The multiple activities of BMPs during spinal cord development

Bone morphogenetic proteins (BMPs) are one of the main classes of multi-faceted secreted factors that drive vertebrate development. A growing body of evidence indicates that BMPs contribute to the formation of the central nervous system throughout its development, from the initial shaping of the neural primordium to the generation and maturation of the different cell types that form the functional adult nervous tissue. In this review, we focus on the multiple activities of BMPs during spinal cord development, paying particular attention to recent results that highlight the complexity of BMP signaling during this process. These findings emphasize the unique capacity of these signals to mediate various functions in the same tissue throughout development, recruiting diverse effectors and strategies to instruct their target cells.     

A comparative study of the differentiation of dissociated nerve cells under different culture conditions

Summary In vitro differentiation of chick embryo brain cells was compared under several culture conditions. Morphological observations and acetylcholinesterase histochemical staining revealed that the development was similar in all conditions tested if cells have been derived from 7 days embryos. Considering the cultures from 11 days embryos, the cell dissociation by trypsin and the plastic surface proved to be the most favourable conditions in contrast to mechanical dissection and collagen surface.M. Sensenbrenner is Maitre de Recherche au CNRS.     

Density gradient centrifugation of tumor cells from needle biopsies and their respective source tumors: a comparison of density distributions

A needle biopsy technique was applied to a murine fibrosarcoma (FSa) tumor system. FSa tumors of 8 mm diameter in size were biopsied and then made into single cell suspensions. Resulting density profiles of cells from both sources were compared following centrifugation in Renografin gradients. In all cases, there was excellent agreement between the density profiles of the material from each of the biopsies and the corresponding solid tumors.     

Genetic visualization of notch signaling in mammalian neurogenesis

Notch signaling plays crucial roles in fate determination and the differentiation of neural stem cells in embryonic and adult brains. It is now clear that the notch pathway is under more complex and dynamic regulation than previously thought. To understand the functional details of notch signaling more precisely, it is important to reveal when, where, and how notch signaling is dynamically communicated between cells, for which the visualization of notch signaling is essential. In this review, we introduce recent technical advances in the visualization of notch signaling during neural development and in the adult brain, and we discuss the physiological significance of dynamic regulation of notch signaling.     

Altered proteostasis in aging and heat shock response in C. elegans revealed by analysis of the global and de novo synthesized proteome

Protein misfolding and aggregation as a consequence of impaired protein homeostasis (proteostasis) not only characterizes numerous age-related diseases but also the aging process itself. Functionally related to the aging process are, among others, ribosomal proteins, suggesting an intimate link between proteostasis and aging. We determined by iTRAQ quantitative proteomic analysis in C. elegans how the proteome changes with age and in response to heat shock. Levels of ribosomal proteins and mitochondrial chaperones were decreased in aged animals, supporting the notion that proteostasis is altered during aging. Mitochondrial enzymes of the tricarboxylic acid cycle and the electron transport chain were also reduced, consistent with an age-associated energy impairment. Moreover, we observed an age-associated decline in the heat shock response. In order to determine how protein synthesis is altered in aging and in response to heat shock, we complemented our global analysis by determining the de novo proteome. For that, we established a novel method that enables both the visualization and identification of de novo synthesized proteins, by incorporating the non-canonical methionine analogue, azidohomoalanine (AHA), into the nascent polypeptides, followed by reacting the azide group of AHA by ‘click chemistry’ with an alkyne-labeled tag. Our analysis of AHA-tagged peptides demonstrated that the decreased abundance of, for example, ribosomal proteins in aged animals is not solely due to degradation but also reflects a relative decrease in their synthesis. Interestingly, although the net rate of protein synthesis is reduced in aged animals, our analyses indicate that the synthesis of certain proteins such as the vitellogenins increases with age.     

Density gradient centrifugation of tumor cells from needle biopsies and their respective source tumors: a comparison of density distributions

Summary A needle biopsy technique was applied to a murine fibrosarcoma (FSa) tumor system. FSa tumors of 8 mm diameter in size were biopsied and then made into single cell suspensions. Resulting density profiles of cells from both sources were compared following centrifugation in Renografin gradients. In all cases, there was excellent agreement between the density profiles of the material from each of the biopsies and the corresponding solid tumors.This investigation was supported by grant CA-18628, awarded by the National Cancer Institute, Department of Health and Human Services.     

Central nervous system stem cells in the embryo and adult

The central nervous system is generated from neural stem cells during embryonic development. These cells are multipotent and generate neurons, astrocytes and oligodendrocytes. The last few years it has been found that there are populations of stem cells also in the adult mammalian brain and spinal cord. In this paper, we review the recent development in the field of embryonic and adult neural stem cells. Received 26 March 1998; received after revision 27 April 1998; accepted 27 April 1998     

Regulation of oligodendrocyte precursor migration during development, in adulthood and in pathology

Oligodendrocytes are the myelin-forming cells in the central nervous system (CNS). These cells originate from oligodendrocyte precursor cells (OPCs) during development, and they migrate extensively from oligodendrogliogenic niches along the neural tube to colonise the entire CNS. Like many other such events, this migratory process is precisely regulated by a battery of positional and signalling cues that act via their corresponding receptors and that are expressed dynamically by OPCs. Here, we will review the cellular and molecular basis of this important event during embryonic and postnatal development, and we will discuss the relevance of the substantial number of OPCs existing in the adult CNS. Similarly, we will consider the behaviour of OPCs in normal and pathological conditions, especially in animal models of demyelination and of the demyelinating disease, multiple sclerosis. The spontaneous remyelination observed after damage in demyelinating pathologies has a limited effect. Understanding the cellular and molecular mechanisms underlying the biology of OPCs, particularly adult OPCs, should help in the design of neuroregenerative strategies to combat multiple sclerosis and other demyelinating diseases.     

Effects of retinoic acid on embryonic development of mice in culture

The effects of all-trans-retinoic acid (RA) (tretinoin) on the craniofacial development of mouse embryos were examined using whole embryo culture. In day 8 embryos cultured for 48 h, embryonic growth was inhibited concentration-dependently by all-trans-RA treatment. Most of the treated embryos exhibited hypoplasia of the primary palatal processes and a reduction in the development of the first visceral arches. In day 10 embryos cultured for 48 h, although embryonic growth was not inhibited at any concentrations of all-trans-RA, median cleft lip (93%), hypoplasia of the primary palatal processes (37%) and limb reduction deformities (48%) occurred commonly. Furthermore, RA treatment greatly reduced the size of the secondary palatal processes. The incorporation of 3H-thymidine in the treated maxillary processes was decreased to 65% of the control value at 1.0 x 10(-7) M all-trans-RA. These findings indicate that all-trans-RA is teratogenic in mouse whole embryo culture.     

In vitro development of rat embryos obtained from diabetic mothers

Rat embryos of 9.5 or 10 days of gestation were removed from control or streptozotocin-diabetic mothers and cultured in normal rat serum (180 mg% glucose) or in diabetic serum (600 mg% glucose). The development of control embryos in normal serum was adequate. Embryos from normal mothers cultured in diabetic serum showed signs of developmental retardation. The development of embryos obtained from diabetic mothers was severely impaired, regardless of the gestational age or the culture medium. These results suggest that a diabetic maternal milieu produces irreversible effects in the embryo very early in gestation.     

Effects of retinoic acid on embryonic development of mice in culture

Summary The effects of all-trans-retinoic acid (RA) (tretinoin) on the craniofacial development of mouse embryos were examined using whole embryo culture. In day 8 embryos cultured for 48 h, embryonic growth was inhibited concentration-dependently by all-trans-RA treatment. Most of the treated embryos exhibited hypoplasia of the primary palatal processes and a reduction in the development of the first viscreral arches. In day 10 embryos cultured for 48 h, although embryonic growth was not inhibited at any concentrations of all-trans-RA, median cleft lip (93%), hypoplasia of the primary palatal processes (37%) and limb reduction deformities (48%) occurred commonly. Furthermore, RA treatment greatly reduced the size of the secondary palatal processes. The incorporation of³H-thymidine in the treated maxillary processes was decreased to 65% of the control value at 1.0×10^–7 M alltrans-RA. These findings indicate that all-trans-RA is teratogenic in mouse whole embryo culture.     

Molecular and Cellular Basis of Regeneration and Tissue Repair

Although early after birth the central nervous system is more plastic than in the adult, it already displays limited regenerative capability. This becomes severely impaired at specific stages of embryonic development; however, the precise cellular and molecular basis of this loss is not fully understood. The chick embryo provides an ideal model for direct comparisons of regenerating and non-regenerating spinal cord within the same species because of its accessibility in ovo, the extensive knowledge of chick neural development and the molecular tools now available. Regenerative ability in the chick is lost at around E13, a relatively advanced stage of spinal cord development. This is most likely due to a complex series of events: there is evidence to suggest that developmentally regulated changes in the early response to injury, expression of inhibitory molecules and neurogenesis may contribute to loss of regenerative capacity in the chick spinal cord.     

Epithelial cell processes in the development of the secondary palate in the mouse.

Ultrastructural studies of palatal shelves of Tuck A mice embryos aged 12.25-14.25 days show discontinuities of the epithelial basement membrane traversed by epithelial cell processes before the onset of midline degenerative changes.     

Epithelial cell processes in the development of the secondary palate in the mouse

Summary Ultrastructural studies of palatal shelves of Tuck A mice embryos aged 12.25–14.25 days show discontinuities of the epithelial basement membrane traversed by epithelial cell processes before the onset of midline degenerative changes.     

Roles of glial cells in synapse development

Brain function relies on communication among neurons via highly specialized contacts, the synapses, and synaptic dysfunction lies at the heart of age-, disease-, and injury-induced defects of the nervous system. For these reasons, the formation—and repair—of synaptic connections is a major focus of neuroscience research. In this review, I summarize recent evidence that synapse development is not a cell-autonomous process and that its distinct phases depend on assistance from the so-called glial cells. The results supporting this view concern synapses in the central nervous system as well as neuromuscular junctions and originate from experimental models ranging from cell cultures to living flies, worms, and mice. Peeking at the future, I will highlight recent technical advances that are likely to revolutionize our views on synapse–glia interactions in the developing, adult and diseased brain.     

Inhibition of endogenous phosphodiesterase 7 promotes oligodendrocyte precursor differentiation and survival

During the development of the central nervous system (CNS), oligodendrocyte precursors (OPCs) are generated in specific sites within the neural tube and then migrate to colonize the entire CNS, where they differentiate into myelin-forming oligodendrocytes. Demyelinating diseases such as multiple sclerosis (MS) are characterized by the death of these cells. The CNS reacts to demyelination and by promoting spontaneous remyelination, an effect mediated by endogenous OPCs, cells that represent approximately 5–7 % of the cells in the adult brain. Numerous factors influence oligodendrogliogenesis and oligodendrocyte differentiation, including morphogens, growth factors, chemotropic molecules, extracellular matrix proteins, and intracellular cAMP levels. Here, we show that during development and in early adulthood, OPCs in the murine cerebral cortex contain phosphodiesterase-7 (PDE7) that metabolizes cAMP. We investigated the effects of different PDE7 inhibitors (the well-known BRL-50481 and two new ones, TC3.6 and VP1.15) on OPC proliferation, survival, and differentiation. While none of the PDE7 inhibitors analyzed altered OPC proliferation, TC3.6 and VP1.15 enhanced OPC survival and differentiation, processes in which ERK intracellular signaling played a key role. PDE7 expression was also observed in OPCs isolated from adult human brains and the differentiation of these OPCs into more mature oligodendroglial phenotypes was accelerated by treatment with both new PDE7 inhibitors. These findings reveal new roles for PDE7 in regulating OPC survival and differentiation during brain development and in adulthood, and they may further our understanding of myelination and facilitate the development of therapeutic remyelination strategies for the treatment of MS.     

Myelination in the hippocampus during development and following lesion

Myelin is crucial for the stabilization of axonal projections in the developing and adult mammalian brain. However, myelin components also act as a non-permissive and repellent substrate for outgrowing axons. Therefore, one major factor which accounts for the lack of axonal regeneration in the mature brain is myelin. Here we report on the appearance of mature, fully myelinated axons during hippocampal development and following entorhinal lesion with the myelin-specific marker Black Gold. Although entorhinal axons enter the hippocampal formation at embryonic day 17, light and ultrastructural analysis revealed that mature myelinated fibers in the hippocampus occur in the second postnatal week. During postnatal development, increasing numbers of myelinated fibers appear and the distribution of myelinated fibers at postnatal day 25 was similar to that found in the adult. After entorhinal cortex lesion, a specific anterograde denervation in the hippocampus takes place, accompanied by a long-lasting loss of myelin. Quantitative analysis of myelin and myelin breakdown products at different time points after lesion revealed a temporally close correlation to the degeneration and reorganization pha-ses in the hippocampus. In contrast, electroconvulsive seizures resulted in brief demyelination and a faster recovery time course. In conclusion, we could show that the appearance of mature axons in the hippocampus is temporally regulated during development. In the adult hippocampus, demyelination was found after anterograde degeneration and also following seizures, suggesting that independent types of insult lead to demyelination. Reappearing mature axons were found in the hippocampus following axonal sprouting. Therefore, our quantitative analysis of mature axons and myelination effectively reflects the readjusted axonal density and possible electrophysiological balance following lesion. Received 22 December 2003; received after revision 11 February 2004; accepted 17 February 2004     

Autotaxin (NPP-2) in the brain: cell type-specific expression and regulation during development and after neurotrauma

Autotaxin is a secreted cell motility-stimulating exo-phosphodiesterase with lysophospholipase D activity that generates bioactive lysophosphatidic acid. Lysophosphatidic acid has been implicated in various neural cell functions such as neurite remodeling, demyelination, survival and inhibition of axon growth. Here, we report on the in vivo expression of autotaxin in the brain during development and following neurotrauma. We found that autotaxin is expressed in the proliferating subventricular and choroid plexus epithelium during embryonic development. After birth, autotaxin is mainly found in white matter areas in the central nervous system. In the adult brain, autotaxin is solely expressed in leptomeningeal cells and oligodendrocyte precursor cells. Following neurotrauma, autotaxin is strongly up-regulated in reactive astrocytes adjacent to the lesion. The present study revealed the cellular distribution of autotaxin in the developing and lesioned brain and implies a function of autotaxin in oligodendrocyte precursor cells and brain injuries. Received 18 September 2006; received after revision 30 October 2006; accepted 4 December 2006     

Dense core vesicles during photoreceptor development

Summary The presence of dense core vesicles in the terminal expansions of photoreceptors in development is decribed in the chick embryo retina, from the 16th to the 18th day of incubation.Acknowledgment. J.M.G.-G. was supported by a grant from the Comision Asesora de Investigacion Cientifica y Tecnica, Madrid, Spain.     

A comparative study of the differentiation of dissociated nerve cells under different culture conditions.

In vitro differentiation of chick embryo brain cells was compared under several culture conditions. Morphological observations and acetylcholinesterase histochemical staining revealed that the development was similar in all conditions tested if cells have been derived from 7 days embryos. Considering the cultures from 11 days embryos, the cell dissociation by trypsin and the plastic surface proved to be the most favourable conditions in contrast to mechanical dissection and collagen surface.