Acetylcholine receptor (AChR) channels undergo a shift from long to brief open times during the development of amphibian and mammalian skeletal muscle and during the regeneration of synapses. A similar change also occurs in glycine receptors and glutamate receptors in developing neural tissue. Although the functional significance of this change is not understood, its conservation across species and receptor types suggests that it is important to synaptogenesis. In addition to the developmental change in gating, AChRs in developing amphibian muscle exhibit a lengthening of open time in response to calcitonin gene-related peptide (CGRP), a neuropeptide released by motor neurons. We have recently cloned cDNAs encoding the beta and epsilon subunits of the Xenopus AChR. These clones, in combination with the previously cloned alpha, gamma, and delta subunits, give us a set of expressible subunit clones for the embryonic and adult forms of the Xenopus nicotinic AChR and open the possibility of studying the structure/function relations of embryonic and adult receptors using molecular techniques. The proposed experiments are aimed at explaining the molecular basis of changes in AChR open time in developing muscle and the modulation of AChR channel open time by CGRP. AChRs will be expressed in oocytes and in developing embryos and will be studied by single channel recording. We expect the results from our study of Xenopus AChRs to be directly relevant to understanding developing mammalian AChRs, because of the strong homologies across species, and to be more broadly relevant to understanding the development of other ligand-gated channels.
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