A Novel and Atypical Nf-Kb Proinflammatory Program Regulated by A Camkii-Proteasome Axis Is Involved in the Early Activation of Muller Glia by High Glucose

Abstract BackgroundDiabetic retinopathy (DR) is a microvascular complication of diabetes with a heavy impact on the life-quality of subjects and with a dramatic burden for health and economic systems on a global scale.Although the pathogenesis of DR is largely unknown, several preclinical data have pointed out to a main role of Muller glia, a cell type which spans across the retina layers providing nourishment and support for Retina Ganglion Cells (RGCs), in sensing glycemia and in acquiring a proinflammatory polarization in response to this insult.ResultsBy using a validated experimental model of DR in vitro, the rMC1 cells challenged with high glucose, we uncovered the induction of an early (within minutes) and atypical NF-kB signalling pathway regulated by a CamKII-proteasome axis. Phosphorylation of proteasome subunit Rpt6 (at serine 120) by CamKII stimulated the accelerated turnover of IkBα (i.e., the natural inhibitor of p65-50 transcription factor), regardless of the phosphorylation at serine 32 which labels canonical NF-kB signalling. This event allowed the p65-p50 heterodimer to migrate into the nucleus and to induce the selective transcription of IL-8, Il-1β and MCP-1. Pharmacological inhibition of CamKII or proteasome stopped this proinflammatory program, whereas introduction of a Rpt6 phospho-dead mutant (Rpt6-S120A) stimulated a paradoxical effect on NF-kB probably through the activation of a compensatory mechanism which may involve phosphorylation of 20S α4 subunit.ConclusionsThis study introduces a novel pathway of MG activation by high glucose and casts some light on the biological relevance of proteasome post-translational modifications in modulating pathways regulated through targeted proteolysis.

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Pressure-dependent modulation of inward-rectifying K+ channels: implications for cation homeostasis and K+ dynamics in glaucoma

AJP Cell Physiology ◽

10.1152/ajpcell.00444.2018 ◽

2019 ◽

Vol 317 (2) ◽

pp. C375-C389

Author(s):

Rachel A. Fischer ◽

Abigail L. Roux ◽

Lauren K. Wareham ◽

Rebecca M. Sappington

Keyword(s):

Ganglion Cells ◽

Primary Cultures ◽

Elevated Pressure ◽

Muller Glia ◽

Müller Glia ◽

Cation Homeostasis ◽

K2p Channels ◽

And Function

Glaucoma is the leading cause of blindness worldwide, resulting from degeneration of retinal ganglion cells (RGCs), which form the optic nerve. Prior to structural degeneration, RGCs exhibit physiological deficits. Müller glia provide homeostatic regulation of ions that supports RGC physiology through a process called K+ siphoning. Recent studies suggest that several retinal conditions, including glaucoma, involve changes in the expression of K+ channels in Müller glia. To clarify whether glaucoma-related stressors directly alter expression and function of K+ channels in Müller glia, we examined changes in the expression of inwardly rectifying K+ (Kir) channels and two-pore domain (K2P) channels in response to elevated intraocular pressure (IOP) in vivo and in vitro in primary cultures of Müller glia exposed to elevated hydrostatic pressure. We then measured outcomes of cell health, cation homeostasis, and cation flux in Müller glia cultures. Transcriptome analysis in a murine model of microbead-induced glaucoma revealed pressure-dependent downregulation of Kir and K2P channels in vivo. Changes in the expression and localization of Kir and K2P channels in response to elevated pressure were also found in Müller glia in vitro. Finally, we found that elevated pressure compromises the plasma membrane of Müller glia and induces cation dyshomeostasis that involves changes in ion flux through cation channels. Pressure-induced changes in cation flux precede both cation dyshomeostasis and membrane compromise. Our findings have implications for Müller glia responses to pressure-related conditions, i.e., glaucoma, and identify cation dyshomeostasis as a potential contributor to electrophysiological impairment observed in RGCs of glaucomatous retina.

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Dexamethasone protects retinal ganglion cells but not Müller glia against hyperglycemia in vitro

PLoS ONE ◽

10.1371/journal.pone.0207913 ◽

2018 ◽

Vol 13 (11) ◽

pp. e0207913 ◽

Cited By ~ 7

Author(s):

Xandra Pereiro ◽

Noelia Ruzafa ◽

Arantxa Acera ◽

Alex Fonollosa ◽

F. David Rodriguez ◽

...

Keyword(s):

Retinal Ganglion Cells ◽

Ganglion Cells ◽

Retinal Ganglion ◽

Muller Glia ◽

Müller Glia

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Comparative lipidomic analysis of mammalian retinal ganglion cells and Müller glia in situ and in vitro using High-Resolution Imaging Mass Spectrometry

Scientific Reports ◽

10.1038/s41598-020-77087-x ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Xandra Pereiro ◽

Roberto Fernández ◽

Gabriel Barreda-Gómez ◽

Noelia Ruzafa ◽

Arantxa Acera ◽

...

Keyword(s):

Retinal Ganglion Cells ◽

Ganglion Cells ◽

Imaging Mass Spectrometry ◽

Cell Types ◽

Retinal Ganglion ◽

Muller Glia ◽

Müller Glia ◽

Ocular Diseases

AbstractIn order to better understand retinal physiology, alterations to which underlie some ocular diseases, we set out to establish the lipid signature of two fundamental cell types in the retina, Müller Glia and Retinal Ganglion Cells (RGCs). Moreover, we compared the lipid signature of these cells in sections (in situ), as well as after culturing the cells and isolating their cell membranes (in vitro). The lipidome of Müller glia and RGCs was analyzed in porcine retinal sections using Matrix Assisted Laser Desorption Ionization Imaging Mass Spectrometry (MALDI-IMS). Isolated membranes, as well as whole cells from primary cell cultures of RGCs and Müller glia, were printed onto glass slides using a non-contact microarrayer (Nano Plotter), and a LTQ-Orbitrap XL analyzer was used to scan the samples in negative ion mode, thereafter identifying the RGCs and Müller cells immunohistochemically. The spectra acquired were aligned and normalized against the total ion current, and a statistical analysis was carried out to select the lipids specific to each cell type in the retinal sections and microarrays. The peaks of interest were identified by MS/MS analysis. A cluster analysis of the MS spectra obtained from the retinal sections identified regions containing RGCs and Müller glia, as confirmed by immunohistochemistry in the same sections. The relative density of certain lipids differed significantly (p-value ≤ 0.05) between the areas containing Müller glia and RGCs. Likewise, different densities of lipids were evident between the RGC and Müller glia cultures in vitro. Finally, a comparative analysis of the lipid profiles in the retinal sections and microarrays identified six peaks that corresponded to a collection of 10 lipids characteristic of retinal cells. These lipids were identified by MS/MS. The analyses performed on the RGC layer of the retina, on RGCs in culture and using cell membrane microarrays of RGCs indicate that the lipid composition of the retina detected in sections is preserved in primary cell cultures. Specific lipid species were found in RGCs and Müller glia, allowing both cell types to be identified by a lipid fingerprint. Further studies into these specific lipids and of their behavior in pathological conditions may well help identify novel therapeutic targets for ocular diseases.

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Muller glia induce retinal progenitor cells to differentiate into retinal ganglion cells

Neuroreport ◽

10.1097/01.wnr.0000227991.23046.b7 ◽

2006 ◽

Vol 17 (12) ◽

pp. 1263-1267 ◽

Cited By ~ 3

Author(s):

Jing Yao ◽

Xinghuai Sun ◽

Yang Wang ◽

Ling Wang

Keyword(s):

Progenitor Cells ◽

Retinal Ganglion Cells ◽

Ganglion Cells ◽

Retinal Ganglion ◽

Muller Glia ◽

Müller Glia ◽

Retinal Progenitor Cells ◽

Retinal Progenitor

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THE CHANGES IN DISTRIBUTION OF NMDA- AND AMPA-RECEPTORS IN RETINOCOLLICULAR SYNAPSES

Fiziolohichnyĭ zhurnal ◽

10.15407/fz67.03.010 ◽

2021 ◽

Vol 67 (3) ◽

pp. 10-16

Author(s):

H.V. Dumanska ◽

◽

O.V. Rikchalsky ◽

N.S. Veselovsky ◽

◽

...

Keyword(s):

Ampa Receptors ◽

In Vitro Model ◽

Ganglion Cells ◽

Signal Transmission ◽

Main Role ◽

Retinal Ganglion ◽

Dynamic Changes ◽

Functional Roles ◽

Vitro Model

We investigated the changes in distribution of the NMDA- and AMPA-receptors in the synapses in the in vitro model of the retinocollicular pathway. The model was a primary coculture of the retinal cells and superficial superior colliculus (SSC) neurons. Evoked postsynaptic currents (ePSCs) were recorded in SSC neurons in response to local electrical extracellular stimulation of the afferent retinal ganglion cells’ (RGC) axons. We analyzed the changes in the kinetic characteristics of ePSCs at different holding potentials during cocultivation. The results obtained reflect that NMDA receptors play an essential role in the formation of the retinocollicular synapses. After the formation of retinocollicular connections, the main role in the sensory signal transmission belongs to AMPA receptors. Thus, the data obtained indicate the specific dynamic changes of the functional roles of NMDA- and AMPA-receptors in the forma- tion and development of the retinocollicular synaptic contacts.

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Proteomic profiling of primary retinal Müller glia cells reveals a shift in expression patterns upon adaptation to in vitro conditions

Glia ◽

10.1002/glia.10292 ◽

2003 ◽

Vol 44 (3) ◽

pp. 251-263 ◽

Cited By ~ 65

Author(s):

Stefanie M. Hauck ◽

Sabine Suppmann ◽

Marius Ueffing

Keyword(s):

Expression Patterns ◽

Muller Glia ◽

Müller Glia ◽

Proteomic Profiling ◽

Glia Cells

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Regulation of cilia abundance in multiciliated cells

eLife ◽

10.7554/elife.44039 ◽

2019 ◽

Vol 8 ◽

Cited By ~ 18

Author(s):

Rashmi Nanjundappa ◽

Dong Kong ◽

Kyuhwan Shim ◽

Tim Stearns ◽

Steven L Brody ◽

...

Keyword(s):

Progenitor Cells ◽

Surface Area ◽

Ex Vivo ◽

Compensatory Mechanism ◽

Multiciliated Cells ◽

Culture Model ◽

A Cell ◽

Dependent Mechanism

Multiciliated cells (MCC) contain hundreds of motile cilia used to propel fluid over their surface. To template these cilia, each MCC produces between 100-600 centrioles by a process termed centriole amplification. Yet, how MCC regulate the precise number of centrioles and cilia remains unknown. Airway progenitor cells contain two parental centrioles (PC) and form structures called deuterosomes that nucleate centrioles during amplification. Using an ex vivo airway culture model, we show that ablation of PC does not perturb deuterosome formation and centriole amplification. In contrast, loss of PC caused an increase in deuterosome and centriole abundance, highlighting the presence of a compensatory mechanism. Quantification of centriole abundance in vitro and in vivo identified a linear relationship between surface area and centriole number. By manipulating cell size, we discovered that centriole number scales with surface area. Our results demonstrate that a cell-intrinsic surface area-dependent mechanism controls centriole and cilia abundance in multiciliated cells.

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Characterization of retinal regeneration in adult zebrafish following multiple rounds of phototoxic lesion

PeerJ ◽

10.7717/peerj.5646 ◽

2018 ◽

Vol 6 ◽

pp. e5646 ◽

Cited By ~ 7

Author(s):

Alexandra H. Ranski ◽

Ashley C. Kramer ◽

Gregory W. Morgan ◽

Jennifer L. Perez ◽

Ryan Thummel

Keyword(s):

Cell Cycle ◽

Cellular Localization ◽

Ganglion Cells ◽

Plexiform Layer ◽

Inner Plexiform Layer ◽

Muller Glia ◽

Müller Glia ◽

Light Damage ◽

Retinal Regeneration ◽

Adult Zebrafish

Müller glia in the zebrafish retina respond to retinal damage by re-entering the cell cycle, which generates large numbers of retinal progenitors that ultimately replace the lost neurons. In this study we compared the regenerative outcomes of adult zebrafish exposed to one round of phototoxic treatment with adult zebrafish exposed to six consecutive rounds of phototoxic treatment. We observed that Müller glia continued to re-enter the cell cycle to produce clusters of retinal progenitors in zebrafish exposed to multiple rounds of phototoxic light. Some abnormalities were noted, however. First, we found that retinas exposed to multiple rounds of damage exhibited a greater loss of photoreceptors at 36 hours of light damage than retinas that were exposed to their first round of light damage. In addition, we found that Müller glia appeared to have an increase in the acute gliotic response in retinas exposed to multiple rounds of light treatment. This was evidenced by cellular hypertrophy, changes in GFAP cellular localization, and transient increases in stat3 and gfap expression. Finally, following the sixth round of phototoxic lesion, we observed a significant increase in mis-localized HuC/D-positive amacrine and ganglion cells in the inner plexiform layer and outer retina, and a decreased number of regenerated blue cone photoreceptors. These data add to recent findings that retinal regeneration in adult zebrafish occurs concomitant with Müller glia reactivity and can result in the generation of aberrant neurons. These data are also the first to demonstrate that Müller glia appear to modify their phenotype in response to multiple rounds of phototoxic lesion, exhibiting an increase in acute gliosis while maintaining a remarkable capacity for long-term regeneration of photoreceptors.

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Action Potentials in the Dendrites of Retinal Ganglion Cells

Journal of Neurophysiology ◽

10.1152/jn.1999.81.3.1412 ◽

1999 ◽

Vol 81 (3) ◽

pp. 1412-1417 ◽

Cited By ~ 52

Author(s):

Toby J. Velte ◽

Richard H. Masland

Keyword(s):

Retinal Ganglion Cells ◽

Action Potentials ◽

Ganglion Cells ◽

Retinal Ganglion ◽

Muller Glia ◽

Müller Glia ◽

Whole Cell ◽

The Absolute ◽

Somatic Action Potentials ◽

Site Of Stimulation

Action potentials in the dendrites of retinal ganglion cells. The somas and dendrites of intact retinal ganglion cells were exposed by enzymatic removal of the overlying endfeet of the Müller glia. Simultaneous whole cell patch recordings were made from a ganglion cell’s dendrite and the cell’s soma. When a dendrite was stimulated with depolarizing current, impulses often propagated to the soma, where they appeared as a mixture of small depolarizations and action potentials. When the soma was stimulated, action potentials always propagated back through the dendrite. The site of initiation of action potentials, as judged by their timing, could be shifted between soma and dendrite by changing the site of stimulation. Applying QX-314 to the soma could eliminate somatic action potentials while leaving dendritic impulses intact. The absolute amplitudes of the dendritic action potentials varied somewhat at different distances from the soma, and it is not clear whether these variations are real or technical. Nonetheless, the qualitative experiments clearly suggest that the dendrites of retinal ganglion cells generate regenerative Na+ action potentials, at least in response to large direct depolarizations.

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