Embryonic development of the nervous system of the temnocephalid flatworm Craspedella pedum

The nervous system of temnocephalid flatworms consists of the brain and three pairs of longitudinal connectives extending into the trunk and tail. The connectives are crosslinked by an invariant number of regularly spaced commissures. Branches of the connectives innervate the tentacles of the head and the sucker organ in the tail. A set of nerve rings encircling the pharynx and connected to the brain and connectives constitute the pharyngeal nervous system. The nervous system is formed during early embryogenesis when the embryo represents a multilayered mesenchymal mass of cells. Gastrulation and the formation of separate epithelial germ layers that characterize most other animal groups are absent. The brain arises as a bilaterally symmetric condensation of postmitotic cells in the deep layers of the anterior region of the embryonic mesenchyme. The pattern of axon outgrowth, visualized by labeling with anti-acetylated tubulin (acTub) antibody, shows marked differences from the pattern observed in other flatworm taxa. in regard to the number of neurons that express the acTub epitope. Acetylated tubulin is only expressed in neurons that form long axon tracts. In other flatworm species, such as the typhloplanoid Mesostoma and the polyclad Imogine, which were investigated by us with the acTub antibody (Hartenstein and Ehlers [2000] Dev. Genes Evol. 210:399-415; Younossi-Hartenstein and Hartenstein [2000] Dev. Genes Evol. 210:383-398), only a small number of pioneer neurons become acTub positive during the embryonic period. By contrast, in temnocephalids, most, if not all, neurons express acTub and form long, large-diameter axons. Initially, the brain commissure, pharyngeal nerve ring, and the connectives are laid down. Commissural tracts and tentacle nerves branching off the connectives appear later. We speculate that the precocious differentiation of the nervous system may be related to the fact that temnocephalids move by muscle action, and possess a massive and complex muscular system when they hatch. In addition, they have muscular specializations such as the anterior tentacles and the posterior sucker that are used as soon as they hatch. By contrast, juveniles of Mesostoma and larvae of polyclads move predominantly by ciliary action that may not require a complex neural circuitry for coordination. (C) 2001 Wiley-Liss, Inc.

Species and regional variations in the effectiveness of antivenom against the in Vitro neurotoxicity of death adder (Acanthophis) venoms

Although viperlike in appearance and habit, death adders belong to the Elapidae family of snakes. Systemic envenomation represents a serious medical problem with antivenom, which is raised against Acanthophis antarcticus venom, representing the primary treatment. This study focused on the major Acanthophis variants from Australia and islands in the Indo-Pacific region. Venoms were profiled using liquid chromatography-mass spectrometry, and analyzed for in vitro neurotoxicity (0.3-10 mug/ml), as well as the effectiveness of antivenom. (1-5 units/ml; 10 min prior to the addition of 10 mug/ml venom). The following death adder venoms were examined: A. antarcticus (from separate populations in New South Wales, Queensland, South Australia, and Western Australia), A. hawkei, A. praelongus, A. pyrrhus, A. rugosus, A. wellsi, and venom from an unnamed species from the Indonesian island of Seram. All venoms abolished indirect twitches of the chick isolated biventer cervicis nerve-muscle preparation in a dose-dependent manner. In addition, all venoms blocked responses to exogenous acetylcholine (1 m-M) and carbachol (20 muM), but not KCl (40 mM), suggesting postsynaptic neurotoxicity. Death adder antivenom (1 unit/ml) prevented the neurotoxic effects of A. pyrrhus, A. praelongus, and A. hawkei venoms, although it was markedly less effective against venoms from A. antarcticus (NSW, SA, WA), A. rugosus, A. wellsi, and A. sp. Scram. However, at 5 units/ml, antivenom was effective against all venoms tested. Death adder venoms, including those from A. antarcticus geographic variants, differed not only in their venom composition but also in their neurotoxic activity and susceptibility to antivenom. For the first time toxicological aspects of A. hawkei, A. wellsi, A. rugosus, and A. sp. Seram venoms were studied. (C) 2001 Academic Press.

The expression of fibroblast growth factors and their receptors in the embryonic and neonatal mouse inner ear

Four different fibroblast growth factor receptors (FGFR) are known, three of which have splice variants (known as the b and c variants) in the FGF-binding domain, to give different patterns of sensitivity to the different FGFs. The expression of the b and c variants of the FGF receptors. together with the expression of the ligands FGF1. FGF2, FGF3, FGF7, FGF8b and FGF8c, was determined by quantitative reverse transcription-polymerase chain reaction in developing whole mouse inner ears, and in dissected components of the postnatal mouse inner ear. At embryonic age (E)10.5 days, when the otocyst is a simple closed sac, the receptor most heavily expressed was FGFR2b, relative to the postnatal day 0 level. Over the period E10.5-E12.5. during which the structures of the inner ear start to form, the expression of the different FGF receptors increased 10(2)-10(4) fold per unit of tissue, and there was a gradual switch towards expression of the 'c' splice variants of FGFR2 and FGFR3 rather than the 'b' variants. At E10.5, the ligands most heavily expressed, relative to the postnatal day 0 level, were FGF3, FGF8b and FGF8c. In the postnatal inner eat. the patterns of expression of receptors and ligands tended to be correlated, such that receptor variants were expressed in the same regions as the ligands that are known to activate them effectively. The neural/sensory region expressed high levels of FGFR3c, and high levels of the ligand FGF8b. The same area also expressed high levels of FGFR1b and FGFR2b, and high levels of FGF3. The lateral wall of the cochlea (including the stria vascularis and the spiral ligament) expressed high levels of FGFR1c and FGF1. 11 is suggested that the different FGF receptors and ligands are expressed in a spatially coordinated pattern to selectively program cochlear development. (C) 2001 Elsevier Science B.V. All rights reserved.

Perceived constraints and social support for active leisure among mothers with young children

Evidence suggests that women who are mothers of young children have lower levels of physical activity than women of similar age who do not have children (Brown, Lee, Mishra, & Bauman, 2000). The purposes of this study were to explore the factors that constrain mothers of young children from being more physically active, and the relationship between physical activity and levels of social support available to the women. The empirical basis for this examination was provided through a study of activity levels and barriers to physical activity experienced by a sample of 543 mothers of young children from differing socioeconomic backgrounds. The data indicate that: (a) more than two-thirds of the mothers were inadequately active in their leisure time for health benefit; (b) while the vast majority of mothers expressed a desire to be more active, they were inhibited in their ability to act out their leisure preferences by a combination of structural (e.g., lack of time, money, energy) and ideological influences (e.g., sense of commitment to others); (c) access to social support (from partners, family, and friends) was seen to place some women in a better position than others to negotiate constraints that inhibit leisure participation; and (d) within groups of varying socioeconomic status (SES) there was wide variation in the amount of time spent each week in active leisure.

Overexpression of a Slit homologue impairs convergent extension of the mesoderm and causes cyclopia in embryonic zebrafish

Slit is expressed in the midline of the central nervous system both in vertebrates and invertebrates. In Drosophila, it is the midline repellent acting as a ligand for the Roundabout (Robo) protein, the repulsive receptor which is expressed on the growth cones of the commissural neurons. We have isolated cDNA fragments of the zebrafish slit2 and slit3 homologues and found that both genes start to be expressed by the midgastrula stage well before the axonogenesis begins in the nervous system, both in the axial mesoderm, and slit2 in the anterior margin of the neural plate and slit3 in the polster at the anterior end of the prechordal mesoderm. Later, expression of slit2 mRNA is detected mainly in midline structures such as the floor plate cells and the hypochord, and in the anterior margins of the neural plates in the zebrafish embryo, while slit3 expression is observed in the anterior margin of the prechordal plate, the floorplate cells in the hindbrain, and the motor neurons both in the hindbrain and the spinal cord. To study the role of Slit in early embryos, we overexpressed Slit2 in the whole embryos either by injection of its mRNA into one-cell stage embryos or by heat-shock treatment of the transgenic embryos which carries the slit2 gene under control of the heat-shock promoter. Overexpression of Slit2 in such ways impaired the convergent extension movement of the mesoderm and the rostral migration of the cells in the dorsal diencephalon and resulted in cyclopia. Our results shed light on a novel aspect of Slit function as a regulatory factor of mesodermal cell movement during gastrulation. (C) 2001 Academic Press.

Begging and food distribution in crimson rosella (Platycercus elegans) broods: why don't hungry chicks beg more?

Altricial nestlings solicit food by begging and engaging in scramble competition. Solicitation displays can thus signal both hunger and competitive ability. I examined nestling solicitation and parental responses in crimson rosellas (Platycercus elegans), a species in which parents engage in complex patterns of food allocation and appear to control the distribution of food. By manipulating the hunger of individual chicks and entire broods, I assessed how chick behaviours and parental food allocation varied with hatching rank, level of hunger, and intensity of nestling competition. Last-hatched chicks begged more than first-hatched chicks irrespective of individual hunger levels. The two parents combined fed individually hungry chicks more, but mothers and fathers varied in their responses to begging chicks: fathers fed last-hatched chicks in proportion to their begging intensity, whereas mothers fed chicks equally. Since fathers generally allocate more food to first-hatched chicks, fathers appear to use begging rates to adjust food allocation to non-preferred chicks within the brood. When I manipulated brood hunger levels, begging rates increased for first- and last-hatched chicks suggesting that chick begging rates are sensitive to the level of competition. This study shows that begging by rosella chicks does not correlate with hunger in a straightforward way and that the primary patterns of food allocation by parents art: not influenced by chick begging. Thus the benefits of increased begging may be limited for nestlings in this species.

neptune, a Kruppel-like transcription factor that participates in primitive erythropoiesis in Xenopus

The specification of the erythroid lineage from hematopoietic stem cells requires the expression and activity of lineage-specific transcription factors. One transcription factor family that has several members involved in hematopoiesis is the Kruppel-like factor (KLF) family [1]. For example, erythroid KLF (EKLF) regulates beta -globin expression during erythroid differentiation [2-6]. KLFs share a highly conserved zinc finger-based DNA binding domain (DBD) that mediates binding to CACCC-box and GC-rich sites, both of which are frequently found in the promoters of hematopoietic genes. Here, we identified a novel Xenopus KLF gene, neptune, which is highly expressed in the ventral blood island (VBI), cranial ganglia, and hatching and cement glands. neptune expression is induced in response to components of the BMP-4 signaling pathway in injected animal cap explants. Similar to its family member, EKLF, Neptune can bind CACCC-box and GC-rich DNA elements. We show that Neptune cooperates with the hematopoietic transcription factor XGATA-1 to enhance globin induction in animal cap explants. A fusion protein comprised of Neptune's DBD and the Drosophila engrailed repressor domain suppresses the induction of globin in ventral marginal zones and in animal caps. These studies demonstrate that Neptune is a positive regulator of primitive erythropoiesis in Xenopus.

Significance of serum early pregnancy factor concentrations during pregnancy and embryonic development in Sminthopsis macroura (Spencer) (Marsupialia : Dasyuridae)

Marsupial pregnancy differs from that in eutherians in duration, placentation and hormonal profile so much so that maternal recognition of pregnancy may not occur in polyovular marsupials. However, a comparison of gravid and non-gravid uteri reveals differences indicative of histological and physiological adaptations to pregnancy. In the present study, the hypothesis that embryo-maternal signalling occurs in polyovular marsupials was tested by examining serum from non-pregnant and pregnant Sminthopsis macroura for the presence of early pregnancy factor (EPF), a serum protein secreted by the ovary in response to the presence of a newly fertilized egg in the oviduct. EPF is detectable in the serum of pregnant, but not in non-pregnant, females in all eutherians studied to date. In the present study, EPF was detected in S. macroura serum by the rosette inhibition test during the first 9 days of the 10.7 day gestation period in this marsupial. However, EPF was not detected on day 10, just before parturition, or in non-pregnant or preovulatory animals. Immunohistochemical analysis of ovaries from gravid and non-gravid animals demonstrates that EPF is found in the capillaries, interstitial spaces and secretory cells of the corpus luteum. It is concluded that the spatiotemporal pattern of EPF activity described strongly indicates that maternal recognition of pregnancy in marsupials is mediated, at least in part, by EPF. Because the endocrinological milieu is the same in pregnant and non-pregnant marsupials, the possibility of using marsupials as an experimental system for studying EPF function unconfounded by hormonal effects is presented.

SOX18 and the transcriptional regulation of blood vessel development

SOX18 is a transcription factor that is transiently expressed in nascent endothelial cells during embryonic development and adult neovascularization. This protein belongs to the SOX family of transcription factors, ih,which are proving to be some of the key regulators of cell-type specification in the vertebrate embryo. Natural mutations in the Sox18 gene have been shown to result to cardiovascular dysfunction, in some cases leading to death. Available evidence thus implicates Sox18 as an important regulator of vascular development, most likely playing a key role in endothelial cell specification. However; the genetic knockout of Sox18 in mice has produced a confounding result that complicates our understanding of the molecular mode of action of the SOX18 protein. We speculate that Sox18 inky act in a redundant fashion with closely related genes such as Sox7 and/or Sox17. (C) 2001, Elsevier Science Inc.

Sox18 expression in blood vessels and feather buds during chicken embryogenesis

Sox18 encodes a transcription factor known to be important for the development of blood vessels and hair follicles in mice. In order to study the functional conservation of this gene through evolution, we have isolated and characterized Sox18 in chickens. cSox18 shows a high degree of sequence homology to both the mouse and human orthologues, particularly in the high mobility group DNA-binding domain and to a lesser extent in the transcriptional activation domain. A region of unusually high sequence conservation at the C-terminus may represent a further, previously unrecognized functional domain. Both the chicken and human proteins appear to be truncated at the N-terminus relative to mouse SOX18. In situ hybridization analyses showed expression in the developing vasculature and feather follicles, consistent with reported expression in the mouse embryo. In addition, cSox18 mRNA was observed in the retina and claw beds. (C) 2001 Elsevier Science B.V. All rights reserved.

Gonad development: Signals for sex

The formation of testes or ovaries in the mammalian embryo is critical in determining sexual identity and the ability to reproduce. Recent studies have begun to illuminate the cellular signalling events required for development of functional testes. (C) 2001 Elsevier Science Ltd. All rights reserved.

Regulation of male sexual development by Sry and Sox9

Sry, a gene from the Y chromosome, is known to initiate testis formation and subsequent male differentiation in mammals. A related gene, Sox9, also plays a critical role in testis determination, possibly in all vertebrates. A number of models have been presented regarding the molecular modes of action of these two genes. However, details regarding their regulation, regulatory target genes, and interacting protein factors and co-factors have not been established with any certainty. In this review, we examine new evidence and re-examine existing evidence bearing on these issues, in an effort to build up an integrative model of the network of gene activity centred around Sry and Sox9. J. Exp. Zool. 290:463-474, 2001. (C) 2001 Wiley-Liss, Inc.

In situ hybridization to mRNA: from black art to guiding light

In situ hybridization to mRNA in embryo sections or wholemount embryos is one of the most powerful analytical tools available to the molecular developmental biologist. For many workers, this procedure provides the first insights into the function of newly isolated genes, allowing the formulation of hypotheses and setting the course for further research. This paper presents a personal historical perspective of the development of in situ hybridization, looks at the theory and practice of the technique, summarizes the current state of the art, and speculates on possible directions for the future as a tool in functional genomics.

Embryogenesis and metamorphosis in a haplosclerid demosponge: gastrulation and transdifferentiation of larval ciliated cells to choanocytes

Early development and metamorphosis of Reniera sp., a haplosclerid demosponge, have been examined to determine how gastrulation occurs in this species, and whether there is an inversion of the primary germ layers at metamorphosis. Embryogenesis occurs by unequal cleavage of blastomeres to form a solid blastula consisting micro- and macromeres; multipolar migration of the micromeres to the surface of the embryo results in a bi-layered embryo and is interpreted as gastrulation. Polarity of the embryo is determined by the movement of pigment-containing micromeres to one pole of the embryo; this pole later becomes the posterior pole of the swimming larva. The bi-layered larva has a fully differentiated monociliated outer cell layer, and a solid interior of various cell types surrounded by dense collagen. The pigmented cells at the posterior pole give rise to long cilia that are capable of responding to environmental stimuli. Larvae settle on their anterior pole. Fluorescent labeling of the monociliated outer cell layer with a cell-lineage marker (CMFDA) demonstrates that the monociliated cells resorb their cilia, migrate inwards, and transdifferentiate into the choanocytes of the juvenile sponge, and into other amoeboid cells. The development of the flagellated choanocytes and other cells in the juvenile from the monociliated outer layer of this sponge's larva is interpreted as the dedifferentiation of fully differentiated larval cells-a process seen during the metamorphosis of other ciliated invertebrate larvae-not as inversion of the primary germ layers. These results suggest that the sequences of development in this haplosclerid demosponge are not very different than those observed in many cnidarians.

Homeotic genes influence the axonal pathway of a Drosophila embryonic sensory neuron

Each abdominal hemisegment of the Drosophila embryo has two sensory neurons intimately associated with a tracheal branch. During embryogenesis, the axons of these sensory neurons, termed the v'td2 neurons, enter the CNS and grow toward the brain with a distinctive pathway change in the third thoracic neuromere. We show that the axons use guidance cues that are under control of the bithorax gene complex (BX-C). Pathway defects in mutants suggest that a drop in Ultrabithorax expression permits the pathway change in the T3 neuromere, while combined Ultrabithorax and abdominal-A expression represses it in the abdominal neuromeres. We propose that the axons do not respond to a particular segmental identity in forming the pathway change; rather they respond to pathfinding cues that come about as a result of a drop in BX-C expression along the antero-posterior axis of the CNS.