Involvement of P2X<sub>7</sub>Receptors in the Hypoxia-Induced Death of Rat Retinal Neurons

Sugiyama, Tetsuya; Oku, Hidehiro; Shibata, Maho; Fukuhara, Masayuki; Yoshida, Hideyo; Ikeda, Tsunehiko

doi:10.1167/iovs.09-4192

Cited by 45 publications

(35 citation statements)

References 41 publications

(32 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…P2X 7 receptors expressed on RGCs are essential for modulating their synaptic responses [133]. While a short term stimulation of P2X 7 receptors induces physiological calcium response, prolonged stimulation is shown to mediate RGC death [134]. Under hypoxic/ ischemic conditions, activation of P2X receptors, specifically the P2X 7 receptor, has been reported to cause death of RGCs [134,135] by enhancing intracellular calcium levels.…”

Section: Hypoxia and Purinergic Receptorsmentioning

confidence: 99%

“…While a short term stimulation of P2X 7 receptors induces physiological calcium response, prolonged stimulation is shown to mediate RGC death [134]. Under hypoxic/ ischemic conditions, activation of P2X receptors, specifically the P2X 7 receptor, has been reported to cause death of RGCs [134,135] by enhancing intracellular calcium levels. However, inhibition of P2X 7 receptor prevented the death of RGCs following ischemic insult [135].…”

Section: Hypoxia and Purinergic Receptorsmentioning

confidence: 99%

See 1 more Smart Citation

Cellular and Molecular Mechanisms of Retinal Ganglion Cell Death in Hypoxic-Ischemic Injuries

Kaur¹,

Rathnasamy²,

Foulds³

et al. 2015

J Neurol Exp Neurosci

View full text Add to dashboard Cite

Loss of retinal ganglion cells (RGCs) occurs in retinal degenerative diseases, such as glaucoma, age-related macular degeneration, diabetic retinopathy, central retinal artery occlusion and ischemic central retinal vein thrombosis in adults and in retinopathy of prematurity in infants. A critical role of hypoxia, which underlies most of the above disorders, has been reported in causing RGC death. Disruption of blood-retinal barrier (BRB) occurs due to increased production of vascular endothelial growth factor and nitric oxide in response to hypoxia resulting in extravasation of serum derived substances and retinal edema and this may be one of important factors mediating RGC death. Activation of microglial cells in response to hypoxia and subsequent increased release of proinflammatory cytokines by them such as tumor-necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) target the TNF-receptor 1 and IL-receptor 1 on RGCs. Excess release of glutamate in retinal tissue under hypoxia activates the ionotropic glutamate receptors causing excitotoxicity through increased influx of calcium into the RGCs. Increased production of TNF-α, IL-1β and enhanced intracellular calcium results in RGC death through activating several pathways such as caspase signaling, mitochondrial dysfunction and oxidative stress. In our investigations, melatonin, an antioxidant, was shown to reduce RGC death in the adult and neonatal hypoxic retina, through suppression of TNF-α, IL-1β and glutamate levels as well as oxidative stress. Moreover, the function and structure of BRB was well preserved in these animals along with reduction in retinal edema. In view of our observations, we suggest that melatonin could be considered as a therapeutic agent to reduce RGC death in various retinal pathologies, in addition to other potential drugs/molecules of therapeutic interest that could render protection to RGCs. However, future in-depth research on the effects of melatonin on the retina under hypoxic conditions needs to be undertaken to explore its full potential in preventing/ameliorating RGC death. Factors Mediating RGC DeathSeveral factors induced or upregulated by hypoxia may be involved in causing RGC death. These factors include, among others, disruption of the blood-retinal barrier, activation of microglial cells and the accompanying enhanced release of inflammatory cytokines and excess release and accumulation of glutamate in the retinal tissues. Hypoxia and the blood-retinal barrierThe BRB regulates the passage of molecules from blood into the retina [11,12]. It consists of an inner BRB formed by the tight junctions (TJ) between capillary endothelial cells and an outer barrier formed by TJ between retinal pigment epithelial cells [12][13][14][15][16]. The TJ are made up of transmembrane proteins such as occludin, claudin-5 and cytoplasmic proteins such as zona occludens (ZO)-1 that are structurally and functionally important for maintaining the integrity of the BRB. The proper functioning of the inner BRB also requires the collective co...

show abstract

Section: Hypoxia and Purinergic Receptorsmentioning

confidence: 99%

Section: Hypoxia and Purinergic Receptorsmentioning

confidence: 99%

Cellular and Molecular Mechanisms of Retinal Ganglion Cell Death in Hypoxic-Ischemic Injuries

Kaur¹,

Rathnasamy²,

Foulds³

et al. 2015

J Neurol Exp Neurosci

View full text Add to dashboard Cite

show abstract

“…Zhang et al [34] reported that activation of P2X7 receptors induces a time-dependent death of ganglion cells in rat retinal cultures. Moreover, Sugiyama et al [48] have shown that P2X7 receptor antagonists are able to inhibit the death of rat retinal neurons in purified cultures. Here, we showed that a decrease in cell viability induced by ATP or BzATP was obtained only in mixed cultures containing glial cells.…”

Section: Discussionmentioning

confidence: 99%

ATP induces the death of developing avian retinal neurons in culture via activation of P2X7 and glutamate receptors

Anccasi

Ornelas

Cossenza

et al. 2012

Purinergic Signalling

View full text Add to dashboard Cite

Previous data suggest that nucleotides are important mitogens in the developing retina. Here, the effect of ATP on the death of cultured chick embryo retina cells was investigated. In cultures obtained from retinas of 7-day-old chick embryos (E7) that were cultivated for 2 days (E7C2), both ATP and BzATP induced a ∼30 % decrease in cell viability that was time-and dose-dependent and that could be blocked by 0.2 mM oxidized ATP or 0.3 μM KN-62. An increase in cleaved caspase-3 levels and in the number of TUNELpositive cells was observed when cultures were incubated with 3 mM ATP and immunolabeling for cleaved-caspase 3 was observed over neurons but not over glial cells. ATPdependent cell death was developmentally regulated, the maximal levels being detected by E7C2-3. Nucleotides were able to increase neuronal ethidium bromide and sulforhodamine B uptake in mixed and purified neuronal cultures, an effect that was blocked by the antagonists Brilliant Blue G and oxidized ATP. In contrast, nucleotide-induced cell death was observed only in mixed cultures, but not in purified cultures of neurons or glia. ATP-induced neuronal death was blocked by the glutamatergic antagonists MK801 and DNQX and activation of P2X7 receptors by ATP decreased the uptake of [ These results suggest that ATP induces apoptosis of chick embryo retinal neurons in culture through activation of P2X7 and glutamate ionotropic receptors. Involvement of a P2X7 receptor-mediated inhibition of the glial uptake of glutamate is suggested.

show abstract

“…The procedure can be divided into three main parts: preparation of samples (Steps 1-15), preparation of stains and sample staining (Steps [16][17]; and data acquisition by microplate cytometry and data analysis (Steps 18-19).…”

Section: Experimental Designmentioning

confidence: 99%

“…Brief stimulation of P2X7 receptor leads to an increase in intracellular calcium influx whereas repeated or prolonged (> 15 min) stimulation of P2X7 receptor induces the formation of a non-selective pore. This pore allows the entry of solutes up to 900 Da in size, which eventually leads to membrane blebbing, release of cytokines and activation of caspases ultimately leading to cell death [14][15][16][17][18][19]. P2X7 receptors activation is responsible for a large number of How to cite this article: Rat P, Olivier E, Tanter C, Wakx A, Dutot M. A fast and reproducible cell-and 96-well plate-based method for the evaluation of P2X7 receptor activation using YO-PRO-1 fluorescent dye.…”

mentioning

confidence: 99%

A fast and reproducible cell- and 96-well plate-based method for the evaluation of P2X7 receptor activation using YO-PRO-1 fluorescent dye

Rat

Olivier

Tanter³

et al. 2017

J Biol Methods

View full text Add to dashboard Cite

A B S T R A C TThe YO-PRO-1 assay provides a quantitative estimation of P2X7 receptor activation. P2X7 receptor is associated to pathological conditions including infectious, inflammatory, neurological, musculoskeletal disorders, pain and cancer. Most primary cells and cell lines from diverse origin may be used thanks to the ubiquitous distribution of P2X7 receptor. To study the activation of P2X7 receptor by chemicals or biological agents, we established a microplate-based cytometry protocol to accurately and rapidly quantify the activation of P2X7 receptor that leads to the formation of large pores in cell membranes. The YO-PRO-1 assay is based on the ability of cells to incorporate and bind YO-PRO-1 dye to DNA after activation of P2X7 receptor through pore formation. Cells are seeded in 96-well plates and incubated with the compound being tested for the appropriate time. The microplate is then incubated for 10 min with YO-PRO-1 staining solution. After the 10 min staining time, fluorescence signal is read using a microplate reader in 1 min. This procedure is easier and requires less handling steps than flow cytometry. 96-well plate based YO-PRO-1 assay is a reproducible and fast method to study both P2X7 receptor activation by toxic agents at subnecrotic concentrations and P2X7 receptor inhibition by antagonists.

show abstract

Involvement of P2X₇Receptors in the Hypoxia-Induced Death of Rat Retinal Neurons

Abstract: The results suggest that the death of retinal neurons can be triggered by hypoxia and that P2X(7) activation is involved in the hypoxia-induced death of retinal neurons. P2X(7) antagonists can prevent hypoxia-induced damage in retinal neurons.

Cited by 45 publications

References 41 publications

Cellular and Molecular Mechanisms of Retinal Ganglion Cell Death in Hypoxic-Ischemic Injuries

Cellular and Molecular Mechanisms of Retinal Ganglion Cell Death in Hypoxic-Ischemic Injuries

ATP induces the death of developing avian retinal neurons in culture via activation of P2X7 and glutamate receptors

A fast and reproducible cell- and 96-well plate-based method for the evaluation of P2X7 receptor activation using YO-PRO-1 fluorescent dye

Contact Info

Product

Resources

About

Involvement of P2X7Receptors in the Hypoxia-Induced Death of Rat Retinal Neurons

Abstract: The results suggest that the death of retinal neurons can be triggered by hypoxia and that P2X(7) activation is involved in the hypoxia-induced death of retinal neurons. P2X(7) antagonists can prevent hypoxia-induced damage in retinal neurons.

Cited by 45 publications

References 41 publications

Cellular and Molecular Mechanisms of Retinal Ganglion Cell Death in Hypoxic-Ischemic Injuries

Cellular and Molecular Mechanisms of Retinal Ganglion Cell Death in Hypoxic-Ischemic Injuries

ATP induces the death of developing avian retinal neurons in culture via activation of P2X7 and glutamate receptors

A fast and reproducible cell- and 96-well plate-based method for the evaluation of P2X7 receptor activation using YO-PRO-1 fluorescent dye

Contact Info

Product

Resources

About

Involvement of P2X₇Receptors in the Hypoxia-Induced Death of Rat Retinal Neurons