- Email: [email protected]
Two types of α-synuclein accumulated abnormal terminals in the α-synuclein transgenic mice
Abstracts affected by this molecule. Resveratrol reduced lipid peroxide in serum, lipid droplets in hepatocytes, and activated phagocytosis of Kupffer cells. In the CNS, resveratrol reduced the appearance of lipofuscin granules, a marker for the aging, enhanced expressions of synaptophysin, superoxide dismutase, Bcl-2 and Bcl-xL connected with activation of neuronal function, and decreased neurofilament proteins, GFAP and Bax involved in aging/neurodegeneration. Together, resveratrol may function as beneficial to the CNS and protecting neurons from aging or degeneration. doi:10.1016/j.neures.2009.09.1385
P3-l10 Rhizoma (Senkyu), one of various components of Yokukansan reduced the ER stress induced cell toxicity resulting in the good effect on the Alzheimer disease associated cell death Toru Hiratsuka 1 , Shinsuke Matsuzaki 2 , Shingo Miyata 1 , Taiichi Katayama 2 , Masaya Tohyama 1,2 1
Dept. Anat&Neurosci, University of Osaka, Osaka, Japan; 2 United Grad. Sch. of Child Develop, Osaka Univ., Kanazawa University, Hamamatsu Univ. Sch. of Med., Osaka, Japan Yokukansan has been administrated to the persons who show such symptoms as nervousness etc. Yokukansan is focused as the one of the possible therapies for Alzheimer’s disease (AD). ER stress plays an important role in the pathogenesis of AD. Thus, we examined the effect of yokukansan on the ER stress induced neurotoxicity and the familial AD-linked PS1 mutation (E9) associated cell death to investigate the effects of yokukansan on the AD linked cell death. As results, we elucidated that yokukansan should upregulate the expression of GRP78 and reduce the expression of CHOP resulting in the reduction of ER stress induced cell death and the AD-linked associated cell death. These results suggest that yokukansan and Rhizoma should be potential medicine to the AD and our findings cast new light on the development of therapy for the AD. doi:10.1016/j.neures.2009.09.1386
P3-l11 Aged Alzheimer mice bear the increased expression of diazepam binding inhibitor in activated glial cells Kota Kimura, Mayu Shimatani, Jun Kaneko, Kaori Taniguchi, Ken Aizawa, Tatsuhiro Hisatsune Dept. Integrated., The University of Tokyo, Tokyo, Japan In Alzheimer disease, the change in glial functions has become a current attention in terms of both pathogenesis and therapeutic applications. In a related study, we also report the increased proliferation of astroglial cells at the dentate gyrus of hippocampal formation in the transgenic mice B6C3-Tg(APPswe,PSEN1dE9)85Dbo/J as a model for Alzheimer disease. In this study, we evaluated the expression of diazepam biding inhibitor (DBI) in the activated astroglial cells. A recent report showed that the over-expression of DBI caused the cognitive decline as assessed by hippocampal dependent behavior task and the deficit in LTP function of hippocampal circuit (Siiskonen et al., 2007). Very interestingly, we observed the increased expression of DBI in astroglial cells at the hilar region of Alzheimer transgenic mice, but not in those of control wild-type littermate, after 12 months of ages. Our results may suggest that the functional changes in hippocampal glial cells would be involved in the cognitive deficit in Alzheimer disease. doi:10.1016/j.neures.2009.09.1387
P3-l12 Identification of protein targets for 15-deoxy-12,14 prostaglandin J2 in neuronal plasma membranes Tatsurou Yagami, Kenkichi Takase, Yasuhiro Yamamoto Dept. Physiol., Facult. Pharmath. Sci., Himeji Dokkyo Univ., Himeji, Japan Deposits of amyloid  protein (A) are generally assumed to contribute to progressive neurodegeneration in the Alzheimer’s disease (AD). In primary cultures of cortical neurons, A significantly generated prostaglandin D2 (PGD2 ) before cell death. Although PGD2 induced neuronal cell death, specific binding sites of [3 H]PGD2 were not detected in plasma membranes. Without enzymes, PGD2 was metabolized to 15-deoxy-12,14 -prostaglandin J2 (15d-12,14 -PGJ2 ). 15d-12,14 -PGJ2 was more neurotoxic than PGD2 . The specific binding sites of [3 H]15d-12,14 -PGJ2 were abundant in plasma membranes. A proteomic approach was used to identify protein targets for 15d-12,14 -PGJ2 in neuronal plasma membranes. By using biotinylated 15d-12,14 -PGJ2 , 21 proteins were identified as biotin-positive targets. They were classified into cytoskeltal proteins, glycolytic enzymes and molecular chaperones. Some of these targets have been reported to interact with A and associate with AD. doi:10.1016/j.neures.2009.09.1388
S245
P3-l13 Molecular mechanisms to form insoluble complex in a mouse model of multiple system atrophy (MSA) Kimiko Nakayama, Yasuyo Suzuki, Ikuru Yazawa Laboratory of Research Resources, National Center for Geriatrics and Gerontology, Obu, Japan MSA is a non-hereditary neurodegenerative disease. Glial cytoplasmic and neuronal inclusions, both composed by ␣-synuclein (␣-syn), are characteristic markers in MSA pathology. Formation of ␣-syn inclusions might be the primary lesions that will eventually compromise neuronal function and viability in MSA. Using the transgenic mice overexpressing ␣-syn selectively in oligodendrocytes, we established primary cultured cells and showed that neuronal accumulation of insoluble mouse ␣-syn and binding ␣-syn to -III tubulin led to neuronal dysfunction. We coexpressed the Snca and Tubb3 genes in COS-7 cells and verified that mouse ␣-syn directly bound to -III tubulin to form insoluble complex. To clarify an initial process in the neuronal accumulation of insoluble complex, we examined whether self-aggregation of ␣-syn may induce the insoluble complex formation. We suggest an initial key process in MSA neurodegeneration. doi:10.1016/j.neures.2009.09.1389
P3-l14 Overexpression of a-synuclein gene caused dopaminergic neuron damage in substantia nigra of rats Lingling Lu, Hui Yang Dept. Neurosci., Capital Medical Univ., China The study was to explore whether overexpression of a-synuclein in rat brain caused selective dopaminergic neuron loss in substantia nigra and aimed to find out a new method to make a rat model of PD. The a-synuclein gene was induced into the rat brain by AAV. Rats behaviors were recorded every 4 weeks after the viral particle injection. TH immunohistochemistry as well as HPLC to detect the DA, DOPAC in striatum were performed. Q-PCR revealed a significant overexpression of a-synuclein in the injected hemisphere. By 8 weeks post injection, a significant loss of the dopaminergic neurons was observed and 34% or 60% were lost after 12 weeks or 16 weeks. The DA and DOPAC levels in the striatum decreased by15% at 12 weeks after injection and 30% decreased at 16 weeks. The rats injected with AAVa-synuclein developed a type of motor impairment, compatible with this magnitude of nigrostriatal damage. All the results showed that overexpression of a-synuclein caused selective dopaminergic neuron loss and mimic asymptom of human PD in rats. doi:10.1016/j.neures.2009.09.1390
P3-l15 Aggresomal property of glial cytoplasmic inclusions Shimada 1 , Shiro Takei 1 , Noriko Yoichi Chiba 1 , Atsuyoshi Kawamura 1 , Sanae Ishii 1 , Ayako Furukawa 1 , Kensuke Sasaki 2 , Toru Iwaki 2 , Masanori Hosokawa 1 1 2
Inst. for Developmental Res., Aichi Human Service Center, Kasugai, Japan; Dept. of Neuropathol., Univ. of Kyushu, Fukuoka, Japan
Aggresomes are proteinaceous inclusions formed in response to proteolytic stress. Several lines of evidence suggest that aggresome-related process contributes to the formation of Lewy bodies, the ␣-synuclein-positive neuronal inclusions. Here we examined whether glial cytoplasmic inclusions (GCIs), another ␣-synuclein-positive inclusions observed in the brains of multiple system atrophy (MSA) patients, had any characteristics of aggresomes. Formalin-fixed paraffin-embedded sections of the pons from 5 MSA patients were immunohistochemically stained using antibodies against ␥-tubulin, histone deacetylase 6 (HDAC6) and proteasome core subunits. These aggresome-related proteins were localized in GCIs, and were colocalized with ␣-synuclein. Immunoelectron microscopy revealed that HDAC6 and ␥-tubulin were present on the ␣-synuclein fibrils, of which GCIs were composed. These results suggest that the aggresome-related process may participate in the pathogenetic process forming GCIs. doi:10.1016/j.neures.2009.09.1391
P3-l16 Two types of ␣-synuclein accumulated abnormal terminals in the ␣-synuclein transgenic mice Akio Sekigawa, Masayo Fujita, Kazunari Sekiyama, Makoto Hashimoto Dept. Chemistry and Metabolism, Tokyo Met. Inst. Neurosci., Tokyo In Parkinson’s disease and other synucleinopathies, accumulation of ␣-synuclein (␣syn) may play a causative role for various neuropathology, including Lewy bodies formation, synaptic loss and neuronal cell death. In this study, to better understand
S246
Abstracts
the role of ␣-syn in the axonal pathology, Thy-1 promoter driven ␣-syn transgenic (Tg) mice were analyzed by histological procedures. Two types of ␣-syn accumulated abnormal axon structures, possibly synaptic swellings, were observed in the brains of 18 months old Tg mice but not in the control mice. One was with about 2 m diameter structure, which was immunopositive for vesicular glutamate transporter. The other had 4 m or more diameter and was immunostained with glutamic-acid decarboxylase. Interestingly, calbindin was positive only in the larger type of swelling. Further ultrastructural characterization of these two types of ␣-syn accumulated terminals is in progress. Our results suggest that at least two types of axonal pathology may be brought about by ␣-syn accumulation in the Tg mice brain. doi:10.1016/j.neures.2009.09.1392
P3-l17 Analysis of the degradation of ␣-synuclein by neurosin Harutsugu Tatebe 1 , Yoshihisa Watanabe 2 , Takashi Kasai 1 , Takahiko Tokuda 1 , Toshiki Mizuno 1 , Masaki Tanaka 2 , Masanori Nakagawa 1 1
Dept. Neurol., Kyoto Pref. Univ. Med., Japan; Geriatr., Kyoto Pref. Univ. Med., Japan
2
Dept. Cell Biol., Res. Inst.
␣-Synuclein (␣-syn) is known to be the major component of Lewy Body, which is commonly observed in dopaminergic neurons of patients in sporadic Parkinson’s disease (PD). Usually abnormal ␣-syn is degraded by certain catabolic pathways. In the present study, we studied the degradation of ␣-syn by neurosin (NS), a member of the kallikrein family of serine proteases, and revealed that NS cleaved within the NAC region of ␣-syn in vitro. We then investigated the localization and proteolytic activity of NS using HEK293T cells. The exogenously expressed NS was predominantly secreted from cells. A fraction of NS was localized to the ER and the Golgi apparatus. Proteolytic activities of intra- and extracellular NS were analyzed by zymography, resulting that extracellular NS had protease activity but intracellular NS did not. This result suggests that NS targets to the extracellular ␣-syn. doi:10.1016/j.neures.2009.09.1393
hyper-oxidized form of DJ-1, namely at C106, binds to TH but not stimulates the activity. doi:10.1016/j.neures.2009.09.1395
P3-l20 Experimental analysis of anti-parkinsonian effects of zonisamide Hideto Miwa, Tomomi Kubo, Kiwa Kobayashi, Yoshinori Kajimoto, Tomoyoshi Kondo Dept. Neurol., Wakayama Med. Univ., Wakayama, Japan Objectives: Zonisamide (ZNS), an anti-epileptic drug, has beneficial efficacy in Parkinson disease. Its pharmacological mechanisms remain unclear. The present study aims to study the effect of ZNS on experimental tremors. Methods: Effect of ZNS on harmaline-induced tremor and tacrine-induced tremulous jaw movements (TJMs) in rats, an experimental model of parkinosnian tremor, was studied. Results: ZNS significantly suppressed both harmaline-induced tremor and tacrineinduced TJMs, and this effect was not lost under conditions of monoamine-depletion. There was no effect of ZNS on harmaline-induced and tacrine-induced c-Fos expression in the primary site of the pharmacological action, such as the olivary nucleus and ventrolateral striatum, respectively. ZNS suppressed the tacrine-induced c-Fos expression in the cortex, the dorsal striatum, and the nucleus accumbens, which are involved in the architecture of the cortico-basal ganglia-thalamocortical circuits. Conclusion: ZNS has the anti-tremor effect that may be due to a broad inhibitory effect on tremor-related structures. doi:10.1016/j.neures.2009.09.1396
P3-m01 STN-DBS affects the REM switch Namiko Nishida 1 , Hiroki Toda 1 , Hidemoto Saiki 2 , Tokiko Murakami 3 , Kunihiro Kadoh 3 , Hideki Hayashi 1 , Keita Ueda 4 , Sadayuki Matsumoto 2 , Jun Takahashi 1 1
P3-l18 Prevention of MPTP-induced parkinsonism by calbindin recruitment into nigral dopaminergic cells Kenichi Inoue 1,2 , Shigehiro Miyachi 2,3 , Katsunori Nishi 1 , Haruo Okado 1 , Atsushi Nambu 4 , Masahiko Takada 1,2 1
Dept. Syst. Neurosci., Tokyo Met. Inst. Neurosci., Tokyo, Japan; 2 CREST, JST, Japan; 3 Dept. Behav. Brain Sci., Pri. Res. Inst., Kyoto Univ., Kyoto, Japan; 4 Div. Syst. Neurophysiol, NIPS, Okazaki, Japan A specific population of dopaminergic cells in the substantia nigra pars compacta that expresses calbindin selectively survives during the cell death period in Parkinson’s disease. Based on this finding, we examined the preventive effect of calbindin (Cb) recruitment into nigral dopaminergic cells, with an adenoviral vector and an lentiviral vector, on parkinsonian insults induced by 1-methyl-4-phenyl-1,2,3,6tetrahydropyridine (MPTP). Our behavioral analysis showed that a series of motor impairments occurred primarily in the limbs ipsilateral to Cb recruitment. Furthermore, our immunohistochemical analysis revealed that the expression level of tyrosine hydroxylase in the nigrostriatal system was preserved to some extent on the Cb-recruited side. The present results indicate that Cb recruitment into nigral dopaminergic cells with recombinant viral vectors may provide an effective therapeutic approach to Parkinson’s disease. doi:10.1016/j.neures.2009.09.1394
P3-l19 Oxidation of DJ-1-dependent regulation of tyrosine hydroxylase Shizuma Ishikawa 1 , Takahiro Taira 2 , Hiroyoshi Ariga 1 , Sanae M.M. Iguchi-Ariga 3 1
Grad. Pharm. Sci. Hokkaido Univ., Japan; Japan; 3 Grad. Agr. Hokkaido. Univ., Japan
2
Grad. Med. Yamanashi. Univ.,
DJ-1 is a multi-functional protein and plays roles in transcriptional regulation and anti-oxidative stress, loss of which is thought to be the onset of Parkinson’s disease. Although tyrosine hydroxylase (TH) is a key enzyme for dopamine biosynthesis, it is not clarified yet whether DJ-1 regulates TH. We therefore examined the effect of DJ-1 on TH activity. When SH-SY5Y cells were exposed to H2 O2 at various concentrations, TH activity increased to the maximal at 10 M H2 O2 , and then decreased to the level lower than that without H2 O2 -treatment. The complex formation between DJ-1 and TH, on the other hand, was increased with the increased amount of H2 O2 in a dose-dependent manner. These results suggest that the degree of DJ-1 oxidation, rather than the amount of the DJ-1–TH complex, affects TH activity: the
Dept. Neurosurg., Kitano Hospital, Osaka, Japan; 2 Dept. Neurol, Kitano Hospital, Japan; 3 Dept. Lab. Med., Kitano Hospital, Japan; 4 Dept. Psychiatry, Kitano Hospital, Japan Sleep disturbances are common in Parkinson’s disease(PD). Neuronal degeneration in PD progresses from caudal brainstem possibly including caudal REM-on and rostral REM-off centers. The aim of this study was to determine the effect of deep brain stimulation of the subthalamic nucleus (STN-DBS) on sleep symptoms. We assessed subjective symptoms with PD sleep scale(PDSS) on 17 patients, and objective symptoms on 7 patients with polysomnography (PSG) before and after surgery. Medications and motor symptoms were assessed by levodopa equivalent daily dose (LEDD) and by Unified PD Rating Scale (UPDRS). PDSS correlated with UPDRS motor part, and improved significantly. PSG showed improved sleep architecture with increase in the REM percentage, which was inversely correlated with PD duration. This study indicates that not only the alleviated motor symptoms but also the viable REM-on neurons could have contributed to the sleep modification. doi:10.1016/j.neures.2009.09.1397
P3-m02 Accumulation of C-terminal ATN1 fragment in DRPLA Yasuyo Suzuki 1 , Kimiko Nakayama 1 , Naohiro Hashimoto 2 , Ikuru Yazawa 1 1 Lab. of Research Resources, Natl. Center for Geriatrics and Gerontology, Obu, Japan; 2 Dep. of Regenerative Med., Natl. Center for Geriatrics and Gerontology, Obu, Japan
Dentatorubral-pallidoluysian atrophy (DRPLA) is a hereditary neurodegenerative disorder. The responsible gene product (DRPLA protein), also known as atrophin1 (ATN1), carry polyglutamine (polyQ) expansion. Nuclear accumulation of ATN1 might be a primary disease process in DRPLA, but the mechanism of neurodegeneration is still unclear. Using COS-7 and Neuro2a cells expressing the ATN1 gene, we demonstrated that ATN1 was accumulated with increasing number of polyQs, and identified a novel C-terminal fragment containing a polyQ tract. The mutant C-terminal fragment with expanded polyQ selectively accumulated in the cells, and it was showed in the brain tissues of patients with DRPLA. Full-length ATN1 and C-terminal fragment displayed individual localization, and the mutant fragment was preferentially found in the membrane fraction. We suggest that the mutant fragment plays a role in DRPLA neurodegeneration. doi:10.1016/j.neures.2009.09.1398
P3-l10 Rhizoma (Senkyu), one of various components of Yokukansan reduced the ER stress induced cell toxicity resulting in the good effect on the Alzheimer disease associated cell death Toru Hiratsuka 1 , Shinsuke Matsuzaki 2 , Shingo Miyata 1 , Taiichi Katayama 2 , Masaya Tohyama 1,2 1
Dept. Anat&Neurosci, University of Osaka, Osaka, Japan; 2 United Grad. Sch. of Child Develop, Osaka Univ., Kanazawa University, Hamamatsu Univ. Sch. of Med., Osaka, Japan Yokukansan has been administrated to the persons who show such symptoms as nervousness etc. Yokukansan is focused as the one of the possible therapies for Alzheimer’s disease (AD). ER stress plays an important role in the pathogenesis of AD. Thus, we examined the effect of yokukansan on the ER stress induced neurotoxicity and the familial AD-linked PS1 mutation (E9) associated cell death to investigate the effects of yokukansan on the AD linked cell death. As results, we elucidated that yokukansan should upregulate the expression of GRP78 and reduce the expression of CHOP resulting in the reduction of ER stress induced cell death and the AD-linked associated cell death. These results suggest that yokukansan and Rhizoma should be potential medicine to the AD and our findings cast new light on the development of therapy for the AD. doi:10.1016/j.neures.2009.09.1386
P3-l11 Aged Alzheimer mice bear the increased expression of diazepam binding inhibitor in activated glial cells Kota Kimura, Mayu Shimatani, Jun Kaneko, Kaori Taniguchi, Ken Aizawa, Tatsuhiro Hisatsune Dept. Integrated., The University of Tokyo, Tokyo, Japan In Alzheimer disease, the change in glial functions has become a current attention in terms of both pathogenesis and therapeutic applications. In a related study, we also report the increased proliferation of astroglial cells at the dentate gyrus of hippocampal formation in the transgenic mice B6C3-Tg(APPswe,PSEN1dE9)85Dbo/J as a model for Alzheimer disease. In this study, we evaluated the expression of diazepam biding inhibitor (DBI) in the activated astroglial cells. A recent report showed that the over-expression of DBI caused the cognitive decline as assessed by hippocampal dependent behavior task and the deficit in LTP function of hippocampal circuit (Siiskonen et al., 2007). Very interestingly, we observed the increased expression of DBI in astroglial cells at the hilar region of Alzheimer transgenic mice, but not in those of control wild-type littermate, after 12 months of ages. Our results may suggest that the functional changes in hippocampal glial cells would be involved in the cognitive deficit in Alzheimer disease. doi:10.1016/j.neures.2009.09.1387
P3-l12 Identification of protein targets for 15-deoxy-12,14 prostaglandin J2 in neuronal plasma membranes Tatsurou Yagami, Kenkichi Takase, Yasuhiro Yamamoto Dept. Physiol., Facult. Pharmath. Sci., Himeji Dokkyo Univ., Himeji, Japan Deposits of amyloid  protein (A) are generally assumed to contribute to progressive neurodegeneration in the Alzheimer’s disease (AD). In primary cultures of cortical neurons, A significantly generated prostaglandin D2 (PGD2 ) before cell death. Although PGD2 induced neuronal cell death, specific binding sites of [3 H]PGD2 were not detected in plasma membranes. Without enzymes, PGD2 was metabolized to 15-deoxy-12,14 -prostaglandin J2 (15d-12,14 -PGJ2 ). 15d-12,14 -PGJ2 was more neurotoxic than PGD2 . The specific binding sites of [3 H]15d-12,14 -PGJ2 were abundant in plasma membranes. A proteomic approach was used to identify protein targets for 15d-12,14 -PGJ2 in neuronal plasma membranes. By using biotinylated 15d-12,14 -PGJ2 , 21 proteins were identified as biotin-positive targets. They were classified into cytoskeltal proteins, glycolytic enzymes and molecular chaperones. Some of these targets have been reported to interact with A and associate with AD. doi:10.1016/j.neures.2009.09.1388
S245
P3-l13 Molecular mechanisms to form insoluble complex in a mouse model of multiple system atrophy (MSA) Kimiko Nakayama, Yasuyo Suzuki, Ikuru Yazawa Laboratory of Research Resources, National Center for Geriatrics and Gerontology, Obu, Japan MSA is a non-hereditary neurodegenerative disease. Glial cytoplasmic and neuronal inclusions, both composed by ␣-synuclein (␣-syn), are characteristic markers in MSA pathology. Formation of ␣-syn inclusions might be the primary lesions that will eventually compromise neuronal function and viability in MSA. Using the transgenic mice overexpressing ␣-syn selectively in oligodendrocytes, we established primary cultured cells and showed that neuronal accumulation of insoluble mouse ␣-syn and binding ␣-syn to -III tubulin led to neuronal dysfunction. We coexpressed the Snca and Tubb3 genes in COS-7 cells and verified that mouse ␣-syn directly bound to -III tubulin to form insoluble complex. To clarify an initial process in the neuronal accumulation of insoluble complex, we examined whether self-aggregation of ␣-syn may induce the insoluble complex formation. We suggest an initial key process in MSA neurodegeneration. doi:10.1016/j.neures.2009.09.1389
P3-l14 Overexpression of a-synuclein gene caused dopaminergic neuron damage in substantia nigra of rats Lingling Lu, Hui Yang Dept. Neurosci., Capital Medical Univ., China The study was to explore whether overexpression of a-synuclein in rat brain caused selective dopaminergic neuron loss in substantia nigra and aimed to find out a new method to make a rat model of PD. The a-synuclein gene was induced into the rat brain by AAV. Rats behaviors were recorded every 4 weeks after the viral particle injection. TH immunohistochemistry as well as HPLC to detect the DA, DOPAC in striatum were performed. Q-PCR revealed a significant overexpression of a-synuclein in the injected hemisphere. By 8 weeks post injection, a significant loss of the dopaminergic neurons was observed and 34% or 60% were lost after 12 weeks or 16 weeks. The DA and DOPAC levels in the striatum decreased by15% at 12 weeks after injection and 30% decreased at 16 weeks. The rats injected with AAVa-synuclein developed a type of motor impairment, compatible with this magnitude of nigrostriatal damage. All the results showed that overexpression of a-synuclein caused selective dopaminergic neuron loss and mimic asymptom of human PD in rats. doi:10.1016/j.neures.2009.09.1390
P3-l15 Aggresomal property of glial cytoplasmic inclusions Shimada 1 , Shiro Takei 1 , Noriko Yoichi Chiba 1 , Atsuyoshi Kawamura 1 , Sanae Ishii 1 , Ayako Furukawa 1 , Kensuke Sasaki 2 , Toru Iwaki 2 , Masanori Hosokawa 1 1 2
Inst. for Developmental Res., Aichi Human Service Center, Kasugai, Japan; Dept. of Neuropathol., Univ. of Kyushu, Fukuoka, Japan
Aggresomes are proteinaceous inclusions formed in response to proteolytic stress. Several lines of evidence suggest that aggresome-related process contributes to the formation of Lewy bodies, the ␣-synuclein-positive neuronal inclusions. Here we examined whether glial cytoplasmic inclusions (GCIs), another ␣-synuclein-positive inclusions observed in the brains of multiple system atrophy (MSA) patients, had any characteristics of aggresomes. Formalin-fixed paraffin-embedded sections of the pons from 5 MSA patients were immunohistochemically stained using antibodies against ␥-tubulin, histone deacetylase 6 (HDAC6) and proteasome core subunits. These aggresome-related proteins were localized in GCIs, and were colocalized with ␣-synuclein. Immunoelectron microscopy revealed that HDAC6 and ␥-tubulin were present on the ␣-synuclein fibrils, of which GCIs were composed. These results suggest that the aggresome-related process may participate in the pathogenetic process forming GCIs. doi:10.1016/j.neures.2009.09.1391
P3-l16 Two types of ␣-synuclein accumulated abnormal terminals in the ␣-synuclein transgenic mice Akio Sekigawa, Masayo Fujita, Kazunari Sekiyama, Makoto Hashimoto Dept. Chemistry and Metabolism, Tokyo Met. Inst. Neurosci., Tokyo In Parkinson’s disease and other synucleinopathies, accumulation of ␣-synuclein (␣syn) may play a causative role for various neuropathology, including Lewy bodies formation, synaptic loss and neuronal cell death. In this study, to better understand
S246
Abstracts
the role of ␣-syn in the axonal pathology, Thy-1 promoter driven ␣-syn transgenic (Tg) mice were analyzed by histological procedures. Two types of ␣-syn accumulated abnormal axon structures, possibly synaptic swellings, were observed in the brains of 18 months old Tg mice but not in the control mice. One was with about 2 m diameter structure, which was immunopositive for vesicular glutamate transporter. The other had 4 m or more diameter and was immunostained with glutamic-acid decarboxylase. Interestingly, calbindin was positive only in the larger type of swelling. Further ultrastructural characterization of these two types of ␣-syn accumulated terminals is in progress. Our results suggest that at least two types of axonal pathology may be brought about by ␣-syn accumulation in the Tg mice brain. doi:10.1016/j.neures.2009.09.1392
P3-l17 Analysis of the degradation of ␣-synuclein by neurosin Harutsugu Tatebe 1 , Yoshihisa Watanabe 2 , Takashi Kasai 1 , Takahiko Tokuda 1 , Toshiki Mizuno 1 , Masaki Tanaka 2 , Masanori Nakagawa 1 1
Dept. Neurol., Kyoto Pref. Univ. Med., Japan; Geriatr., Kyoto Pref. Univ. Med., Japan
2
Dept. Cell Biol., Res. Inst.
␣-Synuclein (␣-syn) is known to be the major component of Lewy Body, which is commonly observed in dopaminergic neurons of patients in sporadic Parkinson’s disease (PD). Usually abnormal ␣-syn is degraded by certain catabolic pathways. In the present study, we studied the degradation of ␣-syn by neurosin (NS), a member of the kallikrein family of serine proteases, and revealed that NS cleaved within the NAC region of ␣-syn in vitro. We then investigated the localization and proteolytic activity of NS using HEK293T cells. The exogenously expressed NS was predominantly secreted from cells. A fraction of NS was localized to the ER and the Golgi apparatus. Proteolytic activities of intra- and extracellular NS were analyzed by zymography, resulting that extracellular NS had protease activity but intracellular NS did not. This result suggests that NS targets to the extracellular ␣-syn. doi:10.1016/j.neures.2009.09.1393
hyper-oxidized form of DJ-1, namely at C106, binds to TH but not stimulates the activity. doi:10.1016/j.neures.2009.09.1395
P3-l20 Experimental analysis of anti-parkinsonian effects of zonisamide Hideto Miwa, Tomomi Kubo, Kiwa Kobayashi, Yoshinori Kajimoto, Tomoyoshi Kondo Dept. Neurol., Wakayama Med. Univ., Wakayama, Japan Objectives: Zonisamide (ZNS), an anti-epileptic drug, has beneficial efficacy in Parkinson disease. Its pharmacological mechanisms remain unclear. The present study aims to study the effect of ZNS on experimental tremors. Methods: Effect of ZNS on harmaline-induced tremor and tacrine-induced tremulous jaw movements (TJMs) in rats, an experimental model of parkinosnian tremor, was studied. Results: ZNS significantly suppressed both harmaline-induced tremor and tacrineinduced TJMs, and this effect was not lost under conditions of monoamine-depletion. There was no effect of ZNS on harmaline-induced and tacrine-induced c-Fos expression in the primary site of the pharmacological action, such as the olivary nucleus and ventrolateral striatum, respectively. ZNS suppressed the tacrine-induced c-Fos expression in the cortex, the dorsal striatum, and the nucleus accumbens, which are involved in the architecture of the cortico-basal ganglia-thalamocortical circuits. Conclusion: ZNS has the anti-tremor effect that may be due to a broad inhibitory effect on tremor-related structures. doi:10.1016/j.neures.2009.09.1396
P3-m01 STN-DBS affects the REM switch Namiko Nishida 1 , Hiroki Toda 1 , Hidemoto Saiki 2 , Tokiko Murakami 3 , Kunihiro Kadoh 3 , Hideki Hayashi 1 , Keita Ueda 4 , Sadayuki Matsumoto 2 , Jun Takahashi 1 1
P3-l18 Prevention of MPTP-induced parkinsonism by calbindin recruitment into nigral dopaminergic cells Kenichi Inoue 1,2 , Shigehiro Miyachi 2,3 , Katsunori Nishi 1 , Haruo Okado 1 , Atsushi Nambu 4 , Masahiko Takada 1,2 1
Dept. Syst. Neurosci., Tokyo Met. Inst. Neurosci., Tokyo, Japan; 2 CREST, JST, Japan; 3 Dept. Behav. Brain Sci., Pri. Res. Inst., Kyoto Univ., Kyoto, Japan; 4 Div. Syst. Neurophysiol, NIPS, Okazaki, Japan A specific population of dopaminergic cells in the substantia nigra pars compacta that expresses calbindin selectively survives during the cell death period in Parkinson’s disease. Based on this finding, we examined the preventive effect of calbindin (Cb) recruitment into nigral dopaminergic cells, with an adenoviral vector and an lentiviral vector, on parkinsonian insults induced by 1-methyl-4-phenyl-1,2,3,6tetrahydropyridine (MPTP). Our behavioral analysis showed that a series of motor impairments occurred primarily in the limbs ipsilateral to Cb recruitment. Furthermore, our immunohistochemical analysis revealed that the expression level of tyrosine hydroxylase in the nigrostriatal system was preserved to some extent on the Cb-recruited side. The present results indicate that Cb recruitment into nigral dopaminergic cells with recombinant viral vectors may provide an effective therapeutic approach to Parkinson’s disease. doi:10.1016/j.neures.2009.09.1394
P3-l19 Oxidation of DJ-1-dependent regulation of tyrosine hydroxylase Shizuma Ishikawa 1 , Takahiro Taira 2 , Hiroyoshi Ariga 1 , Sanae M.M. Iguchi-Ariga 3 1
Grad. Pharm. Sci. Hokkaido Univ., Japan; Japan; 3 Grad. Agr. Hokkaido. Univ., Japan
2
Grad. Med. Yamanashi. Univ.,
DJ-1 is a multi-functional protein and plays roles in transcriptional regulation and anti-oxidative stress, loss of which is thought to be the onset of Parkinson’s disease. Although tyrosine hydroxylase (TH) is a key enzyme for dopamine biosynthesis, it is not clarified yet whether DJ-1 regulates TH. We therefore examined the effect of DJ-1 on TH activity. When SH-SY5Y cells were exposed to H2 O2 at various concentrations, TH activity increased to the maximal at 10 M H2 O2 , and then decreased to the level lower than that without H2 O2 -treatment. The complex formation between DJ-1 and TH, on the other hand, was increased with the increased amount of H2 O2 in a dose-dependent manner. These results suggest that the degree of DJ-1 oxidation, rather than the amount of the DJ-1–TH complex, affects TH activity: the
Dept. Neurosurg., Kitano Hospital, Osaka, Japan; 2 Dept. Neurol, Kitano Hospital, Japan; 3 Dept. Lab. Med., Kitano Hospital, Japan; 4 Dept. Psychiatry, Kitano Hospital, Japan Sleep disturbances are common in Parkinson’s disease(PD). Neuronal degeneration in PD progresses from caudal brainstem possibly including caudal REM-on and rostral REM-off centers. The aim of this study was to determine the effect of deep brain stimulation of the subthalamic nucleus (STN-DBS) on sleep symptoms. We assessed subjective symptoms with PD sleep scale(PDSS) on 17 patients, and objective symptoms on 7 patients with polysomnography (PSG) before and after surgery. Medications and motor symptoms were assessed by levodopa equivalent daily dose (LEDD) and by Unified PD Rating Scale (UPDRS). PDSS correlated with UPDRS motor part, and improved significantly. PSG showed improved sleep architecture with increase in the REM percentage, which was inversely correlated with PD duration. This study indicates that not only the alleviated motor symptoms but also the viable REM-on neurons could have contributed to the sleep modification. doi:10.1016/j.neures.2009.09.1397
P3-m02 Accumulation of C-terminal ATN1 fragment in DRPLA Yasuyo Suzuki 1 , Kimiko Nakayama 1 , Naohiro Hashimoto 2 , Ikuru Yazawa 1 1 Lab. of Research Resources, Natl. Center for Geriatrics and Gerontology, Obu, Japan; 2 Dep. of Regenerative Med., Natl. Center for Geriatrics and Gerontology, Obu, Japan
Dentatorubral-pallidoluysian atrophy (DRPLA) is a hereditary neurodegenerative disorder. The responsible gene product (DRPLA protein), also known as atrophin1 (ATN1), carry polyglutamine (polyQ) expansion. Nuclear accumulation of ATN1 might be a primary disease process in DRPLA, but the mechanism of neurodegeneration is still unclear. Using COS-7 and Neuro2a cells expressing the ATN1 gene, we demonstrated that ATN1 was accumulated with increasing number of polyQs, and identified a novel C-terminal fragment containing a polyQ tract. The mutant C-terminal fragment with expanded polyQ selectively accumulated in the cells, and it was showed in the brain tissues of patients with DRPLA. Full-length ATN1 and C-terminal fragment displayed individual localization, and the mutant fragment was preferentially found in the membrane fraction. We suggest that the mutant fragment plays a role in DRPLA neurodegeneration. doi:10.1016/j.neures.2009.09.1398