Reduced glutamate uptake by retinal glial cells under ischemic/hypoxic conditions

The high-affinity uptake of glutamate by glial cells and neurons of the central nervous system, including the retina, serves to inactivate synaptically released glutamate and maintains glutamate at low concentrations in the extracellular space. This uptake prevents accumulation of glutamate extracellularly and thus minimizes the possibility of glutamate neurotoxicity secondary to ischemic insult. One mechanism whereby glutamate neurotoxicity may occur in ischemic/hypoxic insult is through increased extracellular K+ reversing the electrogenic glutamate uptake into retinal glial (Müller) cells. We investigated glial uptake of the amino acids glutamate, GABA, and D-aspartate in the intact isolated rat retina under high extracellular K+ conditions and under conditions simulating ischemia. Immunocytochemical findings showed that uptake of glutamate and GABA by MIller cells in the intact isolated rat retina continues under conditions simulating ischemia and high extracellular K+ conditions, and uptake of D-aspartate also continues under high K+ conditions. However, under high K+ conditions, the glutamate uptake system saturates at a lower concentration of exogenous glutamate than in the normal K+ condition. These findings provide evidence that disruption of glutamate uptake by Müller cells is likely to be a significant contributing factor to excess glutamate accumulation in the extracellular space which can lead to neurotoxicity.     

Changes of GABA metabolic enzymes in acute retinal ischemia.

It is reported that GABA accumulates in Müller cells in ischemic and diabetic rat retina. To investigate the mechanism of GABA accumulation in Müller cells, we localized GABA and glutamate in ischemic rat retina and measured the activity of GAD and GABA-T, enzymes involved in GABA metabolism. Using general anesthesia, we incised the bulbar conjunctiva of the rat around the limbus and clamped the left optic nerve. A sham operation was performed on the right eyes. Ocular ischemia was sustained for 30, 60 and 90 minutes. Rat eyes were enucleated immediately after ischemia and prepared for immunohistochemistry and enzyme activity measurement. Glutamate-like immunoreactivity (Glu-IR) in the sham-operated rat retina was observed in all retinal layers, showing intense staining in the nerve fiber layer (NFL), ganglion cell layer (GCL), and inner plexiform layer (IPL). Glu-IR increased in the outer plexiform layer (OPL) and outer nuclear layer (ONL) in an ischemic time-dependent manner. GABA-like immunoreactivity (GABA-IR) in sham-operated rat retina was observed in NFL, GCL, IPL and inner nuclear layer (INL). When the ischemic time was extended, GABA-IR intensely stained Müller cells. GAD activity was not changed in ischemic rat retina as compared to normal rat retina, but GABA-T activity was significantly decreased in ischemic rat retina. These results suggested that glutamate was induced by ischemia and was converted to GABA by GAD activity. Increased GABA was not metabolized because GABA-T activity was decreased. GABA accumulation in Müller cells progressed during the change in activity of these metabolic enzymes.     

Neuroprotective effects of D-allose against retinal ischemia-reperfusion injury

PURPOSE: To investigate the effect of D-allose, a rare sugar, against ischemia reperfusion injury in the rat retina. METHODS: Retinal ischemia was induced by increasing intraocular pressure to 130 mm Hg and maintaining that level for 45 minutes. Morphometric studies were performed to study the effect of D-allose on the histologic changes induced by ischemia in the rat retina. Glutamate release from the rat retina and intravitreal P(O2) profiles were monitored during and after ischemia with a microdialysis biosensor and oxygen-sensitive microelectrodes. The release of hydrogen peroxide stained with diaminobenzidine hydrochloride was monitored by an in vitro retinal ischemia model. RESULTS: Seven days after the ischemia, significant reductions in both the number of ganglion cells and the thickness of the inner plexiform layer were observed. Pretreatment with D-allose significantly inhibited the ischemic injury of the inner retina. A large release of glutamate occurred during the ischemia. After the recirculation, glutamate levels were increased again and reached a maximum in approximately 20 minutes. The increases in extracellular glutamate during and after ischemia tend to be suppressed by administration of d-allose. d-Allose attenuated the increase in intravitreal P(O2) during reperfusion. After the ischemia, production of hydrogen peroxide was detected within approximately 30 minutes. D-allose suppressed the production of hydrogen peroxide. CONCLUSIONS: These results suggest that D-allose may protect neurons by decreasing extracellular glutamate and attenuating oxidative stress in ischemic insult.     

Protective effect of trimetazidine in a model of ischemia-reperfusion in the rat retina

Trimetazidine is an anti-ischemic agent which is frequently prescribed as a prophylactic treatment of episodes of angina pectoris and as a symptomatic treatment of vertigo and tinnitus. It has also shown beneficial effects in models of visual dysfunction, but the mechanism(s) by which this occurs is as yet undefined. The present study was intended to evaluate the influence of trimetazidine on retinal damage induced by ischemia-reperfusion in a rat model. Retinal ischemia was induced by increasing intraocular pressure to 160 mm Hg for 60 min. Trimetazidine or buffer controls were administered 3 days before the ischemia or in the postischemic period. The degree of retinal damage was assessed after 15 and 30 days of reperfusion after the ischemic insult by histopathologic study according to Hughes' quantification of ischemic damage. Retinal ischemia led to significant reductions in thickness and cell number, mainly in the inner retinal layers. The results from the study demonstrate that treatment with intraperitoneally injected trimetazidine conferred significant protection against retinal ischemic damage. Better results were obtained in the pretreatment group after 15 days of reperfusion. Trimetazidine protects the rat retina from pressure-induced ischemic injury and might be considered a potential therapeutic modality for combating retinal ischemia.     

Neurochemical changes following postmortem ischemia in the rat retina

Glutamate and gamma-aminobutyric acid (GABA) are the dominant amino acids in the retina and brain. The manufacturing and degradation pathways of both of these amino acids are intricately linked with the tricarboxylic acid cycle leading to rapid redistribution of these amino acids after metabolic insult. Postmortem ischemia in mammalian retina predominantly results in a loss of glutamate and GABA from neurons and accumulation of these amino acids within Müller cells. This accumulation of glutamate and GABA in Müller cells may occur as a result of increased release of these neurotransmitters from neurons, and decreased degradation. Quantification of the semisaturation value (half-maximal response) for glutamate and GABA Müller cell loading during postmortem ischemia indicated a shorter semisaturation value for GABA than glutamate. Such changes are consistent with a single aerobically dependent GABA-degradation pathway, and the existence of multiple glutamate-degradation pathways. Comparison with the in vitro ischemic model showed similar qualitative characteristics, but a markedly increased semisaturation time for glutamate and GABA Müller cell loading (a factor of 5-10) in the postmortem ischemia model. We interpret these differences to indicate that the in vitro condition provides a more immediate and/or severe ischemic insult. In the postmortem ischemia model, the delayed glial cell loading implies the availability of internal stores of both glucose and/or oxygen. Increased glial and neuronal immunoreactivity for the amino acids involved in transamination reactions, aspartate, alanine, leucine, and ornithine was observed, indicating a potential shift in the equilibrium of transamination reactions associated with glutamate production. These findings provide evidence that, in the rat retina, there are multiple pathways subserving glutamate production/degradation that include a multitude of transamination reactions. Further evidence is therefore provided to support a role for all four amino acids in glutamate metabolism within a variety of retinal neurons and glia.     

Downregulation of glutamine synthetase via GLAST suppression induces retinal axonal swelling in a rat ex vivo hydrostatic pressure model

PURPOSE. High levels of glutamate can be toxic to retinal GCs. Thus, effective buffering of extracellular glutamate is important in preserving retinal structure and function. GLAST, a major glutamate transporter in the retina, and glutamine synthetase (GS) regulate extracellular glutamate accumulation and prevent excitotoxicity. This study was an examination of changes in function and expression of GLAST and GS in ex vivo rat retinas exposed to acute increases in ambient pressure. METHODS. Ex vivo rat retinas were exposed to elevated hydrostatic pressure for 24 hours. The expression of GLAST and GS were examined using immunochemistry and real-time PCR analysis. Also examined were the effects of (2S,3S)-3-[3-[4-(trifluoromethyl) benzoylamino] benzyloxy] aspartate (TFB-TBOA), an inhibitor of glutamate transporters, and l-methionine-S-sulfoximine (MSO), an inhibitor of GS. RESULTS. In this acute model, Western blot and real-time RT-PCR analyses revealed that substantially (75 mm Hg), but not moderately (35 mm Hg), elevated pressure depressed GLAST expression, diminished GS activity, and induced axonal swelling between the GC layer and the inner limiting membrane. However, at the moderately elevated pressure (35 mm Hg), administration of either TFB-TBOA or MSO also induced axonal swelling and excitotoxic neuronal damage. MSO did not depress GLAST expression but TFB-TBOA significantly suppressed GS, suggesting that downregulation of GS during pressure loading may result from impaired GLAST expression. CONCLUSIONS. The retina is at risk during acute intraocular pressure elevation due to downregulation of GS activity resulting from depressed GLAST expression.     

Neuroprotective effects of eliprodil in retinal excitotoxicity and ischemia

PURPOSE: To evaluate whether eliprodil (SL82.0715), a NR2B-selective N-methyl-D-aspartate (NMDA) antagonist, is protective of retina subjected to an excitotoxic or ischemic insult. METHODS: To evaluate protection against retinal excitotoxicity, eliprodil was administered intraperitoneally before and after the injection of NMDA (5 microl, 20 nmol) into the vitreous of rats. Integrity of the retina was assessed by counting cells in the retinal ganglion cell layer (GCL) and measuring choline acetyltransferase (ChAT) activity. In a subsequent experiment, total retinal ischemia, as measured by a cessation of electroretinographic (ERG) activity, was induced in anesthetized rabbits by elevating intraocular pressure above systolic blood pressure for 65 minutes. After ischemia, recovery of ERG activity was assessed at 24 and 48 hours in animals treated with vehicle or eliprodil (1.0-10.0 mg/kg). RESULTS: Intravitreal NMDA injection resulted in a dose-related decrease in cells of the GCL and in ChAT activity. Eliprodil administered intraperitoneally at 10 mg/kg completely prevented the loss of ChAT and the loss of cells in the GCL. Twenty-four hours after retinal ischemia, A and B waves of vehicle-treated animals were suppressed by 60% to 70%. Eliprodil administered intraperitoneally at 10 mg/kg ameliorated the A- and B-wave depression throughout the 48-hour experiment. CONCLUSIONS: Eliprodil is neuroprotective of retinae subjected to either an excitotoxic or ischemic challenge and may be useful for treating a variety of retinal and optic nerve head disorders.     

Diltiazem-induced Neuroprotection in Glutamate Excitotoxicity and Ischemic Insult of Retinal Neurons

Purpose: Cell death is often related to an abnormal increase in Ca²⁺ flux. In the retina, Ca²⁺ channels are mainly from the L-type that do not inactivate with time. Under excitotoxic and ischemic conditions, their continuous activation may therefore contribute significantly to the lethal Ca²⁺ influx. To assess this hypothesis, the Ca²⁺ channel blocker, diltiazem, was applied in excitotoxic and ischemic conditions. Methods: To induce excitotoxicity, retinal cell cultures from newborn rats were incubated with glutamate. The toxicity of glutamate was quantified by neuronal immunostaining with an antibody directed against the neuron specific enolase. Glutamate receptor function in vitro was assessed in pig retinal cell cultures by patch clamp recording. Retinal ischemia was induced by raising the intraocular pressure in adult rats. Retinal cell loss was quantified on retinal sections by measuring nuclear cell densities. Results: In retinal cell culture, glutamate application induced a major cell loss. This cell loss was attributed to glutamate excitotoxicity because glutamate receptor blockers like MK-801 and CNQX increased significantly neuronal survival. MK-801 and CNQX, which block NMDA and AMPA/Kainate receptors, respectively, had additive effects. Expression of AMPA/Kainate glutamate receptors in mixed adult retinal cell cultures was attested by patch clamp recording. In newborn rat retinal culture, glutamate excitotoxicity was significantly reduced by addition of the L-type Ca²⁺ channel blocker, diltiazem. In in vivo experiments, the increase in ocular pressure induced a decrease in cell number in the inner nuclear and ganglion cell layers. When animals received diltiazem injections, the ischemic treatment induced a less severe reduction in retinal cells; this neuroprotection was statistically significant in the ganglion cell layer. Conclusion: These results are consistent with previous studies suggesting that Ca²⁺ channel activation contributes to retinal cell death following either glutamate excitotoxicity or retinal ischemia. Under both conditions, the L-type Ca²⁺ channel blocker, diltiazem, can limit cell death. These results extend the potential application of diltiazem in retinal neuroprotection to retinal pathologies involving glutamate excitotoxicity and ischemia.     

Atrial natriuretic peptide inhibits osmotical glial cell swelling in the ischemic rat retina: dependence on glutamatergic-purinergic signaling

Atrial natriuretic peptide (ANP) is a regulator of the water and electrolyte content in the brain which also mediates cell volume homeostasis. Here, we determined whether the expression of ANP in the retina of the rat undergoes changes during ischemia-reperfusion, and whether ANP affects the osmotic swelling of Müller glial cells in postischemic retinas under hypotonic conditions. Transient retinal ischemia was induced by elevation of the intraocular pressure above systolic blood pressure for 1h. At 1 and 3 days after reperfusion, there was an increased content of ANP protein in the retina, as determined by Western blotting. The increase of the retinal ANP content was markedly reduced when triamcinolone acetonide (10 mM in 2 microl vehicle) was intravitreally injected before ischemia. ANP inhibited the osmotic swelling of Müller cell somata in retinal slices. The effect of ANP was mediated by activation of NP receptors expressed by retinal neurons which evoked a release of glutamate. The stimulation of metabotropic glutamate receptors expressed by Müller cells evoked an autocrine purinergic signaling mechanism that resulted in the opening of K(+) and Cl(-) channels; the ion efflux counteracted the osmotic swelling of Müller cells. It is concluded that the expression of ANP is transiently upregulated in the postischemic retina of the rat. The increased expression of ANP may represent a part of the retinal response to transient ischemia and may inhibit cytotoxic glial cell swelling.     

Localization of amino acid neurotransmitters following in vitro ischemia and anoxia in the rat retina

Glutamate and gamma-aminobutyric acid (GABA) are two of the dominant neurotransmitters in the retina and brain. The production/degradation of glutamate and GABA involves an intricate interrelationship between neurons and glia, as well as aerobic and anaerobic metabolic pathways. The aim of this work was to develop an in vitro model of retinal ischemia/anoxia and determine the changes in cellular localization of glutamate and GABA and the time course for such changes. After anoxic/ischemic insult, glutamate and GABA rapidly accumulate within glia with GABA showing a quicker time course and larger magnitude change. The accumulation time constant for both glutamate and GABA under anoxic conditions was dependent upon glucose concentration: high glucose levels resulted in delayed glial amino acid loading. The differences in time constants between GABA and glutamate glial loading most likely reflect the multitude of glutamate degradation pathways compared to the single aerobically dependent GABA pathway. Oxygen availability and reduced glucose (hypoglycemia) lead to an almost immediate increase (within 1 min) of glutamate and GABA labelling within glia. In addition, altered labelling patterns were found under anoxic/ischemic conditions for amino acids involved in glutamate transamination reactions: aspartate, leucine, alanine. and ornithine. These changes are consistent with alterations of equilibria of enzymatic reactions involved in glutamate metabolism, and thus support a role for all four amino acids in glutamate metabolism within a variety of retinal neurons.     

Mechanism of the pathogenesis of glutamate neurotoxicity in retinal ischemia

· Purpose: This study was carried out to examine the involvement of glutamate and nitric oxide neurotoxicity in ischemia/reperfusion-induced retinal injury in vivo. · Methods: We monitored glutamate release from in vivo cat retina during and after pressure-induced ischemia using a microdialysis technique. Morphometric studies were performed to study the effects of MK-801 (dizocilpine), L-NAME (N ^ω-nitro-l-arginine methyl ester), and D-NAME (N ^ω-nitro-d-arginine methyl ester) on the histological changes in the rat retina induced by ischemia or intravitreal injection of NMDA (N-methyl-d-aspartate; 200 nmol). · Results: A large release of glutamate occurred during ischemia, followed by a marked release after reperfusion. Histological changes occurred selectively in the inner part of the retina after ischemia as well as intravitreal injection of NMDA. Pretreatment with intravenous injection of MK-801 or L-NAME significantly inhibited the ischemic injury of the inner retina. Intravitreal injection of L-NAME inhibited NMDA-induced neurotoxicity in the retina. · Conclusion: These findings indicate that nitric oxide mediates neurotoxic actions of glutamate which are responsible for ischemic injury in the retina. Received: 17 November 1997 Revised version received: 3 February 1998 Accepted: 5 February 1998     

Glutamate elevation in rabbit vitreous during transient ischemia-reperfusion

PURPOSE: To investigate the response of the retina to an ischemic insult, we measured the levels of glutamate in the vitreous of rabbit eyes. METHODS: Ocular ischemia was induced in the vitreous of rabbit eyes by simultaneous ligature of the optic nerve, ciliary arteries, and extraocular muscles for 45 minutes. Contralateral eyes were subjected to a conjunctival peritomy to serve as sham-operated controls. Reperfusion was achieved by releasing the ligature. Eyes were enucleated at the end of the ischemic period or 15, 45, or 75 minutes after reperfusion. RESULTS: Analysis of the vitreous free amino acids showed a significant increase in glutamate levels in the operated eyes at the end of the ischemic period (P <.001) and after 15 minutes of reperfusion (P <.05) as compared with the contralateral, sham-operated eyes. Glutamine levels in the vitreous were unchanged throughout the study. CONCLUSIONS: These results show that glutamate, which is considered to be derived from synaptic release of the retinal neurons or accumulation due to a deterioration of glutamate uptake or a degradation system in the retina, was transiently elevated in the vitreous.     

Retinal ischemia and cell proliferation in the rat: the role of soluble mitogens

Temporary retinal ischemia in the rat leads to a proliferation of endothelial cells and glial cells. We tested the hypothesis that this proliferation is caused by a release of soluble mitogens from the ischemic retina. Conditioned media were prepared from normal rat retina and from retina that had undergone 2 h ischemia and 48 h reperfusion, at which time it showed intense mitotic activity. The conditioned media were placed in cultures of fibroblasts, bovine adrenal capillary endothelial cells, and rat brain astrocytes. Cell proliferation in vitro was stimulated by the retinal extracts in all cell cultures. However, the cell proliferation in cultures with conditioned media from normal retina was similar to that in cultures with conditioned media from ischemic and proliferating retina. Although these data are consistent with the presence of soluble growth factors in the retina, they also indicate that release of these growth factors into the surrounding milieu after transient retinal ischemia is not altered to a degree that would explain the dramatic increase in mitosis.Supported by Research to Prevent Blindness Inc., The National Eye Institute (EY05903 and R01-07001), The Adler Foundation, and the American Federation For Aging Research, Inc     

Retinal ischemia and cell proliferation in the rat: the role of soluble mitogens

Temporary retinal ischemia in the rat leads to a proliferation of endothelial cells and glial cells. We tested the hypothesis that this proliferation is caused by a release of soluble mitogens from the ischemic retina. Conditioned media were prepared from normal rat retina and from retina that had undergone 2 h ischemia and 48 h reperfusion, at which time it showed intense mitotic activity. The conditioned media were placed in cultures of fibroblasts, bovine adrenal capillary endothelial cells, and rat brain astrocytes. Cell proliferation in vitro was stimulated by the retinal extracts in all cell cultures. However, the cell proliferation in cultures with conditioned media from normal retina was similar to that in cultures with conditioned media from ischemic and proliferating retina. Although these data are consistent with the presence of soluble growth factors in the retina, they also indicate that release of these growth factors into the surrounding milieu after transient retinal ischemia is not altered to a degree that would explain the dramatic increase in mitosis. Supported by Research to Prevent Blindness Inc., The National Eye Institute (EY05903 and R01-07001), The Adler Foundation, and the American Federation For Aging Research, Inc     

Interleukin-1beta mediates ischemic injury in the rat retina.

Two types of experiment were performed to examine the role of interleukin-1beta in ischemia-induced damage in the rat retina. In the in vivo study, enzyme-linked immunosorbent assay was used to investigate the expression of immunoreactive interleukin-1beta in the rat retina following a hypertension-induced ischemia/reperfusion, while the effect of a recombinant human interleukin-1 receptor antagonist or an anti-interleukin-1beta neutralizing antibody on the ischemia-induced damage was examined histologically. A transient increase in the expression of immunoreactive interleukin-1beta was observed in the retina 3-12 hr after reperfusion, and morphometric evaluation at 7 days after the ischemia showed a decrease in cell numbers in the ganglion cell layer and a decreased thickness of the inner plexiform layer with no change in the other retinal layers. Intravitreal injection of interleukin-1 receptor antagonist (1 or 10 ng per eye) or anti-interleukin-1beta antibody (50 or 500 ng per eye) 5 min before the onset of the ischemia reduced the damage. In the in vitro study, interleukin-1 receptor antagonist (500 ng ml(-1)) significantly reduced glutamate-induced neurotoxicity in rat cultured retinal neurons. These results suggest that interleukin-1 plays an important role in mediating ischemic and excitotoxic damage in the retina, and that interleukin-1 inhibitors may be therapeutically useful against neuronal injury caused by optic nerve or retinal diseases such as glaucoma and central retinal artery or vein occlusion.     

Activation of matrix metalloproteinase-9 via neuronal nitric oxide synthase contributes to NMDA-induced retinal ganglion cell death

PURPOSE: Understanding the mechanism of neuronal cell death in retinal diseases like glaucoma is important for devising new treatments. One factor involves excitatory amino acid stimulation of N-methyl-D-aspartate (NMDA)-type glutamate receptors, excessive Ca2+ influx, and formation of nitric oxide (NO) via neuronal NO synthase (nNOS). Another factor is the abnormal activation of matrix metalloproteinases (MMPs), in particular MMP-9, which triggers an extracellular signaling cascade leading to apoptosis. This study was designed to investigate the mechanism of excitotoxic retinal ganglion cell (RGC) death in vivo and its relationship to MMP activation. METHODS: NMDA and glycine were injected into the vitreous of the eye in rats and in nNOS-deficient mice (nNOS-/-) versus control. Gelatinolytic activity of MMP-9 and MMP-2 by zymography and cellular localization by immunohistochemistry were examined, and the effect of MMP inhibition on NMDA-induced RGC death was tested. RESULTS: NMDA was found to upregulate the proform of MMP-9 in the retina and to increase MMP-9 gelatinolytic activity. Retrograde labeling with aminostilbamidine to identify RGCs confirmed that MMP activity occurred only in these retinal neurons and not in glial or other retinal cell types after excitotoxic insult. Deconvolution fluorescence microscopy revealed that MMP activity colocalized with immunoreactive S-nitrosylated protein. NMDA-induced MMP activation was diminished in the retina of nNOS-/- mice, implying that S-nitrosylation of MMP had indeed occurred. In addition, the broad-spectrum MMP inhibitor GM6001 protected RGCs after intravitreal NMDA injection. CONCLUSIONS: These findings suggest that an extracellular proteolytic pathway in the retina contributes to RGC death via NO-activated MMP-9.     

Riluzole improves functional recovery after ischemia in the rat retina

PURPOSE: Retinal ischemia leads to neuronal death. The effects of riluzole, a drug that protects against the deleterious effect of cerebral ischemia by acting on several types of ion channels and blocking glutamatergic neurotransmission, were investigated in a rat model of retinal ischemic injury. METHODS: Retinal ischemia was induced by increasing intraocular pressure above systolic blood pressure for 30 minutes. Electroretinograms were recorded before ischemia and at different periods of reperfusion. Riluzole was injected or topically applied to the eye before or after ischemia and twice daily during the reperfusion period. Retinas were harvested for histopathology (toluidine blue and silver-impregnation stainings, Tdt-dUTP terminal nick-end labeling [TUNEL] method) and immunohistochemistry for cytoskeletal glial fibrillary acid protein and c-jun NH2-terminal kinase (p-JNK). RESULTS: Ischemia for 30 minutes caused a reduction of a- and b-waves of the electroretinogram. Systemic and topical treatments with riluzole significantly enhanced the recovery of the reduced a- and b-waves after defined reperfusion times. Riluzole also prevented or attenuated ischemia-induced retinal cell death (necrosis and apoptosis) and reduced the activation of p-JNK, c-jun phosphorylation, and the increase of cytoskeletal proteins induced by ischemic injury. CONCLUSIONS: Riluzole acted in vivo as a potent neuroprotective agent against pressure-induced ischemia. Therefore, riluzole may be a major drug for use in protection against retinal injury.     

谷氨酸对视网膜缺血的损害机制和相关治疗策略

视网膜缺血损害的机制复杂,大量的研究资料表明视网膜在缺血及再灌注时谷氨酸的释放量增加,谷氨酸的兴奋性毒性作用在缺血视网膜的病理发展中起到了重要的作用,是引起缺血视网膜神经元死亡的重要因素.通过减少谷氨酸的释放,应用谷氨酸受体拮抗剂限制谷氨酸的生物活性等,可以阻止或减轻缺血视网膜的损害.虽然到目前为止临床上还未出现治疗视网膜缺血的有效药物,但对谷氨酸的损害机制和相关治疗策略的实验性研究,为治疗该疾病提供了广阔的前景.     

Intracellular Ca(2+) changes induced by in vitro ischemia in rat retinal slices

The effect of ischemia on intracellular Ca(2+)concentration [[Ca(2+)](i)] in retinal slices was investigated. [Ca(2+)](i)in each layer of the retina was determined from fluorescence images in rat retinal slices loaded with fura2-AM. Ischemic like conditions were imposed on the retinal slice in vitro by perfusion with an oxygen/glucose deprived solution. All measurements were taken at 25 degrees C except when temperature dependence was examined. In response to 100 m M K(+)or 0.2 m M glutamate under normoxic conditions, the [Ca(2+)](i)increase was higher in the inner retinal layers. Fifteen min ischemia evoked an increase in Ca(2+)levels in all layers of the retina, and the rates of increase were especially high in the outer/inner segment layer and the outer nuclear layer. Ischemia in the absence of extracellular Ca(2+)also induced a Ca(2+)rise, but at lower rates than with standard ischemia. Intermittent ischemia, composed of three 15 min bursts of ischemia at 10 min intervals, promoted the Ca(2+)rise. There was a more marked rise in [Ca(2+)](i)when the temperature was increased to 29 or 33 degrees C. Thus, in the rat retinal slice, in vitro ischemia evoked a more marked Ca(2+)rise in the outer retina, which was in contrast to the Ca(2+)responses to glutamate or high K(+). The rates of increase in [Ca(2+)](i)with ischemia were larger at higher temperatures, and intermittent ischemia also promoted the Ca(2+)rise. These increases appear to be derived from predominant influx of extracellular Ca(2+)rather than release of intracellular Ca(2+)stores.     

Effect of trimetazidine on retinal ischemia/reperfusion injury in rats

PURPOSE: To investigate the effect of trimetazidine (TMZ), an antioxidant agent, on the ischemia/reperfusion (I/R) injury in rat retina histopathologically. METHODS: The retinal I/R model was carried out by the 4-vessel occlusion method on Wistar albino rats. Twenty-one rats were divided into 7 groups, each comprising 3 rats. The animals in groups 1, 2 and 3 underwent 30 min of ischemia + 4 h of reperfusion and were treated by the administration of saline, TMZ before reperfusion and TMZ before ischemia, respectively. The animals in groups 4, 5 and 6 underwent 90 min of ischemia + 4 h of reperfusion and were treated in the same way as those in groups 1, 2 and 3, respectively. The 7th group was sham operated. RESULTS: Thirty and 90 min of ischemia followed by 4 h of reperfusion induced retinal injury in the rat retina. Histopathologically, the inner plexiform and inner nuclear layers were the most affected parts. TMZ was able to reduce almost all retinal I/R damage when administered before ischemia. A cytoprotective effect of TMZ was partly observed in those animals which were treated before reperfusion. CONCLUSION: TMZ seemed to have a protective effect against retinal I/R injury in rats.