Asymmetric N-Cadherin Expression Results in Synapse Dysfunction, Synapse Elimination, and Axon Retraction in Cultured Mouse Neurons

Abstract

Synapse elimination and pruning of axon collaterals are crucial developmental events in the refinement of neuronal circuits. While a control of synapse formation by adhesion molecules is well established, the involvement of adhesion molecules in developmental synapse loss is poorly characterized. To investigate the consequences of mis-match expression of a homophilic synaptic adhesion molecule, we analysed an asymmetric, exclusively postsynaptic expression of N-cadherin. This was induced by transfecting individual neurons in cultures of N-cadherin knockout mouse neurons with a N-cadherin expression vector. 2 days after transfection, patch-clamp analysis of AMPA receptor-mediated miniature postsynaptic currents revealed an impaired synaptic function without a reduction in the number of presynaptic vesicle clusters. Long-term asymmetric expression of N-cadherin for 8 days subsequently led to synapse elimination as indicated by a loss of colocalization of presynaptic vesicles and postsynaptic PSD95 protein. We further studied long-term asymmetric N-cadherin expression by conditional, Cre-induced knockout of N-cadherin in individual neurons in cultures of N-cadherin expressing cortical mouse neurons. This resulted in a strong retraction of axonal processes in individual neurons that lacked N-cadherin protein. Moreover, an in vivo asymmetric expression of N-cadherin in the developmentally transient cortico-tectal projection was indicated by in-situ hybridization with layer V neurons lacking N-cadherin expression. Thus, mis-match expression of N-cadherin might contribute to selective synaptic connectivity.

Release activity-dependent control of vesicle endocytosis by the synaptic adhesion molecule N-cadherin

“…As characterized previously37, loss of N-cadherin at the protein level requires several days following Cre transfection. First, we studied vesicle recycling in Cre-transfected neurons by extracellular stimulation-induced (400 stimuli at 20 Hz) uptake of FM4-64 (10 μM) in vesicles leading to fluorescent FM4-64 puncta on dendrites.…”

“…To further study the role of N-cadherin in regulating vesicle endocytosis, we performed a conditional knockout of N-cadherin in individual cortical neurons in cultures from floxed N-cadherin mice35 by sparse expression of Cre3637 (transfection at 9 DIV, analysis of postsynaptic N-cadherin deficient neurons at 12–15 DIV). As characterized previously37, loss of N-cadherin at the protein level requires several days following Cre transfection.…”

Candidate genes and functional noncoding variants identified in a canine model of obsessive-compulsive disorder

“…Disrupting expression of CDH2 in cultured mouse neurons causes synapse dysfunction, synapse elimination and axon retraction [35]. …”

Cadherins and catenins in dendrite and synapse morphogenesis

“…50 The mechanism underlying this functional ability of N-cadherin remains unclear, but it has been suggested that cis-interactions of N-cadherin with non-cadherin receptors or lack of trans-synaptic N-cadherin signaling promotes retrograde signaling via unknown signaling pathways. Further experimental evidence is necessary to determine if either of these models is pertinent.…”