74 results on '"Hirokazu Hirai"'
Search Results
2. Nurr1 overexpression in the primary motor cortex alleviates motor dysfunction induced by intracerebral hemorrhage in the striatum in mice
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Keita Kinoshita, Kensuke Motomura, Keisuke Ushida, Yuma Hirata, Ayumu Konno, Hirokazu Hirai, Shunsuke Kotani, Natsuko Hitora-Imamura, Yuki Kurauchi, Takahiro Seki, and Hiroshi Katsuki
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Hemorrhagic stroke ,Nurr1 ,Motor dysfunction ,Corticospinal tract injury ,Glial cell line-derived neurotrophic factor ,Neurology. Diseases of the nervous system ,RC346-429 - Abstract
Hemorrhage-induced injury of the corticospinal tract (CST) in the internal capsule (IC) causes severe neurological dysfunction in both human patients and rodent models of intracerebral hemorrhage (ICH). A nuclear receptor Nurr1 (NR4A2) is known to exert anti-inflammatory and neuroprotective effects in several neurological disorders. Previously we showed that Nurr1 ligands prevented CST injury and alleviated neurological deficits after ICH in mice. To prove direct effect of Nurr1 on CST integrity, we examined the effect of Nurr1 overexpression in neurons of the primary motor cortex on pathological consequences of ICH in mice. ICH was induced by intrastriatal injection of collagenase type VII, where hematoma invaded into IC. Neuron-specific overexpression of Nurr1 was induced by microinjection of synapsin I promoter-driven adeno-associated virus (AAV) vector into the primary motor cortex. Nurr1 overexpression significantly alleviated motor dysfunction but showed only modest effect on sensorimotor dysfunction after ICH. Nurr1 overexpression also preserved axonal structures in IC, while having no effect on hematoma-associated inflammatory events, oxidative stress, and neuronal death in the striatum after ICH. Immunostaining revealed that Nurr1 overexpression increased the expression of Ret tyrosine kinase and phosphorylation of Akt and ERK1/2 in neurons in the motor cortex. Moreover, administration of Nurr1 ligands 1,1-bis(3′-indolyl)-1-(p-chlorophenyl)methane or amodiaquine increased phosphorylation levels of Akt and ERK1/2 as well as expression of glial cell line-derived neurotrophic factor and Ret genes in the cerebral cortex. These results suggest that the therapeutic effect of Nurr1 on striatal ICH is attributable to the preservation of CST by acting on cortical neurons.
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- 2024
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3. Improving cell-specific recombination using AAV vectors in the murine CNS by capsid and expression cassette optimization
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Hayato Kawabata, Ayumu Konno, Yasunori Matsuzaki, Yumika Sato, Mika Kawachi, Ryo Aoki, Saki Tsutsumi, Shota Togai, Ryosuke Kobayashi, Takuro Horii, Izuho Hatada, and Hirokazu Hirai
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adeno-associated virus ,PHP.eB ,AAV-F ,glial fibrillary acidic protein ,astrocyte ,floxed mouse ,Genetics ,QH426-470 ,Cytology ,QH573-671 - Abstract
The production of cell-type– and age-specific genetically modified mice is a powerful approach for unraveling unknown gene functions. Here, we present a simple and timesaving method that enables adeno-associated virus (AAV)–mediated cell-type– and age-specific recombination in floxed mice. To achieve astrocyte-specific recombination in floxed Ai14 reporter mice, we intravenously injected blood-brain barrier–penetrating AAV-PHP.eB vectors expressing Cre recombinase (Cre) using the astrocyte-specific mouse glial fibrillary acidic protein (mGfaABC1D) promoter. However, we observed nonspecific neuron-predominant transduction despite the use of an astrocyte-specific promoter. We speculated that subtle but continuous Cre expression in nonastrocytic cells triggers recombination, and that excess production of Cre in astrocytes inhibits recombination by forming Cre-DNA aggregates. Here, we resolved this paradoxical event by dividing a single AAV into two mGfaABC1D-promoter-driven AAV vectors, one expressing codon-optimized flippase (FlpO) and another expressing flippase recognition target–flanked rapidly degrading Cre (dCre), together with switching the neuron-tropic PHP.eB capsid to astrocyte-tropic AAV-F. Moreover, we found that the FlpO-dCre system with a target cell-tropic capsid can also function in neuron-targeting recombination in floxed mice.
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- 2024
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4. Protein kinase Cγ negatively regulates the intrinsic excitability in zebrin-negative cerebellar Purkinje cells
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Masashi Watanave, Mika Kawachi, Ayumu Konno, Ryo Aoki, Yuuki Fukai, Yasunori Matsuzaki, Ryosuke Kaneko, and Hirokazu Hirai
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protein kinase ,cerebellum ,action potential ,Purkinje cells ,aldolase C ,zebrin ,Neurosciences. Biological psychiatry. Neuropsychiatry ,RC321-571 - Abstract
Protein kinase C γ (PKCγ), a neuronal isoform present exclusively in the central nervous system, is most abundantly expressed in cerebellar Purkinje cells (PCs). Targeted deletion of PKCγ causes a climbing fiber synapse elimination in developing PCs and motor deficit. However, physiological roles of PKCγ in adult mouse PCs are little understood. In this study, we aimed to unravel the roles of PKCγ in mature mouse PCs by deleting PKCγ from adult mouse PCs of PKCγfl/fl mice via cerebellar injection of adeno-associated virus (AAV) vectors expressing Cre recombinase under the control of the PC-specific L7-6 promoter. Whole cell patch-clamp recording of PCs showed higher intrinsic excitability in PCs virally lacking PKCγ [PKCγ-conditional knockout (PKCγ-cKO) PCs] than in wild-type (WT) mouse PCs in the zebrin-negative module, but not in the zebrin-positive module. AAV-mediated PKCγ re-expression in PKCγ-deficient mouse PCs in the zebrin-negative module restored the enhanced intrinsic excitability to a level comparable to that of wild-type mouse PCs. In parallel with higher intrinsic excitability, we found larger hyperpolarization-activated cyclic nucleotide-gated (HCN) channel currents in PKCγ-cKO PCs located in the zebrin-negative module, compared with those in WT mouse PCs in the same region. However, pharmacological inhibition of the HCN currents did not restore the enhanced intrinsic excitability in PKCγ-cKO PCs in the zebrin-negative module. These results suggested that PKCγ suppresses the intrinsic excitability in zebrin-negative PCs, which is likely independent of the HCN current inhibition.
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- 2024
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5. Production of Spinocerebellar Ataxia Type 3 Model Mice by Intravenous Injection of AAV-PHP.B Vectors
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Ayumu Konno, Yoichiro Shinohara, and Hirokazu Hirai
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adeno-associated virus ,AAV ,AAV-PHP.B ,Spinocerebellar ataxia type 3 ,SCA3 ,Machado–Joseph disease ,Biology (General) ,QH301-705.5 ,Chemistry ,QD1-999 - Abstract
We aimed to produce a mouse model of spinocerebellar ataxia type 3 (SCA3) using the mouse blood–brain barrier (BBB)-penetrating adeno-associated virus (AAV)-PHP.B. Four-to-five-week-old C57BL/6 mice received injections of high-dose (2.0 × 1011 vg/mouse) or low-dose (5.0 × 1010 vg/mouse) AAV-PHP.B encoding a SCA3 causative gene containing abnormally long 89 CAG repeats [ATXN3(Q89)] under the control of the ubiquitous chicken β-actin hybrid (CBh) promoter. Control mice received high doses of AAV-PHP.B encoding ATXN3 with non-pathogenic 15 CAG repeats [ATXN3(Q15)] or phosphate-buffered saline (PBS) alone. More than half of the mice injected with high doses of AAV-PHP.B encoding ATXN3(Q89) died within 4 weeks after the injection. No mice in other groups died during the 12-week observation period. Mice injected with low doses of AAV-PHP.B encoding ATXN3(Q89) exhibited progressive motor uncoordination starting 4 weeks and a shorter stride in footprint analysis performed at 12 weeks post-AAV injection. Immunohistochemistry showed thinning of the molecular layer and the formation of nuclear inclusions in Purkinje cells from mice injected with low doses of AAV-PHP.B encoding ATXN3(Q89). Moreover, ATXN3(Q89) expression significantly reduced the number of large projection neurons in the cerebellar nuclei to one third of that observed in mice expressing ATXN3(Q15). This AAV-based approach is superior to conventional methods in that the required number of model mice can be created simply by injecting AAV, and the expression levels of the responsible gene can be adjusted by changing the amount of AAV injected. Moreover, this method may be applied to produce SCA3 models in non-human primates.
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- 2024
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6. A blood-brain barrier-penetrating AAV2 mutant created by a brain microvasculature endothelial cell-targeted AAV2 variant
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Hayato Kawabata, Ayumu Konno, Yasunori Matsuzaki, and Hirokazu Hirai
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AAV ,PHP.B ,PHP.eB ,BR1 ,blood-brain barrier ,capsid ,Genetics ,QH426-470 ,Cytology ,QH573-671 - Abstract
Upon systemic administration, adeno-associated virus serotype 9 (AAV9) and the capsid variant PHP.eB show distinct tropism for the central nervous system (CNS), whereas AAV2 and the capsid variant BR1 transduce brain microvascular endothelial cells (BMVECs) with little transcytosis. Here, we show that a single amino acid substitution (from Q to N) in the BR1 capsid at position 587 (designated BR1N) confers a significantly higher blood-brain barrier (BBB) penetration capacity to BR1. Intravenously infused BR1N showed significantly higher CNS tropism than BR1 and AAV9. BR1 and BR1N likely use the same receptor for entry into BMVECs; however, the single amino acid substitution has profound consequences on tropism. This suggests that receptor binding alone does not determine the final outcome in vivo and that further improvements of capsids within predetermined receptor usage are feasible.
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- 2023
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7. Excitatory subtypes of the lateral amygdala neurons are differentially involved in regulation of synaptic plasticity and excitation/inhibition balance in aversive learning in mice
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Mieko Morishima, Sohta Matsumura, Suguru Tohyama, Takashi Nagashima, Ayumu Konno, Hirokazu Hirai, and Ayako M. Watabe
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lateral amygdala (LA) ,medial geniculate nucleus (MGN) ,excitatory cell ,inhibitory cell ,fear conditioning ,experience-dependent plasticity ,Neurosciences. Biological psychiatry. Neuropsychiatry ,RC321-571 - Abstract
The amygdala plays a crucial role in aversive learning. In Pavlovian fear conditioning, sensory information about an emotionally neutral conditioned stimulus (CS) and an innately aversive unconditioned stimulus is associated with the lateral amygdala (LA), and the CS acquires the ability to elicit conditioned responses. Aversive learning induces synaptic plasticity in LA excitatory neurons from CS pathways, such as the medial geniculate nucleus (MGN) of the thalamus. Although LA excitatory cells have traditionally been classified based on their firing patterns, the relationship between the subtypes and functional properties remains largely unknown. In this study, we classified excitatory cells into two subtypes based on whether the after-depolarized potential (ADP) amplitude is expressed in non-ADP cells and ADP cells. Their electrophysiological properties were significantly different. We examined subtype-specific synaptic plasticity in the MGN-LA pathway following aversive learning using optogenetics and found significant experience-dependent plasticity in feed-forward inhibitory responses in fear-conditioned mice compared with control mice. Following aversive learning, the inhibition/excitation (I/E) balance in ADP cells drastically changed, whereas that in non-ADP cells tended to change in the reverse direction. These results suggest that the two LA subtypes are differentially regulated in relation to synaptic plasticity and I/E balance during aversive learning.
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- 2023
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8. State-dependent modulation of positive and negative affective valences by a parabrachial nucleus-to-ventral tegmental area pathway in mice
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Takashi Nagashima, Kaori Mikami, Suguru Tohyama, Ayumu Konno, Hirokazu Hirai, and Ayako M. Watabe
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parabrachial nucleus ,ventral tegmental area ,glutamic acid decarboxylase 65 ,avoidance ,aversive memory ,operant task ,Neurosciences. Biological psychiatry. Neuropsychiatry ,RC321-571 - Abstract
Appropriately responding to various sensory signals in the environment is essential for animal survival. Accordingly, animal behaviors are closely related to external and internal states, which include the positive and negative emotional values of sensory signals triggered by environmental factors. While the lateral parabrachial nucleus (LPB) plays a key role in nociception and supports negative valences, it also transmits signals including positive valences. However, the downstream neuronal mechanisms of positive and negative valences have not been fully explored. In the present study, we investigated the ventral tegmental area (VTA) as a projection target for LPB neurons. Optogenetic activation of LPB-VTA terminals in male mice elicits positive reinforcement in an operant task and induces both avoidance and attraction in a place-conditioning task. Inhibition of glutamic acid decarboxylase (GAD) 65-expressing cells in the VTA promotes avoidance behavior induced by photoactivation of the LPB-VTA pathway. These findings indicate that the LPB-VTA pathway is one of the LPB outputs for the transmission of positive and negative valence signals, at least in part, with GABAergic modification in VTA.
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- 2023
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9. Development of microglia-targeting adeno-associated viral vectors as tools to study microglial behavior in vivo
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Yukihiro Okada, Nobutake Hosoi, Yasunori Matsuzaki, Yuuki Fukai, Akito Hiraga, Junichi Nakai, Keisuke Nitta, Yoichiro Shinohara, Ayumu Konno, and Hirokazu Hirai
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Biology (General) ,QH301-705.5 - Abstract
Microglia-targeting AAV vectors are created which can be utilized to study microglial pathophysiology and microglia-targeted therapies in mice, especially in the regions of striatum and cerebellar cortex.
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- 2022
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10. DOPAnization of tyrosine in α-synuclein by tyrosine hydroxylase leads to the formation of oligomers
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Mingyue Jin, Sakiko Matsumoto, Takashi Ayaki, Hodaka Yamakado, Tomoyuki Taguchi, Natsuko Togawa, Ayumu Konno, Hirokazu Hirai, Hiroshi Nakajima, Shoji Komai, Ryuichi Ishida, Syuhei Chiba, Ryosuke Takahashi, Toshifumi Takao, and Shinji Hirotsune
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Science - Abstract
In this work, the authors show that α-synuclein is posttranslationally dopanized at Tyr136 by tyrosine hydroxylase, which facilitates the formation of oligomers. This modification likely impacts pathogenesis and the selective degeneration of dopaminergic neurons in Parkinson’s disease.
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- 2022
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11. Interaction between Teneurin-2 and microtubules via EB proteins provides a platform for GABAA receptor exocytosis
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Sotaro Ichinose, Yoshihiro Susuki, Nobutake Hosoi, Ryosuke Kaneko, Mizuho Ebihara, Hirokazu Hirai, and Hirohide Iwasaki
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synapse ,microtubules ,adhesion molecule ,Medicine ,Science ,Biology (General) ,QH301-705.5 - Abstract
Neurons form dense neural circuits by connecting to each other via synapses and exchange information through synaptic receptors to sustain brain activities. Excitatory postsynapses form and mature on spines composed predominantly of actin, while inhibitory synapses are formed directly on the shafts of dendrites where both actin and microtubules (MTs) are present. Thus, it is the accumulation of specific proteins that characterizes inhibitory synapses. In this study, we explored the mechanisms that enable efficient protein accumulation at inhibitory postsynapse. We found that some inhibitory synapses function to recruit the plus end of MTs. One of the synaptic organizers, Teneurin-2 (TEN2), tends to localize to such MT-rich synapses and recruits MTs to inhibitory postsynapses via interaction with MT plus-end tracking proteins EBs. This recruitment mechanism provides a platform for the exocytosis of GABAA receptors. These regulatory mechanisms could lead to a better understanding of the pathogenesis of disorders such as schizophrenia and autism, which are caused by excitatory/inhibitory (E/I) imbalances during synaptogenesis.
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- 2023
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12. Astrocytic GPCR-Induced Ca2+ Signaling Is Not Causally Related to Local Cerebral Blood Flow Changes
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Katsuya Ozawa, Masaki Nagao, Ayumu Konno, Youichi Iwai, Marta Vittani, Peter Kusk, Tsuneko Mishima, Hirokazu Hirai, Maiken Nedergaard, and Hajime Hirase
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astrocytes ,optogenetic GPCR ,OptoA1AR ,Ca2+ elevation ,blood flow ,cortex ,Biology (General) ,QH301-705.5 ,Chemistry ,QD1-999 - Abstract
Activation of Gq-type G protein-coupled receptors (GPCRs) gives rise to large cytosolic Ca2+ elevations in astrocytes. Previous in vitro and in vivo studies have indicated that astrocytic Ca2+ elevations are closely associated with diameter changes in the nearby blood vessels, which astrocytes enwrap with their endfeet. However, the causal relationship between astrocytic Ca2+ elevations and blood vessel diameter changes has been questioned, as mice with diminished astrocytic Ca2+ signaling show normal sensory hyperemia. We addressed this controversy by imaging cortical vasculature while optogenetically elevating astrocyte Ca2+ in a novel transgenic mouse line, expressing Opto-Gq-type GPCR Optoα1AR (Astro-Optoα1AR) in astrocytes. Blue light illumination on the surface of the somatosensory cortex induced Ca2+ elevations in cortical astrocytes and their endfeet in mice under anesthesia. Blood vessel diameter did not change significantly with Optoα1AR-induced Ca2+ elevations in astrocytes, while it was increased by forelimb stimulation. Next, we labeled blood plasma with red fluorescence using AAV8-P3-Alb-mScarlet in Astro-Optoα1AR mice. We were able to identify arterioles that display diameter changes in superficial areas of the somatosensory cortex through the thinned skull. Photo-stimulation of astrocytes in the cortical area did not result in noticeable changes in the arteriole diameters compared with their background strain C57BL/6. Together, compelling evidence for astrocytic Gq pathway-induced vasodiameter changes was not observed. Our results support the notion that short-term (2+ signaling.
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- 2023
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13. A cortical cell ensemble in the posterior parietal cortex controls past experience-dependent memory updating
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Akinobu Suzuki, Sakurako Kosugi, Emi Murayama, Eri Sasakawa, Noriaki Ohkawa, Ayumu Konno, Hirokazu Hirai, and Kaoru Inokuchi
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Science - Abstract
Animals refer to related past experiences when processing sensory inputs. The authors show that a cellular ensemble in the posterior parietal cortex that is activated during past experience mediates an interaction between past and current information to update memory through a circuit including the anterior cingulate cortex.
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- 2022
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14. Dynamic modulation of pulsatile activities of oxytocin neurons in lactating wild-type mice.
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Kasane Yaguchi, Mitsue Hagihara, Ayumu Konno, Hirokazu Hirai, Hiroko Yukinaga, and Kazunari Miyamichi
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Medicine ,Science - Abstract
Breastfeeding, which is essential for the survival of mammalian infants, is critically mediated by pulsatile secretion of the pituitary hormone oxytocin from the central oxytocin neurons located in the paraventricular and supraoptic hypothalamic nuclei of mothers. Despite its importance, the molecular and neural circuit mechanisms of the milk ejection reflex remain poorly understood, in part because a mouse model to study lactation was only recently established. In our previous study, we successfully introduced fiber photometry-based chronic imaging of the pulsatile activities of oxytocin neurons during lactation. However, the necessity of Cre recombinase-based double knock-in mice substantially compromised the use of various Cre-dependent neuroscience toolkits. To overcome this obstacle, we developed a simple Cre-free method for monitoring oxytocin neurons by an adeno-associated virus vector driving GCaMP6s under a 2.6 kb mouse oxytocin mini-promoter. Using this method, we monitored calcium ion transients of oxytocin neurons in the paraventricular nucleus in wild-type C57BL/6N and ICR mothers without genetic crossing. By combining this method with video recordings of mothers and pups, we found that the pulsatile activities of oxytocin neurons require physical mother-pup contact for the milk ejection reflex. Notably, the frequencies of photometric signals were dynamically modulated by mother-pup reunions after isolation and during natural weaning stages. Collectively, the present study illuminates the temporal dynamics of pulsatile activities of oxytocin neurons in wild-type mice and provides a tool to characterize maternal oxytocin functions.
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- 2023
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15. Protective roles of MITOL against myocardial senescence and ischemic injury partly via Drp1 regulation
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Takeshi Tokuyama, Hideki Uosaki, Ayumu Sugiura, Gen Nishitai, Keisuke Takeda, Shun Nagashima, Isshin Shiiba, Naoki Ito, Taku Amo, Satoshi Mohri, Akiyuki Nishimura, Motohiro Nishida, Ayumu Konno, Hirokazu Hirai, Satoshi Ishido, Takahiro Yoshizawa, Takayuki Shindo, Shingo Takada, Shintaro Kinugawa, Ryoko Inatome, and Shigeru Yanagi
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Physiology ,Cellular physiology ,Molecular biology ,Developmental biology ,Science - Abstract
Summary: Abnormal mitochondrial fragmentation by dynamin-related protein1 (Drp1) is associated with the progression of aging-associated heart diseases, including heart failure and myocardial infarction (MI). Here, we report a protective role of outer mitochondrial membrane (OMM)-localized E3 ubiquitin ligase MITOL/MARCH5 against cardiac senescence and MI, partly through Drp1 clearance by OMM-associated degradation (OMMAD). Persistent Drp1 accumulation in cardiomyocyte-specific MITOL conditional-knockout mice induced mitochondrial fragmentation and dysfunction, including reduced ATP production and increased ROS generation, ultimately leading to myocardial senescence and chronic heart failure. Furthermore, ischemic stress-induced acute downregulation of MITOL, which permitted mitochondrial accumulation of Drp1, resulted in mitochondrial fragmentation. Adeno-associated virus-mediated delivery of the MITOL gene to cardiomyocytes ameliorated cardiac dysfunction induced by MI. Our findings suggest that OMMAD activation by MITOL can be a therapeutic target for aging-associated heart diseases, including heart failure and MI.
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- 2022
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16. The Ser19Stop single nucleotide polymorphism (SNP) of human PHYHIPL affects the cerebellum in mice
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Hisako Sugimoto, Takuro Horii, Jun-Na Hirota, Yoshitake Sano, Yo Shinoda, Ayumu Konno, Hirokazu Hirai, Yasuki Ishizaki, Hajime Hirase, Izuho Hatada, Teiichi Furuichi, and Tetsushi Sadakata
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PHYHIPL ,PHYHIP ,dbSNP ,HapMap Project ,Cerebellum ,Purkinje cell ,Neurology. Diseases of the nervous system ,RC346-429 - Abstract
Abstract The HapMap Project is a major international research effort to construct a resource to facilitate the discovery of relationships between human genetic variations and health and disease. The Ser19Stop single nucleotide polymorphism (SNP) of human phytanoyl-CoA hydroxylase-interacting protein-like (PHYHIPL) gene was detected in HapMap project and registered in the dbSNP. PHYHIPL gene expression is altered in global ischemia and glioblastoma multiforme. However, the function of PHYHIPL is unknown. We generated PHYHIPL Ser19Stop knock-in mice and found that PHYHIPL impacts the morphology of cerebellar Purkinje cells (PCs), the innervation of climbing fibers to PCs, the inhibitory inputs to PCs from molecular layer interneurons, and motor learning ability. Thus, the Ser19Stop SNP of the PHYHIPL gene may be associated with cerebellum-related diseases.
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- 2021
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17. GABAergic neuron-specific whole-brain transduction by AAV-PHP.B incorporated with a new GAD65 promoter
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Chiaki Hoshino, Ayumu Konno, Nobutake Hosoi, Ryosuke Kaneko, Ryo Mukai, Junichi Nakai, and Hirokazu Hirai
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Interneuron ,Inhibitory neuron ,Chandelier cell ,GAD65 ,Dlx ,GABA ,Neurology. Diseases of the nervous system ,RC346-429 - Abstract
Abstract GABAergic interneurons play a critical role in tuning neural networks in the central nervous system, and their defects are associated with neuropsychiatric disorders. Currently, the mDlx enhancer is solely used for adeno-associated virus (AAV) vector-mediated transgene delivery into cortical interneurons. Here, we developed a new inhibitory neuron-specific promoter (designated as the mGAD65 promoter), with a length of 2.5 kb, from a mouse genome upstream of exon 1 of the Gad2 gene encoding glutamic acid decarboxylase (GAD) 65. Intravenous infusion of blood–brain barrier-penetrating AAV-PHP.B expressing an enhanced green fluorescent protein under the control of the mGAD65 promoter transduced the whole brain in an inhibitory neuron-specific manner. The specificity and efficiency of the mGAD65 promoter for GABAergic interneurons, which was assessed at the motor cortex, were almost identical to or slightly higher than those of the mDlx enhancer. Immunohistochemical analysis revealed that the mGAD65 promoter preferentially transduced parvalbumin (PV)-expressing interneurons. Notably, the mGAD65 promoter transduced chandelier cells more efficiently than the mDlx enhancer and robustly labeled their synaptic boutons, called the cartridge, targeting the axon initial segments of excitatory pyramidal neurons. To test the ability of the mGAD65 promoter to express a functional molecule, we virally expressed G-CaMP, a fluorescent Ca2+ indicator, in the motor cortex, and this enabled us to monitor spontaneous and drug-induced Ca2+ activity in GABAergic inhibitory neurons. These results suggest that the mGAD65 promoter is useful for AAV-mediated targeting and manipulation of GABAergic neurons with the dominance of cortical PV-expressing neurons, including chandelier cells.
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- 2021
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18. Distinct temporal integration of noradrenaline signaling by astrocytic second messengers during vigilance
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Yuki Oe, Xiaowen Wang, Tommaso Patriarchi, Ayumu Konno, Katsuya Ozawa, Kazuko Yahagi, Hirokazu Hirai, Takashi Tsuboi, Tetsuya Kitaguchi, Lin Tian, Thomas J. McHugh, and Hajime Hirase
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Science - Abstract
Astrocytic GPCRs activate Ca2+ and cAMP signaling pathways, however, the in vivo dynamics of the two second messengers have not been fully been characterized. The authors demonstrate distinct noradrenaline-induced astrocytic Ca2+ and cAMP dynamics during startle and fear conditioning.
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- 2020
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19. Intestinal immunomodulatory activity of indigestible glucan in mice and its utilization by intestinal bacteria in vitro
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Ayumu Horinouchi, Hirokazu Hirai, Rika Hirano, Shin Kurihara, Hiroki Takagi, and Kenji Matsumoto
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Indigestible glucan ,Intestinal IgA ,Mucin ,Animal experiment ,Intestinal bacteria ,Prebiotics ,Nutrition. Foods and food supply ,TX341-641 - Abstract
The functional properties of indigestible glucan (IG), a newly developed water-soluble glucose polymer, on intestinal health were investigated. Of high amylose cornstarch, fructooligosaccharides, and IG, IG showed the strongest intestinal IgA induction during the 5-week experiment with BALB/cA mice (3% per diet). Next, separated IG samples, degree of polymerization (DP) ≥ 5 (High-IG) and DP ≤ 5 (Low-IG) fractions, were fed to BALB/cA mice for 9 weeks (3% per diet). High-IG consumption immediately increased the fecal IgA, whereas Low-IG consumption gradually increased it. Fecal mucin was significantly increased in the High-IG group but not in the Low-IG group. To identify utilization of High-IG and Low-IG by intestinal bacteria, in vitro assays were performed using 57 human gut bacterial strains. Few bacteria utilized High-IG, whereas more bacteria, particularly Bifidobacterium strains, utilized Low-IG. These results indicate that IG ingestion benefits the intestinal environment via strengthening the intestinal barrier function and prebiotic effects.
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- 2021
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20. Chronic optogenetic stimulation of Bergman glia leads to dysfunction of EAAT1 and Purkinje cell death, mimicking the events caused by expression of pathogenic ataxin-1
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Anton N. Shuvaev, Olga S. Belozor, Oleg Mozhei, Dariya A. Yakovleva, Ilya V. Potapenko, Andrey N. Shuvaev, Marina V. Smolnikova, Vladimir V. Salmin, Alla B. Salmina, Hirokazu Hirai, Anja G. Teschemacher, and Sergey Kasparov
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Bergmann glia ,Excitatory amino acid transporter type 1 ,Spinocerebellar ataxia type 1 ,Cerebellum ,Purkinje cell ,Neurosciences. Biological psychiatry. Neuropsychiatry ,RC321-571 - Abstract
Bergmann glia (BG) are highly specialized radial astrocytes of the cerebellar cortex, which play a key role in the uptake of synaptic glutamate via the excitatory amino acid transporter EAAT1. Multiple lines of evidence suggest that in cerebellar neurodegenerative diseases reactive BG has a negative impact on neuronal function and survival through compromised EAAT activity. A family of such diseases are those caused by expansion of CAG repeats in genes of the ataxin family, resulting in spinocerebellar ataxias (SCA).We investigated the contribution of BG to the pathogenesis of cerebellar neurodegeneration in a model of SCA1, which was induced by expression of a polyglutamine mutant of ataxin-1 (ATXN1[Q85]) in BG specifically. We compared the outcomes with a novel model where we triggered excitotoxicity by a chronic optogenetic activation of BG with channelrhodopsin-2 (ChR2). In both cases we detected evidence of reduced glutamate uptake manifested by prolongation of excitatory postsynaptic currents in Purkinje cells which is consistent with documented reduction of expression and/or function of EAAT1. In both models we detected astroglyosis and Purkinje cells atrophy. Finally, the same pattern was detected in a knock-in mouse which expresses a polyglutamine mutant ataxin-1 ATXN1[Q154] in a non-cell-selective manner.Our results suggest that ATXN1[Q85] and ChR2-induced insult targeted to BG closely mimics SCA1 pathology, where excessive glutamate signaling appears to be a common feature likely being an important contributor to cerebellar neurodegeneration.
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- 2021
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21. Rapamycin activates mammalian microautophagy
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Masahiro Sato, Takahiro Seki, Ayumu Konno, Hirokazu Hirai, Yuki Kurauchi, Akinori Hisatsune, and Hiroshi Katsuki
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Therapeutics. Pharmacology ,RM1-950 - Abstract
Autophagy-lysosome proteolysis is classified into macroautophagy (MA), microautophagy (mA) and chaperone-mediated autophagy (CMA). In contrast to MA and CMA, mA have been mainly studied in yeast. In 2011, mammalian mA was identified as a pathway to deliver cytosolic proteins into multivesicular bodies. However, its molecular mechanism is quite different from yeast mA. Using a cell-based method to evaluate mA and CMA, we revealed that rapamycin, an activator of yeast mA, significantly activated mammalian mA. Although rapamycin activates MA, mA was also activated by rapamycin in MA-deficient cells. These findings suggest that rapamycin is a first-identified activator of mammalian mA. Keywords: Microautophagy, Chaperone-mediated autophagy, Rapamycin
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- 2019
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22. Pharmacological enhancement of retinoid-related orphan receptor α function mitigates spinocerebellar ataxia type 3 pathology
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Masashi Watanave, Chiaki Hoshino, Ayumu Konno, Yumi Fukuzaki, Yasunori Matsuzaki, Tohru Ishitani, and Hirokazu Hirai
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Cerebellum ,Purkinje cell ,Spinocerebellar ataxia type 3 ,Machado–Joseph disease ,AAV vector ,Retinoid-related orphan receptor alpha ,Neurosciences. Biological psychiatry. Neuropsychiatry ,RC321-571 - Abstract
Cerebellar Purkinje cells (PCs) are the sole output neurons of the cerebellar cortex, and damage to PCs results in motor deficits. Spinocerebellar ataxia type 3 (SCA3, also known as Machado–Joseph disease), a hereditary neurodegenerative disease, is caused by an abnormal expansion of the polyglutamine tract in the causative ATXN3 protein. SCA3 affects a wide range of cells in the central nervous system, including those in the cerebellum. To unravel SCA3 pathology, we used adeno-associated virus serotype 9 (AAV9) vectors to express full-length ATXN3 with an abnormally expanded 89 polyglutamine stretch (ATXN3[Q89]) in cerebellar neurons of mature wild-type mice. Mice expressing ATXN3[Q89] exhibited motor impairment in a manner dependent on the viral titer. Immunohistochemistry of the cerebellum showed ubiquitinated nuclear aggregates in PCs; degeneration of PC dendrites; and a significant decrease in multiple proteins including retinoid-related orphan receptor α (RORα), a transcription factor, and type 1 metabotropic glutamate receptor (mGluR1) signaling molecules. Patch clamp analysis of ATXN3[Q89]-expressing PCs revealed marked defects in mGluR1 signaling. Notably, the emergence of behavioral, morphological, and functional defects was inhibited by a single injection of SR1078, an RORα/γ agonist. These results suggest that RORα plays a key role in mutant ATXN3-mediated aberrant phenotypes and that the pharmacological enhancement of RORα could function as a method for therapeutic intervention in SCA3.
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- 2019
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23. Indirect Negative Effect of Mutant Ataxin-1 on Short- and Long-Term Synaptic Plasticity in Mouse Models of Spinocerebellar Ataxia Type 1
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Anton N. Shuvaev, Olga S. Belozor, Oleg I. Mozhei, Andrey N. Shuvaev, Yana V. Fritsler, Elena D. Khilazheva, Angelina I. Mosyagina, Hirokazu Hirai, Anja G. Teschemacher, and Sergey Kasparov
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spinocerebellar ataxia type 1 ,PPF ,PPD ,DSE ,LTD ,synaptic plasticity ,Cytology ,QH573-671 - Abstract
Spinocerebellar ataxia type 1 (SCA1) is an intractable progressive neurodegenerative disease that leads to a range of movement and motor defects and is eventually lethal. Purkinje cells (PC) are typically the first to show signs of degeneration. SCA1 is caused by an expansion of the polyglutamine tract in the ATXN1 gene and the subsequent buildup of mutant Ataxin-1 protein. In addition to its toxicity, mutant Ataxin-1 protein interferes with gene expression and signal transduction in cells. Recently, it is evident that ATXN1 is not only expressed in neurons but also in glia, however, it is unclear the extent to which either contributes to the overall pathology of SCA1. There are various ways to model SCA1 in mice. Here, functional deficits at cerebellar synapses were investigated in two mouse models of SCA1 in which mutant ATXN1 is either nonspecifically expressed in all cell types of the cerebellum (SCA1 knock-in (KI)), or specifically in Bergmann glia with lentiviral vectors expressing mutant ATXN1 under the control of the astrocyte-specific GFAP promoter. We report impairment of motor performance in both SCA1 models. In both cases, prominent signs of astrocytosis were found using immunohistochemistry. Electrophysiological experiments revealed alteration of presynaptic plasticity at synapses between parallel fibers and PCs, and climbing fibers and PCs in SCA1 KI mice, which is not observed in animals expressing mutant ATXN1 solely in Bergmann glia. In contrast, short- and long-term synaptic plasticity was affected in both SCA1 KI mice and glia-targeted SCA1 mice. Thus, non-neuronal mechanisms may underlie some aspects of SCA1 pathology in the cerebellum. By combining the outcomes of our current work with our previous data from the B05 SCA1 model, we further our understanding of the mechanisms of SCA1.
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- 2022
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24. Lysosomal dysfunction and early glial activation are involved in the pathogenesis of spinocerebellar ataxia type 21 caused by mutant transmembrane protein 240
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Takahiro Seki, Masahiro Sato, Yuki Kibe, Tomoko Ohta, Mutsumi Oshima, Ayumu Konno, Hirokazu Hirai, Yuki Kurauchi, Akinori Hisatsune, and Hiroshi Katsuki
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Spinocerebellar ataxia type 21 ,Transmembrane protein 240 ,Autophagic lysosomal protein degradation ,Cerebellar Purkinje cells ,Motor dysfunction ,Gliosis ,Neurosciences. Biological psychiatry. Neuropsychiatry ,RC321-571 - Abstract
Spinocerebellar ataxia type 21 (SCA21) is caused by missense or nonsense mutations of the transmembrane protein 240 (TMEM240). Molecular mechanisms of SCA21 pathogenesis remain unknown because the functions of TMEM240 have not been elucidated. We aimed to reveal the molecular pathogenesis of SCA21 using cell and mouse models that overexpressed the wild-type and SCA21 mutant TMEM240. In HeLa cells, overexpressed TMEM240 localized around large cytoplasmic vesicles. The SCA21 mutation did not affect this localization. Because these vesicles contained endosomal markers, we evaluated the effect of TMEM240 fused with a FLAG tag (TMEM-FL) on endocytosis and autophagic protein degradation. Wild-type TMEM-FL significantly impaired clathrin-mediated endocytosis, whereas the SCA21 mutants did not. The SCA21 mutant TMEM-FL significantly impaired autophagic lysosomal protein degradation, in contrast to wild-type. Next, we investigated how TMEM240 affects the neural morphology of primary cultured cerebellar Purkinje cells (PCs). The SCA21 mutant TMEM-FL significantly prevented the dendritic development of PCs, in contrast to the wild-type. Finally, we assessed mice that expressed wild-type or SCA21 mutant TMEM-FL in cerebellar neurons using adeno-associated viral vectors. Mice expressing the SCA21 mutant TMEM-FL showed impaired motor coordination. Although the SCA21 mutant TMEM-FL did not trigger neurodegeneration, activation of microglia and astrocytes was induced before motor miscoordination. In addition, immunoblot experiments revealed that autophagic lysosomal protein degradation, especially chaperone-mediated autophagy, was also impaired in the cerebella that expressed the SCA21 mutant TMEM-FL. These dysregulated functions in vitro, and induction of early gliosis and lysosomal impairment in vivo by the SCA21 mutant TMEM240 may contribute to the pathogenesis of SCA21.
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- 2018
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25. Protocol for BATTLE-1EX: A High-Resolution Imaging Method to Visualize Whole Synaptic Structures and their Components in the Nervous System
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Akitoshi Inoue, Takuya Kobayashi, Hirokazu Hirai, Noriko Kanaya, and Keigo Kohara
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Microscopy ,Molecular Biology ,Neuroscience ,Science (General) ,Q1-390 - Abstract
Summary: This protocol describes BATTLE-1EX, which is a combined method of BATTLE-1 and expansion microscopy to obtain high-resolution imaging of whole synaptic structures and their components of hippocampal neural circuits. BATTLE-1 uses two genetically engineered recombinase proteins and competition between two recombinases that can be independently titrated, resulting in a tunable proportion of mCherry+/YFP− and YFP+/mCherry− cells. As a combinational method, BATTLE-1EX has the potential to visualize and dissect whole synaptic structures in numerous regions in the brain.For complete details on the use and execution of this protocol, please refer to Kohara et al. (2020).
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- 2020
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26. D-Cysteine Activates Chaperone-Mediated Autophagy in Cerebellar Purkinje Cells via the Generation of Hydrogen Sulfide and Nrf2 Activation
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Erika Ueda, Tomoko Ohta, Ayumu Konno, Hirokazu Hirai, Yuki Kurauchi, Hiroshi Katsuki, and Takahiro Seki
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aromatic-turmerone ,dopaminergic neurons ,microglia ,Nrf2 ,Cytology ,QH573-671 - Abstract
Chaperone-mediated autophagy (CMA) is a pathway in the autophagy-lysosome protein degradation system. CMA impairment has been implicated to play a role in spinocerebellar ataxia (SCA) pathogenesis. D-cysteine is metabolized by D-amino acid oxidase (DAO), leading to hydrogen sulfide generation in the cerebellum. Although D-cysteine alleviates the disease phenotypes in SCA-model mice, it remains unknown how hydrogen sulfide derived from D-cysteine exerts this effect. In the present study, we investigated the effects of D-cysteine and hydrogen sulfide on CMA activity using a CMA activity marker that we have established. D-cysteine activated CMA in Purkinje cells (PCs) of primary cerebellar cultures where DAO was expressed, while it failed to activate CMA in DAO-deficient AD293 cells. In contrast, Na2S, a hydrogen sulfide donor, activated CMA in both PCs and AD293 cells. Nuclear factor erythroid 2-related factor 2 (Nrf2) is known to be activated by hydrogen sulfide and regulate CMA activity. An Nrf2 inhibitor, ML385, prevented CMA activation triggered by D-cysteine and Na2S. Additionally, long-term treatment with D-cysteine increased the amounts of Nrf2 and LAMP2A, a CMA-related protein, in the mouse cerebellum. These findings suggest that hydrogen sulfide derived from D-cysteine enhances CMA activity via Nrf2 activation.
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- 2022
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27. BATTLE: Genetically Engineered Strategies for Split-Tunable Allocation of Multiple Transgenes in the Nervous System
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Keigo Kohara, Akitoshi Inoue, Yousuke Nakano, Hirokazu Hirai, Takuya Kobayashi, Masato Maruyama, Ryosuke Baba, and Chiho Kawashima
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Molecular Biology ,Molecular Neuroscience ,Neuroanatomy ,Techniques in Neuroscience ,Science - Abstract
Summary: Elucidating fine architectures and functions of cellular and synaptic connections requires development of new flexible methods. Here, we created a concept called the “battle of transgenes,” based on which we generated strategies using genetically engineered battles of multiple recombinases. The strategies enabled split-tunable allocation of multiple transgenes. We demonstrated the versatility of these strategies and technologies in inducing strong and multi-sparse allocations of multiple transgenes. Furthermore, the combination of our transgenic strategy and expansion microscopy enabled three-dimensional high-resolution imaging of whole synaptic structures in the hippocampus with simultaneous visualizations of endogenous synaptic proteins. These strategies and technologies based on the battle of genes may accelerate the analysis of whole synaptic and cellular connections in diverse life science fields.
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- 2020
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28. Mechanical Regulation Underlies Effects of Exercise on Serotonin-Induced Signaling in the Prefrontal Cortex Neurons
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Youngjae Ryu, Takahiro Maekawa, Daisuke Yoshino, Naoyoshi Sakitani, Atsushi Takashima, Takenobu Inoue, Jun Suzurikawa, Jun Toyohara, Tetsuro Tago, Michiru Makuuchi, Naoki Fujita, Keisuke Sawada, Shuhei Murase, Masashi Watanave, Hirokazu Hirai, Takamasa Sakai, Yuki Yoshikawa, Toru Ogata, Masahiro Shinohara, Motoshi Nagao, and Yasuhiro Sawada
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Science - Abstract
Summary: Mechanical forces are known to be involved in various biological processes. However, it remains unclear whether brain functions are mechanically regulated under physiological conditions. Here, we demonstrate that treadmill running and passive head motion (PHM), both of which produce mechanical impact on the head, have similar effects on the hallucinogenic 5-hydroxytryptamine (5-HT) receptor subtype 2A (5-HT2A) signaling in the prefrontal cortex (PFC) of rodents. PHM generates interstitial fluid movement that is estimated to exert shear stress of a few pascals on cells in the PFC. Fluid shear stress of a relevant magnitude on cultured neuronal cells induces ligand-independent internalization of 5-HT2A receptor, which is observed in mouse PFC neurons after treadmill running or PHM. Furthermore, inhibition of interstitial fluid movement by introducing polyethylene glycol hydrogel eliminates the effect of PHM on 5-HT2A receptor signaling in the PFC. Our findings indicate that neuronal cell function can be physiologically regulated by mechanical forces in the brain. : Biological Sciences; Neuroscience; Molecular Neuroscience; Cellular Neuroscience Subject Areas: Biological Sciences, Neuroscience, Molecular Neuroscience, Cellular Neuroscience
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- 2020
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29. Optimal number of endoscopic biopsies for diagnosis of early gastric cancer
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Masaki Nishitani, Naohiro Yoshida, Shigetsugu Tsuji, Teppei Masunaga, Hirokazu Hirai, Saori Miyajima, Akihiro Dejima, Takashi Nakashima, Shigenori Wakita, Kenichi Takemura, Hiroshi Minato, Shuichi Kaneko, and Hisashi Doyama
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Diseases of the digestive system. Gastroenterology ,RC799-869 - Abstract
Background and study aims No recommendations are available for optimal number of endoscopic biopsies for early gastric cancer (GC), and whether detection of early GC is improved by increasing the number of biopsy is unclear. We therefore evaluated the relationship between number of biopsies and diagnostic accuracy. Materials and methods We retrospectively evaluated 858 early GCs (623 from endoscopic submucosal dissection and 235 surgical specimens), which we classified as obtained after one, two, or three or more biopsies. We assessed diagnostic accuracy by number of biopsies, and in subgroups by tumor diameter, gross type, and surface color. Results Almost half the lesions were obtained after one biopsy each, 30 % after two biopsies, and 20 % after three or more biopsies. Although diagnostic accuracy increased with biopsy number, it was significantly greater for the two-biopsy group than the one-biopsy group, (92.5 % vs. 83.9 %, P = 0.0009), but did not significantly differ between the two- and three or more-biopsy groups. This finding was seen when tumors were evaluated by size, but not by elevated type and surface color, for which more biopsies did not improve diagnostic accuracy. Multivariate analysis demonstrated that two or more biopsies was the independent significant factors for diagnostic accuracy. Conclusions Two biopsies are the optimal number required to diagnose early GC.
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- 2019
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30. Minimal Purkinje Cell-Specific PCP2/L7 Promoter Virally Available for Rodents and Non-human Primates
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Keisuke Nitta, Yasunori Matsuzaki, Ayumu Konno, and Hirokazu Hirai
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Purkinje cell ,L7 ,PCP2 ,cerebellum ,lentivirus ,adeno-associated virus ,viral vector ,cell type-specific promoter ,marmoset ,non-human primate ,Genetics ,QH426-470 ,Cytology ,QH573-671 - Abstract
Cell-type-specific promoters in combination with viral vectors and gene-editing technology permit efficient gene manipulation in specific cell populations. Cerebellar Purkinje cells play a pivotal role in cerebellar functions. Although the Purkinje cell-specific L7 promoter is widely used for the generation of transgenic mice, it remains unsuitable for viral vectors because of its large size (3 kb) and exceedingly weak promoter activity. Here, we found that the 0.8-kb region (named here as L7-6) upstream of the transcription initiation codon in the first exon was alone sufficient as a Purkinje cell-specific promoter, presenting a far stronger promoter activity over the original 3-kb L7 promoter with a sustained significant specificity to Purkinje cells. Intravenous injection of adeno-associated virus vectors that are highly permeable to the blood-brain barrier confirmed the Purkinje cell specificity of the L7-6 in the CNS. The features of the L7-6 were also preserved in the marmoset, a non-human primate. The high sequence homology of the L7-6 among mouse, marmoset, and human suggests the preservation of the promoter strength and Purkinje cell specificity features also in humans. These findings suggest that L7-6 will facilitate the cerebellar research targeting the pathophysiology and gene therapy of cerebellar disorders.
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- 2017
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31. Red fluorescent protein-based cAMP indicator applicable to optogenetics and in vivo imaging
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Kazuki Harada, Motoki Ito, Xiaowen Wang, Mika Tanaka, Devina Wongso, Ayumu Konno, Hirokazu Hirai, Hajime Hirase, Takashi Tsuboi, and Tetsuya Kitaguchi
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Medicine ,Science - Abstract
Abstract cAMP is a common second messenger that is involved in various physiological processes. To expand the colour palette of available cAMP indicators, we developed a red cAMP indicator named “Pink Flamindo” (Pink Fluorescent cAMP indicator). The fluorescence intensity of Pink Flamindo increases 4.2-fold in the presence of a saturating dose of cAMP, with excitation and emission peaks at 567 nm and 590 nm, respectively. Live-cell imaging revealed that Pink Flamindo is effective for monitoring the spatio-temporal dynamics of intracellular cAMP generated by photoactivated adenylyl cyclase in response to blue light, and in dual-colour imaging studies using a green Ca2+ indicator (G-GECO). Furthermore, we successfully monitored the elevation of cAMP levels in vivo in cerebral cortical astrocytes by two-photon imaging. We propose that Pink Flamindo will facilitate future in vivo, optogenetic studies of cell signalling and cAMP dynamics.
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- 2017
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32. Contribution of Thyrotropin-Releasing Hormone to Cerebellar Long-Term Depression and Motor Learning
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Masashi Watanave, Yasunori Matsuzaki, Yasuyo Nakajima, Atsushi Ozawa, Masanobu Yamada, and Hirokazu Hirai
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thyrotropin-releasing hormone ,motor learning ,cerebellum ,LTD ,NO ,Neurosciences. Biological psychiatry. Neuropsychiatry ,RC321-571 - Abstract
Thyrotropin-releasing hormone (TRH) regulates various physiological activities through activation of receptors expressed in a broad range of cells in the central nervous system. The cerebellum expresses TRH receptors in granule cells and molecular layer interneurons. However, the function of TRH in the cerebellum remains to be clarified. Here, using TRH knockout (KO) mice we studied the role of TRH in the cerebellum. Immunohistochemistry showed no gross morphological differences between KO mice and wild-type (WT) littermates in the cerebellum. In the rotarod test, the initial performance of KO mice was comparable to that of WT littermates, but the learning speed of KO mice was significantly lower than that of WT littermates, suggesting impaired motor learning. The motor learning deficit in KO mice was rescued by intraperitoneal injection of TRH. Electrophysiology revealed absence of long-term depression (LTD) at parallel fiber-Purkinje cell synapses in KO mice, which was rescued by bath-application of TRH. TRH was shown to increase cyclic guanosine monophosphate (cGMP) content in the cerebellum. Since nitric oxide (NO) stimulates cGMP synthesis in the cerebellum, we examined whether NO-cGMP pathway was involved in TRH-mediated LTD rescue in KO mice. Pharmacological blockade of NO synthase and subsequent cGMP production prevented TRH-induced LTD expression in KO mice, whereas increase in cGMP signal in Purkinje cells by 8-bromoguanosine cyclic 3’,5’-monophosphate, a membrane-permeable cGMP analog, restored LTD without TRH application. These results suggest that TRH is involved in cerebellar LTD presumably by upregulating the basal cGMP level in Purkinje cells, and, consequently, in motor learning.
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- 2018
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33. Calcium imaging reveals glial involvement in transcranial direct current stimulation-induced plasticity in mouse brain
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Hiromu Monai, Masamichi Ohkura, Mika Tanaka, Yuki Oe, Ayumu Konno, Hirokazu Hirai, Katsuhiko Mikoshiba, Shigeyoshi Itohara, Junichi Nakai, Youichi Iwai, and Hajime Hirase
- Subjects
Science - Abstract
While transcranical direct current stimulation (tDCS) is used in clinical setting, its cellular mechanism of action is unclear. Here, Hajime Hirase and colleagues visualize cellular response in mouse brain to tDCS and show robust astrocyte activation that coincide with plasticity changes.
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- 2016
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34. Safety profile of the intravenous administration of brain-targeted stable nucleic acid lipid particles
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Mariana Conceição, Liliana Mendonça, Clévio Nóbrega, Célia Gomes, Pedro Costa, Hirokazu Hirai, João Nuno Moreira, Maria C. Lima, N. Manjunath, and Luís Pereira de Almeida
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Computer applications to medicine. Medical informatics ,R858-859.7 ,Science (General) ,Q1-390 - Abstract
In a clinical setting, where multiple administrations of the therapeutic agent are usually required to improve the therapeutic outcome, it is crucial to assess the immunogenicity of the administered nanoparticles. In this data work, we investigated the safety profile of the repeated intravenous administration of brain-targeted stable nucleic acid lipid particles (RVG-9r-targeted SNALPs). To evaluate local activation of the immune system, we performed analysis of mouse tissue homogenates and sections from cerebellum. To investigate peripheral activation of the immune system, we used serum of mice that were intravenously injected with RVG-9r-targeted SNALPs. These data are related and were discussed in the accompanying research article entitled “Intravenous administration of brain-targeted stable nucleic acid lipid particles alleviates Machado–Joseph disease neurological phenotype” (Conceição et al., in press) [1].
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- 2016
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35. Author Correction: Distinct temporal integration of noradrenaline signaling by astrocytic second messengers during vigilance
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Yuki Oe, Xiaowen Wang, Tommaso Patriarchi, Ayumu Konno, Katsuya Ozawa, Kazuko Yahagi, Hirokazu Hirai, Takashi Tsuboi, Tetsuya Kitaguchi, Lin Tian, Thomas J. McHugh, and Hajime Hirase
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Science - Abstract
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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- 2020
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36. Inhibition gates supralinear Ca2+ signaling in Purkinje cell dendrites during practiced movements
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Michael A Gaffield, Matthew J M Rowan, Samantha B Amat, Hirokazu Hirai, and Jason M Christie
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cerebellum ,inhibition ,calcium ,Medicine ,Science ,Biology (General) ,QH301-705.5 - Abstract
Motor learning involves neural circuit modifications in the cerebellar cortex, likely through re-weighting of parallel fiber inputs onto Purkinje cells (PCs). Climbing fibers instruct these synaptic modifications when they excite PCs in conjunction with parallel fiber activity, a pairing that enhances climbing fiber-evoked Ca2+ signaling in PC dendrites. In vivo, climbing fibers spike continuously, including during movements when parallel fibers are simultaneously conveying sensorimotor information to PCs. Whether parallel fiber activity enhances climbing fiber Ca2+ signaling during motor behaviors is unknown. In mice, we found that inhibitory molecular layer interneurons (MLIs), activated by parallel fibers during practiced movements, suppressed parallel fiber enhancement of climbing fiber Ca2+ signaling in PCs. Similar results were obtained in acute slices for brief parallel fiber stimuli. Interestingly, more prolonged parallel fiber excitation revealed latent supralinear Ca2+ signaling. Therefore, the balance of parallel fiber and MLI input onto PCs regulates concomitant climbing fiber Ca2+ signaling.
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- 2018
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37. Npas4 Regulates Mdm2 and thus Dcx in Experience-Dependent Dendritic Spine Development of Newborn Olfactory Bulb Interneurons
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Sei-ichi Yoshihara, Hiroo Takahashi, Nobushiro Nishimura, Masahito Kinoshita, Ryo Asahina, Michiko Kitsuki, Kana Tatsumi, Yoko Furukawa-Hibi, Hirokazu Hirai, Taku Nagai, Kiyofumi Yamada, and Akio Tsuboi
- Subjects
Biology (General) ,QH301-705.5 - Abstract
Sensory experience regulates the development of various brain structures, including the cortex, hippocampus, and olfactory bulb (OB). Little is known about how sensory experience regulates the dendritic spine development of OB interneurons, such as granule cells (GCs), although it is well studied in mitral/tufted cells. Here, we identify a transcription factor, Npas4, which is expressed in OB GCs immediately after sensory input and is required for dendritic spine formation. Npas4 overexpression in OB GCs increases dendritic spine density, even under sensory deprivation, and rescues reduction of dendrite spine density in the Npas4 knockout OB. Furthermore, loss of Npas4 upregulates expression of the E3-ubiquitin ligase Mdm2, which ubiquitinates a microtubule-associated protein Dcx. This leads to reduction in the dendritic spine density of OB GCs. Together, these findings suggest that Npas4 regulates Mdm2 expression to ubiquitinate and degrade Dcx during dendritic spine development in newborn OB GCs after sensory experience.
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- 2014
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38. Fusion of Human Fetal Mesenchymal Stem Cells with 'Degenerating' Cerebellar Neurons in Spinocerebellar Ataxia Type 1 Model Mice.
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Fathul Huda, Yiping Fan, Mamiko Suzuki, Ayumu Konno, Yasunori Matsuzaki, Nobutaka Takahashi, Jerry K Y Chan, and Hirokazu Hirai
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Medicine ,Science - Abstract
Mesenchymal stem cells (MSCs) migrate to damaged tissues, where they participate in tissue repair. Human fetal MSCs (hfMSCs), compared with adult MSCs, have higher proliferation rates, a greater differentiation capacity and longer telomeres with reduced senescence. Therefore, transplantation of quality controlled hfMSCs is a promising therapeutic intervention. Previous studies have shown that intravenous or intracortical injections of MSCs result in the emergence of binucleated cerebellar Purkinje cells (PCs) containing an MSC-derived marker protein in mice, thus suggesting a fusion event. However, transdifferentiation of MSCs into PCs or transfer of a marker protein from an MSC to a PC cannot be ruled out. In this study, we unequivocally demonstrated the fusion of hfMSCs with murine PCs through a tetracycline-regulated (Tet-off) system with or without a Cre-dependent genetic inversion switch (flip-excision; FLEx). In the FLEx-Tet system, we performed intra-cerebellar injection of viral vectors expressing tetracycline transactivator (tTA) and Cre recombinase into either non-symptomatic (4-week-old) or clearly symptomatic (6-8-month-old) spinocerebellar ataxia type 1 (SCA1) mice. Then, the mice received an injection of 50,000 genetically engineered hfMSCs that expressed GFP only in the presence of Cre recombinase and tTA. We observed a significant emergence of GFP-expressing PCs and interneurons in symptomatic, but not non-symptomatic, SCA1 mice 2 weeks after the MSC injection. These results, together with the results obtained using age-matched wild-type mice, led us to conclude that hfMSCs have the potential to preferentially fuse with degenerating PCs and interneurons but not with healthy neurons.
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- 2016
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39. Viral Vector-Based Dissection of Marmoset GFAP Promoter in Mouse and Marmoset Brains.
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Yoichiro Shinohara, Ayumu Konno, Nobutaka Takahashi, Yasunori Matsuzaki, Shoji Kishi, and Hirokazu Hirai
- Subjects
Medicine ,Science - Abstract
Adeno-associated virus (AAV) vectors are small in diameter, diffuse easily in the brain, and represent a highly efficient means by which to transfer a transgene to the brain of a large animal. A major demerit of AAV vectors is their limited accommodation capacity for transgenes. Thus, a compact promoter is useful when delivering large transgenes via AAV vectors. In the present study, we aimed to identify the shortest astrocyte-specific GFAP promoter region that could be used for AAV-vector-mediated transgene expression in the marmoset brain. The 2.0-kb promoter region upstream of the GFAP gene was cloned from the marmoset genome, and short promoters (1.6 kb, 1.4 kb, 0.6 kb, 0.3 kb and 0.2 kb) were obtained by progressively deleting the original 2.0-kb promoter from the 5' end. The short promoters were screened in the mouse cerebellum in terms of their strength and astrocyte specificity. We found that the 0.3-kb promoter maintained 40% of the strength of the original 2.0-kb promoter, and approximately 90% of its astrocyte specificity. These properties were superior to those of the 1.4-kb, 0.6-kb (20% promoter strength) and 0.2-kb (70% astrocyte specificity) promoters. Then, we verified whether the 0.3-kb GFAP promoter retained astrocyte specificity in the marmoset cerebral cortex. Injection of viral vectors carrying the 0.3-kb marmoset GFAP promoter specifically transduced astrocytes in both the cerebral cortex and cerebellar cortex of the marmoset. These results suggest that the compact 0.3-kb promoter region serves as an astrocyte-specific promoter in the marmoset brain, which permits us to express a large gene by AAV vectors that have a limited accommodation capacity.
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- 2016
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40. Distinct transduction profiles in the CNS via three injection routes of AAV9 and the application to generation of a neurodegenerative mouse model
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Fathul Huda, Ayumu Konno, Yasunori Matsuzaki, Hanna Goenawan, Koichi Miyake, Takashi Shimada, and Hirokazu Hirai
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Genetics ,QH426-470 ,Cytology ,QH573-671 - Abstract
Using single-stranded adeno-associated virus serotype 9 (ssAAV9) vectors containing the neuron-specific synapsin-I promoter, we examined whether different administration routes (direct cerebellar cortical (DC), intrathecal (IT) and intravenous (IV) injections) could elicit specific transduction profiles in the CNS. The DC injection route robustly and exclusively transduced the whole cerebellum, whereas the IT injection route primarily transduced the cerebellar lobules 9 and 10 close to the injection site and the spinal cord. An IV injection in neonatal mice weakly and homogenously transduced broad CNS areas. In the cerebellar cortex, the DC and IT injection routes transduced all neuron types, whereas the IV injection route primarily transduced Purkinje cells. To verify the usefulness of this method, we generated a mouse model of spinocerebellar ataxia type 1 (SCA1). Mice that received a DC injection of the ssAAV9 vector expressing mutant ATXN1, a protein responsible for SCA1, showed the intranuclear aggregation of mutant ATXN1 in Purkinje cells, significant atrophy of the Purkinje cell dendrites and progressive motor deficits, which are characteristics of SCA1. Thus, ssAAV9-mediated transduction areas, levels, and cell types change depending on the route of injection. Moreover, this approach can be used for the generation of different mouse models of CNS/neurodegenerative diseases.
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- 2014
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41. Silencing mutant ataxin-3 rescues motor deficits and neuropathology in Machado-Joseph disease transgenic mice.
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Clévio Nóbrega, Isabel Nascimento-Ferreira, Isabel Onofre, David Albuquerque, Hirokazu Hirai, Nicole Déglon, and Luís Pereira de Almeida
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Medicine ,Science - Abstract
Machado-Joseph disease (MJD) or spinocerebellar ataxia type 3 (SCA3) is an autosomal dominantly-inherited neurodegenerative disorder caused by the over-repetition of a CAG codon in the MJD1 gene. This expansion translates into a polyglutamine tract that confers a toxic gain-of-function to the mutant protein--ataxin-3, leading to neurodegeneration in specific brain regions, with particular severity in the cerebellum. No treatment able to modify the disease progression is available. However, gene silencing by RNA interference has shown promising results. Therefore, in this study we investigated whether lentiviral-mediated allele-specific silencing of the mutant ataxin-3 gene, after disease onset, would rescue the motor behavior deficits and neuropathological features in a severely impaired transgenic mouse model of MJD. For this purpose, we injected lentiviral vectors encoding allele-specific silencing-sequences (shAtx3) into the cerebellum of diseased transgenic mice expressing the targeted C-variant of mutant ataxin-3 present in 70% of MJD patients. This variation permits to discriminate between the wild-type and mutant forms, maintaining the normal function of the wild-type allele and silencing only the mutant form. Quantitative analysis of rotarod performance, footprint and activity patterns revealed significant and robust alleviation of gait, balance (average 3-fold increase of rotarod test time), locomotor and exploratory activity impairments in shAtx3-injected mice, as compared to control ones injected with shGFP. An important improvement of neuropathology was also observed, regarding the number of intranuclear inclusions, calbindin and DARPP-32 immunoreactivity, fluorojade B and Golgi staining and molecular and granular layers thickness. These data demonstrate for the first time the efficacy of gene silencing in blocking the MJD-associated motor-behavior and neuropathological abnormalities after the onset of the disease, supporting the use of this strategy for therapy of MJD.
- Published
- 2013
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42. The murine stem cell virus promoter drives correlated transgene expression in the leukocytes and cerebellar Purkinje cells of transgenic mice.
- Author
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Miho Oue, Hiroshi Handa, Yasunori Matsuzaki, Kazutomo Suzue, Hirokazu Murakami, and Hirokazu Hirai
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Medicine ,Science - Abstract
The murine stem cell virus (MSCV) promoter exhibits activity in mouse hematopoietic cells and embryonic stem cells. We generated transgenic mice that expressed enhanced green fluorescent protein (GFP) under the control of the MSCV promoter. We obtained 12 transgenic founder mice through 2 independent experiments and found that the bodies of 9 of the founder neonates emitted different levels of GFP fluorescence. Flow cytometric analysis of circulating leukocytes revealed that the frequency of GFP-labeled leukocytes among white blood cells ranged from 1.6% to 47.5% across the 12 transgenic mice. The bodies of 9 founder transgenic mice showed various levels of GFP expression. GFP fluorescence was consistently observed in the cerebellum, with faint or almost no fluorescence in other brain regions. In the cerebellum, 10 founders exhibited GFP expression in Purkinje cells at frequencies of 3% to 76%. Of these, 4 mice showed Purkinje cell-specific expression, while 4 and 2 mice expressed GFP in the Bergmann glia and endothelial cells, respectively. The intensity of the GFP fluorescence in the body was relative to the proportion of GFP-positive leukocytes. Moreover, the frequency of the GFP-expressing leukocytes was significantly correlated with the frequency of GFP-expressing Purkinje cells. These results suggest that the MSCV promoter is useful for preferentially expressing a transgene in Purkinje cells. In addition, the proportion of transduced leukocytes in the peripheral circulation reflects the expression level of the transgene in Purkinje cells, which can be used as a way to monitor transgene expression properties in the cerebellum without invasive techniques.
- Published
- 2012
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43. Contribution of Thyrotropin-Releasing Hormone to Cerebellar Long-Term Depression and Motor Learning.
- Author
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Masashi Watanave, Yasunori Matsuzaki, Yasuyo Nakajima, Atsushi Ozawa, Masanobu Yamada, and Hirokazu Hirai
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THYROTROPIN releasing factor ,CYCLIC guanylic acid ,GRANULE cells ,CENTRAL nervous system ,MOTOR learning - Abstract
Thyrotropin-releasing hormone (TRH) regulates various physiological activities through activation of receptors expressed in a broad range of cells in the central nervous system. The cerebellum expresses TRH receptors in granule cells and molecular layer interneurons. However, the function of TRH in the cerebellum remains to be clarified. Here, using TRH knockout (KO) mice we studied the role of TRH in the cerebellum. Immunohistochemistry showed no gross morphological differences between KO mice and wild-type (WT) littermates in the cerebellum. In the rotarod test, the initial performance of KO mice was comparable to that of WT littermates, but the learning speed of KO mice was significantly lower than that of WT littermates, suggesting impaired motor learning. The motor learning deficit in KO mice was rescued by intraperitoneal injection of TRH. Electrophysiology revealed absence of longterm depression (LTD) at parallel fiber-Purkinje cell synapses in KO mice, which was rescued by bath-application of TRH. TRH was shown to increase cyclic guanosine monophosphate (cGMP) content in the cerebellum. Since nitric oxide (NO) stimulates cGMP synthesis in the cerebellum, we examined whether NO-cGMP pathway was involved in TRH-mediated LTD rescue in KO mice. Pharmacological blockade of NO synthase and subsequent cGMP production prevented TRH-induced LTD expression in KO mice, whereas increase in cGMP signal in Purkinje cells by 8-bromoguanosine cyclic 3',5'-monophosphate, a membrane-permeable cGMP analog, restored LTD without TRH application. These results suggest that TRH is involved in cerebellar LTD presumably by upregulating the basal cGMP level in Purkinje cells, and, consequently, in motor learning. [ABSTRACT FROM AUTHOR]
- Published
- 2024
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44. Long-term depression–inductive stimulation causes long-term potentiation in mouse Purkinje cells with a mutant thyroid hormone receptor.
- Author
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Ayane Ninomiya, Izuki Amano, Michifumi Kokubo, Yusuke Takatsuru, Sumiyasu Ishii, Hirokazu Hirai, Nobutake Hosoi, and Noriyuki Koibuchi
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THYROID hormone receptors ,PURKINJE cells ,LONG-term potentiation ,CENTRAL nervous system ,MOTOR ability - Abstract
Thyroid hormones (THs) regulate gene expression by binding to nuclear TH receptors (TRs) in the cell. THs are indispensable for brain development. However, we have little knowledge about how congenital hypothyroidism in neurons affects functions of the central nervous system in adulthood. Here, we report specific TH effects on functional development of the cerebellum by using transgenic mice overexpressing a dominant negative TR (Mf-1) specifically in cerebellar Purkinje cells (PCs). Adult Mf-1 mice displayed impairments in motor coordination and motor learning. Surprisingly, long-term depression (LTD)–inductive stimulation caused long-term potentiation (LTP) at parallel fiber (PF)–PC synapses in adult Mf-1 mice, although there was no abnormality in morphology or basal properties of PF–PC synapses. The LTP phenotype was turned to LTD in Mf-1 mice when the inductive stimulation was applied in an extracellular high Ca
2+ condition. Confocal calcium imaging revealed that dendritic Ca2+ elevation evoked by LTD-inductive stimulation is significantly reduced in Mf-1 PCs but not by PC depolarization only. Single PC messenger RNA quantitative analysis showed reduced expression of SERCA2 and IP3 receptor type 1 in Mf-1 PCs, which are essential for mGluR1-mediated internal calcium release from endoplasmic reticulum in cerebellar PCs. These abnormal changes were not observed in adult-onset PC-specific TH deficiency mice created by adeno-associated virus vectors. Thus, we propose the importance of TH action during neural development in establishing proper cerebellar function in adulthood, independent of its morphology. The present study gives insight into the cellular and molecular mechanisms underlying congenital hypothyroidism–induced dysfunctions of central nervous system and cerebellum. [ABSTRACT FROM AUTHOR]- Published
- 2022
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45. Protein kinase Cγ in cerebellar Purkinje cells regulates Ca2+-activated large-conductance K+ channels and motor coordination.
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Masashi Watanave, Nobutaka Takahashi, Nobutake Hosoi, Ayumu Konno, Hikaru Yamamoto, Hiroyuki Yasui, Mika Kawachi, Takuro Horii, Yasunori Matsuzaki, Izuho Hatada, and Hirokazu Hirai
- Subjects
MOTOR ability ,PURKINJE cells ,PROTEIN kinases ,PROTEIN kinase C ,CENTRAL nervous system ,SOYBEAN meal ,COMMERCIAL products - Abstract
The cerebellum, the site where protein kinase C (PKC) was first discovered, contains the highest amount of PKC in the central nervous system, with PKCγ being the major isoform. Systemic PKCγ- knockout (KO) mice showed impaired motor coordination and deficient pruning of surplus climbing fibers (CFs) from developing cerebellar Purkinje cells (PCs). However, the physiological significance of PKCγ in the mature cerebellum and the cause of motor incoordination remain unknown. Using adeno-associated virus vectors targeting PCs, we showed that impaired motor coordination was restored by re-expression of PKCγ in mature PKCγ-KO mouse PCs in a kinase activity-dependent manner, while normal motor coordination in mature Prkcgfl/fl mice was impaired by the Cre-dependent removal of PKCγ from PCs. Notably, the rescue or removal of PKCγ from mature PKCγ-KO or Prkcgfl/fl mice, respectively, did not affect the CF innervation profile of PCs, suggesting the presence of a mechanism distinct from multiple CF innervation of PCs for the motor defects in PKCγ-deficient mice. We found marked potentiation of Ca2+-activated large-conductance K+ (BK) channel currents in PKCγ-deficient mice, as compared to wild-type mice, which decreased the membrane resistance, resulting in attenuation of the electrical signal during the propagation and significant alterations of the complex spike waveform. These changes in PKCγ-deficient mice were restored by the rescue of PKCγ or pharmacological suppression of BK channels. Our results suggest that PKCγ is a critical regulator that negatively modulates BK currents in PCs, which significantly influences PC output from the cerebellar cortex and, eventually, motor coordination. [ABSTRACT FROM AUTHOR]
- Published
- 2022
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46. White globe appearance is an endoscopic predictive factor for synchronous multiple gastric cancer.
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Teppei Masunaga, Naohiro Yoshida, Shinichiro Akiyama, Gen Sugiyama, Hirokazu Hirai, Saori Miyajima, Shigenori Wakita, Yosuke Kito, Hiroyoshi Nakanishi, Kunihiro Tsuji, Kazuhiro Matsunaga, Shigetsugu Tsuji, Kenichi Takemura, Kazuyoshi Katayanagi, Hiroshi Minato, and Hisashi Doyama
- Subjects
STOMACH cancer ,HELICOBACTER pylori ,UNIVARIATE analysis ,MULTIVARIATE analysis ,ODDS ratio ,HELICOBACTER pylori infections ,DUODENAL ulcers ,ENDOSCOPY - Abstract
Background White globe appearance (WGA) is a small white lesion with a globular shape identified during magnifying endoscopy with narrow-band imaging. However, the association between WGA and synchronous multiple gastric cancer (SMGC) remains unclear. Methods Consecutive patients who underwent endoscopic submucosal dissection for gastric cancer (GC) between July 2013 and April 2015 at our institution were eligible for this study. We excluded patients with a history of gastric tumor or gastrectomy. Patients who had more than 2 GCs in their postoperative pathological evaluation were classified as SMGC-positive, and patients who had at least 1 WGA-positive GC were classified as WGA-positive patients. The primary outcome was a comparison of the prevalence of WGA in patients classified as SMGCpositive and SMGC-negative. Univariate and multivariate analyses were performed using the following variables: WGA, age, sex, atrophy, and Helicobacter pylori (H. pylori) status. Results There were 26 and 181 patients classified as SMGC-positive and SMGC-negative, respectively. Univariate analysis revealed that WGA-positive classification (50% vs. 23%, P=0.008) and male sex (88% vs. 66%, P=0.02) were significant factors associated with SMGC classification, while age ≥65 years (81% vs. 81%, P>0.99), severe atrophy (46% vs. 46%, P>0.99), and H. pylori positivity (69% vs. 65%, P=0.8) were not. In the multivariate analysis, only WGA-positive classification (odds ratio 2.78, 95% confidence interval 1.16-6.67; P=0.02) was a significant independent risk factor for SMGC. Conclusions Our exploratory study showed the possibility of WGA as a predictive factor for SMGC. In cases of WGA-positive gastric cancer, careful examination might be needed to diagnose SMGC. [ABSTRACT FROM AUTHOR]
- Published
- 2021
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47. Deletion of Class II ADP-Ribosylation Factors in Mice Causes Tremor by the Nav1.6 Loss in Cerebellar Purkinje Cell Axon Initial Segments.
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Nobutake Hosoi, Koji Shibasaki, Mayu Hosono, Ayumu Konno, Yo Shinoda, Hiroshi Kiyonari, Kenichi Inoue, Shin-ichi Muramatsu, Yasuki Ishizaki, Hirokazu Hirai, Teiichi Furuichi, and Tetsushi Sadakata
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PURKINJE cells ,PROTHROMBIN ,IMMOBILIZED proteins ,AXONS ,ESSENTIAL tremor - Abstract
ADP-ribosylation factors (ARFs) are a family of small monomeric GTPases comprising six members categorized into three classes: class I (ARF1,2, and 3), class II (ARF4 and 5), and class III (ARF6). In contrast to class I and III ARFs, which are the key regulators in vesicular membrane trafficking, the cellular function of class II ARFs remains unclear. In the present study, we generated class II ARF-deficient mice and found that ARF4 +/-/ARF5-/- mice exhibited essential tremor (ET)-like behaviors. In vivo electrophysiological recordings revealed that ARF4 1 /ARF5 /mice ofboth sexes exhibited abnormal brain activity when moving, raising the possibility of abnormal cerebellar excitability. Slice patch-damp experiments demonstrated the reduced excitability of the cerebellar Purkinje cells (PCs) in ARF4+/-/ARF5-/- mice. Immunohistochemical and electrophysiological analyses revealed a severe and selective decrease of poreforming voltage-dependent NaH channel subunit Navi.6, important for maintaining repetitive action potential firing, in the axon initial segment (AIS) of PCs. Importantly, this decrease in Navi.6 protein localized in the AIS and the consequent tremors in ARF4+/- ARF5 mice could be alleviated by the PC-specific expression of ARF5 using adeno-associated virus vectors. Together, our data demonstrate that the decreased expression of the class IIARF proteins in ARF4+/-/ARF5 mice, leading to a haploinsufficiency of ARF4 in the absence of ARF5, impairs the localization of Navl.6 to the AIS and hence reduces the membrane excitability in PCs, resulting in the ET-like movement disorder. We suggest that class II ARFs function in localizing specific proteins, such as Navl.6, to the AIS. [ABSTRACT FROM AUTHOR]
- Published
- 2019
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- View/download PDF
48. A Subtype of Olfactory Bulb Interneurons Is Required for Odor Detection and Discrimination Behaviors.
- Author
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Hiroo Takahashi, Yoichi Ogawa, Sei-ichi Yoshihara, Ryo Asahina, Masahito Kinoshita, Tatsuro Kitano, Michiko Kitsuki, Kana Tatsumi, Mamiko Okuda, Kouko Tatsumi, Akio Wanaka, Hirokazu Hirai, Stern, Peter L., and Akio Tsuboi
- Subjects
OLFACTORY bulb ,INTERNEURONS ,MEMBRANE proteins ,OPTOGENETICS ,CARCINOEMBRYONIC antigen ,GRANULE cells - Abstract
Neural circuits that undergo reorganization by newborn interneurons in the olfactory bulb (OB) are necessary for odor detection and discrimination, olfactory memory, and innate olfactory responses, including predator avoidance and sexual behaviors. TheOBpossesses many interneurons, including various types of granule cells (GCs); however, the contribution that each type of interneuron makes to olfactory behavioral control remains unknown. Here, we investigated the in vivo functional role of oncofetal trophoblast glycoprotein 5T4, a regulator for dendritic arborization of 5T4-expressing GCs (5T4 GCs), the level of which is reduced in the OB of 5T4 knock-out (KO) mice. Electrophysiological recordings with acute OB slices indicated that external tufted cells (ETCs) can be divided into two types, bursting and nonbursting. Optogenetic stimulation of 5T4 GCs revealed their connection to both bursting and nonbursting ETCs, as well as to mitral cells (MCs). Interestingly, nonbursting ETCs received fewer inhibitory inputs from GCs in 5T4 KO mice than from those in wild-type (WT) mice, whereas bursting ETCs and MCs received similar inputs in both mice. Furthermore, 5T4 GCs received significantly fewer excitatory inputs in 5T4 KO mice. Remarkably, in olfactory behavior tests, 5T4 KO mice had higher odor detection thresholds than the WT, as well as defects in odor discrimination learning. Therefore, the loss of 5T4 attenuates inhibitory inputs from 5T4 GCs to nonbursting ETCs and excitatory inputs to 5T4 GCs, contributing to disturbances in olfactory behavior. Our novel findings suggest that, among the various types of OB interneurons, the 5T4 GC subtype is required for odor detection and discrimination behaviors. [ABSTRACT FROM AUTHOR]
- Published
- 2016
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49. Neuropeptide Y mitigates neuropathology and motor deficits in mouse models of Machado-Joseph disease.
- Author
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Duarte-Neves, Joana, Gonçalves, Nélio, Cunha-Santos, Janete, Simões, Ana Teresa, den Dunnen, Wilfred F. A., Hirokazu Hirai, Kügler, Sebastian, Cavadas, Cláudia, and de Almeida, Luís Pereira
- Published
- 2015
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50. Shp2 in Forebrain Neurons Regulates Synaptic Plasticity, Locomotion, and Memory Formation in Mice.
- Author
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Shinya Kusakari, Fumihito Saitow, Yukio Ago, Koji Shibasaki, Miho Sato-Hashimoto, Yasunori Matsuzaki, Takenori Kotani, Yoji Murata, Hirokazu Hirai, Toshio Matsuda, Hidenori Suzuki, Takashi Matozaki, and Hiroshi Ohnishi
- Subjects
NEURONS ,CELLS ,NEUROPLASTICITY ,PHYSIOLOGICAL adaptation ,LABORATORY mice - Abstract
Shp2 (Src homology 2 domain-containing protein tyrosine phosphatase 2) regulates neural cell differentiation. It is also expressed in postmitotic neurons, however, and mutations of Shp2 are associated with clinical syndromes characterized by mental retardation. Here we show that conditional-knockout (cKO) mice lacking Shp2 specifically in postmitotic forebrain neurons manifest abnormal behavior, including hyperactivity. Novelty-induced expression of immediate-early genes and activation of extracellular-signal-regulated kinase (Erk) were attenuated in the cerebral cortex and hippocampus of Shp2 cKO mice, suggestive of reduced neuronal activity. In contrast, ablation of Shp2 enhanced high-K
+ -induced Erk activation in both cultured cortical neurons and synaptosomes, whereas it inhibited that induced by brain-derived growth factor in cultured neurons. Posttetanic potentiation and paired-pulse facilitation were attenuated and enhanced, respectively, in hippocampal slices from Shp2 cKO mice. The mutant mice also manifested transient impairment of memory formation in the Morris water maze. Our data suggest that Shp2 contributes to regulation of Erk activation and synaptic plasticity in postmitotic forebrain neurons and thereby controls locomotor activity and memory formation. [ABSTRACT FROM AUTHOR]- Published
- 2015
- Full Text
- View/download PDF
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