RESUMEN
In the developing cerebellum, the nascent white matter (WM) serves as an instructive niche for cerebellar cortical inhibitory interneurons. As their Pax2 expressing precursors transit the emerging WM, their laminar fate is programmed. The source(s) and nature of the signals involved remain unknown. Here, we used immunocytochemistry to follow the cellular maturation of the murine cerebellar WM during this critical period. During the first few days of postnatal development, when most Pax2 expressing cells are formed and many of them reach the cerebellar gray matter, only microglial cells can be identified in the territories through which Pax2 cells migrate. From p4 onward, cells expressing the oligodendrocytic or astrocyte markers, CNP-1, MBP or GFAP, started to appear in the nascent WM. Expression of macroglial markers increased with cerebellar differentiation, yet deep nuclei remained GFAP-negative at all ages. The progressive spread of maturing glia did not correlate with the exit of Pax2 cells from the WM, as indicated by the extensive mingling of these cells up to p15. Whereas sonic hedgehog-associated p75NTR expression could be verified in granule cell precursors, postmitotic Pax2 cells are p75NTR negative at all ages analyzed. Thus, if Pax2 cells, like their precursors, are sensitive to sonic hedgehog, this does not affect their expression of p75NTR. Our findings document that subsequently generated sets of Pax2 expressing precursors of inhibitory cerebellar interneurons are confronted with a dynamically changing complement of cerebellar glia. The eventual identification of fate-defining pathways should profit from the covariation with glial maturation predicted by the present findings.
Asunto(s)
Cerebelo/crecimiento & desarrollo , Interneuronas/citología , Neurogénesis/fisiología , Sustancia Blanca/crecimiento & desarrollo , Animales , Cerebelo/citología , Ratones , Ratones Endogámicos C57BL , Sustancia Blanca/citologíaRESUMEN
In contrast to humans and other mammals, zebrafish can successfully regenerate and remyelinate central nervous system (CNS) axons following injury. In addition to common myelin proteins found in mammalian myelin, 36K protein is a major component of teleost fish CNS myelin. Although 36K is one of the most abundant proteins in zebrafish brain, its function remains unknown. Here we investigate the function of 36K using translation-blocking Morpholinos. Morphant larvae showed fewer dorsally migrated oligodendrocyte precursor cells as well as upregulation of Notch ligand. A gamma secretase inhibitor, which prevents activation of Notch, could rescue oligodendrocyte precursor cell numbers in 36K morphants, suggesting that 36K regulates initial myelination through inhibition of Notch signaling. Since 36K like other short chain dehydrogenases might act on lipids, we performed thin layer chromatography and mass spectrometry of lipids and found changes in lipid composition in 36K morphant larvae. Altogether, we suggest that during early development 36K regulates membrane lipid composition, thereby altering the amount of transmembrane Notch ligands and the efficiency of intramembrane gamma secretase processing of Notch and thereby influencing oligodendrocyte precursor cell differentiation and further myelination. Further studies on the role of 36K short chain dehydrogenase in oligodendrocyte precursor cell differentiation during remyelination might open up new strategies for remyelination therapies in human patients.
Asunto(s)
Axones/metabolismo , Proteínas de la Mielina/metabolismo , Vaina de Mielina/metabolismo , Oligodendroglía/citología , Animales , Axones/patología , Encéfalo/metabolismo , Células CHO , Diferenciación Celular/fisiología , Cricetulus , Enfermedades Desmielinizantes/metabolismo , Humanos , Neurogénesis/fisiología , Pez CebraRESUMEN
Efficient coupling of the actin cytoskeleton to the cell membrane is crucial for histogenesis and maintenance of the nervous system. At this critical interface, BAR (Bin-Amphiphysin-Rvs) proteins regulate membrane bending, shown to be instrumental for mobility and morphogenesis of individual cells. Yet, the systemic significance of these proteins remains largely unexplored. Here, we probe the role of a prominent member of this protein family, the inverse-BAR protein Mtss1, for the development and function of a paradigmatic neuronal circuit, the cerebellar cortex. Mtss1-null mice show granule cell ectopias, dysmorphic Purkinje cells, malformed axons, and a protracted neurodegeneration entailing age-dependent motor deficits. In postmitotic granule cells, which transiently express Mtss1 while they migrate and form neurites, Mtss1 impinges on directional persistence and neuritogenesis. The latter effect can be specifically attributed to its exon 12a splice variant. Targeted re-expression of Mtss1 in Mtss1-null animals indicated that these pathologies were largely due to cell type-specific and intrinsic effects. Together, our results provide a mechanistic perspective on Mtss1 function for brain development and degeneration and relate it to structural features of this protein.
Asunto(s)
Axones/metabolismo , Cerebelo/metabolismo , Proteínas de Microfilamentos/metabolismo , Actividad Motora , Neuronas Motoras/metabolismo , Proteínas de Neoplasias/metabolismo , Degeneración Nerviosa , Células de Purkinje/metabolismo , Factores de Edad , Animales , Axones/patología , Células Cultivadas , Cerebelo/patología , Cerebelo/fisiopatología , Exones , Regulación del Desarrollo de la Expresión Génica , Genotipo , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas de Microfilamentos/deficiencia , Proteínas de Microfilamentos/genética , Neuronas Motoras/patología , Proteínas de Neoplasias/deficiencia , Proteínas de Neoplasias/genética , Fenotipo , Isoformas de Proteínas , Células de Purkinje/patología , Prueba de Desempeño de Rotación con Aceleración ConstanteRESUMEN
Maternally derived inflammatory mediators, such as IL-6 and IL-8, contribute to preterm delivery, low birth weight, and respiratory insufficiency, which are routinely treated with oxygen. Premature infants are at risk for developing adult-onset cardiac, metabolic, and pulmonary diseases. Long-term pulmonary consequences of perinatal inflammation are unclear. We tested the hypothesis that a hostile perinatal environment induces profibrotic pathways resulting in pulmonary fibrosis, including persistently altered lung structure and function. Pregnant C3H/HeN mice injected with LPS or saline on embryonic day 16. Offspring were placed in room air (RA) or 85% O(2) for 14 days and then returned to RA. Pulmonary function tests, microCTs, molecular and histological analyses were performed between embryonic day 18 and 8 wk. Alveolarization was most compromised in LPS/O(2)-exposed offspring. Collagen staining and protein levels were increased, and static compliance was decreased only in LPS/O(2)-exposed mice. Three-dimensional microCT reconstruction and quantification revealed increased tissue densities only in LPS/O(2) mice. Diffuse interstitial fibrosis was associated with decreased micro-RNA-29, increased transforming growth factor-ß expression, and phosphorylation of Smad2 during embryonic or early fetal lung development. Systemic maternal LPS administration in combination with neonatal hyperoxic exposure induces activation of profibrotic pathways, impaired alveolarization, and diminished lung function that are associated with prenatal and postnatal suppression of miR-29 expression.
Asunto(s)
Hiperoxia/fisiopatología , Inflamación/fisiopatología , Pulmón/patología , Pulmón/fisiopatología , Efectos Tardíos de la Exposición Prenatal/fisiopatología , Animales , Colágeno/metabolismo , Femenino , Fibrosis , Inflamación/inducido químicamente , Lipopolisacáridos/efectos adversos , Pulmón/embriología , Masculino , Ratones , Ratones Endogámicos C3H , MicroARNs/metabolismo , Modelos Animales , Embarazo , Alveolos Pulmonares/metabolismo , Alveolos Pulmonares/patología , Proteína Smad2/metabolismo , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
The gap junction protein connexin-45 (Cx45) is expressed in the conduction system of the heart and in certain neurons of the retina and brain. General and cardiomyocyte-directed deficiencies of Cx45 in mice lead to lethality on embryonic day 10.5 as a result of cardiovascular defects. Neuron-directed deletion of Cx45 leads to defects in transmission of visual signals. Connexin-36 (Cx36) is co-expressed with Cx45 in certain types of retinal interneurons. To determine whether these two connexins have similar functions and whether Cx36 can compensate for Cx45, we generated knock-in mice in which DNA encoding Cx45 was replaced with that encoding Cx36. Neuron-directed replacement of Cx45 with Cx36 resulted in viable animals. Electroretinographic and neurotransmitter coupling analyses demonstrated functional compensation in the retina. By contrast, general and cardiomyocyte-directed gene replacement led to lethality on embryonic day 11.5. Mutant embryos displayed defects in cardiac morphogenesis and conduction. Thus, functional compensation of Cx45 by Cx36 did not occur during embryonic heart development. These data suggest that Cx45 and Cx36 have similar functions in the retina, whereas Cx45 fulfills special functions in the developing heart that cannot be compensated by Cx36.
Asunto(s)
Conexinas/metabolismo , Embrión de Mamíferos/metabolismo , Corazón/embriología , Neuronas/metabolismo , Retina/metabolismo , Animales , Conexinas/genética , Femenino , Immunoblotting , Inmunoprecipitación , Técnicas In Vitro , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteína delta-6 de Union ComunicanteAsunto(s)
Neurociencias , Interferencia de ARN , ARN Interferente Pequeño/genética , Animales , Células Cultivadas , Cerebelo/citología , Cerebelo/embriología , Ciclofilinas/genética , Embrión de Mamíferos , Silenciador del Gen , Marcación de Gen/métodos , Ratones , Ratones Endogámicos C57BL , Factores de TiempoRESUMEN
Ventricular tachyarrhythmias are the main cause of sudden death in patients after myocardial infarction. Here we show that transplantation of embryonic cardiomyocytes (eCMs) in myocardial infarcts protects against the induction of ventricular tachycardia (VT) in mice. Engraftment of eCMs, but not skeletal myoblasts (SMs), bone marrow cells or cardiac myofibroblasts, markedly decreased the incidence of VT induced by in vivo pacing. eCM engraftment results in improved electrical coupling between the surrounding myocardium and the infarct region, and Ca2+ signals from engrafted eCMs expressing a genetically encoded Ca2+ indicator could be entrained during sinoatrial cardiac activation in vivo. eCM grafts also increased conduction velocity and decreased the incidence of conduction block within the infarct. VT protection is critically dependent on expression of the gap-junction protein connexin 43 (Cx43; also known as Gja1): SMs genetically engineered to express Cx43 conferred a similar protection to that of eCMs against induced VT. Thus, engraftment of Cx43-expressing myocytes has the potential to reduce life-threatening post-infarct arrhythmias through the augmentation of intercellular coupling, suggesting autologous strategies for cardiac cell-based therapy.