This proposal addresses basic questions concerning the electrophysiological and molecular pharmacological mechanisms of the recently identified and essentially uncharacterized heterogeneous populations of nicotinic acetylcholine receptors (AChRs) located in various regions of the brain. The precise knowledge of the functional properties of such an elusive receptor is of major significance for the understanding of the physiology of the brain under normal conditions and in disease states. The novel neurotoxin (+) anatoxin-a (AnTX) and its synthetic analogs including the (-) enantiomer have proven essential to uncover the structural requirements for agonism at the muscle AChR. Furthermore, by virtue of its potency and specificity, (+)AnTX has enabled the recording of AChR currents in the central nervous system (CNS) and to reveal a great degree of heterogeneity of the AChRs, both within the same cell and among different cell types. Thus, the new long-term goal of this project is to investigate the fundamental functional and pharmacological properties of the AChR at different stages of development and under conditions of chronic exposure to selected drugs. In addition to ACh and the AnTX analogs, other selected weak agonists with similar specificity will be used to investigate AChR responses in the CNS. The noncompetitive antagonists that have been documented for the peripheral AChR will now be studied in the CNS, including a novel series of acridine araphane analogs which function as sensitive """"""""rulers"""""""" to define the antagonist sites on the AChR. The effects of these selective toxins will also be studied at the N-methyl-D-aspartate receptor because of the great deal of structural and functional homology that has been observed with peripheral and central AChR. These studies are essential for the understanding of the interrelationships of homologous ligand-gated channels and related disease states such as Alzheimer's dementia. The proposed experiments will utilize a variety of electrophysiological techniques, including a new fast drug perfusion and withdrawal system developed in this laboratory, coupled with ligand binding, fluorescence labelling, and kinetic studies of receptors, and morphological studies of acutely dissociated and tissue cultured neuronal cells to reveal the basic mechanisms underlying the function of these different forms of AChR.
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