ENDOGENOUS REGULATORS OF GLUTAMATE-ACTIVATED CHANNELS

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A basic understanding of many neuropsychiatric disorders has focused on the synapse as a critical site in the disease process; this has led in several examples to design of successful therapeutic strategies. In neurology and psychiatry, much of this emphasis has been on the catecholamine pathways, however it is increasing apparent that excitatory amino acids acting as neurotransmitters and neuromodulators also play key physiological roles. L-glutamate activates several receptor subtypes including the N-methyl-D- aspartate (NMDA) receptor, and is the major excitatory transmitter in many brain regions including the limbic system. The NMDA receptor has recently been shown to underlie a slow excitatory postsynaptic potential at these synapses. This transmitter system has been implicated in neurodegenerative diseases such as Huntington's and Alzheimer's disease, in ischemic brain injury, and in learning and memory. Recent studies, particularly of the NMDA receptor/channel, suggest that glutamate-activated receptors and ion channels have a complex pharmacology including several modulatory sites which dramatically influence the function of the receptor/channel complex; both endogenous and exogenous agents including psychoactive drugs such as phencyclidine (PCP) act on these modulatory sites. However, little is known of the structure or detailed physiological mechanisms of these ligand-gated ion channels, thus the ability to manipulate this system therapeutically is presently limited. The long range goal of this project is relate the function of these receptors and their modulatory sites to their structure; this approach should allow the design of more effective therapeutic agents as well as guide molecular biological approaches to these receptors. The objective of this proposal is to examine in detail a known endogenous regulator of the NMDA channel as a step toward an understanding of channel function. The focus will be on two important, unresolved aspects of glutamate receptor functions: 1. the mechanism of NMDA channel regulation by transition metal ions including zinc; and 2. the kinetic properties of NMDA channels evoked by synaptic stimulation. Two preparations will be used: primary dissociated cultures of rodent hippocampus, and Xenopus oocytes injected with poly (A) + mRNA purified from rat hippocampus. Single channel and voltage clamp electrophysiological methods will be used. Agonist-gated channels will be activated by exogenously applied agonists; as well as by activation of single presynaptic neurons in the cultures hippocampal preparation. It is expected that these results will provide important new information concerning the function and regulation of glutamate receptors in the CNS.