The amino acid glutamate is the major excitatory synaptic transmitter in the vertebrate central nervous system (CNS). Two classes of glutamate- gated ion channels, one responsive to the agonist N-methyl-D-aspartate (NMDA) and another selectively activated by alpha-amino-5-methyl- isoxazole-4-propionate (AMPA), are thought to play crucial roles in activity-dependent synaptic plasticity during cortical development and learning. In order to investigate the molecular mechanisms by which these two receptors exert their function, we have generated three types of transgenic mice carrying dominant-acting subunits of glutamate receptors. The first type of transgene encodes a mutant R1 subunit of the NMDA receptor which effectively blocks Ca2+ influx with little effect on monovalent ion permeability, as determined by expression studies in Xenopus oocytes. This mutation also eliminates the Mg2+ blockade of the ion channel. Expression of this transgene should impair the Ca2+ permeability of NMDA receptors in these mice. The second transgene encodes another mutation within the NMDA-R1 subunit which diminishes the voltage-dependent Mg2+ blockade when expressed in Xenopus oocytes. NMDA receptors expressed in these transgenic mice should exhibit increased permeation of both monovalent and divalent cations. The third transgenic mouse is designed so that a splice variant (the flip form) of an AMPA receptor subunit is ectopically expressed in the CA1 region and dentate gyrus of the hippocampus. The flip form of the receptor subunit generates larger current responses than the flop form normally expressed in these areas. It is predicted that expression of this transgene will lead to increased activity of AMPA receptors in area CA1 and the dentate gyrus of the hippocampus. The following experiments are planned for each mutant line. First, expression of the transgene will be confirmed by both RNA analyses and electrophysiological recordings. Second, the brains of transgenic mice will be examined under light microscopy to identify any morphological abnormalities. Third, processes involving activity-dependent synaptic plasticity will be studied in developing cortex. Fourth, cognitive abilities of the mice will be tested using the Morris water maze. Together, the proposed studies should provide considerable insight into the mechanisms by which glutamate receptor function mediates the synaptic plasticity that accompanies cortical development and cognitive function.
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