Fig. 2.

LiGluR expression downstream in RGCs and upstream in ON-BCs restores light responses in the rd1 mouse retina in vitro (light intensity for all recordings: 27.7 mW/cm² or 6.3 × 10¹⁶ photons per cm⁻²⋅s⁻¹). (A and B) Viral DNA expression cassette. LiGluR is flanked by inverted terminal repeat (ITR) domains and stabilized by a polyadenylation signal sequence (polyA) and a woodchuck hepatitis posttranscriptional regulatory element (WPRE). ONL, outer nuclear layer. LiGluR expression is driven by hsyn-1 (A) or 4xgrm6 (B). (C and E) Schematic of a degenerated rd1 mouse retina with targeted cells highlighted in green. Confocal images of LiGluR expression in RGCs of rd1 mouse retina >4 wk after intravitreal injection of AAV2/2-hSyn-LiGLuR (2-μL volume equal to 5 × 10¹¹ viral genomes) (D and G) or ON-BCs of rd1 mouse retina >6 wk after subretinal injection of AAV2/2-(4YF)-4xgrm6-LiGLuR (2-μL volume equal to 5 × 10¹¹ viral genomes) (F and H). Retinas were stained with an anti-GluK2 antibody (green), and nuclei were stained with DAPI (blue). (Scale bars: 10 μm.) The IPL is shown with indication of on- and off-sublayers. MEA recordings from rd1 mouse retinas expressing LiGluR in RGCs (I and K) or ON-BCs (J and L) in the absence (I and J) or presence (K and L) of MAG0₄₆₀. (Top) Light stimulation protocol: 5× 3 s of blue light and 8 s dark. (Middle) Raster plot with spikes for all light-sensitive RGCs (I and K, n = 130 cells; J and L, n = 46 cells). (Bottom) Peristimulus time histogram (PSTH) with 250-ms bins. Comparison of the average firing rates (500-ms bins) in the light vs. dark for rd1 retinas expressing LiGLuR in RGCs (M) and ON-BCs (N) in the absence (gray) (M: n = 6 retinas, 478 cells; N: n = 4 retinas, 416 cells) and presence (green) (M, n = 5 retinas, 303 cells; N, n = 4 retinas, 332 cells) of MAG0₄₆₀ and population mean (black and blue, respectively).