The functional properties of the muscle acetylcholine (ACh) receptor channel are determined by the type of innervation, the contractile characteristics of the muscle fiber, and the level of muscle membrane activity. The nicotinic ACh receptor on vertebrate skeletal muscle has several functionally distinct forms, and the proportions of these different channel types change during development. Specifically, channels undergo a dramatic decrease in open time during early development, followed at a later time by an increase in single channel conductance and further shortening in open time. How are these distinct changes in ACh receptor properties programmed during development? We will use several different approaches to analyze the developmental changes in ACh receptor function. First, we will identify precisely the structural alterations which impart the different functional properties to the ACh receptor. Xenopus myotomal muscle is uniquely suited for these studies because it exhibits developmental alterations in channel function both in vivo and in cell culture. We will combine single channel recording technique with recombinant DNA techniques to identify structural correlates underlying the physiological changes in channel function. Then, we will prove that these alterations in subunit mRNAs account for the developmental changes in ACh receptor channel function by expressing the full length cDNAs in Xenopus oocyte or heterologous cell expression systems. Blocking developmental changes in functional properties by injection of subunit specific antisense RNA will also be done. We will examine, at the molecular and functional level, the effect of synapse formation on in vitro muscle in order to ascertain whether nerve alters receptor properties by altering gene expression. Additionally, we will test second messengers known to be present in nerve, and which have been shown to alter levels of receptor synthesis, for effectiveness in promoting the developmental changes in ACh receptor channel function. This combination of molecular and physiological approaches to the study of this channel will provide answers to the long standing questions relating to neuronal control of the properties of postsynaptic ACh receptor channels.
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