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The gene for the atypical NOTCH ligand delta-like homologue 1 (Dlk1) encodes membrane-bound and secreted isoforms that function in several developmental processes in vitro and in vivo. Dlk1, a member of a cluster of imprinted genes, is expressed from the paternally inherited chromosome12. Here we show that mice that are deficient in Dlk1 have defects in postnatal neurogenesis in the subventricular zone: a developmental continuum that results in depletion of mature neurons in the olfactory bulb. We show that DLK1 is secreted by niche astrocytes, whereas its membrane-bound isoform is present in neural stem cells (NSCs) and is required for the inductive effect of secreted DLK1 on self-renewal. Notably, we find that there is a requirement for Dlk1 to be expressed from both maternally and paternally inherited chromosomes. Selective absence of Dlk1 imprinting in both NSCs and niche astrocytes is associated with postnatal acquisition of DNA methylation at the germ-line-derived imprinting control region. The results emphasize molecular relationships between NSCs and the niche astrocyte cells of the microenvironment, identifying a signalling system encoded by a single gene that functions coordinately in both cell types. The modulation of genomic imprinting in a stem-cell environment adds a new level of epigenetic regulation to the establishment and maintenance of the niche, raising wider questions about the adaptability, function and evolution of imprinting in specific developmental contexts., The mammalian adult brain is generally postmitotic, but reservoirs of NSCs with features of astroglial cells exist in the hippocampus and subventricular zone (SVZ), supporting lifelong neurogenesis (3,4). The SVZ [...]

Satb2, encoding a nuclear matrix protein, is expressed in branchial arches and in cells of the osteoblast lineage. Studies found that Satb2 mice exhibit both craniofacial abnormalities that resemble those observed in humans carrying a translocation in SATB2 and defects in osteoblast differentiation and function.

Null mutations in Lef1 and Tcf1 create severe defects in differentiating paraxial mesoderm as well as forming additional neural tubes. Gene inactivations of Lef1, Tcf1, or Tcf4 in mice have not produced phenotypes that recreate any known Wnt mutation. LEF-1 and TCF-1 have redundant roles in Wnt signaling in mouse development.

Mice homozygous for a null glial-cell-line-derived neurotrophic factor (GDNF) allele exhibit the absence of kidneys, ureters and enteric nervous system (ENS). The mutants show defects in the sympathetic and nodose neurons, and dorsal root ganglion. The basal forebrain, cerebellum, striatum, neocortex, hippocampus, and the dopaminergic, catecholaminergic and noradrenergic neurons are normal. This indicates that GDNF is necessary for the development of ENS, kidney, and sympathetic and sensory neurons. The homozygous mice die within two days of birth while the heterozygous mice are normal in size.

Study of the nervous system of mice reveals that the absence of neurotrophin-3, which maintains nervous system through neuronal survival and differentiation in mice, leads to chronic sensory and sympathetic deficits. These mice show absence of muscle spindles and exhibit extraordinary limb position. Sympathetic and sensory neurons which require nerve growth factor or neurotrophin factor for survival at a later stage require neurotrophin-3 which acts as a receptor binder.

A study of brain-derived neurotrophic factor (BDNF) null mutation reveals that nervous system cell types such as neurons in multiple sensory ganglia, cerebral cortex, the hippocampus and the striatum require BDNF for normal development. BDNF controls brain differentiation in infants, as indicated by the diminished expression of neuropeptide and calcium binding proteins.

Superior cervical ganglia of postnatal mice with a targeted disruption of the gene for neurotrophin-3 have 50% fewer neurons than those of wild-type mice. In culture, neurotrophin-3 increases the survival of proliferating sympathetic precursors. Both precursor death (W. ElShamy et al., 1996, Development 122, 491-500) and, more recently, neuronal death (S. Wyatt et al., 1997, EMBO J. 16, 3115-3123) have been described in mice lacking NT-3. Consistent with the second report, we found that, in vivo, neurogenesis and precursor survival were unaffected by the absence of neurotrophin-3 but neuronal survival was compromised so that only 50% of the normal number of neurons survived to birth. At the time of neuron loss, neurotrophin-3 expression, assayed with a lacZ reporter, was detected in sympathetic target tissues and blood vessels, including those along which sympathetic axons grow, suggesting it may act as a retrograde neurotrophic factor, similar to nerve growth factor. To explore this possibility, we compared neuron loss in neurotrophin-3-deficient mice with that in nerve growth factor-deficient mice and found that neuronal losses occurred at approximately the same time in both mutants, but were less severe in mice lacking neurotrophin-3. Eliminating one or both neurotrophin-3 alleles in mice that lack nerve growth factor does not further reduce sympathetic neuron number in the superior cervical ganglion at E17.5 but does alter axon outgrowth and decrease salivary gland innervation. Taken together these results suggest that neurotrophin-3 is required for survival of some sympathetic neurons that also require nerve growth factor. Key Words: neurotrophin; sympathetic neuron; survival; cell death.

Determines the essential function of the neurotrophin, brain-derived neurotrophic factor (BDNF) by mutating the BDNF gene. Regulatory function of neurotrophins; Generation of mice having a disrupted BDNF gene; Observation of the mice; Brain development in the BDNF mutant.

We describe a protocol developed/modified by our group for the ex vivo and in vivo assessment of the response to a soluble factor of murine neural stem cells from the adult sub-ventricular zone (SVZ). The procedure includes several experimental options that can be used either independently or in combination. Potential factor effects on self-renewal, survival and proliferation are assayed by means of neurosphere cultures, with the factor administered directly in vitro to the culture plates (Step 1) or infused in vivo immediately before tissue dissociation (Step 3). We also use bromodeoxiuridine (BrdU) retention to label slowly dividing cells in vivo and subsequently perform two different types of experiments. In one set of experiments, the factor is added to primary cultures of stem cells obtained from the BrdU-pulsed animals and effects are tested on label-retaining cells after immunocytochemistry (Step 2). In another set, prolonged intraventricular infusion of the factor in BrdU-pulsed animals is followed by immunohistochemical analysis of BrdU labeling in the intact SVZ (Step 4). The minimum estimated time for the full combined procedure is 45 d.