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| title |
Coordinating morphological and
functional synaptic plasticity at central synapses |
| speaker |
Dr. Yukiko Goda |
| abstract |
We address how the structural organization of
synapses determines the efficacy of synaptic communication and
how changes in synaptic plasticity in turn, govern synapse morphology.
Actin is one of the major component of synaptic cytoskeleton
that regulates synapse integrity. The dynamic state of synapse
adhesion could be communicated to the actin cytoskeleton via
cell adhesion molecules. Similarly, changes in actin dynamics
could regulate the synaptic junction to influence synaptic transmission.
The involvement of actin in regulating neurotransmitter release
property is supported by studies in which actin dynamics is
perturbed by pharmacological agents and by modifying the activities
of ADF/cofilins and gelsolin. How synaptic activity regulates
synapse morphology has been examined by photoconductive stimulation
of neurons grown on silicon wafers. Stimuli that mimic the induction
of a durable form of long-term synaptic plasticity trigger the
formation of new presynaptic actin puncta that contain active
recycling synaptic vesicles. Synaptic actin dynamics, therefore,
is attuned to synaptic activity and can drive major morphological
synaptic changes. |
| reference |
Colicos, M. A., Collins, B. E., Sailor, M. J. and Goda, Y.
(2001). Remodeling of synaptic actin induced by photoconductive
stimulation. Cell 107, 605-616.
Morales, M., Colicos, M. A., Goda, Y. (2000). Actin-dependent
regulation of neurotransmitter release at central synapses.
Neuron 27, 539-550.
Optional reading: Goda, Y. and Davis G. W. (2003). Mechanisms
of synapse assembly and disassembly. Neuron 40, 243-264.
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| title |
Expression mechanism underlying
hippocampal short- and long-term presynaptic plasticity |
| speaker |
Dr. Haruyuki Kamiya |
| abstract |
Multiple forms of use-dependent synaptic plasticity
provide a cellular basis for learning and memory. Though postsynaptic
mechanisms have been studied extensively, little is known about
expression mechanisms for presynaptic plasticity in the brain.
The hippocampal mossy fiber (MF) synapse exhibits short- and
long-term presynaptic plasticity, i.e. paired-pulse facilitation
(PPF) and N-methyl-D-aspartate (NMDA) receptor-independent long-term
potentiation (LTP). Sustained presynaptic potentiation is also
induced by forskolin, an activator of adenylyl cyclase, suggesting
that cAMP signaling is crucial for persistent potentiating mechanism.
Using fluorescence recordings of presynaptic Ca2+ and voltage,
we demonstrated that unusually large PPF at MF synapses is accompanied
by short-term facilitation of presynaptic Ca2+ influx mediated
by activation of kainate autoreceptors. This finding deviates
from typical residual Ca2+ hypothesis of PPF assuming equal
amount of Ca2+ influx per action potential. In contrast, tetanus-induced
LTP at MF synapses is due to increased efficacy of transmitter
release without a change in presynaptic Ca2+ entry. Thus, kainate
receptor-dependent facilitation of presynaptic Ca2+ influx supports
robust short-term plasticity at the hippocampal MF synapses,
while the presynaptic LTP involves modification of molecular
targets downstream from Ca2+ influx. |
| reference |
Kamiya, H., Umeda, K., Ozawa, S., Manabe, T.
Presynaptic Ca2+ entry is unchanged during hippocampal mossy
fiber long-term potentiation. J. Neurosci. 22: 10524-8, 2002.
Kamiya, H., Ozawa, S., Manabe, T. Kainate receptor-dependent
short-term plasticity of presynaptic Ca2+ influx at the hippocampal
mossy fiber synapses. J. Neurosci. 22: 9237-43, 2002.
Optional reading
Schmitz, D., Mellor, J., Nicoll, R.A. Presynaptic kainate receptor
mediation of frequency facilitation at hippocampal mossy fiber
synapses. Science 291: 1972-6, 2001
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| title |
Single cell genetics: a novel approach
to study experience-dependent plasticity in vivo |
| speaker |
Dr. Pavel Osten |
| abstract |
The rodent somatosensory barrel cortex, with
its precise formation of whisker-matched Receptive Fields (RFs)
and large-scale functional plasticity induced by trimming whiskers,
is a well-established system for the study of experience-dependent
plasticity. In this presentation, I will first review some the
proposed molecular mechanisms that may underlie experience-dependent
plasticity. Then, I will introduce a novel method, based on
stereotaxic delivery of lentiviral infectious particles, which
allows us to genetically alter individual neurons in vivo by
means of single cell transgenic expression, siRNA-based gene
silencing and Cre recombinase-based gene knock-out. Importantly,
after the cellular phenotype of the genetic manipulations is
assessed in vitro in acute cortical slices, the RF properties
of the altered neurons are examined in vivo by 2-photon microscopy
targeted whole-cell recordings. In the end, I will present a
work-in-progress report on using the single cell genetics approach
to study the role of dendritic action potential backpropagation
in establishing the RF properties of layer2/3 barrel cortex
neurons. |
| reference |
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| title |
Visualization of spatio-temporal
regulation of Rho-family G proteins in neuronal morphogenesis |
| speaker |
Dr. Takeshi Nakamura |
| abstract |
Neuronal morphogenesis, including axon growth
and dendrite elaboration, is required for the basic function
of neurons to communicate with each other and with effector
cells. Rho GTPases (RhoA, Rac1, and Cdc42), which control actin
dynamics in a diversity of cellular functions, also play key
roles in neuronal morphogenesis during the development of the
neuronal network. It is widely accepted that Rac1/Cdc42 and
RhoA are positive and negative regulators of neurite outgrowth,
respectively. However, the spatio-temporal changes of the activities
of the Rho-family G proteins have not been elucidated in a living
neuron. To determine when and where Rho GTPases are activated
during neuritogenesis, we used in vivo probes based on fluorescence
resonance energy transfer (FRET). In my talk, I will first introduce
the principle of FRET and the FRET-based probes, Raichu-RhoA,
Rac1, and Cdc42. Next, I will describe the regulation of Rac1
and Cdc42 activity during NGF-induced neurite outgrowth in PC12
cells. In the last part, I will present the data showing the
relatively high activity of RhoA in the peripheral domain of
growth cones.
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| reference |
Aoki, K., Nakamura,T., Matsuda, M. (2004). Spatio-temporal
regulation of Rac1 and Cdc42 activity during nerve growth factor-induced
neurite outgrowth in PC12 cells. J. Biol. Chem. 279, 713-719.
Optional readings
Yoshizaki, H. et al. (2003). Activity of Rho-family GTPases
during cell division as visualized with FRET-based probes. J.
Cell. Biol. 162, 223-232.
Itoh, R. E. et al. (2002). Activation of Rac and Cdc42 video
imaged by fluorescent resonance energy transfer-based single-molecule
probes in the membrane of living cells. Mol. Cell. Biol. 22,
6582-6591. |
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| title |
Modeling synaptic Ca2+ dynamics
in hippocampal dendritic spines |
| speaker |
Dr. Pablo d'Alcantara |
| abstract |
Synaptic plasticity is triggered by transient
changes in Ca2+ concentration ([Ca2+]) within dendritic spines.
This in turn induces molecular alterations including changes
in the number and opening probability of various ion channels
in the spine membrane that result in potentiation or depression
of synaptic transmission. While the mechanisms induced by [Ca2+]
variation are being elucidated, how the Ca2+ dynamics is governed
remains mysterious. Ca2+ dynamic is influenced by sources and
sinks in the spines, such as Ca2+ channels and pumps, but also
by Ca2+ buffers and the spatiotemporal effects of diffusion.
Here we investigate Ca2+ dynamics in dendritic spines duringback-propagating
action potentials (BAPs). During depolarisation, voltage sensitive
Ca2+ channels (VSCCs) are activated leading to a [Ca2+] increase.
Using confocal laser scanning microscopy, we image BAP-induced
Ca2+ transients in dendritic spines of CA1 hippocampal neurons
in rat organotypic slices. Analysis of these transients under
different conditions-including dye saturation, optical fluctuation
analysis, and altered dye concentrations-enable us to estimate
the increase and decay in [Ca2+], the number and open probability
of VSCCs, and the endogenous Ca2+ buffering capacity. Our model
predicts that, according both to our data and to previously
published data, much bigger [Ca2+] elevations should be observed,
from which we infer the operation of unknown Ca2+ sink mechanisms.
We show that additional Ca2+ buffering could account for smaller
[Ca2+] transients. However, we also show that, if spatiotemporal
diffusion of Ca2+ is taken into account, very large [Ca2+] elevations
can occur in microdomains although average transients in the
spine are small. By combining experiments with physiology-based
models, we can investigate the minimum elements required to
explain published experimental observations, thereby clarifying
the contribution of various molecular mechanisms to synaptic
function and plasticity. |
| reference |
Maravall et al., Biophys J. 2000 May;78(5):2655-67.
Sabatini and Svoboda, Nature. 2000 Nov 30;408(6812):589-93.
Neher and Augustine, J Physiol. 1992 May;450:273-301.
d'Alcantara et al., Eur J Neurosci. 2003 Jun;17(12):2521-8.
Castellani et al., Proc Natl Acad Sci U S A. 2001 Oct 23;98(22):12772-7
Sharp et al., J Neurosci. 1993 Jul;13(7):3051-63.
Yang et al., J Neurophysiol. 1999 Feb;81(2):781-7.
Cormier et al., J Neurophysiol. 2001 Jan;85(1):399-406. |
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| title |
Modeling of long-term potentiation
and depression of synaptic efficacy using A-Cell |
| speaker |
Dr. Kazuhisa Ichikawa |
| abstract |
Long-term modification of synaptic efficacy is
thought to be a basis for learning and memory. Huge amount of
data ranging from biochemical and electrophysiological to behavioral
level have been accumulated. The main pathway in the molecular
mechanisms for the induction phase involves the activation of
kinases and phosphatases triggered by the increase in [Ca2+]i,
and the balance of activated kinase and phosphatase is thought
to determine the direction of the synaptic modification (potentiated
or depressed). In this lecture, I will present a biochemical
reaction model for the synaptic plasticity using A-Cell, a tool
for modeling and simulation of biochemical reaction and electrical
equivalent circuit in a neuron by GUI-based operation, and show
what is suggested from the model. The model is extended to the
three-dimensional morphology of a spine, and the localization
of activated kinse within a spine head will be discussed. A-Cell
will be introduced and demonstrated in the lecture. |
| reference |
Soderling, T.R. and Derkach, V.A., "Postsynaptic protein
phosphorylation and LTP", TINS Vol.23(2000), 75-80.
Gleason, M.R., Higashijima, S., Dallman, J., Liu, K., Mandel,
G., and Fetcho, J.R., "Translocation of CaM kinase II
to synaptic sites in vivo", Nat.Neurosci. Vol.6(2003),
217-218.
Ichikawa, K., "A-Cell: graphical user interface for
the construction of biochemical reaction models", Bioinformatics
Vol.17(2001), pp.483-484.
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| title |
Probing molecular kinetics with
single-molecule speckle microscopy: Actin turnover and mDia1,
the actin polymerization-driven motor. |
| speaker |
Dr. Naoki Watanabe |
| abstract |
Despite our knowledge of genes and molecules,
much remains unknown about how these molecules behave in cells.
Here I describe single-molecule speckle microscopy, a potent
tool for probing cellular processes at the molecular level.
Fluorescent speckle microscopy (FSM) provides fiduciary marks
on polymers with dilute fluorescent tags, and it is used for
following polymer movement. By further lowering the label concentration,
I found it is possible to detect speckles of single EGFP-actin
in living cells. As only immobilized EGFP-actin gives rise to
discrete "speckle" signals, the single-molecule speckle
microscopy allows actin polymerization mapping as well as actin
filament lifetime analysis. With this method, I observed that
more actin polymerized in the lamellipodium body than at the
leading edge of XTC fibroblasts.
I also present our recent results of single-molecule imaging
of mDia1. mDia1, a Rho effector, is a member of Formin homology
proteins. Recent biochemistry has revealed the binding of Formins
to the fast growing end of actin filaments in addition to their
actin nucleation activity. We captured images of mDia1 moving
along the growing end of actin filaments in living cells, which
suggests that Formins may function as actin polymerization-driven
molecular motors.
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| reference |
Pollard T.D., Blanchoin L. and Mullins R.D.,
Molecular mechanisms controlling actin filament dynamics in
nonmuscle cells. Annu. Rev. Biophys. Biomol. Struct. 29, 545
(2000).
Watanabe N. and Mitchison T. J., Single molecule speckle
analysis of actin filament turnover in lamellipodia. Science
295, 1083 (2002).
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| title |
Multiple Mechanisms by which Sensory Experience
Drives the Structural and Functional Development of the Visual
System |
| speaker |
Dr. Hollis Cline |
| abstract |
Sensory experience is required for the normal
development of brain circuits that process visual information,
however the mechanisms by which visual stimulation controls
circuit development are not yet clear. We approach this issue
by studying the visual system of Xenopus tadpoles using time-lapse
in vivo imaging, electrophysiology and molecular biology techniques.
We recently found that a brief (4 hour) presentation of visual
stimulus to the animal results in a significant increase in
the rate of dendritic arbor growth. The same visual stimulation
also leads to changes in glutamatergic synaptic responses and
neuronal excitability. The synaptic and intrinsic adaptations
function together to make the visual system less responsive
to background activity yet more sensitive to burst stimuli.
We are continuing our multidisciplinary studies of the developing
visual system in an effort to determine the multiple mechanisms
by which sensory inputs regulate visual system development.
|
| reference |
Cline H. T. Dendritic arbor development and synaptogenesis.
Curr Opin Neurobiol 11;118--126 (2001)
Aizenman C. D., Munoz-Elias, G. and Cline H. T. Visually
Driven Modulation of Glutamatergic Synaptic Transmission Is
Mediated by the Regulation of Intracellular Polymines. Neuron
Vol.34 1--20; (2002)
Ruthazer E. S., Akerman C. J. and Cline H. T. Control of
Axon Branch Dynamics by Correlated Activity in Vivo. Science
301 66--70 (2003)
Sin W. C., Haas K., Ruthazer E. S. and Cline H. T. Dendrite
growth increased by visual activity requires NMDA receptor
and Rho GTPases. NATURE 419 475--480 (2002)
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