Visualization of Molecular Mechanism of Memory using New Experimental Methods
The work of the research team led by Professor Tomoo Hirano and Mr. Hiromitsu Tanaka of the Graduate School of Science was published online in Cell Reports.
When we learn and remember something, the efficacy of information transmission between neurons change in the brain. The information transmission occurs at a synapse, where a presynaptic neuron secretes transmitter molecule such as glutamate, and receptor proteins on the postsynaptic membrane capture transmitter and respond. Repeated use of a synapse increased the transmission efficacy for long-term, which is called long-term potentiation (LTP), and contributes to the formation of memory. The increase in number of AMPA type glutamate receptor (AMPAR) is a main molecular mechanism for LTP. However, when and how different types of AMPAR increase in the postsynaptic membrane had been unclear. Therefore, we addressed this issue by developing new experimental methods to visualize the change and movement of AMPAR around postsynaptic membrane.
We thought to use total internal reflection microscopy (TIRFM) in order to observe AMPAR tagged with a fluorescent molecule with a high signal-to-noise ratio. TIRFM detect only the fluorescence signal originating from molecules located very close (<100 nm) to the glass surface, thus reducing the background fluorescence to a very low level. In order to maximally utilize the excellent resolution power of TIRFM for AMPAR observation, we tried to form the postsynaptic membrane directly on the glass. Here, the glass was coated with neurexin molecule, which is known to induce differentiation of postsynaptic membrane, and neurons were cultured on it, resulting in the formation of postsynaptic membrane on the glass. In this study, neurons were prepared from the hippocampus, a brain area essential for the memory formation, and where LTP takes place robustly. Next, we expressed AMPAR tagged with a fluorescent molecule, and LTP was induced. The changes in number of AMPAR, exocytosis and lateral movement were observed during LTP. By the way, each AMPAR is composed of 4 subunits GluA1-4. In the hippocampus, main AMPARs are GluA1/GluA2 hetero-tetramer and GluA2/GluA3 hetero-tetramer. However, how they change during LTP had been elusive. Contribution of GluA1 homo-tetramer to LTP was also proposed, but others challenged this idea. We demonstrated that GluA1 homo-tetramer, GluA1/GluA2 hetero-tetramer and GluA2/GluA3 hetero-tetramer increased in the postsynaptic membrane through distinct pathways during LTP. This study contributes to deeper understanding of the LTP mechanism. We also would like to note that the methods developed in this study, are applicable to studies focusing on the movement and function of membrane bound molecules around specialized regions of cell membrane in general.
This work "Visualization of subunit-specific delivery of glutamate receptors to postsynaptic membrane during hippocampal long-term potentiation" was published online in Cell Reports on March 22, 2012 (doi: 10.1016/j.celrep.2012.02.004). This work was supported by grants-in-aid for scientific research from Japan Society for the Promotion of Science, and from the Ministry of Education, Culture, Sports, Science and Technology in Japan, and also by Global COE program A06 of Kyoto University.
Provided by Kyoto University
