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Activation of GSK-3β and caspase-3 occurs in Nigral dopamine neurons during the development of apoptosis activated by a striatal injection of 6-hydroxydopamine.

Hernandez-Baltazar D, Mendoza-Garrido ME, Martinez-Fong D - PLoS ONE (2013)

Bottom Line: The disappearance of TH(+) cells was associated with a decrease in NeuN and β-III tubulin immunoreactivity and an increase in Apostain, cleaved caspase-3, and GSK-3β pY216 in the SNc.Increased levels of caspase-3 immunoreactivity in TH(+) cells were detected from days 1 to 15, and the levels then decreased to day 30 postlesion.Our results suggest that caspase-3 and GSK-3β pY216 activation might participate in the DA cell death and that the active caspase-3 might also participate in the neuroinflammation caused by the striatal 6-OHDA injection.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, México, DF, México.

ABSTRACT
The 6-Hydroxydopamine (6-OHDA) rat model of Parkinson's disease is essential for a better understanding of the pathological processes underlying the human disease and for the evaluation of promising therapeutic interventions. This work evaluated whether a single striatal injection of 6-OHDA causes progressive apoptosis of dopamine (DA) neurons and activation of glycogen synthase kinase 3β (GSK-3β) and caspase-3 in the substantia nigra compacta (SNc). The loss of DA neurons was shown by three neuron markers; tyrosine hydroxylase (TH), NeuN, and β-III tubulin. Apoptosis activation was determined using Apostain and immunostaining against cleaved caspase-3 and GSK-3β pY216. We also explored the possibility that cleaved caspase-3 is produced by microglia and astrocytes. Our results showed that the 6-OHDA caused loss of nigral TH(+) cells, progressing mainly in rostrocaudal and lateromedial directions. In the neostriatum, a severe loss of TH(+) terminals occurred from day 3 after lesion. The disappearance of TH(+) cells was associated with a decrease in NeuN and β-III tubulin immunoreactivity and an increase in Apostain, cleaved caspase-3, and GSK-3β pY216 in the SNc. Apostain immunoreactivity was observed from days 3 to 21 postlesion. Increased levels of caspase-3 immunoreactivity in TH(+) cells were detected from days 1 to 15, and the levels then decreased to day 30 postlesion. The cleaved caspase-3 also collocated with microglia and astrocytes indicating its participation in glial activation. Our results suggest that caspase-3 and GSK-3β pY216 activation might participate in the DA cell death and that the active caspase-3 might also participate in the neuroinflammation caused by the striatal 6-OHDA injection.

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Related in: MedlinePlus

Representative merged micrographs of mesencephalon immunostained against NeuN and TH.The numbers at the left margin of the figures are the days after 6-OHDA lesion. The primary antibodies were a mouse monoclonal anti-NeuN and a rabbit polyclonal anti-TH. The secondary antibodies were sheep anti-mouse IgG (H+L) FITC conjugated and goat anti-rabbit IgG (H+L) Texas conjugated. The rectangles on the panoramic view show the regions that were amplified 40x (details). Scale bar = 1 mm (central panels) and 50 µm (details).
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pone-0070951-g002: Representative merged micrographs of mesencephalon immunostained against NeuN and TH.The numbers at the left margin of the figures are the days after 6-OHDA lesion. The primary antibodies were a mouse monoclonal anti-NeuN and a rabbit polyclonal anti-TH. The secondary antibodies were sheep anti-mouse IgG (H+L) FITC conjugated and goat anti-rabbit IgG (H+L) Texas conjugated. The rectangles on the panoramic view show the regions that were amplified 40x (details). Scale bar = 1 mm (central panels) and 50 µm (details).

Mentions: The studies of double immunofluorescence against TH and NeuN, a nuclear marker of neuronal lineage [48], showed that the loss of TH(+) cells was related to the disappearance of neuronal bodies after the 6-OHDA lesion. This was qualitatively shown in the medial region of the SNc. During the advance of a 6-OHDA lesion, there was a simultaneous loss of cells with TH and NeuN immunostaining (Fig. 2). In addition to cells with double immunostaining, cells with immunoreactivity only to NeuN are seen in the SNc on the days studied and remained until the end of the study.


Activation of GSK-3β and caspase-3 occurs in Nigral dopamine neurons during the development of apoptosis activated by a striatal injection of 6-hydroxydopamine.

Hernandez-Baltazar D, Mendoza-Garrido ME, Martinez-Fong D - PLoS ONE (2013)

Representative merged micrographs of mesencephalon immunostained against NeuN and TH.The numbers at the left margin of the figures are the days after 6-OHDA lesion. The primary antibodies were a mouse monoclonal anti-NeuN and a rabbit polyclonal anti-TH. The secondary antibodies were sheep anti-mouse IgG (H+L) FITC conjugated and goat anti-rabbit IgG (H+L) Texas conjugated. The rectangles on the panoramic view show the regions that were amplified 40x (details). Scale bar = 1 mm (central panels) and 50 µm (details).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3733721&req=5

pone-0070951-g002: Representative merged micrographs of mesencephalon immunostained against NeuN and TH.The numbers at the left margin of the figures are the days after 6-OHDA lesion. The primary antibodies were a mouse monoclonal anti-NeuN and a rabbit polyclonal anti-TH. The secondary antibodies were sheep anti-mouse IgG (H+L) FITC conjugated and goat anti-rabbit IgG (H+L) Texas conjugated. The rectangles on the panoramic view show the regions that were amplified 40x (details). Scale bar = 1 mm (central panels) and 50 µm (details).
Mentions: The studies of double immunofluorescence against TH and NeuN, a nuclear marker of neuronal lineage [48], showed that the loss of TH(+) cells was related to the disappearance of neuronal bodies after the 6-OHDA lesion. This was qualitatively shown in the medial region of the SNc. During the advance of a 6-OHDA lesion, there was a simultaneous loss of cells with TH and NeuN immunostaining (Fig. 2). In addition to cells with double immunostaining, cells with immunoreactivity only to NeuN are seen in the SNc on the days studied and remained until the end of the study.

Bottom Line: The disappearance of TH(+) cells was associated with a decrease in NeuN and β-III tubulin immunoreactivity and an increase in Apostain, cleaved caspase-3, and GSK-3β pY216 in the SNc.Increased levels of caspase-3 immunoreactivity in TH(+) cells were detected from days 1 to 15, and the levels then decreased to day 30 postlesion.Our results suggest that caspase-3 and GSK-3β pY216 activation might participate in the DA cell death and that the active caspase-3 might also participate in the neuroinflammation caused by the striatal 6-OHDA injection.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Fisiología, Biofísica y Neurociencias, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, México, DF, México.

ABSTRACT
The 6-Hydroxydopamine (6-OHDA) rat model of Parkinson's disease is essential for a better understanding of the pathological processes underlying the human disease and for the evaluation of promising therapeutic interventions. This work evaluated whether a single striatal injection of 6-OHDA causes progressive apoptosis of dopamine (DA) neurons and activation of glycogen synthase kinase 3β (GSK-3β) and caspase-3 in the substantia nigra compacta (SNc). The loss of DA neurons was shown by three neuron markers; tyrosine hydroxylase (TH), NeuN, and β-III tubulin. Apoptosis activation was determined using Apostain and immunostaining against cleaved caspase-3 and GSK-3β pY216. We also explored the possibility that cleaved caspase-3 is produced by microglia and astrocytes. Our results showed that the 6-OHDA caused loss of nigral TH(+) cells, progressing mainly in rostrocaudal and lateromedial directions. In the neostriatum, a severe loss of TH(+) terminals occurred from day 3 after lesion. The disappearance of TH(+) cells was associated with a decrease in NeuN and β-III tubulin immunoreactivity and an increase in Apostain, cleaved caspase-3, and GSK-3β pY216 in the SNc. Apostain immunoreactivity was observed from days 3 to 21 postlesion. Increased levels of caspase-3 immunoreactivity in TH(+) cells were detected from days 1 to 15, and the levels then decreased to day 30 postlesion. The cleaved caspase-3 also collocated with microglia and astrocytes indicating its participation in glial activation. Our results suggest that caspase-3 and GSK-3β pY216 activation might participate in the DA cell death and that the active caspase-3 might also participate in the neuroinflammation caused by the striatal 6-OHDA injection.

Show MeSH
Related in: MedlinePlus