Project: Research project

Project Details


The long-term objective of the research proposed in this application is to
determine, at a molecular level, how motor neurons exert their trophic
influence on rat skeletal muscle acetylcholinesterase (AChE). There is
good evidence that the influence of nerve on muscle AChE is mediated in
part by the activity (action potentials and/or contraction) set up in
muscle by nerve and in part by a factor (AIF) that is delivered to muscle
by nerve. We plan to improve our bioassay for AIF so that we can purify
and characterize this factor efficiently. Muscle cells produce three major
types of AChE; soluble globular forms that are secreted, integral
sarcolemmal globular forms that are externalized with their active sites
exposed, and asymmetric forms that appear to associate with the basal
lamina that ensheathes each muscle fiber. Our working hypothesis is that
each of these forms is assembled intracellularly from the monomeric form of
the enzyme, externalized, and then lost by secretion or local degradative
processes. We propose that the activity set up in muscle by nerve
increases the rate at which all forms of the enzyme are synthesized (we
have good evidence for this) and that innervation selectively increases the
rate at which asymmetric forms are produced at the end-plate and decreases
the rate at which asymmetric forms are degraded at the endplate. AIF
selectively maintains endplate enzyme and probably acts by altering the
metabolism of endplate asymmetric forms in one of the ways postulated.
Using two different methods to evaluate AChE metabolism, we plan to test
these hypotheses in various ways by examining three preparations - cultured
embryonic rat myotubes, organ-cultured rat diaphragm, and model innervated
muscles prepared by explanting phrenic nerve - hemidiaphragm preparations
into organ culture and stimulating the nerve to maintain AChE at innervated
levels. In an effort to extend our analysis to the mRNA level, we will
evaluate the Xenopus oocyte as a system for determining if the increased
rate of AChE synthesis that we have observed in active (fibrillating)
embryonic rat myotubes is correlated with an increased amount of AChE mRNA.
StatusNot started


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