A visually guided swim assay for mouse models of human retinal disease recapitulates the multi-luminance mobility test in humans

Abstract

Purpose: The purpose of this study was to develop a visually guided swim assay (VGSA) for measuring vision in mouse retinal disease models comparable to the multi-luminance mobility test (MLMT) utilized in human clinical trials.

Methods: Three mouse retinal disease models were studied: Bardet-Biedl syndrome type 1 (Bbs1^M390R/M390R), n = 5; Bardet-Biedl syndrome type 10 (Bbs10^-/-), n = 11; and X linked retinoschisis (retinoschisin knockout; Rs1-KO), n = 5. Controls were normally-sighted mice, n = 10. Eyeless Pax6^Sey-Dey mice, n = 4, were used to determine the performance of animals without vision in VGSA.

Results: Eyeless Pax6^Sey-Dey mice had a VGSA time-to-platform (TTP) 7X longer than normally-sighted controls (P < 0.0001). Controls demonstrated no difference in their TTP in both lighting conditions; the same was true for Pax6^Sey-Dey. At 4-6 M, Rs1-KO and Bbs10^-/- had longer TTP in the dark than controls (P = 0.0156 and P = 1.23 × 10^-8, respectively). At 9-11 M, both BBS models had longer TTP than controls in light and dark with times similar to Pax6^Sey-Dey (P < 0.0001), demonstrating progressive vision loss in BBS models, but not in controls nor in Rs1-KO. At 1 M, Bbs10^-/- ERG light-adapted (cone) amplitudes were nonrecordable, resulting in a floor effect. VGSA did not reach a floor until 9-11 M. ERG combined rod/cone b-wave amplitudes were nonrecordable in all three mutant groups at 9-11 M, but VGSA still showed differences in visual function. ERG values correlate non-linearly with VGSA, and VGSA measured the continual decline of vision.

Conclusion: ERG is no longer a useful endpoint once the nonrecordable level is reached. VGSA differentiates between different levels of vision, different ages, and different disease models even after ERG is nonrecordable, similar to the MLMT in humans.

Keywords: Dark-adapted; electroretinogram; functional vision; light-adapted; retinal degeneration; visually guided swim assay.

Figure 1

Visually guided swim assay setup. (a) Schematic of pool setup. Circles determine the possible platform positions. Image to scale. 1 square = 1 inch. The red X denotes the mouse entry location. (b) Photo of experimental assay setup. Measurements follow the schematic to the left. (c) A night vision monocular used by investigators to view the mice during dark-adapted conditions. The individual in the photograph granted permission for its use

Figure 2

Comparison of Pax6^Sey-Dey and control mice. (a) Wild-type mouse with normal eye development. (b) Photo of Pax6^Sey-Dey mouse showing absent eye. (c) Close-up photo of anophthalmic Pax6^Sey-Dey mouse; right and left ocular regions in mice selected for the study were the same. (d) Comparison of time-to-platform (TTP) at 9–11 M for control mice and Pax6^Sey-Dey in both light and dark. Difference in TTP between genotypes at the same light level is significant. There was no difference in TTP between light and dark in mice of the same genotype. Control: Wild-type or heterozygous mice. s = seconds. **** = P < 0.0001. ns = not significant

Figure 3

Three different mouse models of retinal dystrophy compared to controls in the visually guided swim assay. (a and b) Comparison of the control group in light-adapted (LA) and dark-adapted (DA) conditions. No significant difference is shown between the control group at both lighting conditions. (c and d) Comparison of Bardet–Biedl syndrome (Bbs1^M390R/M390R), Bbs10^−/−, and Rs1-KO mouse models in LA and DA conditions at 4–6 M time point. (e and f) Comparison of Bbs1^M390R/M390R, Bbs10^−/−, Rs1-KO, and eyeless Pax6^Sey-Dey mouse models in LA and DA conditions at 9–11 M time point. BBS1^M390R/M390R data were reported previously.[10] Control = Wild type or heterozygous; s = seconds. * = P < 0.01, ** = P < 0.001, **** = P < 0.0001. ns = not significant. BBS: Bardet–Biedl syndrome

Figure 4

Electroretinogram (ERG) amplitude comparison of all mouse models versus control. ERG amplitudes of Bardet–Biedl syndrome (Bbs1^M390R/M390R), Bbs10^−/−, and Rs1-KO mouse models compared to controls at both ages (4–6 M and 9–11 M) in the dark-adapted standard combined response ERG test. Significant difference is shown between all mutant groups compared to controls at both time points. Control = Wild type or heterozygous; s = seconds. **** = P < 0.0001. Amp = Amplitude. μV = Microvolt. BBS: Bardet–Biedl syndrome, SCR: Standard combined response

Figure 5

The relationship between electroretinogram (ERG) and functional vision. (a) The standard combined response b-wave amplitudes of control, Bardet–Biedl syndrome (Bbs1^M390R/M390R), Bbs10^−/−, and Rs1-KO mice at the age of 4–6 M and 9–11 M are depicted against their respective time-to-platform (TTP) in dark-adapted conditions. (b) The difference in ERG b-wave amplitudes in plot (a), above, per 5-second window[shown in gray blocks in (b)] in TTP is illustrated. In the “low-vision” zone, ERG becomes insensitive, as evidenced by the absence of notable changes in ERG amplitudes despite the lengthening of TTP. (c) Comparison of Bbs10^−/− and controls 5 Hz ERG amplitudes in the light-adapted (LA) condition showing constant amplitudes over time for both groups. (d) Comparison of Bbs10^−/− and controls TTP in the LA condition over time showing longer TTP for Bbs10^−/− as they age and constant TTP in controls. s = seconds. M = months. Amp = Amplitude. μV = Microvolt. BBS: Bardet–Biedl syndrome, SCR: Standard combined response