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Li Jia, Wei Li; Cone Photoreceptor Afferents and Dendritic Development of S-cone Bipolar Cells in the Mouse Retina. Invest. Ophthalmol. Vis. Sci. 2014;55(13):3520.
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© ARVO (1962-2015); The Authors (2016-present)
To gain insight into guidance mechanisms that regulate the formation of specific connections between true S cones and S cone bipolar cells (SCBCs) in the retina, we ask whether altering the number and type of cone afferents affects the dendritic development and synapse formation of the postsynaptic SCBCs. To identify factors that distinguish true S cones from M cones for specific synaptic targeting, we set out to purify S and M cones and compare their gene expression profiles.
SCBCs are labeled in a transgenic mouse line expressing Clomeleon (Clm) driven by thy1 promoter. Thrb2-/-, Sop-/- and Clm1 mice were crossed to generate Clm;Thrb2+/+, Clm;Thrb2-/-, Clm;Sop-/- and Clm;Thrb2-/-;Sop-/- mice. Whole mount retinae were immunolabeled with GFP, cone arrestin, Sopsin and then imaged on confocal. The dendritic morphology of SCBCs was analyzed and compared. True S cones were FACS sorted from a BAC transgenic mouse line, Sop-Venus, where Venus (a variant of GFP) is inserted in the endogenous Opn1sw locus on the BAC clone. M cones were sorted from Mop-cre/ZEG mice in which M cones are labeled with EGFP.
Two mouse models with altered densities and types of cones were used. In Thrb2-/- mice, which lack thyroid hormone receptor β2 (TRβ2), M-opsin expression is abolished and all cones become S-cones. In Sopsin-/- mice, the Sopsin gene is knocked out. We obtained Clm;Thrb2+/+, Clm;Thrb2-/-, Clm;Sopsin-/- and Clm;Thrb2-/- ;Sopsin-/- (DKO) mice and compared the dendritic morphology of SCBCs in these mice. We found that the numbers of SCBCs in Thrb2-/-, Sopsin-/- and DKO mice are comparable to that in wildtype. Morphologically, SCBCs in Thrb2-/- and Sopsin-/- mice are indistinguishable from those in wildtype in terms of dendritic length, number of dendritic branches and number of cone contacts. To obtain a pure population of true S cones, we focused on the dorsal third of the Sop-Venus retina where individual cones express either Sopsin or Mopsin exclusively. We have sorted true S cones and M cones using FACS and their expression profiles are being compared using RNAseq.
Our results show that dendritic development of SCBCs does not depend on the expression of Mopsin or Sopsin. Specific connections between true S cones and SCBCs appear to depend on factors other than cone opsin, which we are exploring by comparing gene expression patterns of true S and M cones.
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