The broad objective of the proposed research is to deepen our understanding of the structure, function, and inhibition of the receptor molecules responsible for synaptic transmission. The research will focus on a subtype of glutamate receptor specifically activated by N-methyl-D-aspartate (NMDA). The family of excitatory receptors activated by the neurotransmitter glutamate is responsible for the majority of fast synaptic communications within the vertebrate nervous system. The NMDA receptor is thought to be of fundamental importance in synaptic transmission, synaptic plasticity, and development of the nervous system. It is also directly or indirectly involved in many nervous system diseases, including epilepsy, schizophrenia, ischemia, and a variety of degenerative diseases. Improved understanding of a receptor involved is such a remarkable range of processes in normal and diseased nervous systems will lead to an enhanced ability to prevent, or ameliorate the consequences of, nervous system dysfunction. Possibly because it must function in numerous neural systems, the NMDA receptor is regulated by multiple mechanisms. One mode of NMDA receptor regulation blockade by Mg2+ of the ion-conducting channel formed by the NMDA receptor. Mg2+, both from the extracellular (Mg2+e) and intracellular (Mg2+i) sides of the membrane, can enter the NMDA-activated channel and obstruct the flow of ions. Block by Mg2+i will be studied using the patch-clamp technique to measure current flow across the membranes of cultured neurons and of Chinese hamster ovary (CHO) cells that have been genetically modified to express NMDA receptors. The mechanism by which Mg2+i inhibits neuronal and CHO cell responses will be investigated by simultaneously measuring NMDA responses and the concentration of Mg2+i. Numerous drugs can block the NMDA-activated channel. The effects on humans of these drugs are diverse: some act as hallucinogens and are used as drugs of abuse; others are well tolerated clinically and are used in the treatment of neurodegenerative diseases. The mechanisms by which such drugs inhibit NMDA receptors will be examined in the proposed research. The influence of permanent ions on the interaction between blocker and channel also will be investigated. Finally, the results will be integrated into a kinetic model of the interaction of channel blockers with the NMDA receptor. By modeling and comparing block by several different compounds, basic rules of blocker-channel interactions and their dependence on blocker structure will be developed. The results will improve our understanding of channel function and how it can be influenced by drug binding, and may facilitate the design of clinically useful neuroprotective agents.
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