Purpose: This study was designed to investigate whether the volume of vector used for subretinal injection can be reduced to transfect C57bl/6J mouse whole retina and whether it can restore cone function in a Cngb3 knockout (KO) mouse model.
Methods: C57bl/6J mice and Cngb3 KO mice received a subretinal injection of 0.5 μL or 1 μL of AAV5-smCBAmCherry vector and AAV5-IRBP/GNAT2-hCngb3 vector, respectively. Retinal whole mounts and frozen sections were prepared from the wild-type mouse eyes to evaluate the transfected area. Dark and light-adapted electroretinograms (ERGs) were recorded two months after vector injection in the eyes of Cngb3 KO mice.
Result: In the retina of AAV5-smCBA-mCherry injected wild-type mice, no difference was observed between the injection volumes. mCherry positive retinal pigment epithelial (RPE) and photoreceptor cells were observed throughout the entire retina. In AAV5-IRBP/GNAT2-hCngb3-injected Cngb3 KO mice, 1-μL-injected mice showed a higher average of photopic ERG restoration than 0.5-μL-injected mice. However, the scotopic ERGs were lower in 1-μL-injected mice, indicating that higher injection volumes resulted in more damages.
Conclusion: Reduced volume (0.5 μL) of vector induced fewer damages. However, higher doses of vector (1 μL) restore higher ERG function in Cngb3 KO mouse.