David Bissig

Who Benefits From Memory Training

Cognitive training programs can have significant benefits. However, their efficacy is often reduced for individuals of advanced age or lower cognitive ability. Using older adult subjects, we examined the role of self-initiation of cognitive control in a training program that targets recollection memory. Relative time spent on an open-ended, intentional encoding task that requires the self-initiation of cognitive control was highly predictive of improvement in the training task, and fully accounted for individual differences related to age and crystallized intelligence. Analyzing training programs from the perspective of cognitive theory may help clarify how these programs have their effects and suggest ways to optimize such programs for the individuals who need them most.

Manganese-enhanced MRI of layer-specific activity in the visual cortex from awake and free-moving rats.

Cortical responses to visual stimulation have been studied extensively in the rodent, but often require post-stimulation ex vivo examination of the tissue. Here, we test the hypothesis that visual stimulus-dependent cortical activity from awake and free-moving rats can be encoded following systemically administered MnCl(2), and activity subsequently readout using manganese-enhanced MRI (MEMRI), a technique that can be performed without sacrificing the animal. Unanesthetized Sprague-Dawley rats, with or without systemic injection of MnCl(2), were maintained for 8 h in either a visually stimulating environment or darkness. To identify vision-dependent changes in cortical activity, animals were anesthetized and cortices were examined by 3D RARE MEMRI. Mean signal intensities in sub-cortical regions (e.g., superior colliculus and the lateral geniculate), and cortical regions (primary and accessory visual cortices) were compared. Cortex linearization was performed to aid in layer-specific signal intensity comparisons. Manganese administration alone globally increased signal intensity in the brain (P<0.0001). In visually stimulated and unstimulated rats, layer-specific analysis revealed that stimulated rats had on average significantly (P<0.05) higher signal intensities in layers IV and V of the primary visual cortex, as well as in deeper portions of the superficial superior colliculus, relative to dark adapted rats. Such differences went undetected without layer-specific analysis. We demonstrate, for the first time, the feasibility of layer-specific stimulus-dependant non-invasive MEMRI readout after encoding activity in awake and free moving rats. Future MEMRI studies are envisioned that measure the effects on cortical activity of sensory stimulation, as well as normal development, disease, plasticity, and therapy in longitudinal studies.

Evidence of key tinnitus-related brain regions documented by a unique combination of manganese-enhanced MRI and acoustic startle reflex testing

Animal models continue to improve our understanding of tinnitus pathogenesis and aid in development of new treatments. However, there are no diagnostic biomarkers for tinnitus-related pathophysiology for use in awake, freely moving animals. To address this disparity, two complementary methods were combined to examine reliable tinnitus models (rats repeatedly administered salicylate or exposed to a single noise event): inhibition of acoustic startle and manganese-enhanced MRI. Salicylate-induced tinnitus resulted in wide spread supernormal manganese uptake compared to noise-induced tinnitus. Neither model demonstrated significant differences in the auditory cortex. Only in the dorsal cortex of the inferior colliculus (DCIC) did both models exhibit supernormal uptake. Therefore, abnormal membrane depolarization in the DCIC appears to be important in tinnitus-mediated activity. Our results provide the foundation for future studies correlating the severity and longevity of tinnitus with hearing loss and neuronal activity in specific brain regions and tools for evaluating treatment efficacy across paradigms.

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.

Same-session functional assessment of rat retina and brain with manganese-enhanced MRI.

Manganese-enhanced MRI (MEMRI) is a powerful non-invasive approach for objectively measuring either retina or binocular visual brain activity in vivo. In this study, we investigated the sensitivity of MEMRI to monocular stimulation using a new protocol for providing within-subject functional comparisons in the retina and brain in the same scanning session. Adult Sprague Dawley or Long-Evans rats had one eye covered with an opaque patch. After intraperitoneal Mn(2+) administration on the following day, rats underwent visual stimulation for 8h. Animals were then anesthetized, and the brain and each eye examined by MEMRI. Function was assessed through pairwise comparisons of the patched (dark-adapted) versus unpatched (light-exposed) eyes, and of differentially-stimulated brain structures - the dorsal lateral geniculate nucleus, superior colliculus, and visual cortical regions - contralateral to the patched versus unpatched eye. As expected, Mn(2+) uptake was greater in the outer retina of dark-adapted, relative to light-exposed, eyes (P<0.05). Contralateral to the unpatched eye, significantly more Mn(2+) uptake was found throughout the visual brain regions than in the corresponding structures contralateral to the patched eye (P<0.05). Notably, this regional pattern of activity corresponded well to previous work with monocular stimulation. No stimulation-dependent differences in Mn(2+) uptake were observed in negative control brain regions (P>0.05). Post-hoc assessment of functional data by animal age and strain revealed no significant effects. These results demonstrate, for the first time, the acquisition of functional MRI data from the eye and visual brain regions in a single scanning session.

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.

Loss of Caveolin-1 Impairs Retinal Function Due to Disturbance of Subretinal Microenvironment

Background: Caveolin-1 is widely expressed in the retina and is linked to ocular disease. Results: Loss of caveolin-1 results in defective retinal function and ion homeostasis that is not photoreceptor-intrinsic. Conclusion: Caveolin-1 expressed in non-neuronal cells (e.g. Müller glia, retinal pigment epithelium) supports neuronal function through regulating the subretinal microenvironment. Significance: This study provides key evidence that caveolin-1 maintains retinal homeostasis. Caveolin-1 (Cav-1), an integral component of caveolar membrane domains, is expressed in several retinal cell types, including photoreceptors, retinal vascular endothelial cells, Müller glia, and retinal pigment epithelium (RPE) cells. Recent evidence links Cav-1 to ocular diseases, including autoimmune uveitis, diabetic retinopathy, and primary open angle glaucoma, but its role in normal vision is largely undetermined. In this report, we show that ablation of Cav-1 results in reduced inner and outer retinal function as measured, in vivo, by electroretinography and manganese-enhanced MRI. Somewhat surprisingly, dark current and light sensitivity were normal in individual rods (recorded with suction electrode methods) from Cav-1 knock-out (KO) mice. Although photoreceptor function was largely normal, in vitro, the apparent K+ affinity of the RPE-expressed α1-Na+/K+-ATPase was decreased in Cav-1 KO mice. Cav-1 KO retinas also displayed unusually tight adhesion with the RPE, which could be resolved by brief treatment with hyperosmotic medium, suggesting alterations in outer retinal fluid homeostasis. Collectively, these findings demonstrate that reduced retinal function resulting from Cav-1 ablation is not photoreceptor-intrinsic but rather involves impaired subretinal and/or RPE ion/fluid homeostasis.

Catalase therapy corrects oxidative stress-induced pathophysiology in incipient diabetic retinopathy.

PURPOSE Preclinical studies have highlighted retinal oxidative stress in the pathogenesis of diabetic retinopathy. We evaluated whether a treatment designed to enhance cellular catalase reduces oxidative stress in retinal cells cultured in high glucose and in diabetic mice corrects an imaging biomarker responsive to antioxidant therapy (manganese-enhanced magnetic resonance imaging [MEMRI]). METHODS Human retinal Müller and pigment epithelial cells were chronically exposed to normal or high glucose levels and treated with a cell-penetrating derivative of the peroxisomal enzyme catalase (called CAT-SKL). Hydrogen peroxide (H2O2) levels were measured using a quantitative fluorescence-based assay. For in vivo studies, streptozotocin (STZ)-induced diabetic C57Bl/6 mice were treated subcutaneously once a week for 3 to 4 months with CAT-SKL; untreated age-matched nondiabetic controls and untreated diabetic mice also were studied. MEMRI was used to analytically assess the efficacy of CAT-SKL treatment on diabetes-evoked oxidative stress-related pathophysiology in vivo. Similar analyses were performed with difluoromethylornithine (DFMO), an irreversible inhibitor of ornithine decarboxylase. RESULTS After catalase transduction, high glucose-induced peroxide production was significantly lowered in both human retinal cell lines. In diabetic mice in vivo, subnormal intraretinal uptake of manganese was significantly improved by catalase supplementation. In addition, in the peroxisome-rich liver of treated mice catalase enzyme activity increased and oxidative damage (as measured by lipid peroxidation) declined. On the other hand, DFMO was largely without effect in these in vitro or in vivo assays. CONCLUSIONS This proof-of-concept study raises the possibility that augmentation of catalase is a therapy for treating the retinal oxidative stress associated with diabetic retinopathy.

Evidence for diffuse central retinal edema in vivo in diabetic male Sprague Dawley rats.

Background Investigations into the mechanism of diffuse retinal edema in diabetic subjects have been limited by a lack of animal models and techniques that co-localized retinal thickness and hydration in vivo. In this study we test the hypothesis that a previously reported supernormal central retinal thickness on MRI measured in experimental diabetic retinopathy in vivo represents a persistent and diffuse edema. Methodology/Principal Findings In diabetic and age-matched control rats, and in rats experiencing dilutional hyponatremia (as a positive edema control), whole central retinal thickness, intraretinal water content and apparent diffusion coefficients (ADC, ‘water mobility’) were measured in vivo using quantitative MRI methods. Glycated hemoglobin and retinal thickness ex vivo (histology) were also measured in control and diabetic groups. In the dilutional hyponatremia model, central retinal thickness and water content were supernormal by quantitative MRI, and intraretinal water mobility profiles changed in a manner consistent with intracellular edema. Groups of diabetic (2, 3, 4, 6, and 9 mo of diabetes), and age-matched controls were then investigated with MRI and all diabetic rats showed supernormal whole central retinal thickness. In a separate study in 4 mo diabetic rats (and controls), MRI retinal thickness and water content metrics were significantly greater than normal, and ADC was subnormal in the outer retina; the increase in retinal thickness was not detected histologically on sections of fixed and dehydrated retinas from these rats. Conclusions/Significance Diabetic male Sprague Dawley rats demonstrate a persistent and diffuse retinal edema in vivo, providing, for the first time, an important model for investigating its pathogenesis and treatment. These studies also validate MRI as a powerful approach for investigating mechanisms of diabetic retinal edema in future experimental and clinical investigations.

MRI biomarkers for evaluation of treatment efficacy in preclinical diabetic retinopathy

INTRODUCTION One sober consequence of the current epidemic of diabetes mellitus is that an increasing number of people world-wide will partially or completely lose their sight to diabetic retinopathy. Clinically, the sight-threatening complications of diabetes are diagnosed and treated based on visible retinal lesions (e.g., dot-blot hemorrhages or retinal neovascularization). However, such anatomical microvascular lesions are slow to respond with treatment. Thus, there remains an urgent need for imaging biomarkers that are abnormal before retinal lesions are visibly apparent and are responsive to treatment. AREAS COVERED Here, the development of new MRI methods, such as manganese-enhanced MRI, for evaluating early diabetes-evoked retinal pathophysiology, and its usefulness in guiding new treatments for diabetic retinopathy are reviewed. EXPERT OPINION In diabetic retinopathy, not all important diagnostic and prognostic needs are well served by optical methods. In the absence of gross anatomy changes, critical times when drug intervention is most likely to be successful at reducing vision loss are missed by most light-based methods and thus provide little help in guiding diagnosis and treatment. For example, before clinical symptoms, is there an optimal time to intervene with drug therapy? Is a drug reaching its target? How does one assess optimal drug dose, schedule, and routes? How well do current experimental models mimic the clinical condition? As discussed herein, MRI is as an analytical tool for addressing these unmet needs. Future clinical applications of MRI can be envisioned such as in clinical trials to assess drug treatment efficacy, or as an adjunct approach to refine or clarify a difficult clinical case. New MRI-generated hypotheses about the pathogenesis of diabetic retinopathy and its treatment are discussed. In the coming years, a substantial growth in the development and application of MRI is expected to address relevant question in both the basic sciences and in the clinic.