A series of experiments are proposed to estimate with a higher degree of accuracy the time course and magnitude of the glutamate concentration in the synaptic cleft following exocytosis of a synaptic vesicle. A mathematical model of AMPA receptor kinetics will be refined through a series of kinetic experiments using excised membrane patches from CA1 hippocampal neurons. This model will be combined with recordings of AMPA receptor-mediated synaptic responses in cultured CA1 hippocampal neurons to reach an estimate of the size and shape of the glutamate transient. While similar experiments have been performed with NMDA receptor responses, the faster-AMPA receptor kinetics will result in a substantial improvement in temporal resolution, allowing us to predict more accurately the early phase of the glutamate transient and, therefore, the fraction of postsynaptic AMPA receptors that is activated during a synaptic response. This prediction, which impacts heavily on recently proposed mechanisms of synaptic plasticity and the physiological importance of multivesicular release, will be tested in another set of experiments designed to determine whether AMPA receptors are saturated by a single quantal event and during evoked responses. Understanding the degree to which AMPA receptors are bound and activated during a synaptic response will provide valuable insight into questions regarding the modulatory effects, at the synaptic level, of various mechanisms thought to regulate synaptic efficacy, including long-term potentiation.
