Localization of primary cilia in mouse retina

Abstract

Introduction: The primary cilia are considered as a cellular antennae that sensing and interchanging the information with its extracellular environment and nearly all mammalian cells have a single primary cilium. In the retina, the outer segment (OS) and connecting cilium of photoreceptor is known to be a specialized form of primary cilia, but studies on cilia in other layer of retina is scarce. In this study, we aimed to find out the expression of primary cilia using different ciliary markers in a whole layer of mouse retina.

Methods: C57BL/6 mice were sacrificed on postnatal 26th day and eyes were enucleated from it. The eyes were embedded in paraffin, sectioned to 4㎛ thickness and stained with three different ciliary markers

Arl13b, Acetylated α tubulin and Adenylyl cyclase III. Using immunofluorescence, we evaluated the expression and relative location of these markers in the retina.

Results: Arl13b was expressed in photoreceptor layer (PRL), outer plexiform layer (OPL) and weakly in inner plexiform layer (IPL). Adenylyl cyclase III showed similar patterns with Arl13b and it also showed diffuse reactivity in ganglion cell layer (GCL). Acetylated α tubulin was expressed in PRL, OPL, IPL and also some longitudinal staining patterns were observed throughout the whole thickness which is thought to be the Muller cells. Arl13b and Adenylyl cyclase III showed diffuse staining in inner segment of PRL while Acetylated α tubulin showed speckled patterns in connecting cilium area. In OPL, all three markers showed strong reactivity with feathery patterns along the neuronal synapses. However, we couldnt find any single strand-like shapes that emerge directly from the cell surfaces.

Conclusions: In immunofluorescence with three different ciliary markers in mouse retina, we found positive reaction in PRL, OPL and GCL. We couldnt directly prove the strand-like shape of cilia in those areas, which might more specifically suggest the presence of cilia, but our results suggest the possibility of presence of primary cilia in OPL and GCL in addition to PRL OS. Furthermore, in the outer plexiform layer, all three markers showed intense staining along the neuronal synapses, which suggests that the neuronal processes themselves might share the features of cilia.