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Cell surface heparan sulfate proteoglycan syndecan-2 induces the maturation of dendritic spines in rat hippocampal neurons.

Ethell IM, Yamaguchi Y - J. Cell Biol. (1999)

Bottom Line: We demonstrate that the cell surface heparan sulfate proteoglycan syndecan-2 plays a critical role in spine development.Deletion of the COOH-terminal EFYA motif of syndecan-2, the binding site for PDZ domain proteins, abrogates the spine-promoting activity of syndecan-2.Our results indicate that syndecan-2 plays a direct role in the development of postsynaptic specialization through its interactions with PDZ domain proteins.

View Article: PubMed Central - PubMed

Affiliation: The Burnham Institute, La Jolla, California 92037, USA.

ABSTRACT
Dendritic spines are small protrusions that receive synapses, and changes in spine morphology are thought to be the structural basis for learning and memory. We demonstrate that the cell surface heparan sulfate proteoglycan syndecan-2 plays a critical role in spine development. Syndecan-2 is concentrated at the synapses, specifically on the dendritic spines of cultured hippocampal neurons, and its accumulation occurs concomitant with the morphological maturation of spines from long thin protrusions to stubby and headed shapes. Early introduction of syndecan-2 cDNA into immature hippocampal neurons, by transient transfection, accelerates spine formation from dendritic protrusions. Deletion of the COOH-terminal EFYA motif of syndecan-2, the binding site for PDZ domain proteins, abrogates the spine-promoting activity of syndecan-2. Syndecan-2 clustering on dendritic protrusions does not require the PDZ domain-binding motif, but another portion of the cytoplasmic domain which includes a protein kinase C phosphorylation site. Our results indicate that syndecan-2 plays a direct role in the development of postsynaptic specialization through its interactions with PDZ domain proteins.

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Time course of  heparan sulfate expression  and maturation of dendritic  spines in hippocampal neurons at 1–4 wk in vitro. Hippocampal neurons were  stained with 10E4 anti–heparan sulfate mAb at 1 (A), 2  (B), 3 (C), and 4 (D) wk in  culture. Punctate pattern of  heparan sulfate immunoreactivity became detectable on  the surfaces of cell bodies  and dendrites at 3 wk in  vitro. (E and F) High-power  image of heparan sulfate immunoreactivity along dendrites at 1 and 4 wk in vitro,  correspondingly. (G and H)  Confocal images of proximal  dendrites in DiO-injected  hippocampal neurons at 1  (G) and 4 (H) wk in culture.  The dendritic protrusions at  1 wk in vitro are long, thin  filopodia without heads (G).  At 4 wk the majority of dendritic protrusions have mature mushroom shapes with  thin necks and large heads  (H). Bars, 20 μm in A–D and  3 μm in E–H. (I) Formation  of mature spines with thin  neck and a distinct head over  the course of 1–4 wk in culture. Percentage of spines with heads was counted in 1-, 2-, 3-,  and 4-wk-cultures. A more than twofold increase in number of  headed spines was seen between 2 and 3 wk in culture.
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Figure 1: Time course of heparan sulfate expression and maturation of dendritic spines in hippocampal neurons at 1–4 wk in vitro. Hippocampal neurons were stained with 10E4 anti–heparan sulfate mAb at 1 (A), 2 (B), 3 (C), and 4 (D) wk in culture. Punctate pattern of heparan sulfate immunoreactivity became detectable on the surfaces of cell bodies and dendrites at 3 wk in vitro. (E and F) High-power image of heparan sulfate immunoreactivity along dendrites at 1 and 4 wk in vitro, correspondingly. (G and H) Confocal images of proximal dendrites in DiO-injected hippocampal neurons at 1 (G) and 4 (H) wk in culture. The dendritic protrusions at 1 wk in vitro are long, thin filopodia without heads (G). At 4 wk the majority of dendritic protrusions have mature mushroom shapes with thin necks and large heads (H). Bars, 20 μm in A–D and 3 μm in E–H. (I) Formation of mature spines with thin neck and a distinct head over the course of 1–4 wk in culture. Percentage of spines with heads was counted in 1-, 2-, 3-, and 4-wk-cultures. A more than twofold increase in number of headed spines was seen between 2 and 3 wk in culture.

Mentions: Heparan sulfate immunoreactivity has been found previously in adult rat hippocampus (Goedert et al., 1996; Fuxe et al., 1997). To define the precise localization of heparan sulfate on neuronal cell surfaces, we used glia-free monolayer cultures of E 17–18 rat hippocampal neurons. In these cultures, neurons form synapses and establish neuronal circuits in the course of 3–4 wk in vitro. Immunofluorescent staining for heparan sulfate was performed on these neurons at different stages in culture with the 10E4 mAb that recognizes intact heparan sulfate chains (David et al., 1992; Goedert et al., 1996). The expression of heparan sulfate is weak and diffuse during the first 2 wk in culture (Fig. 1, A and B), but then increases during the following weeks. At 3 wk in vitro, heparan sulfate immunoreactivity was detectable as punctate signals distributed on the cell bodies and dendrites (Fig. 1 C). The punctate staining became even stronger at 4 wk in vitro (Fig. 1, D and F). This timing of heparan sulfate expression temporally coincided with the widespread formation of dendritic spines (Fig. 1, E–H). At 1 wk in vitro, when the majority of dendritic protrusions were long, thin filopodia (Fig. 1 G), heparan sulfate immunostaining was very weak and did not show any distinct pattern of distribution (Fig. 1 E). By 4 wk in vitro, the majority of postsynaptic sites developed into stubby or mushroom-shaped mature spines (Fig. 1 H), morphologically similar to the spines seen in vivo. At the same time, strong heparan sulfate immunoreactivity was detected as puncta along the dendrites (Fig. 1 F). These results suggested that heparan sulfate may be associated with dendritic spines.


Cell surface heparan sulfate proteoglycan syndecan-2 induces the maturation of dendritic spines in rat hippocampal neurons.

Ethell IM, Yamaguchi Y - J. Cell Biol. (1999)

Time course of  heparan sulfate expression  and maturation of dendritic  spines in hippocampal neurons at 1–4 wk in vitro. Hippocampal neurons were  stained with 10E4 anti–heparan sulfate mAb at 1 (A), 2  (B), 3 (C), and 4 (D) wk in  culture. Punctate pattern of  heparan sulfate immunoreactivity became detectable on  the surfaces of cell bodies  and dendrites at 3 wk in  vitro. (E and F) High-power  image of heparan sulfate immunoreactivity along dendrites at 1 and 4 wk in vitro,  correspondingly. (G and H)  Confocal images of proximal  dendrites in DiO-injected  hippocampal neurons at 1  (G) and 4 (H) wk in culture.  The dendritic protrusions at  1 wk in vitro are long, thin  filopodia without heads (G).  At 4 wk the majority of dendritic protrusions have mature mushroom shapes with  thin necks and large heads  (H). Bars, 20 μm in A–D and  3 μm in E–H. (I) Formation  of mature spines with thin  neck and a distinct head over  the course of 1–4 wk in culture. Percentage of spines with heads was counted in 1-, 2-, 3-,  and 4-wk-cultures. A more than twofold increase in number of  headed spines was seen between 2 and 3 wk in culture.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2132915&req=5

Figure 1: Time course of heparan sulfate expression and maturation of dendritic spines in hippocampal neurons at 1–4 wk in vitro. Hippocampal neurons were stained with 10E4 anti–heparan sulfate mAb at 1 (A), 2 (B), 3 (C), and 4 (D) wk in culture. Punctate pattern of heparan sulfate immunoreactivity became detectable on the surfaces of cell bodies and dendrites at 3 wk in vitro. (E and F) High-power image of heparan sulfate immunoreactivity along dendrites at 1 and 4 wk in vitro, correspondingly. (G and H) Confocal images of proximal dendrites in DiO-injected hippocampal neurons at 1 (G) and 4 (H) wk in culture. The dendritic protrusions at 1 wk in vitro are long, thin filopodia without heads (G). At 4 wk the majority of dendritic protrusions have mature mushroom shapes with thin necks and large heads (H). Bars, 20 μm in A–D and 3 μm in E–H. (I) Formation of mature spines with thin neck and a distinct head over the course of 1–4 wk in culture. Percentage of spines with heads was counted in 1-, 2-, 3-, and 4-wk-cultures. A more than twofold increase in number of headed spines was seen between 2 and 3 wk in culture.
Mentions: Heparan sulfate immunoreactivity has been found previously in adult rat hippocampus (Goedert et al., 1996; Fuxe et al., 1997). To define the precise localization of heparan sulfate on neuronal cell surfaces, we used glia-free monolayer cultures of E 17–18 rat hippocampal neurons. In these cultures, neurons form synapses and establish neuronal circuits in the course of 3–4 wk in vitro. Immunofluorescent staining for heparan sulfate was performed on these neurons at different stages in culture with the 10E4 mAb that recognizes intact heparan sulfate chains (David et al., 1992; Goedert et al., 1996). The expression of heparan sulfate is weak and diffuse during the first 2 wk in culture (Fig. 1, A and B), but then increases during the following weeks. At 3 wk in vitro, heparan sulfate immunoreactivity was detectable as punctate signals distributed on the cell bodies and dendrites (Fig. 1 C). The punctate staining became even stronger at 4 wk in vitro (Fig. 1, D and F). This timing of heparan sulfate expression temporally coincided with the widespread formation of dendritic spines (Fig. 1, E–H). At 1 wk in vitro, when the majority of dendritic protrusions were long, thin filopodia (Fig. 1 G), heparan sulfate immunostaining was very weak and did not show any distinct pattern of distribution (Fig. 1 E). By 4 wk in vitro, the majority of postsynaptic sites developed into stubby or mushroom-shaped mature spines (Fig. 1 H), morphologically similar to the spines seen in vivo. At the same time, strong heparan sulfate immunoreactivity was detected as puncta along the dendrites (Fig. 1 F). These results suggested that heparan sulfate may be associated with dendritic spines.

Bottom Line: We demonstrate that the cell surface heparan sulfate proteoglycan syndecan-2 plays a critical role in spine development.Deletion of the COOH-terminal EFYA motif of syndecan-2, the binding site for PDZ domain proteins, abrogates the spine-promoting activity of syndecan-2.Our results indicate that syndecan-2 plays a direct role in the development of postsynaptic specialization through its interactions with PDZ domain proteins.

View Article: PubMed Central - PubMed

Affiliation: The Burnham Institute, La Jolla, California 92037, USA.

ABSTRACT
Dendritic spines are small protrusions that receive synapses, and changes in spine morphology are thought to be the structural basis for learning and memory. We demonstrate that the cell surface heparan sulfate proteoglycan syndecan-2 plays a critical role in spine development. Syndecan-2 is concentrated at the synapses, specifically on the dendritic spines of cultured hippocampal neurons, and its accumulation occurs concomitant with the morphological maturation of spines from long thin protrusions to stubby and headed shapes. Early introduction of syndecan-2 cDNA into immature hippocampal neurons, by transient transfection, accelerates spine formation from dendritic protrusions. Deletion of the COOH-terminal EFYA motif of syndecan-2, the binding site for PDZ domain proteins, abrogates the spine-promoting activity of syndecan-2. Syndecan-2 clustering on dendritic protrusions does not require the PDZ domain-binding motif, but another portion of the cytoplasmic domain which includes a protein kinase C phosphorylation site. Our results indicate that syndecan-2 plays a direct role in the development of postsynaptic specialization through its interactions with PDZ domain proteins.

Show MeSH
Related in: MedlinePlus