scholarly journals Extracellular H+ fluxes from tiger salamander Müller (glial) cells measured using self-referencing H+-selective microelectrodes

2017 ◽  
Vol 118 (6) ◽  
pp. 3132-3143 ◽  
Author(s):  
Matthew A. Kreitzer ◽  
David Swygart ◽  
Meredith Osborn ◽  
Blair Skinner ◽  
Chad Heer ◽  
...  
Keyword(s):  
Glial Cells ◽  
Müller Cells ◽  
Bicarbonate Transport ◽  
Extracellular Potassium ◽  
Tiger Salamander ◽  
Muller Cells ◽  
Extracellular Acidification ◽  
Müller Glial Cells ◽  
Bicarbonate Transporter ◽  
Induced Changes

Self-referencing H+-selective electrodes were used to measure extracellular H+ fluxes from Müller (glial) cells isolated from the tiger salamander retina. A novel chamber enabled stable recordings using H+-selective microelectrodes in a self-referencing format using bicarbonate-based buffer solutions. A small basal H+ flux was observed from the end foot region of quiescent cells bathed in 24 mM bicarbonate-based solutions, and increasing extracellular potassium induced a dose-dependent increase in H+ flux. Barium at 6 mM also increased H+ flux. Potassium-induced extracellular acidifications were abolished when bicarbonate was replaced by 1 mM HEPES. The carbonic anhydrase antagonist benzolamide potentiated the potassium-induced extracellular acidification, while 300 μM DIDS, 300 μM SITS, and 30 μM S0859 significantly reduced the response. Potassium-induced extracellular acidifications persisted in solutions lacking extracellular calcium, although potassium-induced changes in intracellular calcium monitored with Oregon Green were abolished. Exchange of external sodium with choline also eliminated the potassium-induced extracellular acidification. Removal of extracellular sodium by itself induced a transient alkalinization, and replacement of sodium induced a transient acidification, both of which were blocked by 300 μM DIDS. Recordings at the apical portion of the cell showed smaller potassium-induced extracellular H+ fluxes, and removal of the end foot region further decreased the H+ flux, suggesting that the end foot was the major source of acidifications. These studies demonstrate that self-referencing H+-selective electrodes can be used to monitor H+ fluxes from retinal Müller cells in bicarbonate-based solutions and confirm the presence of a sodium-coupled bicarbonate transporter, the activity of which is largely restricted to the end foot of the cell. NEW & NOTEWORTHY The present study uses self-referencing H+-selective electrodes for the first time to measure H+ fluxes from Müller (glial) cells isolated from tiger salamander retina. These studies demonstrate bicarbonate transport as a potent regulator of extracellular levels of acidity around Müller cells and point toward a need for further studies aimed at addressing how such glial cell pH regulatory mechanisms may shape neuronal signaling.

scholarly journals 670nm photobiomodulation modulates bioenergetics and oxidative stress, in rat Müller cells challenged with high glucose

PLoS ONE ◽  
2021 ◽  
Vol 16 (12) ◽  
pp. e0260968
Author(s):  
Hannah J. Nonarath ◽  
Alexandria E. Hall ◽  
Gopika SenthilKumar ◽  
Betsy Abroe ◽  
Janis T. Eells ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Glial Cells ◽  
High Glucose ◽  
Near Infrared ◽  
Müller Cells ◽  
Model System ◽  
Light Treatment ◽  
Muller Cells ◽  
Müller Glial Cells

Diabetic retinopathy (DR), the most common complication of diabetes mellitus, is associated with oxidative stress, nuclear factor-κB (NFκB) activation, and excess production of vascular endothelial growth factor (VEGF) and intracellular adhesion molecule-1 (ICAM-1). Muller glial cells, spanning the entirety of the retina, are involved in DR inflammation. Mitigation of DR pathology currently occurs via invasive, frequently ineffective therapies which can cause adverse effects. The application of far-red to near-infrared (NIR) light (630-1000nm) reduces oxidative stress and inflammation in vitro and in vivo. Thus, we hypothesize that 670nm light treatment will diminish oxidative stress preventing downstream inflammatory mechanisms associated with DR initiated by Muller cells. In this study, we used an in vitro model system of rat Müller glial cells grown under normal (5 mM) or high (25 mM) glucose conditions and treated with a 670 nm light emitting diode array (LED) (4.5 J/cm2) or no light (sham) daily. We report that a single 670 nm light treatment diminished reactive oxygen species (ROS) production and preserved mitochondrial integrity in this in vitro model of early DR. Furthermore, treatment for 3 days in culture reduced NFκB activity to levels observed in normal glucose and prevented the subsequent increase in ICAM-1. The ability of 670nm light treatment to prevent early molecular changes in this in vitro high glucose model system suggests light treatment could mitigate early deleterious effects modulating inflammatory signaling and diminishing oxidative stress.

scholarly journals Metabolic Imbalance Effect on Retinal Müller Glial Cells Reprogramming Capacity: Involvement of Histone Deacetylase SIRT6

2021 ◽  
Vol 12 ◽  
Author(s):  
L Francisco Sanhueza Salas ◽  
Alfredo García-Venzor ◽  
Natalia Beltramone ◽  
Claudia Capurro ◽  
Debra Toiber ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Histone Deacetylase ◽  
Glial Cells ◽  
High Glucose ◽  
Müller Cells ◽  
Enrichment Analysis ◽  
Regulatory Elements ◽  
Muller Cells ◽  
Müller Glial Cells ◽  
And Oxidative Stress

Retinal Müller glial cells (MGs) are among the first to demonstrate metabolic changes during retinal disease and are a potential source of regenerative cells. In response to a harmful stimulus, they can dedifferentiate acquiring neural stem cells properties, proliferate and migrate to the damaged retinal layer and differentiate into lost neurons. However, it is not yet known how this reprogramming process is regulated in mammals. Since glucose and oxygen are important regulatory elements that may help directing stem cell fate, we aimed to study the effect of glucose variations and oxidative stress in Müller cells reprogramming capacity and analyze the participation the histone deacetylase SIRT6, as an epigenetic modulator of this process. We found that the combination of high glucose and oxidative stress induced a decrease in the levels of the marker glutamine synthetase, and an increase in the migration capacity of the cells suggesting that these experimental conditions could induce some degree of dedifferentiation and favor the migration ability. High glucose induced an increase in the levels of the pluripotent factor SOX9 and a decrease in SIRT6 levels accompanied by the increase in the acetylation levels of H3K9. Inhibiting SIRT6 expression by siRNA rendered an increase in SOX9 levels. We also determined SOX9 levels in retinas from mice with a conditional deletion of SIRT6 in the CNS. To further understand the mechanisms that regulate MGs response under metabolic impaired conditions, we evaluated the gene expression profile and performed Gene Ontology enrichment analysis of Müller cells from a murine model of Diabetes. We found several differentially expressed genes and observed that the transcriptomic change involved the enrichment of genes associated with glucose metabolism, cell migration, development and pluripotency. We found that many functional categories affected in cells of diabetic animals were directly related to SIRT6 function. Transcription factors enrichment analysis allowed us to predict several factors, including SOX9, that may be involved in the modulation of the differential expression program observed in diabetic MGs. Our results underline the heterogeneity of Müller cells response and the challenge that the study of metabolic impairment in vivo represents.

ATP-mediated Increase in H+ Flux from Retinal Müller Cells: a Role for Na+/H+ Exchange

2020 ◽  
Author(s):  
Boriana K Tchernookova ◽  
Michael W Gongwer ◽  
Alexis George ◽  
Brock Goeglein ◽  
Alyssa M Powell ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Neurotransmitter Release ◽  
Müller Cells ◽  
Cell Voltage ◽  
Extracellular Atp ◽  
Extracellular Potassium ◽  
Tiger Salamander ◽  
Synaptic Connections ◽  
Muller Cells ◽  
Extracellular Sodium

Small alterations in extracellular H+ can profoundly alter neurotransmitter release by neurons. We examined mechanisms by which extracellular ATP induces an extracellular H+ flux from Müller glial cells, which surround synaptic connections throughout the vertebrate retina. Müller glia were isolated from tiger salamander retinae and H+ fluxes examined using self-referencing H+-selective microelectrodes. Experiments were performed in 1 mM HEPES with no bicarbonate present. Replacement of extracellular sodium by choline decreased H+ efflux induced by 10 µM ATP by 75%. ATP-induced H+ efflux was also reduced by Na+/H+ exchange inhibitors. Amiloride reduced H+ efflux initiated by 10 µM ATP by 60%, while 10 µM cariporide decreased by 37%, and 25 µM zoniporide reduced H+ flux by 32%. ATP-induced H+ fluxes were not significantly altered by the K+/H+ pump blockers SCH28080 or TAK438, and replacement of all extracellular chloride with gluconate was without effect on H+ fluxes. Recordings of ATP-induced H+ efflux from cells simultaneously whole-cell voltage-clamped revealed no effect of membrane potential from -70 mV to 0 mV. Restoration of extracellular potassium after cells were bathed in 0 mM potassium produced a transient alteration in ATP-dependent H+ efflux. The transient response to extracellular potassium occurred only when extracellular sodium was present and was abolished by 1 mM ouabain, suggesting alterations in sodium gradients mediated by Na+/K ATPase activity. Our data indicate that the majority of H+ efflux elicited by extracellular ATP from isolated Müller cells is mediated by Na+/H+ exchange.

A role for aquaporin-4 in fluid regulation in the inner retina

2009 ◽  
Vol 26 (2) ◽  
pp. 159-165 ◽  
Author(s):  
MELINDA J. GOODYEAR ◽  
SHEILA G. CREWTHER ◽  
BARBARA M. JUNGHANS
Keyword(s):  
Water Transport ◽  
Glial Cells ◽  
Müller Cells ◽  
Extracellular Potassium ◽  
Muller Cells ◽  
Inner Retina ◽  
Aqp4 Expression ◽  
Kir Channels ◽  
Light Activity ◽  
Spatial Buffering

AbstractMany diverse retinal disorders are characterized by retinal edema; yet, little experimental attention has been given to understanding the fundamental mechanisms underlying and contributing to these fluid-based disorders. Water transport in and out of cells is achieved by specialized membrane channels, with most rapid water transport regulated by transmembrane water channels known as aquaporins (AQPs). The predominant AQP in the mammalian retina is AQP4, which is expressed on the Müller glial cells. Müller cells have previously been shown to modulate neuronal activity by modifying the concentrations of ions, neurotransmitters, and other neuroactive substances within the extracellular space between the inner and the outer limiting membrane. In doing so, Müller cells maintain extracellular homeostasis, especially with regard to the spatial buffering of extracellular potassium (K+) via inward rectifying K+ channels (Kir channels). Recent studies of water transport and the spatial buffering of K+ through glial cells have highlighted the involvement of both AQP4 and Kir channels in regulating the extracellular environment in the brain and retina. As both glial functions are associated with neuronal activation, controversy exists in the literature as to whether the relationship is functionally dependent. It is argued in this review that as AQP4 channels are likely to be the conduit for facilitating fluid homeostasis in the inner retina during light activation, AQP4 channels are also likely to play a consequent role in the regulation of ocular volume and growth. Recent research has already shown that the level of AQP4 expression is associated with environmentally driven manipulations of light activity on the retina and the development of myopia.

scholarly journals Physiological properties of retinal Müller glial cells from the cynomolgus monkey, Macaca fascicularis—a comparison to human Müller cells

2005 ◽  
Vol 45 (14) ◽  
pp. 1781-1791 ◽  
Author(s):  
Thomas Pannicke ◽  
Bernd Biedermann ◽  
Ortrud Uckermann ◽  
Michael Weick ◽  
Andreas Bringmann ◽  
...  
Keyword(s):  
Glial Cells ◽  
Cynomolgus Monkey ◽  
Macaca Fascicularis ◽  
Müller Cells ◽  
Muller Cells ◽  
Müller Glial Cells ◽  
Physiological Properties

scholarly journals Localization of αA-Crystallin in Rat Retinal Müller Glial Cells and Photoreceptors

2018 ◽  
Vol 24 (5) ◽  
pp. 545-552 ◽  
Author(s):  
Astrid Zayas-Santiago ◽  
David S. Ríos ◽  
Lidia V. Zueva ◽  
Mikhail Y. Inyushin
Keyword(s):  
Glial Cells ◽  
Light Transmission ◽  
Müller Cells ◽  
Cell Types ◽  
Confocal Imaging ◽  
Muller Cells ◽  
Müller Glial Cells ◽  
Fluorescence Confocal Imaging ◽  
Light Guides ◽  
Epithelium Cells

AbstractTransparent cells in the vertebrate optical tract, such as lens fiber cells and corneal epithelium cells, have specialized proteins that somehow permit only a low level of light scattering in their cytoplasm. It has been shown that both cell types contain (1) beaded intermediate filaments as well as (2) α-crystallin globulins. It is known that genetic and chemical alterations to these specialized proteins induce cytoplasmic opaqueness and visual complications. Crystallins were described previously in the retinal Müller cells of frogs. In the present work, using immunocytochemistry, fluorescence confocal imaging, and immuno-electron microscopy, we found that αA-crystallins are present in the cytoplasm of retinal Müller cells and in the photoreceptors of rats. Given that Müller glial cells were recently described as “living light guides” as were photoreceptors previously, we suggest that αA-crystallins, as in other highly transparent cells, allow Müller cells and photoreceptors to minimize intraretinal scattering during retinal light transmission.

scholarly journals Characteristics of Müller glial cells in MNU-induced retinal degeneration

2016 ◽  
Vol 33 ◽  
Author(s):  
MIRIAM REISENHOFER ◽  
THOMAS PANNICKE ◽  
ANDREAS REICHENBACH ◽  
VOLKER ENZMANN
Keyword(s):  
Glial Cells ◽  
Cell Activation ◽  
Ischemia Reperfusion ◽  
Outer Nuclear Layer ◽  
Müller Cells ◽  
Müller Cell ◽  
Reactive Gliosis ◽  
Muller Cells ◽  
Müller Glial Cells ◽  
Muller Cell

AbstractRetinal Müller glial cells have been shown to undergo reactive gliosis in a variety of retinal diseases. Upregulation of glial fibrillary acidic protein (GFAP) is a hallmark of Müller cell activation. Reactive gliosis after retinal detachment or ischemia/reperfusion is characterized by hypertrophy and downregulation of inwardly rectifying K+ (Kir) currents. However, this kind of physiological alteration could not be detected in slowly progressing retinal degenerations. The photoreceptor toxin N-methyl-N-nitrosourea (MNU) leads to the rapid loss of cells in the outer nuclear layer and subsequent Müller cell activation. Here, we investigated whether Müller cells from MNU-treated mice exhibit reactive gliosis. We found that Müller cells showed increased GFAP expression and increased membrane capacitance, indicating hypertrophy. Membrane potential and Kir channel-mediated K+ currents were not significantly altered whereas Kir4.1 mRNA expression and Kir-mediated inward current densities were markedly decreased. This suggests that MNU-induced Müller cell gliosis is characterized by plasma membrane increase without alteration in the membrane content of Kir channels. Taken together, our findings show that Müller cells of MNU-treated mice are reactive and respond with a form of gliosis which is characterized by cellular hypertrophy but no changes in Kir current amplitudes.

Müller Cell Ca2+ Waves Evoked by Purinergic Receptor Agonists in Slices of Rat Retina

2001 ◽  
Vol 85 (2) ◽  
pp. 986-994 ◽  
Author(s):  
Yang Li ◽  
Lynne A. Holtzclaw ◽  
James T. Russell
Keyword(s):  
Glial Cells ◽  
Purinergic Receptor ◽  
Rank Order ◽  
Müller Cells ◽  
Uridine Diphosphate ◽  
Muller Cells ◽  
Wave Amplification ◽  
Dye Loading ◽  
Intracellular Ca ◽  
Retinal Slices

We have measured agonist evoked Ca2+ waves in Müller cells in situ within freshly isolated retinal slices. Using an eye cup dye loading procedure we were able to preferentially fill Müller glial cells in retinal slices with calcium green. Fluorescence microscopy revealed that bath perfusion of slices with purinergic agonists elicits Ca2+ waves in Müller cells, which propagate along their processes. These Ca2+ signals were insensitive to tetrodotoxin (TTX, 1.0 μM) pretreatment. Cells were readily identified as Müller cells by their unique morphology and by subsequent immunocytochemical labeling with glial fibrillary acidic protein antibodies. While cells never exhibited spontaneous Ca2+ oscillations, purinoreceptor agonists, ATP, 2 MeSATP, ADP, 2 MeSADP, and adenosine readily elicited Ca2+ waves. These waves persisted in the absence of [Ca2+]o but were abolished by thapsigargin pretreatment, suggesting that the purinergic agonists tested act by releasing Ca2+ from intracellular Ca2+ stores. The rank order of potency of different purines and pyrimidines for inducing Ca2+ signals was 2 MeSATP = 2MeSADP > ADP > ATP ≫ αβmeATP = uridine triphosphate (UTP) > uridine diphosphate (UDP). The Ca2+signals evoked by ATP, ADP, and 2 MeSATP were inhibited by reactive blue (100 μM) and suramin (200 μM), and the adenosine induced signals were abolished only by 3,7-dimethyl-1-propargylxanthine (200 μM) and not by 1,3-dipropyl-8-(2-amino-4-chlorophenyl)-xanthine) or 8-cyclopentyl-1,3-dipropylxanthine at the same concentration. Based on these pharmacological characteristics and the dose-response relationships for ATP, 2 MeSATP, 2 MeSADP, ADP, and adenosine, we concluded that Müller cells express the P1A2 and P2Y1 subtypes of purinoceptors. Analysis of Ca2+ responses showed that, similar to glial cells in culture, wave propagation occurred by regenerative amplification at specialized Ca2+ release sites (wave amplification sites), where the rate of Ca2+ release was significantly enhanced. These data suggest that Müller cells in the retina may participate in signaling, and this may serve as an extra-neuronal signaling pathway.

scholarly journals A dialogue between glia and neurons in the retina: modulation of neuronal excitability

2004 ◽  
Vol 1 (3) ◽  
pp. 245-252 ◽  
Author(s):  
ERIC A. NEWMAN
Keyword(s):  
Synaptic Transmission ◽  
Glial Cells ◽  
Neuronal Excitability ◽  
Ganglion Cells ◽  
Brain Slices ◽  
Müller Cells ◽  
Muller Cells ◽  
Signaling Mechanisms ◽  
Bidirectional Signaling ◽  
Stimulation Of

Bidirectional signaling between neurons and glial cells has been demonstrated in brain slices and is believed to mediate glial modulation of synaptic transmission in the CNS. Our laboratory has characterized similar neuron–glia signaling in the mammalian retina. We find that light-evoked neuronal activity elicits Ca2+ increases in Müller cells, which are specialized retinal glial cells. Neuron to glia signaling is likely mediated by the release of ATP from neurons and is potentiated by adenosine. Glia to neuron signaling has also been observed and is mediated by several mechanisms. Stimulation of glial cells can result in either facilitation or depression of synaptic transmission. Release of D-serine from Müller cells might also potentiate NMDA receptor transmission. Müller cells directly inhibit ganglion cells by releasing ATP, which, following hydrolysis to adenosine, activates neuronal A1 receptors. The existence of bidirectional signaling mechanisms indicates that glial cells participate in information processing in the retina.

scholarly journals Glial cell regulation of neuronal activity and blood flow in the retina by release of gliotransmitters

2015 ◽  
Vol 370 (1672) ◽  
pp. 20140195 ◽  
Author(s):  
Eric A. Newman
Keyword(s):  
Blood Flow ◽  
Arachidonic Acid ◽  
Glial Cells ◽  
Neuronal Activity ◽  
Neurovascular Coupling ◽  
Müller Cells ◽  
Future Research ◽  
Prostaglandin E ◽  
Sk Channels ◽  
Muller Cells

Astrocytes in the brain release transmitters that actively modulate neuronal excitability and synaptic efficacy. Astrocytes also release vasoactive agents that contribute to neurovascular coupling. As reviewed in this article, Müller cells, the principal retinal glial cells, modulate neuronal activity and blood flow in the retina. Stimulated Müller cells release ATP which, following its conversion to adenosine by ectoenzymes, hyperpolarizes retinal ganglion cells by activation of A1 adenosine receptors. This results in the opening of G protein-coupled inwardly rectifying potassium (GIRK) channels and small conductance Ca 2+ -activated K + (SK) channels. Tonic release of ATP also contributes to the generation of tone in the retinal vasculature by activation of P2X receptors on vascular smooth muscle cells. Vascular tone is lost when glial cells are poisoned with the gliotoxin fluorocitrate. The glial release of vasoactive metabolites of arachidonic acid, including prostaglandin E 2 (PGE 2 ) and epoxyeicosatrienoic acids (EETs), contributes to neurovascular coupling in the retina. Neurovascular coupling is reduced when neuronal stimulation of glial cells is interrupted and when the synthesis of arachidonic acid metabolites is blocked. Neurovascular coupling is compromised in diabetic retinopathy owing to the loss of glial-mediated vasodilation. This loss can be reversed by inhibiting inducible nitric oxide synthase. It is likely that future research will reveal additional important functions of the release of transmitters from glial cells.

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