Functional methods for evaluating the efficacy of retinal optogenetic therapy for vision restoration

Optogenetic therapy is a promising strategy for vision restoration. Functional assays have an important role in assessing the modulation of neural activity in response to light stimulation. Each functional assay needs to be carefully considered and evaluated for the preclinical development of optogenetic approaches to work toward meaningful vision restoration. Each strategy contributes to understanding the efficacy of vision restoration and the physiology of retinal optogenetic therapy. At a molecular level, bioluminescence resonance energy-transfer based and G protein coupling assays can be used. Calcium imaging provides measurements with useful spatial and temporal resolution using fluorescent calcium indicators at the cellular level. Electrophysiological tests can include ex vivo recordings by patch-clamping at single-cell resolution, multielectrode array recordings at the network level, and in vivo recordings at the lateral geniculate nucleus and cortical levels. Behavioural tests such as light avoidance, optomotor response and visual discrimination assess functional restoration in vivo. In this review, each functional assay is discussed in the context of retinal optogenetic therapy with notable examples that have demonstrated vision restoration. The advantages, disadvantages, and limitations of each assay are critically compared to highlight their relative scientific value and applicability across different stages of development. This provides insight into how these methods can be integrated within a translational framework, from molecular validation to behavioural outcomes, to better inform the design of preclinical studies. As clinical trials in optogenetic therapy continue to expand, improved alignment between preclinical functional assays and clinically meaningful endpoints will be essential to maximise translational success.