Manganese-enhanced MRI for preclinical evaluation of retinal degeneration treatments

Retinal degenerative diseases are a leading cause of blindness.1 In healthy individuals, the retinoid cycle mediates the light response in photoreceptor cells, whereby photoisomerization of the cis-retinal chromophore initiates a cycle of enzymatic activities to transduce the signal and regenerate the original chromophore.2 Retinal degeneration, at least in part, is attributed to accumulation of toxic byproducts of the retinoid cycle such as all-trans-retinal (atRAL) and A2E-like derivatives that induce apoptosis of photoreceptor cells.3 Primary amines, including retinylamine, have been shown to modulate the visual cycle and prevent excessive accumulation of atRAL.4–6 Studies in Abca4−/−Rdh8−/− double knockout mice, a light-induced retinal degeneration model, have demonstrated the efficacy of retinylamine to prophylactically preserve photoreceptor and RPE cells and prevent vision loss.7,8 The retinoid cycle relies on Ca2+ channel activity to transduce light-responsive enzymatic activity into electrical signaling.9,10 Regulation of intracellular Ca2+ concentration is critical for polarization of the cell, activation of voltage-gated ion channels, and adaptation of photoreceptor cells to retain responsiveness in different conditions of ambient lighting. In retinal degeneration, photoreceptor cell death results in a loss of capacity for Ca2+ signaling. Thus, when these cells are protected from degeneration by prophylactic treatment with retinylamine, we decided to assess Ca2+-signaling activity in response to this intervention. Manganese-enhanced magnetic resonance imaging (MEMRI) enables direct spatial visualization of Ca2+ channel activity in vivo. In MEMRI, paramagnetic Mn2+ ions are used as T1 contrast agents to image the activity of Ca2+ channels.11,12 In excitable cells, such as neurons and cardiac cells, Mn2+ has been shown to be taken up by active calcium signaling pathways, including voltage-gated channels. Mn2+ then is retained in these cells with a long half-life due to its slow cellular efflux.13,14 Manganese-enhanced magnetic resonance imaging is uniquely suited to image neural activity with the high spatial resolution and soft tissue contrast characteristic of MRI. Manganese-enhanced MRI has been used to study activity in various sensory pathways of the brain, including the auditory tract15 and the olfactory response.16 This technique holds particular appeal to assess retinal health because Mn2+ administration in the eyes is nontoxic at low doses that still fall within detection limits for contrast enhancement.17–19 In the visual system, MEMRI has been employed to study phototransduction under differing conditions20,21 and evaluated in disease models such as glaucoma,22,23 optic nerve crush,24 and diabetic retinopathy.25 With its ability to image anatomical structures and signaling activity simultaneously, we expect that MEMRI will be an effective method to assess the treatment response by measuring Ca2+ signaling activities in the retina. In this work, we investigated the use of MEMRI in Abca4−/−Rdh8−/− double knockout mice to evaluate intense light-induced retinal degeneration and its response to prophylactic treatment with retinylamine, a potent retinoid cycle modulator, based on changes in Ca2+ channel activity in the retina. Thus, we evaluated the ability of MEMRI to assess the structure and function of healthy, drug-treated, and fully degenerated retinas in these mutant mice according to the experimental time line shown in Figure 1. Figure 1 Use of MEMRI to assess retinal integrity and function. Mn2+ was injected directly into the vitreous humor of the right eye. (A) Actively signaling photoreceptor cells take up and retain Mn2+ through voltage-gated calcium channels. This phenomenon is detected ...