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Rod–Cone Dystrophy with Maculopathy in Genetic Glutathione Synthetase Deficiency
Rod–Cone Dystrophy with Maculopathy in Genetic Glutathione Synthetase Deficiency A Morphologic and Electrophysiologic Study Marie S. I. Burstedt, MD, PhD,1 Ellinor Ristoff, MD, PhD,2 Agne Larsson, MD, PhD,2 Lillemor Wachtmeister, MD, PhD1 Purpose: To describe the retinal findings in 2 young adults with glutathione synthetase (GS) deficiency, an autosomal-recessive inborn error of glutathione (GSH) metabolism. Design: Report of 2 cases. Participants: Binocular study in 2 affected siblings. Methods: Two sisters with severe GS deficiency underwent a first ophthalmologic examination including full-field electroretinogram (ERGs). The single flash and flicker ERGs and the oscillatory potentials were measured. The clinical examination was repeated after 1 year with the addition of fluorescein angiography, optical coherence tomography (OCT), and electrooculography (EOG). Main Outcome Measures: Angiograms and the retinal OCTs were analyzed, the morphologic findings compared, and the Arden ratio measured. Results: Myopia decreased in both sisters, and visual acuity remained unchanged. Ophthalmoscopy showed bilateral retinal degenerative changes. Binocular cystic macular edema was present in the fovea and perifoveal areas. Cystic changes were located in the inner nuclear layer and outer plexiform layer. The ERGs showed low or no recordable rod-isolated b-waves, mixed rod–cone a- and b-waves, and cone responses. The oscillatory potentials were subnormal or nonrecordable. The EOG values were subnormal except in 1 eye of the older sister that had a normal Arden ratio. Conclusions: Severe GS deficiency is associated with progressive retinal dystrophy of the rod– cone type, affecting the central retina with advanced macular edema in adulthood. The retinal degenerative changes in GS deficiency may be the result of the increased oxidative stress accumulated generally in the retina and also apparent in the macular area, and an insufficient level of the free radical scavenger GSH. The patients with GS deficiency may represent a model of the retinal response to oxidative stress in humans. Financial Disclosure(s): The authors have no proprietary or commercial interest in any materials discussed in this paper. Ophthalmology 2009;116:324 –331 © 2009 by the American Academy of Ophthalmology.
The retina is among the most vascularized tissues in the body, and has one of the highest oxidative metabolic rates per tissue weight. The high content of polyunsaturated fatty acids in the photoreceptor outer segments also makes the photoreceptors susceptible to damage if high levels of reactive oxygen species are present. In addition, the retina is exposed to light, which promotes the generation of cytotoxic reactive oxygen species, which cause dysfunction and cell death in photoreceptors and other retinal cells. Factors such as high oxygen flux, light, and polyunsaturated fatty acids induce a high susceptibility of the retina to oxidative stress. Protection against this oxidative stress in the retina is provided by a complex defense mechanism involving the major water-soluble antioxidant glutathione (GSH), which is also found in the rest of the eye.1 This antioxidant functions primarily in the cytoplasm and mitochondria, and participates in many of the antioxidative cellular mechanisms that are essential for cellular survival,2 such as detoxification of xenobiotics and regulation of several cascades in cellular signaling. Glutathione synthetase (GS) deficiency is a rare, autosomalrecessive inborn error of metabolism. Patients with GS de-
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© 2009 by the American Academy of Ophthalmology Published by Elsevier Inc.
ficiency have low levels of cellular GSH (5%–20% of the normal mean in erythrocytes and fibroblasts; McKusick #266130). According to the severity of the clinical signs, patients with GS deficiency are divided into 3 subgroups: mild, moderate, or severe GS deficiency. Low levels of GSH result in hemolytic anemia; those with moderate disease also exhibit metabolic acidosis. Patients with severe GS deficiency experience progressive neurologic symptoms with retarded psychomotor development, seizures, spasticity, and cerebellar symptoms. In about 25% of patients with GS deficiency, the eyes are affected with retinitis pigmentosa, retinal dystrophy, crystalline opacities in the lens, and decreased visual acuity.3 Because the retina can be considered a part of the central nervous system, patients with retinal changes are classified as having a severe disease. We have previously reported on 2 sisters with severe GS deficiency who were investigated ophthalmologically as children and then reexamined 20 years later.4,5 In adulthood, they had developed moderate to advanced myopia, slowly progressing sparse lens opacities, and symptoms of defective macular function with modestly decreased visual acuISSN 0161-6420/09/$–see front matter doi:10.1016/j.ophtha.2008.09.007
Burstedt et al 䡠 Rode–Cone Dystrophy ity. The elder sister showed maculopathy with granular hyper- and hypopigmentations in both eyes and the younger sister showed bilateral maculopathy and multiple white spots of atrophy in the temporal area of the anatomic macula. Both sisters showed peripheral degenerative changes with granular, reticular, and bone spiculelike hyperpigmentations in the midperiphery that were more pronounced in the nasal and upper parts of the fundi, corresponding to defects in the visual field. In a previous study5 of the 2 sisters, the central visual pathways were found to be preserved with normal visual evoked potential responses, although the full-field electroretinograms (ERGs) of both sisters were even further affected compared to childhood, more than 20 years earlier.4 Thus, the retinal dystrophy in severe types of GS deficiency seems to advance, albeit slowly.5 We have also reported on 2 brothers with severe GS deficiency who presented retinitis pigmentosa with nearly extinguished ERG in adolescence, and a 3rd young boy with signs of retinal dystrophy and lowered visual acuity at the age of 10 years.3 Thus, there is limited information about the retinal findings in this small but important group of patients who, despite aggravating metabolic effect, can reach adulthood and, in addition to possible neurologic symptoms, develop retinal disease, which may lead to severe visual handicap. This study presents an analysis of retinal function as well as a closer evaluation of the retinal morphology in 2 sisters with severe GS deficiency, using full-field ERGs, electrooculograms (EOGs), fluorescein angiograms, and optical coherence tomography (OCT). We hypothesize that there is a relationship between low levels of GSH in the retina and progressive eye symptoms in GS-deficient patients.
Patients and Methods Patients Two sisters with severe GS deficiency were born to healthy, nonconsanguineous parents. The elder sister was diagnosed with GS deficiency at the age of 1 year and the younger at the age of 1 day. In the neonatal period, both showed characteristic signs of GS deficiency, namely, hemolytic anemia, metabolic acidosis, and
5-oxoprolinuria (14 –20 mol/mol creatinine; control range ⬍0.1 mol/mol creatinine). The GSH levels in erythrocytes were ⬍10% of the normal mean, GS (enzyme commission number [EC] 6.3.2.3) activity was ⬍5% of the normal mean, and GS activity in fibroblasts was ⬍20% of the normal mean. Both patients have been described previously.4 The sisters also showed a 1.4-fold to 4.5-fold increase in the levels of ␥-glutamylcysteine and cysteine in cultured fibroblasts (unpublished results). Other antioxidative enzymes (superoxide dismutase, catalase, and GSH peroxidase) had activities within normal limits in erythrocytes. Both sisters were homozygous for the mutation 77C¡A, Ala26Asp in the GS gene.6 Treatment with acidosis correction was started in the neonatal period. Vitamin E supplementation (10 mg/kg per day) was started before puberty, and vitamin C supplementation (100 mg/kg per day) when the sisters were in their 20s. Both sisters completed high school and found employment. The younger sister had a normal pregnancy and gave birth to a healthy, heterozygous child.7 When tested as adults, their intelligence quotients were within normal limits. Neurologic examination showed brisk stretch reflexes. The sisters had well-compensated hemolytic anemia, metabolic acidosis, and slightly elevated triglycerides, and both had a normal blood pressure. When examined for the present study, the sisters were 31 and 34 years old. The studies of the physiologic consequences of GSH deficiency were approved by the Ethics Committee of Karolinska Institutet, Stockholm, Sweden. The ophthalmologic investigations were considered as routine health care and performed according to standardized protocols and procedures. Informed consent was obtained, and the study followed the tenets of the Declaration of Helsinki.
Clinical and Morphologic Methods Routine clinical ophthalmologic methods were used to examine the anterior and posterior segments of both eyes of the 2 sisters. Monocular visual acuity was tested with the Early Treatment Diabetic Retinopathy Study chart at a distance of 4 m. Lighting conditions were standardized using an Early Treatment Diabetic Retinopathy Study chart illuminator cabinet (cat. no. 2425, Precision Vision). The time elapsed between the first clinical examination including electroretinography and second one including morphologic investigations and electrooculography was within 1 year (the older sister [JG] at 33 and 34 years of age, respectively, and the younger sister [HG] at 30 and 31 years of age, respectively). Fluorescein angiograms of the retinas and
Table 1. Macular Thickness Measurements Using the Stratus Optical Coherence Tomography (OCT3) Retinal Thickness (m) Inner Ring (Ø 3 mm)
Outer Ring (Ø 6 mm)
Right Eye
Left Eye
Right Eye
Left Eye
Right Eye
Left Eye
Right Eye
Left Eye
S301 I270 T289 N279 S456 I377 T412 N416
S364 I334 T354 N214 S410 I376 T399 N393
S235 I209 T215 N239 S264 I254 T244 N270
S238 I189 T222 N214 S247 I261 T252 N254
VA (LogMAR) Case/Age (yrs)
Refraction
Central (Ø 1 mm)
JG/34
⫺6.5 — 1.75 ⫻10 ⫺8.25 — 2.0 ⫻175
0.02
0.24
326
439
HG/31
⫺4.25 — 1 ⫻10 ⫺4.75 — 0.75 ⫻180
0.22
0.22
534
475
I ⫽ inferior; LogMAR ⫽ logarithm of minimum angle of resolution; N ⫽ nasal; S ⫽ superior; T ⫽ temporal; VA ⫽ visual acuity.
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Ophthalmology Volume 116, Number 2, February 2009 photographs of the fundus were taken and analyzed according to standard clinical procedures. The OCT images were examined with Stratus OCT, Version. 3.0 (Carl Zeiss Meditec AG, Oberkochen, Germany). Both eyes of the sisters were examined using 6-mm vertical scan lengths including the fovea and the adjacent perifoveal region. Late-phase angiograms (6 –9 minutes after injection) were compared with the corresponding cross-sectional OCT images.
Electrophysiologic Methods Full-field, single-flash, and flicker ERGs, including the oscillatory potentials, were recorded with a UTAS-E 2000 (LKC Technologies Inc, Gaithersburg, MD) using Burian–Allen bipolar electrodes and according to the recommendations of the International Society for Clinical Electrophysiology of Vision.8 The isolated rod responses were measured during standard dark-adapted conditions (20 minutes) using full-field white flashes of low intensity, 24 dB attenuation (0.010 cd s/m2). The dark-adapted mixed rod– cone responses were obtained using stimulation with flashes of maximum intensity, 0 dB (2.5 cd s/m2). The oscillatory potentials were recorded in dark adaptation using an interstimulus interval of 30 seconds and in response to stimulus flashes of maximal intensity. The cone responses were elicited in light adaptation (white background illumination; 480 lumen/m2) using maximum flash stimulation. The flicker ERGs (30 Hz) were recorded in light-adapted eyes with an averaging technique (n ⫽ 10) and in response to maximum intensity flashes. Amplitudes and peak times were compared with those previously determined in control subjects of the same age group. Parts of the electrophysiologic recordings (mixed rod– cone ERG responses) have been published elsewhere.5 The EOGs were recorded bitemporally with a UTAS-E 3000 (LKC Technologies Inc), then amplified and digitized at a sampling rate of 1000 Hz; measurements were performed in accordance with International Society for Clinical Electrophysiology of Vision recommendations.9
Results Clinical and Morphologic Findings The results of the present study are shown in Table 1. In comparison with the previously published ophthalmologic examinations,5 the older sister (JG, age 34) showed less advanced myopia, a slightly decreased visual acuity with best correction in both eyes, and bilateral unchanged thin central disc-shaped, zonularlike lens opacities. The younger sister (HG, age 31) showed a lessened degree of her moderate myopia, reduced visual acuity at the same level as before, and unchanged peripheral opacities around the lens nuclei. Funduscopy of both sisters revealed unchanged normal optic nerve heads and peripapillary atrophy bilaterally (Fig 1A, Fig 2A). The elder sister (JG) had maculopathy with essentially unchanged granular hyper- and hypopigmentations but now with a significant macular edema in the foveal zones of both eyes (Fig 1A). The younger sister (HG) presented bilateral maculopathy, now with an established edema that had not been present at the previous examination (Fig 2A). As before, she also had multiple white spots of atrophy in the temporal area of the anatomic macula in both eyes. Both sisters had granular, reticular, and bone spiculelike hyperpigmentations in the midperiphery, most pronounced in the nasal and upper parts of the fundi, as previously. Arteriovenous fluorescein angiograms, which visualize patterns of hyperfluorescence as a result of dye pooling in cystoid
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Figure 1. Fundus photo and fluorescein angiogram of the older sister (JG). A, Fundus photo showing maculopathy with macular edema and granular hyperpigmentations and hypopigmentations. B, Late arteriovenous phase fluorescin angiogram (8 minutes) showing cystoid hyperfluorescence in a petalloid pattern in the foveal area with a honeycomb pattern perifoveally. A diffuse, mottled background hyperfluorescence is present in the periphery of the retina. There is also a leakage at the vascular arcades and peripapillary.
spaces, showed a bilateral petalloid hyperfluorescence in the foveal area in both sisters (Fig 1B; Fig 2B), most distinctly in the younger of the two. Perifoveal changes with honeycomblike hyperfluorescence were present (except nasally) and became more apparent in the late phases of the angiograms (Fig 1B, Fig 2B, C, top frame). The arcade areas and optic disc showed leakage. There was also a diffuse, mottled background hyperfluorescence in the periphery of the retina. The OCT retinal maps in the older sister (JG) showed an increased thickness in the central parts (1- to 3-mm rings) of the macula, though the perifoveal (outer 6-mm ring) area thickness was within normal limits in both eyes (Table 1). The younger sister (HG) showed a more pronounced as well as a more general increased thickness of the macular region (all 1- to 6-mm rings), including the foveal as well as the adjacent perifoveal regions (Table 1; Fig 2C middle and bottom frames). On OCT vertical scan, the swelling involved almost the entire retinal thickness and was most obvious in the center of the foveal area (Fig 2C middle and bottom frames). As can be seen
Burstedt et al 䡠 Rode–Cone Dystrophy
Figure 2. Fundus photo, fluorescein angiograms and optical coherence tomography (OCT) scans of the younger sister (HG). A, Fundus photo showing maculopathy with central macular edema and retinal thinning in the centre of the fovea surrounded by swollen “edges.” Multiple white spots of atrophy are seen in the temporal area of the anatomic macula. B, Late arteriovenous phase angiogram (6 minutes) of the central area of the fundus, showing cystoid hyperfluorescence in a petalloid pattern in the fovea with perifoveal leakage of a honeycomb type. A diffuse, mottled background hyperfluorescence is present in the periphery of the retina. There is also a leakage at the vascular arcades and peripapillary. C, Top frame: The late-phase fluorescein angiogram (9 minutes) is shown to facilitate comparison. The angiogram of the central area of the fundus shows cystoid hyperfluorescence, in a petalloid pattern in the foveal area (inner circle) and a honeycomb pattern in the perifoveal area (outer circle). Middle and bottom frames: The OCT scans show reduced foveolar depression with large cystic spaces (white asterisk). Smaller cystic formations most evident perifoveally are seen in the inner nuclear layer (white arrow). There is also a retinal swelling of the outer plexiform layer (yellow asterisk). Over the central area with cystic spaces there is generally a thinning of the inner and outer layers of the retina.
in Figure 2C, the younger sister (HG) showed cystic spaces with low reflectivity of accumulated intraretinal fluid; the larger ones appeared centrally (middle frame) and the smaller ones more perifoveally (bottom frame). These cysts appeared in both the inner nuclear layer and the outer plexiform layer in both eyes of both sisters. The large cystic spaces of the OCT scans corresponded to the dye pooling in the angiograms. There were primarily small cystic spaces in the inner nuclear layer over the central foveal area. Thinning of the outer and inner retinal layers was also present.
Electrophysiologic Findings The results of the full-field single flash ERGs, including the oscillatory potentials, are shown in Figures 3 and 4. In the older
sister (JG), the amplitudes of the rod-isolated b-waves were subnormal in both eyes and the peak times were prolonged (Fig 3A). The rod-isolated b-wave was extinguished in both eyes of the younger sister (HG; Fig 4A). The amplitudes of the mixed rod– cone b-waves were subnormal in both sisters, and again the peak times were prolonged (Figs 3A and 4A). The amplitudes of the mixed rod– cone a-waves, the cone b-waves, and the 30-Hz flicker ERGs were low in the right eye and subnormal in the left eye of the older sister (Fig 3A). The younger sister showed subnormal values of the mixed rod– cone a-waves in the right eye, and the a-wave was nonrecordable in the left eye (Fig 4A). The cone b-waves and the 30-Hz flicker ERGs were subnormal in both eyes. The recorded mixed rod– cone a-waves and the cone b-waves were delayed with increased peak times in both sisters. In the older sister (JG), the summed amplitudes of the individual oscillatory peaks
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Figure 3. Electroretinograms (ERGs) and electrooculograms (EOGs) of the older sister (JG) and a normal control (49 years) old. A, Dark- and light-adapted ERG responses. B, The oscillatory responses of both eyes showing subnormal responses in the right eye and a nonrecordable response in the left eye. C, The EOGs show a normal response of the right eye (RE) and a subnormal value in the left eye (LE).
(interstimulus interval of 30 seconds) showed subnormal values in the right eye, and the oscillatory response was extinguished in the left eye (Fig 3B). There were no recordable oscillatory potentials in either eye in the younger sister (HG).
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The Arden ratio of the EOG of the older sister (JG) was normal (2.2) in the right eye but was subnormal (1.4) in the left (Fig 3C). Both eyes of the younger sister (HG) showed subnormal EOG values (1.6 and 1.3; Fig 4B).
Burstedt et al 䡠 Rode–Cone Dystrophy
Figure 4. Electroretinograms and electrooculograms of the younger sister (HG) and a normal control (49 years old). A, Dark- and light-adapted ERG responses. B, The EOG shows subnormal values in both eyes.
Discussion The causes and the development of retinal degeneration are not yet fully understood, although multiple factors are implicated, including oxidative stress. In GS deficiency, the levels of the scavenger GSH are extremely low, and reactive oxygen species production probably exceeds the antioxidant capacity of cells; this situation may lead to oxidative stress in tissues such as the retina. Both of the GS-deficient sisters in this study had subnormal visual acuity and significant bilateral macular edemas, which may explain why refractive
changes leading to less myopia were found. The cystic changes in the fovea and perifoveal areas, measured with OCT, were located predominately in the inner nuclear layer and the outer plexiform layer, which correlated with the angiograms. Similar findings have been described in diabetic macular edema.10 Although the 2 sisters had no family history of retinitis pigmentosa, their angiograms showed similarities to this condition. The cystic macular edema in hereditary retinal disorders is believed to be caused by alterations of the blood–retinal barrier, initially at the level of the retinal pigment epithelium (RPE) with leakage in the
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Ophthalmology Volume 116, Number 2, February 2009 macula, optic disc, and arcade areas,11 a situation also observed in these GS-deficient patients. Normally, GSH are enriched in RPE and Müller cells of the retina,12,13 which offer a large capacity for GSH synthesis. The findings of RPE affection in GS deficiency is supported by the mottled hyperfluorescent areas noted in these sisters, indicating multiple window defects in the RPE on the angiogram. The EOG findings may indicate the existence of degenerative changes of the RPE. However, EOG is almost always abnormal in the presence of significant ERG abnormalities, which was the case in the sisters. An interesting observation is the preserved EOG results in 1 eye of the elder sister, which seems to contradict the theory that the retinal degeneration in GS deficiency is primarily due to problems with the RPE. Furthermore, the fluorescein dye accumulation without cystic spaces suggested that there had been some effect on the glial (Müller) cells,14 and the failure of the Müller cell GSH synthesis may be especially harmful to the entire retina.15 The electrophysiologic results with low or nonrecordable rod-isolated b-waves and mixed rod– cone b-waves of the ERG indicate reduced amount of and/or lost rod photoreceptor function as well as affected Müller cells and/or ON (pathway of activity of bipolar cells) bipolar cells, the latter being second-order neurons in the on-pathway in the neuroretina.16 –19 The cone responses were also found to be markedly reduced. Thus, the progress of retinal dystrophy in GS deficiency involves the function of the cones, which are the major photoreceptors in the foveal area. This supports the morphologic changes with pronounced edema in the cone-rich macula region and the affected visual acuity, reflecting a disturbed function of the cones in the fovea. The findings also corroborate the view that retinal dystrophy in GS deficiency belongs to the rod– cone dystrophy group affecting the rods and subsequently the cones.4,5 The summed amplitudes of the individual oscillatory peaks showed subnormal to nonrecordable values in both patients, reflecting affected and/or extinguished neuronal activity at the level of the inner plexiform layer.20 The oscillatory potentials represent both scotopic and photopic activities or an interaction between the rods and cones,20,21 and a loss of the rapid oscillatory activity of the ERG is therefore expected because there was not only pronounced reduction of rod function, but also diminished cone activity. Furthermore, because there are metabolic disturbances (acidosis) present in the sisters, one cannot exclude an effect of acidic transient and acid-sensing ion channels in the retina, which have recently been suggested as being involved in the generation of the oscillatory response.21,22 Also of interest is the partially preserved function of the right eye of the older sister, which had better visual acuity and less affected electrophysiologic cone-derived responses than the other eye. However, the funduscopy, angiography, and OCT findings showed no obvious differences between the eyes except a somewhat larger foveal avascular zone in the less-preserved eye. Finally, we suggest that these findings comprise an in vivo model for studies of the retinal response to oxidative stress in humans. The retina is an accessible part of the brain; it is easily observed by ophthalmoscopy, its morphology can be readily analyzed at a comparatively high reso-
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lution level, and its function can be reliably tested both subjectively and objectively. In summary, 2 sisters with severe GS deficiency developed maculopathy with bilateral macular edema in early adulthood in addition to previous peripheral and subtle macular degenerative changes of the retina. Induced apoptosis of the photoreceptors and neural dysfunction at the level of the inner plexiform layer both seem to be associated with GS deficiency. Our findings corroborate the view that retinal dystrophy in GS deficiency is of the rod– cone type, and also present a good in vivo model for studying the effect of oxidative stress of the retina in humans. Acknowledgment. The authors thank Dr P. Pohjanen, Sweden, for referring these patients.
References 1. Handelman GJ, Dratz EA. The role of antioxidants in the retina and retinal pigment epithelium and the nature of prooxidantinduced damage. Adv Free Radic Biol Med 1986;2:1– 89. 2. Dringen R, Gutterer JM, Hirrlinger J. Glutathione metabolism in brain metabolic interaction between astrocytes and neurons in the defense against reactive oxygen species. Eur J Biochem 2000;267:4912– 6. 3. Ristoff E, Mayatepek E, Larsson A. Long-term clinical outcome in patients with glutathione synthetase deficiency. J Pediatr 2001;139:79 – 84. 4. Larsson A, Wachtmeister L, von Wendt L, et al. Ophthalmological, psychometric and therapeutic investigation in two sisters with hereditary glutathione synthetase deficiency (5oxoprolinuria). Neuropediatrics 1985;16:131– 6. 5. Ristoff E, Burstedt M, Larsson A, Wachtmeister L. Progressive retinal dystrophy in two sisters with glutathione synthetase (GS) deficiency. J Inherit Metab Dis 2007;30:102. 6. Dahl N, Pigg M, Ristoff E, et al. Missense mutations in the human glutathione synthetase gene result in severe metabolic acidosis, 5-oxoprolinuria, hemolytic anemia and neurological dysfunction. Hum Mol Genet 1997;6:1147–52. 7. Ristoff E, Augustson C, Larsson A. Generalized glutathione synthetase deficiency and pregnancy. J Inherit Metab Dis 1999;22:758 –9. 8. Marmor MF, Holder GE, Seeliger MW, Yamamoto S. Standard for clinical electroretinography (2004 update). Doc Ophthalmol 2004;108:107–14. 9. Brown M, Marmor M, Vaegan, et al. ISCEV standard for clinical electro-oculography (EOG) 2006. Doc Ophthalmol 2006;113:205–12. 10. Otani T, Kishi S. Correlation between optical coherence tomography and fluorescein angiography findings in diabetic macular edema. Ophthalmology 2007;114:104 –7. 11. Newsome DA. Retinal fluorescein leakage in retinitis pigmentosa. Am J Ophthalmol 1986;101:354 – 60. 12. Pow DV, Crook DK. Immunocytochemical evidence for the presence of high levels of reduced glutathione in radial glial cells and horizontal cells in the rabbit retina. Neurosci Lett 1995;193:25– 8. 13. Huster D, Hjelle OP, Haug FM, et al. Subcellular compartmentation of glutathione and glutathione precursors: a high resolution immunogold analysis of the outer retina of guinea pig. Anat Embryol (Berl) 1998;198:277– 87. 14. Fine BS, Brucker AJ. Macular edema and cystoid macular edema. Am J Ophthalmol 1981;92:466 – 81.
Burstedt et al 䡠 Rode–Cone Dystrophy 15. Huster D, Reichenbach A, Reichelt W. The glutathione content of retinal Müller (glial) cells: effect of pathological conditions. Neurochem Int 2000;36:461–9. 16. Miller RF, Dowling JE. Intracellular responses of the Müller (glial) cells of mudpuppy retina: their relation to b-wave of the electroretinogram. J Neurophysiol 1970;33:323– 41. 17. Xu X, Karwoski CJ. Current source density analysis of retinal field potentials. II. Pharmacological analysis of the b-wave and M-wave. J Neurophysiol 1994;72:96 –105. 18. Rangaswamy NV, Hood DC, Frishman LJ. Regional variations in local contributions to the primate photopic flash ERG: revealed using the slow-sequence mfERG. Invest Ophthalmol Vis Sci 2003;44:3233– 47.
19. Friedburg C, Allen CP, Mason PJ, Lamb TD. Contribution of cone photoreceptors and post-receptoral mechanisms to the human photopic electroretinogram. J Physiol 2004;556:819 – 34. 20. Wachtmeister L. Oscillatory potentials in the retina: what do they reveal. Prog Retin Eye Res 1998;17:485–521. 21. Lundstrom AL, Wang L, Wachtmeister L. Neuronal adaptation in the human retina: a study of the single oscillatory response in dark adaptation and mesopic background illumination. Acta Ophthalmol Scand 2007;85:756 – 63. 22. Ettaiche M, Deval E, Cougnon M, et al. Silencing acid-sensing ion channel 1a alters cone-mediated retinal function. J Neurosci 2006;26:5800 –9.
Footnotes and Financial Disclosures Originally received: April 28, 2008. Final revision: August 20, 2008. Accepted: September 4, 2008.
Manuscript no. 2008-522.
1
Department of Clinical Sciences/Ophthalmology, University of Umeå, Umeå, Sweden.
2
Department of Pediatrics, Karolinska Institutet, Stockholm, Sweden.
Financial Disclosure(s): The authors have no proprietary or commercial interest in any materials discussed in this paper.
Supported by the grants from the Research Foundation of Sight Preferment, Crown Princess Margaretha’s Foundation for Vision Research (KMA), the Kempe Foundation, the Swedish Medical Society, the Swedish Research Council (4792), the Free Masons in Stockholm for Children’s Welfare, the HRH Crown Princess Lovisa Foundation, the Åke Wiberg Foundation, the Lennanders Foundation, and the Samariten Foundation, Sweden. Correspondence: Marie Burstedt, MD, PhD, Department of Clinical Sciences/Ophthalmology, Umeå University, SE-901 85 Umeå, Sweden. E-mail: Marie.Burstedt@ ophthal.umu.se.
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The retina is among the most vascularized tissues in the body, and has one of the highest oxidative metabolic rates per tissue weight. The high content of polyunsaturated fatty acids in the photoreceptor outer segments also makes the photoreceptors susceptible to damage if high levels of reactive oxygen species are present. In addition, the retina is exposed to light, which promotes the generation of cytotoxic reactive oxygen species, which cause dysfunction and cell death in photoreceptors and other retinal cells. Factors such as high oxygen flux, light, and polyunsaturated fatty acids induce a high susceptibility of the retina to oxidative stress. Protection against this oxidative stress in the retina is provided by a complex defense mechanism involving the major water-soluble antioxidant glutathione (GSH), which is also found in the rest of the eye.1 This antioxidant functions primarily in the cytoplasm and mitochondria, and participates in many of the antioxidative cellular mechanisms that are essential for cellular survival,2 such as detoxification of xenobiotics and regulation of several cascades in cellular signaling. Glutathione synthetase (GS) deficiency is a rare, autosomalrecessive inborn error of metabolism. Patients with GS de-
324
© 2009 by the American Academy of Ophthalmology Published by Elsevier Inc.
ficiency have low levels of cellular GSH (5%–20% of the normal mean in erythrocytes and fibroblasts; McKusick #266130). According to the severity of the clinical signs, patients with GS deficiency are divided into 3 subgroups: mild, moderate, or severe GS deficiency. Low levels of GSH result in hemolytic anemia; those with moderate disease also exhibit metabolic acidosis. Patients with severe GS deficiency experience progressive neurologic symptoms with retarded psychomotor development, seizures, spasticity, and cerebellar symptoms. In about 25% of patients with GS deficiency, the eyes are affected with retinitis pigmentosa, retinal dystrophy, crystalline opacities in the lens, and decreased visual acuity.3 Because the retina can be considered a part of the central nervous system, patients with retinal changes are classified as having a severe disease. We have previously reported on 2 sisters with severe GS deficiency who were investigated ophthalmologically as children and then reexamined 20 years later.4,5 In adulthood, they had developed moderate to advanced myopia, slowly progressing sparse lens opacities, and symptoms of defective macular function with modestly decreased visual acuISSN 0161-6420/09/$–see front matter doi:10.1016/j.ophtha.2008.09.007
Burstedt et al 䡠 Rode–Cone Dystrophy ity. The elder sister showed maculopathy with granular hyper- and hypopigmentations in both eyes and the younger sister showed bilateral maculopathy and multiple white spots of atrophy in the temporal area of the anatomic macula. Both sisters showed peripheral degenerative changes with granular, reticular, and bone spiculelike hyperpigmentations in the midperiphery that were more pronounced in the nasal and upper parts of the fundi, corresponding to defects in the visual field. In a previous study5 of the 2 sisters, the central visual pathways were found to be preserved with normal visual evoked potential responses, although the full-field electroretinograms (ERGs) of both sisters were even further affected compared to childhood, more than 20 years earlier.4 Thus, the retinal dystrophy in severe types of GS deficiency seems to advance, albeit slowly.5 We have also reported on 2 brothers with severe GS deficiency who presented retinitis pigmentosa with nearly extinguished ERG in adolescence, and a 3rd young boy with signs of retinal dystrophy and lowered visual acuity at the age of 10 years.3 Thus, there is limited information about the retinal findings in this small but important group of patients who, despite aggravating metabolic effect, can reach adulthood and, in addition to possible neurologic symptoms, develop retinal disease, which may lead to severe visual handicap. This study presents an analysis of retinal function as well as a closer evaluation of the retinal morphology in 2 sisters with severe GS deficiency, using full-field ERGs, electrooculograms (EOGs), fluorescein angiograms, and optical coherence tomography (OCT). We hypothesize that there is a relationship between low levels of GSH in the retina and progressive eye symptoms in GS-deficient patients.
Patients and Methods Patients Two sisters with severe GS deficiency were born to healthy, nonconsanguineous parents. The elder sister was diagnosed with GS deficiency at the age of 1 year and the younger at the age of 1 day. In the neonatal period, both showed characteristic signs of GS deficiency, namely, hemolytic anemia, metabolic acidosis, and
5-oxoprolinuria (14 –20 mol/mol creatinine; control range ⬍0.1 mol/mol creatinine). The GSH levels in erythrocytes were ⬍10% of the normal mean, GS (enzyme commission number [EC] 6.3.2.3) activity was ⬍5% of the normal mean, and GS activity in fibroblasts was ⬍20% of the normal mean. Both patients have been described previously.4 The sisters also showed a 1.4-fold to 4.5-fold increase in the levels of ␥-glutamylcysteine and cysteine in cultured fibroblasts (unpublished results). Other antioxidative enzymes (superoxide dismutase, catalase, and GSH peroxidase) had activities within normal limits in erythrocytes. Both sisters were homozygous for the mutation 77C¡A, Ala26Asp in the GS gene.6 Treatment with acidosis correction was started in the neonatal period. Vitamin E supplementation (10 mg/kg per day) was started before puberty, and vitamin C supplementation (100 mg/kg per day) when the sisters were in their 20s. Both sisters completed high school and found employment. The younger sister had a normal pregnancy and gave birth to a healthy, heterozygous child.7 When tested as adults, their intelligence quotients were within normal limits. Neurologic examination showed brisk stretch reflexes. The sisters had well-compensated hemolytic anemia, metabolic acidosis, and slightly elevated triglycerides, and both had a normal blood pressure. When examined for the present study, the sisters were 31 and 34 years old. The studies of the physiologic consequences of GSH deficiency were approved by the Ethics Committee of Karolinska Institutet, Stockholm, Sweden. The ophthalmologic investigations were considered as routine health care and performed according to standardized protocols and procedures. Informed consent was obtained, and the study followed the tenets of the Declaration of Helsinki.
Clinical and Morphologic Methods Routine clinical ophthalmologic methods were used to examine the anterior and posterior segments of both eyes of the 2 sisters. Monocular visual acuity was tested with the Early Treatment Diabetic Retinopathy Study chart at a distance of 4 m. Lighting conditions were standardized using an Early Treatment Diabetic Retinopathy Study chart illuminator cabinet (cat. no. 2425, Precision Vision). The time elapsed between the first clinical examination including electroretinography and second one including morphologic investigations and electrooculography was within 1 year (the older sister [JG] at 33 and 34 years of age, respectively, and the younger sister [HG] at 30 and 31 years of age, respectively). Fluorescein angiograms of the retinas and
Table 1. Macular Thickness Measurements Using the Stratus Optical Coherence Tomography (OCT3) Retinal Thickness (m) Inner Ring (Ø 3 mm)
Outer Ring (Ø 6 mm)
Right Eye
Left Eye
Right Eye
Left Eye
Right Eye
Left Eye
Right Eye
Left Eye
S301 I270 T289 N279 S456 I377 T412 N416
S364 I334 T354 N214 S410 I376 T399 N393
S235 I209 T215 N239 S264 I254 T244 N270
S238 I189 T222 N214 S247 I261 T252 N254
VA (LogMAR) Case/Age (yrs)
Refraction
Central (Ø 1 mm)
JG/34
⫺6.5 — 1.75 ⫻10 ⫺8.25 — 2.0 ⫻175
0.02
0.24
326
439
HG/31
⫺4.25 — 1 ⫻10 ⫺4.75 — 0.75 ⫻180
0.22
0.22
534
475
I ⫽ inferior; LogMAR ⫽ logarithm of minimum angle of resolution; N ⫽ nasal; S ⫽ superior; T ⫽ temporal; VA ⫽ visual acuity.
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Ophthalmology Volume 116, Number 2, February 2009 photographs of the fundus were taken and analyzed according to standard clinical procedures. The OCT images were examined with Stratus OCT, Version. 3.0 (Carl Zeiss Meditec AG, Oberkochen, Germany). Both eyes of the sisters were examined using 6-mm vertical scan lengths including the fovea and the adjacent perifoveal region. Late-phase angiograms (6 –9 minutes after injection) were compared with the corresponding cross-sectional OCT images.
Electrophysiologic Methods Full-field, single-flash, and flicker ERGs, including the oscillatory potentials, were recorded with a UTAS-E 2000 (LKC Technologies Inc, Gaithersburg, MD) using Burian–Allen bipolar electrodes and according to the recommendations of the International Society for Clinical Electrophysiology of Vision.8 The isolated rod responses were measured during standard dark-adapted conditions (20 minutes) using full-field white flashes of low intensity, 24 dB attenuation (0.010 cd s/m2). The dark-adapted mixed rod– cone responses were obtained using stimulation with flashes of maximum intensity, 0 dB (2.5 cd s/m2). The oscillatory potentials were recorded in dark adaptation using an interstimulus interval of 30 seconds and in response to stimulus flashes of maximal intensity. The cone responses were elicited in light adaptation (white background illumination; 480 lumen/m2) using maximum flash stimulation. The flicker ERGs (30 Hz) were recorded in light-adapted eyes with an averaging technique (n ⫽ 10) and in response to maximum intensity flashes. Amplitudes and peak times were compared with those previously determined in control subjects of the same age group. Parts of the electrophysiologic recordings (mixed rod– cone ERG responses) have been published elsewhere.5 The EOGs were recorded bitemporally with a UTAS-E 3000 (LKC Technologies Inc), then amplified and digitized at a sampling rate of 1000 Hz; measurements were performed in accordance with International Society for Clinical Electrophysiology of Vision recommendations.9
Results Clinical and Morphologic Findings The results of the present study are shown in Table 1. In comparison with the previously published ophthalmologic examinations,5 the older sister (JG, age 34) showed less advanced myopia, a slightly decreased visual acuity with best correction in both eyes, and bilateral unchanged thin central disc-shaped, zonularlike lens opacities. The younger sister (HG, age 31) showed a lessened degree of her moderate myopia, reduced visual acuity at the same level as before, and unchanged peripheral opacities around the lens nuclei. Funduscopy of both sisters revealed unchanged normal optic nerve heads and peripapillary atrophy bilaterally (Fig 1A, Fig 2A). The elder sister (JG) had maculopathy with essentially unchanged granular hyper- and hypopigmentations but now with a significant macular edema in the foveal zones of both eyes (Fig 1A). The younger sister (HG) presented bilateral maculopathy, now with an established edema that had not been present at the previous examination (Fig 2A). As before, she also had multiple white spots of atrophy in the temporal area of the anatomic macula in both eyes. Both sisters had granular, reticular, and bone spiculelike hyperpigmentations in the midperiphery, most pronounced in the nasal and upper parts of the fundi, as previously. Arteriovenous fluorescein angiograms, which visualize patterns of hyperfluorescence as a result of dye pooling in cystoid
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Figure 1. Fundus photo and fluorescein angiogram of the older sister (JG). A, Fundus photo showing maculopathy with macular edema and granular hyperpigmentations and hypopigmentations. B, Late arteriovenous phase fluorescin angiogram (8 minutes) showing cystoid hyperfluorescence in a petalloid pattern in the foveal area with a honeycomb pattern perifoveally. A diffuse, mottled background hyperfluorescence is present in the periphery of the retina. There is also a leakage at the vascular arcades and peripapillary.
spaces, showed a bilateral petalloid hyperfluorescence in the foveal area in both sisters (Fig 1B; Fig 2B), most distinctly in the younger of the two. Perifoveal changes with honeycomblike hyperfluorescence were present (except nasally) and became more apparent in the late phases of the angiograms (Fig 1B, Fig 2B, C, top frame). The arcade areas and optic disc showed leakage. There was also a diffuse, mottled background hyperfluorescence in the periphery of the retina. The OCT retinal maps in the older sister (JG) showed an increased thickness in the central parts (1- to 3-mm rings) of the macula, though the perifoveal (outer 6-mm ring) area thickness was within normal limits in both eyes (Table 1). The younger sister (HG) showed a more pronounced as well as a more general increased thickness of the macular region (all 1- to 6-mm rings), including the foveal as well as the adjacent perifoveal regions (Table 1; Fig 2C middle and bottom frames). On OCT vertical scan, the swelling involved almost the entire retinal thickness and was most obvious in the center of the foveal area (Fig 2C middle and bottom frames). As can be seen
Burstedt et al 䡠 Rode–Cone Dystrophy
Figure 2. Fundus photo, fluorescein angiograms and optical coherence tomography (OCT) scans of the younger sister (HG). A, Fundus photo showing maculopathy with central macular edema and retinal thinning in the centre of the fovea surrounded by swollen “edges.” Multiple white spots of atrophy are seen in the temporal area of the anatomic macula. B, Late arteriovenous phase angiogram (6 minutes) of the central area of the fundus, showing cystoid hyperfluorescence in a petalloid pattern in the fovea with perifoveal leakage of a honeycomb type. A diffuse, mottled background hyperfluorescence is present in the periphery of the retina. There is also a leakage at the vascular arcades and peripapillary. C, Top frame: The late-phase fluorescein angiogram (9 minutes) is shown to facilitate comparison. The angiogram of the central area of the fundus shows cystoid hyperfluorescence, in a petalloid pattern in the foveal area (inner circle) and a honeycomb pattern in the perifoveal area (outer circle). Middle and bottom frames: The OCT scans show reduced foveolar depression with large cystic spaces (white asterisk). Smaller cystic formations most evident perifoveally are seen in the inner nuclear layer (white arrow). There is also a retinal swelling of the outer plexiform layer (yellow asterisk). Over the central area with cystic spaces there is generally a thinning of the inner and outer layers of the retina.
in Figure 2C, the younger sister (HG) showed cystic spaces with low reflectivity of accumulated intraretinal fluid; the larger ones appeared centrally (middle frame) and the smaller ones more perifoveally (bottom frame). These cysts appeared in both the inner nuclear layer and the outer plexiform layer in both eyes of both sisters. The large cystic spaces of the OCT scans corresponded to the dye pooling in the angiograms. There were primarily small cystic spaces in the inner nuclear layer over the central foveal area. Thinning of the outer and inner retinal layers was also present.
Electrophysiologic Findings The results of the full-field single flash ERGs, including the oscillatory potentials, are shown in Figures 3 and 4. In the older
sister (JG), the amplitudes of the rod-isolated b-waves were subnormal in both eyes and the peak times were prolonged (Fig 3A). The rod-isolated b-wave was extinguished in both eyes of the younger sister (HG; Fig 4A). The amplitudes of the mixed rod– cone b-waves were subnormal in both sisters, and again the peak times were prolonged (Figs 3A and 4A). The amplitudes of the mixed rod– cone a-waves, the cone b-waves, and the 30-Hz flicker ERGs were low in the right eye and subnormal in the left eye of the older sister (Fig 3A). The younger sister showed subnormal values of the mixed rod– cone a-waves in the right eye, and the a-wave was nonrecordable in the left eye (Fig 4A). The cone b-waves and the 30-Hz flicker ERGs were subnormal in both eyes. The recorded mixed rod– cone a-waves and the cone b-waves were delayed with increased peak times in both sisters. In the older sister (JG), the summed amplitudes of the individual oscillatory peaks
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Ophthalmology Volume 116, Number 2, February 2009
Figure 3. Electroretinograms (ERGs) and electrooculograms (EOGs) of the older sister (JG) and a normal control (49 years) old. A, Dark- and light-adapted ERG responses. B, The oscillatory responses of both eyes showing subnormal responses in the right eye and a nonrecordable response in the left eye. C, The EOGs show a normal response of the right eye (RE) and a subnormal value in the left eye (LE).
(interstimulus interval of 30 seconds) showed subnormal values in the right eye, and the oscillatory response was extinguished in the left eye (Fig 3B). There were no recordable oscillatory potentials in either eye in the younger sister (HG).
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The Arden ratio of the EOG of the older sister (JG) was normal (2.2) in the right eye but was subnormal (1.4) in the left (Fig 3C). Both eyes of the younger sister (HG) showed subnormal EOG values (1.6 and 1.3; Fig 4B).
Burstedt et al 䡠 Rode–Cone Dystrophy
Figure 4. Electroretinograms and electrooculograms of the younger sister (HG) and a normal control (49 years old). A, Dark- and light-adapted ERG responses. B, The EOG shows subnormal values in both eyes.
Discussion The causes and the development of retinal degeneration are not yet fully understood, although multiple factors are implicated, including oxidative stress. In GS deficiency, the levels of the scavenger GSH are extremely low, and reactive oxygen species production probably exceeds the antioxidant capacity of cells; this situation may lead to oxidative stress in tissues such as the retina. Both of the GS-deficient sisters in this study had subnormal visual acuity and significant bilateral macular edemas, which may explain why refractive
changes leading to less myopia were found. The cystic changes in the fovea and perifoveal areas, measured with OCT, were located predominately in the inner nuclear layer and the outer plexiform layer, which correlated with the angiograms. Similar findings have been described in diabetic macular edema.10 Although the 2 sisters had no family history of retinitis pigmentosa, their angiograms showed similarities to this condition. The cystic macular edema in hereditary retinal disorders is believed to be caused by alterations of the blood–retinal barrier, initially at the level of the retinal pigment epithelium (RPE) with leakage in the
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Ophthalmology Volume 116, Number 2, February 2009 macula, optic disc, and arcade areas,11 a situation also observed in these GS-deficient patients. Normally, GSH are enriched in RPE and Müller cells of the retina,12,13 which offer a large capacity for GSH synthesis. The findings of RPE affection in GS deficiency is supported by the mottled hyperfluorescent areas noted in these sisters, indicating multiple window defects in the RPE on the angiogram. The EOG findings may indicate the existence of degenerative changes of the RPE. However, EOG is almost always abnormal in the presence of significant ERG abnormalities, which was the case in the sisters. An interesting observation is the preserved EOG results in 1 eye of the elder sister, which seems to contradict the theory that the retinal degeneration in GS deficiency is primarily due to problems with the RPE. Furthermore, the fluorescein dye accumulation without cystic spaces suggested that there had been some effect on the glial (Müller) cells,14 and the failure of the Müller cell GSH synthesis may be especially harmful to the entire retina.15 The electrophysiologic results with low or nonrecordable rod-isolated b-waves and mixed rod– cone b-waves of the ERG indicate reduced amount of and/or lost rod photoreceptor function as well as affected Müller cells and/or ON (pathway of activity of bipolar cells) bipolar cells, the latter being second-order neurons in the on-pathway in the neuroretina.16 –19 The cone responses were also found to be markedly reduced. Thus, the progress of retinal dystrophy in GS deficiency involves the function of the cones, which are the major photoreceptors in the foveal area. This supports the morphologic changes with pronounced edema in the cone-rich macula region and the affected visual acuity, reflecting a disturbed function of the cones in the fovea. The findings also corroborate the view that retinal dystrophy in GS deficiency belongs to the rod– cone dystrophy group affecting the rods and subsequently the cones.4,5 The summed amplitudes of the individual oscillatory peaks showed subnormal to nonrecordable values in both patients, reflecting affected and/or extinguished neuronal activity at the level of the inner plexiform layer.20 The oscillatory potentials represent both scotopic and photopic activities or an interaction between the rods and cones,20,21 and a loss of the rapid oscillatory activity of the ERG is therefore expected because there was not only pronounced reduction of rod function, but also diminished cone activity. Furthermore, because there are metabolic disturbances (acidosis) present in the sisters, one cannot exclude an effect of acidic transient and acid-sensing ion channels in the retina, which have recently been suggested as being involved in the generation of the oscillatory response.21,22 Also of interest is the partially preserved function of the right eye of the older sister, which had better visual acuity and less affected electrophysiologic cone-derived responses than the other eye. However, the funduscopy, angiography, and OCT findings showed no obvious differences between the eyes except a somewhat larger foveal avascular zone in the less-preserved eye. Finally, we suggest that these findings comprise an in vivo model for studies of the retinal response to oxidative stress in humans. The retina is an accessible part of the brain; it is easily observed by ophthalmoscopy, its morphology can be readily analyzed at a comparatively high reso-
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lution level, and its function can be reliably tested both subjectively and objectively. In summary, 2 sisters with severe GS deficiency developed maculopathy with bilateral macular edema in early adulthood in addition to previous peripheral and subtle macular degenerative changes of the retina. Induced apoptosis of the photoreceptors and neural dysfunction at the level of the inner plexiform layer both seem to be associated with GS deficiency. Our findings corroborate the view that retinal dystrophy in GS deficiency is of the rod– cone type, and also present a good in vivo model for studying the effect of oxidative stress of the retina in humans. Acknowledgment. The authors thank Dr P. Pohjanen, Sweden, for referring these patients.
References 1. Handelman GJ, Dratz EA. The role of antioxidants in the retina and retinal pigment epithelium and the nature of prooxidantinduced damage. Adv Free Radic Biol Med 1986;2:1– 89. 2. Dringen R, Gutterer JM, Hirrlinger J. Glutathione metabolism in brain metabolic interaction between astrocytes and neurons in the defense against reactive oxygen species. Eur J Biochem 2000;267:4912– 6. 3. Ristoff E, Mayatepek E, Larsson A. Long-term clinical outcome in patients with glutathione synthetase deficiency. J Pediatr 2001;139:79 – 84. 4. Larsson A, Wachtmeister L, von Wendt L, et al. Ophthalmological, psychometric and therapeutic investigation in two sisters with hereditary glutathione synthetase deficiency (5oxoprolinuria). Neuropediatrics 1985;16:131– 6. 5. Ristoff E, Burstedt M, Larsson A, Wachtmeister L. Progressive retinal dystrophy in two sisters with glutathione synthetase (GS) deficiency. J Inherit Metab Dis 2007;30:102. 6. Dahl N, Pigg M, Ristoff E, et al. Missense mutations in the human glutathione synthetase gene result in severe metabolic acidosis, 5-oxoprolinuria, hemolytic anemia and neurological dysfunction. Hum Mol Genet 1997;6:1147–52. 7. Ristoff E, Augustson C, Larsson A. Generalized glutathione synthetase deficiency and pregnancy. J Inherit Metab Dis 1999;22:758 –9. 8. Marmor MF, Holder GE, Seeliger MW, Yamamoto S. Standard for clinical electroretinography (2004 update). Doc Ophthalmol 2004;108:107–14. 9. Brown M, Marmor M, Vaegan, et al. ISCEV standard for clinical electro-oculography (EOG) 2006. Doc Ophthalmol 2006;113:205–12. 10. Otani T, Kishi S. Correlation between optical coherence tomography and fluorescein angiography findings in diabetic macular edema. Ophthalmology 2007;114:104 –7. 11. Newsome DA. Retinal fluorescein leakage in retinitis pigmentosa. Am J Ophthalmol 1986;101:354 – 60. 12. Pow DV, Crook DK. Immunocytochemical evidence for the presence of high levels of reduced glutathione in radial glial cells and horizontal cells in the rabbit retina. Neurosci Lett 1995;193:25– 8. 13. Huster D, Hjelle OP, Haug FM, et al. Subcellular compartmentation of glutathione and glutathione precursors: a high resolution immunogold analysis of the outer retina of guinea pig. Anat Embryol (Berl) 1998;198:277– 87. 14. Fine BS, Brucker AJ. Macular edema and cystoid macular edema. Am J Ophthalmol 1981;92:466 – 81.
Burstedt et al 䡠 Rode–Cone Dystrophy 15. Huster D, Reichenbach A, Reichelt W. The glutathione content of retinal Müller (glial) cells: effect of pathological conditions. Neurochem Int 2000;36:461–9. 16. Miller RF, Dowling JE. Intracellular responses of the Müller (glial) cells of mudpuppy retina: their relation to b-wave of the electroretinogram. J Neurophysiol 1970;33:323– 41. 17. Xu X, Karwoski CJ. Current source density analysis of retinal field potentials. II. Pharmacological analysis of the b-wave and M-wave. J Neurophysiol 1994;72:96 –105. 18. Rangaswamy NV, Hood DC, Frishman LJ. Regional variations in local contributions to the primate photopic flash ERG: revealed using the slow-sequence mfERG. Invest Ophthalmol Vis Sci 2003;44:3233– 47.
19. Friedburg C, Allen CP, Mason PJ, Lamb TD. Contribution of cone photoreceptors and post-receptoral mechanisms to the human photopic electroretinogram. J Physiol 2004;556:819 – 34. 20. Wachtmeister L. Oscillatory potentials in the retina: what do they reveal. Prog Retin Eye Res 1998;17:485–521. 21. Lundstrom AL, Wang L, Wachtmeister L. Neuronal adaptation in the human retina: a study of the single oscillatory response in dark adaptation and mesopic background illumination. Acta Ophthalmol Scand 2007;85:756 – 63. 22. Ettaiche M, Deval E, Cougnon M, et al. Silencing acid-sensing ion channel 1a alters cone-mediated retinal function. J Neurosci 2006;26:5800 –9.
Footnotes and Financial Disclosures Originally received: April 28, 2008. Final revision: August 20, 2008. Accepted: September 4, 2008.
Manuscript no. 2008-522.
1
Department of Clinical Sciences/Ophthalmology, University of Umeå, Umeå, Sweden.
2
Department of Pediatrics, Karolinska Institutet, Stockholm, Sweden.
Financial Disclosure(s): The authors have no proprietary or commercial interest in any materials discussed in this paper.
Supported by the grants from the Research Foundation of Sight Preferment, Crown Princess Margaretha’s Foundation for Vision Research (KMA), the Kempe Foundation, the Swedish Medical Society, the Swedish Research Council (4792), the Free Masons in Stockholm for Children’s Welfare, the HRH Crown Princess Lovisa Foundation, the Åke Wiberg Foundation, the Lennanders Foundation, and the Samariten Foundation, Sweden. Correspondence: Marie Burstedt, MD, PhD, Department of Clinical Sciences/Ophthalmology, Umeå University, SE-901 85 Umeå, Sweden. E-mail: Marie.Burstedt@ ophthal.umu.se.
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