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Postnatal NG2 proteoglycan-expressing progenitor cells are intrinsically multipotent and generate functional neurons.

Belachew S, Chittajallu R, Aguirre AA, Yuan X, Kirby M, Anderson S, Gallo V - J. Cell Biol. (2003)

Bottom Line: The fast kinetics and the high rate of multipotent fate of these NG2+ progenitors in vitro reflect an intrinsic property, rather than reprogramming.We demonstrate in the hippocampus in vivo that a sizeable fraction of postnatal NG2+ progenitor cells are proliferative precursors whose progeny appears to differentiate into GABAergic neurons capable of propagating action potentials and displaying functional synaptic inputs.These data show that at least a subpopulation of postnatal NG2-expressing cells are CNS multipotent precursors that may underlie adult hippocampal neurogenesis.

View Article: PubMed Central - PubMed

Affiliation: Center for Neuroscience Research, Children's Research Institute, Children's National Medical Center, Washington, DC 20010-2970, USA.

ABSTRACT
Neurogenesis is known to persist in the adult mammalian central nervous system (CNS). The identity of the cells that generate new neurons in the postnatal CNS has become a crucial but elusive issue. Using a transgenic mouse, we show that NG2 proteoglycan-positive progenitor cells that express the 2',3'-cyclic nucleotide 3'-phosphodiesterase gene display a multipotent phenotype in vitro and generate electrically excitable neurons, as well as astrocytes and oligodendrocytes. The fast kinetics and the high rate of multipotent fate of these NG2+ progenitors in vitro reflect an intrinsic property, rather than reprogramming. We demonstrate in the hippocampus in vivo that a sizeable fraction of postnatal NG2+ progenitor cells are proliferative precursors whose progeny appears to differentiate into GABAergic neurons capable of propagating action potentials and displaying functional synaptic inputs. These data show that at least a subpopulation of postnatal NG2-expressing cells are CNS multipotent precursors that may underlie adult hippocampal neurogenesis.

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The existence of NG2+/TOAD-64+/CNP-GFP+ immature hippocampal neurons establishes a lineage continuum between NG2+/CNP-GFP+ progenitor cells and NG2−/NeuN+/CNP-GFP+ neurons in the adult dentate gyrus. Confocal photomicrographs (merged images from 4–6 optical sections of 0.5 μm each) of three representative fields of P30 hippocampal slices in which CNP-GFP+ cells were immunolabeled for either TOAD-64 and NeuN (A–D, dentate gyrus), TOAD-64 and NG2 (E–H, dentate gyrus), or NeuN and NG2 (I–L, CA3). 0.5-μm thin single plane high magnification images of single cells are shown for each panel as insets located in the upper right corners (A–L). In A–D and E–H, white arrows point to NG2+/TOAD-64+/CNP-GFP+ early post-mitotic neurons that expressed levels of GFP higher than differentiated NG2−/NeuN+/CNP-GFP+ neurons (A–D, arrowheads; I–L, arrows). Red arrows in A–D and E–H depict TOAD-64+ neuronal cells that did not express CNP-GFP. Bars, 50 μm for all panels.
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fig8: The existence of NG2+/TOAD-64+/CNP-GFP+ immature hippocampal neurons establishes a lineage continuum between NG2+/CNP-GFP+ progenitor cells and NG2−/NeuN+/CNP-GFP+ neurons in the adult dentate gyrus. Confocal photomicrographs (merged images from 4–6 optical sections of 0.5 μm each) of three representative fields of P30 hippocampal slices in which CNP-GFP+ cells were immunolabeled for either TOAD-64 and NeuN (A–D, dentate gyrus), TOAD-64 and NG2 (E–H, dentate gyrus), or NeuN and NG2 (I–L, CA3). 0.5-μm thin single plane high magnification images of single cells are shown for each panel as insets located in the upper right corners (A–L). In A–D and E–H, white arrows point to NG2+/TOAD-64+/CNP-GFP+ early post-mitotic neurons that expressed levels of GFP higher than differentiated NG2−/NeuN+/CNP-GFP+ neurons (A–D, arrowheads; I–L, arrows). Red arrows in A–D and E–H depict TOAD-64+ neuronal cells that did not express CNP-GFP. Bars, 50 μm for all panels.

Mentions: In the dentate gyrus of adult (P30) hippocampus, we observed that some CNP-GFP+ cells also expressed TOAD-64 (turned on after division, 64 kD) protein (Fig. 8 , A–H), a transient marker of early post-mitotic neurons that is expressed before most neuron-specific proteins during neurogenesis (Minturn et al., 1995a, 1995b; Cameron and McKay, 2001). TOAD-64 is also referred to as collapsin response-mediated protein 4 (Seki, 2002). In P30 dentate gyrus and CA3 areas, 11.3 ± 3.8% of CNP-GFP+ cells were TOAD-64+ (mean ± SEM, n = 244 total counted cells, two independent experiments). Hippocampal TOAD-64+/CNP-GFP+ immature neurons (Fig. 8, A–H, white arrows) reliably expressed levels of GFP fluorescence higher than NeuN+/CNP-GFP+-differentiated neurons (Fig. 8, A–D, arrowheads), consistent with the hypothesis that a gradual loss of CNP gene expression correlates with neuronal differentiation of CNP-GFP+ cells. Moreover, the vast majority of TOAD-64+/CNP-GFP+ immature neurons were found to be NG2+ (90%, total cells counted = 262) in the adult dentate gyrus (Fig. 8, E–H, white arrows), whereas NeuN+/CNP-GFP+ neurons were NG2− (Fig. 8, I–L, arrows), except for a small percentage of cells (5%; total counted cells = 754). However, in these cells, NG2 staining appeared ambiguous and suggested either a low level of NG2 proteoglycan expression or a close enwrapping of the cells within an intricate network of NG2+ processes from neighboring CNP-GFP+ cells (Fig. 8, I–L, arrowhead). Colocalization of TOAD-64 and NeuN expression in CNP-GFP+ cells was only observed in cells displaying a weak nuclear pattern of TOAD-64 expression (Fig. 8, A–D, white arrowheads) different from the classical perinuclear distribution of this protein (Fig. 8 B, white arrows). We could also visualize TOAD-64+ adult hippocampal neurons that were CNP-GFP− (Fig. 8, A–H, red arrows) and consistently NG2− (Fig. 8, E–H, red arrows).


Postnatal NG2 proteoglycan-expressing progenitor cells are intrinsically multipotent and generate functional neurons.

Belachew S, Chittajallu R, Aguirre AA, Yuan X, Kirby M, Anderson S, Gallo V - J. Cell Biol. (2003)

The existence of NG2+/TOAD-64+/CNP-GFP+ immature hippocampal neurons establishes a lineage continuum between NG2+/CNP-GFP+ progenitor cells and NG2−/NeuN+/CNP-GFP+ neurons in the adult dentate gyrus. Confocal photomicrographs (merged images from 4–6 optical sections of 0.5 μm each) of three representative fields of P30 hippocampal slices in which CNP-GFP+ cells were immunolabeled for either TOAD-64 and NeuN (A–D, dentate gyrus), TOAD-64 and NG2 (E–H, dentate gyrus), or NeuN and NG2 (I–L, CA3). 0.5-μm thin single plane high magnification images of single cells are shown for each panel as insets located in the upper right corners (A–L). In A–D and E–H, white arrows point to NG2+/TOAD-64+/CNP-GFP+ early post-mitotic neurons that expressed levels of GFP higher than differentiated NG2−/NeuN+/CNP-GFP+ neurons (A–D, arrowheads; I–L, arrows). Red arrows in A–D and E–H depict TOAD-64+ neuronal cells that did not express CNP-GFP. Bars, 50 μm for all panels.
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fig8: The existence of NG2+/TOAD-64+/CNP-GFP+ immature hippocampal neurons establishes a lineage continuum between NG2+/CNP-GFP+ progenitor cells and NG2−/NeuN+/CNP-GFP+ neurons in the adult dentate gyrus. Confocal photomicrographs (merged images from 4–6 optical sections of 0.5 μm each) of three representative fields of P30 hippocampal slices in which CNP-GFP+ cells were immunolabeled for either TOAD-64 and NeuN (A–D, dentate gyrus), TOAD-64 and NG2 (E–H, dentate gyrus), or NeuN and NG2 (I–L, CA3). 0.5-μm thin single plane high magnification images of single cells are shown for each panel as insets located in the upper right corners (A–L). In A–D and E–H, white arrows point to NG2+/TOAD-64+/CNP-GFP+ early post-mitotic neurons that expressed levels of GFP higher than differentiated NG2−/NeuN+/CNP-GFP+ neurons (A–D, arrowheads; I–L, arrows). Red arrows in A–D and E–H depict TOAD-64+ neuronal cells that did not express CNP-GFP. Bars, 50 μm for all panels.
Mentions: In the dentate gyrus of adult (P30) hippocampus, we observed that some CNP-GFP+ cells also expressed TOAD-64 (turned on after division, 64 kD) protein (Fig. 8 , A–H), a transient marker of early post-mitotic neurons that is expressed before most neuron-specific proteins during neurogenesis (Minturn et al., 1995a, 1995b; Cameron and McKay, 2001). TOAD-64 is also referred to as collapsin response-mediated protein 4 (Seki, 2002). In P30 dentate gyrus and CA3 areas, 11.3 ± 3.8% of CNP-GFP+ cells were TOAD-64+ (mean ± SEM, n = 244 total counted cells, two independent experiments). Hippocampal TOAD-64+/CNP-GFP+ immature neurons (Fig. 8, A–H, white arrows) reliably expressed levels of GFP fluorescence higher than NeuN+/CNP-GFP+-differentiated neurons (Fig. 8, A–D, arrowheads), consistent with the hypothesis that a gradual loss of CNP gene expression correlates with neuronal differentiation of CNP-GFP+ cells. Moreover, the vast majority of TOAD-64+/CNP-GFP+ immature neurons were found to be NG2+ (90%, total cells counted = 262) in the adult dentate gyrus (Fig. 8, E–H, white arrows), whereas NeuN+/CNP-GFP+ neurons were NG2− (Fig. 8, I–L, arrows), except for a small percentage of cells (5%; total counted cells = 754). However, in these cells, NG2 staining appeared ambiguous and suggested either a low level of NG2 proteoglycan expression or a close enwrapping of the cells within an intricate network of NG2+ processes from neighboring CNP-GFP+ cells (Fig. 8, I–L, arrowhead). Colocalization of TOAD-64 and NeuN expression in CNP-GFP+ cells was only observed in cells displaying a weak nuclear pattern of TOAD-64 expression (Fig. 8, A–D, white arrowheads) different from the classical perinuclear distribution of this protein (Fig. 8 B, white arrows). We could also visualize TOAD-64+ adult hippocampal neurons that were CNP-GFP− (Fig. 8, A–H, red arrows) and consistently NG2− (Fig. 8, E–H, red arrows).

Bottom Line: The fast kinetics and the high rate of multipotent fate of these NG2+ progenitors in vitro reflect an intrinsic property, rather than reprogramming.We demonstrate in the hippocampus in vivo that a sizeable fraction of postnatal NG2+ progenitor cells are proliferative precursors whose progeny appears to differentiate into GABAergic neurons capable of propagating action potentials and displaying functional synaptic inputs.These data show that at least a subpopulation of postnatal NG2-expressing cells are CNS multipotent precursors that may underlie adult hippocampal neurogenesis.

View Article: PubMed Central - PubMed

Affiliation: Center for Neuroscience Research, Children's Research Institute, Children's National Medical Center, Washington, DC 20010-2970, USA.

ABSTRACT
Neurogenesis is known to persist in the adult mammalian central nervous system (CNS). The identity of the cells that generate new neurons in the postnatal CNS has become a crucial but elusive issue. Using a transgenic mouse, we show that NG2 proteoglycan-positive progenitor cells that express the 2',3'-cyclic nucleotide 3'-phosphodiesterase gene display a multipotent phenotype in vitro and generate electrically excitable neurons, as well as astrocytes and oligodendrocytes. The fast kinetics and the high rate of multipotent fate of these NG2+ progenitors in vitro reflect an intrinsic property, rather than reprogramming. We demonstrate in the hippocampus in vivo that a sizeable fraction of postnatal NG2+ progenitor cells are proliferative precursors whose progeny appears to differentiate into GABAergic neurons capable of propagating action potentials and displaying functional synaptic inputs. These data show that at least a subpopulation of postnatal NG2-expressing cells are CNS multipotent precursors that may underlie adult hippocampal neurogenesis.

Show MeSH
Related in: MedlinePlus