Ianors Iandiev

Cellular signaling and factors involved in Müller cell gliosis: Neuroprotective and detrimental effects

Müller cells are active players in normal retinal function and in virtually all forms of retinal injury and disease. Reactive Müller cells protect the tissue from further damage and preserve tissue function by the release of antioxidants and neurotrophic factors, and may contribute to retinal regeneration by the generation of neural progenitor/stem cells. However, Müller cell gliosis can also contribute to neurodegeneration and impedes regenerative processes in the retinal tissue by the formation of glial scars. This article provides an overview of the neuroprotective and detrimental effects of Müller cell gliosis, with accounts on the cellular signal transduction mechanisms and factors which are implicated in Müller cell-mediated neuroprotection, immunomodulation, regulation of Müller cell proliferation, upregulation of intermediate filaments, glial scar formation, and the generation of neural progenitor/stem cells. A proper understanding of the signaling mechanisms implicated in gliotic alterations of Müller cells is essential for the development of efficient therapeutic strategies that increase the supportive/protective and decrease the destructive roles of gliosis.

A potassium channel-linked mechanism of glial cell swelling in the postischemic retina.

The cellular mechanisms underlying glial cell swelling, a central cause of edema formation in the brain and retina, are not yet known. Here, we show that glial cells in the postischemic rat retina, but not in control retina, swell upon hypotonic stress. Swelling of control cells could be evoked when their K(+) channels were blocked. After transient ischemia, glial cells strongly downregulated their K(+) conductance and their prominent Kir4.1 protein expression at blood vessels and the vitreous body. In contrast, the expression of the aquaporin-4 (AQP4) (water channel) protein was only slightly altered after ischemia. Activation of D(2) dopaminergic receptors prevents the hypotonic glial cell swelling. The present results elucidate the coupling of transmembraneous water fluxes to K(+) currents in glial cells and reveal the role of altered K(+) channel expression in the development of cytotoxic edema. We propose a mechanism of postischemic glial cell swelling where a downregulation of their K(+) conductance prevents the emission of intracellularly accumulated K(+) ions, resulting in osmotically driven water fluxes from the blood into the glial cells via aquaporins. Inhibition of these water fluxes may be beneficial to prevent ischemia-evoked glial cell swelling.

Role of retinal glial cells in neurotransmitter uptake and metabolism

In addition to photoreceptors and neurons, glial cells (in particular Müller cells) contribute to the removal and metabolization of neurotransmitters in the neural retina. This review summarizes the present knowledge regarding the role of retinal glial cells in the uptake of glutamate, N-acetylaspartylglutamate, gamma-aminobutyric acid, glycine, and d-serine, as well as the degradation and removal of purinergic receptor agonists. Some major pathways of glutamate metabolism in Müller cells are described; these pathways are involved in the glutamate-glutamine cycle of the retina, in the defense against oxidative and nitrosative stress via the production of glutathione, and in the production of substrates for the neuronal energy metabolism. In addition, the developmental regulation of the major glial glutamate transporter, GLAST, and of the glia-specific enzyme glutamine synthetase is described, as well as the importance of a malfunction and even reversal of glial glutamate transporters, and a downregulation of the glutamine synthetase, as pathogenic factors in different retinopathies.

Müller cells as players in retinal degeneration and edema

BackgroundUnder normal conditions, Müller cells support neuronal activity and the integrity of the blood-retinal barrier, whereas gliotic alterations of Müller cells under pathological conditions may contribute to retinal degeneration and edema formation. A major function of Müller cells is the fluid absorption from the retinal tissue, which is mediated by transcellular water transport coupled to currents through potassium channels.MethodsAlterations of retinal Müller cells under pathological conditions were investigated by immunohistochemistry and recording their behavior under osmotic stress.ResultsIn animal models of various retinopathies, e.g., retinal ischemia, ocular inflammation, retinal detachment, and diabetes, it was found that Müller cells decrease the expression of their major potassium channel (Kir4.1). This alteration is associated with an impairment of the rapid water transport across Müller cell membranes, as recognizable in the induction of cellular swelling under hypoosmolar conditions. Osmotic swelling of Müller cells is also induced by oxidative stress and by inflammatory mediators such as arachidonic acid and prostaglandins.ConclusionsThe data suggest that a disturbed fluid transport through Müller cells is (in addition to vascular leakage) a pathogenic factor contributing to the development of retinal edema. Pharmacological re-activation of the retinal water clearance by Müller cells may represent an approach to the development of new edema-resolving drugs. Triamcinolone acetonide, which is clinically used to resolve edema, prevents osmotic swelling of Müller cells as it induces the release of endogenous adenosine and subsequent A1 receptor activation which results in the opening of ion channels. Apparently, triamcinolone resolves edema by both inhibition of vascular leakage and stimulation of retinal fluid clearance by Müller cells.

Reactive glial cells: increased stiffness correlates with increased intermediate filament expression

Increased stiffness of reactive glial cells may impede neurite growth and contribute to the poor regenerative capabilities of the mammalian central nervous system. We induced reactive gliosis in rodent retina by ischemia‐reperfusion and assessed intermediate filament (IF) expression and the viscoelastic properties of dissociated single glial cells in wild‐type mice, mice lacking glial fibrillary acidic protein and vimentin (GFAP−/−Vim−/−) in which glial cells are consequently devoid of IFs, and normal Long‐Evans rats. In response to ischemia‐reperfusion, glial cells stiffened significantly in wild‐type mice and rats but were unchanged in GFAP−/− Vim−/− mice. Cell stiffness (elastic modulus) correlated with the density of IFs. These results support the hypothesis that rigid glial scars impair nerve regeneration and that IFs are important determinants of cellular viscoelasticity in reactive glia. Thus, therapeutic suppression of IF up‐regulation in reactive glial cells may facilitate neuroregeneration.—Lu, Y.‐B., Iandiev, I., Hollborn, M., Korber, N., Ulbricht, E., Hirrlinger, P. G., Pannicke, T., Wei, E.‐Q., Bringmann, A., Wol‐burg, H., Wilhelmsson, U., Pekny, M., Wiedemann, P., Reichenbach, A., Kas, J. A. Reactive glial cells: increased stiffness correlates with increased intermediate filament expression. FASEB J. 25, 624–631 (2011). www.fasebj.org

Retinal gene expression and Müller cell responses after branch retinal vein occlusion in the rat.

PURPOSE In a rat model of branch retinal vein occlusion (BRVO), changes in gene expression of factors implicated in the development of retinal edema and alterations in the properties of Müller cells were determined. METHODS In adult Long-Evans rats, BRVO was induced by laser photocoagulation of retinal veins; untreated eyes served as controls. The mRNA levels of after factors were determined with real-time RT-PCR in the neural retina and retinal pigment epithelium after 1 and 3 days of BRVO: VEGF-A, pigment epithelium-derived factor (PEDF), tissue factor, prothrombin, the potassium channel Kir4.1, and aquaporins 1 and 4. Potassium currents were recorded in isolated Müller cells, and cellular swelling was assessed in retinal slices. RESULTS In the neural retina, the expression of VEGF was upregulated within 1 day of BRVO and returned to the control level after 3 days. PEDF was upregulated in the neuroretina and retinal pigment epithelium after 3 days of BRVO. Prothrombin, Kir4.1, and both aquaporins were downregulated in the neuroretina. After BRVO, Müller cells displayed a decrease in their potassium currents and an altered distribution of Kir4.1 protein, an increase in the size of their somata, and cellular swelling under hypoosmotic stress that was not observed in control tissues. CONCLUSIONS BRVO results in a rapid transient increase in the expression of VEGF and a delayed increase in the expression of PEDF. The downregulation of Kir4.1 and aquaporins, the mislocation of Kir4.1 protein, and the osmotic swelling of Müller cells may contribute to the development of edema and neuronal degeneration.

Tandem‐pore domain potassium channels are functionally expressed in retinal (Müller) glial cells

Tandem‐pore domain (2P‐domain) K+‐channels regulate neuronal excitability, but their function in glia, particularly, in retinal glial cells, is unclear. We have previously demonstrated the immunocytochemical localization of the 2P‐domain K+ channels TASK‐1 and TASK‐2 in retinal Müller glial cells of amphibians. The purpose of the present study was to determine whether these channels were functional, by employing whole‐cell recording from frog and mammalian (guinea pig, rat and mouse) Müller cells and confocal microscopy to monitor swelling in rat Müller cells. TASK‐like immunolabel was localized in these cells. The currents mediated by 2P‐domain channels were studied in isolation after blocking Kir, KA, KD, and BK channels. The remaining cell conductance was mostly outward and was depressed by acid pH, bupivacaine, methanandamide, quinine, and clofilium, and activated by alkaline pH in a manner consistent with that described for TASK channels. Arachidonic acid (an activator of TREK channels) had no effect on this conductance. Blockade of the conductance with bupivacaine depolarized the Müller cell membrane potential by about 50%. In slices of the rat retina, adenosine inhibited osmotic glial cell swelling via activation of A1 receptors and subsequent opening of 2P‐domain K+ channels. The swelling was strongly increased by clofilium and quinine (inhibitors of 2P‐domain K+ channels). These data suggest that 2P‐domain K+ channels are involved in homeostasis of glial cell volume, in activity‐dependent spatial K+ buffering and may play a role in maintenance of a hyperpolarized membrane potential especially in conditions where Kir channels are blocked or downregulated.

Selective Staining by Vital Dyes of Muller Glial Cells in Retinal Wholemounts

Müller glial cells within the retina may respond to different signaling molecules with an elevation of their intracellular free calcium. To prove the localization of the recorded calcium responses in Müller cells within acutely isolated retinal wholemounts, retinal pieces from adult animals and humans were exposed to different vital dyes just after the calcium imaging records were finished. The dyes, Mitotracker Orange, Mitotracker Green, Celltracker Orange, Celltracker Green, and monochlorobimane, are all selectively taken up by Müller glial cells, while neuronal cells remain largely devoid of the dyes. By using this method, it can be demonstrated that the free calcium alterations within the wholemounts indeed occur within Müller cells. Moreover, the cross‐sectional areas of (dye‐filled) Müller glial cell bodies, as well as of (dye‐free) neuronal cell bodies, can be measured in retinal wholemounts, and the spatial densities of both types of cells can be determined. The vital dye loading of Müller cells may facilitate investigations of stimulus‐induced alterations of retinal glial cell physiology and morphology.

Ocular inflammation alters swelling and membrane characteristics of rat Müller glial cells.

Ocular inflammation is a common cause of retinal edema that may involve swelling of Müller glial cells. In order to investigate whether endotoxin-induced ocular inflammation in rats alters the swelling and membrane characteristics of Müller cells, lipopolysaccharide (LPS; 0.5%) was intravitreally injected. At 3 and 7 days after treatment, hypotonic challenge induced swelling of Müller cell somata that was not observed in non-treated control eyes. Müller cells of LPS-treated eyes displayed a downregulation of inward K(+) currents and upregulation of A-type K(+) currents that was associated with a decreased expression of Kir4.1 protein in retinal slices. The data suggest that ocular inflammation induces alterations of both the swelling characteristics and the K(+) channel expression of Müller cells.

Altered membrane physiology in Müller glial cells after transient ischemia of the rat retina.

Inwardly rectifying K+ (Kir) channels have been implicated in the mediation of retinal K+ homeostasis by Müller glial cells. To assess possible involvement of altered glial K+ channel expression in ischemia‐reperfusion injury, transient retinal ischemia was induced in rat eyes. Acutely isolated Müller cells from postischemic retinae displayed a fast downregulation of their Kir currents, which began within 1 day and reached a maximum at 3 days of reperfusion, with a peak decrease to 20% as compared with control. This strong decrease of Kir currents was accompanied by an increase of the incidence of cells which displayed depolarization‐evoked fast transient (A‐type) K+ currents. While no cell from untreated control rats expressed A‐type K+ currents, all cells investigated from 3‐ and 7‐day postischemic retinae displayed such currents. An increased incidence of cells displaying fast transient Na+ currents was observed at 7 days after ischemia. These results suggest a role of altered glial Kir channel expression in postischemic neuronal degeneration via disturbance of retinal K+ siphoning.