MOLECULAR CHARACTERIZATION OF MUSCARINIC ACH RECEPTORS

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A 5-year molecular-level strategy is proposed for attacking an important problem in vertebrate neurodevelopment: "How do CNS neurons create postsynaptic zones specialized for proper signal transduction?" The elementary activity of the brain is signaling between cells -- mechanisms underlying development, maintenance and plasticity of signal transduction are at the foundation of brain function. Muscarinic acetylcholine receptors have been selected as prototypes for studying CNS postsynaptic specialization. These receptors are implicated in a host of physiological and behavioral processes, including conscious arousal, learning and memory. Moreover, because Alzheimer's disease is associated with the death of ACh- containing cells, new studies of cholinoceptive postsynaptic cells could have considerable relevance to the public health. The proposed work pursues fundamental questions stimulated by three significant discoveries: (1) The CNS (but not heart) expresses a novel 86 kDa "embryonic" type MAChR, abundant during neurite growth, that developmentally down-regulates and is supplanted by a 72 kDa "adult type" molecules; (2) In vivo, neurons stringently regulate the placement of receptors, with all molecules localized to dendrites even before synapses have formed; (3) Developing neurons are "plastic" with respect to receptor placement and expression of 86K/72K molecular types. These new and important observations were made in studies of the chicken CNS, a widely-used model for vertebrate neurodevelopment. Three specific aims have been selected as most critical for expanding these findings: (1) To establish the origin and structure of the 86 kDa and 72 kDa receptor molecules by characterizing chicken receptor genes and their transcription in the developing CNS. (2) To determine where the 86K and 72K molecules are localized relative to synaptic junctions, assessing possible mechanisms underlying placement and plasticity. (3) To compare the function of 86K and 72K receptors, investigating possible novel growth-associated roles during the period of neurite arborization and synaptogenesis.