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. 2014 Mar 25;55(3):1859-66.
doi: 10.1167/iovs.13-13661.

Photoreceptor Cells With Profound Structural Deficits Can Support Useful Vision in Mice

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Free PMC article

Photoreceptor Cells With Profound Structural Deficits Can Support Useful Vision in Mice

Stewart Thompson et al. Invest Ophthalmol Vis Sci. .
Free PMC article

Abstract

Purpose: In animal models of degenerative photoreceptor disease, there has been some success in restoring photoreception by transplanting stem cell-derived photoreceptor cells into the subretinal space. However, only a small proportion of transplanted cells develop extended outer segments, considered critical for photoreceptor cell function. The purpose of this study was to determine whether photoreceptor cells that lack a fully formed outer segment could usefully contribute to vision.

Methods: Retinal and visual function was tested in wild-type and Rds mice at 90 days of age (Rds(P90)). Photoreceptor cells of mice homozygous for the Rds mutation in peripherin 2 never develop a fully formed outer segment. The electroretinogram and multielectrode recording of retinal ganglion cells were used to test retinal responses to light. Three distinct visual behaviors were used to assess visual capabilities: the optokinetic tracking response, the discrimination-based visual water task, and a measure of the effect of vision on wheel running.

Results: Rds(P90) mice had reduced but measurable electroretinogram responses to light, and exhibited light-evoked responses in multiple types of retinal ganglion cells, the output neurons of the retina. In optokinetic and discrimination-based tests, acuity was measurable but reduced, most notably when contrast was decreased. The wheel running test showed that Rds(P90) mice needed 3 log units brighter luminance than wild type to support useful vision (10 cd/m(2)).

Conclusions: Photoreceptors that lack fully formed outer segments can support useful vision. This challenges the idea that normal cellular structure needs to be completely reproduced for transplanted cells to contribute to useful vision.

Keywords: mouse; photoreceptor; retinal degeneration; stem cell; visual behavior.

Figures

Figure 1
Figure 1
Photoreceptors in Rds mice. (A) Hematoxylin- and eosin-stained sections show that the RdsP90 outer nuclear layer (ONL) is thinner but most photoreceptors are intact. (B) Anti-rhodopsin labeling (red) shows the efficient compartmentalization of opsin to the photoreceptor outer segments (OS) in wild type, and the lack of a defined outer segment in RdsP90. Nuclei are labeled with DAPI (blue). (C) Electron microscope images show the stacked discs in outer segments (OS) of wild-type photoreceptor cells. In RdsP90 the inner segments (IS) are adjacent to the RPE. No stacked disc-containing structures are apparent. Inset: higher-magnification view of an example of a basal body (BB) with a truncated cilium. (D) Stem cell–derived photoreceptor cells (dsRed positive) transplanted into a retina with advanced degeneration express outer segment structural proteins (Rom1), but most do not develop an outer segment.
Figure 2
Figure 2
Electroretinogram measurement of gross retinal function. (A) Representative electroretinogram waveforms are shown for wild-type, RdsP90, and RdsP365 mice. (B) Mean and standard deviation of the b-wave amplitude to a dark-adapted bright flash or “combined maximal response” stimulus is shown for groups of animals.
Figure 3
Figure 3
Light-evoked responses of retinal ganglion cells in wild-type and RdsP90 mice. (AC) Responses are shown in columns for ON, sustained ON, OFF, and ON–OFF types of retinal ganglion cells. Example 10-trial raster plots and corresponding peristimulus time histograms are shown for these retinal ganglion cell types in (A) wild-type and (B) RdsP90 mice. Timing of the 1-second full-field flash of light is indicated by white background. Note that for a given response type, the scale is the same for both wild type and RdsP90, but scales are different for each cell type. (C) The mean percentage of cells of a given type is shown below the histograms (mean and SD per array). (D) The percentage of cells identified by baseline action potential activity that had an identifiable response to light is shown for wild-type and RdsP90 retinas (mean and SD per array). (E) The amplitude of change in activity is shown for wild type and RdsP90 (mean and SD of all cells with an identified response to light). (F) The distribution of response amplitudes is shown for wild type and RdsP90. Cells are grouped in bins of change in spike rate. Note that the distribution of amplitudes is not Gaussian, so bin sizes are not equal.
Figure 4
Figure 4
Optokinetic tracking responses. The threshold for tracking responses (mean and SD) in an optokinetic test is shown for (A) acuity tested at fixed contrast levels and (B) contrast sensitivity tested at fixed grating sizes (cyc/deg). Grating size is the width of the vertical bars presented, in cyc/deg. Contrast is the difference in shading between the light and dark bars of the stimulus. Where no animals responded to a stimulus, data point is shown on a gray square.
Figure 5
Figure 5
Visual water task. Performance in the stimulus discrimination–based visual water task test is shown for wild-type and RdsP90 mice. The percent of trials in which animals positively identified the stimulus versus an equal-luminance control gray screen is shown for (A) stimuli with different spatial frequencies and (C) stimuli with different levels of percent contrast. Data points are the mean of wild-type and RdsP90 mice. Performance below 70% correct indicates the animals could not distinguish the positive stimulus from the equal-luminance gray screen. The threshold for individual animals is then shown for (B) spatial frequency or grating (acuity) and (D) contrast sensitivity.
Figure 6
Figure 6
Vision augmented wheel running. The effect of light on wheel running activity is shown for wild-type, RdsP90, and RdsP365 mice at different luminance levels. Responses are defined as a percentage of baseline activity in complete darkness. Significance is indicated for the dimmest light level generating a increase in activity (****P < 0.0001; ***P < 0.001).

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