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D Summerbell

Publications and source records attributed to D Summerbell.

At least 19 recordsLinked to original sources
PubMed1991

Retinoic acid stimulates neurite outgrowth in the amphibian spinal cord.

There is increasing evidence that retinoic acid (RA), a vitamin A metabolite, plays a role in the development of the nervous system. Here we specifically test this notion by examining the effect of RA on neurite outgrowth from explanted segments of the axolotl spinal cord. We show that there is a threshold concentration in the region of 0.1-1 nM above which neurite outgrowth is stimulated 4-5 fold. Retinol, by contrast, only stimulated the migration of glial cells from the explants. Using HPLC we demonstrate that RA and retinol are present endogenously in the axolotl spinal cord. In addition, we have identified by immunocytochemistry with antipeptide antibodies the cells of the spinal cord that contain the binding proteins for RA (cellular RA-binding protein; CRABP) and retinol (cellular retinol-binding protein; CRBP). CRABP is found in the axons and CRBP is found in the ependyma and glial cells. These results provide strong evidence for a role for RA in the developing nervous system, and we propose a specific hypothesis involving CRBP, CRABP, retinol, and RA in the control of axon outgrowth in the spinal cord.

Ambystoma↗
PubMed1991

The respecification of limb pattern by new synthetic retinoids and their interaction with cellular retinoic acid-binding protein.

We describe here experiments to examine the role of cellular retinoic-acid-binding protein (CRABP) during the induction of limb duplication in the chick limb bud and regenerating axolotl limb by retinoids. A newly synthesised class of retinoic acid analogues have been used because among them, some have been specifically designed with the property of binding to the retinoic acid receptors, but not to CRABP. We can thus test whether binding to CRABP is an obligatory step during limb respecification. The binding of four of these compounds to chick limb bud and axolotl CRABP was tested in sucrose density gradient assays and then their potencies at inducing limb duplications tested. Two of the four compounds do not bind to limb CRABP and yet are able to induce limb duplications, suggesting that an interaction with CRABP is not an obligatory step in the process. However, the two compounds which do bind to CRABP are more potent than the two which do not, suggesting that an interaction with CRABP may, nevertheless, increase the potency of a retinoid.

Ambystoma mexicanum↗
PubMed1991

Retinoic acid-binding protein, rhombomeres and the neural crest.

We have investigated by immunocytochemistry the spatial and temporal distribution of cellular retinoic acid-binding protein (CRABP) in the developing nervous system of the chick embryo in order to answer two specific questions: do neural crest cells contain CRABP and where and when do CRABP-positive neuroblasts first arise in the neural tube? With regard to the neural crest, we have compared CRABP staining with HNK-1 staining (a marker of migrating neural crest) and found that they do indeed co-localise, but cephalic and trunk crest behave slightly differently. In the cephalic region in tissues such as the frontonasal mass and branchial arches, HNK-1 immunoreactivity is intense at early stages, but it disappears as CRABP immunoreactivity appears. Thus the two staining patterns do not overlap, but are complementary. In the trunk, HNK-1 and CRABP stain the same cell populations at the same time, such as those migrating through the anterior halves of the somites. In the neural tube, CRABP-positive neuroblasts first appear in the rhombencephalon just after the neural folds close and then a particular pattern of immunoreactivity appears within the rhombomeres of the hindbrain. Labelled cells are present in the future spinal cord, the posterior rhombencephalon up to rhombomere 6 and in rhombomere 4 thus producing a single stripe pattern. This pattern is dynamic and gradually changes as anterior rhombomeres begin to label. The similarity of this initial pattern to the arrangement of certain homeobox genes in the mouse stimulated us to examine the expression of the chicken Hox-2.9 gene. We show that at stage 15 the pattern of expression of this gene is closely related to that of CRABP. The relationship between retinoic acid, CRABP and homeobox genes is discussed.

Animals↗
PubMed1990

Retinoic acid, a developmental signalling molecule.

Retinoic acid has been used as a tool both by embryologists studying the spatial organization of cells in the embryo and by molecular biologists studying the control of gene expression in the nucleus. Embryologists have shown that retinoic acid can modify the pattern of cell differentiation so as to duplicate complete parts of the embryo in a well-organized way; molecular biologists have shown that retinoic acid can act as the switch starting the sequence of differential gene expression that results in cell differentiation. In the past year these two approaches have converged so that there now seems a real possibility that we may soon for the first time understand how a particular vertebrate development system works.

Animals↗
PubMed1989

Cellular retinoic acid-binding protein and the role of retinoic acid in the development of the chick embryo.

The distribution of cellular retinoic acid-binding protein (CRABP) in four stages of chick development is described using an affinity-purified antibody against rat CRABP. CRABP is the protein to which retinoic acid (RA) binds when it enters cells and may reflect the requirement of those cells for RA. We found several discrete cell populations which showed high levels of immunoreactivity. Some were in the neural tube such as the commissural neurons and the dorsal roof plate. Some were of neural crest origin such as the dorsal root ganglia, sensory axons, sympathetic ganglia, and enteric ganglia. The remaining populations were certain connective tissue cells, limb bud cells, and the myotome. These results suggest that certain organ systems, particularly the nervous system, have a requirement for RA during development and they may further our understanding of the teratogenic effects of retinoids on the embryo.

Age Factors↗
PubMed1989

The role of retinoid-binding proteins in the generation of pattern in the developing limb, the regenerating limb and the nervous system.

We summarize existing data and describe new information on the levels and distribution of cellular retinoic acid-binding protein (CRABP) and cellular retinol-binding protein (CRBP) in the regenerating axolotl limb, the developing chick limb bud and the nervous system of the chick embryo in the light of the known morphogenetic effects of retinoids on these systems. In the regenerating limb, levels of CRABP rise 3- to 4-fold during regeneration, peaking at the time when retinoic acid (RA) is most effective at causing pattern duplications. The levels of CRBP are low. The potency of various retinoids in causing pattern respecification correlates well with the ability of these compounds to bind to CRABP. In the chick limb bud, the levels of CRABP are high and the levels of CRBP are low. Again the binding of various retinoids to CRABP correlates well with their ability to cause pattern duplications. By immunocytochemistry, we show that CRABP is present at high levels in the progress zone of the limb bud and is distributed across the anteroposterior axis in a gradient with the high point at the anterior margin. In the chick embryo, CRABP levels are high and CRBP levels are low. By immunocytochemistry, CRABP is localised primarily to the developing nervous system, labelling cells and axons in the mantle layer of the neural tube. These become the neurons of the commissural system. Also sensory axons label intensely with CRABP whereas motor axons do not and in the mixed nerves at the brachial plexus sensory and motor components can be distinguished on this basis. In the neural tube, CRBP only stains the ventral floor plate. Since the ventral floor plate may be a source of chemoattractant for commissural axons, we suggest on the basis of these staining patterns that RA may fulfill this role and thus be involved morphogenetically in the developing nervous system.

Ambystoma↗
PubMed1988

Spatial distribution of cellular protein binding to retinoic acid in the chick limb bud.

Retinoic acid may be the natural morphogen used to generate digit pattern in the chick limb bud. It has been proposed that retinoic acid acts by binding to a cellular retinoic acid-binding protein (CRABP) and then entering the nucleus to alter the pattern of gene activity. High-affinity receptors that bind both retinoic acid and DNA and are analogous to the steroid receptors have been identified. But the concentration of endogenous retinoic acid in the limb and the binding coefficient of the nuclear receptors indicate that they are saturated throughout the limb. Here we investigate the CRABP distribution in the developing chick limb bud. We find CRABP in the area of intense morphogenetic activity at the tip, with a differential distribution across the anteroposterior axis, the high point being at the anterior margin. Retinoic acid also forms a concentration gradient across the limb bud, but is highest on the posterior side. We propose that CRABP could be reducing the effective concentration of retinoic acid reaching the nucleus to a level appropriate for the differential regulation of gene transcription, providing a spatially modulated morphogenetic gradient of information for digit formation.

Animals↗
PubMed1988

Specific guidance of motor axons to duplicated muscles in the developing amniote limb.

The effect of alteration of limb pattern upon motor axon guidance has been investigated in chick embryos. Following grafting of the zone of polarizing activity (ZPA) into the anterior margin of the early limb bud, limbs develop with forearms duplicated about the anteroposterior axis. The position of motoneurones innervating the duplicated posterior forearm extensor EMU was mapped by retrograde transport of horse radish peroxidase (HRP). The motor pool labelled from injection into the anteriorly duplicated EMU muscle is consistently similar to that supplying the posterior EMU muscle on the unoperated side of the embryo. In those cases where the axons are well filled, their trajectories from the injection site are observed to change position within the radial nerve to specifically innervate the duplicated muscle. The axons modify their trajectories proximal to the level of limb duplication in a region where there is no change in the pattern of overt differentiation of the limb cells. This suggests that axons may use a cell's positional value to navigate and provides significant support for the theory of positional information.

Animals↗
PubMed1987

The specificity of motor innervation of the chick wing does not depend upon the segmental origin of muscles.

In vertebrate embryos, motor axons originating from a particular craniocaudal position in the neural tube innervate limb muscles derived from myoblasts of the same segmental level. We have investigated whether this relationship is important for the formation of specific nerve-muscle connections, by altering the segmental origin of muscles and examining their resulting innervation. First, by grafting quail wing somites to a new craniocaudal position opposite the chick wing, we established that the segmental origin of a muscle can be altered: presumptive muscle cells migrated according to their new, rather than their original, somitic level, colonizing a different subset of muscles. However, after reversal of a length of brachial somitic mesoderm along the craniocaudal axis, or exchange or shift of brachial somites, the craniocaudal position of wing muscle motoneurone pools within the spinal cord was undisturbed, despite the new segmental origin of the muscles themselves. While not excluding the possibility that muscles and their motor nerves are labelled segmentally, we conclude that specific motor axon guidance in the wing does not depend upon the existence of such labels.

Animals↗
PubMed1986

Timing of ovulation by determination of the urinary luteinizing hormone surge with an enzyme-linked monoclonal antibody dipstick (OvuStick).

An enzyme-linked double monoclonal antibody dipstick for measuring luteinizing hormone in urine was tested for timing ovulation in thrice daily urine samples collected for several days around mid-cycle in 24 women undergoing artificial insemination. The assay produced information comparable to single daily serum LH measurements. The dipsticks could be used by untrained people to test their own urine as an aid to the detection of ovulation for the timing of artificial insemination or of sexual intercourse to promote or avoid conception.

Antibodies, Monoclonal↗
PubMed1986

Retinoic acid-binding protein in the chick limb bud: identification at developmental stages and binding affinities of various retinoids.

The application of retinoic acid (RA) to the developing chick limb bud causes 6-digit double posterior limbs to form instead of the normal 3-digit limb. As an attempt to begin a molecular analysis of this phenomenon we have identified and characterized a soluble cytoplasmic receptor for RA, namely cytoplasmic retinoic acid-binding protein (CRABP), from the cells of the chick limb bud. It is present from stages 20-35 at similar levels and has an apparent Kd of 140-280 nM. In competition experiments with other retinoids Ro 13-7410 was found to be the most effective at competing for sites on CRABP followed by all-trans-RA, 13-cis-RA, Ro 10-1670 and retinal. Retinol, retinyl palmitate, retinyl acetate, etretinate and arotinoid showed low or no affinity for CRABP. Specificity for binding was thus demonstrated since analogues with an acid end group competed effectively, the aldehyde competed less effectively and the ester or alcohol groups did not compete. At the concentration of RA that needs to be administered to cause duplications in the pattern of the limb bud, we estimate that 4% of the CRABP present in the limb bud has RA bound. The similarities between steroid receptors in the mediation of steroid hormone action and CRABP in the mediation of RA action is discussed. In this regard we note that while there are 10(4) steroid receptors per cell in other cell types we estimate that there are about 10(5) RA receptors per cell in the chick limb bud.

Animals↗
PubMed1985

The behaviour of growing axons invading developing chick wing buds with dorsoventral or anteroposterior axis reversed.

The trajectories of motor axons innervating chick wings reversed about the DV or AP axis before axon invasion were analysed after retrograde filling by HRP injection into biceps or triceps muscles. Chick-quail chimaeras showed that the plane of reversal for flank operations was proximal to the confluence of the 14th, 15th, and 16th spinal roots as they form the plexus. The shoulder reversal plane was distal to the plexus. In dorsoventral (DV) reversed wings at both shoulder or flank level, the motor axons do not alter their course as they enter the graft. They therefore innervate by passive deployment any target that they encounter. In anteroposterior (AP) reversed wings at both shoulder or flank level, the motor axons clearly corrected their position in the nerve tract after entering the graft and innervated appropriate targets. The innervation of appropriate targets in AP shoulder reversals shows that axons are sensitive to AP mismatch distal to the plexus. Since axons were displaced similar distances from their normal routes in flank DV and AP reversals, the difference in behaviour suggests that they respond to mismatch in the AP but not the DV axis.

Animals↗
PubMed1985

Maps of strength of positional signalling activity in the developing chick wing bud.

Tissue from the posterior margin of the developing limb bud, when grafted to the anterior margin, evokes the formation of a mirror-image limb duplication from the host tissue. We present maps of the spatial and temporal distribution of this signalling activity in the chick wing bud based on a bioassay that provides a quantitative measure of the completeness of the additional structures (the strength of activity index). Activity is first detected prior to the initial appearance of the limb primordium as early as Hamburger & Hamilton stage 14. It reaches a maximum during early outgrowth of the bud at stages 19 to 25. It then declines as the limb starts to differentiate into its final morphological pattern. The design of the experiment provides serendipitous data showing that two operators can consistently perform grafts with high reproducibility between them while variability between embryos is somewhat higher. The maps of activity are of particular practical value in precisely defining for the experimental embryologist and molecular biologist those positions and stages at which peak signalling activity resides.

Animals↗
PubMed1984

Mathematical modelling of the growth processes in the developing chick wing bud.

This paper illustrates how a simple geometric model resembling the shape of the chick wing bud at an early growth stage can be mathematically expanded to simulate subsequent growth characteristics of the developing bud. The model was tested against several sets of experimental data and gave an acceptable representation of growth over the range considered. Representing growth patterns in this form enables the determination of differential growth characteristics in different parts of the bud and provides boundary constraints which will play an important part in the eventual evaluation of internal growth mechanisms.

Animals↗
PubMed1983

The effect of local application of retinoic acid to the anterior margin of the developing chick limb.

Local application of retinoic acid to the chick limb bud produces effects that are dose and/or stage dependent. Low doses and/or old stages tend to give normal limbs or perhaps one or two supernumerary digits of a more anterior character. Medium doses and/or intermediate stages tend to give full mirror-image supernumeraries with two or even three extra digits including particularly digits of a posterior character. High doses and/or early stages give limbs in which supernumerary digits fail to form or are lost, and in which even host skeletal elements are missing or reduced. The effects are graded over the full dose and/or stage range. Various explanations are discussed in the context of the current hypotheses of limb development. We conclude that one should not necessarily interpret the results as evidence that retinoids normally play a part in the control of development or regeneration.

Abnormalities, Drug-Induced↗