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Guillermo Lehmann-Mántaras, Ryan Schreiner, Erwin de la Fuente, Enrique Rodriguez-Boulan; Expression of RPE primary cilia in adult mice. Invest. Ophthalmol. Vis. Sci. 2013;54(15):6084.
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Primary cilia (PC) are antenna-like organelles that emanate from the apical membrane of epithelial cells. PC are a major signaling center involved in the transduction of environmental stimuli and in organization of signaling events during development. It is currently believed that RPE cells express PC in newborn mice and that PC are lost after P15. We hypothesized that loss of RPE might be an artifact, i.e., a consequence of the peeling off of the neural retina normally utilized to expose the RPE. To test this hypothesis, we utilized several approaches that minimize the damage caused by peeling of neural retina or eliminate the requirement for it.
To analyze the presence of RPE PC in adult mice, we utilized RPE flatmounts prepared under experimental conditions of low retinal adherence (dark adaptation, low divalent cation medium, etc). In addition, we obtained vibratome sections of transcardially fixed eyecups. A subset of mice was subjected to subretinal electroporation of the first 201 amino acids of Nphp3, a reported PC marker with a mouse codon-optimized version of red fluorescent protein mKate2. The samples were processed for immunofluorescence and confocal analysis with antibodies specific for PC components.
Anti-acetylated tubulin antibody and Nphp3-mKate fusion protein detected the presence of PC in the apical membrane of RPE in either newborn or adult mice. Analysis of the detached neural retina during RPE flatmount preparation detected RPE PC attached to the photoreceptors. The length and position of the RPE PC were variable, with areas where the PC was short and positioned close to the tight junctions, and areas with long PC positioned above the nuclei.
Our results provide evidence that adult RPE express PC, suggesting a possible role in the normal physiology of RPE and developmental processes controlled by RPE.
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