Common Origins of Hippocampal Ivy and Nitric Oxide Synthase Expressing Neurogliaform Cells

GABAergic interneurons critically regulate cortical computation through exquisite spatio-temporal control over excitatory networks. Precision of this inhibitory control requires a remarkable diversity within interneuron populations that is largely specified during embryogenesis. Although nNOS+ interneurons constitute the largest hippocampal interneuron cohort their origin and specification remain unknown. Thus, as neurogliaform (NGC) and Ivy cells (IvC) represent the main nNOS+ interneurons we investigated their developmental origins. Although considered distinct interneuron subtypes NGCs and IvCs exhibited similar neurochemical and electrophysiological signatures including NPY expression and late-spiking. Moreover, lineage analyses, including loss-of-function experiments and inducible fate-mapping, indicated that nNOS+ IvCs and NGCs are both derived from medial ganglionic eminence (MGE) progenitors under control of the transcription factor Nkx2-1. Surprisingly, a subset of NGCs lacking nNOS arises from caudal ganglionic eminence (CGE) progenitors. Thus, while nNOS+ NGCs and IvCs arise from MGE progenitors, a CGE origin distinguishes a discrete population of nNOS-NGCs

Characterization of Nkx6-2-Derived Neocortical Interneuron Lineages

Ventral telencephalic progenitors expressing the homeodomain transcription factor Nkx6-2 have been shown to give rise to a multitude of cortical interneuron subtypes usually associated with origin in either the medial ganglionic eminence or the caudal ganglionic eminence. The function of Nkx6-2 in directing the fate of those progenitors has, however, not been thoroughly analyzed. We used a combination of genetic inducible fate mapping and in vivo loss-of-function to analyze the requirement of Nkx6-2 in determining the fate of cortical interneurons. We have found that interneuron subtypes are born with a characteristic temporal pattern. Furthermore, we extend the characterization of interneurons from the Nkx6-2 lineage through the application of electrophysiological methods. Analysis of these populations in Nkx6-2 null mice suggests that there is a small and partially penetrant loss of delayed non-fast spiking somatostatin/calretinin double positive cortical interneurons in the absence of Nkx6-2 gene function

Experience-dependent MeCP2 expression in the excitatory cells of mouse visual thalamus

<div><p>Loss or gain of copy number of the gene encoding the transcription factor methyl-CpG-binding protein 2 (MeCP2) leads to neurodevelopmental disorders (Rett and MeCP2 duplication syndrome), indicating that precisely regulated MeCP2 expression during development is critical for mental health. Consistent with this idea, MeCP2 null mutants exhibit synaptic regression in the dorsal lateral geniculate nucleus (dLGN), the visual relay center in the thalamus, a phenotype resembling that of animals reared in the dark during the visual sensitive period. It remains unclear how MeCP2 expression is regulated during circuit formation and maturation, especially in excitatory and inhibitory populations of neurons. We found that, concomitant with the initiation of the dark-rearing sensitive period, MeCP2 protein levels were elevated in glutamatergic but not GABAergic neurons of the dLGN. Moreover, MeCP2 expression in glutamatergic populations was selectively reduced by dark-rearing. Therefore, we propose that visual experience–dependent MeCP2 induction in glutamatergic populations is essential for synaptic maturation within the dLGN.</p></div

Postnatal MeCP2 expression in excitatory and inhibitory dLGN neurons.

<p>MeCP2 level is dramatically increased in glutamatergic neurons of the dLGN during the visual sensitive period. Visual deprivation (dark-rearing) during the sensitive period (SP) or post-SP prevents MeCP2 up-regulation in glutamatergic neurons. MeCP2 expression in GABAergic neurons is stably maintained, and is not affected by visual deprivation.</p

MeCP2 expression in excitatory versus inhibitory cells of the developing dLGN.

<p><b>(A)</b> Identification of MeCP2-immunopositive cells (green) in the dLGN during development. Glutamatergic means GAD- / Nissl+ cells and GABAergic means GAD+ / Nissl+ cells. Red arrowheads, GABAergic neurons; blue arrowheads, glutamatergic neurons. Scale bar, 20 μm. <b>(B)</b> Changes in MeCP2-immunofluorescence intensity in GABAergic neurons during development. SP, sensitive period. P20: 123.4% ± 10.0% vs. P30: 137.6% ± 10.4%, P > 0.05. Statistical analysis for each developmental period was performed using the Steel-Dwass test. P10: 98 cells for 5 animals, P20: 56 cells for 6 animals. P30: 61 cells for 6 animals. P50: 44 cells for 5 animals. <b>(C)</b> Changes in MeCP2-immunofluorescence intensity in glutamatergic neurons during development. SP, sensitive period. P20: 118% ± 9.6% vs. P30: 168.3% ± 3.8%, **P < 0.01; Statistical analysis for each developmental period was performed using the Steel-Dwass test. NS, not significant. P10, 645 cells for 5 animals, P20: 738 cells for 6 animals. P30: 774 cells for 6 animals. P50: 463 cells for 5 animals. <b>(D)</b> The number of GABAergic and glutamatergic neuron in 10000 μm² decreased between P10 and P20. GABAergic neuron, P10: 1.01 ± 0.12 vs P20: 0.72 ± 0.17, *P < 0.05. Glutamatergic neuron, P10: 13.23 ± 0.66 vs. P20: 8.82±2.1, **P < 0.01. Statistical analysis for each developmental period was performed using the Steel-Dwass test. N = 9 slides for 3 animals. <b>(E)</b> Proportion of MeCP2+ cells increased among glutamataargic neurons (blue), but not among GABAergic neurons (red), before entering the SP. Glutamatergic neurons, P10: 22.8% ± 6.2% vs. P20: 83.1% ± 2.2%, **P < 0.01; Statistical analysis for each developmental period was performed using the Steel-Dwass test. NS, not significant. N = 9 slides for 3 animals.</p

Dark-rearing during the visual sensitive period decreases MeCP2 expression within the dLGN.

<p><b>(A)</b> Experimental design of dark-rearing and analysis. Dark-rearing (DR) was performed in each of the three developmental stages studied, and brains were collected at the end of each stage: Pre-SP, P11–P20; SP, P21–P30; and Post-SP, P41–P50. <b>(B)</b> Western blots for MeCP2 from dLGN of normally reared (control) and DR mice. <b>(C)</b> Quantification of MeCP2 expression levels. The MeCP2 level of each control sample was defined as 100%. Pre-SP: 92.0% ± 15.6% vs. normally reared control group, P > 0.05. SP: 54.2% ± 8.6% vs. control group, *P < 0.05. Post-SP: 74.5% ± 20.0% vs. control group, *P < 0.05; Wilcoxon test. NS, not significant. n = 6 samples for 18 animals. <b>(D)</b> Images of MeCP2 immunostaining in dLGN. Blue dotted lines, dLGN. Scale bar, 500 μm. <b>(E)</b> Quantification of MeCP2 immunofluorescence intensity in the dLGN. The MeCP2 level of each control sample was defined as 100%. SP: 85.3% ± 2.9% against control group, *P < 0.05; Statistical analysis for Ctrl vs DR was performed using the Wilcoxon test. NS, not significant. <b>(F)</b> Images of MeCP2 immunostaining in the retina from normally reared and DR mice. Scale bar, 50 μm. <b>(G)</b> Quantification of MeCP2 intensity in the GCL or INL in the retina. The MeCP2 level of each control sample was defined as 100%. ONL, outer nuclear layer; OPL, outer plexiform layer; INL, inner nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer. Statistical analysis for Ctrl vs DR was performed using the Wilcoxon test. NS, not significant Pre-SP, SP, Post-SP: n = 16 sections from 4 mice.</p

MeCP2 expression in glutamatergic dLGN neurons decreases upon dark-rearing.

<p><b>(A)</b> Distribution of MeCP2-immunopositive cells (green) in the dLGN of normally reared control and dark-reared (DR) mice. Red arrowhead, GABAergic neurons (GAD+, Nissl+); blue arrowhead, glutamatergic neurons (GAD-, Nissl+). Scale bar, 20 μm. <b>(B)</b> Immunofluorescence intensity of MeCP2 signals in GABAergic neurons of the dLGN in DR mice at different developmental stages. <b>(C)</b> Proportion of MeCP2-positive cells among GABAergic neurons. NS, not significant. <b>(D) I</b>mmunofluorescence intensity of MeCP2 in glutamatergic neurons in the dLGN of DR mice. SP: 84.9% ± 5.9% vs. control group, **P < 0.01; Post-SP: 87.5% ± 3.9% vs. control group, **P < 0.01. Statistical analysis for Ctrl vs DR was performed using the Wilcoxon test. <b>(E)</b> Proportion of MeCP2-positive cells among glutamatergic neurons. SP: 88.7% ± 1.1% vs. normally reared control group, *P < 0.05. Statistical analysis for Ctrl vs DR was performed using the Wilcoxon test.</p