Evaluating the Role of Acetylcholinesterase (AChE) in Axonal Outgrowth Using AChE Knockout Mice

Johns Hopkins University, Baltimore MD

Investigators

Abstract

0112111 Jett Abstract The morphology of neurons is a primary determinant of connectivity patterns in the adult nervous system, and the formation of functional neural circuits depends upon precise regulation of the outgrowth of axons from neurons during development of the nervous system. It is clear that developing neurons use specific environmental cues to regulate the growth of axons in space and time. One cue that is emerging as a potentially important factor in controlling axonal growth is acetylcholinesterase (AChE). While AChE is best known as the enzyme that metabolizes the neurotransmitter acetylcholine, recent evidence suggests that AChE also promotes axon outgrowth. Thus, the spatiotemporal expression of AChE in some neuronal cell types is consistent with its proposed role in axonal outgrowth. Functional studies using pharmacological inhibitors of AChE or genetic manipulations of AChE levels in cultured cells further support the hypothesis that AChE promotes axonal outgrowth, and suggest that the morphogenic activity of AChE is not dependent on its catalytic activity. It has been proposed that AChE promotes axonal growth via adhesive mechanisms since analyses of its structure indicate significant homology with cell adhesion molecules known to function in neuronal morphogenesis. However the relevance of these observations to regulation of axonal outgrowth in laboratory animals has yet to be directly tested. Dr. Jett and co-workers will address this by examining axonal morphology in the developing central and peripheral nervous systems of the recently described transgenic mouse that lacks the gene for AChE. Dr. Jett will: (1) Determine if the targeted disruption of the AChE gene alters axonal growth patterns in the nervous system of mice; and (2) Determine if aberrations in axonal morphology can be rescued by addition of exogenous AChE. The first specific aim will be accomplished by comparing spatial and temporal patterns of axonal growth of neurons in AChE knockout mice to those in wild-type mice. Axonal outgrowth will be visualized using specific labeling dyes and immunocytochemical localization. The gain-of-function studies proposed in the second specific aim will be performed by adding recombinant AChE to cultures of DRG neurons derived from AChE knockout mice. Axonal morphology will be quantified with respect to number, length and branching using computerized morphometric analyses of neuronal cultures immunostained for neurofilaments. It is anticipated that results from these studies will further our understanding of the basic mechanisms that control neuronal connectivity in the developing nervous system.