The role of N-methyl-D-aspartate receptor activation in homocysteine-induced death of retinal ganglion cells

Preethi S. Ganapathy, Richard E. White, Yonju Ha, B. Renee Bozard, Paul L. McNeil, R. William Caldwell, Sanjiv Kumar, Stephen M. Black, Sylvia B. Smith

Research output: Contribution to journalArticle

50 Citations (Scopus)

Abstract

Purpose. Elevated plasma homocysteine has been implicated in glaucoma, a vision disorder characterized by retinal ganglion cell death. The toxic potential of homocysteine to ganglion cells is known, but the mechanisms are not clear. A mechanism of homocysteine-induced death of cerebral neurons is via Nmethyl- D-aspartate (NMDA) receptor overstimulation, leading to excess calcium influx and oxidative stress. This study examined the role of the NMDA receptor in homocysteine-mediated ganglion cell death. Methods. Primary mouse ganglion cells were used for these experiments. NMDA receptor stimulation by homocysteine was determined by patch clamp analysis and fluorescent detection of intracellular calcium. NMDA receptor involvement in homocysteine-mediated cell death was determined through assessment of lactate dehydrogenase release and TUNEL analysis. These experiments used the NMDA receptor blocker MK-801. Induction of reactive species superoxide, nitric oxide, and peroxynitrite was measured by electron paramagnetic resonance spectroscopy, chemiluminescent nitric oxide detection, and immunoblotting for nitrotyrosine, respectively. Results. 50 μM homocysteine stimulated the NMDA receptor in presence of 100 μM glycine. Homocysteine induced 59.67 ± 4.89% ganglion cell death that was reduced to 19.87 ± 3.03% with cotreatment of 250 nM MK-801. Homocysteine elevated intracellular calcium ~7-fold, which was completely prevented by MK-801. Homocysteine treatment increased superoxide and nitric oxide levels by ~40% and ~90%, respectively, after 6 hours. Homocysteine treatment elevated peroxynitrite by ~85% after 9 hours. Conclusions. These experiments provide compelling evidence that homocysteine induces retinal ganglion cell toxicity through direct NMDA receptor stimulation and implicate, for the first time, the induction of oxidative stress as a potent mechanism of homocysteine-mediated ganglion cell death.

Original languageEnglish (US)
Pages (from-to)5515-5524
Number of pages10
JournalInvestigative Ophthalmology and Visual Science
Volume52
Issue number8
DOIs
StatePublished - Jul 2011
Externally publishedYes

Fingerprint

Retinal Ganglion Cells
Homocysteine
N-Methyl-D-Aspartate Receptors
D-Aspartic Acid
Ganglia
Cell Death
Dizocilpine Maleate
Nitric Oxide
Peroxynitrous Acid
Calcium
Superoxides
Oxidative Stress
Poisons
Vision Disorders
In Situ Nick-End Labeling
Electron Spin Resonance Spectroscopy
aspartic acid receptor
L-Lactate Dehydrogenase
Immunoblotting
Glaucoma

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience
  • Medicine(all)

Cite this

Ganapathy, P. S., White, R. E., Ha, Y., Renee Bozard, B., McNeil, P. L., William Caldwell, R., ... Smith, S. B. (2011). The role of N-methyl-D-aspartate receptor activation in homocysteine-induced death of retinal ganglion cells. Investigative Ophthalmology and Visual Science, 52(8), 5515-5524. https://doi.org/10.1167/iovs.10-6870

The role of N-methyl-D-aspartate receptor activation in homocysteine-induced death of retinal ganglion cells. / Ganapathy, Preethi S.; White, Richard E.; Ha, Yonju; Renee Bozard, B.; McNeil, Paul L.; William Caldwell, R.; Kumar, Sanjiv; Black, Stephen M.; Smith, Sylvia B.

In: Investigative Ophthalmology and Visual Science, Vol. 52, No. 8, 07.2011, p. 5515-5524.

Research output: Contribution to journalArticle

Ganapathy, PS, White, RE, Ha, Y, Renee Bozard, B, McNeil, PL, William Caldwell, R, Kumar, S, Black, SM & Smith, SB 2011, 'The role of N-methyl-D-aspartate receptor activation in homocysteine-induced death of retinal ganglion cells', Investigative Ophthalmology and Visual Science, vol. 52, no. 8, pp. 5515-5524. https://doi.org/10.1167/iovs.10-6870
Ganapathy, Preethi S. ; White, Richard E. ; Ha, Yonju ; Renee Bozard, B. ; McNeil, Paul L. ; William Caldwell, R. ; Kumar, Sanjiv ; Black, Stephen M. ; Smith, Sylvia B. / The role of N-methyl-D-aspartate receptor activation in homocysteine-induced death of retinal ganglion cells. In: Investigative Ophthalmology and Visual Science. 2011 ; Vol. 52, No. 8. pp. 5515-5524.
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abstract = "Purpose. Elevated plasma homocysteine has been implicated in glaucoma, a vision disorder characterized by retinal ganglion cell death. The toxic potential of homocysteine to ganglion cells is known, but the mechanisms are not clear. A mechanism of homocysteine-induced death of cerebral neurons is via Nmethyl- D-aspartate (NMDA) receptor overstimulation, leading to excess calcium influx and oxidative stress. This study examined the role of the NMDA receptor in homocysteine-mediated ganglion cell death. Methods. Primary mouse ganglion cells were used for these experiments. NMDA receptor stimulation by homocysteine was determined by patch clamp analysis and fluorescent detection of intracellular calcium. NMDA receptor involvement in homocysteine-mediated cell death was determined through assessment of lactate dehydrogenase release and TUNEL analysis. These experiments used the NMDA receptor blocker MK-801. Induction of reactive species superoxide, nitric oxide, and peroxynitrite was measured by electron paramagnetic resonance spectroscopy, chemiluminescent nitric oxide detection, and immunoblotting for nitrotyrosine, respectively. Results. 50 μM homocysteine stimulated the NMDA receptor in presence of 100 μM glycine. Homocysteine induced 59.67 ± 4.89{\%} ganglion cell death that was reduced to 19.87 ± 3.03{\%} with cotreatment of 250 nM MK-801. Homocysteine elevated intracellular calcium ~7-fold, which was completely prevented by MK-801. Homocysteine treatment increased superoxide and nitric oxide levels by ~40{\%} and ~90{\%}, respectively, after 6 hours. Homocysteine treatment elevated peroxynitrite by ~85{\%} after 9 hours. Conclusions. These experiments provide compelling evidence that homocysteine induces retinal ganglion cell toxicity through direct NMDA receptor stimulation and implicate, for the first time, the induction of oxidative stress as a potent mechanism of homocysteine-mediated ganglion cell death.",
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AU - McNeil, Paul L.

AU - William Caldwell, R.

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