Mapping protein distribution in the canine photoreceptor sensory cilium and calyceal processes by ultrastructure expansion microscopy

Photoreceptors are highly polarized sensory neurons, possessing a unique ciliary structure known as the photoreceptor sensory cilium (PSC). Vertebrates have two subtypes of photoreceptors: rods, which are responsible for night vision, and cones, which support daylight vision and color perception. Despite identifying functional and morphological differences between these subtypes, ultrastructural analyses of the PSC molecular architecture in rods and cones are still lacking. In this study, we employed ultrastructure expansion microscopy (U-ExM) to characterize the molecular architecture of the PSC in canine retina. We demonstrated that U-ExM is applicable to both non-frozen and cryopreserved retinal tissues with standard paraformaldehyde fixation. Using this validated U-ExM protocol, we revealed the molecular localization of numerous ciliopathy-related proteins in canine photoreceptors. Furthermore, we identified significant architectural differences in the PSC, ciliary rootlet, and calyceal processes between canine rods and cones. These findings pave the way for a better understanding of alterations in the molecular architecture of the PSC in canine models of retinal ciliopathies.