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Summary: Yu Tian Wang conducts investigations into the molecular mechanisms underlying AMPA receptor trafficking and synaptic plasticity.

Signaling and communication between cells is a pervasive and core issue in the biological sciences. Neurons in the mammalian brain accomplish these functions through a process known as synaptic transmission. It is known that the strength of synaptic transmission is not static but rather subject to dynamic regulation; plastic changes in synaptic strength are believed to be intimately associated with learning and memory, developmental refinement of neuronal circuitry, and the pathogenesis of a large number of brain disorders. Thus, understanding how the strength of synaptic transmission is controlled has been the subject of intense investigation and has been challenging neurobiologists for decades. The efficiency of synaptic transmission at any given synapse can be modified by altering the quantity of neurotransmitter released from the presynaptic terminal or by changing the activity of postsynaptic receptors. The goal of the long-term research program in my laboratory is to understand mechanisms controlling the efficacy of synaptic transmission via the regulation of postsynaptic receptors and the manner in which disease processes alter these mechanisms.

Study of the mechanisms governing membrane trafficking of postsynaptic ionotropic receptors is critical to our understanding of synaptic plasticity, and hence brain physiology and pathology, and therefore will continue to be the major research focus of our laboratory over the next five years. In particular, we will focus on excitatory amino acid (EAA) and type A γ-aminobutyric acid (GABA_A) receptors, given that these are the principal receptors mediating excitatory and inhibitory synaptic transmission in the brain. Expression of recombinant receptors in heterologous cell lines will allow us to use various molecular techniques and characterize in detail the molecular mechanisms controlling the processes by which these receptors are trafficked. We will then study these mechanisms in a physiological context by extending our results to neurons in primary dissociated cultures, brain slices, and the brains of animals as a function of behavior. The present project will focus on the mechanisms underlying the regulation of EAA-mediated synaptic transmission.

EAAs, such as glutamate, mediate fast synaptic transmission at a vast majority of excitatory synapses in the mammalian CNS. They act primarily on two major subfamilies of postsynaptic, ligand-gated glutamate receptors: AMPAR (α-amino-3-hydroxyl-5-methyl-4-isoxazolepropionic acid receptor) and NMDAR (N-methyl-D-aspartate receptor). AMPARs mediate most of the excitatory postsynaptic current (EPSC) at resting membrane potentials. Given the two prominent characteristics associated with their channels (voltage-dependent Mg²⁺ blockade and high calcium permeability), NMDARs make little contribution to basal synaptic transmission but are critically important for the production of various forms of synaptic plasticity in AMPAR-mediated synaptic transmission. Such plastic changes in synaptic strength are thought to be intimately involved in various normal physiological brain functions, including learning and memory and neuronal circuit development, as well as many neuropathological disorders, such as the neurotoxicity associated with stroke. As a result, the mechanisms underlying synaptic plasticity have been a subject of intensive research in the neuroscience field.

The best-studied examples of such synaptic plasticity are the long-term changes in synaptic efficacy observed at the glutamatergic synapses of the CA1 region of the hippocampus. High-frequency stimulation (HFS) of the Schaffer-collateral-commissural pathway induces long-term potentiation (LTP) while prolonged lower-frequency stimulation of the same pathway often causes long-term depression (LTD) of synaptic transmission at these synapses; both LTP and LTD are considered candidates for cellular mechanisms of learning and memory. Although activation of the NMDAR is required for the induction of these forms of synaptic plasticity, it is the AMPA component that primarily expresses the plastic changes. How the AMPA component is altered is hotly debated; the alteration likely involves both a presynaptic component (altering the structure of presynaptic terminals and/or the amount of glutamate released from these terminals) and a postsynaptic component (long-term modifications of the postsynaptic AMPA receptors).

Traditionally, functional changes in ionotropic receptors have been thought to be achieved mainly by altering the properties of existing receptors through the modulation of channel gating and conductance. However, evidence from electrophysiological studies of the last few years has led to the recent formulation of a promising hypothesis—the “silent synapse.” This hypothesis proposes that some synapses contain functional NMDARs but lack functional AMPARs and are therefore nonfunctional under basal synaptic transmission. Further, it is proposed that such synapses can be converted into functional ones by recruiting AMPARs, thus increasing synaptic strength. However, evidence for such AMPAR recruitment and its achievement was not available until recently. In 1997 we made the novel discovery that insulin produces a long-lasting potentiation of GABA_A receptor–mediated inhibitory postsynaptic currents (IPSCs) in hippocampal CA1 cells by a rapid translocation of intracellular GABA_A receptors to the postsynaptic domain. The study thereby provided the first evidence that ligand-gated ion channel receptors could be rapidly recruited into or away from the postsynaptic domain, and that such a rapid change in the number of postsynaptic receptors is a simple and powerful way to regulate synaptic efficacy. A large body of evidence has since accumulated supporting the view that similar postsynaptic trafficking and translocation of AMPARs is also a critical mechanism underlying LTP and LTD. Thus, as a result of facilitated receptor insertion (exocytosis) into the postsynaptic domain, increased postsynaptic AMPARs contribute to the expression of LTP, whereas decreased numbers of AMPARs mediate certain forms of LTD in the brain owing to enhanced endocytosis of AMPARs away from the postsynaptic domain. However, the details of signaling mechanisms underlying AMPAR trafficking, and hence synaptic plasticity, remain largely unknown.

In our most recent study, we found that LTP-producing protocols lead to selective activation of both the PI3K that is physically associated with AMPARs and Akt, a down-stream serine/threonine kinase of the PI3K signaling pathway. In addition, we found that the intracellular carboxyl tail of GluR1 contains a weak Akt consensus site and can be phosphorylated in vitro by active Akt. These results led to our working hypothesis that Akt-mediated direct phosphorylation of AMPA receptors is a necessary step leading to AMPAR insertion and hence LTP. The main purpose of this project is to test this hypothesis by answering the following questions: 1) Is activation of Akt a necessary and sufficient step in LTP expression? 2) Is Akt activity increased by LTP-producing stimuli and does this increase lead to enhanced in situ GluR1 phosphorylation at the site identified in vitro? 3) If so, is GluR1 phosphorylation by Akt required for AMPAR insertion and LTP expression? These questions will be investigated using a combination of electrophysiological recording of AMPAR-mediated excitatory postsynaptic currents, biochemical characterization of Akt activity, and in situ GluR1 phosphorylation in hippocampal neurons maintained in primary culture or brain slices.

The proposed investigation may reveal a fundamental mechanism for the regulation of synaptic strength within the CNS and is thus expected to contribute significantly to the understanding of a broad range of normal brain physiological processes. Furthermore, as evidence implicating AMPA receptor involvement in excitotoxicity grows, we anticipate that our findings will prove useful for the development of novel therapeutic strategies to afford neuroprotection against many CNS neurodegenerative disorders.

Last updated August 2008