Biomedical subjects

J C Glover

Publications and source records attributed to J C Glover.

At least 19 recordsLinked to original sources

PubMed2006

C fragment of tetanus toxin hybrid proteins evaluated for muscle-specific transsynaptic mapping of spinal motor circuitry in the newborn mouse.

We investigated whether the non-toxic C fragment of tetanus toxin (TTC) fused to either beta-galactosidase or green fluorescent protein could be utilized to transsynaptically trace muscle-specific spinal circuitry in the neonatal mouse after i.m. injection into a single hindlimb muscle. We found that even with careful low volume injection (0.2-1.0 microl) into a single muscle (medial gastrocnemius), the TTC hybrid proteins spread rapidly to many other hindlimb muscles and to trunk musculature such that retrograde labeling of motoneurons could not be constrained to a single motoneuron pool. Retrogradely labeled motoneurons in the lower lumbar segments harboring the medial gastrocnemius motoneuron pool were first observed two hours after the medial gastrocnemius injection. Within the next 10 h, additional lumbar and lower thoracic motoneurons became labeled, and punctate labeling in the neuropil surrounding the motoneurons appeared. Many of the TTC hybrid protein-labeled puncta in the neuropil co-localized synaptotagmin, indicating that they represent presynaptic axon terminals onto motoneurons. Although this is consistent with retrograde transsynaptic passage, we found no evidence that the TTC hybrid proteins were transported further along premotor axons to label interneuron somata. The i.m. TTC injection procedure described here therefore provides an important tool for the study of presynaptic terminals onto motoneurons. However, additional technical modifications will be required to utilize TTC tracers for transsynaptic mapping of muscle-specific spinal motor circuitry in the neonatal mouse. We provide here a set of criteria for assessing the i.m. delivery of TTC tracers as a basis for future improvements in this technique.

Acetylthiocholine↗

PubMed2001

[New knowledge of brain repair].

BACKGROUND: Injury to the brain and spinal cord has long been considered particularly refractory to recovery because of the distinct lack of regenerative capacity in the central nervous system. In recent years, however, new knowledge has been gained about what limits this capacity, and increasing evidence has indicated that the potential for regrowth and functional reorganization is much greater than previously deduced. MATERIAL AND METHODS: This review presents historical background and a selection of recent studies that illustrate some of the emerging principles and the outlook for future clinical developments. RESULTS AND INTERPRETATION: Two potential restitutive mechanisms have been identified in the adult brain that open new horizons for the clinical promotion of brain repair. These art the presence of proliferative neural stem cells and the capacity for cortical reorganization within and across functional modalities.

Adult↗

PubMed2001

Quantitative assessment of retinoid signaling pathways in the developing eye and retina of the chicken embryo.

Retinoid signaling has been implicated as an important regulator of retinal development and differentiation. We have used state of the art high-pressure liquid chromatography to identify and quantitate biologically active retinoids, immunohistochemistry to localize the retinoic acid synthetic enzyme retinaldehyde dehydrogenase 2 (RALDH2), and nucleic acid assays to quantitate and localize retinoid receptor gene transcripts in the developing eye and retina of the chicken. Our results demonstrate spatial distinctions in retinoid synthesis and signaling that may be related to laminar differentiation in the developing retina. Retinoic acids (RAs) and their precursor retinols (ROHs) are the predominant retinoids in the developing eye. All-trans-RA and all-trans-3,4-didehydro-RA are present in the neuroepithelium in approximately equal amounts from early stages of neurogenesis until shortly before hatching. The retinoid X receptor (RXR) ligand 9-cis-RA is undetectable at all stages; if present, it cannot exceed a small percentage of the total RA content. RAs are not detected in the pigment epithelium. All-trans-ROH is present in the neuroepithelium and pigment epithelium, whereas all-trans-3,4-didehydro-ROH is detected only in the pigment epithelium and/or the choroid and sclera. RALDH2 immunoreactivity is intense in the choroid, low or absent in the pigment epithelium, and moderate in the neuroepithelium, where it is highest in the outer layers. Transcripts of all five chicken retinoid receptor genes are present in the neural retina and eye throughout development. During the period of neurogenesis, at least three of the receptors (RAR gamma, RXR gamma, RXRalpha), exhibit dynamic patterns of differential localization within the depths of the neural retina.

Aldehyde Oxidoreductases↗

PubMed2001

Quantitative axial profiles of retinoic acid in the embryonic mouse spinal cord: 9-cis retinoic acid only detected after all-trans-retinoic acid levels are super-elevated experimentally.

Studies using bioassays in normal mice and gene activation in transgenic reporter mice have demonstrated peaks of retinoic acid receptor (RAR) signaling in the brachial and lumbar regions of the spinal cord. Recently, Solomin et al. (Solomin et al. [1998] Nature 395:398-402) detected a retinoid X receptor (RXR) signal in the same region of the developing spinal cord at a slightly later stage than the RAR signal. This finding raises the question of which retinoid ligands underlie RAR and RXR signaling in this part of the embryo. Quantitative measurements of regional differences in retinoid profiles have not been reported previously due to limitation in the sensitivity and specificity of available retinoid detection methods. Here, by using a recently developed ultrasensitive HPLC technique (Sakhi et al. [1998] J. Chromatogr. A 828:451-460), we address this question in an attempt to identify definitively the endogenous retinoids present in different regions of the spinal cord at the stages when regional differences in RAR and RXR signaling have been reported. We find a bimodal distribution of all-trans retinoic acid (at-RA), the ligand for RARs, and relate this to the expression of several retinoid-synthesizing enzymes. However, we do not detect 9-cis-retinoic acid (9-cis-RA), the putative RXR ligand, in any region of the spinal cord unless retinoid levels are massively increased experimentally by gavage feeding pregnant mice with teratogenic doses of at-RA. This study provides for the first time quantitative profiles of endogenous retinoids along the axis of the developing spinal cord, thereby establishing a foundation for more definitive studies of retinoid function in the future. It sets definite limits on how much 9-cis-RA potentially is present and demonstrates that at-RA predominates over 9-cis-RA by at least 30- to 180-fold in different spinal cord regions.

Aldehyde Oxidoreductases↗

PubMed2001

Correlated patterns of neuron differentiation and Hox gene expression in the hindbrain: a comparative analysis.

Hindbrain neurons are organized into coherent subpopulations with characteristic projection patterns and functions. Many of these serve vital functions that have been conserved throughout the vertebrate radiation, but diversification to modified or highly specialized functions has also occurred. The differentiation of identifiable neuron groups in specific spatial domains must involve the regional expression of determinants within the hindbrain neuroepithelium. The Hox genes are involved in longitudinal regionalization of the neural tube, and their expression patterns in the hindbrain are closely related to the rhombomeres which partition the hindbrain into morphogenetic units. Hox gene expression also exhibits conserved patterning as well as phylogenetic variation. One plausible mechanism that may have contributed to evolutionary diversification in hindbrain neuron populations is therefore the emergence of species-specific differences in Hox gene expression. This article presents a comparative overview of the regional patterning of selected Hox genes and hindbrain neuron populations in several embryologically important species. Although tantalizing correlations exist, the relationship between Hox genes and neuronal patterning is complex, and complicated by dynamic features in each. Much more comparative and developmental data must be obtained before the link between Hox gene expression and hindbrain neuron patterning can be elucidated satisfactorily in an evolutionary context.

Animals↗

PubMed2000

RXR gamma gene is expressed by discrete cell columns within the alar plate of the brainstem of the chicken embryo.

With in situ hybridization assays, we mapped the distribution of retinoid X receptor gamma (RXR gamma) gene transcripts in the central nervous system of the chicken embryo. Previous studies have demonstrated the presence of RXR gamma transcripts in migrating neural crest and in neural crest derivatives throughout the peripheral nervous system, implicating RXR gamma as an early pan-neural crest marker (Rowe et al. 1991. Development 111:771-778), and in the retina (Hoover et al. 1998. J Comp Neurol 391:204-213). Here we report the presence of RXR gamma transcripts in discrete regions of the developing neural tube, within the hindbrain, the cerebellar plate, the optic tectum, and the diencephalon. At stage 10, when migrating neural crest expresses RXR gamma transcripts, we detect no transcripts in the neural tube. By stage 13, RXR gamma transcripts accumulate to detectable levels along the midline of the posterior optic tectum, where the neural crest-derived sensory neurons of the mesencephalic trigeminal nucleus are located. By stage 15, RXR gamma transcripts also appear in an intermittent longitudinal cell column within the mantle zone of the alar plate of the hindbrain, eventually extending into the cerebellar plate rostrally and into the cervical spinal cord caudally, with a gap at about rhombomere 3. By stage 19, transcripts appear in a discrete population of cells within the diencephalon. Expression in these cell populations continues until at least stage 22.5, when many neuron populations have been generated in the hindbrain. The localization of the RXR gamma-positive cells to the mantle zone suggests that they are postmitotic and are probably neurons. Their specific alar locations indicate that they reside within sensory columns and potential downstream targets, evidently corresponding to some of the central components of the trigeminal system.

Animals↗

PubMed2000

Neuroepithelial 'compartments' and the specification of vestibular projections.

The implication that there exist coherent vestibulo-ocular neuron pools with specific functions may provide new insight into how conjugate eye movements are synthesized within the vestibulo-ocular reflex. The systematic relationship between pool position and synergistic principle terminations, the 'hodological mosaic' suggests, moreover, a determinate groundplan established by developmental mechanisms operative at early stages in the hindbrain neuroepithelium. From such a groundplan, evolutionary and use-dependent modifications could mold connectivity patterns functionally appropriate for each species and individual. How the expression of developmentally regulatory genes contributes to establishing the mosaic organization of the vestibular system is the current focus of our research.

Animals↗

PubMed2000

Development of specific connectivity between premotor neurons and motoneurons in the brain stem and spinal cord.

Astounding progress has been made during the past decade in understanding the general principles governing the development of the nervous system. An area of prime physiological interest that is being elucidated is how the neural circuitry that governs movement is established. The concerted application of molecular biological, anatomical, and electrophysiological techniques to this problem is yielding gratifying insight into how motoneuron, interneuron, and sensory neuron identities are determined, how these different neuron types establish specific axonal projections, and how they recognize and synapse upon each other in patterns that enable the nervous system to exercise precise control over skeletal musculature. This review is an attempt to convey to the physiologist some of the exciting discoveries that have been made, within a context that is intended to link molecular mechanism to behavioral realization. The focus is restricted to the development of monosynaptic connections onto skeletal motoneurons. Principal topics include the inductive mechanisms that pattern the placement and differentiation of motoneurons, Ia sensory afferents, and premotor interneurons; the molecular guidance mechanisms that pattern the projection of premotor axons in the brain stem and spinal cord; and the precision with which initial synaptic connections onto motoneurons are established, with emphasis on the relative roles played by cellular recognition versus electrical activity. It is hoped that this review will provide a guide to understanding both the existing literature and the advances that await this rapidly developing topic.

Afferent Pathways↗

PubMed1999

Regional patterning of reticulospinal and vestibulospinal neurons in the hindbrain of mouse and rat embryos.

The dispositions and axonal trajectories of bulbospinal neurons in the pons and medulla of mouse and rat embryos is described from the earliest times these projections can be labelled retrogradely from the cervical spinal cord. Reticulospinal and vestibulospinal neurons are clustered into identifiable groups, each with a characteristic combination of spatial domain and axon trajectory. The various groups can be labelled retrogradely in a specific developmental sequence. The position of some groups shifts from medial to lateral with development, apparently through cell migration. These observations show that the basic regional organization of the reticulospinal and vestibulospinal projections is similar in mouse and rat and is already established during early stages of axon outgrowth.

Animals↗

PubMed1998

The relationship between rhombomeres and vestibular neuron populations as assessed in quail-chicken chimeras.

The aim of this study was to evaluate the role segmentation plays in the determination of neuronal identity in the hindbrain. We focused on two specific sets of hindbrain neurons, namely, the vestibulospinal and vestibulo-ocular neurons, which comprise distinct groups that can be identified and distinguished by virtue of their axonal projection pathways. The relationship between rhombomeres and the vestibular neuron groups was assessed by a combination of quail-chicken chimeric grafting and selective retrograde axonal tracing. Individual quail hemirhombomeres were transplanted homotopically and isochronically into a chicken embryo host. Subsequently, vestibulospinal and vestibulo-ocular neurons with specific axon trajectories were labeled retrogradely with biotin-conjugated dextran-amines. The relationship between the spatial domains of the vestibular neuron groups and rhombomere-derived domains had the following features: (1) some groups were derived from single rhombomeres; (2) some groups were derived from multiple contiguous rhombomeres; (3) two groups occupied domains that could not be defined in terms of whole rhombomere lengths; (4) some groups spanning multiple rhombomeres exhibited an internal cytoarchitectonic organization that related to individual rhombomeres; and (5) some groups exhibited limited boundary violation. These results support the notion that positional information within defined domains of the neural tube provides a groundplan for the regional determination of neuronal identity and axon pathfinding, and that hindbrain segmentation contributes to this process. But they also indicate that segmentation is not the only mechanism that defines the rostrocaudal domains of neuron types. Moreover, they emphasize that the relationship between rhombomeres and neuronal determination cannot be couched simply in terms of segmental iteration or of bimeric (paired rule) specification.

Animals↗

PubMed1998

Regional pattern of retinoid X receptor-alpha gene expression in the central nervous system of the chicken embryo and its up-regulation by exposure to 9-cis retinoic acid.

We have investigated the expression of the retinoid X receptor-alpha (RXRalpha) gene in the developing chicken embryo by using nonradioactive wholemount in situ hybridization. At the earliest stage of development examined (stage 9; Hamburger and Hamilton [1951] J. Morphol. 88:49-92), we detect RXRalpha transcripts in a stretch of neuroepithelium corresponding roughly to the presumptive caudal hindbrain. Upon formation of the rhombomeres at stage 12, a strongly RXRalpha-positive region extends from a sharp rostral limit at the boundary between rhombomeres 6 and 7 caudad to at least the level of somite 9. This pattern of highest expression continues at least until stage 22 but with some variability in the caudal extent. A lower level of expression extends throughout the spinal cord. Transverse sections show that RXRalpha transcripts are expressed in a gradient, with the highest levels near the roof plate and decreasing toward the floor plate. At later stages, the level of expression is highest in the proliferative ventricular zone. However, at reduced levels, RXRalpha transcripts are also detectable in the mantle zone as well as outside the developing central nervous system, for example, in the neural crest and the limb buds. Nine-cis-retinoic acid up-regulates RXRalpha transcripts at stages 19.5-22.0 within a few hours, augmenting but not expanding the expression pattern. Northern blots demonstrate the potential expression of multiple RXRalpha isoforms in the central nervous system at posthatch stages. These results implicate the RXRalpha receptor in both rostrocaudal and transverse patterning of the neural tube.

Alitretinoin↗

PubMed1998

Segment-specific pattern of sympathetic preganglionic projections in the chicken embryo spinal cord is altered by retinoids.

Sympathetic preganglionic neurons exhibit segment-specific projections. Preganglionic neurons located in rostral spinal segments project rostrally within the sympathetic chain, those located in caudal spinal segments project caudally, and those in midthoracic segments project either rostrally or caudally in segmentally graded proportions. Moreover, rostrally and caudally projecting preganglionic neurons are skewed toward the rostral and caudal regions, respectively, of each midthoracic segment. The mechanisms that establish these segment-specific projections are unknown. Here we show that experimental manipulation of retinoid signaling in the chicken embryo alters the segment-specific pattern of sympathetic preganglionic projections and that this effect is mediated by the somitic mesoderm. Application of exogenous retinoic acid to a single rostral thoracic somite decreases the number of rostrally projecting preganglionic neurons at that level. Conversely, disrupting endogenous synthesis of retinoic acid in a single caudal thoracic somite increases the number of rostrally projecting preganglionic neurons at that level. The number of caudally projecting neurons does not change in either case, indicating that the effect is specific for rostrally projecting preganglionic neurons. These results indicate that the sizes of the rostrally and caudally projecting populations may be independently regulated by different factors. Opposing gradients of such factors along the longitudinal axis of the thoracic region of the embryo could be sufficient, in combination, to determine the segment-specific identity of preganglionic projections.

Animals↗

PubMed1998

Retinoid X receptor gamma gene transcripts are expressed by a subset of early generated retinal cells and eventually restricted to photoreceptors.

We have examined the distribution of the retinoid X receptor gamma (RXRgamma) in the developing chicken retina by using in situ hybridization and RNase protection assays. We detected RXRgamma transcripts as early as 4 days of embryonic development (d4) in central regions of the retina, spreading to more peripheral regions by d8. The first few RXRgamma-positive cells were scattered within the depth of the retinal neuroepithelium, but as they increased in number they became localized predominantly to the apical (outer, ventricular) layer. The identity of the RXRgamma-positive cells at these stages is unknown, due to the lack of cell type-specific markers. By d10, when photoreceptors and ganglion cells have been generated and begun to establish their definitive layers, RXRgamma-positive cells were virtually restricted to the photoreceptor layer, and maintained this distribution to posthatch stages. RNase protection assays were performed on whole retinae to verify the temporal pattern of in situ hybridization results and showed that between d5 and d16 there was a significant increase in the mRNA levels of the RXRgamma2 isoform. Between d16 and early posthatch stages the level of RXRgamma2 mRNA did not change significantly. Consistent with previous studies, mRNA levels of the RXRgamma1 isoform were substantially lower than mRNA levels of the RXRgamma2 isoform at all time points examined. These results demonstrate that RXRgamma mRNA is expressed in photoreceptors in the developing chicken retina and implicate RXRgamma as the earliest marker of photoreceptor differentiation documented to date.

Animals↗

PubMed1997

Developmental dynamics of functionally specific primary sensory afferent projections in the chicken embryo.

The central projections of specific subpopulations of lumbar primary afferents were selectively labeled with the lipophilic tracer DiI in fixed preparations of the chicken embryo. Muscle or cutaneous afferents were selectively labeled by applying DiI to identified peripheral nerves. Medial or lateral afferent populations were selectively labeled by partially lesioning the dorsal root. Muscle and cutaneous afferent populations each contribute to both the medial and the lateral afferent populations. Medial muscle afferents terminate in the intermediate zone and lateral motor column proximally, but only in the intermediate zone distally. Lateral muscle afferents terminate in a ventrolateral region of the dorsal horn both proximally and distally. Medial cutaneous afferents terminate predominantly in lamina III, but a few terminate in the medial region of the intermediate zone. Lateral cutaneous afferents terminate in lamina II and in a ventrolateral region of the dorsal horn. On the basis of the principle termination patterns, specific termination fields were defined and related to the classical cytoarchitectonics of the spinal gray matter. Differential retrograde tracing from the spinal cord with fluorescent dextran-amines demonstrated that the medial afferents originate from the earlier-generated ventrolateral population of large sensory cell bodies, while the lateral afferents originate from the later-generated dorsomedial population of small sensory cell bodies. The medial afferents establish their central projections earlier than the lateral afferents, but for each subpopulation the initial pattern of termination prefigures the mature pattern, throughout the segmental range of the collaterals. Birthdating with 3H-thymidine showed that potential target neurons in the different terminal fields within the dorsal horn are born at different times. In particular, interneurons in lamina II are born after those in lamina III, paralleling the early and late termination of cutaneous afferents in these laminae. Our observations support the notion that primary afferents recognize specific cues in the spinal cord, but also implicate the relative timing of afferent and target differentiation as an important determinant of primary afferent termination patterns.

Afferent Pathways↗

PubMed1996

Evidence that intersegmental competition influences the segmental distribution of the central terminals of muscle sensory afferents in the chicken embryo.

The possibility that muscle sensory afferents that enter the spinal cord at different segmental levels compete for terminal space in the lateral motor column was tested by unilaterally ablating neural crest cells destined to generate dorsal root ganglia (DRGs). Newly migrating neural crest was surgically removed from several lumbosacral segments on one side of the spinal cord on day 3 of embryogenesis (d3). The resultant experimental preparations developed without DRGs on the operated side at the segmental levels where the ablation was performed. At d18 the central projection from the intact DRG immediately caudal to the operated segments was anatomically mapped with a lipophilic axonal tracer. The central projection from the same DRG on the opposite, unoperated side was mapped as a control. The density of terminals along the longitudinal axis of the lateral motor column was quantified in serial sections with an image analysis program. On both the operated and control sides, the majority of terminals were found within the segment of entry and the rostral and caudal neighboring spinal segments. However, the rostral neighboring segment received a larger proportion of terminals on the operated side compared to the control side. Thus, the absence of primary afferents entering the spinal cord at specific segments leads to a shift of the afferent projection to the lateral motor column from the neighboring segments towards the deafferented segments.

Animals↗

PubMed1996

Development of an identified spinal commissural interneuron population in an amniote: neurons of the avian Hofmann nuclei.

The commissural interneurons of the Hofmann nuclei (HN) of the avian spinal cord (The axonal projections of the Hofmann nuclei in the spinal cord of the late stage chicken embryo, Anat Embryol (Berl), A.L. Eide, 1996, Vol 193, pp 543-557) provide a unique opportunity to describe the development of an identified spinal commissural axon projection and its terminal collaterals in an amniote vertebrate. Here, we use the lipophilic tracer Dil to label these and other commissural projections anterogradely and retrogradely from the time the HN neurons are born. [3H]thymidine birthdating shows that the final mitoses of HN neurons occur at stages 21-24 [developmental day (d) 4]. By direct comparison, this follows the generation of motoneurons and of large, dorsally located commissural interneurons. The first HN neurons reach the ventrolateral margin of the spinal cord by d6 by a radial migration through the ventral horn. Radial migration occurs after the extension of HN axons across the midline. Thus, HN neurons are determined to be commissural interneurons before attaining their definitive locations. The HN neurons subsequently aggregate into segmentally iterated clusters at the ventrolateral margin of the spinal cord by d8. Also by d8 their logitudinal axons attain mature extent in the ventral funiculus of the contralateral side and begin to sprout collaterals. The collaterals are directed predominantly toward the medial aspect of the ventral horn at all stages, forming by d12 a dense thicket of terminals that thins out over several segments to each side of the HN of origin. The initial direction of collateral outgrowth is largely appropriate for the mature termination pattern of the HN. Terminal arbors, however, are less focused at early developmental stages than at later stages.

Animals↗

PubMed1996

Development of second-order vestibular projections in the chicken embryo.

The results reviewed here demonstrate that vestibulospinal and vestibulo-ocular interneurons in the chicken embryo are organized into coherent clusters, each of which occupies a unique spatial domain and has a characteristic axon trajectory. Moreover, the vestibulo-ocular clusters have specific termination patterns in the extraocular motoneuron pools and may to some extent have specific functions. The axon trajectories and termination patterns are established accurately during early development. This suggests the presence of selective mechanisms that ensure the generation of an appropriately specific connectivity pattern. The correlation of cluster domains with unique fields of regulatory gene expression at early stages suggests a mechanistic link between the position of a vestibular interneuron and the differentiation of its axon trajectory, termination pattern, and neurotransmitter phenotype.

Animals↗

PubMed1996

Appropriate pattern formation following regulative regeneration in the hindbrain neural tube.

Patterns of neuronal differentiation and axon outgrowth were examined in regenerated neural tissue of the hindbrain of the chicken embryo. Specific regions of the hindbrain neural tube corresponding to identified rhombomeres were selectively removed unilaterally by microsurgery. Of the embryos that survived, about 35% exhibited regulative regeneration, wherein the missing hemi-rhombomere was reconstituted to normal size and morphology through compensatory proliferation and migration from adjacent tissue. The capacity for regeneration depended on the embryonic stage at which the ablation was performed and on whether the midline floor plate was included in the ablation. Ablations performed prior to the formation of morphologically overt rhombomere boundaries led to significantly lower frequencies of regulative regeneration than ablations performed after the formation of boundaries. Ablations that included the floor plate led to significantly lower frequencies of regulative regeneration than ablations sparing the floor plate. Regulative regeneration was also observed at low frequency following excision of an entire rhombomere. Within the regenerated hemi-rhombomere, identified neuron groups appeared in their normal positions and differentiated normal patterns of migration and axon outgrowth. The fidelity of this patterning, however, depended on the mediolateral position of the neuron group, being more reliable for neuron groups in the basal plate than in the alar plate. These results confirm the extensive regulative potential of the neural tube and demonstrate the capacity for appropriately patterned cellular differentiation within the regenerated tissue.

Animals↗