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.

QUEST MRI assessment of fetal brain oxidative stress in utero.

PURPOSE: To achieve sufficient precision of R1 (=1/T1) maps of the fetal brain in utero to perform QUEnch-assiSTed (QUEST) MRI in which a significant anti-oxidant-induced reduction in R1 indicates oxidative stress. METHODS: C57BL/6 mouse fetuses in utero were gently and non-surgically isolated and secured using a homemade 3D printed clip. Using a commercial receive-only surface coil, brain maps of R1, an index sensitive to excessive and continuous free radical production, were collected using either a conventional Cartesian or a non-Cartesian (periodically rotated overlapping parallel lines with enhanced reconstruction) progressive saturation sequence. Data were normalized to the shortest TR time to remove bias. To assess oxidative stress, brain R1 maps were acquired on the lipopolysaccharide (LPS) model of preterm birth ±â€¯rosiglitazone (ROSI, which has anti-oxidant properties); phosphate buffered saline (PBS) controls ±â€¯ROSI were similarly studied. RESULTS: Sufficient quality R1 maps were generated by a combination of the 3D printed clip, surface coil detection, non-Cartesian sequence, and normalization scheme ensuring minimal fetal movement, good detection sensitivity, reduced motion artifacts, and minimal baseline variations, respectively. In the LPS group, the combined caudate-putamen and thalamus region R1 was reduced (p < 0.05) with ROSI treatment consistent with brain oxidative stress; no evidence for oxidative stress was found in the pons region. In the PBS control group, brain R1's did not change with ROSI treatment. CONCLUSION: The sensitivity and reproducibility of the combined approaches described herein enabled first-time demonstration of regional oxidative stress measurements of the fetal brain in utero using QUEST MRI.

Outer Retinal Oxidative Stress Measured In Vivo Using QUEnch-assiSTed (QUEST) OCT.

Purpose: To test the hypothesis that oxidative stress in the outer retina (OR = distance from external limiting membrane to the retinal pigment epithelium-choroid boundary) can be detected by using antioxidants (AOs) to correct an impaired light-evoked response as measured by optical coherence tomography (OCT). Methods: C57BL/6J mice were maintained in the dark for ∼20 hours and studied by OCT before and after 1 hour of light exposure. OR thickness in dark or light was measured, and the light-dark difference (i.e., the photoresponse) was calculated. Subgroups of mice were given either saline or d-cis-diltiazem (an inducer of transient and nondamaging OR oxidative stress) ± methylene blue (24 hours before examination) and α-lipoic acid (1 hour before examination); one group was kept only in the dark and given only AOs. Results: In uninjected or saline-injected control mice, the OR showed a similar and reproducible light-induced expansion; dark-adapted mice given AOs did not increase dark-adapted OR thickness. The d-cis-diltiazem-treated mice had no photoresponse (P > 0.05). The d-cis-diltiazem-treated mice given AOs corrected (P < 0.05) the suppressed OR photoresponse, indicating the presence of oxidative stress. Conclusions: QUEnch-assiSTed (QUEST) OCT reproduced results from previous gold standard assays, showing that oxidative stress impairs the OR photoresponse and that d-cis-diltiazem produces OR oxidative stress. We envision future applications of QUEST OCT in a range of oxidative stress-based retinopathies.