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Expression of mutant huntingtin in glial cells contributes to neuronal excitotoxicity.

Shin JY, Fang ZH, Yu ZX, Wang CE, Li SH, Li XJ - J. Cell Biol. (2005)

Bottom Line: Here, we report that mutant huntingtin accumulates in glial nuclei in HD brains and decreases the expression of glutamate transporters.Mutant htt in cultured astrocytes decreased their protection of neurons against glutamate excitotoxicity.These findings suggest that decreased glutamate uptake caused by glial mutant htt may critically contribute to neuronal excitotoxicity in HD.

View Article: PubMed Central - PubMed

Affiliation: Department of Human Genetics, Emory University School of Medicine, Atlanta, GA 30322, USA.

ABSTRACT
Huntington disease (HD) is characterized by the preferential loss of striatal medium-sized spiny neurons (MSNs) in the brain. Because MSNs receive abundant glutamatergic input, their vulnerability to excitotoxicity may be largely influenced by the capacity of glial cells to remove extracellular glutamate. However, little is known about the role of glia in HD neuropathology. Here, we report that mutant huntingtin accumulates in glial nuclei in HD brains and decreases the expression of glutamate transporters. As a result, mutant huntingtin (htt) reduces glutamate uptake in cultured astrocytes and HD mouse brains. In a neuron-glia coculture system, wild-type glial cells protected neurons against mutant htt-mediated neurotoxicity, whereas glial cells expressing mutant htt increased neuronal vulnerability. Mutant htt in cultured astrocytes decreased their protection of neurons against glutamate excitotoxicity. These findings suggest that decreased glutamate uptake caused by glial mutant htt may critically contribute to neuronal excitotoxicity in HD.

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Htt-mediated neurotoxicity in glia–neuron coculture. (A) Cultured cortical neurons were infected with adenoviral htt-23Q or htt-130Q for 24 h. The infected neurons were then cultured with or without wild-type glial cells or MK801 (10 μM) and labeled by antibodies to htt (top), MAP2 (middle), and Hoechst (bottom). Htt-130Q–infected neurons show decreased MAP2-staining (arrows). (B) The percentage of MAP2-positive neurons and apoptotic neurons with nuclear DNA fragmentation in the presence or absence of wild-type glial cells or MK801. (C) Cultured glial cells (4–6 wk) infected with adenoviral htt-23Q or htt-130Q were cocultured with wild-type cortical neurons. EM48 immunofluorescence staining of glia–neuron coculture shows that htt-23Q is distributed in the cytoplasm whereas htt-130Q accumulates in the nuclei (arrows) of infected glial cells. The size of nuclei of cultured glial cells is often larger than that of cultured cortical neurons. There is a decrease in the number of MAP2-positive neurons in the coculture with htt-130Q–infected glial cells. Nuclei were stained with Hoechst (blue). (D) The percentage of MAP2-positive neurons and apoptotic cells in the presence of adenoviral infected glial cells. Neurons were treated with or without MK801 (10 μM). The data (mean ± SEM) were obtained by counting the number of degenerated cells and the total number of nuclei per image. **, P < 0.01 compared with neurons cocultured with glial cells. Bars, 10 μm.
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fig8: Htt-mediated neurotoxicity in glia–neuron coculture. (A) Cultured cortical neurons were infected with adenoviral htt-23Q or htt-130Q for 24 h. The infected neurons were then cultured with or without wild-type glial cells or MK801 (10 μM) and labeled by antibodies to htt (top), MAP2 (middle), and Hoechst (bottom). Htt-130Q–infected neurons show decreased MAP2-staining (arrows). (B) The percentage of MAP2-positive neurons and apoptotic neurons with nuclear DNA fragmentation in the presence or absence of wild-type glial cells or MK801. (C) Cultured glial cells (4–6 wk) infected with adenoviral htt-23Q or htt-130Q were cocultured with wild-type cortical neurons. EM48 immunofluorescence staining of glia–neuron coculture shows that htt-23Q is distributed in the cytoplasm whereas htt-130Q accumulates in the nuclei (arrows) of infected glial cells. The size of nuclei of cultured glial cells is often larger than that of cultured cortical neurons. There is a decrease in the number of MAP2-positive neurons in the coculture with htt-130Q–infected glial cells. Nuclei were stained with Hoechst (blue). (D) The percentage of MAP2-positive neurons and apoptotic cells in the presence of adenoviral infected glial cells. Neurons were treated with or without MK801 (10 μM). The data (mean ± SEM) were obtained by counting the number of degenerated cells and the total number of nuclei per image. **, P < 0.01 compared with neurons cocultured with glial cells. Bars, 10 μm.

Mentions: To determine whether glia can protect against neuronal htt cytotoxicity, we infected cultured neurons with adenoviral GFP, htt-23Q, and htt-130Q constructs, and then cocultured them with wild-type astrocytes. In the absence of glial cells, htt-130Q neurons generally showed a decrease in the staining of microtubule-associated protein 2 (MAP2) (Fig. 8 A), a neuron-specific protein whose decrease reflects early neurodegeneration (Matesic and Lin, 1994). When htt-130Q neurons were cocultured with wild-type glial cells, the number of MAP2-positive neurons was significantly increased (Fig. 8 B). The NMDA receptor blocker MK801 (dizocilpine maleate), which increases the survival of cultured neurons (Driscoll et al., 1991), also increased the number of MAP2-positive neurons expressing htt-130Q (Fig. 8 B).


Expression of mutant huntingtin in glial cells contributes to neuronal excitotoxicity.

Shin JY, Fang ZH, Yu ZX, Wang CE, Li SH, Li XJ - J. Cell Biol. (2005)

Htt-mediated neurotoxicity in glia–neuron coculture. (A) Cultured cortical neurons were infected with adenoviral htt-23Q or htt-130Q for 24 h. The infected neurons were then cultured with or without wild-type glial cells or MK801 (10 μM) and labeled by antibodies to htt (top), MAP2 (middle), and Hoechst (bottom). Htt-130Q–infected neurons show decreased MAP2-staining (arrows). (B) The percentage of MAP2-positive neurons and apoptotic neurons with nuclear DNA fragmentation in the presence or absence of wild-type glial cells or MK801. (C) Cultured glial cells (4–6 wk) infected with adenoviral htt-23Q or htt-130Q were cocultured with wild-type cortical neurons. EM48 immunofluorescence staining of glia–neuron coculture shows that htt-23Q is distributed in the cytoplasm whereas htt-130Q accumulates in the nuclei (arrows) of infected glial cells. The size of nuclei of cultured glial cells is often larger than that of cultured cortical neurons. There is a decrease in the number of MAP2-positive neurons in the coculture with htt-130Q–infected glial cells. Nuclei were stained with Hoechst (blue). (D) The percentage of MAP2-positive neurons and apoptotic cells in the presence of adenoviral infected glial cells. Neurons were treated with or without MK801 (10 μM). The data (mean ± SEM) were obtained by counting the number of degenerated cells and the total number of nuclei per image. **, P < 0.01 compared with neurons cocultured with glial cells. Bars, 10 μm.
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Related In: Results  -  Collection

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fig8: Htt-mediated neurotoxicity in glia–neuron coculture. (A) Cultured cortical neurons were infected with adenoviral htt-23Q or htt-130Q for 24 h. The infected neurons were then cultured with or without wild-type glial cells or MK801 (10 μM) and labeled by antibodies to htt (top), MAP2 (middle), and Hoechst (bottom). Htt-130Q–infected neurons show decreased MAP2-staining (arrows). (B) The percentage of MAP2-positive neurons and apoptotic neurons with nuclear DNA fragmentation in the presence or absence of wild-type glial cells or MK801. (C) Cultured glial cells (4–6 wk) infected with adenoviral htt-23Q or htt-130Q were cocultured with wild-type cortical neurons. EM48 immunofluorescence staining of glia–neuron coculture shows that htt-23Q is distributed in the cytoplasm whereas htt-130Q accumulates in the nuclei (arrows) of infected glial cells. The size of nuclei of cultured glial cells is often larger than that of cultured cortical neurons. There is a decrease in the number of MAP2-positive neurons in the coculture with htt-130Q–infected glial cells. Nuclei were stained with Hoechst (blue). (D) The percentage of MAP2-positive neurons and apoptotic cells in the presence of adenoviral infected glial cells. Neurons were treated with or without MK801 (10 μM). The data (mean ± SEM) were obtained by counting the number of degenerated cells and the total number of nuclei per image. **, P < 0.01 compared with neurons cocultured with glial cells. Bars, 10 μm.
Mentions: To determine whether glia can protect against neuronal htt cytotoxicity, we infected cultured neurons with adenoviral GFP, htt-23Q, and htt-130Q constructs, and then cocultured them with wild-type astrocytes. In the absence of glial cells, htt-130Q neurons generally showed a decrease in the staining of microtubule-associated protein 2 (MAP2) (Fig. 8 A), a neuron-specific protein whose decrease reflects early neurodegeneration (Matesic and Lin, 1994). When htt-130Q neurons were cocultured with wild-type glial cells, the number of MAP2-positive neurons was significantly increased (Fig. 8 B). The NMDA receptor blocker MK801 (dizocilpine maleate), which increases the survival of cultured neurons (Driscoll et al., 1991), also increased the number of MAP2-positive neurons expressing htt-130Q (Fig. 8 B).

Bottom Line: Here, we report that mutant huntingtin accumulates in glial nuclei in HD brains and decreases the expression of glutamate transporters.Mutant htt in cultured astrocytes decreased their protection of neurons against glutamate excitotoxicity.These findings suggest that decreased glutamate uptake caused by glial mutant htt may critically contribute to neuronal excitotoxicity in HD.

View Article: PubMed Central - PubMed

Affiliation: Department of Human Genetics, Emory University School of Medicine, Atlanta, GA 30322, USA.

ABSTRACT
Huntington disease (HD) is characterized by the preferential loss of striatal medium-sized spiny neurons (MSNs) in the brain. Because MSNs receive abundant glutamatergic input, their vulnerability to excitotoxicity may be largely influenced by the capacity of glial cells to remove extracellular glutamate. However, little is known about the role of glia in HD neuropathology. Here, we report that mutant huntingtin accumulates in glial nuclei in HD brains and decreases the expression of glutamate transporters. As a result, mutant huntingtin (htt) reduces glutamate uptake in cultured astrocytes and HD mouse brains. In a neuron-glia coculture system, wild-type glial cells protected neurons against mutant htt-mediated neurotoxicity, whereas glial cells expressing mutant htt increased neuronal vulnerability. Mutant htt in cultured astrocytes decreased their protection of neurons against glutamate excitotoxicity. These findings suggest that decreased glutamate uptake caused by glial mutant htt may critically contribute to neuronal excitotoxicity in HD.

Show MeSH
Related in: MedlinePlus