The Wnts are a family of glycoproteins that regulate cell proliferation, fate decisions, and differentiation. In our study, we examined the contribution of Wnts to the development of ventral midbrain (VM) dopaminergic (DA) neurons. Our results show that ␤-catenin is expressed in DA precursor cells and that ␤-catenin signaling takes place in these cells, as assessed in TOPGAL [Tcf optimal-promoter ␤-galactosidase] reporter mice. We also found that Wnt-1, -3a, and -5a expression is differentially regulated during development and that partially purified Wnts distinctively regulate VM development. Wnt-3a promoted the proliferation of precursor cells expressing the orphan nuclear receptor-related factor 1 (Nurr1) but did not increase the number of tyrosine hydroxylase-positive neurons. Instead, Wnt-1 and -5a increased the number of rat midbrain DA neurons in rat embryonic day 14.5 precursor cultures by two distinct mechanisms. Wnt-1 predominantly increased the proliferation of Nurr1؉ precursors, up-regulated cyclins D1 and D3, and down-regulated p27 and p57 mRNAs. In contrast, Wnt-5a primarily increased the proportion of Nurr1؉ precursors that acquired a neuronal DA phenotype and up-regulated the expression of Ptx3 and c-ret mRNA. Moreover, the soluble cysteine-rich domain of Frizzled-8 (a Wnt inhibitor) blocked endogenous Wnts and the effects of Wnt-1 and -5a on proliferation and the acquisition of a DA phenotype in precursor cultures. These findings indicate that Wnts are key regulators of proliferation and differentiation of DA precursors during VM neurogenesis and that different Wnts have specific and unique activity profiles.T he development of midbrain dopaminergic (DA) neurons requires a complex combination of transcriptional regulators and diffusible signals to control both the acquisition and maintenance of a neurotransmitter-specific phenotype. The orphan nuclear receptor-related factor 1 (Nurr1, also known as NR4A2) is the only factor known to be required by midbrain precursor cells for the acquisition of a midbrain DA phenotype (1-4). Null mutations in other transcriptional regulators expressed in DA neurons, such as the homeodomain proteins Lmx1b and Ptx3, result in the loss of midbrain DA neurons after their birth (5-7). With regard to soluble diffusible signals, intersections of Shh (ventrally) and FGF8 (in the isthmus) create sites for the induction of DA neurons (8). Members of the Wnt family of secreted glycoproteins are also expressed in the midbrain (9) and are known to regulate precursor proliferation (10-12), fate decisions (13-17), and neuronal differentiation (18)(19)(20) in the nervous system. Interestingly, deletion of Wnt-1 results in the loss of DA neurons (21) and of the entire midbrain-hindbrain junction (22,23). Another mutant mouse with a similar phenotype in the midbrain is the LRP6 (low-density lipoprotein receptor-related protein 6) null (24), which lacks a receptor necessary for Wnt signaling. Combined, these findings suggest an important role for Wnts during the development of mid...

Proneural genes are crucial regulators of neurogenesis and subtype specification in many areas of the nervous system; however, their function in dopaminergic neuron development is unknown. We report that proneural genes have an intricate pattern of expression in the ventricular zone of the ventral midbrain, where mesencephalic dopaminergic neurons are generated. Neurogenin 2(Ngn2) and Mash1 are expressed in the ventral midline, while Ngn1, Ngn2 and Mash1 are co-localized more laterally in the ventricular zone. Ngn2 is also expressed in an intermediate zone immediately adjacent to the ventricular zone at the ventral midline. To examine the function of these genes, we analyzed mutant mice in which one or two of these genes were deleted (Ngn1, Ngn2 and Mash1) or substituted (Mash1 in the Ngn2 locus). Our results demonstrate that Ngn2 is required for the differentiation of Sox2+ ventricular zone progenitors into Nurr1+postmitotic dopaminergic neuron precursors in the intermediate zone, and that it is also likely to be required for their subsequent differentiation into tyrosine hydroxylase-positive dopaminergic neurons in the marginal zone. Although Mash1 normally has no detectable function in dopaminergic neuron development, it could partially rescue the generation of dopaminergic neuron precursors in the absence of Ngn2. These results demonstrate that Ngn2 is uniquely required for the development of midbrain dopaminergic neurons.

Nerve Growth Factor (NGF)-induced neuronal differentiation requires the activation of members of the Rho family of small GTPases. However, the molecular mechanisms through which NGF regulates cytoskeletal changes and neurite outgrowth are not totally understood. In this work, we identify the Rac1-specific guanine exchange factor (GEF) Tiam1 as a novel mediator of NGF/TrkA-dependent neurite elongation. In particular, we report that knockdown of Tiam1 causes a significant reduction in Rac1 activity and neurite outgrowth induced by NGF. Physical interaction between Tiam1 and active Ras (Ras-GTP), but not tyrosine phosphorylation of Tiam1, plays a central role in Rac1 activation by NGF. In addition, our findings indicate that Ras is required to associate Tiam1 with Rac1 and promote Rac1 activation upon NGF stimulation. Taken together, these findings define a novel molecular mechanism through which Tiam1 mediates TrkA signaling and neurite outgrowth induced by NGF.

Gene expression profiling identifies transcription factors involved in maturation of the blood-brain barrier.

VEGF-B ablation in pancreatic β-cells upregulates insulin expression without affecting glucose homeostasis or islet lipid uptake.

Secreted Frizzled related proteins (sFRPs) are a family of proteins that modulate Wnt signaling, which in turn regulates multiple aspects of ventral midbrain (VM) and dopamine (DA) neuron development. However, it is not known which Wnt signaling branch and what aspects of midbrain DA neuron development are regulated by sFRPs. Here, we show that sFRP1 and sFRP2 activate the Wnt/planar-cellpolarity/Rac1 pathway in DA cells. In the developing VM, sFRP1 and sFRP2 are expressed at low levels, and sFRP12/ 2 or sFRP22/2 mice had no detectable phenotype. However, compound sFRP12/2;sFRP22/2 mutants revealed a Wnt/PCP phenotype similar to that previously described for Wnt5a2/2 mice. This included an anteroposterior shortening of the VM, a lateral expansion of the Shh domain and DA lineage markers (Lmx1a and Th), as well as an accumulation of Nurr11 precursors in the VM. In vitro experiments showed that, while very high concentrations of SFRP1 had a negative effect on cell survival, low/medium concentrations of sFRP1 or sFRP2 promoted the DA differentiation of progenitors derived from primary VM cultures or mouse embryonic stem cells (ESCs), mimicking the effects of Wnt5a. We thus conclude that the main function of sFRP1 and sFRP2 is to enhance Wnt/PCP signaling in DA cells and to regulate Wnt/PCP-dependent functions in midbrain development. Moreover, we suggest that lowmedium concentrations of sFRPs may be used to enhance the DA differentiation of ESCs and improve their therapeutic application. STEM CELLS 2012;30:865-875 Disclosure of potential conflicts of interest is found at the end of this article.

No abstract

Highlights d Irradiation (IR) triggers transient microglial activation in the hippocampus d Microglia undergo a series of temporally regulated transcriptomic changes after IR d Single-cell RNA-seq reveals heterogeneous microglial populations after IR d Microglia upregulate pro-and anti-inflammatory genes simultaneously after IR