This project employs the hippocampal formation of the rat brain to investigate mechanisms of glutamate transmission. The present proposal focuses on the physiological role of proline in regulating glutamate transmission and on the pathophysiological consequences of elevated proline concentrations in the genetic disorders known as hyperprolinemia. Proline is an excitatory and excitotoxic imino acid that can block memory formation and induce retrograde amnesia. Moreover, hyperprolinemia is associated with a much higher than normal incidence of childhood seizures and mental retardation. A Na+/Cl--dependent, high affinity transporter specific for pro-line is expressed exclusively by a subset of glutamate pathways. Proline, at a concentration typical of human cerebrospinal fluid, potentiates transmission in one such pathway: the Schaffer collateral-commissural projection to area CA1. "Proline-induced potentiation" is NMDA receptor-dependent and long-lasting. These observations suggest that some glutamate synapses in vivo normally exist in a partially potentiated state due to the continuous occupation of a small fraction of NMDA receptors by proline. Mechanistic studies of proline-induced potentiation will be carried out with use of field potential and whole cell patch clamp recordings from CA1 pyramidal cells. Specific objectives include determining to what extent proline-induced potentiation utilizes mechanisms that overlap those involved in electrically-evoked long-term potentiation, whether low concentrations of proline actually evoke a measurable NMDA current in these cells, whether proline-induced potentiation involves presynaptic as well as postsynaptic mechanisms, whether both AMPA and NMDA components of the postsynaptic response are potentiated and whether proline modifies GABA inhibition. Another set of studies will examine the role of the proline transporter in proline-induced potentiation. The objective is to determine whether proline must be taken up for potentiation to occur or whether transport serves simply to regulate the local extracellular proline concentration. These studies will take advantage of findings that the tetrapeptide gly- gly-phe-leu potently and selectively inhibits high affinity proline transport and that the lateral and medial divisions of the hippocampal perforant path differ markedly in their ability to take up proline. A final study will investigate the mechanism by which proline depresses glutamate release. This effect is observed with concentrations of proline that are present in the cerebrospinal fluid of persons having hyperprolinemia type II, but not with normal proline concentrations. Findings from the proposed studies will elucidate a novel regulation of glutamate transmission that may be an attractive target for drug development. They may also prove relevant to synaptic mechanisms of memory and learning.