Amyloid-beta (Aβ) is a group of aggregation-prone, 38- to 43-amino acid peptides generated in the eye and other organs. Numerous studies suggest that the excessive build-up of low-molecular-weight soluble oligomers of Aβ plays a role in the progression of Alzheimer’s disease and other brain degenerative diseases. Recent studies raise the hypothesis that excessive Aβ levels may contribute also to certain retinal degenerative diseases. These findings, together with evidence that a major portion of Aβ is released as monomer into the extracellular space, raise the possibility that a technology enabling the enzymatic break-down of monomeric Aβ in the living eye under physiological conditions could prove useful for research on ocular Aβ physiology and, perhaps ultimately, for therapeutic applications. Neprilysin (NEP), an endopeptidase known to cleave Aβ monomer into inactive products, is a membrane-associated protein. However, sNEP, a recombinant form of the NEP catalytic domain, is soluble in aqueous medium. With the aim of determining the Aβ-cleaving activity of exogenous sNEP in the microenvironment of the intact eye, we analyzed the effect of intra-vitreally delivered sNEP on ocular Aβ levels in mice that exhibit readily measurable, aqueous buffer-extractable Aβ40 and Aβ42, two principal forms of Aβ. Anesthetized 10-month wild-type (C57BL/6J) and 2–3-month 5XFAD transgenic mice received intra-vitreal injections of sNEP (0.004 – 10 μg) in one eye and were sacrificed at defined post-treatment times (30 min – 12 weeks). Eye tissues (combined lens, vitreous, retina, RPE and choroid) were homogenized in phosphate-buffered saline, and analyzed for Aβ40 and Aβ42 (ELISA) and for total protein (Bradford assay). The fellow, untreated eye of each mouse served as control, and concentrations of Aβ (pmol/g protein) in the treated eye were normalized to that of the untreated control eye. In C57BL/6J mice, as measured at 2 hr after sNEP treatment, increasing amounts of injected sNEP yielded progressively greater reductions of Aβ40, ranging from 12% ± 3% (mean ± SEM; n=3) with 4 ng sNEP to 85% ± 13% (n=5) with 10 μg sNEP. At 4 ng sNEP the average Aβ40 reduction reached >70% by 24 hr following treatment and remained near this level for about 8 weeks. In 5XFAD mice, 10 μg sNEP produced an Aβ40 decrease of 99% ± 1% (n=4) and a substantial although smaller decrease in Aβ42 (42% ± 36%; n=4) within 24 hr. Electroretinograms (ERGs) were recorded from eyes of C57BL/6J and 5XFAD mice at 9 days following treatment with 4 ng or 10 μg sNEP, conditions that on average led, respectively, to an 82% and 91% Aβ40 reduction in C57BL/6J eyes, an 87% and 92% Aβ40 reduction in 5XFAD eyes, and a 23% and 52% Aβ42 reduction in 5XFAD eyes. In all cases, sNEP-treated eyes exhibited robust ERG responses, consistent with a general tolerance of the posterior eye tissues to the investigated conditions of sNEP treatment. The sNEP-mediated decrease of ocular Aβ levels reported here represents a possible approach for determining effects of Aβ reduction in normal...

Corneal potential maps obtained in response to full-field flash stimuli are altered in eyes with scotomas in the central and far-peripheral retina. The meERG approach yields useful spatial information following a single brief flash, analogous to body-surface potential maps used to evaluate heart and brain.

PurposeIt has been known for several decades that the magnitude of the corneal electroretinogram (ERG) varies with position on the eye surface, especially in the presence of focal or asymmetric stimuli or retinal lesions. However, this phenomenon has not been well-characterized using simultaneous measurements at multiple locations on the cornea. This work provides the first characterization of spatial differences in the ERG across the rat cornea.MethodsA contact lens electrode array was employed to record ERG potentials at 25 corneal locations simultaneously following brief full-field flash stimuli in normally sighted Long-Evans rats. These multi-electrode electroretinogram (meERG) responses were analyzed for spatial differences in a-wave and b-wave amplitudes and implicit times.ResultsSpatially distinct ERG potentials could be recorded reliably. Comparing relative amplitudes across the corneal locations suggested a slight non-uniform distribution when using full-field, near-saturating stimuli. Amplitudes of a- and b-waves were approximately 3 % lower in the inferior quadrant than in the superior quadrant of the cornea.ConclusionsThe present results comprise the start of the first normative meERG database for rat eyes and provide a basis for comparison of results from eyes with functional deficit. Robust measures of spatial differences in corneal potentials will also support optimization and validation of computational source models of the ERG. To fully utilize the information contained in the meERG data, a detailed understanding of the roles of the many determinants of local corneal potentials will eventually be required.

This model can now be used for optimization of ERG electrode design, and to support functional mapping of the retina from meERG data via solving the inverse bioelectric source problem.

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