Sigma-1 receptor stimulation protects retinal ganglion cells from ischemia-like insult through the activation of extracellular-signal-regulated kinases 1/2

Brett H. Mueller, Yong Park, Hai Ying Ma, Adnan Dibas, Dorette Zoe Ellis, Abbot Clark, Thomas Yorio

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Abstract

Sigma-1 receptor (σ-1) activation and mitogen-activated protein kinases (MAPKs) have been shown to protect retinal ganglion cells (RGCs) from cell death. The purpose of this study was to determine if σ-1 receptor stimulation with pentazocine could promote neuroprotection under conditions of an ischemia-like insult (oxygen glucose deprivation (OGD)) through the phosphorylation of extracellular signal regulated kinase (pERK)1/2. Primary RGCs were isolated from P3-P7 Sprague-Dawley rats and purified by sequential immunopanning using Thy1.1 antibodies. RGCs were cultured for 7 days before subjecting the cells to an OGD insult (0.5% oxygen in glucose-free medium) for 6h. During the OGD, RGCs were treated with pentazocine (σ-1 receptor agonist) with or without BD 1047 (σ-1 receptor antagonist). In other experiments, primary RGCs were treated with pentazocine in the presence or absence of an MEK1/2 inhibitor, PD098059. Cell survival/death was assessed by staining with the calcein-AM/ethidium homodimer reagent. Levels of pERK1/2, total ERK1/2, and beta tubulin expression were determined by immunoblotting and immunofluorescence staining. RGCs subjected to OGD for 6h induced 50% cell death in primary RGCs (. p<0.001) and inhibited pERK1/2 expression by 65% (. p<0.001). Cell death was attenuated when RGCs were treated with pentazocine under OGD (. p<0.001) and pERK1/2 expression was increased by 1.6 fold (. p<0.05) compared to OGD treated RGCs without pentazocine treatment. The co-treatment of PD098059 (MEK1/2 inhibitor) with pentazocine significantly abolished the protective effects of pentazocine on the RGCs during this OGD insult. Activation of the σ-1 receptor is a neuroprotective target that can protect RGCs from an ischemia-like insult. These results also established a direct relationship between σ-1 receptor stimulation and the neuroprotective effects of the ERK1/2 pathway in purified RGCs subjected to OGD. These findings suggest that activation of the σ-1 receptor may be a therapeutic target for neuroprotection particularly relevant to ocular neurodegenerative diseases that effect RGCs.

Original languageEnglish
Pages (from-to)156-169
Number of pages14
JournalExperimental Eye Research
Volume128
DOIs
StatePublished - 1 Nov 2014

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Mitogen-Activated Protein Kinase 3
Retinal Ganglion Cells
Mitogen-Activated Protein Kinase 1
Ischemia
Pentazocine
Oxygen
Glucose
Cell Death
sigma-1 receptor
Staining and Labeling
Eye Diseases
MAP Kinase Signaling System
Neuroprotective Agents
Tubulin
Mitogen-Activated Protein Kinases
Immunoblotting
Neurodegenerative Diseases
Fluorescent Antibody Technique
Sprague Dawley Rats
Cell Survival

Keywords

  • Extracellular signal-regulated kinases 1/2
  • Neuroprotection
  • Primary retinal ganglion cells
  • Sigma-1 receptor

Cite this

@article{2b780d5ac5cb409e9c9e6c874dc570fb,
title = "Sigma-1 receptor stimulation protects retinal ganglion cells from ischemia-like insult through the activation of extracellular-signal-regulated kinases 1/2",
abstract = "Sigma-1 receptor (σ-1) activation and mitogen-activated protein kinases (MAPKs) have been shown to protect retinal ganglion cells (RGCs) from cell death. The purpose of this study was to determine if σ-1 receptor stimulation with pentazocine could promote neuroprotection under conditions of an ischemia-like insult (oxygen glucose deprivation (OGD)) through the phosphorylation of extracellular signal regulated kinase (pERK)1/2. Primary RGCs were isolated from P3-P7 Sprague-Dawley rats and purified by sequential immunopanning using Thy1.1 antibodies. RGCs were cultured for 7 days before subjecting the cells to an OGD insult (0.5{\%} oxygen in glucose-free medium) for 6h. During the OGD, RGCs were treated with pentazocine (σ-1 receptor agonist) with or without BD 1047 (σ-1 receptor antagonist). In other experiments, primary RGCs were treated with pentazocine in the presence or absence of an MEK1/2 inhibitor, PD098059. Cell survival/death was assessed by staining with the calcein-AM/ethidium homodimer reagent. Levels of pERK1/2, total ERK1/2, and beta tubulin expression were determined by immunoblotting and immunofluorescence staining. RGCs subjected to OGD for 6h induced 50{\%} cell death in primary RGCs (. p<0.001) and inhibited pERK1/2 expression by 65{\%} (. p<0.001). Cell death was attenuated when RGCs were treated with pentazocine under OGD (. p<0.001) and pERK1/2 expression was increased by 1.6 fold (. p<0.05) compared to OGD treated RGCs without pentazocine treatment. The co-treatment of PD098059 (MEK1/2 inhibitor) with pentazocine significantly abolished the protective effects of pentazocine on the RGCs during this OGD insult. Activation of the σ-1 receptor is a neuroprotective target that can protect RGCs from an ischemia-like insult. These results also established a direct relationship between σ-1 receptor stimulation and the neuroprotective effects of the ERK1/2 pathway in purified RGCs subjected to OGD. These findings suggest that activation of the σ-1 receptor may be a therapeutic target for neuroprotection particularly relevant to ocular neurodegenerative diseases that effect RGCs.",
keywords = "Extracellular signal-regulated kinases 1/2, Neuroprotection, Primary retinal ganglion cells, Sigma-1 receptor",
author = "Mueller, {Brett H.} and Yong Park and Ma, {Hai Ying} and Adnan Dibas and Ellis, {Dorette Zoe} and Abbot Clark and Thomas Yorio",
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journal = "Experimental Eye Research",
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Sigma-1 receptor stimulation protects retinal ganglion cells from ischemia-like insult through the activation of extracellular-signal-regulated kinases 1/2. / Mueller, Brett H.; Park, Yong; Ma, Hai Ying; Dibas, Adnan; Ellis, Dorette Zoe; Clark, Abbot; Yorio, Thomas.

In: Experimental Eye Research, Vol. 128, 01.11.2014, p. 156-169.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - Sigma-1 receptor stimulation protects retinal ganglion cells from ischemia-like insult through the activation of extracellular-signal-regulated kinases 1/2

AU - Mueller, Brett H.

AU - Park, Yong

AU - Ma, Hai Ying

AU - Dibas, Adnan

AU - Ellis, Dorette Zoe

AU - Clark, Abbot

AU - Yorio, Thomas

PY - 2014/11/1

Y1 - 2014/11/1

N2 - Sigma-1 receptor (σ-1) activation and mitogen-activated protein kinases (MAPKs) have been shown to protect retinal ganglion cells (RGCs) from cell death. The purpose of this study was to determine if σ-1 receptor stimulation with pentazocine could promote neuroprotection under conditions of an ischemia-like insult (oxygen glucose deprivation (OGD)) through the phosphorylation of extracellular signal regulated kinase (pERK)1/2. Primary RGCs were isolated from P3-P7 Sprague-Dawley rats and purified by sequential immunopanning using Thy1.1 antibodies. RGCs were cultured for 7 days before subjecting the cells to an OGD insult (0.5% oxygen in glucose-free medium) for 6h. During the OGD, RGCs were treated with pentazocine (σ-1 receptor agonist) with or without BD 1047 (σ-1 receptor antagonist). In other experiments, primary RGCs were treated with pentazocine in the presence or absence of an MEK1/2 inhibitor, PD098059. Cell survival/death was assessed by staining with the calcein-AM/ethidium homodimer reagent. Levels of pERK1/2, total ERK1/2, and beta tubulin expression were determined by immunoblotting and immunofluorescence staining. RGCs subjected to OGD for 6h induced 50% cell death in primary RGCs (. p<0.001) and inhibited pERK1/2 expression by 65% (. p<0.001). Cell death was attenuated when RGCs were treated with pentazocine under OGD (. p<0.001) and pERK1/2 expression was increased by 1.6 fold (. p<0.05) compared to OGD treated RGCs without pentazocine treatment. The co-treatment of PD098059 (MEK1/2 inhibitor) with pentazocine significantly abolished the protective effects of pentazocine on the RGCs during this OGD insult. Activation of the σ-1 receptor is a neuroprotective target that can protect RGCs from an ischemia-like insult. These results also established a direct relationship between σ-1 receptor stimulation and the neuroprotective effects of the ERK1/2 pathway in purified RGCs subjected to OGD. These findings suggest that activation of the σ-1 receptor may be a therapeutic target for neuroprotection particularly relevant to ocular neurodegenerative diseases that effect RGCs.

AB - Sigma-1 receptor (σ-1) activation and mitogen-activated protein kinases (MAPKs) have been shown to protect retinal ganglion cells (RGCs) from cell death. The purpose of this study was to determine if σ-1 receptor stimulation with pentazocine could promote neuroprotection under conditions of an ischemia-like insult (oxygen glucose deprivation (OGD)) through the phosphorylation of extracellular signal regulated kinase (pERK)1/2. Primary RGCs were isolated from P3-P7 Sprague-Dawley rats and purified by sequential immunopanning using Thy1.1 antibodies. RGCs were cultured for 7 days before subjecting the cells to an OGD insult (0.5% oxygen in glucose-free medium) for 6h. During the OGD, RGCs were treated with pentazocine (σ-1 receptor agonist) with or without BD 1047 (σ-1 receptor antagonist). In other experiments, primary RGCs were treated with pentazocine in the presence or absence of an MEK1/2 inhibitor, PD098059. Cell survival/death was assessed by staining with the calcein-AM/ethidium homodimer reagent. Levels of pERK1/2, total ERK1/2, and beta tubulin expression were determined by immunoblotting and immunofluorescence staining. RGCs subjected to OGD for 6h induced 50% cell death in primary RGCs (. p<0.001) and inhibited pERK1/2 expression by 65% (. p<0.001). Cell death was attenuated when RGCs were treated with pentazocine under OGD (. p<0.001) and pERK1/2 expression was increased by 1.6 fold (. p<0.05) compared to OGD treated RGCs without pentazocine treatment. The co-treatment of PD098059 (MEK1/2 inhibitor) with pentazocine significantly abolished the protective effects of pentazocine on the RGCs during this OGD insult. Activation of the σ-1 receptor is a neuroprotective target that can protect RGCs from an ischemia-like insult. These results also established a direct relationship between σ-1 receptor stimulation and the neuroprotective effects of the ERK1/2 pathway in purified RGCs subjected to OGD. These findings suggest that activation of the σ-1 receptor may be a therapeutic target for neuroprotection particularly relevant to ocular neurodegenerative diseases that effect RGCs.

KW - Extracellular signal-regulated kinases 1/2

KW - Neuroprotection

KW - Primary retinal ganglion cells

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DO - 10.1016/j.exer.2014.10.007

M3 - Article

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JO - Experimental Eye Research

JF - Experimental Eye Research

SN - 0014-4835

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