Restoration of mGluR6 Localization Following AAV-Mediated Delivery in a Mouse Model of Congenital Stationary Night Blindness

Abstract

Complete congenital stationary night blindness (cCSNB) is an incurable inherited retinal disorder characterized by an ON-bipolar cell (ON-BC) defect. GRM6 mutations are the third most prevalent cause of cCSNB. The Grm6 −/− mouse model mimics the human phenotype, showing no b-wave in the electroretinogram (ERG) and a loss of mGluR6 and other proteins of the same cascade at the outer plexiform layer (OPL). Our aim was to restore protein localization and function in Grm6 −/− adult mice targeting specifically ON-BCs or the whole retina. METHODS. Adeno-associated virus-encoding Grm6 under two different promoters (GRM6-Grm6 and CAG-Grm6) were injected intravitreally in P15 Grm6 −/− mice. ERG recordings at 2 and 4 months were performed in Grm6 +/+ , untreated and treated Grm6 −/− mice. Similarly, immunolocalization studies were performed on retinal slices before or after treatment using antibodies against mGluR6, TRPM1, GPR179, RGS7, RGS11, Gβ5, and dystrophin. RESULTS. Following treatment, mGluR6 was localized to the dendritic tips of ON-BCs when expressed with either promoter. The relocalization efficiency in mGluR6-transduced retinas at the OPL was 2.5% versus 11% when the GRM6-Grm6 and CAG-Grm6 were used, respectively. Albeit no functional rescue was seen in ERGs, relocalization of TRPM1, GPR179, and Gβ5 was also noted using both constructs. The restoration of the localization of RGS7, RGS11, and dystrophin was more obvious in retinas treated with GRM6-Grm6 than in retinas treated with CAG-Grm6. CONCLUSIONS. Our findings show the potential of treating cCSNB with GRM6 mutations; however, it appears that the transduction rate must be improved to restore visual function.

Functional Availability of ON-Bipolar Cells in the Degenerated Retina: Timing and Longevity of an Optogenetic Gene Therapy

“…Moreover, mGluR6 cascade activation may promote the preservation of OBC integrity. It is known from the literature that retinal remodeling is caused by deafferentation due to lack of input [ 42 ]; however, reintroducing such an input in OBCs may lead to the modulation and expression of mGluR6 components [ 15 , 67 ].…”

“…Indeed, in the last decade the effort of many research groups propelled the field of vision restoration into new, previously unimaginable dimensions [ 9 , 10 , 11 ]. In the early stages of retinal degeneration, with a still relatively intact retinal architecture, gene replacement [ 12 , 13 , 14 , 15 ] and pharmacology [ 16 , 17 , 18 ] are employed. In the case of complete photoreceptor loss, the treatment strategies include electronic prostheses [ 19 , 20 ], stem cells and cell transplantation [ 21 ], photo-switchable ligands [ 22 , 23 ], and optogenetics [ 24 , 25 , 26 , 27 ].…”

A New Mouse Model for Complete Congenital Stationary Night Blindness Due to Gpr179 Deficiency

“…Together, we delivered herein a novel mouse model mimicking the phenotype of patients with cCSNB due to mutations in GPR179, which delivers the basis to better understand the pathophysiology of the disease and develop therapeutic approaches to treat cCSNB. Indeed, partial restoration of the proteins underlying cCSNB and the phenotype was obtained using AAV-mediated gene therapy approaches for mice lacking Nyx , Lrit3, and Grm6 [ 40 , 55 , 56 ] (Varin, 2021, under revision). However, the bottleneck of an AAV-gene therapy approach for mice lacking Gpr179 will be the larger size of the coding sequence of Gpr179 (>6 kb), making it too large to pack it into an AAV.…”

“…Independently, another group identified mutations in GPR179 in patients with cCSNB and provided another mouse model with this gene defect, Gpr179 nob5 [ 32 ]. Therapeutic approaches using mice lacking functional NYX, GRM6, and LRIT3 partially restored the cCSNB phenotype [ 40 , 55 , 56 ] (Varin et al, 2021, under revision). Here, we describe the generation and characterization of a novel Gpr179 −/− mouse model which allows the study of the gene defect in an animal model to investigate the role of GPR179 in the signaling cascade in respect to other molecules implicated herein and which delivers the basis to study therapeutic approaches.…”

Substantial restoration of night vision in adult mice with congenital stationary night blindness

“… 23 After gene therapy in adult mice lacking Grm6 , mGluR6 and other proteins of the same cascade were localized at the dendritic tips of ON-BCs in the absence of functional restoration. 24 In addition, in all models, localization of TRPM1 at the dendritic tips of ON-BCs was observed, 22 , 23 , 24 while no restoration of the b-wave under photopic conditions could be obtained. 16 , 22 , 23 Here, we aimed to achieve a robust functional rescue in mature Lrit3 −/− retinas using different AAV-promoter combinations targeting ON-BCs, both rod and cone PRs, or the OPL.…”