Development of the blue cone bipolar cell in the mouse retina

神经过程如何发展并与其突触伙伴建立适当的连接是神经科学的基本问题之一。脊椎动物视网膜是研究树突发育和神经元连接的杰出模型系统。其中一个关键的视觉功能，色觉，需要视网膜神经元的精确连接。在小鼠视网膜中，有两种类型的视锥细胞光感受器，短波敏感视锥细胞（s -cone），只表达s -视蛋白，和长波敏感视锥细胞（m -cone），其中许多共表达s -视蛋白。为了产生颜色对应性，来自这两种视锥细胞的信号必须在神经节细胞水平上进行对比之前被分离。s型球果只占总球果总数的2-5%。因此，下游的s锥双极细胞（scbc）面临着从大多数m锥中寻找非常稀疏的s锥的艰巨任务。结果是，造血干细胞发育出一种非常独特的树突乔木，具有细长的树突，与少量s锥接触。s -cone和SCBCs之间的这种独特联系使其成为研究突触前神经元如何影响树突发育和突触后神经元突触靶向的良好模型系统。

How neuronal processes develop and establish proper wirings with their synaptic partners is one of the fundamental questions of neuroscience. The vertebrate retina is an outstanding model system for studying dendritic development and neuronal connections. One of the critical visual functions, color vision, requires precise wiring of retinal neurons. In the mouse retina, there are two types of cone photoreceptors, the short wavelength sensitive cones (S-cones), which only express S-opsin, and the long wavelength sensitive cones (M-cones), many of which co-express S-opsin. In order to generate color opponency, signals from these two types of cones have to be segregated before they are contrasted at the ganglion cell level. S-cones only account for 2-5% of the total cone population. Thus, the downstream S-cone bipolar cells (SCBCs) face the daunting task of seeking out very sparse S-cones from a majority of M-cones. The outcome is that SCBCs develop a very unique dendritic arbor with long, meager dendrites that contact a handful of S-cones. This distinctive connection between S-cones and SCBCs makes it an excellent model system to study how presynaptic neurons affect the dendritic development and synaptic targeting of postsynaptic neurons. SCBs are labeled in a transgenic mouse line expressing Clomeleon (Clm) driven by thy1 promoter. Two mouse models were used to change the density and type of cones. In Thrb2-/- mice which lack thyroid hormone receptor 2 (TR2), M-opsin expression is abolished and all M-cones are turned into S-cones32. In S-opsin-/- mice, the S-opsin gene is knocked out. We crossed two lines and generated a double knockout (DKO) with neither S- nor M-opsin expression. We found that the numbers of SCBs in Thrb2-/-, S-opsin-/- and DKO mice are similar to those in wildtype. Morphologically, SCBs in Thrb2-/- and S-opsin-/- mice were indistinguishable from those in wildtype in terms of number of dendritic branches and cone contacts. SCBs in these mice appear to target specific true S-cones, even though the type of opsin expression is identical across all cones. Our results suggest that 1) dendritic development of SCBs appears to be intrinsic; 2) S-cone identity may be specified by factors other than S-opsin expression a hypothesis that we will investigate in the future. In order to compare the transcriptome of S- and M-cones, we need to isolate true S-cones. An existing transgenic mouse line that expresses EGFP driven by an S-opsin promoter is not reliable in that some S-opsin negative cones also express GFP. This is likely due to the short S-opsin promoter region used in generating the mouse line. Due to low percentage of true S-cones (5%), this contamination from M-cones will be ruinous for RNAseq. We thus are currently working on generating our own mouse line using the Bacterial Artificial Chromosome (BAC) recombination approach to include longer upstream and downstream regulatory sequences at the endogenous S-opsin gene locus. We are sceening mouse lines that express yellow fluorescent protein YFP (Venus) only in S-opsin expressing cones.