Download Session 2. Synaptic Plasticity (Chair, H. Kamiguchi)

The 2nd Japan-Korea Neuroscience Symposium
(A Satellite Symposium to Neuro2007)
“Cutting Edge of Neuroscience”
Date: September 13, 2007
Venue: Room 503, Pacifico Yokohama, Japan
Organizer: Japan Neuroscience Society
Korean Society of Brain and Neural Science
RIKEN Brain Science Institute
Organizing Committee: Tadaharu Tsumoto (RIKEN Brain Science Institute)
Masao Ito (RIKEN Brain Science Institute)
Yoo-Hun Suh (Seoul National University)
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Program
Session 1. Growth and Differentiation of Neurons (Chair, K. Yamaguchi)
9:30-10:00 Hiroyuki Kamiguchi (RIKEN/Brain Science Institute, Japan)
“The Role of Ca2+ signals in axon guidance”
10:00-10:30 Kyong Tai Kim (Pohang University of Science & Technology, Korea)
“Regulation of nerve differentiation by VRK3”
10:30-11:00 Kozo Kaibuchi (Nagoya University Graduate School of Medicine, Japan)
“Neuronal polarity and axonal vesicle transport”
11:00-11:15
Coffee Break
Session 2. Synaptic Plasticity (Chair, H. Kamiguchi)
11:15-11:45 Kazuhiko Yamaguchi (RIKEN/Brain Science Institute, Japan)
“Regulatory roles of actin dynamics in constitutive and activity-dependent trafficking of
AMPA-receptors in cerebellar Purkinje cell”
11:45-12:15 Bong-Kiun Kaang (Seoul National University, Korea)
“Synaptic plasticity and transcription factors: Role of a retrograde transcription factor
CAMAP in synaptic plasticity”
12:15-12:45 Toshiya Manabe (Tokyo University, Institute of Medical Sciences, Japan)
“The role of tyrosine phosphorylation of the NMDA receptor in synaptic plasticity and
higher brain functions”
12:45-14:00
Lunch
Session 3. Neuroscience of Amyotrophic Lateral Sclerosis (Chair, T. Saido)
14:00-14:30 Koji Yamanaka (RIKEN/Brain Science Institute, Japan)
"Onset and progression in inherited ALS determined by motor neurons and their
neighboring glial cells"
14:30-15:00 Kwang-Woo Lee (Seoul National University, Korea)
“Pathomechanism of ALS and neuroprotective effects of the novel dihydroxy bile acid
ursodeoxycholic acid derivate (Yoo's solution) in ALS”
15:00-15:30 Shin Kwak (Tokyo University Graduate School of Medicine, Japan)
“RNA editing and motor neuron diseases”
15:30-15:45
Coffee Break
Session 4. Neuroscience of Alzheimer’s Disease (Chair, K. Yamanaka)
15:45-16:15 Takaomi Saido (RIKEN/Brain Science Institute, Japan)
“Metabolism of amyloid-β peptide and Alzheimer's disease”
16:15-16:45 Yoo-Hun Suh (Seoul National University, Korea)
“Molecular pathogenesis and peripheral markers of α-synucleinopathy”
16:45-17:15 Takeshi Iwatsubo (University of Tokyo, Japan)
"Molecular pathology of Alzheimer's disease: β-amyloid and γ-secretase"
18:00-20:00
Reception
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Session 1. Growth and Differentiation of Neurons
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The role of Ca2+ signals in axon guidance
Hiroyuki Kamiguchi
Laboratory for Neuronal Growth Mechanisms
RIKEN Brain Science Institute, Japan
Axonal growth cones migrate along the correct paths during development, not only directed by
diffusible guidance cues but also contacted by local environment via cell adhesion molecules
(CAMs). Extracellular gradients of many guidance cues attract or repel the growth cone via
an asymmetric elevation of cytosolic free Ca2+ concentration ([Ca2+]c). Interestingly, a [Ca2+]c
gradient across the growth cone can trigger turning to the side with higher [Ca2+]c (attraction)
as well as to the side with lower [Ca2+]c (repulsion). Then, what determines the growth cone
responses to the [Ca2+]c gradient?
In this symposium, I will present evidence that the turning direction of growth cones
depends on the occurrence of Ca2+-induced Ca2+ release (CICR) through the ryanodine receptor
type 3 (RyR3): Ca2+ signals that are accompanied by RyR3-mediated CICR trigger growth cone
attraction whereas Ca2+ signals without CICR induce growth cone repulsion. The activity of
RyR3 is regulated by local environment contacting the growth cone: CAMs such as L1 and
N-cadherin facilitate RyR3-mediated CICR by activating cAMP-dependent protein kinase
(PKA), whereas extracellular matrix molecules such as laminin inactivate RyR3 by
down-regulating PKA. In this way, axon-guiding and CAM-derived signals are integrated at
the level of RyR3, which serves as a key regulator of growth cone guidance. Next I will
examine the molecular machinery downstream of Ca2+ signals and present evidence that
asymmetric membrane trafficking in the growth cone mediates attractive axon guidance. A
localized elevation of [Ca2+]c together with CICR (attractive Ca2+ signals) on one side of the
growth cone facilitates microtubule-dependent centrifugal transport of vesicles towards the
leading front and subsequent vesicle-associated membrane-protein 2 (VAMP2)-mediated
exocytosis on the side with elevated Ca2+. In contrast, Ca2+ signals without CICR (repulsive
Ca2+ signals) have no effect on the vesicle transport. Furthermore, pharmacological inhibition
of VAMP2-mediated exocytosis prevents growth cone attraction but not repulsion. Our study
provides a simple mechanisms for attractive axon guidance: the growth cone turns by
preferentially supplying membrane components and associated molecules to the side facing the
new direction. Our results also suggest that growth cone attraction and repulsion are driven
by distinct mechanisms, rather than using the same molecular machinery with opposing
polarities.
Ooashi N, Futatsugi A, Yoshihara F, Mikoshiba K, Kamiguchi H: Cell adhesion molecules
regulate Ca2+-mediated steering of growth cones via cyclic AMP and ryanodine receptor type 3.
J Cell Biol 170: 1159-1167, 2005
Tojima T, Akiyama H, Itofusa R, Li Y, Katayama H, Miyawaki A, Kamiguchi H: Attractive
axon guidance involves asymmetric membrane transport and exocytosis in the growth cone.
Nat Neurosci 10: 58-66, 2007
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Regulation of Nerve Differentiation by VRK3
Kyong-Tai Kim
Department of Life Science, Pohang University of Science and Technology, Pohang, Korea
Extracellular signal regulated kinases (ERKs) represent a signalling hub in many physiological
responses and have pivotal functions in cell proliferation, differentiation, development and
death, as well as in synaptic plasticity. Mitogen-activated protein kinase phosphatases (MKPs)
selectively
inactivate
ERKs
by
dephosphorylating
critical
phosphothreonine
and
phosphotyrosine residues. Transcriptional induction of MKP expression and posttranscriptional
stabilization of MKP mRNA are well-documented as negative-feedback mechanisms for ERK
signalling. Vaccinia-related kinase 3 (VRK3) is a member of the novel VRK family, but its
function has not been defined. Here, we show that VRK3 suppresses ERK activity through
direct binding to one of the MKPs, vaccinia H1-related (VHR), which specifically
dephosphorylates and inactivates ERK in the nucleus. Notably, VRK3 enhances the
phosphatase activity of VHR by a mechanism independent of its kinase activity. VRK3 is
therefore a member of a new class of phosphatase-activating kinases that regulate the activity
of ERK. Our findings show that direct interaction of VHR with VRK3 posttranslationally
regulates ERK signalling. ERK regulates nerve growth factor (NGF)-induced neurite outgrowth
in PC12 cells. Transient expression of VRK3 strongly suppressed both ERK activation and
neurite outgrowth induced by NGF in PC12 cells. Phorbol 12-myristate 13-acetate (PMA), a
well-known activator of protein kinase C (PKC), has been shown to induce accumulation of
ERK in the nucleus and neurite outgrowth in neuroblastoma HT22 cells. Overexpression of
VRK3 also inhibited PMA-induced nurite outgrowth in HT22 cells. Consistently, knockdown
of VRK3 promoted neurite outgrowth in both NGF-treated PC12 cells and PMA-treated HT22
cells. Our study provides evidence for a negative-feedback mechanism of MAPK signalling,
where VRK3 modulates MKP activity. This emphasizes the multiplicity and importance of
precise control of MAPK signaling in neuronal differentiation.
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Neuronal polarity and axonal vesicle transport
Kozo Kaibuchi
Nagoya University Graduate School of Medicine, Japan
Neurons are highly polarized cells that have axons and dendrites, both of which are
differentiated from common immature neurites in cultured hippocampal neurons.
One of the
key events for neuronal polarization is the directional trafficking of signaling molecules
including a neurotrophin receptor (Trk), which plays a critical role in axon specification.
Here we report that collapsin response mediator protein-2 (CRMP-2) links the vesicles
containing TrkB to Kinesin-1 through synaptotagmin-like protein 1 (Slp1) and Rab27, and
regulates the anterograde transport of TrkB in growing axons. The velocity of TrkB transport
is much higher in growing axons than in the immature neurites. Glycogen synthase kinase-3ß
(GSK-3ß) phosphorylates CRMP-2, prevents the association of CRMP-2 with Kinesin-1 and
Slp1, and impairs TrkB transport and neuronal polarization. Because the activity GSK-3ß
lower in the distal part of growing axons than in other immature neurites, we propose that local
inactivation of GSK-3ß in growing axons promotes the complex formation of
Kinesin-1/CRMP-2/Slp1/Rab27 and anterograde TrkB transport for neuronal polarization.
Reference
Arimura, N and Kaibuchi, K. Nat Rev Neurosci, 8, 194-205, 2007
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Session 2. Synaptic Plasticity
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Regulatory roles of actin dynamics in constitutive and activity-dependent
trafficking of AMPA-receptors in cerebellar Purkinje cell
Kazuhiko Yamaguchi
Lab. for Memory and Learning, RIKEN Brain Science Institute, Japan
In mammalian cerebral neurons, the balance between exocytosis and endocytosis maintains
postsynaptic expression of AMPA-type glutamate receptors (AMPA-Rs), and any
activity-dependent shift in this balance is the basis for postsynaptic type of plasticity. In the
cerebellar Purkinje cell, relationship between constitutive and activity-dependent trafficking of
AMPA-Rs is not clear. We analyzed this relationship in parallel fiber (PF) synapses of
cerebellar Purkinje cells by recording EPSC using a whole-cell patch-clamp
method. Constitutive endocytosis of AMPA-Rs was observed as a rapid decrease in synaptic
current when we blocked exocytosis by tetanus toxin (TeTx). Some populations of AMPA-Rs
were TeTx-resistant, and this TeTx-resistant pool of AMPA-Rs may be the stabilized
AMPA-Rs seen at the PF-synapse. We examined the role of actin in stabilizing AMPA-Rs at
the PF-synapse. Latrunculin A (Lat), a blocker of actin-polymerization, reduced the stable pool
of AMPA-Rs while jasplakinolide (Jas), a blocker of actin-depolymerization, enhanced this
pool of AMPA-Rs, indicating that actin dynamics regulated AMPA-Rs stabilization at
PF-synapse.
To examine whether constitutive endocytosis of AMPA-Rs and long-term depression
(LTD) of EPSC share the same mechanism, we induced LTD at the steady state of
TeTx-induced EPSC-reduction. LTD did not occlude constitutive endocytosis of AMPA-Rs.
Jas blocked LTD and Lat enhanced it, suggesting the involvement of actin-depolymerization in
the induction of LTD. Toxin B of Clostridium difficile, an inhibitor for Rho-family monomeric
GTPases (Rho, Rac and Cdc42), suppressed LTD induction, while exoenzyme C3, a
Rho-specific blocker, did not. Thus, Cdc42 and/or Rac most likely mediate LTD-induction.
Internalization of surface-expressed GluR2 was visualized using monoclonal antibody
recognizing GluR2 N-terminus. Stimulation with glutamate and KCl caused a substantial
increase in the internalization of surface-expressed GluR2, and Lat enhanced this
internalization. Taken together, these results suggest that actin-depolymerization is involved in
LTD-induction in cerebellar Purkinje cell.
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Synaptic plasticity and transcription factors: Role of a retrograde
transcription factor CAMAP in synaptic plasticity
Bong-Kiun Kaang, Ph.D.
Department of Biological Sciences, College of Natural Sciences,
Seoul National University, Seoul, Korea
The formation of long-term memory requires both new RNA and protein synthesis, whereas
short-term memory requires only covalent modifications of constitutively expressed preexisting
proteins. The core molecular features of the transcriptional regulation involved in long-term
memory is to be evolutionally conserved in Aplysia and Drosophila and in the mouse. A
growing body of evidence indicates that gene regulation by different combinations of
transcriptional factors may be involved in specific forms of long-term memory. In the marine
snail Aplysia, the molecular mechanisms of long-term memory have been extensively studied
in sensory neuron-to-motor neuron synapses of the gill-withdrawal reflex. Multiple pulses of
5-hydroxytryptamine (5-HT) produce long-term synaptic facilitation (LTF) and depends on
transcription and translation. These pulses of 5-HT upregulate the levels of cAMP within the
sensory cell via G-protein coupled receptors, and activate protein kinase A (PKA) and
mitogen-activated protein kinase (MAPK). Both kinases then translocate into the nucleus
where they can activate transcription factors such as ApCREB1, ApCREB2, and ApAF,
expressed in sensory neurons. Repeated pulses of serotonin (5-HT) induce long-term
facilitation (LTF) of the synapses between sensory and motor neurons of the gill-withdrawal
reflex in Aplysia. To explore how the apCAM down-regulation at the plasma membrane and
the CREB-mediated transcription in the nucleus, both of which are required for the formation
of LTF, might relate to each other, we have cloned an apCAM-associated protein (CAMAP) by
yeast two-hybrid screening. We found that 5-HT signaling at the synapse activates PKA which
in turn phosphorylates CAMAP to induce the dissociation of CAMAP from apCAM and the
subsequent translocation of CAMAP into the nucleus of sensory neurons. In the nucleus,
CAMAP acts as a transcriptional co-activator for CREB1 essential for the activation of the
immediate early gene ApC/EBP. Combined, our data suggest that CAMAP is one of the critical
retrograde signaling components that translocates from activated synapses to the nucleus during
synapse-specific LTF.
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The role of tyrosine phosphorylation of the NMDA receptor in synaptic
plasticity and higher brain functions
Toshiya Manabe
Division of Neuronal Network, Institute of Medical Science, University of Tokyo, Tokyo,
Japan
Long-lasting synaptic modification, such as long-term potentiation (LTP) of synaptic
transmission, has been thought to be involved in memory formation in the central nervous
system. The hippocampus plays a critical role in the memory of facts and episodes, while the
amygdala is associated with the memory of emotions such as fear. The expression of LTP is
usually mediated by long-lasting modification of intracellular biochemical processes in neurons.
Phosphorylation of neural proteins in response to a variety of external stimuli is one of the
main mechanisms underlying dynamic changes in the neural circuitry. The NR2B (GluRε2)
subunit of the N-methyl-D-aspartate (NMDA) receptor is tyrosine-phosphorylated in the brain,
with Tyr-1472 its major phosphorylation site. We have generated mutant mice with a knockin
mutation of the Tyr-1472 site to phenylalanine (Y1472F). These mutant mice (YF/YF mice)
show that Tyr-1472 phosphorylation is essential for the functions of the amygdala. The YF/YF
mice show impaired auditory fear conditioning and reduced LTP in the lateral nucleus of the
amygdala. In addition, the calcium/calmodulin-dependent protein kinase II (CaMKII) signaling
mediated by NMDA receptor activation is impaired in YF/YF mice. Furthermore,
electron-microscopic analyses reveal that the Y1472F mutant of the NR2B subunit shows
improper localization on postsynaptic spines at synapses. We have thus identified Tyr-1472
phosphorylation of the NR2B subunit as a key mediator of synaptic plasticity in the amygdala
and fear-related learning.
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Session 3. Neuroscience of Amyotrophic Lateral Sclerosis
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Onset and progression in inherited ALS determined by motor neurons and
their neighboring glial cells.
Koji Yamanaka
Yamanaka Research Unit, RIKEN Brain Science Institute, Japan
Dominant mutations in the ubiquitously expressed Cu/Zn superoxide dismutase (SOD1) lead to
amyotrophic lateral sclerosis (ALS), a neurodegenerative disease affecting adult motor neurons.
Although ubiquitous expression of mutant SOD1 provokes progressive, selective motor neuron
degeneration in human and rodents due to an acquired toxic property(ies) of the mutant, the cell
types that contribute to the onset and progression of the motor neuron disease are not known.
To test whether mutant SOD1 toxicity within specific cell types contributes to motor
neuron degeneration, we have generated a mouse ubiquitously expressing a “floxed” mutant
SOD1G37R transgene (LoxSOD1G37R) which can be removed within specific cell populations by
the action of Cre recombinase. To eliminate mutant SOD1 within motor neurons or
non-neuronal neighboring cells, these lox SOD1G37R mice were mated to Islet1-Cre,
CD11b-Cre, or GFAP-Cre that express Cre specifically in motor neurons, microglia, or
astrocytes, respectively. Removing mutant SOD1 from motor neurons slowed the timing of
disease onset and early disease progression, indicating mutant action in neurons as an initiating
factor in triggering disease. More importantly, silencing of SOD1 mutant expression selectively
within microglial cells or astrocytes has minimal effect on age of disease onset, but sharply
slows disease progression.
Thus, onset and progression represent distinct disease phases defined by mutant action
within different cell types to generate non-cell-autonomous killing of motor neurons, findings
that validate therapies, including cell replacement, targeted to the glial cells.
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Pathomechanism of ALS and neuroprotective effects of the novel dihydroxy
bile acid ursodeoxycholic acid derivate (Yoo's solution) in ALS
Kwang-Woo Lee, MD, PhD
Department of Neurology and Neuroscience Research Institute, College of Medicine, Seoul
National University, Seoul, Korea
Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease characterized
by selective death of motor neurons. Approximately 10% of ALS cases are familial, and about
20% of these familial cases (FALS) are associated with dominantly inherited mutations in the
gene encoding the enzyme Cu, Zn-superoxide dismutase (SOD1). Although the eitiology of
ALS pathology is not fully understood, the observed death of motoneurons is thought to occur
through oxidative stress-induced apoptosis.
The dihydroxy bile acid ursodeoxycholic acid (UDCA) is hydrophilic bile acid that
has been in widespread clinical use for the past 20 years for the treatment of chronic cholestatic
liver disease. Recent evidence have reported that UDCA is neuroprotective in
pharmachological and transgenic animal model of Huntington disease, but also in acute
ischemic stroke through reducing infarct size and improving neurological function. UDCA
plays a unique role in modulating the apoptotic threshold in both heapatic and nonhepatic cell
including neuronal cells through interaction with the mitochondrial membrane to prevent
mitochondrial membrane depolarization. UDCA also modulates the mitogen-activated protein
kinase and phosphoinositol-3-kinase survival pathways.
Yoo's solution is an aqueous clear solution that is composed of intact UDCA and
aqueous soluble starch which has low Dextrose Equivalency. UDCA in Yoo's solution has
about 30,000 times higher aqueous solubility than that of commercizlized UDCA (solubility
about; 3.5 mg/I) without any chemical modification of original UDCA molecule. UDCA of
Yoo's solution is solubilized in water with assistance of aqueous soluble strarch. Because of
coexistence with soluble starch, UDCA in Yoo's solution should be protected from
biotransformation to hydrophobic bile acid. Yoo's solution does not produce any precipitation
at any pH conditions, peculiary in acidic environment. Therefore, free UDCA or its protonated
form can freely penetrate various membranes including BBB.
Treatment with Yoo's solution increased significantly the proliferation of motor
neuronal cells expressing mutant SOD1 (A4V, G93A), while the effect was not significant in
cells expressing wild type SOD1 (WT). The Yoo's solution-induced proliferation of mutant
cells was attenuated by pretreatment with PD98050, the inhibitor for MAP kinase. In addition,
we investigated neuroprotective effect of Yoo's solution against oxidative stress. Exogenous
nitric oxide did reduce the viability of mutant cells, which was attenuated by Yoo's solution
treatment. We also investigated effects of Yoo's solution on G93A SOD1 transgenic mice,
which are the most widely used animal model of familial ALS. G93A transgenic mice were
treated with Yoo's solution daily from 70days of age (500mg/kg). It prolonged the average
lifespan, and improved motor perfomance of G93A mice.
Our data suggests that Yoo's solution may contribute to the reduction of degeneration
of motoneuron with the SOD1 mutation through activation of survival signal pathways or
action as an anti-apoptotic agent. Therefore, it may provide a potentially useful treatment in
FALS patients.
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RNA editing and motor neuron diseases
Shin Kwak
Department of Neurology, Graduate School of Medicine,
The University of Tokyo, Japan
Amyotrophic lateral sclerosis (ALS) is the most common motor neuron disease, leading
patients to inevitable death from respiratory failure within a few years. AMPA
receptor-mediated excitotoxity has been proposed to play a major role in the selective motor
neuron death in ALS. We recently demonstrated that a significant proportion of mRNA of the
AMPA receptor subunit GluR2 was unedited at the Q/R site in motor neurons of sporadic ALS,
the most common motor neuron disease. This molecular change occurs in a region-selective
and disease-specific manner and has been demonstrated to be a direct cause of neuronal death
in animal experiments, hence is highly relevant to ALS pathogenesis. We have found that the
efficiency of RNA editing at the GluR2 Q/R site was reduced in all the sporadic ALS cases
irrespective to the phenotypic variation but not in motor neurons of the rat modes of
SOD1-associated familial ALS or those of bulbar and spinal muscular atrophy (SBMA).
Therefore, underediting of GluR2 at the Q/R site may be the molecular change that leads
neurons to death in sporadic ALS, but not in familial ALS associated with mutated SOD1 or
spinal muscular atrophy. Because RNA editing at this site is specifically catalyzed by
adenosine deaminase acting on RNA type 2 (ADAR2), and because the expression level of
ADAR2 mRNA was reduced in the ventral gray of the spinal cord, it is likely that a reduction
of ADAR2 activity in motor neurons caused death of motor neurons in sporadic ALS. The role
of deficient GluR2 underediting in pathogenesis of sporadic ALS will be discussed.
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Session 4. Neuroscience of Alzheimer’s Disease
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Metabolism of amyloid  peptide and Alzheimer’s disease
Takaomi C. Saido
Laboratory for Proteolytic Neuroscience, RIKEN Brain Science Institute
Wako-shi, Saitama 351-0198, JAPAN
The conversion of normal brain aging to Alzheimer’s disease (AD) via a transition state
appears to be a continuous and chronic process primarily caused by aging-dependent
accumulation of amyloid ß peptide (Aß) in the brain. This notion gives us a hope that, by
manipulating the Aß levels in the brain, we may be able not only to prevent and cure the
disease but also to partially control some very significant aspects of brain aging. Aß is
constantly produced from its precursor and immediately catabolized under normal conditions,
whereas dysmetabolism of Aß seems to lead to pathological deposition upon aging. We have
focused our attention on elucidation of the unresolved mechanism of Aß catabolism in the
brain. We discovered neprilysin as a major Aß-degrading enzyme, the aging-dependent
reduction of which is likely to account for pathogenesis of sporadic AD. This peptidase stands
out as a unique Aß-degrading enzyme among other candidate mechanisms because only
neprilysin can degrade Aß at neuronal synapses. Moreover, deficiency of neprilysin resulted in
generation of metabolically stable and pathologically mature Aß3(pE)-42 via conversion of Aß1-42
to Aß3(E)-42. It should be noted that Aß starting with pyroglultamate (pE) at position 3, Aß3(pE)-42,
represents the predominant species deposited in Alzheimer’s disease (AD) brains, whereas
transgenic mice overexpressing amyloid precursor protein (APP) accumulate mostly full-length
Aß1-40 and Aß1-42. We also found that calpain is deeply involved in regulation AD pathology,
such as Aß amyloidosis, tau phosphorylation, microgliosis and somato-dendritic dystrophy.
Combining medications targeting at up-, mid- and down-stream processes of disease cascade
shall be the most effective strategy for prevention and treatment of AD.
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Molecular pathogenesis and peripheral markers of α-Synucleinopathy
Yoo-Hun Suh
Department of Pharmacology, College of Medicine, The National Creative Research Initiative
Center for Alzheimer’s Dementia and Neuroscience Research Institute, MRC, Seoul National
University, Seoul, Korea
α-Synuclein (α-SN) is a ubiquitous protein that is especially abundant in the brain and has been
postulated to play a central role in the pathogenesis of Parkinson’s disease (PD), Alzheimer’s
disease (AD), and other neurodegenerative disorders. However, little is known about the
neuronal functions of α-SN and the molecular and cellular mechanisms underlying neuronal
loss and immune abnormalities in PD. Here, we show that α-SN plays dual roles of
neuroprotection and neurotoxicity depending on its concentration or level of expression. At
nanomolar concentrations, α-SN protected neurons against serum deprivation, oxidative stress,
and excitotoxicity through the PI3/Akt signaling pathway. At both low micromolar and
overexpressed levels in the cell, α-SN resulted in cytotoxicity, through decreased Bcl-xL and
increased bax expression, followed by cytochrome c release and caspase activation and also by
microglia-mediated inflammatory responses. In addition, our study shows that -SN is
differentially expressed in human peripheral blood mononuclear cells. PD patients expressed
more -SN than healthy controls. Those expressions were shown to be correlated with
glucocorticoid sensitive apoptosis possibly caused by caspase activations (casapase-8,
caspase-9), CD95 up-regulation, and reactive oxygen radical (ROS) production.
And we hypothesize that α-SN may activate microglia to migrate into the SNpc
affecting neuronal cytotoxicity. We demonstrate that α-SN induces the CD44 expression on
microglia which participates in cell adhesion with surrounding extracellular matrix (ECM) and
it also enhances MT1-MMP (membrane-type 1 matrix metalloproteinase) to degrade ECM
opening the migratory pathway. Same as in the transfectants, extracellularly treated α-SN also
induces CD44 and MT1-MMP expressions. All those events are probably regulated through
ERK1/2 pathway. We confirmed α-SN induced cell migration in the brain of both A53T α-SN
transgenic mice and human PD patients. In this way, α-SN forms a functional link with
microglial migration in Parkinson’s disease.
We also demonstrated that nuclear translocation of α –SN that was mediated by the
karyopherin α6 and regulated by the SUMO and in nucleus, α-synuclein attend to DNA repair
by PCNA binding for neuronal cell survival.
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Molecular pathology of Alzheimer's disease: β-amyloid and γ-secretase
Takeshi Iwatsubo
University of Tokyo, Graduate Schools of Medicine and Pharmaceutical Sciences
Alzheimer’s disease (AD) is characterized by the deposition of amyloid ß peptides (Aß) as
senile plaques in brains. Aß is a 40-42 amino acid fragment of its precursor APP, and Aß42
species ending at position 42 forms amyloid fibrils much faster than Aß40. Mutations in APP
(Suzuki et al. 1994) as well as presenilin (PS) genes, that are major causative genes for familial
AD, enhance the production of Aß42, by shifting the preferred γ-secretase cleavage site from
position 40 to 42, resulting in an increase in Aß deposition in brains, supporting the pathogenic
significance of Aß42. PS has been demonstrated to comprise the catalytic center of γ-secretase,
that is responsible for the intramembrane proteolysis of APP, Notch and other type I membrane
proteins. These findings highlighted γ-secretase as one of the prime therapeutic targets for the
“disease-modifying therapy” of AD.
Protein chemical as well as genetic studies identified three “cofactor” proteins, i.e.,
nicastrin, APH-1 and PEN-2, that are essential to the formation and function of γ-secretase
complex. Reconstitution studies in cells suggested that the four proteins, i.e., PS, nicastrin,
APH-1 and PEN-2, are the minimal set of components that constitute the framework of
catalytically “active” γ-secretase complex. Furthermore, structural analysis of γ-secretase
complex by cysteine scanning unequivocally demonstrated the presence of a water-permeable
pore-like structure around the catalytic center of γ-secretase, providing structural basis for the
intramembrane proteolysis.
In view of the emerging new therapies for AD, e.g. secretase inhibitors and Aß
immunotherapy, an establishment of methods to monitor the progression of AD using imaging
and biofluid surrogate markers would be vital to the successful completion of clinical trials. In
this regard, a large-scale longitudinal clinical study, e.g. AD neuroimaging initiative (ADNI),
should be crucial to the development of effective disease-modifying therapies for AD.
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