The goal of the work is to understand, at the molecular level, how the interaction of excitable cells results in the formation and muturation of sites of neurotransmission. The approach to this problem is to understand the structure and function of the acetylcholine receptor and the mechanism by which the innervating neuron controls the properties of this neurotransmitter. In the past, the research has focussed upon the acetylcholine receptor at the neuromuscular junction, but recent advances in the molecular biology of the receptor now make it possible to study the nicotinic acetylcholine receptor and cholinergic transmission in the central nervous system. We now have several cDNA clones which encode proteins that we propose are neuronal nicotinic acetylcholine receptor Alpha-subunits. We will determine whether these clones encode a protein which can assemble with or without additional subunits to form a ligand gated ion channel. Initially these experiments will be performed using expression in oocytes or mamallian cell lines in conjection with our available clones encoding the muscle receptor subunits and will provide physiological access to the neuronal nicotinic acetylcholine receptors. We will isolate the neuronal analogues of the muscle Beta-, Gamma-, and Delta-subunits for use in similar expression studies over the long term. Hybrid Alpha-subunits composed of portions of the muscle and neural Alpha-subunits will be constructed to determine which portions of the Alpha-subunits contribure which functions to the receptor oligomer. Antibodies against the several neural Alpha-subunits will be made and used to determine the distribution of nicotinic cholinergic transmission in the central nervous system and, in conjunction with in situ hybridization, will allow us to study the formation and maturation of synapses in the central nervous system.
Showing the most recent 10 out of 33 publications
