levels of TH+ fibre density in the dorsolateral (area a) and ventromedial (area b) parts of the striatum are shown for the ungrafted lesion control (open bars, n = 8), Pitx3WT/GFP (grey bars, n = 8) and Pitx3GFP/GFP (black bars, n = 7) graft groups (F). The figures are given as a percentage of the TH+ density values obtained in corresponding regions from the intact striatum. In the dorsolateral striatum (area a), the TH+ fibre density was significantly greater in the Pitx3WT/GFP grafted group compared to the lesion control or Pitx3GFP/GFP graft groups (*P < 0.005). There was no significant difference in the level of TH+ fibre density in the ventromedial striatum (area b) between the three groups. CPu = caudate-putamen unit; NAc = nucleus accumbens; cc = corpus callosum; G = graft; SN = substantia nigra; VTA = ventral tegmental area. See main text for details of statistical analyses. Scale: A–C, 2 mm; D and E, 200 µm.
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Figure 6: Connectivity of Pitx3WT/GFP and Pitx3GFP/GFP grafts. Immunohistochemistry for TH in representative coronal sections from the ungrafted lesion control (A), Pitx3WT/GFP graft (B), and Pitx3GFP/GFP graft (C) groups, 12 weeks after transplantation. Two sections through the forebrain (∼bregma + 3.0 mm, left section and −2.0 mm, middle section) and one section through the midbrain are shown. The left-hand side of the lesion control brain shows the intact nigrostriatal projection system, while the right-hand side illustrates the substantial loss of TH+ neurons from the substantia nigra and reduction of TH+ terminal staining in the dorsolateral striatum resulting from the terminal 6-hydroxydopamine lesion (A). Dashed lines in (A) identify the dorsolateral (a) and ventromedial (b) quadrants of the striatum used to quantify TH+ fibre density in the three groups. Boxed areas are enlarged and illustrate the level of TH+ terminal staining in the dorsolateral striatum from each group. Part of the grafts, containing intensely TH+ cell bodies, can be seen on the left side of the boxed panels from the two grafted groups (B, C). Darkfield photographs illustrate immunohistochemical detection of GFP in the dorsolateral striatum in Pitx3WT/GFP graft (D) and Pitx3GFP/GFP graft (E) groups. The graft core (G) can be seen in the lower left part of these panels. The average levels of TH+ fibre density in the dorsolateral (area a) and ventromedial (area b) parts of the striatum are shown for the ungrafted lesion control (open bars, n = 8), Pitx3WT/GFP (grey bars, n = 8) and Pitx3GFP/GFP (black bars, n = 7) graft groups (F). The figures are given as a percentage of the TH+ density values obtained in corresponding regions from the intact striatum. In the dorsolateral striatum (area a), the TH+ fibre density was significantly greater in the Pitx3WT/GFP grafted group compared to the lesion control or Pitx3GFP/GFP graft groups (*P < 0.005). There was no significant difference in the level of TH+ fibre density in the ventromedial striatum (area b) between the three groups. CPu = caudate-putamen unit; NAc = nucleus accumbens; cc = corpus callosum; G = graft; SN = substantia nigra; VTA = ventral tegmental area. See main text for details of statistical analyses. Scale: A–C, 2 mm; D and E, 200 µm.

Mentions: The area of re-innervated striatum around the grafts was calculated in the dorsolateral or ventromedial quadrant from two tyrosine hydroxylase-stained sections (+0.20 and −0.30 mm from bregma), as depicted in Fig. 6, using Image J software (Version 1.32j, National Institutes of Health, USA). The entire striatum was divided into quarters, and in sections where cell bodies were visible, the core of the graft was excluded from the analysis. The measured values were corrected for non-specific background staining by subtracting values obtained from the corpus callosum. The data are expressed as optical density as a percentage of the corresponding area from the intact side, and values from both sections were combined to provide a single value for each region.Figure 2


The A9 dopamine neuron component in grafts of ventral mesencephalon is an important determinant for recovery of motor function in a rat model of Parkinson's disease.

Grealish S, Jönsson ME, Li M, Kirik D, Björklund A, Thompson LH - Brain (2010)

Connectivity of Pitx3WT/GFP and Pitx3GFP/GFP grafts. Immunohistochemistry for TH in representative coronal sections from the ungrafted lesion control (A), Pitx3WT/GFP graft (B), and Pitx3GFP/GFP graft (C) groups, 12 weeks after transplantation. Two sections through the forebrain (∼bregma + 3.0 mm, left section and −2.0 mm, middle section) and one section through the midbrain are shown. The left-hand side of the lesion control brain shows the intact nigrostriatal projection system, while the right-hand side illustrates the substantial loss of TH+ neurons from the substantia nigra and reduction of TH+ terminal staining in the dorsolateral striatum resulting from the terminal 6-hydroxydopamine lesion (A). Dashed lines in (A) identify the dorsolateral (a) and ventromedial (b) quadrants of the striatum used to quantify TH+ fibre density in the three groups. Boxed areas are enlarged and illustrate the level of TH+ terminal staining in the dorsolateral striatum from each group. Part of the grafts, containing intensely TH+ cell bodies, can be seen on the left side of the boxed panels from the two grafted groups (B, C). Darkfield photographs illustrate immunohistochemical detection of GFP in the dorsolateral striatum in Pitx3WT/GFP graft (D) and Pitx3GFP/GFP graft (E) groups. The graft core (G) can be seen in the lower left part of these panels. The average levels of TH+ fibre density in the dorsolateral (area a) and ventromedial (area b) parts of the striatum are shown for the ungrafted lesion control (open bars, n = 8), Pitx3WT/GFP (grey bars, n = 8) and Pitx3GFP/GFP (black bars, n = 7) graft groups (F). The figures are given as a percentage of the TH+ density values obtained in corresponding regions from the intact striatum. In the dorsolateral striatum (area a), the TH+ fibre density was significantly greater in the Pitx3WT/GFP grafted group compared to the lesion control or Pitx3GFP/GFP graft groups (*P < 0.005). There was no significant difference in the level of TH+ fibre density in the ventromedial striatum (area b) between the three groups. CPu = caudate-putamen unit; NAc = nucleus accumbens; cc = corpus callosum; G = graft; SN = substantia nigra; VTA = ventral tegmental area. See main text for details of statistical analyses. Scale: A–C, 2 mm; D and E, 200 µm.
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Figure 6: Connectivity of Pitx3WT/GFP and Pitx3GFP/GFP grafts. Immunohistochemistry for TH in representative coronal sections from the ungrafted lesion control (A), Pitx3WT/GFP graft (B), and Pitx3GFP/GFP graft (C) groups, 12 weeks after transplantation. Two sections through the forebrain (∼bregma + 3.0 mm, left section and −2.0 mm, middle section) and one section through the midbrain are shown. The left-hand side of the lesion control brain shows the intact nigrostriatal projection system, while the right-hand side illustrates the substantial loss of TH+ neurons from the substantia nigra and reduction of TH+ terminal staining in the dorsolateral striatum resulting from the terminal 6-hydroxydopamine lesion (A). Dashed lines in (A) identify the dorsolateral (a) and ventromedial (b) quadrants of the striatum used to quantify TH+ fibre density in the three groups. Boxed areas are enlarged and illustrate the level of TH+ terminal staining in the dorsolateral striatum from each group. Part of the grafts, containing intensely TH+ cell bodies, can be seen on the left side of the boxed panels from the two grafted groups (B, C). Darkfield photographs illustrate immunohistochemical detection of GFP in the dorsolateral striatum in Pitx3WT/GFP graft (D) and Pitx3GFP/GFP graft (E) groups. The graft core (G) can be seen in the lower left part of these panels. The average levels of TH+ fibre density in the dorsolateral (area a) and ventromedial (area b) parts of the striatum are shown for the ungrafted lesion control (open bars, n = 8), Pitx3WT/GFP (grey bars, n = 8) and Pitx3GFP/GFP (black bars, n = 7) graft groups (F). The figures are given as a percentage of the TH+ density values obtained in corresponding regions from the intact striatum. In the dorsolateral striatum (area a), the TH+ fibre density was significantly greater in the Pitx3WT/GFP grafted group compared to the lesion control or Pitx3GFP/GFP graft groups (*P < 0.005). There was no significant difference in the level of TH+ fibre density in the ventromedial striatum (area b) between the three groups. CPu = caudate-putamen unit; NAc = nucleus accumbens; cc = corpus callosum; G = graft; SN = substantia nigra; VTA = ventral tegmental area. See main text for details of statistical analyses. Scale: A–C, 2 mm; D and E, 200 µm.
Mentions: The area of re-innervated striatum around the grafts was calculated in the dorsolateral or ventromedial quadrant from two tyrosine hydroxylase-stained sections (+0.20 and −0.30 mm from bregma), as depicted in Fig. 6, using Image J software (Version 1.32j, National Institutes of Health, USA). The entire striatum was divided into quarters, and in sections where cell bodies were visible, the core of the graft was excluded from the analysis. The measured values were corrected for non-specific background staining by subtracting values obtained from the corpus callosum. The data are expressed as optical density as a percentage of the corresponding area from the intact side, and values from both sections were combined to provide a single value for each region.Figure 2

Bottom Line: Here, we report results from a series of grafting experiments where the anatomical and functional properties of grafts either selectively lacking in A9 neurons, or with a typical A9/A10 composition were compared.The findings highlight dopamine neuronal subtype composition as a potentially important parameter to monitor in order to understand the variable nature of functional outcome better in transplantation studies.Furthermore, the results have interesting implications for current efforts in this field to generate well-characterized and standardized preparations of transplantable dopamine neuronal progenitors from stem cells.

View Article: PubMed Central - PubMed

Affiliation: Wallenberg Neuroscience Centre, Lund University, Lund, Sweden.

ABSTRACT
Grafts of foetal ventral mesencephalon, used in cell replacement therapy for Parkinson's disease, are known to contain a mix of dopamine neuronal subtypes including the A9 neurons of the substantia nigra and the A10 neurons of the ventral tegmental area. However, the relative importance of these subtypes for functional repair of the brain affected by Parkinson's disease has not been studied thoroughly. Here, we report results from a series of grafting experiments where the anatomical and functional properties of grafts either selectively lacking in A9 neurons, or with a typical A9/A10 composition were compared. The results show that the A9 component of intrastriatal grafts is of critical importance for recovery in tests on motor performance, in a rodent model of Parkinson's disease. Analysis at the histological level indicates that this is likely to be due to the unique ability of A9 neurons to innervate and functionally activate their target structure, the dorsolateral region of the host striatum. The findings highlight dopamine neuronal subtype composition as a potentially important parameter to monitor in order to understand the variable nature of functional outcome better in transplantation studies. Furthermore, the results have interesting implications for current efforts in this field to generate well-characterized and standardized preparations of transplantable dopamine neuronal progenitors from stem cells.

Show MeSH
Related in: MedlinePlus