A new Opn4cre recombinase mouse line to target intrinsically photosensitive retinal ganglion cells (ipRGCs).

Intrinsically photosensitive retinal ganglion cells (ipRGCs) play a crucial role in several physiological light responses. In this study we generate a new Opn4 cre knock-in allele ( Opn4 cre(DSO) ), in which cre is placed immediately downstream of the Opn4 start codon. This approach aims to faithfully reproduce endogenous Opn4 expression and improve compatibility with widely used reporters. We evaluated the efficacy and sensitivity of Opn4 cre(DSO ) for labeling in retina and brain, and provide an in-depth comparison with the extensively utilized Opn4 cre(Saha ) line. Through this characterization, Opn4 cre(DSO) demonstrated higher specificity in labeling ipRGCs, with minimal recombination escape. Leveraging a combination of electrophysiological, molecular, and morphological analyses, we confirmed its sensitivity in detecting all ipRGC types (M1-M6). Using this new tool, we describe the topographical distributions of ipRGC types across the retinal landscape, uncovering distinct ventronasal biases for M5 and M6 types, setting them apart from their M1-M4 counterparts. In the brain, we find vastly different labeling patterns between lines, with Opn4 cre(DSO) only labeling ipRGC axonal projections to their targets. The combination of off-target effects of Opn4 cre(Saha) across the retina and brain, coupled with diminished efficiencies of both Cre lines when coupled to less sensitive reporters, underscores the need for careful consideration in experimental design and validation with any Opn4 cre driver. Overall, the Opn4 cre(DSO ) mouse line represents an improved tool for studying ipRGC function and distribution, offering a means to selectively target these cells to study light-regulated behaviors and physiology.