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Biomedical subjects

A Lumsden

Publications and source records attributed to A Lumsden.

At least 73 records · Page 4Linked to original sources
PubMed1996

Rhombencephalic neural crest segmentation is preserved throughout craniofacial ontogeny.

To investigate the influence of hindbrain segmentation on craniofacial patterning we have studied the long term fate of neural crest (NC) subpopulations of individual rhombomeres (r), using quail-chick chimeras. Mapping of all skeletal and muscle connective tissues developing from these small regions revealed several novel features of the cranial neural crest. First, the mandibular arch skeleton has a composite origin in which the proximal elements are r1+r2 derived, whereas more distal ones are exclusively midbrain derived. The most proximal region of the lower jaw is derived from second arch (r4) NC. Second, both the lower jaw and tongue skeleton display an organisation which precisely reflects the rostrocaudal order of segmental crest deployment from the embryonic hindbrain. Third, cryptic intraskeletal boundaries, which do not correspond to anatomical landmarks, form sharply defined interfaces between r1+r2, r4 and r6+r7 crest. Cells that survive the early apoptotic elimination of premigratory NC in r3 and r5 are restricted to tiny contributions within the 2nd arch (r4) skeleton. Fourth, a highly constrained pattern of cranial skeletomuscular connectivity was found that precisely respects the positional origin of its constitutive crest: each rhombomeric population remains coherent throughout ontogeny, forming both the connective tissues of specific muscles and their respective attachment sites onto the neuro- and viscerocranium. Finally, focal clusters of crest cells, confined to the attachment sites of branchial muscles, intrude into the otherwise mesodermal cranial base. In the viscerocranium, an equally strict, rhombomere-specific matching of muscle connective tissues and their attachment sites is found for all branchial and tongue (hypoglossal) muscles. This coherence of segmental crest populations explains how cranial skeletomuscular pattern can be implemented and conserved despite evolutionary changes in the shapes of skeletal elements.

Animals↗
PubMed1996

Interactions between rhombomeres modulate Krox-20 and follistatin expression in the chick embryo hindbrain.

The rhombomeres of the embryonic hindbrain display compartment properties, including cell lineage restriction, genetic definition and modular anatomical phenotype. Consistent with the idea that rhombomeres are autonomous developmental units, previous studies have shown that certain aspects of rhombomere phenotype are determined early, at the time of rhombomere formation. By contrast, the apoptotic depletion of neural crest from rhombomeres 3 and 5 is due to an interaction with their neighbouring rhombomeres, involving the signalling molecule Bmp4. In this paper, we have examined whether inter-rhombomere interactions control further aspects of rhombomere phenotype. We find that the expression of Krox-20 and the repression of follistatin in r3 is dependent upon neighbour interaction, whereas these genes are expressed autonomously in r5. We further demonstrate that modulation of Krox-20 and follistatin expression is not dependent on Bmp4, indicating the existence of multiple pathways of interaction between adjacent rhombomeres. We also show that, although some phenotypic aspects of r3 are controlled by neighbour interactions, the axial identity of the segment is intrinsically determined.

Amino Acid Sequence↗
PubMed1996

Late effects of retinoic acid on neural crest and aspects of rhombomere.

We exposed st.10 chicks to retinoic acid (RA), both globally, and locally to individual rhombomeres, to look at its role in specification of various aspects of hindbrain derived morphology. Previous studies have looked at RA exposure at earlier stages, during axial specification. Stage 10 is the time of morphological segmentation of the hindbrain and is just prior to neural crest migration. Rhombomere 4 localised RA injections result in specific alterations of pathways some crest cells that normally migrate to sites of differentiation of neurogenic derivatives. The r4 crest cells that give rise to mesenchymal derivatives are unaffected. In addition, r4 gene expression is also partially altered by RA; within 6 hours of r4 exposure to RA, ectopic expression of Krox-20 is seen in r4 and Hoxb-1 expression is lost while Hoxa-2 expression continues normally. When we examined these RA-treated animals later in development, they showed an anterior displacement of the facial ganglion in addition to a mis-direction of the extensions of its distal axons and a dramatic decrease in the number of contralateral vestibuloacoustic neurons normally seen in r4. Only this r4-specific neuronal type is affected in r4; the motor neuron projections seem normal in experimental animals. The specificity of this result, combined with the loss of Hoxb-1 expression in r4 and the work by Krumlauf and co-workers showing gain of contralateral neurons co-localised with ectopic Hoxb-1 expression, indicates a role for Hoxb-1 and RA in the specification of this cell type in normal development. These results suggest that RA, at st.10, is able to affect some aspects of segment identity while leaving others unchanged.

Animals↗
PubMed1996

Persistence of rhombomeric organisation in the postsegmental hindbrain.

Rhombomeres are morphological varicosities of the neural tube that are present between embryonic day (E) 1.5 and E5 and are characterised by compartment organisation, segmentally neuronal organisation and spatially restricted patterns of gene expression. After E5, the segmented origins of the hindbrain become indistinct, while the adult hindbrain has an longitudinal columnar nuclear organisation. In order to assess the impact of the early transverse pattern on later longitudinal organisation, we have used orthotopic quail grafts and in situ hybridisation to investigate the long-term fate of rhombomeres in the embryonic chick hindbrain. The uniformity of mixing between quail and chick cells was first verified using short-term aggregation cultures. The dispersal of the progeny of individual rhombomeres (r) was then assessed by the unilateral, isochronic and orthotopic transplantation of either r2, r3, r4, r5 or r6 from quail to chick at embryonic day E2. In addition, orthotopic, partial rhombomere grafts, encompassing an inter-rhombomere boundary and adjacent rhombomere bodies were used to assess cell mixing within rhombomeres. Operated embryos were incubated to either E7 or E10 when chimaeric brains were removed. Quail cells were identified in whole mounts or serial sections using the quail-specific antibody QCPN. Subsequently, radial glia morphology was assessed either by immunohistochemistry or DiI labelling. A series of fixed hindbrains between E6 and E9 were probed for transcripts of Hoxa-2 and Hoxb-1. Fate-mapping reveals that the progeny of individual rhombomeres form stripes of cells running dorsoventrally through the hindbrain. This pattern of dispersal precisely parallels the array of radial glia. Although the postmitotic progeny of adjacent rhombomeres spread to some extent into each others' territory in intermediate and marginal zones, there is little or no mixing between rhombomeres in the ventricular zone, which thus remains compartmentalised long after the rhombomeric morphology disappears. Segmental gene expression within this layer is also maintained after E5. A more detailed analysis of mixing between proliferating cells, using partial rhombomere grafts, reveals that both mixing and growth are non-uniform within the ventricular layer, suggesting, in particular, that longitudinal expansion within this layer is restricted. Together, these observations suggest that rhombomeres do not disappear at E5, as has previously been supposed, rather they persist in the ventricular zone to at least E9, ensuring a continuity in the presumed segmental cues that specify neuroepithelial cells in the hindbrain.

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PubMed1996

Late emigrating neural crest cells migrate specifically to the exit points of cranial branchiomotor nerves.

Morphological segmentation of the avian hindbrain into rhombomeres is also reflected by the emergent organisation of branchiomotor nerves. In each case, the motor neurons of these nerves lie in two adjacent rhombomeres (e.g. of the Vth nerve in r2 and r3, VIIth in r4 and r5 etc.), and their outgrowing axons emerge into the periphery through defined exit points in rhombomeres r2, r4 and r6, respectively. Sensory axons of the cranial ganglia also enter the neuroepithelium at the same points. Motor axon outgrowth through experimentally rotated rhombomeres has suggested that a chemoattractive mechanism, involving the exit points, may form a component of their guidance. Yet so far, nothing is known about the establishment of the exit points or the identity of the cells that form them. In this study, we describe a group of late emigrating cranial neural crest cells which populate specifically the prospective exit points. Using chimaeras in which premigratory chick neural crest had been replaced orthotopically by quail cells, a population of neural crest was found to leave the cranial neural tube from about stage 10+ onwards and to migrate directly to the prospective exit points. These cells define the exit points by stage 12+, long before either motor or sensory axons have grown through them. The entire neural crest population of exit point cells expresses the recently described cell adhesion molecule c-cad7. Further, heterotopic grafting experiments show that midbrain and spinal cord crest, grafted at late stages in place of r4 crest, share the same migratory behaviour to the facial nerve exit points and express the same markers as cells contributed by the native r4 crest. It was not possible to generate new exit points in odd numbered rhombomeres simply by experimentally increasing their (normally insignificant) amount of crest production. Initiation of the exit point region probably lies, therefore, in the neuroepithelium.

Animals↗
PubMed1996

Patterning the cranial neural crest.

The pattern of skeletal elements and musculature in the branchial region of the chick embryo is profoundly influenced by the neural crest which migrates in separate streams. Neural crest emigration from the hindbrain into the branchial arches is discontinuous, and regions of crest production are separated from each other by two regions of crest apoptosis, rhombomeres (r) 3 and 5. Both r3 and r5 produce large numbers of crest cells when they are cultured in isolation; but, when co-cultured with their neighbouring segment neural crest apoptosis is restored. The expression of the signalling molecule Bmp-4 and the gene msx-2 in apoptotic domains of r3 and r5 is also dependent upon neighbour interactions and the addition of recombinant Bmp-4 to explant cultures of r3/r5 upregulates both Bmp-4 and msx-2 expression and restores apoptosis. This interactive mechanism acts to ensure the sculpting of crest outflow into non-mixing streams and, consequently, the fidelity of pattern transfer into the branchial arches.

Animals↗
PubMed1995

Rhythm generation in the segmented hindbrain of chick embryos.

1. The embryonic hindbrain of chick is segmented until stage 24, when it starts to generate rhythmic activities in cranial nerves. In order to recognize a possible influence of segmentation on the organization of neuronal systems generating motor rhythms, the activity of trigeminal, facial, glossopharyngeal, vagal and hypoglossal nerves was studied during embryonic stages 24-36, by simultaneous recording of different cranial nerves in an isolated, superfused chick hindbrain preparation. 2. Highly correlated recurrent episodes of cyclical burst discharges occurred in all nerves studied (correlation coefficients, 0.8 +/- 0.1) throughout stages 24-36. 3. Such coactivation is unlikely to be due to monosynaptic connections between widely divergent premotor neurons and motoneurons, or between motoneurons themselves, because no short-term correlation was apparent in the millisecond range between activities of different motor nerves. 4. Complete transverse or midsagittal sectioning of the hindbrain disrupted coactivation of nerves located at distinct rostrocaudal levels or occupying an ipsi- or contralateral position, respectively, while sparing the ability of individual nerves to generate rhythmic activity. Each hindbrain segment thus contains bilaterally the motor nuclei together with their own rhythm generator. Coactivation of motor patterns appears to result from intersegmental and cross-median connections between these rhythm generators. 5. The results are in keeping with the hypothesis of a segmental organization of the primordial hindbrain rhythm generator and give further support to the early determination of both the anatomical and the functional fate of neurons in this region of the vertebrate central nervous system.

Animals↗
PubMed1995

A role for FGF-8 in the initiation and maintenance of vertebrate limb bud outgrowth.

BACKGROUND: The outgrowth of the vertebrate limb bud is the result of a reciprocal interaction between the mesenchyme and a specialized region of the ectoderm, the apical ectodermal ridge (AER), which overlies it. Signals emanating from the AER act to maintain the underlying mesenchyme, called the progress zone, in a highly proliferative and undifferentiated state. Removal of the AER results in the cessation of limb bud growth, thus causing limb truncation. The best candidates for this AER-derived signal are members of the fibroblast growth factor (FGF) family, in particular FGF-4, which can maintain limb bud outgrowth following removal of the AER. However, FGF-4 is only expressed after considerable outgrowth has occurred and a well-developed limb bud has formed, and then only in the posterior part of the AER. Likewise, the other FGFs studied to date are not candidates for this activity. RESULTS: We report evidence that a recently identified member of this family, FGF-8, is expressed in the ectoderm of the prospective limb territory prior to morphological outgrowth of the limb bud in both mouse and chick. Thereafter, expression is maintained throughout the AER during limb development. We have produced and purified the FGF-8 protein, and shown that it will substitute for the AER in maintaining limb bud outgrowth in mouse embryos from which the AER has been surgically removed. FGF-8 does not, however, maintain expression of the sonic hedgehog gene. CONCLUSIONS: These results indicate that FGF-8 is an AER-derived mitogen that stimulates limb bud outgrowth. Moreover, our data suggest that FGF-8 may also be an ectodermally derived mitogen that stimulates the onset of limb bud outgrowth (budding) in the absence of a morphological AER, and indicate the possible involvement of FGF-8 in the establishment of the limb field.

Amino Acid Sequence↗
PubMed1995

Independent assignment of antero-posterior and dorso-ventral positional values in the developing chick hindbrain.

BACKGROUND: Cell patterning in the developing central nervous system seem to involve a coordinate system of positional information, in which specific fates are assigned to multipotent precursor cells by positional signals acting on the antero-posterior and dorso-ventral axes of the neural tube. Before neurons differentiate in the hindbrain, it becomes subdivided antero-posteriorly into a series of developmental compartments, the rhombomeres. When the rhombomeres are delineated from each other by interfaces at which cell mixing is transiently restricted, they are determined for expression of specific selector Hox genes that may encode aspects of their individual identity. To assess whether the phenotypic identities of the rhombomeres are also determined at this stage, we have analyzed the capacity of individual rhombomeres to realize specific neuronal fates when grafted heterotopically along both antero-posterior and dorso-ventral axes. RESULTS: When rhombomere 4 (r4) is grafted unilaterally to the r2 position, both facial motor neurons and contralateral vestibulo-acoustic efferent neurons differentiate, as normal, in the ventral region of the graft. These aspects of phenotypic identity therefore appear to have been determined at or before the time of grafting. When r4 is grafted to the r2 position with its dorso-ventral polarity inverted, both types of neuron again develop, but in the ventral region of the graft, in a position appropriate to the dorso-ventral pattern of the host, rather than their original dorso-ventral position. The change in fate of these cells is restricted, however, to the repertoire characteristic of the antero-posterior position of origin, in this case r4. CONCLUSIONS: Cells seem to 'know' details of their presumptive fate before more general features. At this stage of development, precursor cells in r4 seem to have been assigned an 'r4 fate', but remain multipotent in their choice of r4-specific cell type. Precursor cells seem to be committed to their fates according to position on an orthogonal grid, the coordinates of which are set (or read) independently and sequentially. Thus, at the 7-10 somite stage, dorso-ventral positional values are still labile, whereas antero-posterior values are already fixed.

Animals↗
PubMed1995

Cell and matrix specialisations of rhombomere boundaries.

Hindbrain segments, rhombomeres, define distinct cellular and molecular domains which furnish the ground plan for important aspects of neural and cranial development. In this study, further evidence is presented that the interfaces between rhombomeres, rhombomere boundaries, contain both cells and extracellular matrix with specialised characteristics. Cells at rhombomere boundaries show temporally and spatially distinct expression patterns of developmentally important genes. Towards the end of the developmental period when rhombomeres are present, a fan-shaped array of cells at rhombomere boundaries, that constitute the ventricular ridge, shows decreased expression of two genes (Hoxb-1 and Krox-20), which earlier in development were expressed in all cells of specific rhombomeres. In contrast, these boundary cells show increased expression of another gene, Pax-6, which earlier in development has a rhombomere-specific expression pattern. A specialised identity for boundary cells is further suggested by increased labelling with an anti-vimentin antibody at rhombomere boundaries, indicating that at least some boundary cells are radial glia or glial precursors. In addition to distinct cellular properties, the extracellular domain at rhombomere boundaries is also specialised. Chondroitin sulphate proteoglycan (CSPG) immunoreactivity is increased and, as revealed by immuno-electron microscopy, localised to extracellular spaces. CSPG is also enriched in boundaries regenerated after ablation, or boundaries generated ectopically by rhombomere transplantation. We propose that rhombomere boundaries form their characteristic morphology at the interface between groups of cells with differing molecular characteristics, representing different cell states. A specialised band of cells then develops at the interface. Both the boundary cells and extracellur matrix have characteristics which could be important in later events of neural development such as axon guidance and cell migration.

Animals↗
PubMed1995

Development expression of Hox11 and specification of splenic cell fate.

Hox11 is the first member of a novel class of orphan homeobox genes. We report that Hox11 is expressed in a discrete temporal and spatially segmented pattern during embryonic development and appears critical for the specification of splenic cell fate. Expression is first observed in the developing muscle plates of branchial arches 1, 2, 3 and 4/6, and subsequently within motor neurons of cranial nerves V, VII, IX, and X, which innervate these muscles. Hox11 serves as a molecular maker distinguishing branchial from somatic motor nuclei. Additionally, Hox11 is expressed in the surface ectoderm of the first branchial arch in the region destined to become the tongue and teeth and then in ganglia innervating this area. However, Hox11-deficient mice have no apparent morphological of functional defects within these structures. Notably the closely related homeobox genes, Hox11L.1 and Hox1L1.2, were not expressed in a redundant pattern. Neither Hox11L1 nor Hox11L2 was expressed in the branchial arches or their motor nuclei within wild-type or Hox11-/- mice. Beginning at E11.5, Hox11 is normally expressed at a single site in the abdomen within splanchnic mesoderm destined to form the spleen, and Hox11-/- mice have no spleen. We noted no increase in cell death within the dorsal mesogastrium of Hox11-deficient mice. Instead the dorsal mesogastrium fails to separate from the stomach. Hox11-/- mice display a larger stomach and possibly pancreas, suggesting that these mesodermal cells now contribute to other organs.

Animals↗
PubMed1994

The signalling molecule BMP4 mediates apoptosis in the rhombencephalic neural crest.

The pattern of skeletal structures and muscles in the branchial region of the head is profoundly influenced by the neural crest, whose cells arise at discrete segmental levels of the chick hindbrain: specifically, rhombomeres (r)1+2, r4 and r6, whereas r3 and r5 are crest-depleted. We have demonstrated that an interaction between even-numbered rhombomeres and r3/r5 effects this depletion of neural crest, resulting in the sculpting of discrete migratory streams of neural crest. This mechanism acts through increased expression of msx2 and the induction of apoptosis in dorsal cells of r3 and r5 (ref. 3) (Fig. 1A). Here we demonstrate that the signalling molecule Bmp4 is expressed in r3 and r5 and is dependent on the neighbouring rhombomeres. Addition of recombinant BMP4 protein to explant cultures of r3 or r5, which produce neural crest when isolated from their neighbouring rhombomeres, upregulates msx2 and reinstates apoptosis in the neural crest population.

Animals↗
PubMed1994

Onset and maturation of branchio-motor activities in the chick hindbrain.

Branchio-motor activities from trigeminal, facial, glossopharyngeal and vagal nerves, in chick embryos have been recorded using suction electrodes on an isolated preparation of the hindbrain in vitro between developmental stages 20 and 36. They were composed of recurring episodes of cyclical burst discharges first identified at stage 24, therefore constituting one of the earliest organized activities generated in the chick central nervous system. Between stage 24 and 36, both the period between episodes and the number of bursts per episode were increased. This maturation sequence was preserved for several hours in vitro in the absence of supraspinal and sensory inputs. Results are in agreement with rhythmogenic properties constituting an early functional commitment of neuronal networks in this particular region of the neuroepithelium.

Animals↗
PubMed1994

Regulation of SC1/DM-GRASP during the migration of motor neurons in the chick embryo brain stem.

The hindbrain of the chick embryo contains three classes of motor neurons: somatic, visceral, and branchial motor. During development, somata of neurons in the last two classes undergo a laterally directed migration within the neuroepithelium; somata translocate towards the nerve exit points, through which motor axons are beginning to extend into the periphery. All classes of motor neuron are immunopositive for the SC1/DM-GRASP cell surface glycoprotein. We have examined the relationship between patterns of motor neuron migration, axon outgrowth, and expression of the SC1/DM-GRASP mRNA and protein, using anterograde or retrograde axonal tracing, immunohistochemistry, and in situ hybridization. We find that as motor neurons migrate laterally, SC1/DM-GRASP is down-regulated, both on neuronal somata and axonal surfaces. Within individual motor nuclei, these lateral, more mature neurons are found to possess longer axons than the young, medial cells of the population. Labelling of sensory or motor axons growing into the second branchial arch also shows that motor axons reach the muscle plate first, and that SC1/DM-GRASP is expressed on the muscle at the time growth cones arrive.

Activated-Leukocyte Cell Adhesion Molecule↗
PubMed1994

Characterization of thrombin receptor expression during vascular lesion formation.

Blood vessels respond to injury by initiating cell proliferation and migration that result in vascular lesion formation. To determine the roles of thrombin and the thrombin receptor in this process, we characterized thrombin receptor expression in normal and injured arteries, thrombin receptor-mediated smooth muscle cell mitogenesis, and the regulation of thrombin receptor mRNA expression in vitro. Thrombin receptor mRNA was not detected in normal rat or baboon arteries by in situ hybridization. Immunohistochemistry using an antithrombin receptor antibody (TR-R9), directed against the thrombin cleavage site of the rat aortic smooth muscle cell thrombin receptor, revealed low-level staining for thrombin receptor protein in endothelial cells and smooth muscle cells of normal arteries. In contrast, balloon catheter injury increased thrombin mRNA expression in medial smooth muscle cells within 6 hours. This increased thrombin receptor expression continued within the media and in neointimal cells throughout vascular lesion formation, predominantly in areas of active cell proliferation. In vitro, alpha-thrombin stimulates rat aortic smooth muscle cell proliferation in a concentration-dependent manner. That thrombin receptor activation is required for the mitogenic response was confirmed by demonstrating that the polyclonal antibody TR-R9 inhibits thrombin-induced cell proliferation. Thrombin receptor mRNA synthesis was induced by both basic fibroblast growth factor (maximal stimulation of 1.8-fold at 1 hour) and platelet-derived growth factor (maximal stimulation of 2.4-fold at 8 and 24 hours) in quiesced cultured rat aortic smooth muscle cells. In summary, upregulation of smooth muscle cell thrombin receptor expression occurs very early after vascular injury and continues throughout neointimal development.(ABSTRACT TRUNCATED AT 250 WORDS)

Angioplasty↗
PubMed1994

Hoxa-2 expression in normal and transposed rhombomeres: independent regulation in the neural tube and neural crest.

In this study we have cloned the chick Hoxa-2 gene and analysed its expression during early development. We find that Hoxa-2 has a rostral limit of expression in the rhombencephalic neural tube corresponding precisely to the boundary between rhombomeres (r)1 and 2; a limit further rostral than any other Hox gene reported to date. Neural crest migrates from r2 to populate the first branchial arch, yet although Hoxa-2 is expressed down the full dorsoventral extent of r2 during the phase of neural crest emigration, there is no Hoxa-2 expression in either the emergent neural crest or in the first branchial arch. Conversely, at the level of r4, both the neural tube and the neural crest cells, which migrate out of this rhombomere to populate the second branchial arch, express Hoxa-2. Other Hox genes expressed in the rhombencephalic neural tube demonstrate a transfer of expression from neural tube to neural crest at all axial levels of expression. Hoxa-2 is thus unusual in demonstrating separate anterior expression limits in neural tube and neural crest; this allowed us to test whether Hox gene expression patterns in neural crest are determined by migratory pathways or are prespecified by the site of origin in the neuroepithelium. Grafting experiments in which pairs of rhombomeres were transplanted to ectopic sites at the time of rhombomere boundary formation reveal a prepatterning of the neural crest with respect to Hoxa-2 expression. The decision to down-regulate Hoxa-2 expression in r2-derived neural crest, but to maintain Hoxa-2 expression in r4-derived neural crest is intrinsic to the premigratory crest cell population. Thus, following grafting of r4 to the r2 site and vice-versa, Hoxa-2 expression is maintained in r4-derived neural crest, but lost in r2-derived neural crest.

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