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Yasunori Hayashi, M.D., Ph.D.
Assistant Professor of Neurobiology
Department of Brain and Cognitive Sciences
Building: 46-4243
Email: yhayashi@mit.edu
Assistant Professor of Neurobiology
Department of Brain and Cognitive Sciences
Building: 46-4243
Email: yhayashi@mit.edu
Synaptic Plasticity and Glutamate Receptor
Topic 1: Molecular Mechanism of Synaptic Plasticity
In the mammalian hippocampus, a brief tetanic stimulation of input fibers
leads to a long-term potentiation (LTP) of excitatory synaptic transmission.
LTP has been widely studied as a cellular and synaptic model for learning
and memory. The LTP induction in hippocampal CA1 region requires post-synaptic
NMDA receptor activation and a resultant influx of Ca2+ ions. One consequence
of this rise in postsynaptic Ca2+ concentration is to trigger an increase
of the transmission mediated by AMPA receptors via an activation of
various Ca2+-dependent protein kinases. However, it has not been clear
how this increase in transmission is attained. We suggested that an
activity-induced translocation of the AMPA receptor from the extrasynaptic
location to the postsynaptic site explains an increase in AMPA receptor
transmission after LTP induction. We demonstrated this by using GFP
and electrophysiologically tagged AMPA receptor molecules. On the other
hand, recent application of the yeast two-hybrid screening method disclosed
a complex network of protein-protein interaction underlying excitatory
synapse. This structural meshwork is not rigid but is dynamically regulated
by various stimuli. It is therefore highly likely that the dynamics
of AMPA receptors which themselves are embedded in this meshwork are
regulated as a part of this dynamism of the meshwork. The principal
aim of our research project is to understand the protein-protein interactions
regulating AMPA receptor dynamics during LTP. For this purpose, we employ
multidisciplinary approaches including biochemical isolation of glutamate
receptor binding protein, construction of various mutants of glutamate
receptor itself and binding proteins, gene introduction into neurons,
and electrophysiological recording of those neurons.
Topic 2: Molecular mechanism spine formation
Spines are major site of excitatory synaptic transmission in hippocampal
pyramidal cells. Their peculiar shape was first described as early as
19th century but the knowledge how it is formed and how its number and
shapes are regulated are still limited. This is mainly due to a technical
difficulty in accessing this structure. However, recent advance in imaging
technique, mainly two-photon microscope, enabled direct visualization
of spines in living neurons. The number and shape of the spines is regulated
during development as well as by synaptic activity. Hippocampal CA1
pyramidal cells are virtually aspiny during the first two postnatal
weeks except for filopodial structures, which are much longer and thinner
than typical spine structure. After this period, typical spines with
head and neck structure start emerging. In addition, tetanic stimulation
induces new spines. In order to understand the mechanism involved in
spine formation, we express various proteins found in spines and to
see if they can form spine by themselves or change the morphology of
spines. The spines are visualized with green fluorescent protein (GFP)
under two-photon microscope. When GFP is expressed in neuron, it fills
cytosol and thus depict the tiny protrusions of the cells including
spines as if Golgi staining. Electrophysiological recording and Ca2+
imaging will be combined with GFP imaging to assess functional aspect
of spines.
Topic 3: Structural approach to understand postsynapse
To understand the biophysical properties of synaptic protein from different
aspect, we started a project to reveal 3D structures of those proteins.
We will express various postsynaptic proteins in bacteria in large quantity,
purify them, and make crystals. The resultant protein crystal will be
analyzed with X-ray defraction. This approach is auxiliary the functional
approach described above. For example, once functional unit of a protein
is identified the physiological experiment, the relevant fragment will
be used for crystallization trials. On the other hand, once we solved
the structure and find a motif which seems to have functional significance,
we can make mutants for further physiological analyses.
Topic 4: Role of adult neurogenesis in learning and memory
One interesting feature of hippocampus is a continual regeneration of
neurons that takes place in adulthood. Placing an animal in a rich environment
(for example, provided with a place to hide etc.) increases neurogenesis.
The neurogenesis also takes place in olfactory bulb. However, there
are no direct proof that the newly generated neurons are actually involved
in formation of neuronal circuit and hence in a process of learning
and memory. In order to test this, we are currently making a transgenic
animal model where neuronal stem cell can be removed by infusion of
cytotoxic substance. The resulting animal will be tested for memory
paradigms such as Morris water maze task or pregnancy block.
Topic 5: Further application of our knowledge to clinical medicine
Amyotrophic lateral sclerosis (ALS) and motor neuron disease are neurological
diseases that affect over 350,000 of the world’s population, and
kill over 100,000 every year. In the patients, spinal motor neurons
degenerate resulting in a loss of control of muscles and, eventually,
in a tragic death due to inability of respiration. Of the total ALS
cases, 90% is sporadic and 10% has family history. Some populations
of familial ALS cases have mutation in cytosolic Cu/Zn superoxide dismutase,
an enzyme that converts free radical oO2 to H2O2 and O2. However, it
accounts for only 15-20% of the familial cases and the cause the rest
of familial and also most of sporadic cases is not known. We are interested
in looking for gene involved in such familial ALS cases which cannot
be attributed to SOD mutation. The discovery of a gene responsible for
ALS will help to understand the pathogenesis of ALS and motor neuron
diseases, or even neurodegenerative disease in general.
Recent original papers
Nishi, M., Hinds, H., Lu, H. P., Kawata, M., and Hayashi, Y. (2001). Motoneuron-specific expression of NR3B, a novel NMDA-type glutamate receptor subunit that works in a dominant-negative manner, J Neurosci 21 , RC185.
Sala, C., Futai, K., Yamamoto, K., Worley, P. F., Hayashi, Y. , Sheng, M. (2003) Inhibition of dendritic spine morphogenesis and synaptic transmission by activity-inducible protein Homer1a. J Neurosci 23 , 6327-6337.
Okamoto, K., Nagai, T., Miyawaki, A. & Hayashi, Y. (2004) Rapid and persistent modulation of actin dynamics regulates postsynaptic reorganization underlying bidirectional plasticity. Nat Neurosci 7:1104-1112.
Nakagawa, T., Futai, K., Lashuel, H.A., Lo, I., Okamoto, K., Walz, T., Hayashi, Y. , Sheng, M. Quaternary structure, protein dynamics and synaptic function of SAP97 controlled by L27 domain interactions. Neuron 44:453-467.
Li, Z., Okamoto, K., Hayashi, Y ., and Sheng, M. (2004). The importance of dendritic mitochondria in the morphogenesis and plasticity of spines and synapses. Cell 119, 873-887.
Takao, K., Okamoto, K., Nakagawa, T., Neve, R. L., Nagai, T., Miyawaki, A., Hashikawa, T., Kobayashi, S., and Hayashi, Y. (2005). Visualization of synaptic Ca 2+ /calmodulin-dependent protein kinase II activity in living neurons. J Neurosci 25, 3107-3112.
Fukaya, M., Hayashi, Y. , and Watanabe, M. (2005). NR2 to NR3B subunit switchover of NMDA receptors in early postnatal motoneurons. Eur J Neurosci 21, 1432-1436.
Howarth, M., Takao, K., Hayashi, Y., and Ting, A. Y. (2005) Targeting quantum dots to surface proteins in living cells by using biotin ligase. Proc Natl Acad Sci 102, 7583-7588
Hayashi, MK, Ames , H., and Hayashi, Y. (2006) Tetrameric hub structure of postsynaptic scaffolding protein Homer. J Neurosci 23, 8492-8501
Okamoto, K. and Hayashi, Y. (2006) Visualization of F-actin and G-actin equilibrium using fluorescence resonance energy transfer (FRET) in cultured cells and neurons in slices. Nature Protocols 1, 912-919 .
Futai, K., Kim, KJ., Hashikawa, T., Sheng, M., Scheiffele, M., Hayashi, Y. (2007) Retrograde modulation of presynaptic release probability through PSD-95-neuroligin mediated signaling. Nat. Neurosci. 10, 186-195.
Okamoto, K.-I., Narayanan, R., Lee, S.-H., Murata, K., Hayashi, Y. The role of CaMKII as an F-actin bundling protein crucial for maintenance of dendritic spine structure. Proceedings of the National Academy of Science USA 104: 6418-6423 (2007).
Review
Hayashi, Y. Majewska, A. (2005) Dendritic spine geometry: Functional implication and regulation. Neuron 44, 529-532
