Demonstration of a somatostatin-like activity in retinal cells of the rat

Summary A somatostatin-like substance is demonstrated by light microscopic immunohistochemistry (PAP-method) in perikarya and cell processes of the retina of adult and infant rats. These perikarya are identified according to their size, arrangement and distribution. Each of the first two neuronal orders (receptors, bipolar cells, ganglionic cells) of the visual pathway can be associated with retinal cells reacting positively with anti-somatostatin. In the adult rat, perikarya and processes of (i) horizontal cells, (ii) amacrine cells and (iii) large neurons in the ganglionic layer are specifically labeled. The staining of middle-sized and small ganglion cells is probably caused by the close attachment of labeled fibers to non-reacting cells. Postnatally, the immunoreactive elements develop in parallel to the differentiation of the corresponding retinal layers. It is discussed whether the three types of retinal cells containing a somatostatin-like substance provide an inhibitory system to each of the two orders of retinal neurons.Supported by the Deutsche Forschungsgemeinschaft (Grant Nr. Kr 569/2) and Stiftung Volkswagenwerk     

Restoration of retino-tectal connections in frogs during regeneration of the optic nerve

At various times after unilateral division of the optic nerve in the frogRana temporaria L. evoked potentials in response to electrical stimulation of the optic nerve were investigated in a segment distal to the site of operation, spike activity was recorded from endings of regenerating and intertectal axons when stimuli of different shapes were placed in the field of vision, and the distribution of axonal bulbs of growth by depth in the tectum mesencephal was studied electron-microscopically. During regeneration of the axons the responses of the retinal ganglionic cells to visual stimuli retained most of their individual features. Myelinated axons of the retinal ganglionic cells regenerate first (starting on the 21st day after operation). Myelination of these fibers lags significantly behind their growth and is complete more than 100 days after the operation. Unmyelinated axons of the retinal ganglionic cells grow up toward the tectum mesencephali after myelinated axons (80 or more days after the operation). Axonal bulbs of growth in the initial periods after the operation are located close to the pial surface and the level of spread of the myelinated axons of the retinal ganglionic cells differs significantly from their normal level of localization. Intertectal connections persist after division of the nerve and are activated by visual stimuli during regeneration of the axons of the retinal ganglionic cells. Connections were found mainly between intertectal fibers terminating superficially and retinal ganglionic cells belonging to class 1 and 2 detectors. Axons of the retinal ganglionic cells grow up toward the caudal region of the tectum mesencephali later than toward the rostral region.A. N. Severtsov Institute of Evolutionary Morphology and Ecology of Animals, Academy of Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 5, No. 6, pp. 611–620, November–December, 1973.     

Neural stem cell lineages are regionally specified, but not committed, within distinct compartments of the developing brain.

Regional patterning in the developing mammalian brain is partially regulated by restricted gene expression patterns within the germinal zone, which is composed of stem cells and their progenitor cell progeny. Whether or not neural stem cells, which are considered at the top of the neural lineage hierarchy, are regionally specified remains unknown. Here we show that the cardinal properties of neural stem cells (self-renewal and multipotentiality) are conserved among embryonic cortex, ganglionic eminence and midbrain/hindbrain, but that these different stem cells express separate molecular markers of regional identity in vitro, even after passaging. Neural stem cell progeny derived from ganglionic eminence but not from other regions are specified to respond to local environmental cues to migrate ventrolaterally, when initially deposited on the germinal layer of ganglionic eminence in organotypic slice cultures. Cues exclusively from the ventral forebrain in a 5 day co-culture paradigm could induce both early onset and late onset marker gene expression of regional identity in neural stem cell colonies derived from both the dorsal and ventral forebrain as well as from the midbrain/hindbrain. Thus, neural stem cells and their progeny are regionally specified in the developing brain, but this regional identity can be altered by local inductive cues.     

Effects of cadmium on photoreceptors and ganglionic cells of retinal layer in mice embryo--an ultrastructural study

Cadmium (Cd) is one of the environmental contaminant and because of its non-decomposable character, it can damage nature. In this study, TEM was used in order to assess the ultrastructural effects of Cd on photorececptor and ganglionic cells of mouse retinal layer. Apoptotic nuclei, heterochromatic nuclei, deletion of nucleus membrane, invisible nucleolus, and apoptotic cells with mitochondrial changes were observed in mice embryo (days 15 of gestation) following CdCl2 injection to mothers on day 9 of gestation. Cadmium exposure caused apoptotic changes both in photoreceptors and ganglionic cells.     

Changes in the cerebellar cortex of rabbits subjected to an electrical current on the posterior hypothalamus

After 3, 15 and 30 sessions of electrical stimulation of the hypothalamus by means of stereotaxically implanted bipolar electrodes into the area hypothalamica posterior, in the cortex of the cerebellar hemispheres, predominantly in the ganglionic layer, certain reactive, as well as poorly and intensively manifested pathological changes develop in neural and glial cells. Mosaicism of Purkinje's cells lesions within the layer, the state of their cytoplasmic matrix, granular endoplasmic reticulum and mitochondria, essential shifts in activity of oxidation-reduction enzymes at a simultaneous disorder in blood circulation and decreased content of catecholamines in the neural structures of the cerebellar vessels make it possible to estimate the process as centrogenic and neurodistrophic.     

Light-optic, electron microscopic and electrophysiological study of centrifugal fibers of the retina in the lamprey Lampetra fluviatilis

Light and electronmicroscopic studies have been made on retinal structures in the lamprey labeled by horseradish peroxidase injected into the peripheral end of the cut optic nerve or to the midbrain tectum. On total retinal preparations, labeled axons were revealed together with dendrites and ganglionic cell bodies, as well as branching (presumably retinopetal) fibers, fine endings of which come closely to the labeled dendrites of the ganglionic cells. Electron microscopic data indicate that the labeled terminations of afferent fibers from synapses with both labeled and unlabeled dendrites, as well as with unlabeled neuronal bodies. It is concluded that centrifugal fibers in lamprey retina form contacts with the bodies and dendrites of the amacrine cells and dendrites of the ganglionic cells. Results of intracellular registration of responses of various retinal elements to the electrical stimulation of the optic nerve support this conclusion.     

Ciliary neurotrophic factor: a survival and differentiation inducer in human retinal progenitors

Retinitis pigmentosa, age-related macular degeneration, and Parkinson’s disease remain major problems in the field of medicine. Some of the strategies being explored for treatment include replacement of damaged tissue by transplantation of healthy tissues or progenitor cells and delivery of neurotrophins to rescue degenerating tissue. One of the neurotrophins with promise is the ciliary neurotrophic factor (CNTF). In this study, we report the role played by CNTF in retinal cell differentiation and survival in retinal progenitors. We found that CNTF is a survival factor for multipotential human retinal cells and increased cell survival by 50%, over a 7-d period, under serum-free conditions, as determined by apoptotic assays (immunohistochemistry and flow cytometry). This effect is dose dependent with a maximum survival at a CNTF concentration of 20 ng/ml. We also report that CNTF might be a cell commitment factor, directing the differentiation mainly toward large multipolar cells with ganglionic and amacrine phenotype. These cells express tyrosine hydroxylase (amacrine cells) as well as, thy 1.1 and neuron-specific enolase (ganglionic cells). Additionally, there was also an increase in protein kinase C alpha, a protein expressed in rod and cone bipolars as well as cone photoreceptors and calbindin, a protein expressed in cone photoreceptors and horizontal cells. In our studies, CNTF doubled the number of cells with ganglionic phenotypes, and basic fibroblast growth factor doubled the number of cells with photoreceptor phenotype. Additionally, CNTF induced a subset of progenitors to undergo multiple rounds of cell division before acquiring the large multipolar ganglionic phenotype. Our conclusion is that CNTF could be an agent that has therapeutic potential and possibly induces differentiation of large multipolar ganglionic phenotype in a subset of progenitors.     

Retinal remodeling in inherited photoreceptor degenerations

Photoreceptor degenerations initiated in rods or the retinal pigmented epithelium usually evoke secondary cone death and sensory deafferentation of the surviving neural retina. In the mature central nervous system, deafferentation evokes atrophy and connective re-patterning. It has been assumed that the neural retina does not remodel, and that it is a passive survivor. Screening of advanced stages of human and rodent retinal degenerations with computational molecular phenotyping has exposed a prolonged period of aggressive negative remodeling in which neurons migrate along aberrant glial columns and seals, restructuring the adult neural retina (1). Many neurons die, but survivors rewire the remnant inner plexiform layer (IPL), forming thousands of novel ectopic microneuromas in the remnant inner nuclear layer (INL). Bipolar and amacrine cells engage in new circuits that are most likely corruptive. Remodeling in human and rodent retinas emerges regardless of the molecular defects that initially trigger retinal degenerations. Although remodeling may constrain therapeutic intervals for molecular, cellular, or bionic rescue, the exposure of intrinsic retinal remodeling by the removal of sensory control in retinal degenerations suggests that neuronal organization in the normal retina may be more plastic than previously believed.     

Light-Induced CREB Phosphorylation and Gene Expression in Rat Retinal Cells

Abstract: The signal pathway for light-induced expression of c- fos and the neuropeptide somatostatin (SS) in rat retinal cells was investigated. Flashing light induced c- fos and SS mRNA in the inner nuclear layer and the ganglion cell layer. As both c- fos and SS genes have a cyclic AMP response element (CRE) in their promoters, CRE-binding protein (CREB) phosphorylation in retinal cells was examined with a phospho-CREB-specific antibody. Both flashing light and administration of the L-type Ca²⁺ channel activator Bay K 8644 induced phosphorylation of CREB in the nuclei of the amacrine cells and the ganglion cells where c- fos /SS mRNAs were expressed. These cells could be double-stained with anti-calmodulin kinase II (anti-CaM kinase II) monoclonal antibody and phospho-CREB-specific polyclonal antiserum after Bay K 8644 administration, indicating the colocalization of phosphorylated CREB at Ser¹³³ and CaM kinase II in the neural retina.     

Neural computations in the tiger salamander and mudpuppy outer retinae and an analysis of GABA action from horizontal cells

 A neural network architecture based on the neural anatomy and function of retinal neurons in tiger salamander and mudpuppy retinae is proposed to study basic aspects of early visual information processing. The model predictions for the main response characteristics of retinal neurons are found to be in agreement with neurophysiological data, including the antagonistic role of horizontal cells in the outer plexiform layer. The examination of possible γ-aminobutyric acid (GABA) action from horizontal cells suggests that GABA_A alone, GABA_B alone, or their weighted combination can generate the response characteristics observed in bipolar cells. Received: 25 June 2002 / Accepted: 28 January 2003 / Published online: 20 May 2003 Acknowledgements. The authors would like to thank an anonymous reviewer for valuable comments. Correspondence to: S. X. Yang (e-mail: syang@uoguelph.ca)     

Differentiation of the interstitial cell line in hydrozoan planulae

Repopulation of epithelial (colchicine-treated) planular tissue by interstitial cells, nematoblasts/nematocytes, and ganglionic cells was examined via grafting. Seventy-two-hour epithelial planular head pieces were grafted to 72-hour control labelled planular tail pieces, left in contact for 24 h, separated, and the head pieces were analyzed for interstitial cells and their derivatives. The reciprocal experiment of grafting 72-hour epithelial planular tails to 72-hour control labelled planular heads was also done and the tail pieces were examined. Repopulated planular head pieces contained interstitial cells, ganglionic cells and a reforming neural plexus but few nematoblasts/nematocytes. Reconstituted planular tail pieces contained interstitial cells and nematoblasts/nematocytes but no ganglionic cells. Results possibly suggest that the migrating interstitial cell population of 72-hour planulae is rich in committed precursors.     

Connexin 40 expression in bovine and rat retinas

Retinal neurons are extensively coupled through gap junction intercellular channels, but few connexin subtypes have been identified in mammalian retinal neurons. Based on previous findings that retinal gap junctional coupling is modulated by both dopamine and nitric oxide, presumably through connexin phosphorylation, we examined whether the connexin phosphoprotein subtype, connexin 40 (Cx40), was expressed in mammalian retinas. Immunostaining of rat and bovine retinas using Cx40-specific antibodies from two independent sources showed punctate staining between cells in the outer nuclear layer (ONL) and a sublayer of cells within the inner nuclear layer (INL). In addition, sparse punctate staining was detected in the ganglion cell/axon fiber layers (GCL/AFL). No punctate staining was observed in the outer (OS) or inner segment (IS) layers, and rarely in the outer plexiform layer (OPL) or inner plexiform layer (IPL). Double immunostaining of bovine retinas with antibodies to G(o), which stains bipolar cells, and to Cx40, showed little overlap, suggesting these bipolar cells do not express Cx40. Western blot analysis of alkaline-extracted bovine retinal membranes revealed Cx40 immunopositive bands of about 40 kD (monomer) and 80 kD (dimer). In both locations (monomer and dimer), the bands appeared as doublets, and their immunoreactivity was abolished when the antibody was pre-adsorbed with immunogenic Cx40 peptide. The doublet at 40 kD co-migrated with an immunopositive doublet present in heart membranes. Treatment with alkaline phosphatase altered the banding pattern of Cx40. The results suggest that the connexin phosphoprotein subtype, Cx40, is expressed within the neural layers of the mammalian retina.     

Sympathetic and sensory innervation of small intensely fluorescent (SIF) cells in rat superior cervical ganglion

The sympathetic ganglion contains small intensely fluorescent (SIF) cells derived from the neural crest. We morphologically characterize SIF cells and focus on their relationship with ganglionic cells, preganglionic nerve fibers and sensory nerve endings. SIF cells stained intensely for tyrosine hydroxylase (TH), with a few cells also being immunoreactive for dopamine β-hydroxylase (DBH). Vesicular acetylcholine transporter (VAChT)-immunoreactive puncta were distributed around some clusters of SIF cells, whereas some SIF cells closely abutted DBH-immunoreactive ganglionic cells. SIF cells contained bassoon-immunoreactive products beneath the cell membrane at the attachments and on opposite sites to the ganglionic cells. Ganglion neurons and SIF cells were immunoreactive to dopamine D2 receptors. Immunohistochemistry for P2X3 revealed ramified nerve endings with P2X3 immunoreactivity around SIF cells. Triple-labeling for P2X3, TH and VAChT allowed the classification of SIF cells into three types based on their innervation: (1) with only VAChT-immunoreactive puncta, (2) with only P2X3-immunoreactive nerve endings, (3) with both P2X3-immunoreactive nerve endings and VAChT-immunoreactive puncta. The results of retrograde tracing with fast blue dye indicated that most of these nerve endings originated from the petrosal ganglion. Thus, SIF cells in the superior cervical ganglion are innervated by preganglionic fibers and glossopharyngeal sensory nerve endings and can be classified into three types. SIF cells might modulate sympathetic activity in the superior cervical ganglion.     

A molecular phenotype atlas of the zebrafish retina

The rasborine cyprinid Danio rerio (the zebrafish) has become a popular model of retinal function and development. Its value depends, in part, on validation of homologies with retinal cell populations of cyprinine cyprinids. This atlas provides raw and interpreted molecular phenotype data derived from computationally classified sets of small molecule signals from different cell types in the zebrafish retina: L-alanine, L-aspartate, L-glutamine, L-glutamate, glutathione, glycine, taurine and γ-aminobutyrate. This basis set yields an 8-dimensional signature for every retinal cell and formally establishes molecular signature homologies with retinal neurons, glia, epithelia and endothelia of other cyprinids. Zebrafish photoreceptor classes have been characterized previously: we now show their metabolic profiles to be identical to those of the corresponding photoreceptors in goldfish. The inner nuclear layer is partitioned into precise horizontal, bipolar and amacrine cell layers. The horizontal cell layer contains at least three and perhaps all four known classes of cyprinine horizontal cells. Homologues of cyprinid glutamatergic ON-center and OFF-center mixed rod-cone bipolar cells are present and it appears likely that all five classes are present in zebrafish. The cone bipolar cells defy simple analysis but comprise the largest fraction of bipolar cells, as in all cyprinids. Signature analysis reveals six molecular phenotypes in the bipolar cell cohort: most are superclasses. The amacrine cell layer is composed of ≈64% GABA+ and 35% glycine+ amacrine cells, with the remainder being sparse dopaminergic interplexiform cells and other rare unidentified neurons. These different amacrine cell types are completely distinct in the dark adapted retina, but light adapted retinas display weak leakage of GABA signals into many glycinergic amacrine cells, suggesting widespread heterocellular coupling. The composition of the zebrafish ganglion cell layer is metabolically indistinguishable from that in other cyprinids, and the signatures of glial and non-neuronal cells display strong homologies with those in mammals. As in most vertebrates, zebrafish Müller cells possess a high glutamine, low glutamate signature and contain the dominant pool of glutathione in the neural retina. The retinal pigmented epithelium shows a general mammalian signature but also has exceptional glutathione content (5–10 mM), perhaps required by the unusually high oxygen tensions of teleost retinas. The optic nerve and the marginal zone of the retina reveal characteristic metabolic specializations. The marginal zone is strongly laminated and its nascent neurons display their characteristic signatures before taking their place in the retina proper.     

The Relationship between Visuospatial Impairments and Retinal and Cortical Thickness in Parkinson’s Disease

The paper touches upon current views on the pathophysiology of visuospatial impairments in Parkinson’s disease (PD). To assess thickness of retina’s ganglionic layer, retinal nerve fiber layer, and macular map, optical coherence tomography method was used. Brain MRI was also performed followed by evaluation of the cortical thickness. Patients underwent neuropsychological tests, including those for assessment of visuospatial perception and cognitive functions. We found certain retinal regions and areas of visual cortex with significant changes in PD patients on different stages. Our findings allowed us to speculate on the role of changes in the peripheral and central structures of the visual sensory system in the pathophysiology of visuospatial impairments in PD.     

Differential effect of dopamine on mitosis in early postnatal albino and pigmented rat retinae

Insufficient levels of L-DOPA, released from the retinal pigment epithelium (RPE), in albino animals are considered responsible for the abnormal development of the underlying neural retina. L-DOPA normalizes retinal neurogenesis by reducing levels of cell proliferation either by acting on the cells directly or by being converted into dopamine. Here we report the effects of dopamine on mitosis in early postnatal neural retinae from albino and pigmented rats, using 4D (x, y, z and time) confocal microscopy. Exogenous dopamine significantly prolongs mitosis in retinae from albino, but not pigmented, animals. As fewer cells move into and divide in the ventricular zone (VZ) in the presence of dopamine, we conclude that the overall cell cycle is affected. The D1 receptor blocker, SCH 23390, inhibits these effects. Thus, the differential effects of dopamine on neural retinae from pigmented and albino rats in vitro must result from the activation of D1 receptors, which are present in the retina from birth. Immunohistochemical labeling of D1 receptors shows that the pattern of their distribution is similar between pigmentation phenotypes, but levels of expression may be elevated in albinos. Labeling is most intense in the inner plexiform layer but is present throughout the neuroblastic layer. These findings are discussed in light of previous reports of reduced catecholamine levels in the albino retina.     

The transition zone in Hirschsprung's bowel contains abnormal hybrid ganglia with characteristics of extrinsic nerves

The aganglionic bowel in short-segment Hirschsprung's disease is characterized both by the absence of enteric ganglia and the presence of extrinsic thickened nerve bundles (TNBs). The relationship between the TNBs and the loss of enteric ganglia is unknown. Previous studies have described decreasing numbers of ganglia with increasing density of TNBs within the transition zone (TZ) between ganglionic and aganglionic gut, and there is some evidence of spatial contact between them in this region. To determine the cellular interactions involved, we have analysed the expression of perineurial markers of TNBs and enteric ganglionic markers for both neural cells and their ensheathing telocytes across four cranio-caudal segments consisting of most proximal ganglionic to most distal aganglionic from pull-through resected colon. We show that in the TZ, enteric ganglia are abnormal, being surrounded by perineurium cells characteristic of TNBs. Furthermore, short processes of ganglionic neurons extend caudally towards the aganglionic region, where telocytes in the TNB are located between the perineurium and nerve fibres into which they project telopodes. Thus, enteric ganglia within the TZ have abnormal structural characteristics, the cellular relationships of which are shared by the TNBs. These findings will help towards elucidation of the cellular mechanisms involved in the aetiology of Hirschsprung's disease.     

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. © 1993 John Wiley & Sons, Inc.     

Pax-6 expression during retinal regeneration in the adult newt

The present study examined the expression of Pax-6 during retinal regeneration in adult newts using in situ hybridization. In a normal retina, Pax-6 is expressed in the ciliary marginal zone, the inner part of the inner nuclear layer, and the ganglion cell layer. After surgical removal of the neural retina, retinal pigment epithelial cells proliferate into retinal precursor cells and regenerate a fully functional retina. At the beginning of retinal regeneration, Pax-6 was expressed in all retinal precursor cells. As regeneration proceeded, differentiating cells appeared at the scleral and vitreal margins of the regenerating retina, which had no distinct plexiform layers. In this stage, the expression of Pax-6 was localized in a strip of cells along the vitreal margin of the regenerating retina. In the late stage of regeneration, when the layer structure was completed, the expression pattern of Pax-6 became similar to that of a normal retina. It was found that Pax-6 is expressed in the retinal precursor cells in the early regenerating retina and that the expression pattern of Pax-6 changed as cell differentiation proceeded during retinal regeneration.     

Selective expression of the 180-kD component of the neural cell adhesion molecule N-CAM during development

The rodent neural cell adhesion molecule (N-CAM) consists of three glycoprotein chains of 180, 140, and 120 kD in their adult forms. Although the proportions of the three components are known to change during development and differ between brain regions, their individual distribution and function are unknown. Here we report studies carried out with a monoclonal antibody that specifically recognizes the 180-kD component of mouse N-CAM (N-CAM180) in its highly sialylated embryonic and less glycosylated adult forms. In primary cerebellar cell cultures, N-CAM180 antibody reacts intracellularly with all types of neural cells including astrocytes, oligodendrocytes, and neurons. During cerebellar, telencephalic, and retinal development N-CAM180 is detectable by indirect immunohistology in differentiated neural cells, but, in contrast to total N-CAM, not in their proliferating precursors in the ventricular zone and primordial and early postnatal external granular layer. In monolayer cultures of C1300 neuroblastoma cells, N-CAM180 appears by immunofluorescence more concentrated at contact points between adjacent cells, while N-CAM comprising the 180- and 140-kD component shows a more uniform distribution at the plasma membrane. Treatment of neuroblastoma cells with dimethylsulfoxide, which promotes differentiation, induces a shift toward the predominant expression of N- CAM180. These observations support the notion that N-CAM180 is expressed selectively in more differentiated neural cells and suggest a differential role of N-CAM180 in the stabilization of cell contacts.