Excessive Expression of Acetylcholinesterase Impairs Glutamatergic Synaptogenesis in Hippocampal Neurons

Abstract

Acetylcholinesterase (AChE) exerts noncatalytic activities on neural cell differentiation, adhesion, and neuritogenesis independently of its catalytic function. The noncatalytic functions of AChE have been attributed to its peripheral anionic site (PAS)-mediated proteinprotein interactions. Structurally, AChE is highly homologous to the extracellular domain of neuroligin, a postsynaptic transmembrane molecule that interacts with presynaptic ␤-neurexins, thus facilitating synaptic formation and maturation. Potential effects of AChE expression on synaptic transmission, however, remain unknown. Using electrophysiology, immunocytochemistry, and molecular biological approaches, this study investigated the role of AChE in the regulation of synaptic formation and functions. We found that AChE was highly expressed in cultured embryonic hippocampal neurons at early culture days, particularly in dendritic compartments including the growth cone. Subsequently, the expression level of AChE declined, whereas synaptic activity and synaptic proteins progressively increased. Chronic blockade of the PAS of AChE with specific inhibitors selectively impaired glutamatergic functions and excitatory synaptic structures independently of cholinergic activation, while inducing AChE overexpression. Moreover, the PAS blockade-induced glutamatergic impairments were associated with a depressed expression of ␤-neurexins and an accumulation of other synaptic proteins, including neuroligins, and were mostly preventable by antisense suppression of AChE expression. Our findings demonstrate that interference with the nonenzymatic features of AChE alters AChE expression, which impairs excitatory synaptic structure and functions.

Differential localization of acetylcholinesterase in neuronal and non‐neuronal cells

“…It was also surprising that CGN express such a high level of catalytic activity and protein, because CGN are primarily glutamatergic, however express cholinergic receptors, but have not previously been shown to express high levels of AChE [Schousboe et al, 1985;Fucile et al, 2004]. AChE has been shown to be involved transiently in neuritogenesis, and the CGN may represent immature neurons that express AChE prior to other more mature glutamatergic markers [Layer, 1990;Dong et al, 2004].…”

Peripheral Site Acetylcholinesterase Blockade Induces RACK1-Associated Neuronal Remodeling

“…Primary Cell Culture Dissociated embryonic hippocampal neurons were grown as reported [14] . Neurons were treated at the 3rd or 5th day in culture (DIC), for 3 days with 3-10 M of the positively-charged PAS ligand, 1,5-bis (4-allyldimethylammoniumphenyl) pentan-3-one dibromide (BW284c51/BW, Sigma, St. Louis, Mo., USA), the active site AChE inhibitor physostigmine or the rat AChE antisense EN101 (12 g/ml) or inverse oligonucleotide INV101 (12 g/ml) together with BW [22] .…”

“…Therefore, an extracellular PAS blockade, which affects Neuroligin 1 expression, could be predicted to interfere with neuritogenesis and/or modify neuronal cytoarchitecture and synaptic activity. Indeed, our recent study found that inhibition of the peripheral site of AChE induces overexpression of this molecule accompanied by impaired glutamatergic synaptogenesis [14] .…”

Neurexin–neuroligin signaling in synapse development

“…The major extracellular domain of neuroligins is homologous to acetylcholinesterase (AChE) but lacks cholinesterase activity and mediates binding to neurexins. Curiously, overexpression of AChE reduces levels of β-neurexins in vivo and in culture, and impairs genesis of glutamatergic synapses in culture, indicating that there is cross-talk between the two proteins [13,14]. Neuroligins form homomultimers through the AChE-homologous domain, a structural association that is important for neuroligin function [15,16].…”