Retinal degeneration induced by N-methyl-N-nitrosourea in Syrian golden hamsters

BACKGROUND: The sequential retinal changes in Syrian golden hamsters induced by N-methyl-N-nitrosourea (MNU) have not been studied. METHODS: Female hamsters received a single intraperitoneal injection of 90 mg/kg MNU at 50 days of age, and the retina was examined light and electron microscopically, immunohistochemically and by the TdT-mediated dUTP-digoxigenin nick end labeling (TUNEL) method until 20 weeks after the treatment. RESULTS: The retinal changes were as follows: (1) Photoreceptor apoptosis occurred 1 day after the treatment and resulted in photoreceptor loss at day 7. During the degeneration, Müller cell proliferation was conspicuous at day 5. (2) After the photoreceptor cell loss, migration of the pigment epithelial cells in all layers of the retina which were in contact with blood vessels occurred. Due to the Müller cell proliferation, gliosis was prominent at the later stage. CONCLUSIONS: The MNU injection caused photoreceptor apoptosis followed by pigment epithelial cell migration around the blood vessels, accompanied by gliosis. The primary event and the course of this disease closely resemble those of retinitis pigmentosa in humans.     

Late proliferation of retinal Müller cell progenitors facilitates preferential targeting with retroviral vectors in vitro

During vertebrate neural retina development, the relationship between mitotic activity in progenitor cells and the acquisition of a mature cell phenotype remains an area of controversy. The Müller glial cell has long been recognized as one of the last cell types of the retina to mature, which occurs under the influence of cell-cell interactions. In this report we examine the acquisition of the Müller cell phenotype in relation to mitotic activity. Using immunohistochemical markers, we demonstrate that a gene product characteristic of mature Müller cells, the 2M6 antigen, is expressed in mitotically active cells, even after all the major retina architectural features have been laid down. Furthermore, we show that retroviral infection, a process that requires mitotically active cells, preferentially targets Müller cell progenitors when late embryonic retina is infected in vitro. The two lines of evidence are consistent with a model for Müller cell differentiation that includes a mitotically active progenitor that has already begun to express specific differentiation gene products.     

Tractional force generation by porcine Müller cells. Development and differential stimulation by growth factors

PURPOSE: To assess the ability of retinal Müller cells to generate tractional forces during dedifferentiation in culture and to assess their responsiveness to contraction-stimulating growth factors. METHODS: Müller cells were isolated from papain-DNase-digested porcine retina. The identity of the isolated cells was confirmed by immunodetection of carbonic anhydrase II (CA-II), cellular retinaldehyde-binding protein (CRALBP), glial fibrillary acidic protein (GFAP), vimentin, and alpha smooth muscle actin (alpha SMA). Tractional force generation was assessed as a function of Müller cell contraction of collagenous extracellular matrices in vitro. The effects of potential promoters were assessed by addition directly to culture medium. The contributions of specific promoting to the contraction-promoting activity in serum were assessed by adding neutralizing antibodies and measuring loss of stimulatory activity. RESULTS: Freshly isolated Müller cells did not generate substantial matrix contraction. However, this activity increased 150-fold within 12 days in culture and continued to increase during the next 21 days. Development of the capacity for extracellular matrix contraction coincided with the acquisition of immunodetectable alpha SMA and loss of GFAP. Matrix contraction by Müller cells was stimulated in a dose-dependent fashion by human serum, platelet-derived growth factor (PDGF), and insulin-like growth factor-I (IGF-I). Müller cells were not stimulated by transforming growth factor beta 1 (TGF beta 1), transforming growth factor beta 2 (TGF beta 2), or endothelin-1 (E1). Neutralizing antibodies against PDGF and IGF-I reduced the activity in human serum by 37% and 58%, respectively, and 87% when added together. CONCLUSIONS: Porcine Müller cells in culture acquire the ability to contract extracellular matrices and thus generate tractional forces. Acquisition of this activity coincides with alpha SMA expression and loss of GFAP. Further, this activity is dependent on the presence of exogenous promoters, including PDGF or IGF-I.     

The NIDDK liver transplantation database

Gap junctional communication between glial cells is thought to play a role in K+ spatial buffering, in the propagation of inter-astrocytic Ca2+ waves, and in glial-neuronal signaling. In the present study, we characterize dye coupling between astrocytes, and between astrocytes and Müller cells, in the isolated rat retina. Whole-cell patch recordings were obtained from retinal astrocytes and Müller cells and the cells filled with Lucifer Yellow and neurobiotin. Spread of Lucifer Yellow to two to ten neighboring astrocytes occurred in 90% of the astrocyte recordings. After fixation and incubation of the retina with fluorescent conjugated streptavidin, neurobiotin was seen to label clusters of 13-88 astrocytes, as well as > 100 Müller cells. In contrast, when Müller cells were filled with Lucifer Yellow and neurobiotin, both tracers were confined solely to the recorded Müller cell. The uncoupling agents octanol, halothane, and doxyl-stearic acid were tested for their ability to uncouple retinal glia in situ. All three agents eliminated the visible spread of Lucifer Yellow from the injected astrocyte and the spread of neurobiotin into Müller cells. However, only doxyl-stearic acid combined with octanol eliminated the spread of neurobiotin between astrocytes. These results demonstrate that astrocytes in the rat retina are coupled to each other and to Müller cells. The astrocyte-to-Müller cell coupling is asymmetric, allowing transfer of the tracer in the forward direction only. In addition, astrocyte-to-Müller cell coupling is more sensitive to the uncoupling agents tested than is astrocyte-to-astrocyte coupling.     

Distribution of mitochondria within Müller cells--I. Correlation with retinal vascularization in different mammalian species

The distribution of mitochondria within retinal glial (Müller) cells and neurons was studied by electron microscopy, by confocal microscopy of a mitochondrial dye and by immunocytochemical demonstration of the mitochondrial enzyme GABA transaminase (GABA-T). We studied sections and enzymatically dissociated cells from adult vascularized (human, pig and rat) and avascular or pseudangiotic (guinea-pig and rabbit) mammalian retinae. The following main observations were made. (1) Müller cells in adult euangiotic (totally vascularized) retinae contain mitochondria throughout their length. (2) Müller cells from the periphery of avascular retinae display mitochondria only within the sclerad-most end of Müller cell processes. (3) Müller cells from the vascularized retinal rim around the optic nerve head in guinea-pigs contain mitochondria throughout their length. (4) Müller cells from the peripapillar myelinated region ('medullary rays') of the pseudangiotic rabbit retina contain mitochondria up to their soma. In living dissociated Müller cells from guinea-pig retina, there was no indication of low intracellular pH where the mitochondria were clustered. These data support the hypothesis that Müller cells display mitochondria only at locations of their cytoplasm where the local O2 pressure (pO2) exceeds a certain threshold. In contrast, retinal ganglion cells of guinea-pig and rabbit retinae display many mitochondria although the local pO2 in the inner (vitread) retinal layers has been reported to be extremely low. It is probable that the alignment of mitochondria and the expression of mitochondrial enzymes are regulated by different mechanisms in various types of retinal neurons and glial cells.     

Retinal FABP principally localizes to neurons and not to glial cells

The presence of fatty acid-binding protein (FABP) in the embryonic chick retina may be linked to the demand for polyunsaturated fatty acids in this developing neural tissue. There is a decline in the overall level of FABP as the retina matures, suggesting a role for FABP in cellular differentiation. However, this pattern is not present in the chick brain, indicating a unique function for FABP in the retina. Immunohistochemical staining of paraffin sections of chick retina from embryonic day 21 revealed immunopositive photoreceptor inner segments, outer nuclear layer, 'radial processes' in the inner nuclear layer, a subpopulation of cells in the ganglion cell layer, and inner limiting membrane. This pattern suggested that FABP positive cells were photoreceptors, Müller (glial) cells, and possibly ganglion cells. Staining of sections for glutamine synthetase, an enzyme specific for Müller cells, was similar but not identical to the pattern observed with FABP; thus identification of these cells as FABP-positive was not conclusive. However, in retinal cells dissociated from day E14 embryos and cultured for one week, staining with FABP was more intense in the neurons than in the 'flat' cells (presumed to be derived from the Müller cells). Retinal FABP thus appears to be localized predominantly in neurons, and may serve to sequester fatty acids in preparation for neurite outgrowth as the retinal cells differentiate.     

The Müller (glial) cell in normal and diseased retina: a case for single-cell electrophysiology

In the retina of most vertebrates there exists only one type of macroglia, the Müller cell. Müller cells express voltage-gated ion channels, neurotransmitter receptors and various uptake carrier systems. These properties enable the Müller cells to control the activity of retinal neurons by regulating the extracellular concentration of neuroactive substances such as K+, GABA and glutamate. We show here how electrophysiological recordings from enzymatically dissociated mammalian Müller cells can be used to study these mechanisms. Müller cells from various species have Na(+)-dependent GABA uptake carriers, but only cells from primates have additional GABA receptors that activate Cl- channels. Application of glutamate analogues causes enhanced membrane currents recorded from Müller cells in situ but not from isolated cells. We show that mammalian Müller cells have no ionotropic glutamate receptors but respond to increased K+ release from glutamate-stimulated retinal neurons. This response is involved in extracellular K+ clearance and is mediated by voltage-gated (inwardly rectifying) K+ channels which are abundantly expressed by healthy Müller cells. In various cases of human retinal pathology, currents through these channels are strongly reduced or even extinguished. Another type of voltage-gated ion channels, observed in Müller cells from many mammalian species, are Na+ channels. In Müller cells from diseased human retinae, voltage-dependent Na+ currents were significantly increased in comparison to cells from control donors. Thus, the expression of glial ion channels seems to be controlled by neuronal signals. This interaction may be involved in the pathogenesis of retinal gliosis which inevitably accompanies any degeneration of retinal neurons. In particular, Müller cell proliferation may be triggered by mechanisms requiring the activation of Ca(2+)-dependent K+ channels. Ca(2+)-dependent K+ currents are easily elicitable in Müller cells from degenerating retinae and can be blocked by 1 mM TEA (tetraethylammonium). In purified Müller cell cultures, the application of 1 mM TEA greatly reduces the proliferative activity of the cells. These data clearly show that Müller cells are altered in cases of neuronal degeneration and may be crucially involved in pathogenetic mechanisms of the retina.     

Müller cells in the retina of the cane toad, Bufo marinus, express neuropeptide Y-like immunoreactivity

We have used light- and electron-microscopic immunohistochemistry to identify the presence of immunoreactivity to neuropeptide Y (NPY) within Müller cells in the retina of the cane toad, Bufo marinus. Müller cells containing NPY-like immunoreactivity (NPY-LI) were identified at the light-microscopic level by the coexistence with immunoreactivity to glial fibrillary acidic protein (GFAP) and at the ultrastructural level by their characteristic relationship to neuron cell bodies and processes. At the light-microscopic level, those cells which contained both NPY-LI and GFAP-LI usually had small cell bodies in the inner nuclear layer, while those cells which contained only NPY-LI were identified as large and small amacrine cells. The radially oriented primary processes in the inner plexiform layer and the vitreal end feet of GFAP-LI Müller cells also expressed NPY-LI. At the ultrastructural level, thin lamellar processes of Müller cells with NPY-LI enclosed some amacrine cell bodies in the inner nuclear layer and amacrine cell dendrites in the inner plexiform layer. These observations suggest that NPY-LI is localized in Müller cells in addition to two types of amacrine cells previously identified in the Bufo retina. This study provides the first evidence that glial elements in the vertebrate retina express NPY-LI.     

Alterations in the Ha-ras-1 and the p53 pathway genes in the progression of N-methyl-N-nitrosourea-induced rat mammary tumors

Glutamate is the most prominent excitatory neurotransmitter in the retina and brain. It has become clear that the physiology of many glial cells, including retinal Müller cells, is modified by a host of neurotransmitters, including glutamate. The experiments presented here demonstrate that Müller cells isolated from the tiger salamander retina have metabotropic glutamate receptors that, when activated, lead to the release of calcium ions (Ca2+) from intracellular stores. The Ca2+-sensitive fluorescent dye, Fura-2, and video imaging microscopy were used to monitor changes in cytosolic calcium ion concentration ([Ca2+]i) evoked by glutamate (30-50 microM), (1S,3R)-ACPD (50-200 microM), quisqualate (10-50 microM), and L-AP4 (5-100 microM). Bath application of each of these metabotropic receptor agonists in the absence of extracellular Ca2+ resulted in an increase in [Ca2+]i that often began in the distal end of the cell and occurred later in the endfoot. This wavelike increase in [Ca2+]i is reminiscent of the Ca2+ waves evoked in these cells by other Ca2+ releasing agents such as ryanodine and caffeine. Extracellular application ofATP also evoked increases in [Ca2+] in Müller cells. The presence on Müller cells of receptors for retinal neurotransmitters, such as glutamate and ATP, demonstrates that these glial cells can respond to changes in the retinal extracellular environment and hence neuronal activity. Since Müller cells span almost all layers of the retina, they are likely to be exposed to most retinal neurotransmitters. The Ca2+ waves evoked in Müller cells by neurotransmitters could represent a form of signaling from the outer retinal layers to the inner ones.     

Preconditioning with bright light evokes a protective response against light damage in the rat retina

Constant exposure to bright light induces photoreceptor degeneration and at the same time upregulates the expression of several neurotrophic factors in the retina. At issue is whether the induced neurotrophic factors protect photoreceptors. We used a preconditioning paradigm to show that animals preconditioned with bright light became resistant to subsequent light damage. The preconditioning consisted of a 12-48 hr preexposure, followed by a 48 hr "rest phase" of normal cyclic lighting. The greatest protection was achieved by a 12 hr preexposure. Preconditioning induces a prolonged increase in two endogenous neurotrophic factors: basic fibroblast growth factor (bFGF) and ciliary neurotrophic factor (CNTF). It also stimulates the phosphorylation of extracellular signal-regulated protein kinases (Erks) in both photoreceptors and Müller cells. These findings indicate that exposure to bright light initiates two opposing processes: a fast degenerative process that kills photoreceptors and a relatively slower process that leads to the protection of photoreceptors. The extent of light damage, therefore, depends on the interaction of the two processes. These results also suggest a role of endogenous bFGF and CNTF in photoreceptor protection and the importance of Erk activation in photoreceptor survival.     

C-reactive protein as an outcome predictor for maintenance hemodialysis patients

We produced the monoclonal antibody RT10F7, characterized its antigenic specificity and expression in the adult and developing retina, in cultured retinal cells and in other parts of the central nervous system. In metabolically-labelled retinal cultures RT10F7 immunoprecipitated a protein of approximately 36,000 mol. wt. In the adult, RT10F7 stained endfeet of Müller cells in the ganglion cell layer, four horizontal bands in the inner plexiform layer, and radial fibres in the outer plexiform layer which terminated at the outer limiting membrane. In the inner nuclear layer, most somata were underlined by Müller processes that wrapped around them, but some cell bodies were immunoreactive for RT10F7 in the cytoplasm. During development, postnatal day 21 was the first age at which the adult pattern of immunoreactivity was present, although a fourth band in the inner plexiform layer was less clear than for the adult. By 14 and eight days after birth, the pattern of RT10F7 immunoreactivity approximated that of the adult; however, only three bands and one band were present, respectively, in the inner plexiform layer. At earlier ages, postnatal days 4, 1 and embryonic ages 19 and 15, the monoclonal antibody stained Müller cell endfeet and radial fibres, from the inner plexiform layer through the neuroblastic layer to the outer limiting membrane. At these ages, the immunoreactivity was more prominent at the level of Müller cell endfeet. The monoclonal antibody stained glia in preparations of dissociated retinal cells maintained in culture but not astrocytes or oligodendrocytes from optic nerve cultures. In brain sections, tanycytes exhibited RT10F7 immunoreactivity. The monoclonal antibody RT10F7 recognized a specific cell type in the retina, the Müller cell. In the adult and developing retina, RT10F7 recognized an antigen that is present primarily in Müller cell processes. This feature allowed us to follow the maturation of the Müller cell and correlate it with developmental events in the retina. RT10F7 is a specific marker for Müller cells in vivo and in vitro and may be useful for studies of function of Müller cells after ablation or after injuries that are known to activate Müller cells.     

Regulation of insulin-like growth factors I and II and their binding proteins in human bone marrow stromal cells by dexamethasone

Müller cells are highly permeable to potassium ions and play a major role in maintaining potassium homeostasis in the vertebrate retina during light-evoked neuronal activity. Potassium fluxes across the Müller cell's membrane are believed to underlie the light-evoked responses of these cells. We studied the potassium currents of turtle Müller cells in the retinal slice and in dissociated cell preparations and their role in the genesis of the light-evoked responses of these cells. In either preparation, the I-V curve, measured under voltage-clamp conditions, consisted of inward and outward currents. A mixture of cesium ions, TEA, and 4-AP blocked the inward current but had no effect on the outward current. Extracellular cesium ions alone blocked the inward current but exerted no effect on the photoresponses. Extracellular barium ions blocked both inward and outward currents, induced substantial depolarization, and augmented the light-evoked responses, especially the OFF component. Exposing isolated Müller cells to a high potassium concentration did not cause any current or voltage responses when barium ions were present. In contrast, application of glutamate in the presence of barium ions induced a small inward current that was associated with a substantially augmented depolarizing wave relative to that observed under control conditions. This observation suggests a role for an electrogenic glutamate transporter in generating the OFF component of the turtle Müller cell photoresponse.     

Prolonged delivery of brain-derived neurotrophic factor by adenovirus-infected Müller cells temporarily rescues injured retinal ganglion cells

In this study, we demonstrate that: (i) injection of an adenovirus (Ad) vector containing the brain-derived neurotrophic factor (BDNF) gene (Ad.BDNF) into the vitreous chamber of adult rats results in selective transgene expression by Müller cells; (ii) in vitro, Müller cells infected with Ad.BDNF secrete BDNF that enhances neuronal survival; (iii) in vivo, Ad-mediated expression of functional BDNF by Müller cells, temporarily extends the survival of axotomized retinal ganglion cells (RGCs); 16 days after axotomy, injured retinas treated with Ad.BDNF showed a 4.5-fold increase in surviving RGCs compared with control retinas; (iv) the transient expression of the BDNF transgene, which lasted approximately 10 days, can be prolonged with immunosuppression for at least 30 days, and such Ad-mediated BDNF remains biologically active, (v) persistent expression of BDNF by infected Müller cells does not further enhance the survival of injured RGCs, indicating that the effect of this neurotrophin on RGC survival is limited by changes induced by the lesion within 10-16 days after optic nerve transection rather than the availability of BDNF. Thus, Ad-transduced Müller cells are a novel pathway for sustained delivery of BDNF to acutely-injured RGCs. Because these cells span the entire thickness of the retina, Ad-mediated gene delivery to Müller cells may also be useful to influence photoreceptors and other retinal neurons.     

Müller glia cells and their possible roles during retina differentiation in vivo and in vitro

Müller cells are astrocyte-like radial glia cells which are formed exclusively in the retina. Here we present evidence that Müller cells are crucially involved in the development of the retina's architecture and circuitry. There is increasing evidence that Müller cells are present from the very early beginning of retinogenesis. We postulate the "gradual maturation hypothesis of Müller cells". According to this hypothesis, Müller cells are continuously generated by a gradual transition of neuroepithelial stem cells into mature Müller cells. This process may be partly reversible. Müller cells, or their immature precursors, are able to subserve different functions. They are primary candidates for stabilizing the complex retinal architecture and for providing an orientation scaffold. Thereby, they introduce a reference system for the migration and correct allocation of neurons. Moreover, they may provide spatial information and microenvironmental cues for differentiating neurons, and may also be important for the segregation of cell and fibre layers. Additionally, they seem to be involved in the guidance of axonal fibres both in radial and in lateral directions, as they are involved in the support and stabilization of synapses.     

Rapid increase in cytosolic calcium ion concentration mediated by acetylcholine receptors in cultured retinal neurons and Müller cells

PURPOSE: This study was conducted to detect the presence of muscarinic or nicotinic receptors in cultured retinal neurons and Müller cells. METHODS: Pure Müller cell cultures and cocultures of retinal neurons and Müller cells were used; the former, obtained from adult rabbit retinas, and the latter, retinal neurons from neonatal rats, were cocultured with Müller cells. Intracellular calcium ion concentration ([Ca2+]i) following the administration of acetylcholine, a cholinesterase inhibitor (trichlorfon), nicotine or muscarinic agonist with or without a receptor antagonist was monitored using the calcium ion indicator, fura-2. RESULTS: Acetylcholine and trichlorfon induced rapid increase in [Ca2+]i in half of either cell type. Trichlorfon induced positive response in coculture but not in the pure Müller cell cultures. This positive response was blocked only partially in the presence of atropine. Approximately 30-40% of neurons responded to nicotine at 5 microM, which was significantly blocked by alpha-bungarotoxin at 50 nM. No response to nicotine could be detected in Müller cells. Approximately 50% of neurons responded to muscarine at 50 microM, but 500 microM was required for the formation of calcium transients in 50% of Müller cells. The muscarine inducement of rapid increase in [Ca2+]i was blocked by atropine. The agonist of M1 (a muscarinic receptor subtype), McN-A-343, at 0.5 microM induced the most significant and rapid increase in [Ca2+]i both in neurons and Müller cells. McN-A-343 administration at 0.05 microM induced positive response in half the neurons, but only in approximately 10% of Müller cells. Such positive response was not observed following preincubation with the M1 antagonist, pirenzepine, at 50 microM. CONCLUSIONS: Cocultured retinal neurons enhance the release of acetylcholine following anticholinesterase administration, and approximately half the neurons were found to possess muscarinic and nicotinic receptors. However, Müller cells appeared to possess only the less sensitive muscarinic receptor. Muscarinic receptor subtypes on either type of cell contained at least M1.     

Effects of clonidine, dexmedetomidine and xylazine on thermal antinociception in rhesus monkeys

The antinociceptive effects of the s.c. administration of the alpha-2 agonists clonidine (0.0032-1.0 mg/kg), dexmedetomidine (0.001-0.032 mg/kg) and xylazine (0.1-3.2 mg/kg) were examined in the warm-water tail withdrawal assay in rhesus monkeys. The three agonists were dose-dependently effective in this assay; their potency order being dexmedetomidine > clonidine > xylazine. The alpha-2 antagonist idazoxan (0.1-3.2 mg/kg) caused dose-dependent and roughly parallel rightward shifts in the dose-effect curves for the three agonists. Apparent pA2 analysis with idazoxan yielded homogeneous values for the three agonists, supporting the notion that similar receptors mediate their antinociceptive effects. The opioid antagonist quadazocine (1.0 mg/kg) did not antagonize the antinociceptive effects of clonidine and xylazine, indicating that opioid receptors do not participate in the effects of the compounds in this assay. At dose ranges found to be effective in the antinociceptive assay, clonidine, dexmedetomidine and xylazine also dose-dependently caused sedation, muscle relaxation, bradycardia and moderate respiratory depression. The sedative, muscle relaxant and respiratory depressant effects of xylazine could be antagonized by idazoxan, suggesting that these effects may be mediated through alpha-2 receptors. These data indicate that the three imidazoline alpha-2 agonists, clonidine, dexmedetomidine and xylazine are effective s.c. in the warm-water tail withdrawal assay in rhesus monkeys, but only at doses that produce other behavioral and physiological effects.     

Relationship between lipophilicity and binding to human serum albumin of arylpropionic acid non-steroidal anti-inflammatory drugs

In cases of retinal light damage, glaucoma, or senile macula degeneration, the loss of retinal neurons is thought to cause alterations of glial cells. We performed immunocytochemical studies on retinae of (i) healthy rats and human donors, (ii) rats exposed to enhanced illumination for 24 months, a procedure which leads to complete loss of photoreceptor cells, (iii) a human donor who had suffered from senile macula (photoreceptor cell) degeneration, and (iv) human donors who had suffered from glaucoma, known to be accompanied by a loss of ganglion cells and other retinal neurons. Furthermore, Müller cells were enzymatically isolated from human glaucomatous retinae. All preparations were subjected to immunocytochemistry for CD44 antigen and Apolipoprotein E (ApoE). In normal rat and human retinae, CD44 immunoreactivity was observed in the microvillous sclerad processes of Müller cells: in human retinae, perivascular (astro-)glial cell processes were also CD44 immunopositive. ApoE immunoreactivity was only found in some perivascular (astro-)glial cell processes of human retinae. Both rat and human Müller cells respond to photoreceptor cell damage by increased, and ectopic, expression of the CD44 antigen. Increased ApoE immunoreactivity was found in Müller cells from degenerative human retinae, but rarely in light-damaged rat retinae. It is concluded that degeneration-related reorganization involves enhanced expression of the glial cell adhesion molecule CD44 as well as elevated activity of the glial lipid transport molecule ApoE.     

Replication slippage may cause parallel evolution in the secondary structures of mitochondrial transfer RNAs

We observed the three dimensional structure of cellular aggregates formed from chick retinal cells in a floating culture system for 2 months. The aggregated cells partially mimicked the structure of the retina and showed differentiation of photoreceptor cells and Müller cells with numerous synapses. Immunohistochemical studies showed the number of anti-rhodopsin positive cells increasing over time. In the long-term culture, increasing anti-crystalline positive cells appeared late in the culture, indicative of differentiation of lens epithelial cells. Nerve, epidermal, and basic fibroblast growth factors, and co-culture with retinal pigment epithelial cells stimulated to some degree the growth of dendrites in retinal cellular aggregates. Epidermal growth factor, in particular, promoted the production of rhodopsin in photoreceptor cells. Retinal cellular aggregates in a floating culture system could be used to examine the effect of various factors on differentiation of the neuroretina.     

Na+ channels are expressed by mammalian retinal glial (Müller) cells

Müller cells constitute the principal glia of the vertebrate retina. Unlike other types of neuroglial cells such as astrocytes and Schwann cells, Müller cells have not yet been demonstrated to express Na+ channels. Here we present first evidence of Müller cell Na+ currents from voltage-clamp studies in enzymatically isolated cells. Some cells from retinae of cats and dogs, but none from rabbit or guinea-pig retinae, revealed fast and rapidly inactivating inward currents in response to depolarizing voltage steps. The currents reversibly disappeared in Na+ free solutions or under tetrodotoxin (TTX, 1 microM). Activation and inactivation characteristics of these currents were strikingly similar to those of neurone-type Na+ channels.     

Elevation of soluble IL-2 receptor and IL-4, and nonelevation of IFN-gamma in sera from patients with allergic asthma

This study is the first demonstration of glial fibrillary acidic protein (GFAP)-immunoreactivity in the retina of the lamprey Lampetra fluviatilis. This immunoreactivity is expressed on one hand, in radial processes and somata which belong to Müller cells and, on the other hand, in horizontal fibers in the intermediate plexus between horizontal cells. The tracing of these fibers to Müller cells or horizontal cells is discussed.