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T A Reh

Publications and source records attributed to T A Reh.

At least 55 records · Page 3Linked to original sources
PubMed1994

Retinoic acid promotes differentiation of photoreceptors in vitro.

The results of several recent studies have demonstrated that cell commitment and differentiation in the developing vertebrate retina are influenced by cell-cell interactions within the microenvironment. Retinoic acid has been shown to influence cell fates during development of the nervous system, and retinoic acid has been detected in the embryonic retina. To determine whether retinoic acid mediates the differentiation of specific neuronal phenotypes during retinal histogenesis, we treated dissociated cell cultures of embryonic and neonatal rat retina with varying concentrations of all-trans or 9-cis retinoic acid and analyzed the effects on cell fate using neuron and photoreceptor-specific antibodies. Addition of exogenous retinoic acid caused a dose-dependent, specific increase in the number of cells that developed as photoreceptors in culture throughout the period of retinal neurogenesis. In the same cultures, retinoic acid also caused a dose-dependent decrease in the number of cells that developed as amacrine cells. Also, results of double-labeled immunohistochemical studies using bromodeoxyuridine demonstrated that the primary effect of retinoic acid was to influence progenitor cells to develop as newly generated rod photoreceptors. Since retinoic acid and at least one of the retinoic acid receptors (RAR alpha) have been localized to the developing neural retina, these results suggest that retinoic acid may play a role in the normal development of photoreceptor cells in vivo.

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PubMed1994

Intrinsic and extrinsic signals in the developing vertebrate and fly eyes: viewing vertebrate and invertebrate eyes in the same light.

The development of complex tissues, such as the nervous system, requires coordinated interactions between cells, and defining the molecular basis of these interactions is a primary goal of developmental neurobiology. Classic experimental embryological studies of the developmental potential of embryonic neural tissues have more recently been augmented by genetic and molecular approaches to yield a picture of the mechanisms by which such complex neural structures arise during development. Perhaps coincidentally, two of the most well-studied structures in this regard have been the eyes of Drosophila and vertebrates. In the past few years, a number of laboratories have focused on the development of the eye, of both Drosophila and vertebrates in order to understand the role of cell-cell interactions in the generation of organized neural structures. Recent evidence from both the vertebrate and Drosophila models suggests that common mechanisms apply in both systems, particularly in the role of tyrosine kinases in regulating phenotypic choices during histogenesis. Thus, although the eyes of flies and vertebrates have very different appearances, we examine the possibility that similar developmental processes are used to produce tissues with analogous functions.

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PubMed1994

Regional differences in glial-derived factors that promote dendritic outgrowth from mouse cortical neurons in vitro.

To determine whether glia from different CNS regions differ in their ability to support axons or dendrites, embryonic (E18) mouse cortical neurons were cocultured with early postnatal (P4) rat astroglial derived from cortex, retina, olfactory bulb, mesencephalon, striatum, and spinal cord. After 5 d in vitro, axon and dendrite outgrowth from isolated neurons was quantified with double-labeling immunohistochemical techniques. Whereas axonal growth was similar on the various monolayers, total primary dendritic outgrowth was nearly threefold greater on glia derived from the cortex, retina, and olfactory bulb than on glia derived from mesencephalon, striatum, or spinal cord. This effect was principally on the number of primary dendrites rather than the elongation of individual dendrites. Similar morphological differences were observed when cortical neurons were grown on polylysine in a noncontact coculture system with glia continuously conditioning the media. This selective promotion of dendrite growth was independent of neuron survival. These results indicate that there are regional differences in the ability of CNS glia to support dendritic growth and that this effect is due, in part, to release of a diffusable factor.

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PubMed1994

Cell adhesion molecules regulating neurite growth from amacrine and rod photoreceptor cells.

A great deal is now known about the cell adhesion molecules (CAMs) that are responsible for promoting the growth of ganglion cell axons as they project out of the retina through the optic nerve and finally to distant targets in the brain. However, the CAMs important for regulating axon outgrowth from nonprojection neurons, such as amacrine cells and rods, are not known. Such local circuit neurons extend their neurites rather short distances on cellular surfaces not normally encountered by the ganglion cell axons. To study the mechanisms regulating axon or dendrite growth from local circuit neurons, neurite outgrowth from amacrine cells and rod photoreceptor cells derived from the rat was examined in vitro on immunopurified forms of NCAM, L1, and N-cadherin, three well-characterized adhesive molecules found in the developing retina. Either early (P3) or late (P10) postnatal amacrine cells grew neurites on all three CAMs, but there were significant differences in the percentage of the amacrine cells that responded to each CAM. None of the CAMs supported neurite outgrowth from early postnatal rods, but, surprisingly, NCAM stimulated vigorous neurite extension from rods isolated at postnatal day 10. Postnatal ganglion cells were also examined for comparison and were found not to grow neurites on NCAM, but did grow extensive processes on L1 and N-cadherin. These results show that NCAM, L1, and N-cadherin can promote neurite outgrowth from local circuit neurons, but that the effectiveness of any particular CAM is dependent on the cell type and the developmental period.

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PubMed1993

Developmental study of the expression of B50/GAP-43 in rat retina.

B50/GAP-43 has been implicated in neural plasticity, development, and regeneration. Several studies of axonally transported proteins in the optic nerve have shown that this protein is synthesized by developing and regenerating retinal ganglion cells in mammals, amphibians, and fish. However, previous studies using immunohistochemistry to localize B50/GAP-43 in retina have shown that this protein is found in the inner plexiform layer in adults. Since the inner plexiform layer contains the processes of amacrine cells, ganglion cells, and bipolar cells to determine which cells in the retina express B50/GAP-43, we have now used in situ hybridization to localize the mRNA that codes for this protein in the developing rat retina. We have found that B50/GAP-43 is expressed primarily by cells in the retinal ganglion cell layer as early as embryonic day 15, and until 3 weeks postnatal. Some cells in the inner nuclear layer, possibly a subclass of amacrine cells, also express B50/GAP-43 protein and mRNA; however, the other retinal neurons-bipolar cells, photoreceptors, and horizontal cells express little, if any, B50/GAP-43 at any stage in their development. Early in development, the protein appears in the somata and axons of ganglion cells, while later in development, B50/GAP-43 becomes concentrated in the inner plexiform layer, where it continues to be expressed in adult animals. These results are discussed in terms of previous proposals as to the functions of this molecule.

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PubMed1993

Hair-cell regeneration in organ cultures of the postnatal chicken inner ear.

The sensory epithelium of the avian inner ear retains into adulthood progenitor cells for inner-ear hair cells and other cell types in the epithelium. Hair cells are produced normally on an ongoing basis in the vestibular sensory epithelium, and hair-cell production is increased after insult in both auditory and vestibular sensory epithelia. The details of postnatal hair-cell production are not understood. In particular, molecular factors involved in the initiation and regulation of hair-cell genesis and differentiation are not known. Studies of this phenomena have been hampered by the lack of cell culture models. An organ culture system was developed which encourages generation and differentiation of hair cells in mature inner-ear sensory epithelia. Continuous labeling with tritiated thymidine showed genesis of both supporting cells and hair cells in normal vestibular epithelia grown in culture, and an increase in hair-cell and supporting-cell proliferation in damaged sensory epithelia grown in culture as compared to undamaged controls. This demonstrates, in vitro, both the division and differentiation of hair-cell progenitor cells in normal vestibular epithelia, and the maintenance of the hair-cell regeneration process in damaged inner-ear epithelia. This culture system should be useful for studies of hair-cell genesis and differentiation as well as studies of hair-cell and supporting-cell functioning in general.

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PubMed1992

Cellular interactions determine neuronal phenotypes in rodent retinal cultures.

Progenitor cells isolated from early rat embryo retinas differentiate into phenotypes normally generated early in retinal development (e.g., ganglion cells), whereas progenitors isolated from postnatal retinas differentiate into later-generated retinal cell types (e.g., rod photoreceptors; Reh and Kljavin, J. Neurosci. 9:4179-4189; 1989; Adler and Hatlee, 1989; Science 243:391-393; Sparrow, Hicks, and Barnstable, 1990, Dev. Brain Res. 51:69-84). To determine whether this change in committment is intrinsic to the progenitor cells, or alternatively can be modified by interactions with their developing environment, I co-cultured mouse and rat retinal cells, from different developmental stages, and identified the resulting phenotypes with species-specific and cell class-specific antibodies. I found that the phenotypes into which mouse neuroepithelial cells differentiate depends on the phenotypes of the rat cells that surround them. Retinal precursor cells from embryonic day (E) 10-12 will adopt the rod photoreceptor phenotype only when close to cells expressing this phenotype. By contrast, when the E10-12 retinal progenitor cells are cultured with cells from the cerebral cortex, they differentiate primarily into large multipolar neurons, similar in their morphology and antigen expression to retinal ganglion cells. These results indicate that interactions among the cells of the developing retina are important in the determination of cell fate.

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PubMed1991

Common mechanisms of retinal regeneration in the larval frog and embryonic chick.

Amphibians and embryonic chicks possess the ability to regenerate retinal neurons by the transdifferentiation of pigment epithelium into neuronal progenitors. Recent studies have begun to identify the molecular factors involved in this process. Laminin (a component of the extracellular matrix) has been shown to be important in the process of retinal regeneration in the larval frog both in vitro and in vivo and basic fibroblast growth factor (bFGF) stimulates the same process in chicks in vivo. To determine the mechanisms by which these factors induce retinal regeneration we studied their effects on cultured chick pigment epithelium cells. bFGF was added to enzymically dissociated chick embryo pigment epithelial cells plated at several different densities on various substrates including laminin. We found that bFGF stimulated proliferation but although the cells lost pigmentation and demonstrated distinct morphological changes, no definitive transdifferentiation could be demonstrated using several neuron-specific antibodies as markers. When the pigment epithelium was cultured as aggregates on a shaker device which prevented flattening and spreading on the substrate a large number of retinal progenitor cells were generated from the pigment epithelium treated with bFGF. The ability of chick pigment epithelium to undergo transdifferentiation thus appears to be dependent on the physical configuration of the cells.

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PubMed1991

Optic nerve tissue sections derived from myelin-deficient mutant mice as culture substrata for fibroblast adhesion and spreading.

The substrate properties were compared between normal and myelin-deficient central nervous system (CNS) tissues by an in vitro assay of cell attachment and spreading. Fibroblasts (3T3) were plated onto culture substrata consisting of optic nerve tissue sections cut from normal or two myelin-deficient mutant mice, Shiverer and Quaking. Optic nerve sections from either of the mutant animals supported more 3T3 fibroblast spreading and adhesion than sections derived from animals with normal myelin. These results demonstrate that CNS myelin influences the ability of cells to attach and spread and that it is the actual presence of myelin which is inhibitory rather than the presence of optic nerve axons or oligodendrocytes.

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PubMed1991

EGF and TGF-alpha stimulate retinal neuroepithelial cell proliferation in vitro.

Peptide growth factors have been shown to have diverse effects on cells of the CNS, such as promoting neuronal survival, neurite outgrowth, and several other aspects of neuronal differentiation. In addition, some of these factors have been shown to be mitogenic for particular classes of glial cells within the brain and optic nerve, and recently two peptide growth factors, fibroblast growth factor and nerve growth factor, have been shown to have mitogenic activity on the CNS neuronal progenitors. We now report that two members of another peptide growth factor, epidermal growth factor and transforming growth factor-alpha, are mitogenic for retinal neuroepithelial cells in primary cultures and provide evidence for the presence of both of these factors in normal developing rat retina.

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PubMed1991

Basic fibroblast growth factor induces retinal pigment epithelium to generate neural retina in vitro.

During embryogenesis, the cells of the eye primordium are initially capable of giving rise to either neural retina or pigmented epithelium (PE), but become restricted to one of these potential cell fates. However, following surgical removal of the retina in embryonic chicks and larval amphibians, new neural retina is generated by the transdifferentiation, or phenotypic switching, of PE cells into neuronal progenitors. A recent study has shown that basic fibroblast growth factor (bFGF) stimulates this process in chicks in vivo. To characterize further the mechanisms by which this factor regulates the phenotype of retinal tissues, we added bFGF to enzymatically dissociated chick embryo PE. We found that bFGF stimulated proliferation and caused several morphological changes in the PE, including the loss of pigmentation; however, no transdifferentiation to neuronal phenotypes was observed. By contrast, when small sheets of PE were cultured as aggregates on a shaker device, preventing flattening and spreading on the substratum, we found that a large number of retinal progenitor cells were generated from the PE treated with bFGF. These results indicate that bFGF promotes retinal regeneration in vitro, as well as in ovo, and suggest that the ability of chick PE to undergo transdifferentiation to neuronal progenitors appears to be dependent on the physical configuration of the cells.

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PubMed1991

Müller cells are a preferred substrate for in vitro neurite extension by rod photoreceptor cells.

To define the factors important in photoreceptor cell morphogenesis, we have examined the ability of rods to extend neurites in vitro. Retinas from neonatal rats were dissociated and plated onto substrate-bound extracellular matrix (ECM) components or cell monolayers. When rods, identified with monoclonal antibodies to opsin, were in contact exclusively with purified ECM (e.g., laminin, fibronectin, type I collagen, or Matrigel), neurite outgrowth was extremely limited. By contrast, rods extended long neurites on Müller cells. Retinal or brain astrocytes, endothelial cells, 3T3 fibroblasts, or other retinal neurons were less supportive of rod process outgrowth. These data demonstrate regional specificity in the promotion of neurite outgrowth by glia and suggest that not all neurons within the retina require the same morphogenic factors.

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PubMed1990

Induction of differentiation of rat retinal, germinal, neuroepithelial cells by dbcAMP.

Little is known about the factors that regulate the production of neurons during the development of the vertebrate central nervous system (CNS); however, evidence from several neuronal cell lines suggests that an increase in intracellular cAMP might trigger the process of differentiation. To determine if a similar process is involved in differentiation during normal CNS neurogenesis, we raised the intracellular level of cAMP in primary cultures of mitotically active, germinal neuroepithelial cells from fetal and postnatal rat retina. This treatment induced differentiation of the CNS precursors, causing the cells to cease DNA synthesis and increase their expression of proteins normally found in differentiated retinal cells. These results indicate that germinal neuroepithelial cell differentiation can be controlled through the cAMP second messenger system, and that the regulation of this system may in part determine the numbers and ratios of the various classes of neurons during the normal development of the CNS.

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PubMed1989

Age of differentiation determines rat retinal germinal cell phenotype: induction of differentiation by dissociation.

We are interested in the mechanisms that control cell phenotype during the development of the CNS. Since different neuronal types arise at different times during neurogenesis in the retina, we predicted that the factors that determine cell type must be developmentally regulated as well. To test this hypothesis, we induced retinal germinal cells to differentiate at different ages by dissociating the retina into single cells and culturing them on a variety of substrates. Prior to dissociation, the S-phase germinal cells were labeled with 3H-thymidine so that their fate could be specifically followed. We found that our culture conditions promoted the differentiation of the majority of the germinal cells and that these cells differentiated into different neuronal types depending on the age of the animal from which the retina had been taken; embryonic day 14 germinal cells differentiated primarily into ganglion cells, and never produced rods, while germinal cells from postnatal day 1 retina differentiated into rods, but not ganglion cells. These results are consistent with the hypothesis that temporally regulated factors determine cell phenotype during the development of the retina.

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PubMed1988

Immunocytochemical and morphological evidence for a retinopetal projection in anuran amphibians.

The centrifugal projection to the retina in anuran amphibians (Rana catesbeiana and Xenopus laevis) has been investigated by immunocytochemistry, HRP transport, and optic nerve lesionings. FMRFamide- and N-terminal substance P-immunoreactive (Fa-ir and SP(3-7)-ir) fibers were abundant in the normal retina and optic nerve but almost absent distal to an optic nerve section or crush after 7-14 days survival. Fa-ir and SP(3-7)-ir fibers were traced to the optic nerve from the lamina terminalis (or the septopreoptic junctional area), where there are many Fa-ir and SP(3-7)-ir perikarya. After application of HRP to the optic nerve and survival for 9-10 days, retrogradely labeled neurons were observed in the lamina terminalis. Conversely, following HRP injection into the septal and preoptic area, labeled fibers were observed in the optic nerve. These results suggest that Fa-ir and SP(3-7)-ir efferent fibers project from the lamina terminalis to the retina. But in anurans, unlike teleosts, these fibers are not gonadotropin-releasing-hormone (GnRH)-ir. The morphological relations of this retinopetal pathway with the GnRH-ir nervus terminalis are discussed.

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PubMed1988

A role for the extracellular matrix in retinal neurogenesis in vitro.

The germinal neuroepithelial cells that give rise to the majority of neurons in the vertebrate central nervous system are in contact with the basement membrane that surrounds the neural tube from the very earliest stages. The effect of removing this basement membrane on the organization and proliferative potential of these cells was examined in a new slice culture preparation of developing Rana tadpole retina. The results indicate that the germinal neuroepithelium, like other epithelial tissues, requires contact with a basement membrane for the maintenance of its structure and a normal degree of cell proliferation.

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PubMed1987

A possible role for the vascular membrane in retinal regeneration in Rana catesbienna tadpoles.

We have studied the process of retinal regeneration in Rana catesbienna tadpoles using a recently developed monoclonal antibody (2D3) directed against frog neurons and germinitive neuroepithelium. We have found that, following retinal degeneration induced by devascularization, new retina is generated in the posterior eye from transdifferentiating pigment epithelial (RPE) cells and in the anterior eye from increased proliferation at the normal growth zone in the ora serrata. This demonstrates that the anuran retina regenerates in a manner similar to that observed previously in urodeles. In addition, the use of MAb-2D3 has allowed us to study the process of RPE transdifferentiation more accurately than was previously possible, and consequently we have found a high degree of association of migratory pigment cells with the retinal vascular membrane at the time of the initial RPE transdifferentiation to retinal neuroblasts.

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