A newOpn4crerecombinase mouse line to target intrinsically photosensitive retinal ganglion cells (ipRGCs)

Abstract

ABSTRACTIntrinsically photosensitive retinal ganglion cells (ipRGCs) play a crucial role in several physiological light responses. In this study we generate a newOpn4creknock-in allele (Opn4cre(DSO)), in which cre is placed immediately downstream of theOpn4start codon. This approach aims to faithfully reproduce endogenousOpn4expression and improve compatibility with widely used reporters. We evaluated the efficacy and sensitivity ofOpn4cre(DSO)for labeling in retina and brain, and provide an in-depth comparison with the extensively utilizedOpn4cre(Saha)line. Through this characterization,Opn4cre(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, withOpn4cre(DSO)only labeling ipRGC axonal projections to their targets. The combination of off-target effects ofOpn4cre(Saha)across the retina and brain, coupled with diminished efficiencies of bothCrelines when coupled to less sensitive reporters, underscores the need for careful consideration in experimental design and validation with anyOpn4credriver. Overall, theOpn4cre(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.