The deep cerebellar nuclei (DCN) are the main output centers of the cerebellum, but little is known about their development. Using transcription factors as cell type-specific markers, we found that DCN neurons in mice are produced in the rhombic lip and migrate rostrally in a subpial stream to the nuclear transitory zone (NTZ). The rhombic lip-derived cells express transcription factors Pax6, Tbr2, and Tbr1 sequentially as they enter the NTZ. A subset of rhombic lip-derived cells also express reelin, a key regulator of Purkinje cell migrations. In organotypic slice cultures, the rhombic lip was necessary and sufficient to produce cells that migrate in the subpial stream, enter the NTZ, and express Pax6, Tbr2, Tbr1, and reelin. In later stages of development, the subpial stream is replaced by the external granular layer, and the NTZ organizes into distinct DCN nuclei. Tbr1 expression persists to adulthood in a subset of medial DCN projection neurons. In reeler mutant mice, which have a severe cerebellar malformation, rhombic lip-derived cells migrated to the NTZ, despite reelin deficiency. Studies in Tbr1 mutant mice suggested that Tbr1 plays a role in DCN morphogenesis but is not required for reelin expression, glutamatergic differentiation, or the initial formation of efferent axon pathways. Our findings reveal underlying similarities in the transcriptional programs for glutamatergic neuron production in the DCN and the cerebral cortex, and they support a model of cerebellar neurogenesis in which glutamatergic and GABAergic neurons are produced from separate progenitor compartments.

Inheritance of APOE alleles is associated with varying clinical outcomes in several neurodegenerative diseases that are associated with innate immune response in brain. We tested the hypothesis that inheritance of different APOE alleles would significantly modulate neurotoxicity arising from glial innate immune response. We first used dissociated cultures of wild-type (wt) murine neurons and glia derived from mice with targeted replacement (TR) of the epsilon2, epsilon3, or, epsilon4 APOE allele. Our results showed that the vast majority of bystander damage to wt neurons derived from microglia was greatest with TR APOE4 glia, intermediate from TR APOE3 glia, and least from TR APOE2 glia and preceded detectable NO secretion. Microglial p38MAPK-dependent cytokine secretion followed a similar pattern of TR APOE dependence. In hippocampal slice cultures, innate immune activation had a similar pattern of TR APOE-dependence and produced postsynaptic neuronal damage in TR APOE4 and TR APOE3 but not TR APOE2 cultures that was p38MAPK dependent. These findings suggest a new mechanism by which inheritance of different APOE alleles may influence the outcome of neurodegenerative diseases associated with microglial innate immune response.

The mechanisms that regulate the transition between the initial priming phase and DNA replication in liver regeneration are poorly understood. To study this transition, we compared events occurring after standard two-thirds partial hepatectomy, which elicits full regeneration, with response to a reduced hepatectomy, onethird partial hepatectomy (1/3PH), which leads to little DNA replication. Although the initial response to partial hepatectomy at the priming phase appeared to be similar between the two procedures, cell cycle progression was significantly blunted in 1/3PH mice. Among the main defects observed in 1/3PH mice were an almost complete deficiency in retinoblastoma phosphorylation and the lack of increase in kinase activity associated with cyclin E. We report that, in two-thirds partial hepatectomy mice, the expression of heparin-binding epidermal growth factor-like growth factor (HB-EGF) preceded the start of DNA replication and was not detectable in 1/3PH animals. Injection of HB-EGF into 1/3PH mice resulted in a >15-fold increase in DNA replication. Moreover, we show that hepatocyte DNA replication was delayed in HB-EGF knock-out mice. In summary, we show that HB-EGF is a key factor for hepatocyte progression through G 1 /S transition during liver regeneration.

Edited by Gerald W. HartViral infection is an exacerbating factor contributing to chronic airway diseases, such as asthma, via mechanisms that are still unclear. Polyinosine-polycytidylic acid (poly(I:C)), a Toll-like receptor 3 (TLR3) agonist used as a mimetic to study viral infection, has been shown to elicit inflammatory responses in lungs and to exacerbate pulmonary allergic reactions in animal models. Previously, we have shown that poly(I:C) stimulates lung fibroblasts to accumulate an extracellular matrix (ECM), enriched in hyaluronan (HA) and its binding partner versican, which promotes monocyte adhesion. In the current study, we aimed to determine the in vivo role of versican in mediating inflammatory responses in poly(I:C)-induced lung inflammation using a tamoxifen-inducible versican-deficient mouse model (Vcan ؊/؊ mice). In C57Bl/6 mice, poly(I:C) instillation significantly increased accumulation of versican and HA, especially in the perivascular and peribronchial regions, which were enriched in infiltrating leukocytes. In contrast, versican-deficient (Vcan ؊/؊ ) lungs did not exhibit increases in versican or HA in these regions and had strikingly reduced numbers of leukocytes in the bronchoalveolar lavage fluid and lower expression of inflammatory chemokines and cytokines. Poly(I:C) stimulation of lung fibroblasts isolated from control mice generated HA-enriched cable structures in the ECM, providing a substrate for monocytic cells in vitro, whereas lung fibroblasts from Vcan ؊/؊ mice did not. Moreover, increases in proinflammatory cytokine expression were also greatly attenuated in the Vcan ؊/؊ lung fibroblasts. These findings provide strong evidence that versican is a critical inflammatory mediator during poly(I:C)-induced acute lung injury and, in association with HA, generates an ECM that promotes leukocyte infiltration and adhesion.

Cerebral palsy is a common birth disorder that frequently involves ischemic-type injury to developing white matter (WM). Dead glial cells are a common feature of this injury and here we describe a novel form of acute ischemic cell death in developing WM astrocytes. Ischemia, modeled by the withdrawal of oxygen and glucose, evoked [Ca2+]i increases and cell death in astrocytes in post-natal day 10 (P10) rat optic nerve (RON). Removing extracellular Ca2+ prevented increases in [Ca2+]i but increased the amount of cell death. Astrocytes showed rapid [Na+]i increases during ischemia and cell death was reduced to control levels by substitution of extracellular Na+ or Cl- or by perfusion with bumetanide, a selective Na-K-Cl cotransport (NKCC) blocker. Astrocytes showed marked swelling during ischemia in the absence of extracellular Ca2+, which was blocked by bumetanide. Raising the extracellular osmolarity to limit water uptake reduced ischemic astrocyte death to control levels. Ultrastructural examination showed that post-ischemic astrocytes had lost their processes and frequently were necrotic, effects partially prevented by bumetanide. At this point in development, therefore, NKCC activation in astrocytes during ischemia produces an osmo-regulatory challenge. Astrocytes can subsequently regulate their cell volume in a Ca2+-dependent fashion but this will require ATP hydrolysis and does not protect the cells against acute cell death.