Marius Gradianu

Retinal Ion Regulation in a Mouse Model of Diabetic Retinopathy: Natural History and the Effect of Cu/Zn Superoxide Dismutase Overexpression

PURPOSE To test the hypotheses that manganese-enhanced MRI (MEMRI) is useful in evaluating intraretinal ion dysregulation in wild-type (WT) and Cu/Zn superoxide dismutase (SOD1) overexpressor mice. METHODS Central intraretinal ion activity and retinal thickness were measured from high-resolution data of light- and dark-adapted WT C57BL/6 mice (to gauge MEMRI sensitivity to normal visual processing in mice) and dark-adapted diabetic and nondiabetic WT and Cu/Zn superoxide dismutase overexpressor (SOD1OE) mice. Glycated hemoglobin and retinal vascular histopathology were also determined. RESULTS In WT mice, light adaptation reduced outer retinal manganese uptake compared with that in dark adaptation; no effect on inner retinal uptake was found. In diabetic WT mice, intraretinal manganese uptake became subnormal between 1.5 and 4 months of diabetes onset and then relatively increased. Central retinal thickness, as determined with MEMRI, decreased as a function of age in diabetic mice but remained constant in control mice. Nondiabetic SOD1OE mice had normal retinal manganese uptake but subnormal retinal thickness and supernormal acellular capillary density. At 4.2 months of diabetes, SOD1OE mice had normal manganese uptake and no further thinning; acellular capillaries frequency did not increase by 9 to 10 months of diabetes. CONCLUSIONS In emerging diabetic retinopathy, MEMRI provided an analytic measure of an ionic dysregulatory pattern that was sensitive to SOD1 overexpression. The potential benefit of SOD1 overexpression to inhibit retinal abnormality in this model is limited by the retinal and vascular degeneration that develops independently of diabetes.

Manganese-Enhanced MRI of the DBA/2J Mouse Model of Hereditary Glaucoma

PURPOSE To test the hypothesis that manganese-enhanced magnetic resonance imaging (MEMRI) is a sensitive approach for measuring of age-related ocular changes in experimental pigmentary glaucoma. METHODS Four groups of light-adapted mice were studied using MEMRI: young (2-3 months), C57BL/6 (negative controls), and DBA/2J mice and aged (10-11 months) C57BL/6 and DBA/2J mice. In all mice, eye perimeter, optic nerve head width, iridocorneal angle, ciliary body area, and total and inner retinal thickness, and a surrogate of retinal ion regulation (intraretinal uptake of manganese) were assessed from MEMRI data and compared. Axon counts were obtained from optic nerves harvested from MEMRI-assessed eyes. RESULTS As the C57BL/6 and DBA/2J mice aged, differential and significant changes in ocular perimeter, retinal thickness, iridocorneal angle, ciliary body area, and optic nerve head width were readily measured from MEMRI data (P < 0.05). In C57BL/6 mice, only inner retinal thickness and perimeter were correlated. In DBA/2J mice, ocular perimeter was correlated with total and inner retinal thickness, ciliary body area, optic nerve head width, and iridocorneal angle. Comparison of young and aged mice revealed a subnormal intraretinal manganese uptake (P < 0.05) in aged DBA/2J mice, but not in aged C57BL/6 mice. Manganese uptake did not correlate with the ocular perimeter. Axon density in the optic nerve correlated with MEMRI-measured optic nerve head width (P < 0.05). CONCLUSIONS These studies provide a baseline of noninvasive MEMRI-detectable changes associated with age in a common animal model of hereditary glaucoma that may be useful in the longitudinal evaluation of therapeutic success.

Quantitative mapping of ion channel regulation by visual cycle activity in rodent photoreceptors in vivo.

PURPOSE To test the hypothesis that the extent of outer retina uptake of manganese, measured noninvasively with manganese-enhanced MRI (MEMRI), is a quantitative biomarker of photoreceptor ion channel regulation by visual cycle activity. METHODS Four groups of animals were studied: control rats adapted to three different background light intensities, dark-adapted control mice systemically pretreated with retinylamine, and dark-adapted mice with a nonsense mutation in exon 3 of the RPE65 gene (RPE65(rd12)) with and without systemic 11-cis-retinal pretreatment. In all cases, rodents were anesthetized and studied with MEMRI 4 hours after manganese administration IP. Central retinal thickness and intraretinal ion channel regulation were measured from the MEMRI data. RESULT No differences (P>0.05) in retinal thickness were noted within any arm of this study. In rats, manganese uptake was inversely proportional to the background light intensity in the outer retina but not in the inner retina. Specific inhibition at the level of RPE65 activity, either acutely with retinylamine or chronically in RPE65(rd12) mice, similarly reduced (P<0.05) outer retinal manganese uptake compared with that in control mice. In RPE65(rd12) mice, outer retinal manganese uptake returned to normal (P>0.05) after 11-cis retinal treatment. Inner retinal uptake was supernormal (P<0.05) in retinylamine-treated mice but normal in untreated or 11-cis treated RPE65(rd12) mice. CONCLUSIONS The present data support measuring the extent of manganese uptake in the outer retina as an analytic noninvasive metric of visual cycle regulation of photoreceptor ion channel activity in vivo.

Ionic Dysregulatory Phenotyping of Pathologic Retinal Thinning with Manganese-Enhanced MRI

PURPOSE To test the hypothesis that manganese-enhanced MRI (MEMRI) provides a sensitive and robust measure of an important retinal ionic dysregulatory phenotype in pathologic retinal thinning. METHODS Four hours after intraperitoneal MnCl(2) injection, high-resolution MEMRI data were collected from overnight dark-adapted male control Sprague-Dawley and albino Royal College of Surgeons rats before (at development stage postnatal day [P] 17) and during photoreceptor degeneration (P36 and P57). In separate experiments, control rats, with and without repetitive hypoxic preconditioning, were subjected to high IOP (100 mm Hg) for 60 minutes followed by 24 hours or 7 days of reperfusion (e.g., ischemia/reperfusion). Central retinal thickness and intraretinal ion activity were measured from the MEMRI data. Histology examination was also performed to confirm retinal damage. RESULTS In two different neurodegenerative models, MEMRI revealed first-time evidence for changes (P < 0.05) in intraretinal ion regulation before and during pathologic, but not (P > 0.05) developmental, retinal thinning. This phenotype was significantly altered by a neuroprotective repetitive hypoxic preconditioning protocol. CONCLUSIONS MEMRI and a nontoxic systemic dose of MnCl(2) provided an objective, noninvasive measure of an ionic deregulatory phenotype that appears useful for improved early diagnosis and treatment prognosis in a range of neurodegenerative diseases and their treatment.