Developing cortical neurons express a tightly choreographed sequence of cytoskeletal and transmembrane proteins to form and strengthen specific synaptic connections during circuit formation. Nectin-3 is a cell-adhesion molecule with previously described roles in synapse formation and maintenance. This protein and its binding partner, nectin-1, are selectively expressed in upper-layer neurons of mouse visual cortex, but their role in the development of cortical circuits is unknown. Here we block nectin-3 expression (via shRNA) or overexpress nectin-3 in developing layer 2/3 visual cortical neurons using in utero electroporation. We then assay dendritic spine densities at three developmental time points: eye opening (postnatal day (P)14), one week following eye opening after a period of heightened synaptogenesis (P21), and at the close of the critical period for ocular dominance plasticity (P35). Knockdown of nectin-3 beginning at E15.5 or ~ P19 increased dendritic spine densities at P21 or P35, respectively. Conversely, overexpressing full length nectin-3 at E15.5 decreased dendritic spine densities when all ages were considered together. The effects of nectin-3 knockdown and overexpression on dendritic spine densities were most significant on proximal secondary apical dendrites. Interestingly, an even greater decrease in dendritic spine densities, particularly on basal dendrites at P21, was observed when we overexpressed nectin-3 lacking its afadin binding domain. These data collectively suggest that the proper levels and functioning of nectin-3 facilitate normal synapse formation after eye opening on apical and basal dendrites in layer 2/3 of visual cortex.

中文翻译：

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出生后视觉皮层突触的正确形成需要精确的 nectin-3 水平


发育中的皮质神经元表达严格编排的细胞骨架和跨膜蛋白序列，以在电路形成过程中形成和加强特定的突触连接。 Nectin-3 是一种细胞粘附分子，在突触形成和维持中具有先前描述的作用。这种蛋白及其结合伙伴 nectin-1 在小鼠视觉皮层的上层神经元中选择性表达，但它们在皮层回路发育中的作用尚不清楚。在这里，我们使用子宫内电穿孔在发育中的 2/3 层视觉皮层神经元中阻断 nectin-3 表达（通过 shRNA）或过度表达 nectin-3。然后，我们在三个发育时间点测定树突棘密度：睁眼（出生后天（P）14）、突触发生增强期后睁眼后一周（P21）以及眼优势可塑性关键期结束时（P35）。从 E15.5 或 ~ P19 开始敲低 nectin-3，分别增加 P21 或 P35 处的树突棘密度。相反，当所有年龄一起考虑时，在 E15.5 过表达全长 nectin-3 会降低树突棘密度。 nectin-3 敲低和过度表达对树突棘密度的影响在近端次级顶端树突上最为显着。有趣的是，当我们过度表达缺乏 afadin 结合域的 nectin-3 时，观察到树突棘密度的更大下降，特别是 P21 处的基底树突。这些数据共同表明，nectin-3 的适当水平和功能有助于睁眼后视觉皮层 2/3 层顶端和基底树突上的正常突触形成。