Measurement of amino acid levels in the vitreous humor of rats after chronic intraocular pressure elevation or optic nerve transection.
ABSTRACT To investigate whether the levels of free amino acids and protein in the vitreous of rat eyes are altered with chronic intraocular pressure (IOP) elevation or after optic nerve transection.
The concentrations of 20 amino acids in the vitreous humor were measured by high-performance liquid chromatography in both eyes of 41 rats with unilateral IOP elevation induced by translimbal photocoagulation. Eyes were studied 1 day and 1, 2, 4, and 9 weeks after initial IOP elevation. The same amino acids were measured in 41 rats 1 day and 2, 4, and 9 weeks after unilateral transection of the orbital optic nerve. The intravitreal protein level was assayed in additional 22 rats with IOP elevation and 12 rats after nerve transection. Two masked observers evaluated the amount of optic nerve damage with a semiquantitative, light-microscopic technique.
In rats with experimental glaucoma, amino acid concentrations were unchanged 1 day after treatment. At 1 week, 4 of 20 amino acids (aspartate, proline, alanine, and lysine) were higher than in control eyes ( < or = 0.01), but this difference was nonsignificant after Bonferroni correction for multiple simultaneous amino acid comparisons (none achieved < 0.0025). No amino acid was significantly different from control in the nerve transection groups (all > 0.05). Vitreous protein level was significantly higher in glaucomatous eyes than their paired controls at 1 day ( < 0.0001) and 1 week ( < 0.002). One day and 1 week after optic nerve transection, vitreal proteins were significantly elevated compared with control eyes from untreated animals ( < 0.0020 and < 0.0022, respectively), though not compared with their fellow eyes ( = 0.25 and 0.10).
Chronic experimental glaucoma and transection of the optic nerve increase the amount of protein in the rat vitreous above control levels. In the vitreous of rats with experimental glaucoma, a number of free amino acids were transiently elevated to a modest degree, but no significant difference in vitreous glutamate concentration was detected ( > 0.01).
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ABSTRACT: To induce chronic intraocular pressure (IOP) elevation in rat eyes by circumlimbal suture. Anesthetized (isoflurane) Long-Evans rats underwent unilateral circumlimbal suture implantation while the fellow eyes served as untreated controls (n=15). A sham group (n=8) received the same procedure except the suture was loosely tied. IOP, electroretinography (ERG) and optical coherence tomography (OCT) were monitored for 15 weeks, after which retinal histology and immunofluorescence staining for glial fibrillary acidic protein (GFAP) and ionized calcium binding adapter molecule-1 (Iba-1) were undertaken. Both IOP and ERG remained unaltered in the sham and all control eyes over 15 weeks. In the ocular hypertensive eye, IOP spiked from 17±1 to 58±3 mmHg immediately after suture application, recovering to 32±2 mmHg by 24 hours, and remained elevated by 7-10 mmHg above baseline for 15 weeks. At week 2, there was a small reduction of ERG components involving the photoreceptor a-wave, bipolar cell b-wave, and ganglion cell mediated Scotopic Threshold Response (pSTR). The reduction in a- and b-wave remained stable, while the pSTR became more affected from week 8 onwards (p<0.05). By weeks 12, there was progressive retinal nerve fiber layer (RNFL) thinning. At week 15, GFAP expression was upregulated in inner retina and on Müller cells. The ganglion cell dysfunction was associated with RNFL thinning and cell loss in the ganglion cell layer. Circumlimbal suture provides a simple and cost-effective way to induce mild chronic ocular hypertension in rat eyes. This model produces preferential ganglion cell dysfunction and RNFL reduction. Copyright © 2015 by Association for Research in Vision and Ophthalmology.Investigative ophthalmology & visual science 03/2015; DOI:10.1167/iovs.14-16009 · 3.66 Impact Factor
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ABSTRACT: 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. 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. 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. 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.Molecular vision 05/2014; 20:606-15. · 2.25 Impact Factor
North American journal of medicine & science 01/2008; 1(1). DOI:10.7156/v1i1p001