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Hyperbaric pressure and increased susceptibility to glutamate toxicity in retinal ganglion cells in vitro.

Aihara M, Chen YN, Uchida S, Nakayama M, Araie M - Mol. Vis. (2014)

Bottom Line: The effects of N-Methyl-D-aspartic acid (NMDA) and 2-amino-3-(5-methyl-3-oxo-1,2- oxazol-4-yl)propanoic acid (AMPA)/kainate receptor antagonists, MK-801, and 6,7-dinitroquinoxaline-2,3-dione (DNQX), on cell survival were assessed.MK-801 and DNQX significantly reduced glutamate-induced RGC death, and DNQX was more effective than MK-801.In a rat RGC culture, hyperbaric pressure alone did not induce RGC death but increased RGC susceptibility to glutamate toxicity, which may be of relevance to ocular diseases with pressure-induced RGC death.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, University of Tokyo School of Medicine, Tokyo, Japan.

ABSTRACT

Purpose: To investigate the effect of hyperbaric pressure on purified retinal ganglion cells (RGCs) and the additive effect of hyperbaric pressure on glutamate-induced RGC death.

Methods: An RGC primary culture from 8-day-old Wistar rats was prepared and cultured in a hyperbaric chamber. The RGC survival rate under various pressure conditions and with 5 or 25 µM of glutamate stimulation was determined and compared with that of RGCs under isobaric conditions. First, RGCs were cultured at atmospheric pressure (0 mmHg) and under hyperbaric pressure (+30 and +90 mmHg, with pressure fluctuations varying from 0 to +30 or +60 mmHg). Next, RGCs were cultured at +15, +30, and +90 mmHg with the addition of 5 or 25 µM of glutamate. The effects of N-Methyl-D-aspartic acid (NMDA) and 2-amino-3-(5-methyl-3-oxo-1,2- oxazol-4-yl)propanoic acid (AMPA)/kainate receptor antagonists, MK-801, and 6,7-dinitroquinoxaline-2,3-dione (DNQX), on cell survival were assessed. Additionally, types of cell death and the induction of Bcl-2-associated X protein (BAX) leading to apoptosis were studied under hyperbaric pressure conditions and/or with 5 µM of glutamate.

Results: RGC death was not induced under increasing or fluctuating pressure conditions. RGC death was induced by 25 µM of glutamate and increased as pressure increased. RGC death was not induced by 5 µM of glutamate but was induced by and increased with increasing pressure. MK-801 and DNQX significantly reduced glutamate-induced RGC death, and DNQX was more effective than MK-801. Under hyperbaric pressure conditions, the addition of 5 µM of glutamate resulted in the induction of apoptosis and BAX, which did not occur under hyperbaric pressure conditions or with the addition of glutamate alone.

Conclusion: In a rat RGC culture, hyperbaric pressure alone did not induce RGC death but increased RGC susceptibility to glutamate toxicity, which may be of relevance to ocular diseases with pressure-induced RGC death.

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Related in: MedlinePlus

The effect of constant or variable pressure on RGC survival. A: RGCs were cultured under constant +30 and +90 mmHg hyperbaric pressure and were compared to the atmospheric pressure in 72 h. A constant hyperbaric pressure had no effect on RGC survival. B: RGCs were cultured under +0 and +60 mmHg of variable pressure with an 8 h cycle and under +30 mmHg pressure as a control. Repeated variable pressure also had no effect on RGC survival. The data indicates a mean±SD, which is not significantly different from the control (white bar), using Dunnett’s test, n=6. C: RGCs were cultured under +0 and +30 mmHg of variable pressure with an 8 h cycle and under +15 mmHg of pressure as a control. Repeated variable pressure also showed no effect on RGC survival. The data indicates a mean±SD, which is not significantly different from the control (white bar), using Dunnett’s test, n=6.
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f1: The effect of constant or variable pressure on RGC survival. A: RGCs were cultured under constant +30 and +90 mmHg hyperbaric pressure and were compared to the atmospheric pressure in 72 h. A constant hyperbaric pressure had no effect on RGC survival. B: RGCs were cultured under +0 and +60 mmHg of variable pressure with an 8 h cycle and under +30 mmHg pressure as a control. Repeated variable pressure also had no effect on RGC survival. The data indicates a mean±SD, which is not significantly different from the control (white bar), using Dunnett’s test, n=6. C: RGCs were cultured under +0 and +30 mmHg of variable pressure with an 8 h cycle and under +15 mmHg of pressure as a control. Repeated variable pressure also showed no effect on RGC survival. The data indicates a mean±SD, which is not significantly different from the control (white bar), using Dunnett’s test, n=6.

Mentions: The pressure variable chamber was purchased from Hirayama Company (Chiba, Japan) and was placed in the normal incubator (Figure 1 shows the effects of pressure and not the chamber setup). This type of pressure chamber contains a pressurized pipe allowing variations in the chamber’s inner pressure where the culture wells are placed. To avoid pressure-dependent increases in O2 concentration in the culture media, the inflated O2 concentration was automatically controlled based on Henry’s law as follows: the inflated O2 concentration=20% × 760 mmHg/incubating pressure.


Hyperbaric pressure and increased susceptibility to glutamate toxicity in retinal ganglion cells in vitro.

Aihara M, Chen YN, Uchida S, Nakayama M, Araie M - Mol. Vis. (2014)

The effect of constant or variable pressure on RGC survival. A: RGCs were cultured under constant +30 and +90 mmHg hyperbaric pressure and were compared to the atmospheric pressure in 72 h. A constant hyperbaric pressure had no effect on RGC survival. B: RGCs were cultured under +0 and +60 mmHg of variable pressure with an 8 h cycle and under +30 mmHg pressure as a control. Repeated variable pressure also had no effect on RGC survival. The data indicates a mean±SD, which is not significantly different from the control (white bar), using Dunnett’s test, n=6. C: RGCs were cultured under +0 and +30 mmHg of variable pressure with an 8 h cycle and under +15 mmHg of pressure as a control. Repeated variable pressure also showed no effect on RGC survival. The data indicates a mean±SD, which is not significantly different from the control (white bar), using Dunnett’s test, n=6.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4016804&req=5

f1: The effect of constant or variable pressure on RGC survival. A: RGCs were cultured under constant +30 and +90 mmHg hyperbaric pressure and were compared to the atmospheric pressure in 72 h. A constant hyperbaric pressure had no effect on RGC survival. B: RGCs were cultured under +0 and +60 mmHg of variable pressure with an 8 h cycle and under +30 mmHg pressure as a control. Repeated variable pressure also had no effect on RGC survival. The data indicates a mean±SD, which is not significantly different from the control (white bar), using Dunnett’s test, n=6. C: RGCs were cultured under +0 and +30 mmHg of variable pressure with an 8 h cycle and under +15 mmHg of pressure as a control. Repeated variable pressure also showed no effect on RGC survival. The data indicates a mean±SD, which is not significantly different from the control (white bar), using Dunnett’s test, n=6.
Mentions: The pressure variable chamber was purchased from Hirayama Company (Chiba, Japan) and was placed in the normal incubator (Figure 1 shows the effects of pressure and not the chamber setup). This type of pressure chamber contains a pressurized pipe allowing variations in the chamber’s inner pressure where the culture wells are placed. To avoid pressure-dependent increases in O2 concentration in the culture media, the inflated O2 concentration was automatically controlled based on Henry’s law as follows: the inflated O2 concentration=20% × 760 mmHg/incubating pressure.

Bottom Line: The effects of N-Methyl-D-aspartic acid (NMDA) and 2-amino-3-(5-methyl-3-oxo-1,2- oxazol-4-yl)propanoic acid (AMPA)/kainate receptor antagonists, MK-801, and 6,7-dinitroquinoxaline-2,3-dione (DNQX), on cell survival were assessed.MK-801 and DNQX significantly reduced glutamate-induced RGC death, and DNQX was more effective than MK-801.In a rat RGC culture, hyperbaric pressure alone did not induce RGC death but increased RGC susceptibility to glutamate toxicity, which may be of relevance to ocular diseases with pressure-induced RGC death.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, University of Tokyo School of Medicine, Tokyo, Japan.

ABSTRACT

Purpose: To investigate the effect of hyperbaric pressure on purified retinal ganglion cells (RGCs) and the additive effect of hyperbaric pressure on glutamate-induced RGC death.

Methods: An RGC primary culture from 8-day-old Wistar rats was prepared and cultured in a hyperbaric chamber. The RGC survival rate under various pressure conditions and with 5 or 25 µM of glutamate stimulation was determined and compared with that of RGCs under isobaric conditions. First, RGCs were cultured at atmospheric pressure (0 mmHg) and under hyperbaric pressure (+30 and +90 mmHg, with pressure fluctuations varying from 0 to +30 or +60 mmHg). Next, RGCs were cultured at +15, +30, and +90 mmHg with the addition of 5 or 25 µM of glutamate. The effects of N-Methyl-D-aspartic acid (NMDA) and 2-amino-3-(5-methyl-3-oxo-1,2- oxazol-4-yl)propanoic acid (AMPA)/kainate receptor antagonists, MK-801, and 6,7-dinitroquinoxaline-2,3-dione (DNQX), on cell survival were assessed. Additionally, types of cell death and the induction of Bcl-2-associated X protein (BAX) leading to apoptosis were studied under hyperbaric pressure conditions and/or with 5 µM of glutamate.

Results: RGC death was not induced under increasing or fluctuating pressure conditions. RGC death was induced by 25 µM of glutamate and increased as pressure increased. RGC death was not induced by 5 µM of glutamate but was induced by and increased with increasing pressure. MK-801 and DNQX significantly reduced glutamate-induced RGC death, and DNQX was more effective than MK-801. Under hyperbaric pressure conditions, the addition of 5 µM of glutamate resulted in the induction of apoptosis and BAX, which did not occur under hyperbaric pressure conditions or with the addition of glutamate alone.

Conclusion: In a rat RGC culture, hyperbaric pressure alone did not induce RGC death but increased RGC susceptibility to glutamate toxicity, which may be of relevance to ocular diseases with pressure-induced RGC death.

Show MeSH
Related in: MedlinePlus