Correcting QUEST Magnetic Resonance Imaging-Sensitive Free Radical Production in the Outer Retina In Vivo Does Not Correct Reduced Visual Performance in 24-Month-Old C57BL/6J Mice.

Purpose: To test the hypothesis that acutely correcting a sustained presence of outer retina free radicals measured in vivo in 24-month-old mice corrects their reduced visual performance. Methods: Male C57BL/6J mice two and 24 months old were noninvasively evaluated for unremitted production of paramagnetic free radicals based on whether 1/T1 in retinal laminae are reduced after acute antioxidant administration (QUEnch-assiSTed [QUEST] magnetic resonance imaging [MRI]). Superoxide production was measured in freshly excised retina (lucigenin assay). Combining acute antioxidant administration with optical coherence tomography (i.e., QUEST OCT) tested for excessive free radical-induced shrinkage of the subretinal space volume. Combining antioxidant administration with optokinetic tracking tested for a contribution of uncontrolled free radical production to cone-based visual performance declines. Results: At two months, antioxidants had no effect on 1/T1 in vivo in any retinal layer. At 24 months, antioxidants reduced 1/T1 only in superior outer retina. No age-related change in retinal superoxide production was measured ex vivo, suggesting that free radical species other than superoxide contributed to the positive QUEST MRI signal at 24 months. Also, subretinal space volume did not show evidence for age-related shrinkage and was unresponsive to antioxidants. Finally, visual performance declined with age and was not restored by antioxidants that were effective per QUEST MRI. Conclusions: An ongoing uncontrolled production of outer retina free radicals as measured in vivo in 24 mo C57BL/6J mice appears to be insufficient to explain reductions in visual performance.

Sildenafil-evoked photoreceptor oxidative stress in vivo is unrelated to impaired visual performance in mice.

PURPOSE: The phosphodiesterase inhibitor sildenafil is a promising treatment for neurodegenerative disease, but it can cause oxidative stress in photoreceptors ex vivo and degrade visual performance in humans. Here, we test the hypotheses that in wildtype mice sildenafil causes i) wide-spread photoreceptor oxidative stress in vivo that is linked with ii) impaired vision. METHODS: In dark or light-adapted C57BL/6 mice ± sildenafil treatment, the presence of oxidative stress was evaluated in retina laminae in vivo by QUEnch-assiSTed (QUEST) magnetic resonance imaging, in the subretinal space in vivo by QUEST optical coherence tomography, and in freshly excised retina by a dichlorofluorescein assay. Visual performance indices were also evaluated by QUEST optokinetic tracking. RESULTS: In light-adapted mice, 1 hr post-sildenafil administration, oxidative stress was most evident in the superior peripheral outer retina on both in vivo and ex vivo examinations; little evidence was noted for central retina oxidative stress in vivo and ex vivo. In dark-adapted mice 1 hr after sildenafil, no evidence for outer retina oxidative stress was found in vivo. Evidence for sildenafil-induced central retina rod cGMP accumulation was suggested as a panretinally thinner, dark-like subretinal space thickness in light-adapted mice at 1 hr but not 5 hr post-sildenafil. Cone-based visual performance was impaired by 5 hr post-sildenafil and not corrected with anti-oxidants; vision was normal at 1 hr and 24 hr post-sildenafil. CONCLUSIONS: The sildenafil-induced spatiotemporal pattern of oxidative stress in photoreceptors dominated by rods was unrelated to impairment of cone-based visual performance in wildtype mice.

Rod Photoreceptor Neuroprotection in Dark-Reared Pde6brd10 Mice.

Purpose: The purpose of this study was to test the hypothesis that anti-oxidant and / or anti-inflammation drugs that suppress rod death in cyclic light-reared Pde6brd10 mice are also effective in dark-reared Pde6brd10 mice. Methods: In untreated dark-reared Pde6brd10 mice at post-natal (P) days 23 to 24, we measured the outer nuclear layer (ONL) thickness (histology) and dark-light thickness difference in external limiting membrane-retinal pigment epithelium (ELM-RPE) (optical coherence tomography [OCT]), retina layer oxidative stress (QUEnch-assiSTed [QUEST] magnetic resonance imaging [MRI]); and microglia/macrophage-driven inflammation (immunohistology). In dark-reared P50 Pde6brd10 mice, ONL thickness was measured (OCT) in groups given normal chow or chow admixed with methylene blue (MB) + Norgestrel (anti-oxidant, anti-inflammatory), or MB or Norgestrel separately. Results: P24 Pde6brd10 mice showed no significant dark-light ELM-RPE response in superior and inferior retina consistent with high cGMP levels. Norgestrel did not significantly suppress the oxidative stress of Pde6brd10 mice that is only found in superior central outer retina of males at P23. Overt rod degeneration with microglia/macrophage activation was observed but only in the far peripheral superior retina in male and female P23 Pde6brd10 mice. Significant rod protection was measured in female P50 Pde6brd10 mice given 5 mg/kg/day MB + Norgestrel diet; no significant benefit was seen with MB chow or Norgestrel chow alone, nor in similarly treated male mice. Conclusions: In early rod degeneration in dark-reared Pde6brd10 mice, little evidence is found in central retina for spatial associations among biomarkers of the PDE6B mutation, oxidative stress, and rod death; neuroprotection at P50 was limited to a combination of anti-oxidant/anti-inflammation treatment in a sex-specific manner.

Age-related murine hippocampal CA1 laminae oxidative stress measured in vivo by QUEnch-assiSTed (QUEST) MRI: impact of isoflurane anesthesia.

Age-related impairments in spatial learning and memory often precede non-familial neurodegenerative disease. Ex vivo studies suggest that physiologic age-related oxidative stress in hippocampus area CA1 may contribute to prodromal spatial disorientation and to morbidity. Yet, conventional blood or cerebrospinal fluid assays appear insufficient for early detection or management of oxidative stress within CA1 sub-regions in vivo. Here, we address this biomarker problem using a non-invasive MRI index of CA1 laminae oxidative stress based on reduction in R1 (= 1/T1) after anti-oxidant administration. An R1 reduction reflects quenching of continuous and excessive production of endogenous paramagnetic free radicals. Careful motion-correction image acquisition, and avoiding repeated exposure to isoflurane, facilitates detection of hippocampus CA1 laminae oxidative stress with QUEnch-assiSTed (QUEST) MRI. Intriguingly, age- and isoflurane-related oxidative stress is localized to the stratum lacunosum of the CA1 region. Our data raise the possibility of using QUEST MRI and FDA-approved anti-oxidants to remediate spatial disorientation and later neurodegeneration with age in animals and humans.