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  • Formation of perineuronal nets in organotypic mouse brain slice cultures is independent of neuronal glutamatergic activity. 17561838

    Perineuronal nets (PNs) are a specialized form of the extracellular matrix and cover specific sets of neurons in distinct brain areas. Animal experiments on sensory visual deprivation have demonstrated that the generation of PNs around neurons of the visual cortex is dependent on neuronal activity during the critical period of visual experience. The importance of the activity of specific neurotransmitter systems for PN formation has, however, not yet been demonstrated. Based on the predominantly glutamatergic innervation of the visual cortex we hypothesized that reduced glutamatergic activity impairs the development of PNs. To address this question, genetic mouse models with compromised glutamate release [Munc13-1-knockout (KO) and Munc13-1/2 double-KO (DKO)] and chronic pharmacological treatments interfering with specific steps of glutamatergic transmission were used. Under experimental conditions of glutamatergic hypofunction PN formation was studied in organotypic brain slice cultures with Wisteria floribunda lectin binding and with aggrecan immunohistochemistry. After cultivation for 21 days a regular PN formation was observed in brain slices (i) derived from Munc13-1-KO and Munc13-1/2-DKO mice, (ii) after blockade of metabotropic and ionotropic glutamate receptors with MCPG and kynurenate, and (iii) after suppression of glutamate release by blockade of presynaptic Ca++ channels with riluzole. Nonselective suppression of neuronal activity by blockade of voltage-gated sodium channels with tetrodotoxin clearly inhibited PN formation. These results indicate that neuronal activity is required but that the glutamatergic system is not essential for PN development.
    Tipo de documento:
    Referencia
    Referencia del producto:
    AB1031
    Nombre del producto:
    Anti-Aggrecan Antibody
  • Hepatocyte growth factor favors monocyte differentiation into regulatory interleukin (IL)-10++IL-12low/neg accessory cells with dendritic-cell features. 16527888

    Several hematopoietic growth factors, including interleukin-10 (IL-10) and transforming growth factor-beta1 (TGF-beta1), promote the differentiation of tolerogenic dendritic cells (DCs). Hepatocyte growth factor (HGF) is a pleiotropic cytokine whose effects on human DC differentiation and function have not been investigated. Monocytes cultured with HGF (HGFMo) differentiated into accessory cells with DC-like morphology, released low amounts of IL-12p70 and up-regulated IL-10 both at the mRNA and at the protein level. Upon activation with HGFMo, allogeneic CD4+CD25- T cells expressed the T regulatory (Treg)-associated transcription factor FoxP3, proliferated poorly, and released high levels of IL-10. Interestingly, blockade of surface immunoglobulin-like transcript 3 (ILT3) on HGFMo or neutralization of secreted IL-10 translated into partial restoration of T-cell proliferation. Secondary stimulation of HGFMo-primed CD4+ T cells with immunogenic DCs differentiated with granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-4 from monocytes of the same donor resulted in measurable T-cell proliferation. HGFMo-primed CD4+ T cells significantly inhibited the proliferation of naive CD4+CD25- T cells in a cell-contact-dependent manner. Finally, DNA microarray analysis revealed a unique gene-expression profile of HGF-activated monocytes. Collectively, our findings point to a novel role for HGF in the regulation of monocyte/DC functions that might be exploited therapeutically.
    Tipo de documento:
    Referencia
    Referencia del producto:
    MAB5412
    Nombre del producto:
    Anti-Indoleamine 2,3-dioxygenase Antibody, clone 10.1
  • Brain-derived neurotrophic factor mediates activity-dependent dendritic growth in nonpyramidal neocortical interneurons in developing organotypic cultures. 12843269

    Brain-derived neurotrophic factor (BDNF) promotes postnatal maturation of GABAergic inhibition in the cerebral and cerebellar cortices, and its expression and release are enhanced by neuronal activity, suggesting that it acts in a feedback manner to maintain a balance between excitation and inhibition during development. BDNF promotes differentiation of cerebellar, hippocampal, and neostriatal inhibitory neurons, but its effects on the dendritic development of neocortical inhibitory interneurons remain unknown. Here, we show that BDNF mediates depolarization-induced dendritic growth and branching in neocortical interneurons. To visualize inhibitory interneurons, we biolistically transfected organotypic cortical slice cultures from neonatal mice with green fluorescent protein (GFP) driven by the glutamic acid decarboxylase (GAD)67 promoter. Nearly all GAD67-GFP-expressing neurons were nonpyramidal, many contained GABA, and some expressed markers of neurochemically defined GABAergic subtypes, indicating that GAD67-GFP-expressing neurons were GABAergic. We traced dendritic trees from confocal images of the same GAD67-GFP-expressing neurons before and after a 5 d growth period, and quantified the change in total dendritic length (TDL) and total dendritic branch points (TDBPs) for each neuron. GAD67-GFP-expressing neurons growing in control medium exhibited a 20% increase in TDL, but in 200 ng/ml BDNF or 10 mm KCl, this increase nearly doubled and was accompanied by a significant increase in TDBPs. Blocking action potentials with TTX did not prevent the BDNF-induced growth, but antibodies against BDNF blocked the growth-promoting effect of KCl. We conclude that BDNF, released by neocortical pyramidal neurons in response to depolarization, enhances dendritic growth and branching in nearby inhibitory interneurons.
    Tipo de documento:
    Referencia
    Referencia del producto:
    AB1778
  • A Critical role for Bim in retinal ganglion cell death. 17442051

    Optic nerve transection results in the death of retinal ganglion cells (RGCs) by apoptosis. Apoptosis is regulated by the Bcl-2 family of proteins, of which the Bcl-2 homology (BH3) -only proteins forms a subset. As BH3-only proteins have been shown to play a significant role in regulating cell death in the central nervous system, we wished to investigate the role of Bcl-2 interacting mediator of cell death (Bim), a prominent member of this protein family in the regulation of cell death in the RGC layer using in vitro retinal explants. In this study, we use an innovative retinal shaving procedure to isolate the cells of the ganglion cell layer to use for western blotting. Members of the BH3-only protein family are down-regulated during retinal development and are not normally expressed in the adult retina. Using this procedure, we demonstrate that Bim is re-expressed and its expression is increased over time following axotomy. Expression of Bad and Bik decreases over the same time course, whereas there is no indication that Bid and Puma are re-expressed. We show that explants from Bim knockout mice are resistant to axotomy-induced death when compared with their wild-type counterparts. Genetic deletion of Bim also prevents caspase 3 cleavage. The activity of Bim can be negatively regulated by phosphorylation. We show that the decrease of Bim phosphorylation correlates with a decrease in expression of survival kinases such as pAkt and pERK over the same time course. These results implicate Bim re-expression as being essential for axotomy-induced death of RGCs and that phosphorylation of Bim negatively regulates its activity in RGCs.
    Tipo de documento:
    Referencia
    Referencia del producto:
    AB9016
    Nombre del producto:
    Anti-Chx10 Antibody, NT