Abstract
BackgroundDeveloping 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.MethodsHere 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).ResultsKnockdown 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.ConclusionThese 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.
Highlights
Developing cortical neurons express a tightly choreographed sequence of cytoskeletal and transmembrane proteins to form and strengthen specific synaptic connections during circuit formation
Overexpressing Nec3Δafadin produced an even greater decrease in dendritic spine densities than full-length nectin-3, on basal dendrites at P21. These results collectively indicate that the neuronal regulation of synaptic nectin-3 may mediate both the formation and removal of synapses in layer 2/3 (L2/3) visual cortex during critical periods of development
In situ hybridization (ISH) revealed that nectin-3 and nectin-1 had specific expression patterns in hippocampus, where nectin-3 is enriched in CA3 and nectin-1 is enriched in the dentate gyrus, and in cortex, where nectin-3 and nectin-1 are both enriched in upper layer cortical neurons (P16; Fig. 1b)
Summary
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. The development of mature brain circuits requires the selective formation, elimination, and strengthening of synaptic connections between neurons [1, 2] This process requires the timed expression and spatial localization of proteins that are just beginning to be elucidated [3,4,5]. We sought to characterize the role of the cell-adhesion molecule nectin-3 in spine density profiles in developing layer 2/3 (L2/3) visual cortical neurons
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