Autism spectrum disorder-like behaviors induced by hyper-glutamatergic NMDA receptor signaling through hypo-serotonergic 5-HT1A receptor signaling in the prefrontal cortex in mice exposed to prenatal valproic acid

Hitomi Kurahashi, Kazuo Kunisawa, Kenji F. Tanaka, Hisayoshi Kubota, Masaya Hasegawa, Mai Miyachi, Yuka Moriya, Yoichi Hasegawa, Taku Nagai, Kuniaki Saito, Toshitaka Nabeshima, Akihiro Mouri

Neuropsychopharmacology 2024年10月11日

Abstract Autism spectrum disorder (ASD) is a neurodevelopmental disorder characterized by repetitive behaviors, social deficits, and cognitive impairments. Maternal use of valproic acid (VPA) during pregnancy is associated with an increased risk of ASD in offspring. The prevailing pathophysiological hypothesis for ASD involves excitation/inhibition (E/I) imbalances and serotonergic dysfunction. Here, we investigated the association between glutamatergic-serotonergic neuronal interactions and ASD-like behaviors in mice exposed to prenatal VPA. Prenatal VPA exposure induced excessive repetitive self-grooming behavior and impaired social behavior and object recognition memory in young adult period. Prenatal VPA mice showed hyper-glutamatergic function (increase in basal extracellular glutamate levels and CaMKII phosphorylation) and hypo-serotonergic function (decrease in 5-hydroxyindoleacetic acid and stimulation-induced serotonin [5-HT] release, but an increase in 5-HT transporter expression) in the prefrontal cortex. Treatment with a low-affinity NMDA receptor antagonist (memantine), a selective 5-HT reuptake inhibitor (fluoxetine), and a 5-HT_1A receptor agonist (tandospirone) attenuated both the increase in CaMKII phosphorylation and ASD-like behavior of prenatal VPA mice. Opto-genetic activation of the serotonergic neuronal system attenuated impairments in social behavior and object recognition memory in prenatal VPA mice. WAY-100635—a 5-HT_1A receptor antagonist—antagonized the effect of fluoxetine on impaired social behavior and object recognition memory. These results suggest that E/I imbalance and ASD-like behavior are associated with hypo-serotonergic receptor signaling through 5-HT_1A receptors in prenatal VPA mice.
Signal flow in the NMDA receptor-dependent phosphoproteome regulates postsynaptic plasticity for aversive learning.

Yasuhiro Funahashi, Rijwan Uddin Ahammad, Xinjian Zhang, Emran Hossen, Masahiro Kawatani, Shinichi Nakamuta, Akira Yoshimi, Minhua Wu, Huanhuan Wang, Mengya Wu, Xu Li, Md Omar Faruk, Md Hasanuzzaman Shohag, You-Hsin Lin, Daisuke Tsuboi, Tomoki Nishioka, Keisuke Kuroda, Mutsuki Amano, Yukihiko Noda, Kiyofumi Yamada, Kenji Sakimura, Taku Nagai, Takayuki Yamashita, Shigeo Uchino, Kozo Kaibuchi

Science signaling 17(853) eado9852 2024年9月10日

Structural plasticity of dendritic spines in the nucleus accumbens (NAc) is crucial for learning from aversive experiences. Activation of NMDA receptors (NMDARs) stimulates Ca2+-dependent signaling that leads to changes in the actin cytoskeleton, mediated by the Rho family of GTPases, resulting in postsynaptic remodeling essential for learning. We investigated how phosphorylation events downstream of NMDAR activation drive the changes in synaptic morphology that underlie aversive learning. Large-scale phosphoproteomic analyses of protein kinase targets in mouse striatal/accumbal slices revealed that NMDAR activation resulted in the phosphorylation of 194 proteins, including RhoA regulators such as ARHGEF2 and ARHGAP21. Phosphorylation of ARHGEF2 by the Ca2+-dependent protein kinase CaMKII enhanced its RhoGEF activity, thereby activating RhoA and its downstream effector Rho-associated kinase (ROCK/Rho-kinase). Further phosphoproteomic analysis identified 221 ROCK targets, including the postsynaptic scaffolding protein SHANK3, which is crucial for its interaction with NMDARs and other postsynaptic scaffolding proteins. ROCK-mediated phosphorylation of SHANK3 in the NAc was essential for spine growth and aversive learning. These findings demonstrate that NMDAR activation initiates a phosphorylation cascade crucial for learning and memory.
Pramipexole Hyperactivates the External Globus Pallidus and Impairs Decision-Making in a Mouse Model of Parkinson’s Disease

Hisayoshi Kubota, Xinzhu Zhou, Xinjian Zhang, Hirohisa Watanabe, Taku Nagai

International Journal of Molecular Sciences 25(16) 8849-8849 2024年8月14日

In patients with Parkinson’s disease (PD), dopamine replacement therapy with dopamine D2/D3 receptor agonists induces impairments in decision-making, including pathological gambling. The neurobiological mechanisms underlying these adverse effects remain elusive. Here, in a mouse model of PD, we investigated the effects of the dopamine D3 receptor (D3R)-preferring agonist pramipexole (PPX) on decision-making. PD model mice were generated using a bilateral injection of the toxin 6-hydroxydopamine into the dorsolateral striatum. Subsequent treatment with PPX increased disadvantageous choices characterized by a high-risk/high-reward in the touchscreen-based Iowa Gambling Task. This effect was blocked by treatment with the selective D3R antagonist PG-01037. In model mice treated with PPX, the number of c-Fos-positive cells was increased in the external globus pallidus (GPe), indicating dysregulation of the indirect pathway in the corticothalamic-basal ganglia circuitry. In accordance, chemogenetic inhibition of the GPe restored normal c-Fos activation and rescued PPX-induced disadvantageous choices. These findings demonstrate that the hyperactivation of GPe neurons in the indirect pathway impairs decision-making in PD model mice. The results provide a candidate mechanism and therapeutic target for pathological gambling observed during D2/D3 receptor pharmacotherapy in PD patients.
KANPHOS: Kinase-associated neural phospho-signaling database for data-driven research

Takayuki Kannon, Satoshi Murashige, Tomoki Nishioka, Mutsuki Amano, Yasuhiro Funahashi, Daisuke Tsuboi, Yukie Yamahashi, Taku Nagai, Kozo Kaibuchi, Junichiro Yoshimoto

Frontiers in Molecular Neuroscience 17 2024年4月2日

Protein phosphorylation, a key regulator of cellular processes, plays a central role in brain function and is implicated in neurological disorders. Information on protein phosphorylation is expected to be a clue for understanding various neuropsychiatric disorders and developing therapeutic strategies. Nonetheless, existing databases lack a specific focus on phosphorylation events in the brain, which are crucial for investigating the downstream pathway regulated by neurotransmitters. To overcome the gap, we have developed a web-based database named “Kinase-Associated Neural PHOspho-Signaling (KANPHOS).” This paper presents the design concept, detailed features, and a series of improvements for KANPHOS. KANPHOS is designed to support data-driven research by fulfilling three key objectives: (1) enabling the search for protein kinases and their substrates related to extracellular signals or diseases; (2) facilitating a consolidated search for information encompassing phosphorylated substrate genes, proteins, mutant mice, diseases, and more; and (3) offering integrated functionalities to support pathway and network analysis. KANPHOS is also equipped with API functionality to interact with external databases and analysis tools, enhancing its utility in data-driven investigations. Those key features represent a critical step toward unraveling the complex landscape of protein phosphorylation in the brain, with implications for elucidating the molecular mechanisms underlying neurological disorders. KANPHOS is freely accessible to all researchers at https://kanphos.jp.
Phenotypes for general behavior, activity, and body temperature in 3q29 deletion model mice

Daisuke Mori, Ryosuke Ikeda, Masahito Sawahata, Sho Yamaguchi, Akiko Kodama, Takashi Hirao, Yuko Arioka, Hiroki Okumura, Chihiro Inami, Toshiaki Suzuki, Yu Hayashi, Hidekazu Kato, Yoshihiro Nawa, Seiko Miyata, Hiroki Kimura, Itaru Kushima, Branko Aleksic, Hiroyuki Mizoguchi, Taku Nagai, Takanobu Nakazawa, Ryota Hashimoto, Kozo Kaibuchi, Kazuhiko Kume, Kiyofumi Yamada, Norio Ozaki

Translational Psychiatry 14(1) 2024年3月7日査読有り

Abstract Whole genome analysis has identified rare copy number variations (CNV) that are strongly involved in the pathogenesis of psychiatric disorders, and 3q29 deletion has been found to have the largest effect size. The 3q29 deletion mice model (3q29-del mice) has been established as a good pathological model for schizophrenia based on phenotypic analysis; however, circadian rhythm and sleep, which are also closely related to neuropsychiatric disorders, have not been investigated. In this study, our aims were to reevaluate the pathogenesis of 3q29-del by recreating model mice and analyzing their behavior and to identify novel new insights into the temporal activity and temperature fluctuations of the mouse model using a recently developed small implantable accelerometer chip, Nano-tag. We generated 3q29-del mice using genome editing technology and reevaluated common behavioral phenotypes. We next implanted Nano-tag in the abdominal cavity of mice for continuous measurements of long-time activity and body temperature. Our model mice exhibited weight loss similar to that of other mice reported previously. A general behavioral battery test in the model mice revealed phenotypes similar to those observed in mouse models of schizophrenia, including increased rearing frequency. Intraperitoneal implantation of Nano-tag, a miniature acceleration sensor, resulted in hypersensitive and rapid increases in the activity and body temperature of 3q29-del mice upon switching to lights-off condition. Similar to the 3q29-del mice reported previously, these mice are a promising model animals for schizophrenia. Successive quantitative analysis may provide results that could help in treating sleep disorders closely associated with neuropsychiatric disorders.

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アメリカにおける医療用麻薬乱用防止製剤の審査—The review system for abuse deterrent opioid formulations in the United States

松﨑哲郎, 奥村啓樹, 永井拓, 山田清文

日本アルコール・薬物医学会雑誌 56(2) 31-38 2021年4月
Shati/Nat8l deficiency disrupts adult neurogenesis and causes attentional impairment through dopaminergic neuronal dysfunction in the dentate gyrus.

Bolati Wulaer, Kazuo Kunisawa, Kazuhiro Hada, Willy Jaya Suento, Hisayoshi Kubota, Tsubasa Iida, Aika Kosuge, Taku Nagai, Kiyofumi Yamada, Atsumi Nitta, Yasuko Yamamoto, Kuniaki Saito, Akihiro Mouri, Toshitaka Nabeshima

Journal of neurochemistry 157(3) 642-655 2020年4月10日

Successful completion of daily activities relies on the ability to select the relevant features of the environment for memory and recall. Disruption to these processes can lead to various disorders, such as attention-deficit hyperactivity disorder (ADHD). Dopamine is a neurotransmitter implicated in the regulation of several processes, including attention. In addition to the higher-order brain function, dopamine is implicated in the regulation of adult neurogenesis. Previously, we generated mice lacking Shati, an N-acetyltransferase-8-like protein on a C57BL/6J genetic background (Shati/Nat8l-/- ). These mice showed a series of changes in the dopamine system and ADHD-like behavioral phenotypes. Therefore, we hypothesized that deficiency of Shati/Nat8l would affect neurogenesis and attentional behavior in mice. We found aberrant morphology of neurons and impaired neurogenesis in the dentate gyrus of Shati/Nat8l-/- mice. Additionally, research has suggested that impaired neurogenesis might be because of the reduction of dopamine in the hippocampus. Galantamine (GAL) attenuated the attentional impairment observed in the object-based attention test via increasing the dopamine release in the hippocampus of Shati/Nat8l-/- mice. The α7 nicotinic acetylcholine receptor antagonist, methyllycaconitine, and dopamine D1 receptor antagonist, SCH23390, blocked the ameliorating effect of GAL on attentional impairment in Shati/Nat8l-/- mice. These results suggest that the ameliorating effect of GAL on Shati/Nat8l-/- attentional impairment is associated with activation of D1 receptors following increased dopamine release in the hippocampus via α7 nicotinic acetylcholine receptor. In summary, Shati/Nat8l is important in both morphogenesis and neurogenesis in the dentate gyrus and attention, possible via modulation of dopaminergic transmission.
Methylation analysis for postpartum depression: a case control study.

Yukako Nakamura, Masahiro Nakatochi, Shohko Kunimoto, Takashi Okada, Branko Aleksic, Miho Toyama, Tomoko Shiino, Mako Morikawa, Aya Yamauchi, Akira Yoshimi, Yoko Furukawa-Hibi, Taku Nagai, Masako Ohara, Chika Kubota, Kiyofumi Yamada, Masahiko Ando, Norio Ozaki

BMC psychiatry 19(1) 190-190 2019年6月20日

BACKGROUND: Postpartum depression (PPD) is a major depressive disorder that occurs after childbirth. Objective diagnostic and predictive methods for PPD are important for early detection and appropriate intervention. DNA methylation has been recognized as a potential biomarker for major depressive disorder. In this study, we used methylation analysis and peripheral blood to search for biomarkers that could to lead to the development a predictive method for PPD. METHODS: Study participants included 36 pregnant women (18 cases and 18 controls determined after childbirth). Genome-wide DNA methylation profiles were obtained by analysis with an Infinium Human Methylation 450BeadChip. The association of DNA methylation status at each DNA methylation site with PPD was assessed using linear regression analysis. We also conducted functional enrichment analysis of PPD using The Database for Annotation, Visualization and Integrated Discovery 6.8 to explore enriched functional-related gene groups for PPD. RESULTS: In the analysis with postpartum depressed state as an independent variable, the difference in methylation frequency between the postpartum non-depressed group and the postpartum depressed group was small, and sites with genome-wide significant differences were not confirmed. After analysis by The Database for Annotation, Visualization and Integrated Discovery 6.8, we revealed four gene ontology terms, including axon guidance, related to postpartum depression. CONCLUSIONS: These findings may help with the development of an objective predictive method for PPD.
Proteomic analysis of lymphoblastoid cell lines from schizophrenic patients.

Akira Yoshimi, Shinnosuke Yamada, Shohko Kunimoto, Branko Aleksic, Akihiro Hirakawa, Mitsuki Ohashi, Yurie Matsumoto, Kazuhiro Hada, Norimichi Itoh, Yuko Arioka, Hiroki Kimura, Itaru Kushima, Yukako Nakamura, Tomoko Shiino, Daisuke Mori, Satoshi Tanaka, Shuko Hamada, Yukihiro Noda, Taku Nagai, Kiyofumi Yamada, Norio Ozaki

Translational psychiatry 9(1) 126-126 2019年4月22日

Although a number of studies have identified several convincing candidate genes or molecules, the pathophysiology of schizophrenia (SCZ) has not been completely elucidated. Therapeutic optimization based on pathophysiology should be performed as early as possible to improve functional outcomes and prognosis; to detect useful biomarkers for SCZ, which reflect pathophysiology and can be utilized for timely diagnosis and effective therapy. To explore biomarkers for SCZ, we employed fluorescence two-dimensional differential gel electrophoresis (2D-DIGE) of lymphoblastoid cell lines (LCLs) (1st sample set: 30 SCZ and 30 CON). Differentially expressed proteins were sequenced by liquid chromatography tandem-mass spectrometry (LC-MS/MS) and identified proteins were confirmed by western blotting (WB) (1st and 2nd sample set: 60 SCZ and 60 CON). Multivariate logistic regression analysis was performed to identify an optimal combination of biomarkers to create a prediction model for SCZ. Twenty protein spots were differentially expressed between SCZ and CON in 2D-DIGE analysis and 22 unique proteins were identified by LC-MS/MS. Differential expression of eight of 22 proteins was confirmed by WB. Among the eight candidate proteins (HSPA4L, MX1, GLRX3, UROD, MAPRE1, TBCB, IGHM, and GART), we successfully constructed logistic regression models comprised of 4- and 6-markers with good discriminative ability between SCZ and CON. In both WB and gene expression analysis of LCL, MX1 showed reproducibly significant associations. Moreover, Mx1 and its related proinflamatory genes (Mx2, Il1b, and Tnf) were also up-regulated in poly I:C-treated mice. Differentially expressed proteins might be associated with molecular pathophysiology of SCZ, including dysregulation of immunological reactions and potentially provide diagnostic and prognostic biomarkers.
【ストレス(モデル)と分子】末梢性免疫炎症反応に伴うアストロサイトにおけるMHC Class I分子の発現誘導が脳機能に及ぼす影響

祖父江顕, 永井拓, 山田清文

ストレス科学 33(3) 233-241 2019年3月

主要組織適合遺伝子複合体クラスI(major histocompatibility complex class I;MHCI)は中枢神経系において主に神経細胞に発現し、スパインの刈り込みなどに関与している。グリア細胞におけるMHCIの病態生理学的役割については不明な点が多いことから、我々は末梢性免疫炎症反応による中枢神経系におけるMHCIの発現変化、ならびに前頭前皮質のアストロサイトにおける膜貫通型MHCI/H-2Dあるいは分泌型MHCI/sH-2Dの発現が周囲の細胞と高次脳機能に及ぼす影響に着目して研究を行ってきた。成熟マウスにpolyriboinosinic-polyribocytidilic acid(polyI:C)を腹腔内投与あるいはIFN-γ遺伝子を尾静脈から導入し、大脳皮質を摘出してMHCIのmRNA量を測定した。コントロール群と比較して、polyI:C処置群では、MHCIのmRNAレベルが有意に上昇し、同様の結果がIFN-γ遺伝子導入マウスでも認められた。MHCIのmRNA発現については神経細胞およびアストロサイトで増加することをin situ hybridization法により確認した。H-2DあるいはsH-2D遺伝子を導入したアストロサイトにおいて、同蛋白質は主にエキソソームに局在し、細胞外へ分泌されることが示唆された。アデノ随伴ウイルスベクターを用いてアストロサイト特異的にH-2DあるいはsH-2Dを前頭前皮質に発現させたマウスではミクログリアの活性化、パルブアルブミン陽性細胞数の減少、スパイン密度の低下などが認められ、社会性行動と物体認知記憶の障害が観察された。これらの障害はエキソソーム合成阻害剤GW4869により改善したことから、アストロサイトにおけるMHCIはエキソソームを介して近傍の細胞に影響し脳機能障害を惹起することが示唆された。(著者抄録)