Mechanisms Of Early Development Research Articles

Proteome of mouse oocytes at different developmental stages

The mammalian oocyte possesses powerful reprogramming factors, which can reprogram terminally differentiated germ cells (sperm) or somatic cells within a few cell cycles. Although it has been suggested that use of oocyte-derived transcripts may enhance the generation of induced pluripotent stem cells, the reprogramming factors in oocytes are undetermined, and even the identified proteins composition of oocytes is very limited. In the present study, 7,000 mouse oocytes at different developmental stages, including the germinal vesicle stage, the metaphase II (MII) stage, and the fertilized oocytes (zygotes), were collected. We successfully identified 2,781 proteins present in germinal vesicle oocytes, 2,973 proteins in MII oocytes, and 2,082 proteins in zygotes through semiquantitative MS analysis. Furthermore, the results of the bioinformatics analysis indicated that different protein compositions are correlated with oocyte characteristics at different developmental stages. For example, specific transcription factors and chromatin remodeling factors are more abundant in MII oocytes, which may be crucial for the epigenetic reprogramming of sperm or somatic nuclei. These results provided important knowledge to better understand the molecular mechanisms in early development and may improve the generation of induced pluripotent stem cells.

Multi-system disorders of glycosphingolipid and ganglioside metabolism

Glycosphingolipids (GSLs) and gangliosides are a group of bioactive glycolipids that include cerebrosides, globosides, and gangliosides. These lipids play major roles in signal transduction, cell adhesion, modulating growth factor/hormone receptor, antigen recognition, and protein trafficking. Specific genetic defects in lysosomal hydrolases disrupt normal GSL and ganglioside metabolism leading to their excess accumulation in cellular compartments, particularly in the lysosome, i.e., lysosomal storage diseases (LSDs). The storage diseases of GSLs and gangliosides affect all organ systems, but the central nervous system (CNS) is primarily involved in many. Current treatments can attenuate the visceral disease, but the management of CNS involvement remains an unmet medical need. Early interventions that alter the CNS disease have shown promise in delaying neurologic involvement in several CNS LSDs. Consequently, effective treatment for such devastating inherited diseases requires an understanding of the early developmental and pathological mechanisms of GSL and ganglioside flux (synthesis and degradation) that underlie the CNS diseases. These are the focus of this review.

Editorial: Special Issue Neurovision

Visual system research has a longstanding tradition at the Ruhr University in Bochum and it benefitted from continuous support by the German Research Foundation (Deutsche Forschungsgemeinschaft, DFG) granted first for a Research Unit and then the Collaborative Research Center (CRC) ‘‘Neurovision’’ between 1990 and 2007. This special issue of EBR named ‘‘Neurovision’’ concludes this period and summarizes our achievements. The joint venture in vision research in Bochum started in 1990 with the DFG Research Unit ‘‘Neurobiology of the visual system—Neurovision’’ and continued from 1996 onwards in the Collaborative Research Center ‘‘Neuronal mechanisms of vision—Neurovision’’. The Center was devoted to research in visual neuroscience from early developmental mechanisms to cellular and systems properties and further up to higher and visuomotor brain functions, with a wide range of methodical approaches including, molecular biology, cellular physiology, systems neurophysiology, neuroanatomy, and computational neuroscience. After 12 years of continuous and generous funding by the DFG, this CRC had to come to an end. In this issue of EBR, we want to present some of the highlights from the research, which was funded over the last triannual period in the CRC Neurovision. Naturally, most of the results from the previous periods have already been published and papers are given in the reference lists. The development of specific research themes and the projects of the CRC over the last 12 years are represented in the reviews in this volume, and the newest results accumulated during the last funding period are described in the original research articles. With this special issue, we want to especially acknowledge the support we received from the German Research Foundation.

Out With the Old, In With the New?

President Obama's recent announcement of a new policy for Federal Funding of stem cell research changes the landscape for this important area of biomedicine in the United States. His Executive Order and the subsequent draft guidelines of the National Institutes of Health (NIH) remove a major existing limitation on federal support for research using pluripotent human embryonic stem cells (hESCs). These stem cells can specialize into all body cell types. Thus, in principle they can be used to produce a wide range of tissues for therapeutic use. In addition, their study will very likely generate new insights into human early developmental mechanisms. The new Obama policy promises to speed research by making many new hESC lines eligible for federally funded studies. But the draft NIH guidelines accompanying the new policy need revision, because their donor consent rules for embryos used to generate hESCs would make some—and perhaps even all—of the previously approved hESCs ineligible for further federal funding. This would needlessly hinder progress in the stem cell field, and reasonable exceptions should be made to correct this unintended outcome.

Hypoxanthine-guanine phosphoribosyl transferase regulates early developmental programming of dopamine neurons: implications for Lesch-Nyhan disease pathogenesis

Hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficiency results in Lesch-Nyhan disease (LND), where affected individuals exhibit a characteristic neurobehavioral disorder that has been linked with dysfunction of dopaminergic pathways of the basal ganglia. Since the functions of HPRT, a housekeeping enzyme responsible for recycling purines, have no direct relationships with the dopaminergic pathways, the mechanisms whereby HPRT deficiency affect them remain unknown. The current studies demonstrate that HPRT deficiency influences early developmental processes controlling the dopaminergic phenotype, using several different cell models for HPRT deficiency. Microarray methods and quantitative PCR were applied to 10 different HPRT-deficient (HPRT(-)) sublines derived from the MN9D cell line. Despite the variation inherent in such mutant sublines, several consistent abnormalities were evident. Most notable were increases in the mRNAs for engrailed 1 and 2, transcription factors known to play a key role in the specification and survival of dopamine neurons. The increases in mRNAs were accompanied by increases in engrailed proteins, and restoration of HPRT reverted engrailed expression towards normal levels, demonstrating a functional relationship between HPRT and engrailed. The functional relevance of the abnormal developmental molecular signature of the HPRT(-) MN9D cells was evident in impoverished neurite outgrowth when the cells were forced to differentiate chemically. To verify that these abnormalities were not idiosyncratic to the MN9D line, HPRT(-) sublines from the SK-N-BE(2) M17 human neuroblastoma line were evaluated and an increased expression of engrailed mRNAs was also seen. Over-expression of engrailed occurred even in primary fibroblasts from patients with LND in a manner that suggested a correlation with disease severity. These results provide novel evidence that HPRT deficiency may affect dopaminergic neurons by influencing early developmental mechanisms.

Production of different phenotypes from the same genotype in the same environment by developmental variation

The phenotype of an organism is determined by the genes, the environment and stochastic developmental events. Although recognized as a basic biological principle influencing life history, susceptibility to diseases, and probably evolution, developmental variation (DV) has been only poorly investigated due to the lack of a suitable model organism. This obstacle could be overcome by using the recently detected, robust and highly fecund parthenogenetic marbled crayfish as an experimental animal. Batch-mates of this clonal crayfish, which were shown to be isogenic by analysis of nuclear microsatellite loci, exhibited surprisingly broad ranges of variation in coloration, growth, life-span, reproduction, behaviour and number of sense organs, even when reared under identical conditions. Maximal variation was observed for the marmorated coloration, the pattern of which was unique in each of the several hundred individuals examined. Variation among identically raised batch-mates was also found with respect to fluctuating asymmetry, a traditional indicator of the epigenetic part of the phenotype, and global DNA methylation, an overall molecular marker of an animal's epigenetic state. Developmental variation was produced in all life stages, probably by reaction-diffusion-like patterning mechanisms in early development and non-linear, self-reinforcing circuitries involving behaviour and metabolism in later stages. Our data indicate that, despite being raised in the same environment, individual genotypes can map to numerous phenotypes via DV, thus generating variability among clone-mates and individuality in a parthenogenetic species. Our results further show that DV, an apparently ubiquitous phenomenon in animals and plants, can introduce components of randomness into life histories, modifying individual fitness and population dynamics. Possible perspectives of DV for evolutionary biology are discussed.

Amphibians as Model Organisms for Studying the Dynamics of Eukaryote Genetic Material Architecture

Amphibians have played a key role in the elucidation of the mechanisms of early development over the last century. Much of our knowledge about the mechanisms of vertebrate early development comes from studies using Xenopus laevis. Xenopus sp. is a major contributor to our understanding of cell biological and biochemical processes, including: (1) chromosome replication; (2) chromatin, cytoskeleton and nuclear assembly; (3) cell cycle progression and (4) intracellular signaling. Amphibian embryos remained the embryos of choice for experimental embryology for many decades. European embryologists used predominantly urodele embryos (such as Triturus) and embryos of the frog Rana temporaria, which is related to the North American species Rana pipiens. Using light, fluorescence, transmission electron microscopy (TEM) and molecular investigations, some peculiar aspects of chromatin and chromosome organization and evolution in oogenesis and spermatogenesis of amphibians were investigated. We have focused our investigations on dynamics of the chromatin structure in different stages of development. Keywords: amphibians, chromatin remodelling, gametogenesis, DNA methylation, lampbrush chromosomes, ribosomal gene amplification.

Vascular Development, Pulse Pressure, and the Mechanisms of Hypertension

For a given cardiac function, the cyclic blood pressure (BP) curve results from 2 different phenotypes: the mean arterial pressure (MAP), a steady component reflecting the resistance of the microvascular network, and pulse pressure (PP), another component corresponding to large artery stiffness and wave reflections. Around birth, cardiovascular (CV) survival is critically influenced by the coupling between the heart and thoracic aorta, and hence, the adequacy of the Windkessel function, the magnitude of aortic elastin accumulation and the PP level. The maturation of the aortic trunk and its branches results from adaptative mechanisms involving shear and tensile stress, with major potential consequences on heart rate control, transit of wave reflections, and coronary perfusion. An adequate optimization of the Windkessel function, and hence PP, diastolic coronary perfusion and CV survival needs a critical MAP level to be reached in each individual during the postnatal period. The achievement of this MAP level requires the development of multiple resistance segments of the microvascular network, particularly within the kidney. Translated in adult populations, this pathophysiological process gives rise to a Gaussian BP distribution, with individuals remaining in the same BP percentile from birth onward (BP tracking). We suggest that hypertension results from early developmental vascular mechanisms that direct BP toward the higher percentiles of the Gaussian distribution curve.

Developmental Activation of the Rb–E2F Pathway and Establishment of Cell Cycle-regulated Cyclin-dependent Kinase Activity during Embryonic Stem Cell Differentiation

To understand cell cycle control mechanisms in early development and how they change during differentiation, we used embryonic stem cells to model embryonic events. Our results demonstrate that as pluripotent cells differentiate, the length of G(1) phase increases substantially. At the molecular level, this is associated with a significant change in the size of active cyclin-dependent kinase (Cdk) complexes, the establishment of cell cycle-regulated Cdk2 activity and the activation of a functional Rb-E2F pathway. The switch from constitutive to cell cycle-dependent Cdk2 activity coincides with temporal changes in cyclin A2 and E1 protein levels during the cell cycle. Transcriptional mechanisms underpin the down-regulation of cyclin levels and the establishment of their periodicity during differentiation. As pluripotent cells differentiate and pRb/p107 kinase activities become cell cycle dependent, the E2F-pRb pathway is activated and imposes cell cycle-regulated transcriptional control on E2F target genes, such as cyclin E1. These results suggest the existence of a feedback loop where Cdk2 controls its own activity through regulation of cyclin E1 transcription. Changes in rates of cell division, cell cycle structure and the establishment of cell cycle-regulated Cdk2 activity can therefore be explained by activation of the E2F-pRb pathway.

Two developmental switches in GABAergic signalling: the K+–Cl− cotransporter KCC2 and carbonic anhydrase CAVII

GABAergic signalling has the unique property of 'ionic plasticity', which is based on short-term and long-term changes in the Cl(-) and HCO(3)(-) ion concentrations in the postsynaptic neurones. While short-term ionic plasticity is caused by activity-dependent, channel-mediated anion shifts, long-term ionic plasticity depends on changes in the expression patterns and kinetic regulation of molecules involved in anion homeostasis. During development the efficacy and also the qualitative nature (depolarization/excitation versus hyperpolarization/inhibition) of GABAergic transmission is influenced by the neuronal expression of two key molecules: the chloride-extruding K(+)-Cl(-) cotransporter KCC2, and the cytosolic carbonic anhydrase (CA) isoform CAVII. In rat hippocampal pyramidal neurones, a steep up-regulation of KCC2 accounts for the 'developmental switch', which converts depolarizing and excitatory GABA responses of immature neurones to classical hyperpolarizing inhibition by the end of the second postnatal week. The immature hippocampus generates large-scale network activity, which is abolished in parallel by the up-regulation of KCC2 and the consequent increase in the efficacy of neuronal Cl(-) extrusion. At around postnatal day 12 (P12), an abrupt, steep increase in intrapyramidal CAVII expression takes place, promoting excitatory responses evoked by intense GABAergic activity. This is largely caused by a GABAergic potassium transient resulting in spatially widespread neuronal depolarization and synchronous spike discharges. These facts point to CAVII as a putative target of CA inhibitors that are used as antiepileptic drugs. KCC2 expression in adult rat neurones is down-regulated following epileptiform activity and/or neuronal damage by BDNF/TrkB signalling. The lifetime of membrane-associated KCC2 is very short, in the range of tens of minutes, which makes KCC2 ideally suited for mediating GABAergic ionic plasticity. In addition, factors influencing the trafficking and kinetic modulation of KCC2 as well as activation/deactivation of CAVII are obvious candidates in the ionic modulation of GABAergic responses. The down-regulation of KCC2 under pathophysiological conditions (epilepsy, damage) in mature neurones seems to reflect a 'recapitulation' of early developmental mechanisms, which may be a prerequisite for the re-establishment of connectivity in damaged brain tissue.

Generation of mutants with developmental defects in zebrafish by ENU mutagenesis

As a good model for studying early development of vertebrates, zebrafish (Danio rerio) is attracting more and more attention. Following ENU mutagenesis, 320 F2 families were established. Mutants, which showed defects in epiboly, axis, somite, head, and cardiac and blood systems, were identified by observing morphological changes in F3 embryos. So far, 35 mutant lines have been established, the majority of which showed anomalies in axis and somite formation. These mutant lines provide useful genetic resources for cloning of the mutant genes and for studying mechanisms of early development of vertebrate embryos.

N‐MYC Downstream‐Regulated Gene 1 Is Mutated In Hereditary Motor And Sensory Neuropathy‐LOM

Hereditary motor and sensory neuropathies, to which Charcot‐Marie‐Tooth (CMT) disease belongs, are a common cause of disability in adulthood. Growing awareness that axonal loss, rather than demyelination per se, is responsible for the neurological deficit in demyelinating CMT disease has focused research on the mechanisms of early development, cell differentiation, and cell‐cell interactions in the peripheral nervous system. Autosomal recessive peripheral neuropathies are relatively rare but are clinically more severe than autosomal dominant forms of CMT, and understanding their molecular basis may provide a new perspective on these mechanisms. Here we report the identification of the gene responsible for hereditary motor and sensory neuropathy‐Lom (HMSNL). HMSNL shows features of Schwann‐cell dysfunction and a concomitant early axonal involvement, suggesting that impaired axon‐glia interactions play a major role in its pathogenesis. The gene was previously mapped to 8q24.3, where conserved disease haplotypes suggested genetic homogeneity and a single founder mutation. We have reduced the HMSNL interval to 200 kb and have characterized it by means of large‐scale genomic sequencing. Sequence analysis of two genes located in the critical region identified the founder HMSNL mutation: a premature‐termination codon at position 148 of the N‐myc downstream‐regulated gene 1 (NDRG1). NDRG1 is ubiquitously expressed and has been proposed to play a role in growth arrest and cell differentiation, possibly as a signaling protein shuttling between the cytoplasm and the nucleus. We have studied expression in peripheral nerve and have detected particularly high levels in the Schwann cell. Taken together, these findings point to NDRG1 having a role in the peripheral nervous system, possibly in the Schwann‐cell signaling necessary for axonal survival.

N-myc Downstream-Regulated Gene 1 Is Mutated in Hereditary Motor and Sensory Neuropathy–Lom

Hereditary motor and sensory neuropathies, to which Charcot-Marie-Tooth (CMT) disease belongs, are a common cause of disability in adulthood. Growing awareness that axonal loss, rather than demyelination per se, is responsible for the neurological deficit in demyelinating CMT disease has focused research on the mechanisms of early development, cell differentiation, and cell-cell interactions in the peripheral nervous system. Autosomal recessive peripheral neuropathies are relatively rare but are clinically more severe than autosomal dominant forms of CMT, and understanding their molecular basis may provide a new perspective on these mechanisms. Here we report the identification of the gene responsible for hereditary motor and sensory neuropathy-Lom (HMSNL). HMSNL shows features of Schwann-cell dysfunction and a concomitant early axonal involvement, suggesting that impaired axon-glia interactions play a major role in its pathogenesis. The gene was previously mapped to 8q24.3, where conserved disease haplotypes suggested genetic homogeneity and a single founder mutation. We have reduced the HMSNL interval to 200 kb and have characterized it by means of large-scale genomic sequencing. Sequence analysis of two genes located in the critical region identified the founder HMSNL mutation: a premature-termination codon at position 148 of the N-myc downstream-regulated gene 1 (NDRG1). NDRG1 is ubiquitously expressed and has been proposed to play a role in growth arrest and cell differentiation, possibly as a signaling protein shuttling between the cytoplasm and the nucleus. We have studied expression in peripheral nerve and have detected particularly high levels in the Schwann cell. Taken together, these findings point to NDRG1 having a role in the peripheral nervous system, possibly in the Schwann-cell signaling necessary for axonal survival.

Embryonic development of connections in Turtle Pallium

We are interested in similarities and conserved mechanisms in early development of the reptilian and mammalian thalamocortical connections. We set out to analyse connectivity in embryonic turtle brains (Pseudemys scripta elegans, between stages 17 and 25), by using carbocyanine dye tracing. From the earliest stages studied, labelling from dorsal and ventral thalamus revealed backlabelled cells among developing thalamic fibres within the lateral forebrain bundle and striatum, which had similar morphology to backlabelled internal capsule cells in embryonic rat (Molnár and Cordery, 1999). However, thalamic crystal placements did not label cells in the dorsal ventricular ridge (DVR) at any stage examined. Crystal placements into both dorsal and lateral cortex labelled cells in the DVR and, reciprocally, DVR crystal placements labelled cells in the dorsal and lateral cortices. Retrograde labelling revealed that thalamic fibres arrive in the DVR and dorsal cortex by stage 19. The DVR received projections from the nucleus rotundus and the dorsal cortex exclusively from the perirotundal complex (including lateral geniculate nucleus). Thalamic fibres show this remarkable degree of specificity from the earliest stage we could examine with selective retrograde labelling (stage 19). Our study demonstrates that axons of similar cells are among the first to reach dorsal and ventral thalamus in mammals and reptiles. Our connectional analysis in turtle suggests that some cells of the mammalian primitive internal capsule are homologous to a cell group within the reptilian lateral forebrain bundle and striatum and that diverse vertebrate brains might use a highly conserved pattern of early thalamocortical development.

Vertebrate evolution: the developmental origins of adult variation.

Many biologists assume, as Darwin did, that natural selection acts mainly on late embryonic or postnatal development. This view is consistent with von Baer's observations of morphological divergence at late stages. It is also suggested by the conserved morphology and common molecular genetic mechanisms of pattern formation seen in embryos. I argue here, however, that differences in adult morphology may be generated at a variety of stages. Natural selection may have a major action on developmental mechanisms during the organogenetic period, because this is when many adult traits are specified. Evolutionary changes in these early developmental mechanisms probably include subtle shifts in the timing of gene expression. Changes of this kind have little or no gross effect on the anatomy of the embryo; they are only phenotypically expressed, or readily detected, when amplified at later stages. The phylotypic stage, the developmental hourglass, modularity, and von Baerian divergence are reassessed in terms of these arguments.

Proliferation and the monoclonal origins of atherosclerotic lesions.

Benditt's observation of the monoclonal origin of the atherosclerotic lesion has been controversial because it appeared to conflict with conventional wisdom. A new method based on a polymerase chain reaction amplification of the DNA of an X-inactivated gene from microdissected tissue confirms that Benditt was correct. However, this monoclonal expansion can also be found in nonatherosclerotic intima and media. These new data suggest that plaque clonality may represent expansion of preexisting patches of cells arising during development of the media. This developmental view does not conflict with other recent evidence that plaque expansion is associated with mutation or viral events. However, if plaques arise from patches, then early developmental mechanisms may be critical to the later evolution of the lesions.

Behavioral states and their ontogeny: human studies.

Abstract Despite numerous attempts in describing the many underlying events correlated with changes in behavioral states, the biological significance of state alternation remains unknown. There does not yet exist a satisfactory explanation for the biological need for such state changes, nor does there exist sufficient physiological evidence to explain the underlying processes leading to these state transitions. Nonetheless, the study of the ontogeny of behavioral states is important because it shows the progressive ability of the organism to integrate multiple neuronal systems. Available literature suggests that some rudimentary form of behavioral state is established in fetal life throughout the animal kingdom, although there is substantial difference in the degree and rate of development of organized behavioral states among the different animal species. Highly organized behavioral states that involve concurrence of many physiologic systems are present in the third trimester in the fetal lamb, while they are not seen until 2 to 3 weeks postnatally in the rat, rabbit, and cat. Subsequent maturation consists primarily of increased cycle length in all species. In addition, there is some evidence that a circadian rhythm may be superimposed on the ultradian rhythm of behavioral states in immature animals, but there is currently no information on the ontogenesis of this circadian rhythm. It is currently hypothesized that behavioral states cycling in the adult is generated by the interaction of multiple, anatomically distributed sets of neurons. 40 Electrophysiological and pharmacological studies have suggested that both the cholinergic and aminergic neuronal systems play important roles in the maintenance of state cycling. Studies in the fetal lamb also support these proposed mechanisms in early development. 41, 42 However, there is little evidence to indicate that changes in transmitter release or receptor sensitivity occur in the course of sleep deprivation in the adult. A number of other neuropeptides may also be involved in this complicated control system. The progressive organization of behavioral states in early development and the agerelated changes in cycle duration may provide valuable insight in understanding the neurobiological events that underlie state alternation and homeostasis.

Determinative Mechanisms in Early Development. Papers Presented at a Meeting of the British Society for Developmental Biology at the University of East Anglia, April 1986.C. C. Wylie

Previous articleNext article No AccessNew Biological BooksDeterminative Mechanisms in Early Development. Papers Presented at a Meeting of the British Society for Developmental Biology at the University of East Anglia, April 1986. C. C. Wylie Mary BownesMary Bownes Search for more articles by this author PDFPDF PLUS Add to favoritesDownload CitationTrack CitationsPermissionsReprints Share onFacebookTwitterLinkedInRedditEmail SectionsMoreDetailsFiguresReferencesCited by The Quarterly Review of Biology Volume 62, Number 3Sep., 1987 Published in association with Stony Brook University Article DOIhttps://doi.org/10.1086/415549 Copyright 1987 Stony Brook Foundation, Inc.PDF download Crossref reports no articles citing this article.