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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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CNP-GFP+ postnatal hippocampal neurons are mostly GABAergic and receive functional synaptic inputs. Representative confocal images from CA3 (A–C) and dentate gyrus (D–F and G–I; merged images of 4–6 optical sections of 0.5–0.75 μm); three adjacent fields of the same P30 hippocampal slice showed the typical spatial distribution of CNP-GFP+ cells immunoreactive for GAD-67 (examples depicted by arrowheads). 0.5-μm thin single plane high magnification images of a single cell are shown in A–C as insets located in the lower right corners. All GAD-67+/CNP-GFP+ neurons displayed relatively low levels of GFP fluorescence. Bar, 50 μm for all panels A–I. (J) Continuous recording (total = 2.4 s) of spontaneous post-synaptic currents under voltage clamp (−60 mV) in a CNP-GFP+ neuron of the hilar dentate gyrus area (Scale: 0.5 s, 50 pA). (K) Synaptic activity of dentate gyrus CNP-GFP+ neurons was blocked by application of 1 μM TTX and 20 μM DNQX (Scale: 30 s, 50 pA). Spontaneous synaptic currents recorded in baseline conditions were compared with those of the same cell (bottom traces) 2 min after drug application.
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fig9: CNP-GFP+ postnatal hippocampal neurons are mostly GABAergic and receive functional synaptic inputs. Representative confocal images from CA3 (A–C) and dentate gyrus (D–F and G–I; merged images of 4–6 optical sections of 0.5–0.75 μm); three adjacent fields of the same P30 hippocampal slice showed the typical spatial distribution of CNP-GFP+ cells immunoreactive for GAD-67 (examples depicted by arrowheads). 0.5-μm thin single plane high magnification images of a single cell are shown in A–C as insets located in the lower right corners. All GAD-67+/CNP-GFP+ neurons displayed relatively low levels of GFP fluorescence. Bar, 50 μm for all panels A–I. (J) Continuous recording (total = 2.4 s) of spontaneous post-synaptic currents under voltage clamp (−60 mV) in a CNP-GFP+ neuron of the hilar dentate gyrus area (Scale: 0.5 s, 50 pA). (K) Synaptic activity of dentate gyrus CNP-GFP+ neurons was blocked by application of 1 μM TTX and 20 μM DNQX (Scale: 30 s, 50 pA). Spontaneous synaptic currents recorded in baseline conditions were compared with those of the same cell (bottom traces) 2 min after drug application.

Mentions: To determine whether CNP-GFP+ neurons found in the adult hippocampus may express phenotypic markers of functional neuronal differentiation, we performed immunohistochemical stainings for glutamate decarboxylase-67 (GAD-67), a γ-aminobutyric acid (GABA) synthesizing enzyme. In the hippocampus, GAD-67 is mostly expressed by GABAergic interneurons (Stone et al., 1999). We observed that a significant percentage of hippocampal adult CNP-GFP+ cells expressed GAD-67, and all displayed low levels of GFP fluorescence (Fig. 9 , A–I, arrowheads). At P30, these GAD-67+/CNP-GFP+ neurons were particularly concentrated in the inner granular zone of the dentate gyrus (Fig. 9, D–I, arrowheads), but few scattered cells were also seen in CA3 (Fig. 9, A–C, arrow and arrowheads) and CA1 areas. In the adult dentate gyrus (P30), a large proportion (∼90%) of NeuN+/CNP-GFP+ hippocampal neurons were GAD-67+, and all GAD-67+/CNP-GFP+ were found to be NeuN+ (unpublished data). Mature neurons expressing GAD-67 and low levels of GFP were consistently CNP protein-negative (Fig. S3). Even though keeping a detectable level of transcriptional activation of the CNP gene, these GABAergic neurons have thus completely down-regulated CNP protein expression.


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)

CNP-GFP+ postnatal hippocampal neurons are mostly GABAergic and receive functional synaptic inputs. Representative confocal images from CA3 (A–C) and dentate gyrus (D–F and G–I; merged images of 4–6 optical sections of 0.5–0.75 μm); three adjacent fields of the same P30 hippocampal slice showed the typical spatial distribution of CNP-GFP+ cells immunoreactive for GAD-67 (examples depicted by arrowheads). 0.5-μm thin single plane high magnification images of a single cell are shown in A–C as insets located in the lower right corners. All GAD-67+/CNP-GFP+ neurons displayed relatively low levels of GFP fluorescence. Bar, 50 μm for all panels A–I. (J) Continuous recording (total = 2.4 s) of spontaneous post-synaptic currents under voltage clamp (−60 mV) in a CNP-GFP+ neuron of the hilar dentate gyrus area (Scale: 0.5 s, 50 pA). (K) Synaptic activity of dentate gyrus CNP-GFP+ neurons was blocked by application of 1 μM TTX and 20 μM DNQX (Scale: 30 s, 50 pA). Spontaneous synaptic currents recorded in baseline conditions were compared with those of the same cell (bottom traces) 2 min after drug application.
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Related In: Results  -  Collection

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fig9: CNP-GFP+ postnatal hippocampal neurons are mostly GABAergic and receive functional synaptic inputs. Representative confocal images from CA3 (A–C) and dentate gyrus (D–F and G–I; merged images of 4–6 optical sections of 0.5–0.75 μm); three adjacent fields of the same P30 hippocampal slice showed the typical spatial distribution of CNP-GFP+ cells immunoreactive for GAD-67 (examples depicted by arrowheads). 0.5-μm thin single plane high magnification images of a single cell are shown in A–C as insets located in the lower right corners. All GAD-67+/CNP-GFP+ neurons displayed relatively low levels of GFP fluorescence. Bar, 50 μm for all panels A–I. (J) Continuous recording (total = 2.4 s) of spontaneous post-synaptic currents under voltage clamp (−60 mV) in a CNP-GFP+ neuron of the hilar dentate gyrus area (Scale: 0.5 s, 50 pA). (K) Synaptic activity of dentate gyrus CNP-GFP+ neurons was blocked by application of 1 μM TTX and 20 μM DNQX (Scale: 30 s, 50 pA). Spontaneous synaptic currents recorded in baseline conditions were compared with those of the same cell (bottom traces) 2 min after drug application.
Mentions: To determine whether CNP-GFP+ neurons found in the adult hippocampus may express phenotypic markers of functional neuronal differentiation, we performed immunohistochemical stainings for glutamate decarboxylase-67 (GAD-67), a γ-aminobutyric acid (GABA) synthesizing enzyme. In the hippocampus, GAD-67 is mostly expressed by GABAergic interneurons (Stone et al., 1999). We observed that a significant percentage of hippocampal adult CNP-GFP+ cells expressed GAD-67, and all displayed low levels of GFP fluorescence (Fig. 9 , A–I, arrowheads). At P30, these GAD-67+/CNP-GFP+ neurons were particularly concentrated in the inner granular zone of the dentate gyrus (Fig. 9, D–I, arrowheads), but few scattered cells were also seen in CA3 (Fig. 9, A–C, arrow and arrowheads) and CA1 areas. In the adult dentate gyrus (P30), a large proportion (∼90%) of NeuN+/CNP-GFP+ hippocampal neurons were GAD-67+, and all GAD-67+/CNP-GFP+ were found to be NeuN+ (unpublished data). Mature neurons expressing GAD-67 and low levels of GFP were consistently CNP protein-negative (Fig. S3). Even though keeping a detectable level of transcriptional activation of the CNP gene, these GABAergic neurons have thus completely down-regulated CNP protein expression.

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