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ポスター D. 恒常性と神経内分泌システム D. Homeostatic and Neuroendocrine Systems

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-359 Insular responses to 1-Hz transcutaneous auricular vagus nerve stimulation (taVNS) from human intracranial recordings Chan Hui-Ling(1)、Harada Tokiko(1)、Katagiri Masaya (2)、Iida Koji(2,3,4)、Kentaro Ono(1)、 Fermin Alan(1)、Machizawa Maro(1)、Yamawaki Shigeto(1) Hui-Ling Chan(1), Tokiko Harada(1), Masaya Katagiri(2), Koji Iida(2,3,4), Ono Kentaro(1), Alan R. Fermin(1), Maro G. Machizawa(1), Shigeto Yamawaki(1) 1. Center for Brain, Mind and KANSEI Sciences Research, Hiroshima University, Japan, 2. Department of Neurosurgery, Hiroshima University Hospital, Japan, 3. Graduate School of Biomedical and Health Sciences, Hiroshima University, Japan, 4. Epilepsy Center, Hiroshima University Hospital, Japan Keyword: Transcutaneous auricular vagus nerve stimulation (taVNS) , interoception, insula, intracranial electroencephalography (iEEG) Transcutaneous auricular vagus nerve stimulation (taVNS) has been reported to show therapeutic effects for refractory depression and epilepsy. Previous fMRI studies have shown that signals of taVNS are projected to the brain interoceptive network (BIN) via the brainstem. Moreover, several neurophysiological evidence suggest that taVNS modulates interoceptive processing and thought to improve interoceptive accuracy. However, causal neurophysiological mechanism of the taVNS in the BIN has been unclear to date. Here, we directly investigated cortical responses to taVNS using intracranial EEG recording in the left insula, a core region of BIN, of a male patient at a preoperative session. The patient suffered from intractable epilepsy with a subtle lesion suspected as cortical dysplasia around the posterior cingulate cortex. It was hypothesized that (1) active taVNS would elicit larger insular activity than sham taVNS and (2) the neural responses for the active taVNS inputs would propagate from posterior (pINS) to anterior (aINS) insula. Stimulation amplitudes for active and sham stimulations at the tragus and earlobe, respectively, were determined by a randomized staircase method to control the sensation below-median pain level. Each trial consisted of 5-sec resting, 60-sec 1-Hz taVNS with 500-μs pulse width, a 50-sec recovery period and followed by subjective rating of pleasantness. For each ear, a total of 12 trials (6 trials/condition) were conducted in a randomized order. A total of 117 electrode contacts were implanted across temporal, central, and posterior perisylvian regions, including a depth electrode with seven contacts over pINS to aINS. As was expected, compared to the sham condition, active taVNS induced significantly strong activity in pINS during 80-100 ms and aINS after 200 ms under left and right ear stimulations. Time-frequency responses of pINS for the active taVNS showed stronger gamma oscillations and weaker beta oscillations than the sham condition. Furthermore, we observed shift of activation peaks of taVNS-evoked potentials propagating from pINS to middle INS to anterior INS from 100 ms to 130 ms to 180 ms in the active condition. Altogether, our results provide the first direct evidence for the modulatory effect of taVNS on BIN and for the causal neural mechanism underlying our interoception. Thus, taVNS affects interceptive neural processing via pINS and aINS, which may further affect autonomic regulation.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-360 オレキシン受容体拮抗薬スボレキサントが呼吸調節に与える影響 Effects of a Dual Orexin Receptor Blocker Suvorexant on Ventilatory Control 福士勇人(1,2)、横田茂文(3)、武田湖太郎(1,4)、寺田二郎(5)、梅田啓(6)、吉沢雅史(1,7)、河野洋介(1,7)、長谷部洋平(1,7)、鬼丸洋(8)、Pokorski Mieczyslaw(9)、岡田泰昌(1) 1. 独立行政法人国立病院機構村山医療センター臨床研究部、2. 植草学園大学保健医療学部、3. 島根大学医学部医学科解剖学（神経科学）、4. 藤田医科大学保健衛生学部リハビリテーション学科、5. 国際医療福祉大学医学部呼吸器内科学、6. 国際医療福祉大学塩谷病院、7. 山梨大学医学部小児科学、8. 昭和大学医学部生理学、9. Institute of Health Sciences, University of Opole, Opole, Poland Isato Fukushi(1,2), Shigefumi Yokota(3), Kotaro Takeda(1,4), Jiro Terada(5), Akira Umeda(6), Masashi Yoshizawa(1,7), Yosuke Kono(1,7), Yohei Hasebe(1,7), Hiroshi Onimaru(8), Mieczyslaw Pokorski(9), Yasumasa Okada(1) 1. Clinical Research Center, Murayama Medical Center, Musashimurayama, Japan, 2. Faculty of Health Sciences, Uekusa Gakuen University, Chiba, Japan, 3. Department of Anatomy and Neuroscience, Shimane University School of Medicine, Izumo, Japan, 4. Faculty of Rehabilitation, School of Healthcare, Fujita Health University, Toyoake, Japan, 5. Department of Pulmonary Medicine, International University of Health and Welfare, School of Medicine, Narita, Japan, 6. Department of General Medicine, School of Medicine, IUHW Shioya Hospital, International University of Health and Welfare, Yaita, Japan, 7. Department of Pediatrics, Faculty of Medicine, University of Yamanashi, Yamanashi, Japan, 8. Department of Physiology, Showa University School of Medicine, Tokyo, Japan, 9. Institute of Health Sciences, University of Opole, Opole, Poland Keyword: orexin, suvorexant, hypercapnic ventilatory response, hypoxic ventilatory response Objectives: Suvorexant (Belsomra ®), a dual orexin receptor blocker, inhibits the arousal system in the central nervous system providing succor for sleep, which makes it of therapeutic value in insomnia therapy. However, the drug’s ventilatory effects have not been understood. In this study we aimed to investigate the expression of orexin receptors in respiratory neurons and the effects of suvorexant on ventilation. Methods: Immunohistology of brainstem orexin receptor OX2R expression was performed in adult mice (n=4) in (1) rostral ventral respiratory group (rVRG) neurons projecting onto the phrenic nucleus (PhN), (2) neurons immunoreactive for paired like homeobox 2b (Phox2b) in the parafacial respiratory group/retrotrapezoid nucleus (pFRG/RTN), and (3) neurons immunoreactive for neurokinin 1 receptor (NK1R) and somatostatin (SST) in the preBötzinger complex (preBötC). Additionally, we measured in vivo ventilatory responses to hypercapnia (5% CO₂) and hypoxia (10% O₂) before and after suvorexant pretreatment (10 and cumulative 100 mg/kg) in unrestrained mice (n=10) by whole body plethysmography. Results: We found the OX2R immunoreactive materials in pFRG/RTN Phox2b and preBötC NK1R/SST immunoreactive neurons, which suggests the involvement of orexin in respiratory control. Further, suvorexant expressly suppressed the hypercapnic ventilatory augmentation, otherwise unaffecting ventilation. Conclusion: We conclude that suvorexant inhibits hypercapnic ventilatory augmentation. However, either dose of suvorexant did not affect ventilation in room air or hypoxic ventilatory responses. Suvorexant may have to be carefully used in patients with hypercapnic respiratory failure or sleep related hypoventilation.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-361 橋排尿中枢バリントン核から脊髄に対する下行性投射の光遺伝学的検討 Downstream projection of Barrington’s nucleus to the spinal cord in mice 河谷昌泰(1,2,3,4)、William Chet de Groat(5)、井樋慶一(6)、内田克哉(6)、崎村建司(7)、山中章弘(1,2)、山下貴之(4)、河谷正仁(3) 1. 名古屋大学環境医学研究所、2. 名古屋大学医学系研究科、3. 秋田大学医学系研究科、4. 藤田医科大学医学部、5. ピッツバーグ大学、6. 東北大学医学系研究科、7. 新潟大学脳研究所 Masahiro Kawatani(1,2,3,4), William Chet de Groat(5), Keiichi Itoi(6), Katsuya Uchida(6), Kenji Sakimura(7), Akihiro Yamanaka(1,2), Takayuki Yamashita(4), Masahito Kawatani(3) 1. Research Institute of Environmental Medicine, Nagoya University, Nagoya, Japan, 2. Graduate School of Medicine, Nagoya University, Nagoya, Japan, 3. Graduate School of Medicine, Akita University, Akita, Japan, 4. School of Medicine, Fujita Health University, Toyoake, Japan., 5. University of Pittsburgh, Pittsburgh, Pennsylvania., 6. Graduate School of Medicine, Tohoku University, Sendai, Japan, 7. Brain Research Institute, Niigata University, Niigata, Japan Keyword: Sacral para-sympathetic nucleus, Pontin micturition center/ Barrington's nucleus, Spinal cord, Autonomic nervous systems Barrington’s nucleus (Bar) controls the micturition reflex (rhythmic bladder contraction and sphincter relaxation) through downstream projection to the spinal cord (SC). However, the synaptic organization of the Bar→SC projection is poorly understood. In the Bar, two glutamatergic neuronal populations, which express corticotrophin-releasing hormone (Bar^CRH) or estrogen receptor alpha (Bar^ESR1), are thought to provide functionally distinct descending signals to the SC. In this study, using Cre-expressing mouse lines and viral injections, we specifically introduced the expression of channelrhodopsin 2 in Bar^CRHor Bar^ESR1 neurons and analyzed synaptic responses evoked by photo-stimulation in lumbosacral (LS) spinal slices. In the lumbosacral intermediolateral region (LS-IML), photo-stimulation of both Bar^CRH and Bar^ESR1 axons evoked excitatory postsynaptic currents (EPSCs) in DiI-labeled preganglionic neurons (LS-PGNs), which were abolished by bath-application of an AMPA receptor antagonist. In LS-PGNs, the amplitudes of Bar^ESR1-evoked EPSCs were significantly larger than those of Bar^CRH-evoked EPSCs. All Bar^CRH-LS-PGN EPSCs were monosynaptic, whereas a fraction (27%) of Bar^ESR1-LS-PGN EPSCs were non-monosynaptic (i.e., polysynaptic) responses. Furthermore, photo-stimulation of Bar^ESR1, but not Bar^CRH, axons additionally elicited polysynaptic inhibitory postsynaptic currents (IPSCs) in a subset of LS-PGNs. In the lumbosacral dorsal commissure nucleus (LS-DCN), photo-stimulation of both Bar^CRH and Bar^ESR1 axons similarly elicited monosynaptic EPSCs and polysynaptic IPSCs. These results suggest that excitatory signals of Bar^ESR1 have more extensive impacts on the LS-PGNs than those of Bar^CRH, but not on the LS-DCN neurons. In addition, capsaicin-induced excitation of sensory afferent inputs did not alter photo-evoked EPSCs in LS-PGNs, yet augmented their photo-evoked firings through baseline depolarization by increasing spontaneous EPSCs. Thus, both Bar^CRH and Bar^ESR1 directly innervate LS-PGNs and LS-DCN neurons, which operate independently of afferent inputs, enabling integration of descending and ascending signals by the SC circuits.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-362 ウェアラブル端末による簡易心拍解析とその検証 Simple Heart Rate Analysis and Verification using a Wearable Device 藤江博幸(1)、田中靖人(1,2,3) 1. 株式会社　三城ホールディングス R&D、2. 神経数理学研究所、3. 大阪大学 Hiroyuki Fujie(1), Yasuto Tanaka(1,2,3) 1. Paris Miki Holdings Inc. R&D, 2. Neuromathematics Laboratory, 3. Osaka University Keyword: ECG, Chaos Analysis, RR Interval, LF/HF Ratio, Lyapunov index With the spread of mobile terminals and wearable device linked to them, heart rate, blood oxygen concentration, pulse rate, etc. can be easily recorded. Mobile terminals made by the same company are used to link data with the wearable device used for this measurement, and systems such as Android and iOS are mainly used. Data check, simple analysis, and advice will be performed by the mobile terminal. In addition, application development on mobile terminals enables some specialized advice and data transfer by its network connection enables more detailed analysis and advice by the specialist of cardiology doctor. In this verification, simple ECG data acquisition and analysis were performed by Apple's apple-Watch (512Hz) and iPhone, and the results were compared with the ECG measurement data acquisition and analysis by Nihon Kohden's EEG1200 (1000Hz) at the same time and same objects. Since the ECG measurement time set on the wearable device is 30 seconds, we decided to measure three times in a row and combine them to obtain 90 seconds of data and use it enough for our analysis method. Since the PDF image is displayed on the mobile terminal transffered from wearable device, the data was generated extracted from this PDF image and RRI(RR Interval)-LF / HF analysis, RRI-chaos analysis, RRI-Lyapunov index and ECG-chaos analysis were performed using by our original script of MATLAB. This analysis method is based on the detailed analysis of heartbeat components obtained from the data of ultra fine eye movements experiment that we have traditionally performed. We found the chaos components using such wearable device which is relevant with the data obtained with traditional ECG signals. And a similarities of chaos characteristic were compared and discussed.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-363 外側手綱核の興奮は中脳モノアミン領域を介して心血管応答を惹起する Midbrain monoaminergic areas are involved in the cardiovascular response induced by the activation of the lateral habenula in rats 佐藤優真(1,2)、Tri Huu Doan(1)、松本正幸(3,4)、小金澤禎史(1,4) 1. 筑波大・医学医療系・神経生理学、2. 筑波大院・人間総合科学・医学、3. 筑波大・医学医療系・認知行動神経科学、4. 筑波大・トランスボーダー医学研究センター Yuma Sato(1,2), Tri Huu Doan(1), Masayuki Matsumoto(3,4), Tadachika Koganezawa(1,4) 1. Dept Neurophysiol, Fac Med, Univ Tsukuba, Tsukuba, Ibaraki, Japan, 2. Dr Prog Biomed Sci, Grad Sch Comp Human Sci, Univ Tsukuba, Ibaraki, Japan, 3. Dept Cogn Bahav Neurosci, Fac Med, Univ Tsukuba, Tsukuba, Ibaraki, Japan, 4. Transborder Med Res Ctr, Univ Tsukuba, Ibaraki, Japan Keyword: habenula, cardiovascular function, dopamine, serotonin The lateral habenula (LHb) plays a crucial role in behavioral responses to cope with stressful events. Neurons in the LHb descend to the midbrain monoaminergic system, such as the dopaminergic and serotonergic systems. The monoaminergic system is known as one of the components involved in neural processes related to stress. Although neurons in midbrain monoaminergic areas have anatomical connections to the autonomic cardiovascular center in the medulla, little is known to about the neural circuit that is originated from the LHb and mediated by the monoaminergic system in cardiovascular regulation. To address this issue, we examined the mediateing brain area, which includes the dopaminergic and serotonergic systems, in the cardiovascular response induced by the activation of the LHb. In this study, we used Wistar male rats (250 ~ 330 g) anesthetized by urethane (1 ~ 1.25 g/kg, i.p.). Heart rate (HR) was counted from the electrocardiography of rats. Mean arterial pressure (MAP) was also recorded from a catheter inserted into the femoral artery. We performed electrical stimulations (300 µA, 0.5 ms duration, 100 Hz, for 10 s) to the LHb with clozapine (1 mg/kg, i.v.), a nonselective dopamine receptor antagonist, or methysergide (1 mg/kg, i.v.), a serotonin receptor antagonist with the observation of HR and MAP. Electrical stimulation to the LHb caused a decrease in HR and an increase in MAP. Moreover, the administration of clozapine or methysergide attenuated both bradycardia and a pressor response. We next tried to identify the midbrain monoaminergic area, which participates in the neural system from the LHb to the cardiovascular center in the medulla. The LHb has neural connections with the midbrain monoaminergic areas. Therefore, we performed the electrical stimulation to the LHb with infusion of muscimol, GABAA receptor agonist, into the ventrolateral periaqueductal gray (vlPAG) and the dorsal raphe nuclei (DRN) to inactivate each brain area, which contains the monoaminergic neurons. Inactivation of the vlPAG caused an enhancement of bradycardia, but a pressor response didn’t change. On the other hand, the pressor response was attenuated by the inactivation of the DRN. These results indicated that dopamine neurons in the vlPAG may inhibitorily regulate the HR response induced by the activation of the LHb, and serotonin neurons in the DRN may excitatorily modulate the MAP response.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-364 全身周回性の神経回路を介した腸の脂質代謝が線虫Caenorhabditis elegansの温度馴化を制御する Intestinal lipid metabolism via whole body circulatory neural circuit regulates temperature acclimation in Caenorhabditis elegans 太田茜(1)、本村晴佳(1)、村上一寿(1)、五百藏誠(1)、久原篤(1,2) 1. 甲南大学統合ニューロバイオロジー研究所、2. PRIME, AMED Akane Ohta(1), Haruka Motomura(1), Kazutoshi Motomura(1), Makoto Ioroi(1), Atsushi Kuhara(1,2) 1. Konan University, 2. PRIME, AMED Keyword: C. elegans, temperature acclimation, whole body circuit, brain-gut coupling Environmental temperature is a dynamic environmental factor that constantly affects survive and physiology of animal. The nematode C. elegans changes its cold tolerance in response to environmental temperature-changes within only three hours, which is called temperature acclimation. Three pairs of sensory neurons in the C. elegans head, ASJ, ADL, and ASG, have been shown to sense temperature and involved in temperature acclimation (Ohta et al., *Nature commun, 2014; Okahata et al., Science advances, 2019; Takagaki et al., EMBO rep, 2020), however, the downstream neural circuit of the three pairs neurons were unknown. We show here that cell-specific rescue analysis of CREB mutants with abnormal temperature acclimation suggested the involvement of the tail interneuron PVQ and its downstream head hub interneuron RMG in temperature acclimation. Ca²⁺ imaging revealed that ASJ activates PVQ in a temperature-stimulation-dependent manner, and PVQ activates RMG through glutamate signaling. These indicate that a whole-body neural circuit containing ASJ-PVQ-RMG is required for temperature acclimation (Motomura, Ohta et al., under review). 　To identify downstream signaling of the whole-body neural circuit, we measured temperature acclimation of many neuro-peptide and their receptor mutants. As a result, mutations in a neuropeptide-like molecule FLP and its receptor showed severe abnormalities in temperature acclimation. This FLP receptor is expressed in intestine, and intestinal fatty acid metabolism via a lipase for the lipid Triglyceride (TG) were decreased in the FLP and its receptor mutants. We quantitatively measured intestinal fat storage of these mutants by staining a dye, oil Red O. The gut fat storage were changed depending on the cultivation temperature, and FLP and its receptor mutants showed abnormality in the phenomena. Together with these results, a whole body circulatory neural circuit containing ASJ-PVQ-RMG releases a neuropeptide FLP that is received by the intestine, which alters gut fat storage, resulting on changes in cold tolerance depending on cultivation temperature. Namely, a new experimental system of brain-gut coupling was proposed (Motomura, Ohta et al., under review*).

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-365 脳海馬が合成する男性・女性ホルモンは記憶シナプスをノンゲノミックに制御する Hippocampus-synthesized androgen and estrogen non-genomically modulate synaptic plasticity 相馬ミカ(1,2)、池田真理(1,2)、髙須絢子(1)、斎藤稔(1)、川戸佳(1,2) 1. 日本大学文理学部生命科学科、2. 順天堂大学大学院医学研究科泌尿器外科 Mika Soma(1,2), Mari Ikeda(1,2), Ayako Takasu(1), Minoru Saito(1), Suguru Kawato(1,2) 1. Dept of Biosciences, Col of Humanities and Sciences, Nihon Univ, Tokyo, Japan, 2. Dept of Urology, Juntendo Univ, Tokyo, Japan Keyword: ANDROGEN AND ESTROGEN, KINASE, DENDRITIC SPINE, LTP We observed that sex-steroids are rapidly synthesized (within 30 min) in the hippocampal glutamatergic neurons (Kimoto et al., 2001; Kawato et al., 2002; Hojo et al., 2004, 2009). We have investigated rapid and non-genomical functions of hippocampus-synthesized androgen and estrogen. By using super-resolution confocal microscopy and Spiso-3D image analysis software (Mukai et al., 2011), we demonstrated that testosterone (T), dihydrotestosterone (DHT) and estradiol (E2) rapidly increased (within 2 h) hippocampal dendritic spines (= postsynapses) in isolated hippocampal slices. Signaling pathways include synaptic (membrane bound) AR (androgen receptor), mER (estrogen receptor), kinase networks (LIMK, MAPK, PKA, PKC) and actin-binding proteins (cofilin, cortactin), resulting in spine increase. Selective inhibitors of these kinases suppressed the spine increase by T, DHT and E2. By using multi-electrode electrophysiology, we demonstrated that E2 induced the long-term potentiation (E2-LTP) of hippocampal neurons. E2 rapidly drives PKA, PKC, and MAPK through synaptic (membrane bound) ER. These kinases may phosphorylate NR2B subunit of NMDA receptor, resulting in induction of LTP. Synaptic AR and ER are membrane bound via palmitic acid. Rapid kinase-dependent signaling is non-genomic, and is completely different from slow genomic signaling which includes translocation of nuclear steroid receptors (AR, ER) into nucleus and protein synthesis. Detailed results are described in (Tozzi et al., 2019; Soma et al., 2018; Hojo and Kawato, 2018; Murakami et al., 2018; Hatanaka et al., 2015; Hasegawa et al., 2015; Kato et al., 2013, 2020).		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-366 ゼブラフィッシュ成魚視床下部における神経ペプチド発現アトラスー神経亜核の提案と哺乳類との比較ー Neuropeptide expression atlas in the adult zebrafish hypothalamus – definition of subnuclei and comparison with mammals – 梶山十和子(1)、吉原良浩(1) 1. 理化学研究所 Towako Kajiyama(1), Yoshihiro Yoshihara(1) 1. RIKEN Keyword: Neuropeptide, Gene expression atlas, Zebrafish, Comparative neuroanatomy Zebrafish has become one of the most useful model animals in neuroscience. However, gene expression atlas in the adult brain has not yet been developed for zebrafish. In this study, 36 neuropeptide genes and 2 neurotransmitter marker genes were comprehensively mapped in the zebrafish hypothalamus by in situ hybridization and assembled into an atlas. Based on the obtained results, we conducted two detailed analyses as follows. 1) Definition of subnuclei. In the most popularly used zebrafish brain atlas (Wulliman et al., 1996), the hypothalamus (including the preoptic area) is divided into nine nuclei. However, the expression patterns of neuropeptide genes revealed that several nuclei can be divided into multiple sub-regions with significantly different sets of neuropeptide gene expression. For example, the nucleus Hv (ventral zone of the periventricular hypothalamus) can be clearly divided into two subnuclei. Many neuropeptides (agrp, asip2b, cart2, cart3, ccka, crha, crhb, gnrh3, npb, penka, penkb, pomcb, vipa) are expressed only in the ventral part of Hv (vHv), whereas several neuropeptides (pdyn, tac1, tac3a, tac3b) are expressed only in the dorsal part of Hv (dHv). Therefore, we propose vHv and dHv as the subnuclei in Hv. 2) Comparison with mammals. Although anatomical, molecular, and functional homologies of brain nuclei between teleost fish and mammals have been reported previously, only little is known about comparative neuroanatomy of the hypothalamus. We compared our neuropeptide expression data in the zebrafish with those of mice that are available in Allen Mouse Brain Atlas and other literatures. We found that the expression patterns of several neuropeptides are similar in some particular nuclei in both zebrafish and mice. The similarities in the neuropeptide expression patterns in the nuclei suggest that their functions may also be closely related. The comprehensive expression atlas and detailed neuroanatomical analyses in this study will contribute to future neuroscience research using zebrafish.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-367 授乳期の野生型マウスにおけるオキシトシンパルスの測定を実現するミニプロモーター駆動型AAV Recording of pulsatile activities of Oxytocin neurons in lactating wild-type mice via a minipromoter driven AAV vector 矢口花紗音(1,2)、幸長弘子(1)、今野歩(3)、平井宏和(3)、宮道和成(1) 1. 理化学研究所生命機能科学研究センター、2. 京都大学大学院生命科学研究科、3. 群馬大学大学院医学系研究科 Kasane Yaguchi(1,2), Hiroko Yukinaga(1), Ayumu Konno(3), Hirokazu Hirai(3), Kazunari Miyamichi(1) 1. RIKEN Center for Biosystems Dynamics Research, 2. Graduate School of BIOSTUDIES, Kyoto University, Kyoto, Japan, 3. Graduate School of Medicine, Gunma University, Gunma, Japan Keyword: Oxytocin, Hypothalamus, Paraventricular hypothalamic nucleus, Lactation Successful breastfeeding is essential for the survival of mammals. The pulsatile release of the neural hormone oxytocin (OT) mediates milk letdown during lactation, which is regulated by the coincident activities of the central OT neurons located in the paraventricular and supraoptic nucleus of the hypothalamus. Molecular and neural circuit mechanisms by which these pulsatile activities of OT neurons are shaped remain mostly elusive. As a first step to address this issue, our recent study established fiber-photometry-based chronic in vivo Ca²⁺ imaging of pulsatile activities from genetically-defined OT neurons in parturient and lactating mother mice by using OT-Cre crossed with a Cre-dependent GCaMP6s driver (Yukinaga H et al. bioRxiv, 2021.2007.2026.453888). However, the necessity of Cre-based double transgenic mice in this method not only limited the throughput of the data collection but also compromised the use of various Cre-dependent toolkits for dissecting molecular and neural circuit functions. To overcome this difficulty, a simple Cre-free method for monitoring OT neural activities is awaited. Here we show that an AAV vector driving GCaMP6s under a 2.6 kb mouse OT promotor is sufficient to selectively monitor pulsatile activities of OT neurons in lactating wild-type mice. By using this method, we illuminated the temporal dynamics of pulsatile activities of OT neurons in wild-type mice throughout the lactation period. Our strategy can be applied to characterize lactation of various wild-type mouse strains and genetic mutants. By Cre-based viral-genetic toolkits, we expect to facilitate studies to dissect neural circuits underpinning pulsatile activities of OT neurons. How is the coincident activation of OT neurons achieved during the milk-ejection reflex? A prevailing view assumes a paracrine-like positive feedback mechanism utilizing OT receptor (OTR) expressed by the OT neurons; once a certain number of OT neurons fire, the OT ligands released from the cell bodies and dendrites activate other OT neurons via Gq-coupled OTR. However, our ultrasensitive histochemical assay failed to detect any OTR mRNA in OT neurons of lactating mother mice. Moreover, we found that OT neuron-specific OTR conditional knockout (cKO) mothers show no abnormalities in lactation. The function of OTR in shaping pulsatile activities of OT neurons will be discussed with special respect to monitoring activities of OT neurons in OTR cKO mother mice.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-368 新規バイオセンサーを用いた神経内分泌細胞の発火パターンとコルチコトロピン放出ホルモン放出の関係解明 Understanding the relationship between neural activity patterns and the release of corticotropin releasing hormone using novel biosensors. 五十嵐敬幸(1)、Rithwik Ramachandran(1)、岡本賢一(2)、Julio Martinez-Trujillo(1)、井上渉(1) Hiroyuki Igarashi(1), Rithwik Ramachandran(1), Kenichi Okamoto(2), Julio Martinez-Trujillo(1), Wataru Inoue(1) 1. Dept of Physiol and Pharmacol, Western Univ, London, Canada, 2. Dept of Molecular Genetics, Univ of Toronto, Toronto, Canada Keyword: Neuroendocrine stress response, CRH neuron, Excitation secretion coupling, Sniffer cell The neuroendocrine stress response is mediated by the release of corticotropin-releasing hormone (CRH) from axons terminating at the median eminence. However, the relationship between the neural activities and the neuroendocrine CRH release is poorly understood because of the lack of sensitive tools to detect small amount of endogenous CRH at the median eminence. Here, we developed a novel "sniffer cell-line" which can be seeded onto acute brain slices and detect endogenous CRH release. CRH sniffer cells stably express a combination of the human CRH receptor 1 (CRHR1) and Epac-based single polypeptide FRET reporter (CEY) which reports cAMP levels. Upon ligand binding, CRHR1 activates adenylyl cyclase and cAMP production, which leads to the change of FRET signal of CEY in clonally expanded sniffer cells. Fitting of a concentration-effect curve estimated an EC50 of 451 pM. When seeded on acute brain slices containing the median eminence, these cells responded to the endogenous CRH release triggered by the electrical stimulations. To study the electrical stimulation-release relationship with relevance to stress hormone response, we used two distinct modes of firing patterns adopted from our recent in vivo single-unit recordings of CRH neurons in the paraventricular nucleus of the hypothalamus. Specifically, under non-stress conditions, CRH neurons generated rhythmic brief bursts (RB) consisted of 2-5 spikes around 200Hz. The RB firing mode constrains the overall firing rate at low levels because of long (>1 s) and mostly silent inter-burst intervals. On the other hand, stress rapidly switched firing patterns from RB to single spiking (SS) persisted around 10Hz. Importantly, this SS firing mode permits sustained spiking that lasts for several seconds, and consequently increases the number of spikes. In acute slices, GCaMP6s Ca²⁺ imaging focusing at CRH neuron’s axon terminals at the median eminence showed significantly larger response to the electrical stimulation mimicking the SS over the RB. In line with the Ca²⁺ imaging results, we found that the SS induced greater CRH release than the RB pattern. This highly sensitive reporter system optimized for ex vivo live imaging enables us to study the activity-dependent secretion mechanism of CRH in situ.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-369 Ultramicronized palmitoylethanolamide counteracts neuroinflammation in obese mice: the role of PPAR-alpha Lama Adriano(1)、Pirozzi Claudio(1)、Severi Ilenia(3)、Annunziata Chiara(1)、Comella Federica(1)、Del Piano Filomena(2)、Senzacqua Martina(3)、Melini Stefania(1)、Opallo Nicola(1)、Meli Rosaria(1)、Giordano Antonio(3)、Mattace Raso Giuseppina(1) Adriano Lama(1), Claudio Pirozzi(1), Ilenia Severi(3), Chiara Annunziata(1), Federica Comella(1), Filomena Del Piano(2), Martina Senzacqua(3), Stefania Melini(1), Nicola Opallo(1), Rosaria Meli(1), Antonio Giordano(3), Giuseppina Mattace Raso(1) 1. Department of Pharmacy, University of Naples Federico II, Naples, Italy, 2. Department of Veterinary Medicine and Animal Productions, University of Naples Federico II, Naples, Italy, 3. Department of Experimental and Clinical Medicine, Marche Polytechnic University, Ancona, Italy Keyword: Inflammation, Obesity, N-acylethanolamines, Central disorders The high-fat diet (HFD) consumption impacts on brain function by different mechanisms involving the gut-brain axis. Notably, metabolic inflammation induced by HFD overnutrition contributes to neuroinflammation, indicating a strict interplay between periphery and CNS in the pathogenic mechanisms of obesity-related central disorders. The excessive fatty acids reach the hypothalamus which is the managing centre of food intake. Its alteration induces the impairment of all related neuronal circuitries of other brain area (i.e. hippocampus and amygdala) involved in the regulation of mood. Palmitoylethanolamide (PEA) is an endogenous peroxisome proliferator-activated receptor (PPAR)-α agonist, whose metabolic and anti-depressive effects are well-known. This study aimed to elucidate the PEA effect in limiting obesity-induced neuroinflammation, blood-brain barrier (BBB) disruption, and brain immune cell involvement in mice fed an HFD. Male C57Bl/6J (6 weeks of age) mice were divided into 3 groups: control receiving standard diet (STD), HFD and HFD orally treated with ultramicronized PEA (PEA-um, 30 mg/kg/die). The treatment started after 12 weeks of HFD feeding and lasted 7 weeks, along with HFD. To gain mechanistic insight, we performed in vitro experiments on human neuroblastoma SH-SY5Y cells insulted by a mix of glucosamine and glucose and treated with PEA-um and the antagonist of PPAR-α, GW6471. Here, inflammatory pathways and bioenergetic parameters (i.e., mitochondrial stress test) by Seahorse Analyser were determined. PEA-um limited the overall inflammatory state, reducing serum and central pro-inflammatory mediators and astrogliosis and microgliosis in both hypothalamus and hippocampus. The simultaneous reduction of hippocampal mast cell markers (chymase 1 and tryptase β2) indicated the attenuation of immune cell activation underlying the neuroprotective effect of PEA-um. HFD-induced neuroinflammation was also associated with the disruption of BBB. PEA-um limited the albumin extravasation and restored tight junction transcription of obese mice. In a neuronal model of metabolic injury, PEA directly counteracted inflammation and mitochondrial dysfunction in a PPAR-α-dependent manner since receptor antagonist GW6471 blunted its effects. Our results strengthen the therapeutic potential of PEA in obesity-related central comorbidities, managing neuroinflammation, BBB disruption, and immune alterations.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-370 Divergent electrophysiological and morphological properties of hypothalamic paraventricular nucleus neurons between the common marmoset and the mouse Julia Sunstrum(1,2), Sam Mestern(1,2), Rebecca Przy(2), Yihang Qin(2), Stefan Everling(2,3), Julio Martinez-Trujillo(2,3), Wataru Inoue(2,3) 1. Neuroscience Graduate Program, Schulich School of Medicine and Dentistry, Western Univ, London, Ontario, Canada , 2. Robarts Research Institute, London, Ontario, Canada, 3. Dept of Physiology and Pharmacology, Western Univ, London, Ontario, Canada Keyword:* Paraventricular nucleus, marmoset, HCN The release of stress hormones via the hypothalamic-pituitary-adrenal (HPA) axis is thought to be preserved across mammals. The apex of the HPA axis is formed by parvocellular neuroendocrine neurons in the paraventricular nucleus of the hypothalamus (PVN) that release corticotropin releasing hormone (CRH). While PVN-CRH neurons have been extensively studied in rodents, studies in primates remain scarce. It is possible that the diurnal activity cycle and complex social lives of primates have exposed them to different stressors, and PVN-CRH neurons have evolved new features under evolutionary pressures. In this exploratory study, we compared PVN neurons between mice and the common marmoset (Callithrix jacchus), a New World primate that has emerged as a promising model in neuroscience. Using patch clamp electrophysiology in acute brain slices, combined with post-hoc morphology reconstruction and immunohistochemistry, we characterized marmoset PVN neurons (N = 14, n= 364) and compared them to their mouse counterparts (N = 24, n = 184). Marmoset PVN parvocellular neurons showed stereotyped electrophysiological features well-established in rodents and were distinct from other PVN neuron-types. However, marmoset PVN parvocellular neurons exhibited substantially larger sag-current and shifted membrane resonance due to hyperpolarization-activated cyclic nucleotide-gated (HCN) channels. Interestingly, female (N = 7) parvocellular neurons exhibited greater voltage sag compared to male (N=6) in marmosets but not mice. Further, morphological analysis revealed that marmoset PVN parvocellular neurons exhibit more dendritic spines compared to mice. These distinct features of marmoset PVN parvocellular neurons may reflect fundamental differences in synaptic integration adaptive for the lifestyle and stressors of primates.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-371 養子移入による視神経脊髄炎関連疾患（NMOSD）の新規動物モデル A new adoptive transfer model for neuromyelitis optica spectrum disorders (NMOSD) 阿部陽一郎(1)、大松澤知加(1,2)、チャウサイモン(1)、宮坂佳樹(3)、廣瀬美嘉子(2)、麻野珠都(2)、真下知士(4)、三澤日出巳(2)、安井正人(1) 1. 慶應義塾大学医学部、2. 慶應義塾大学薬学部、3. 大阪大学医学部、4. 東京大学医科研 Yoichiro Abe(1), Chika Omatsuzawa(1,2), Simon Chau(1), Yoshiki Miyasaka(3), Mikako Hirose(2), Mito Asano(2), Tomoji Mashimo(4), Hidemi Misawa(2), Masato Yasui(1) 1. Sch Med, Keio Univ, Tokyo, Japan, 2. Faculty of Pharm, Keio Univ, Tokyo, Japan, 3. Osaka Univ, Med Sch, Osaka, Japan, 4. Inst Med Sci, Univ of Tokyo, Tokyo, Japan Keyword: neuromyelitis optica spectrum disorder, aquaporin-4, autoantibody, disease model Neuromyelitis optica spectrum disorder (NMOSD) is an inflammatory autoimmune disease of the central nervous system characterized by the existence of an autoantibody called NMO-IgG that binds to the extracellular domains of aquaporin-4 (AQP4), a water channel densely expressed in the astrocytic end-feet, in patients’ plasma. Biding of NMO-IgG to AQP4 causes complement-dependent disruption of astrocytes, which leads to demyelination with an unknown mechanism. Recently, complement-independent cytotoxic effects of NMO-IgG have also been suggested. To understand detailed pathogenesis of NMOSD as well as to develop new therapeutic drugs for NMOSD, animal models that faithfully reproduce the disease are required. One problem is that there are three amino-acid substitutions in the extracellular domains of AQP4 between humans and rats and this difference diminishes the binding of some NMO-IgGs to rat AQP4. Thus, it is difficult to produce NMOSD-like pathology in rodents using NMO-IgG. To overcome the problem, we established a new rat model for NMOSD. First, we generated “AQP4-humanized” rats by replacing the extracellular domains with those of human AQP4 as well as AQP4 knockout rats by deleting 295 nucleotides from exon 1 of the rat AQP4 gene using a CRISPR/Cas9 genome editing technology. We also produced recombinant rat AQP4 protein having the extracellular domains of human AQP4 (humanized rat AQP4) using a baculovirus-insect cell expression system. We immunized AQP4 knockout rats with humanized rat AQP4 protein. Eleven days after immunization, we detected antibodies against human AQP4 in the immunized rats as determined by ELISA. Then we collected lymphocytes from the immunized rats and transferred them to “AQP4-humanized” rats. Approximately six days after the lymphocyte transfer, the recipient rats started to show tail weakness and the clinical score became worse during next several days. Histopathologically, loss of markers for astrocytes in the brains and spinal cord was observed. Serum anti-AQP4 antibodies were still elevated in the recipient rats and sometimes showed higher titers as compared with that in donor rats, indicating that the AQP4-specific T and B cells survived and were continuously activated in the recipient rats. This is the first rat model showing NMOSD-like pathology triggered only by immune response to human AQP4 and will contribute to develop new therapeutic strategies for NMOSD.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-372 脈絡叢生体内イメージングで辺縁系脳炎の炎症波及をリアルタイムに観察する Real-time observation of inflammation spreads to limbic encephalitis by the choroid plexus imaging in vivo 進藤麻理子(1,2)、辻貴宏(1)、加藤大輔(1,2)、和氣弘明(1,2) 1. 名古屋大学大学院医学系研究科、2. 生理学研究所　多細胞回路動態研究部門 Mariko Shindo(1,2), Takahiro Tsuji(1), Daisuke Kato(1,2), Hiroaki Wake(1,2) 1. Grad Sch Med, Nagoya Univ , Aichi, Japan, 2. Division of Multicellular Circuit Dynamics, NIPS, Aichi, Japan Keyword: choroid plexus, two-photon imaging Systemic lupus erythematosus (SLE) is an autoimmune disease characterized chronic systemic inflammation caused by immune activation. Patients with SLE often present neuropsychiatric symptoms such as cognitive dysfunction and mood disorders, referred to as neuropsychiatric SLE (NPSLE). SLE patients also expressed inflammatory CSF (increased lymphocytes and IL-6 levels) and limbic encephalitis, and it has been supposed that inflammation spills over from the CSF into areas anatomically adjacent to the ventricles, causing neuronal damage. On the other hand, the pathological mechanism that cause SLE-induced systemic inflammation from the circulatory system to the CSF and limbic system remains unclear. We hypothesized that the choroid plexus, a spinal fluid-producing organ, plays a role in the spillover of systemic inflammation to the limbic system, and analyzed the pathway of inflammation in the NPSLE model. Behavioral tests (open field test, Y-maze) were performed on various SLE models at sequential time points to identify models with behavioral abnormalities. These SLE models with behavioral abnormalities were used as NPSLE models for subsequent functional analysis. In the NPSLE model, we found increased lymphocyte numbers in the limbic system and choroid plexus. In order to visualize inflammation spreading through the choroid plexus, we have established a technique for in vivo imaging of the choroid plexus in living mice using a two-photon microscopy. This novel technique allowed us to detect the behavior of macrophages and T cells, and changes in vascular permeability in the NPSLE model. Our data support the involvement of the choroid plexus in inflammatory spillover in the NPSLE model. Identification of the pathways of inflammatory spillover may be useful in developing specific therapies and elucidating pathogenesis. In the future, the function of the choroid plexus in inflammatory spreading will be evaluated by modifying choroid plexus function.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-373 様々な食餌条件下での性成熟期メスマウスのキスペプチンニューロン活動動態 Dynamics of kisspeptin neuronal activities in the pubertal female mice under various food conditions 後藤哲平(1)、阿部高也(1)、清成寛(1)、宮道和成(1) 1. 理化学研究所生命機能科学研究センター Teppei Goto(1), Takaya Abe(1), Hiroshi Kiyonari(1), Kazunari Miyamichi(1) 1. RIKEN BDR Keyword: Kisspeptin, fiber photometry, Puberty, Agrp Mammalian females reach puberty when their body weights exceed a certain threshold with sufficient nutrition from the diet. It is known that food restriction (FR) during the pubertal period impairs sexual maturation in rodent models, which is reversibly restored if FR is removed. After sexual maturation, the secretion of gonadotropin is primarily controlled by kisspeptin neurons in the hypothalamic arcuate nucleus. However, the activity dynamics of kisspeptin neurons during the pubertal period remain uncharacterized. To address this under various food conditions in female mice, we herein combined fiber photometry with a viral-genetic strategy to target GCaMP6s calcium indicator selectively to kisspeptin neurons. Under FR (70% of their regular diet), female mice failed to show sexual maturation without gain of body weight, yet we detected pulsatile activities of kisspeptin neurons at postnatal day 37, around which ad lib fed female mice reach sexual maturation. This data suggest that neural mechanisms to form kisspeptin neural pulses were fully developed even under the FR condition. However, we noticed the frequency of kisspeptin neural pulses was significantly lower in FRed mice compared with ad lib fed mice. Our chronic imaging revealed that, upon removing FR, the frequency of kisspeptin neural pulsatile activities was quickly restored to a level of the ad lib feeding condition. These data suggest the presence of acute and negative regulation of kisspeptin neurons by hunger-sensitive neural systems.Agouti-related peptide (Agrp)-expressing neurons in the arcuate nucleus is one of the candidates that suppress kisspeptin neurons by hunger state, as they are GABAergic neurons known to be active during starvation and mono-synaptically connect to kisspeptin neurons. To manipulate Agrp neurons while monitoring kisspeptin neural activities, we newly developed Agrp-Flpo mouse line. The potential neural circuit mechanisms by which kisspeptin neural pulsatile activities are suppressed in underfed conditions will be discussed with respect to the function of Agrp neurons in the pubertal period of mice.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-374 マウスの母体におけるオキシトシン神経活性の解析 Recording and neural circuit-based manipulation of the maternal oxytocin pulses in mice 幸長弘子(1,2)、萩原光恵(2)、辻本和子(2)、江暁玲(2)、宮道和成(2) 1. 京都大学、2. 理化学研究所 Hiroko Yukinaga(1,2), Mitsue Hagihara(2), Kazuko Tsujimoto(2), Hsiao-Ling Chiang(2), Kazunari Miyamichi(2) 1. Kyoto University, 2. RIKEN Keyword: Oxytocin, Parturition, Breastfeeding For mammals, successful parturition and breastfeeding are critical to the survival of offspring. Pulsatile release of the hormone oxytocin (OT) mediates uterine contraction during parturition and milk ejection during lactation. These OT pulses are generated by unique activity patterns of the central neuroendocrine OT neurons located in the paraventricular (PVH) and supraoptic hypothalamus. Due to technical limitations, however, the identity of OT neurons in the previous studies was speculative based on their electrophysiological characteristics and axonal projection to the posterior pituitary, not on OT gene expression. Although cell-type-specific recoding and manipulations of the OT system have been reported in the studies of OT-mediated emotions and social behaviors, maternal functions of OT remain uncharacterized well by such modern neuroscience toolkits. This motivated us to introduce in vivo chronic Ca²⁺ imaging to characterize maternal pulsatile activities of OT neurons during parturition and lactation. Optical fiber was implanted above the PVH of OT-Cre; Ai162 double heterozygote female mice. Fiber photometry data showed rhythmical pulsatile episodes of Ca²⁺ activities of OT neurons (OT-PAs) in parturition and lactation, although the shape and interval of OT-PAs significantly differed between parturition and lactation. When we chronically monitored OT-PAs in the early and later lactation periods, we noticed that individual peaks became much higher as mothers experienced lactation. There could be two possibilities. Enhanced OT-PAs in the experienced mothers may have been caused by stronger nipple sucking by elder pups. Alternatively, this change may be intrinsic to maternal OT systems independent of the sucking skills of pups. Several experiments revealed that enhanced OT-PAs in late mothers are not simply due to the greater nutritional requests by pups, which suggests the presence of plasticity in the OT neural system. To reveal how are OT-PAs generated or modulated by afferent neural circuitry, we identify mono-synaptic input neurons that can regulate the activities of OT neurons, and demonstrate the pharmaco-genetic manipulation of OT pulses during lactation via activating a prominent pre-synaptic structure of OT neurons. Collectively, our study reveals the basic properties, temporal dynamics, and a modulatory circuit of maternal OT pulses in mice and opens a new avenue for the neuroscience of maternal neuroendocrine functions.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-375 REM睡眠開始における扁桃体基底外側核のドーパミンシグナルの役割 Rapid eye movement sleep is initiated by basolateral amygdala dopamine signaling in mice 長谷川恵美(1,2)、宮坂藍(2)、櫻井勝康(2)、シェラスヨアン(2)、李毓龙(3)、櫻井武(1,2,4) 1. 筑波大学医学医療系、2. 筑波大学国際統合睡眠医科学研究機構、3. State Key Laboratory of Membrane Biology, Peking University School of Life Sciences、4. 筑波大学生存ダイナミクス研究センター Emi Hasegawa(1,2), Ai Miyasaka(2), Katsuyasu Sakurai(2), Yoan Cherasse(2), Yulong Li(3), Takeshi Sakurai(1,2,4) 1. Faculty of Medicine, University of Tsukuba, Ibaraki, Japan, 2. International Institute for Integrative Sleep medicine, University of Tsukuba, Ibaraki, Japan, 3. State Key Laboratory of Membrane Biology, Peking University School of Life Sciences, Beijing, China, 4. Life Science Center for Tsukuba Advanced Research Alliance (TARA), University of Tsukuba, Ibaraki, Japan Keyword: REM Sleep, Dopamine, Amygdala, Cataplexy The sleep cycle alternates between REM (rapid eye movement) and NREM (non-rapid movement) sleep, which is a highly characteristic feature of sleep. However, the mechanisms by which this cycle is generated are totally unknown. We found that a periodic transient increase of dopamine (DA) level in the basolateral amygdala (BLA) during non-rapid eye movement (NREM) sleep terminates NREM sleep and initiates REM sleep. DA acts on dopamine receptor D2 (Drd2)-expressing neurons in the BLA to induce a transition from NREM to REM sleep. This mechanism also plays a role in cataplectic attack, which is a pathological intrusion of REM sleep into wakefulness in narcoleptics. These results show a critical role of DA signaling in the amygdala in REM sleep regulation and provide a neuronal basis of sleep cycle generation.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-376 オレキシン神経の活動記録と睡眠中の活動の役割 Activity recording from orexin neurons and physiological role of activity during sleep 伊藤洋人(1,2,3)、深津紀暁(1,2)、ラハマン　セイクミザン(1,2)、伊澤俊太郎(1,2)、向井康敬(1,2)、小野大輔(1,2)、 Kilduff Thomas (4)、山中章弘(1,2) 1. 名古屋大学　環境医学研究所　神経系分野2、2. 名古屋大学大学院医学系研究科　神経性調節学分野、3. 日本学術振興会　特別研究員(DC1)、4. Center for Neuroscience, Biosciences Division, SRI International, Menlo Park, California 94025, USA Hiroto Ito(1,2,3), Noriaki Fukatsu(1,2), Sheikh Mizanur Rahaman(1,2), Shuntaro Izawa(1,2), Yasutaka Mukai(1,2), Daisuke Ono(1,2), Thomas S. Kilduff(4), Akihiro Yamanaka(1,2) 1. Department of Neuroscience II, Research Institute of Environmental Medicine, Nagoya University, 2. Department of Neural Regulation, Nagoya University Graduate School of Medicine, 3. JSPS Research Fellowship for Young Scientists (DC1), 4. Center for Neuroscience, Biosciences Division, SRI International, Menlo Park, California 94025, USA Keyword: Narcolepsy, orexin, cataplexy Type I narcolepsy is caused by a specific degeneration of orexin neurons, which produce the orexin peptides. Orexin neurons have been considered as "wake active and wake-promoting ". Thus, most studies to date have not focused on the activity dynamics of orexin neurons across vigilance states. Moreover, it has been frequently proposed that orexin neurons should receive excitatory input and be activated to prevent the triggering of cataplexy. However, the activity of orexin neurons during cataplexy has not been directly measured in mouse models of narcolepsy in which the orexin peptides are absent. Therefore, the purpose of this study was to clarify activity of orexin neurons across vigilance states and its physiological role. In order to address this question, we used phenotypically normal transgenic mice and orexin knock-out narcolepsy model mice that enable inducing of genes exclusively in orexin neurons. We conducted Ca²⁺ imaging in freely behaving mice, using fiberphotometry to measure total population activity and microendoscopy (nVista, inscopix) which enables imaging at single cell resolution. As expected, orexin neurons were highly active during wake. However, these neurons showed intermittent synchronous activity during non-rapid eye movement (NREM) sleep, and became low activity during the transition from NREM to rapid eye movement (REM) sleep. During REM sleep, orexin neurons showed weak activity by a subpopulation. In the narcolepsy model mice, the activity of orexin neurons lacking orexin peptides quickly dropped with activity by subpopulation at initiation of cataplexy. Optogenetic inhibition of orexin neurons promoted NREM-REM sleep transitions in NREM sleep. Taken together, dynamics of orexin neurons during NREM sleep regulate the transition to REM sleep. Orexin neurons lacking orexin peptides were almost silent during cataplexy.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-377 非選択的陽イオンチャネルNalcn遺伝子改変マウスを用いたレム睡眠制御機構の解析 NALCN in the forebrain and pons-medulla regions have distinct roles in REM sleep regulation 藤山知之(1)、管野里美(1)、堀田典子(1)、柿﨑美代(1)、一久綾(1)、恒岡洋右(2)、阿部学(4)、望月綾子(1)、セオーネコラッゾパトリシア(1)、宮崎峻弘(1)、水野聖哉(3)、三好千香(1)、上田壮志(1)、渡辺雅彦(6)、崎村建司(4)、高橋智(3)、船戸弘正(1,2)、柳沢正史(1,5) 1. 筑波大学国際統合睡眠医科学研究機構、2. 東邦大学医学部解剖学講座、3. 筑波大学生命科学動物資源センター、4. 新潟大学脳研、5. テキサス大学サウスウェスタン医学センター、6. 北海道大学解剖発生学分野 Tomoyuki Fujiyama(1), Satomi Kanno(1), Noriko Hirashima Hotta(1), Miyo Kakizaki(1), Aya Ikkyu(1), Yosuke Tsuneoka(2), Manabu Abe(4), Ayako Mochizuki(1), Patricia Seoane-Collazo(1), Takehiro Miyazaki(1), Seiya Mizuno(3), Chika Miyoshi(1), Takeshi Kanda(1), Masahiko Watanabe(6), Kenji Sakimura(4), Satoru Takahashi(3), Hiromasa Funato(1,2), Masashi Yanagisawa(1,5) 1. International Institute for Integrative Sleep Medicine, The University of Tsukuba, 2. Dept of Anat, Toho Univ., 3. LARC, Univ. of Tsukuba, 4. Dept of Cell Neurobiol, Niigata Univ., 5. UT Southwestern Medical Center, 6. Dep of Anat & Embryol, Hokkaido Univ. Keyword: REM sleep, Mouse, NALCN Although REM sleep (REMS) is ubiquitous in mammals, the molecular/neural mechanism of REMS regulation remains unknown. We previously established the Dreamless mutant mice exhibiting ~50% reduction in total REMS time. We identified an SNP specific to Dreamless mice within the Nalcn gene, which leads to a single amino acid change (N315K) of NALCN, a non-selective leak cation channel. To elucidate the responsible brain regions / neuronal subtypes through which NALCN regulates REMS, we generated flox and FLEx (flip-excision) knock-in mice bearing Cre-dependent loss-of-function and gain-of-function Nalcn alleles, respectively. In Nalcn-FLEx mice, we confirmed that the mice crossed with a systemic Cre-expressing line Actb-iCre phenocopied the Dreamless mice on electroencephalogram and electromyogram (EEG/EMG) analyses. In Nalcn-flox mice, we confirmed a neuronal subtype-specific deletion of Nalcn mRNA in adult brain tissues. Recently we observed that NALCN has distinct roles in forebrain and pons-medulla regions for REM sleep regulation, by using Foxg1-IRES-Cre or En1-Cre lines. Now I am analyzing the sleep phenotype of Nalcn genetically-modified mice with detailed sleep stage scoring.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-378 慢性的な睡眠の断片化は前駆期パーキンソン病モデルマウスの症状出現を加速する Chronic sleep fragmentation accelerates the onset of prodromal symptoms in Parkinson’s disease model mice 宮崎将行(1,2,3)、八木原紘子(1)、藤田寛美(4)、山門穂高(5)、和田圭司(4)、皆川栄子(1,5) 1. 国立精神・神経医療研究センター神経研究所モデル動物開発研究部、2. 東京医科歯科大学医歯学総合研究科、3. 日本学術振興会特別研究員DC2、4. 国立精神・神経医療研究センター神経研究所疾病研究第四部、5. 京都大学大学院医学研究科 Masayuki Miyazaki(1,2,3), Hiroko Yagihara(1), Hiromi Fujita(4), Hodaka Yamakado(5), Keiji Wada(4), Eiko N Minakawa(1,5) 1. Dept Neurophys, Natl Inst Neurosci, Natl Ctr of Neurol and Psychiatr, Tokyo, Japan, 2. Grad Sch Med Dent Sci, Tokyo Med and Dent Univ, Tokyo, Japan, 3. Research Fellow of Japan Society for the Promotion of Science, Japan Society for the Promotion of Science, 4. Dept Degen Neurol Dis, Natl Inst Neurosci, Natl Ctr of Neurol and Psychiatr, Tokyo, Japan, 5. Dept Neurol, Grad Sch Med, Kyoto Univ, Kyoto, Japan Keyword: Sleep fragmentation, Parkinson's disease, Prodromal symptoms Parkinson’s disease (PD) is the second most common neurodegenerative disease after Alzheimer’s disease (AD). Patients with PD exhibit various non-motor symptoms including constipation, hyposmia, and sleep disturbance in addition to classical motor symptoms such as bradykinesia, muscle rigidity, and rest tremor. The onset of non-motor symptoms often precedes that of motor symptoms, when patients are typically diagnosed as PD. This prodromal phase of PD, which lasts for more than a decade, is an important time window for potential disease modification to prevent or delay the onset of motor symptoms. Sleep fragmentation due to frequent nocturnal arousals is a key feature of insomnia, the most prevalent sleep disorder among patients with PD. While various PD-related etiologies induce sleep fragmentation, recent epidemiological studies suggested that sleep fragmentation itself could exacerbate the disease course of PD. However, the precise causal relationship between PD and sleep fragmentation remains unknown. Meanwhile, we previously demonstrated that sleep fragmentation, which is also prevalent among patients with AD, exacerbates AD pathology. Considering that neurodegenerative diseases share a common disease mechanism of neuronal cell death due to accumulation of misfolded neurotoxic proteins, we aimed to test whether chronic sleep fragmentation exacerbates the disease course of PD. A53T SNCA bacterial artificial chromosome transgenic mice, a prodromal PD model that recapitulates non-motor symptoms, were housed in either of the following cages from the age when these mice exhibit neither constipation nor hyposmia: (1) sleep disturbance (SD) cage (SW-15SD; Melquest), a running-wheel-based device that induces chronic sleep fragmentation similar to PD patients (SD group), (2) wheel cage (WC), a control cage with a running wheel but with no sleep disruption (WC group), or (3) normal cage (NC), another control cage without running wheel (NC group). After 5 or 6 weeks, mice were evaluated for constipation or hyposmia, respectively. The SD group showed significantly worse constipation and hyposmia compared to the WC group, and significantly worse constipation and a tendency towards worse hyposmia compared to the NC group. These results suggest that chronic sleep fragmentation accelerates the onset of prodromal symptoms in PD model mice. Our findings implicate that sleep fragmentation could serve as a potential target for treatments to delay the progression of PD.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-379 乳酸菌が産生する新規睡眠促進物質の同定 Identification of novel sleep-promoting substances produced by lactic acid bacteria. 中川久子(1,2)、小林俊二郎(1,2)、上川内あづさ(3)、石元広志(1) 1. 名古屋大学大学院理学研究科附属ニューロサイエンス研究センター、2. 雪印メグミルク株式会社ミルクサイエンス研究所、3. 名古屋大学大学院理学研究科生命理学 Hisako Nakagawa(1,2), Syunjirou Kobayashi(1,2), Azusa Kamikouchi(3), Hiroshi Ishimoto(1) 1. Neuroscience Institute, Graduate School of Science, Nagoya University, 2. Milk science Research Institute, MEGMILK SNOW BRAND Co., Ltd.,, 3. Division of Biological Science, Graduate School of Science, Nagoya University Keyword: Lactic acid bacteria, Sleep Fruit fly Drosophila harbors symbiotic microorganisms, such as lactic acid bacteria (LAB) in their gut, and microorganisms taken up by feeding might affect the host behavior. We have reported that a LAB, Lactobacillus plantarum SBT2227 significantly promotes fly sleep by feeding. Administration of SBT2227 primarily affected the increase of sleep amount and the decrease of sleep latency at the beginning of night-time. The sleep effect of SBT2227 did not disappear after heat-killing or physical destruction. Hence, it is possible that the metabolites of SBT2227 may contain factors responsible for the sleep effect. In this study, we identified novel molecules, which promote fly sleep from active metabolites of SBT2227. The sleep action of SBT2227 does not require interaction with existing gut bacteria. In other words, it promotes host sleep by a different mechanism of action than probiotics and prebiotics. We examined the sleep effects of the cell fractions of SBT2227. Isolated DNA and RNA did not affect on the fly sleep. Supernatant of crushed bacteria promoted sleep even with protease-treatment, suggesting that macromolecular proteins were not the candidates. Analysis of the action of the centrifugal fraction of crushed SBT2227 on fly sleep behavior showed that small molecules contained in the cell membrane and cytoplasm promoted the sleep of flies. In order to concentrate and recover the small molecules from the SBT2227 strain, molecular adsorption was performed using synthetic resin. After confirming that the resin extracts promoted the sleep of flies, LC/MS analysis was conducted on the resin extracts to identify the small molecular substances contained in them. We then identified novel sleep-promoting substances. Unexpectedly, we also found a novel group of substances that promote arousal from SBT2227 extracts.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-380 アミロイドβ1-40の海馬実質投与はマウスの睡眠覚醒を変化させる Hippocampal injection of amyloid-β1-40 affects sleep/wakefulness state in mice 常松友美(1,2,3)、奥村正樹(2)、佐栁友規(2) 1. 東北大学大学院生命科学研究科、2. 東北大学学際科学フロンティア研究所、3. JSTさきがけ Tomomi Tsunematsu(1,2,3), Masaki Okumura(2), Tomomi Sanagi(2) 1. Grad Sch Life Sci, Tohoku University, 2. FRIS, Tohoku University, 3. PRESTO, JST Keyword: amyloid-β, Sleep, EEG Alzheimer’s disease (AD), a type of dementia, is characterized by the accumulation of the amyloid-β (Aβ) peptides and tau neurofibrillary tangles in the brain. Furthermore, sleep disorders commonly occur in AD patients, and they can precede the onset of disease. In this study, to establish a biomarker using the alternation of sleep, we mimicked AD by administering Aβ into the brain of wild-type mice without using genetically modified mice. Fibril Aβ1-40 (50 µM or 100 µM) was bilaterally injected into the hippocampal CA1 region of 3-month-old wild-type male mice. To determine sleep/wakefulness stage, electroencephalograms (EEGs) and electromyograms were recorded continuously from 3 to 14 days after administration of Aβ1-40. The mice in which buffer was injected into the hippocampus were used as the control. The immunohistochemical studies revealed that the formation of Aβ plaques in the hippocampus and the activation of astrocytes and microglia in the surrounding area. Next, we examined the progressive changes in sleep/wakefulness stage and the component of EEGs of the mice. The results showed that the total time of wakefulness in light period decreased gradually, total time of REM sleep in dark period decreased gradually, and total time of NREM sleep in light period increased gradually after the Aβ injection. Episode duration and episode bouts had no effect. Focusing on the component of EEGs, it became clear that it changed significantly, especially during wakefulness. In other words, the normalized delta component increased, and the normalized theta decreased significantly. Our study has the potential to be a biomarker of AD by using sleep/wakefulness stage and EEG changes. In addition, since the type of misfolded protein to be administered, the brain region in which it accumulates, and its pathogenesis timing can be freely selected, it would be possible to study the effects and mechanisms of each misfolded protein on the pathology.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-381 加齢マウスにおける老化細胞除去による概日リズム障害の改善 Amelioration of age-associated circadian rhythm disorder by removing senescent cells 渡邉綾乃(1,3)、平野有沙(2,3) 1. 筑波大学ヒューマニクス学位プログラム、2. 筑波大学医学医療系、3. 筑波大学国際統合睡眠医科学研究機構 Ayano Watanabe(1,3), Arisa Hirano(2,3) 1. Ph.D. Program in Humanics, Univ of Tsukuba, Ibaraki, Japan, 2. Faculty of Medicine, Univ of Tsukuba, 3. WPI-IIIS, Univ of Tsukuba Keyword: CIRCADIAN RHYTHM, SENESCENT CELL, GLS1 When cells are subjected to various stresses, they acquire resistance to proliferation-promoting stimuli and become senescent cells with stable cell cycle arrest. Characteristics of senescent cells include morphological changes, metabolic changes, chromatin reorganization, changes in gene expression, and a pro-inflammatory phenotype called SASP (senescence-associated secretory phenotype). While cellular senescence is thought to be a mechanism that prevents damaged cells from undergoing malignant transformation and becoming cancerous, senescent cells are involved in a variety of age-related pathologies such as tissue degeneration, inflammatory diseases, and cancer. Existing studies have identified GLS1, which is involved in glutamate metabolism, as an enzyme essential for the survival of senescent cells (Johmura et al., Science, 2021). Inhibition of this GLS1 eliminates senescent cells, and has also been shown to be effective in ameliorating the aging process in aged mice and improving symptoms of atherosclerosis and nonalcoholic steatohepatitis (NASH). Here, we focused on age-related changes in mouse behavior regulated by the central nervous system such as decreased sleep quality and circadian activity rhythms. We tried to improve them in the old animals by inhibiting GLS1. Adeno-associated virus vectors expressing shRNA was used to silence Gls1 mainly in the central nervous system in old mice, and the amount of their activity was measured. Bimodal behavioral rhythm is one of the tendencies seen in young mice but attenuated with aging. We observed bimodal behavioral rhythms during the dark period in some of shRNA-expressing old mice, while control mice did not show. These results may contribute to the development of therapies for increasing the QOL of the elderly and extending healthy life span by removing senescent cells.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-382 SIK3のPKAリン酸化部位欠損が及ぼす睡眠/覚醒行動異常 Sleep/wake behavior of mice lacking PKA phosphorylation site in SIK3 PARK MINJEONG(1)、本多隆利(1)、三好千香(1)、藤山知之(1)、柿﨑美代(1)、一久綾(1)、水野聖哉(2)、高橋智(2)、船戸弘正(1,3)、柳沢正史(1) 1. 筑波大学国際統合睡眠医科学研究機構、2. 筑波大学生命科学動物資源センター、3. 東邦大学医学部 MINJEONG PARK(1), Takato Honda(1), Chika Miyoshi(1), Tomoyuki Fujiyama(1), Miyo Kakizaki(1), Aya Ikkyu(1), Seiya Mizuno(2), Satoru Takahashi(2), Hiromasa Funato(1,3), Masashi Yanagisawa(1) 1. International Institute for Integrative Sleep Medicine, University of Tsukuba, 2. Laboratory Animal Resource Center, University of Tsukuba, 3. Department of Anatomy, Toho University School of Medicine Keyword: Sleep/wake behavior, SIK3, Phosphorylation, PKA We previously identified a kinase, SIK3, as an important sleep regulator through screening of randomly mutagenized mice. Mice that express mutant SIK3 lacking the 52 amino acids encoded by exon 13 showed a decrease in wake time and an increase in NREM sleep time. SIK3 is an AMPK-family protein kinase containing a well-conserved protein kinase A (PKA)-phosphorylation site, serine 551. The skipping of exon 13 results in a deletion of 52 amino acids including S551. Also, Sik3 S551A knock-in mice showed reduced total wake time and increased sleep need. These results suggest that the existence of S551, a PKA recognition site, is crucial for the normal sleep/wake regulation and maintenance of daily sleep need. In SIK3, there are three PKA recognition sites, threonine 469, serine 551, and serine 674. To examine whether the phosphorylation of T469 and S674 of SIK3 is required for proper sleep/wake behavior, we generated mutant mice in which SIK3 T469 and SIK3 S674 were substituted by alanine through the CRISPR/Cas9 method. Sik3 T469A mice showed increased NREM sleep time and NREM sleep delta power, an index for sleep need. Sik3 S674A mice showed no changes in NREM sleep time and NREM sleep delta power. These findings indicate the PKA recognition sites of SIK3, especially T469 and S551 are required for the regulation of sleep/wake behavior. In addition, we generated Flag-tagged Sik3 point mutant mice, and obtained mice brain protein. Using these protein samples, we also identify which proteins bind with SIK3. It is known that SIKs are activated by LKB1-dependent phosphorylation and phosphorylate CRTC family members and Class 2a HDACs. In our results, 14-3-3 was detected clearly in WT and S674A mutant but not detected in T469A and S551A mutants. These results are linked with sleep/wake behavior.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-383 視交叉上核AVP細胞は、視交叉上核全体が発振する概日リズムの周期を制御する AVP neurons control the ensemble period of the central circadian clock of the SCN 津野祐輔(1)、彭雨波(1)、大黒多希子(2)、堀家慎一(2)、山形要人(3)、前島隆司(1)、三枝理博(1) 1. 金沢大学医薬保健研究域医学系、2. 金沢大学疾患モデル総合研究センター、3. 東京都医学総合研究所 Yusuke Tsuno(1), Yubo Peng(1), Takiko Daikoku(2), Shin-ichi Horike(2), Kanato Yamagata(3), Takashi Maejima(1), Michihiro Mieda(1) 1. Grad Sch Med, Kanazawa Univ, Kanazawa, Japan, 2. Research Center for Experimental Modeling of Human Disease, Kanazawa Univ, Kanazawa, Japan, 3. Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan Keyword: SUPRACHIASMATIC NUCLEUS, CIRCADIAN RHYTHM, CALCIUM, FIBER PHOTOMETRY The suprachiasmatic nucleus (SCN), the central circadian clock of mammals, is a network consisting of various types of GABAergic neurons, which can be differentiated by the co-expression of specific peptides. We previously demonstrated that lengthened period of cellular clocks in arginine vasopressin (AVP) neurons, by the deletion of casein kinase 1 delta (CK1δ) (Avp-CK1δ^−/−), elongated the free-running period of circadian behavior, suggesting that AVP neurons act as an essential component of circadian pacemaker cells in the SCN (Mieda et al., 2016, Curr Biol). To examine how much AVP neurons contribute to the circadian period-setting, we compared the behavioral free-running period of mice lacking CK1δ in the whole SCN (Camk2a-CK1δ^−/−) with that of Avp-CK1δ^−/− mice, resulting in no significant difference. However, PER2::LUC reporter rhythm in SCN slices of Avp-CK1δ^-/- mice did not fully recapitulate the period lengthening, underscoring the importance of in vivo analysis. Thus, we aimed to measure in vivo the cellular periods of AVP neurons and VIP neurons in Avp-CK1δ^-/- mice by recording intracellular Ca²⁺ ([Ca²⁺]_i) rhythms using fiber photometry. To do this, we first generated transgenic mice in which tTA (tetracycline transactivator) is expressed specifically in VIP neurons (Vip-tTA mice). Next, these mice were crossed to Avp-CK1δ^-/- mice. Then, we expressed the fluorescent Ca²⁺ indicator jGCaMP7s specifically in AVP neurons or VIP neurons by focally injecting Cre-dependent or tTA-dependent AAV expression vectors, into the SCN. [Ca²⁺]_i in VIP neurons, as well as that in AVP neurons, demonstrated circadian rhythms with periods similar to that of behavioral rhythm in both Avp-CK1δ^-/-;Vip-tTA and control mice. This result indicates that cellular clocks of AVP neurons are capable to control the cellular periods of VIP neurons in vivo, underscoring the critical role of SCN AVP neurons as circadian pacemaker cells.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-384 ラット海馬スライスカルバコール誘導β振動中の高周波振動（HFO）発生機構 Generation mechanism of high-frequency oscillation (HFO) during carbachol-induced oscillation in rat hippocampal slices 夏目季代久(1)、澤田豊宏(1)、前田龍生(1) 1. 九州工業大学 Kiyohisa Natsume(1), Toyohiro Sawada(1), Ryusei Maeda(1) 1. Kyushu Institute of Technology Keyword: High frequency oscillation, Beta rhythm, Hippocampus, Carbachol There are several neuronal rhythms in the rodent brain. Beta rhythm has the frequency of 13-30 Hz and is related to memory retrieval, and theta rhythm has the frequency of 4-12 Hz, and is related to memory encoding. Theta rhythm is induced accompanying with high-frequency oscillation (HFO; 110-160 Hz) in hippocampus. The theta-HFO coupling of rat increases during the decision-making period of a T-maze task. Whether hippocampal beta rhythm accompanies with HFO remains unknown. Carbachol-induced beta oscillation (CIBO) is induced in rat hippocampal slices, and is a model for beta rhythm. Carbachol is a cholinergic agonist. In the previous study, we found HFOs were induced in the generation of CIBO, and they diminished with the suppression of CIBO with the application of glutamate receptor antagonist or muscarinic receptor antagonists. In the present study we studied the source of HFO in CIBO. Eighteen hippocampal slices with 450 μm thick from six rats were used. CIBO was induced in the slices with the application of 30 μM carbachol. CIBO was recorded extracellularly using glass microelectrodes (1 MΩ, 1M NaCl). The electrode was put in the stratum pyramidale of CA3 region in hippocampal slices. The signals were band-pass filtered between 80 and 500 Hz to extract HFOs. To extract CIBO they were band-pass filtered between 13 and 30 Hz. The peak frequency and powers of HFOs were calculated using fast Fourier transformation. The following results were obtained. 1) HFOs were measured as the main sharp negative deflections recorded in stratum pyramidale. HFOs measured in stratum radiatum had the main sharp positive component. The amplitude of HFOs were significantly larger in stratum pyramidale than in stratum radiatum synaptic layer. When dentate gyrus was isolated from CA3 network, HFOs and CIBO could be recorded even in the CA3 mini slices. The results suggest that HFOs will be the spikes from the pyramidal and interneurons in stratum pyramidale and be induced in the local CA3 network. 2) CIBO recorded at stratum pyramidale had the slow positive components while CIBO had the negative components at stratum radiatum. The results suggest CIBO will reflect populational excitatory post-synaptic potential at the recurrent synapse of CA3 pyramidal cells. 3) The phase-coupling analysis of HFOs in CIBO showed that the HFO started to generate from the trough of CIBO and was mainly generated at the rising slope of CIBO. The results suggest that HFO will be induced by the firing of the pyramidal and interneurons locally located at stratum pyramidale, in turn may activate CA3 pyramidal cells populationally and finally CIBO will be induced.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-385 Distinguishing Effects of Circadian Clock Desynchronization on Senescent Cell Accumulation in Peripheral Tissues of Mice Nicholas Grooms(1), Yoshikazu Johmura(2), Makoto Nakanishi(2), Arisa Hirano (3) 1. Graduate School of Comprehensive Human Sciences, International Institute for Integrative Sleep Medicine, University of Tsukuba, Tsukuba, Japan, 2. Institute of Medical Science, University of Tokyo, Tokyo, Japan, 3. International Institute for Integrative Sleep Medicine, University of Tsukuba, Tsukuba, Japan Keyword:* Circadian Clock and Rhythms, Cellular Senescence , Aging, Reporter Mouse Model Circadian rhythms facilitate a critical role in maintaining health throughout various organisms, and many important circadian clock genes have been increasingly evaluated for key effects on aging and cellular senescence. However, despite the current evidence which suggests a contribution from key clock genes towards senescence in age-related conditions such as oncogenesis and immune system degradation and dysfunction, it remains unclear what the contribution of the circadian clock itself is in the cellular aging process when the circadian rhythms are disrupted but clock genes are left functionally intact. Thus, the aim of this research project is to clarify the role of the circadian clock in the accumulation of senescent cells by establishing a method to desynchronize the circadian rhythm entrainment of peripheral tissues through inhibiting the suprachiasmatic nucleus (SCN), the site of the master circadian clock, without manipulating peripheral clock genes in mouse models. This method employs SCN targeted adeno-associated virus (AAV) injection surgeries with both tetanus toxin (TeNT) and green fluorescent protein (GFP) control constructs using p16-Cre-ER/-td-Tomato reporter mouse model, which allows for senescence visualization due to the expression of red fluorescence in the presence of tamoxifen when p16, an established genetic marker of cellular senescence, is activated and transcribed. This is followed by behavioral recording for analysis of circadian rhythm and period disruptions, measurements for mouse metabolism, and finally collection of blood/serum and tissue samples, including the brain, liver, and kidneys, for measurements of cellular senescence and tissue health. Furthermore, in vitro experiments using primary p16-Cre-ER/-td-Tomato reporter mouse embryonic fibroblasts (MEFs) to examine the manipulation of important circadian clock proteins and the accumulation of senescent positive cells both with and without senescence-inducing treatments were conducted for additional analysis. The methods and results presented will provide much needed insights into the nature of how the circadian clock itself and senescence are connected. By using a direct approach, these methods may also be useful to expand further studies in uncovering the pathological mechanisms of and develop treatments for both circadian-related disorders and age-related diseases respectively.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-386 ビフィズス菌SBT2786株の睡眠促進作用に必要な脳神経系トランスポーターの分子遺伝学的解析 Molecular genetic analysis of brain transporters required for the sleep-promoting effect of Bifidobacterium adolescentis SBT2786 in Drosophila. 松本菜々恵(1,2)、上川内あづさ(1,3)、石元広志(1) 1. 名古屋大学大学院　理学研究科　附属ニューロサイエンス研究センター、2. 雪印メグミルク株式会社　ミルクサイエンス研究所、3. 名古屋大学大学院　理学研究科　生命理学 Nanae Matsumoto(1,2), Azusa Kamikouchi(1,3), Hiroshi Ishimoto(1) 1. Neuroscience Institute, Graduate School of Science, Nagoya University, 2. Milk science Research Institute, MEGMILK SNOW BRAND Co., Ltd., 3. Division of Biological Science, Graduate School of Science, Nagoya University Keyword: Sleep, Drosophila, Bifidobacteria Lactic acid bacteria (LAB) and bifidobacteria are known as symbiotic bacteria in the gut, and affect various aspects of brain functions both in vertebrates and invertebrates. By using high-throughput sleep-behavior screening of Drosophila melanogaster, we identified strains with sleep-promoting effects from 46 species of LABs and bifidobacteria. In particular, the sleep-promoting effect of Bifidobacterium adolescentis strain SBT2786 was the most prominent, but the evolutionarily different genus Lactobacillus also had sleep-promoting effects. One of the common features of these bacterial strains was a biological effect that increased the amount of fly sleep at the onset of the nighttime. It is assumed that there are common molecular mechanisms for the biological effects of these bacterial strains, but it still remains unclear. Our previous studies have shown that the sleep effects of these strains do not require interaction with other intestinal bacteria, i.e., they do not act as prebiotics or probiotics, but as substances retained by the bacteria themselves. Here, we focused on the transporter molecules, which transport biomolecules of LABs, and analyzed their role in sleep regulation by using molecular genetics of Drosophila melanogaster. These analyses revealed that two transporter molecules positively regulate sleep in flies, and may also mediate the sleep-promoting effects of SBT2786. We would like to discuss these results at this conference.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-387 Opto-chemical control of sleep in the nucleus accumbens using a photocaged adenosine A2A receptor allosteric modulator Koustav Roy(1), Shuji Ioka(1), Mao Amezawa(1), Yoan Cherasse(1), Hiroshi Nagase(1), Masashi Yanagisawa(1), Kaspar Vogt(1), Manabu Abe(2), Tsuyoshi Saitoh & Michael Lazarus(1) 1. University of Tsukuba, 2. Hiroshima University Keyword:* Slow wave Sleep (SWS), Photopharmacology, Adenosine A2A Receptor , Nucleus Accumbens (NAc) Photopharmacology may offer the possibility of curing diseases and alleviating symptoms while preventing uncontrolled drug activity, i.e., the drug is active only at the times and places where it exerts its therapeutic effect. Although chemical photo switches have been used extensively in vitro, their use in vivo has been slow, largely because of the difficulties in applying these probes in mammalian models. We revealed a prominent role of indirect pathway neurons in the nucleus accumbens (NAc) in sleep/wake regulation and proposed that the NAc links motivation and sleep. This brain circuit may explain why we feel sleepy in the absence of motivating stimuli, i.e., when we are bored. Adenosine is a plausible candidate molecule for activating NAc indirect pathway neurons to induce slow-wave sleep (SWS) because caffeine, the most widely consumed psychostimulant in the world, produces its arousal effect in the NAc by blocking adenosine A2A receptors (A2AR) on indirect pathway neurons. However, the ability of adenosine in controlling NAc indirect pathway neurons for sleep induction remains to be elucidated. We recently reported the first positive allosteric A2AR modulator, named A2AR PAM, that evokes A2AR responses in the brain and developed a visible-light photoactivatable derivative of A2AR PAM (opto-A2AR PAM). Opto-A2AR PAM showed remarkable water solubility (>10 mM) and has an absorption maximum at 415 nm in aqueous solution. SWS was significantly increased for 5h in wild-type mice after intraperitoneal administration of opto-A2AR PAM and stimulation with violet light (405 nm) for 1 h after drug treatment, whereas no effect was observed in the absence of light exposure or in A2AR knockout mice. By using opto-A2AR PAM, we induced sleep for the first time in freely behaving mice by photopharmacologic allosteric A2AR modulation, suggesting that extracellular adenosine is involved in the regulation of sleep in the NAc.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-388 ランニングホイールによる自発活動増加負荷下で飼育した過眠モデルマウスSleepyの成長過程における睡眠覚醒および体重の変化 Sleep/wakefulness and body weight of Sleepy mutant mouse, a mouse model of hypersomnia, grown in the presence of a running wheel that increases spontaneous activity 堀田-平島範子(1)、三好千香(1)、一久綾(1)、柿崎美代(1)、管野里美(1)、柳沢正史(1,2,3)、船戸弘正(1,4) 1. 筑波大・WPI-IIIS、2. テキサス大・サウスウェスタン医学センター、3. 筑波大・TARA、4. 東邦大・医・解剖微細形態 Noriko Hotta-Hirashima(1), Chika Miyoshi(1), Aya Ikkyu(1), Miyo Kakizaki(1), Satomi Kanno(1), Masashi Yanagisawa(1,2,3), Hiromasa Funato(1,4) 1. WPI-IIIS, Univ. of Tsukuba, Ibaraki, Japan, 2. Dept. of Mol. Genet., Univ. of Texas Southwestern Med. Cent., Dallas, USA, 3. TARA, Univ. of Tsukuba, Ibaraki, Japan, 4. Dept. of Anat., Fac. of Med., Toho Univ., Tokyo, Japan Keyword: Sleep, Spontaneous activity, Body weight, EEG/EMG It is known that physical activity during wakefulness affects not only body weight but also sleep, but the mechanism is not fully understood yet. It has reported that physical activity increases non-REM sleep and decreases REM sleep in human studies. However, due to the difficulty of maintaining the experimental　environment, there have been few reports of long-term studies that include the growing period. In this study, we examined the effects of a constant exercise environment on weight gain and sleep/wakefulness in mice during growth. We installed a rotating wheel that increases the amount of spontaneous activity in the home cage, and kept male C57BL/6N mice in the environment from birth (wheel group). Body weight measurements were continuously taken from 4 to 30 weeks of age, and it was confirmed that the weight gain with aging in the wheel group was about 30% less than that in the normally housed mice (non-wheel group) at 30 weeks of age. We performed body weight measurements in the presence of a rotating wheel in SIK3 Sleepy mutant (SLP) mice, a mouse model of hypersomnia and obesity recently established in our laboratory. The littermate wild type (WT) wheel group showed approximately 30% less weight gain than the non-wheel group at 30 weeks of age. On the other hand, the SLP wheel group had about 20% less weight gain than the SLP-non-wheel group. However, at 10 weeks of age, the WT-wheel group showed a 10% decrease in weight gain, whereas the SLP-wheel group showed a 40% decrease. In addition, we performed EEG/EMG based sleep recording at 10-13 weeks and 30-33 weeks of age. Both SLP and WT mice showed a decrease in arousal time with age, but the wheel group showed an increase in arousal time at both ages of weeks. The awakening time of SLP-wheel group was shorter than that of WT-non-wheel group at 10-13 weeks but did not differ from that of WT-non-wheel group at 30-33 weeks. These results confirm that an environment that allows long-term spontaneous activity, including during growth, has an ameliorating effect on genetic hypersomnia and obesity. We confirmed that the strength of the effect differed depending on whether it was in adolescence or middle age in terms of body weight and sleep/wakefulness.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-389 起床時コルチゾール反応(CAR)は人間以外の哺乳類でもみられるのか Does Cortisol Awaking Response occur in non-human mammals? 南部美友(1)、田中和正(1) 1. 沖縄科学技術大学院大学 Miyu F Nambu(1), Kazumasa Z Tanaka(1) 1. Okinawa Institution of Science and Technology Keyword: Hippocampus, Cortisol, Cortisol Awaking Response , Memory There are still many memory processes that have not been understood, and its importance makes it an area that requires further research. In recent years, much research has been carried out into the effects of stress hormones on memory. In humans, cortisol, also known as the stress hormone, is superimposed on circadian rhythms and rises sharply 30-40 minutes after waking. This increase after waking is called the cortisol arousal response (CAR) and is characteristic of the hypothalamic-pituitary-adrenal (HPA) axis. Since there are many cortisol receptors in the hippocampus, it is thought that the hippocampus regulates cortisol levels through negative feedback. Various studies have shown a link between cortisol/corticosterone and memory; however, it is unknown the physiological significance of CAR. A previous study showed the decrease in CAR of human patients whose hippocampus had decreased in size due to diabetes. This finding led us to think that there might be some relationship between CAR and the hippocampus. In this study, we will examine 1) whether CAR also occurs in mice as same as humans, and 2) whether CAR in mice depends on the hippocampus. Firstly, we will establish a method to measure corticosterone levels in freely moving mice throughout the day. This technique will allow us to examine the stress hormone's fluctuation with the minimum effect caused by the experimenter's handling. Secondly, we will examine the rise in corticosterone levels when mice with hippocampal inactivation awaken. The current study will test a hypothesis that CAR is present across species and reveal a novel interaction between stress and memory.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-390 視索前野の慢性的な細胞除去が睡眠覚醒周期に及ぼす影響 Effects of chronic cell ablation in the preoptic area on sleep-wake cycles 今村彩子(1,2)、宮竹功一(2,3)、庵原亜貴子(2)、本城咲季子(2) 1. 筑波大学グローバル教育院ヒューマニクス学位プログラム、2. 筑波大学国際統合睡眠医科学研究機構、3. 学術振興会特別研究員 Ayako Imamura(1,2), Koichi Miyatake(2,3), Akiko Ihara(2), Sakiko Honjoh(2) 1. Ph.D. Program in Humanics, University of Tsukuba, 2. IIIS, University of Tsukuba, 3. JSPS Research Fellow Keyword: SLEEP, PREOPTIC AREA, CELL ABLATION Sleep is an essential physiological phenomenon for organisms with a nervous system. Various brain regions responsible for wakefulness or sleep have been identified by classical elimination of brain regions or by acute manipulation of neural activity. However, acute manipulation of these regions often has only a temporary effect on the sleep-wake cycles and chronic inhibition of neural activity in these areas has little effect. These may have been an adaptation and compensation by other brain regions. In this study, we focused on the preoptic area (POA), which has been reported to promote sleep. It is known that sleep is significantly reduced when the POA is irreversibly removed by electrolytic lesions. The sleep-suppressing effects of POA inhibition are not temporary, but they are not necessarily specific to POA neurons, since axons passing through the area are also affected by the electrode-based technique. The aim of this study is to investigate the effect of brain region-specific cell ablation in the POA on the sleep-wake cycle. Galanin positive cells in the POA were reported to be important in promoting sleep by acute activation using optogenetics, so we focused on Galanin-positive cells. First, we attempted to perform neuron-specific cell ablation in the whole brain during development by crossing Gal-Cre mice, which specifically express Cre recombinase for Galanin-positive cells, with NSE-DTA mice, which express diphtheria toxin A (DTA) specifically for neurons in a Cre-dependent manner. In addition, we locally injected AAV, which expresses DTA in a Cre-dependent manner, into the POA of adult Gal-Cre mice for brain region-specific cell ablation after completion of the brain development. In both experiments, there was no effect on the sleep-wake cycle. The immunostaining results showed that the cell ablation efficiency was low. Thus, we examined the effect of taCasp3 AAV, which overexpresses procaspase-3 in a Cre-dependent manner, on the induction of cell apoptosis. We found that taCasp3 AAV could induce apoptosis more efficiently than DTA in the POA. Therefore, using taCasp3 AAV, we performed POA-specific cell ablation. As a result, fragmentation of the sleep-wake cycle was observed, the frequency of transitions from wakefulness to sleep was increased. This effect was sustained for more than four weeks, and no compensation by other brain regions was observed. These results suggest that POA is necessary for consolidated sleep-wake cycles.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-391 レム睡眠制御の分子機構：近位依存性ビオチン標識法BioIDを用いた電位非依存性陽イオンチャネルNALCNのタンパク質間相互作用因子の探索 Molecular mechanisms of REM sleep regulation: identifying protein-protein interactions of NALCN channel through BioID technique 山元ひかり(1)、藤山知之(1)、北園智弘(1)、一久綾(1)、管野里美(1)、柿﨑美代(1)、Jinhwan Choi (1)、中田慎也(1)、村田知弥(2)、水野聖哉(2)、杉山文博(2)、高橋智(2)、船戸弘正(1,3)、柳沢正史(1) 1. 筑波大学国際統合睡眠医科学研究機構、2. 筑波大学生命科学動物資源センター、3. 東邦大学医学部解剖学講座 Hikari Yamamoto(1), Tomoyuki Fujiyama(1), Tomohiro Kitazono(1), Aya Ikkyu(1), Satomi Kanno(1), Miyo Kakizaki(1), Jinhwan Choi (1), Shiya Nakata(1), Kazuya Murata(2), Seiya Mizuno(2), Fumihiro Sugiyama(2), Satoru Takahashi(2), Hiromasa Funato(1,3), Masashi Yanagisawa(1) 1. International Institute for Integrative Sleep Medicine, Univ of Tsukuba, Ibaraki, Japan, 2. LARC, Univ of Tsukuba, Ibaraki, Japan, 3. Department of Anatomy, Faculty of Medicine, Univ of Toho, Tokyo, Japan Keyword: NALCN, BioID, REM sleep Sleep is a ubiquitous behavior that conserved from vertebrates to invertebrates. REM sleep is unique to animals with highly advanced brain structures. Molecular/cellular mechanisms of REM sleep regulation are largely unclear. The Dreamless mutant mice with a single amino acid substitution (N315K) in the NALCN protein show abnormalities in homeostasis of REM sleep (Funato et al, *Nature, 2016). NALCN is a voltage-independent cation channel known to be involved in the circadian rhythm and respiratory regulation, and NALCN channel complexes such as UNC80, UNC79, and NALF-1 have been identified. However, the components of NALCN channel complex and molecular mechanisms of NALCN regulation including channel opening/closing in REM sleep homeostasis remain unknown. To elucidate the molecular mechanisms of REM sleep regulation through NALCN, we are trying to comprehensively search for unknown protein-protein interactions of NALCN by using biotin-based proximity labeling technique, BioID. Recently, the MAC-tag (consisting of HA-tag, strepⅢ-tag and promiscuous mutant biotin ligase BirA) was developed, that enables parallel analyses of both BioID and conventional affinity purified mass spectrometry (AP-MS). In this study, we generated several lines of knock-in mice, in which a MAC-tag was fused to NALCN, UNC80 or NALF-1, currently known components of the NALCN channel complex. We confirmed no expression changes in the target alleles at the mRNA and protein levels. Using these mice, we identified several interacting proteins of each target by AP-MS. In the future, we will perform comprehensive mapping of NALCN protein interactions by proteomic analyses through BioID, in order to identify novel proteins involved in the molecular regulation of NALCN that are relevant for REM sleep homeostasis.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-392 Changes in sleep/wake by acute disruption in Hdac4 signaling Kim Staci(1)、Nakata Shinya(1)、Asama Nodoka(1)、Ehosainy Asmaa(1)、Hotta-Hirashima Noriko(1)、Kakizaki Miyo(1)、Ikkyu Aya(1)、Kanno Satomi(1)、Mizuno Seiya(2)、Takahashi Satoru(2)、Miyoshi Chika(1)、Funato Hiromasa(1,3)、Yanagisawa Masashi(1,4,5) Staci Jakyong Kim(1), Shinya Nakata(1), Nodoka Asama(1), Asmaa Ehosainy(1), Noriko Hotta-Hirashima(1), Miyo Kakizaki(1), Aya Ikkyu(1), Satomi Kanno(1), Seiya Mizuno(2), Satoru Takahashi(2), Chika Miyoshi(1), Hiromasa Funato(1,3), Masashi Yanagisawa(1,4,5) 1. Intl. Inst. for Integrative Sleep Med. (WPI-IIIS), Univ. of Tsukuba, Tsukuba, Japan, 2. Transborder Med. Res. Ctr., Univ. of Tsukuba, Tsukuba, Japan, 3. Dept. of Anatomy, Fac. of Med., Toho Univ., Tokyo, Japan, 4. Life Sci. Center, Tsukuba Advanced Res. Alliance, Univ. of Tsukuba, Tsukuba, Japan, 5. Dept. of Mol. Genet., Univ. of Texas Southwestern Med. Ctr., Dallas, TX Keyword: Kinase activity, Sleep homeostsis Sleep is ubiquitous behavior observed in various species from nematodes to mammals, where properly maintained sleep is essential for the general well-being of the organism. Recent studies have shown distinct groups of neuronal population that are involved in the transitions between sleep and wake states. These networks of neurons can drive vigilance stage switches and changes in neuronal activity related to sleep and wakefulness. Despite the growing interest and findings in the maintenance of sleep/wakefulness at the circuit level, the intracellular regulatory mechanism of the response to sleep need changes remains largely unknown. We conducted EEG/EMG-based dominant screening sleep/wake behavior in randomly mutagenized and successfully identified several sleep regulatory genes. We previously reported a long-sleep pedigree carrying a gain-of-function mutant allele, Sleepy, in the Sik3 gene and showed SIK3 as a central regulator of sleep homeostasis. Sik3-Sleepy mice showed marked increase in daily total non-REM sleep (NREMS) sleep, as well as inherent increase in slow-wave activity during NREMS. Our effort in elucidating the key component of sleep/wake modulation has continued with discoveries of other sleep regulating genes. Sleepy2 is one of such mutant pedigrees with a loss-of-function of HDAC4. The heterozygous mutant mice showed increase in both daily NREMS time and NREMS slow-wave activity. Importantly, HDAC4 is known as a primary target of SIK3, where the phosphorylation by SIK3 of a conserved serine residue is essential in the nuclear-to-cytoplasmic transfer of HDAC4. Indeed, mice with a phosphodeficient mutation in this residue of HDAC4 showed an opposite phenotype: these mice showed increase in daily wake time and significant decrease in the slow-wave activity during NREMS. Taken together, sleep/wake maintenance and proper response to sleep need changes require canonical signaling of SIK3-HDAC4. In parallel with the systemic mutant mice, we adopted two approaches for acute and brain-specific inhibition of HDAC4 activity by (1) intracerebroventricular injection of selective HDAC4 inhibitor, LMK-235; and (2) AAV-mediated CNS transduction of the phosphodeficient HDAC4 mutant allele. Here we present the detailed analysis of sleep/wake changes induced by perturbed HDAC4-mediated intracellular signaling.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-393 逆行性ウイルスベクターによるオレキシンニューロンの入出力機構の解明 Visualizing input-output architecture of orexin neurons with retrograde tracing vectors 齊藤夕貴(1,2)、Cherasse Yoan(1,2)、丹羽康貴(1,2)、櫻井武(1,2) 1. 筑波大学医学医療系、2. 筑波大学国際統合睡眠医科学研究機構 Yuki Cleo Saito(1,2), Yoan Cherasse(1,2), Yasutaka Niwa(1,2), Takeshi Sakurai(1,2) 1. University of Tsukuba, Ibaraki, Japan, 2. WPI-IIIS, Univ of Tsukuba, Ibaraki, Japan Keyword: orexin, neural circuit, monoamine Orexin-producing neurons (orexin neurons), which are located in the lateral hypothalamic area (LHA) play a highly important role in the maintenance of wakefulness. Orexin neurons send widespread projections to nuclei containing monoaminergic neurons such as ventral tegmental area (VTA), locus coeruleus (LC), tuberomammillary nucleus (TMN), and raphe nuclei (Nambu, et al., 1999), all of which contain monoaminergic neurons. We previously identified input neurons that make direct synaptic contacts to orexin neurons with modified rabies vector-based retrograde tracing (Saito, et al., 2018). This study showed orexin neurons receive input from a variety of regions of the brain. It has been still unknown whether orexin neuron populations that send projections to particular regions receive biased input. In this study, to identify the neuronal inputs to the orexin neurons with projections to particular regions, we used projection sites-specific rabies monosynaptic retrograde tracing (Piñol, et al., 2018). We used newly generated orexin-iCre KI mice to analyze the input-output relationship of orexin neuronal circuits using the modified multi-color simple-cTRIO method. This new method allowed us to detect more than two different input-output pathways in the same brain. This study revealed that orexin neurons projecting to each output region also send projections to all brain regions we examined. Orexin neurons integrate information from broad regions of the brain and broadcast to all monoaminergic nuclei and other regions. However, we found some biased input and output architectures.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-394 睡眠覚醒サイクルを通じた神経活動ダイナミクス Neural activity dynamics across vigilance states 本城咲季子(1)、宮竹功一(1,2)、庵原亜貴子(1) 1. 筑波大学、2. 日本学術振興会 Sakiko Honjoh(1), Koichi Miyatake(1,2), Akiko Ihara(1) 1. University of Tsukuba, 2. Japan Spciety for the Promotion of Science Keyword: Sleep, Firing rate , Thalamus, Cortex Sleep is a homeostatically regulated biological process. As we stay awake longer, “sleep need” accumulates in the brain and we feel sleepier and sleepier, resulting in a marked decline in our cognitive ability. However, the fundamental functions of sleep at the cellular level still remain controversial. What is the molecular identity of “sleep need”? How does it interfere with our cognitive ability? How “sleep need” gets dissipated during sleep? To address these questions, we investigate sleep/wake history-dependent changes in neural activity. In this study, we focused on the thalamo-cortical system, which plays critical roles in high-order cognitive functions. We implanted EEG and EMG electrodes for vigilance state analysis and microelectrodes in multiple thalamo-cortical regions for single unit recording, simultaneously. 24 hours of baseline, 6 hours of sleep deprivation, and 18 hours of subsequent recovery sleep were recorded in freely behaving mice. We analyzed single unit activities using wave_clus, a method of unsupervised spike sorting with wavelets and superparamagnetic clustering. Our sleep scoring and single unit analyses, that were performed independently, revealed that thalamic neurons show a consistent switch-like behavior, with high and low firing rates in wakefulness and in NREM sleep, respectively. In contrast, cortical neurons showed heterogeneous activity patterns. Interestingly, we found that neurons in a higher cortical area tend to fire more often during sleep than they do during wakefulness. These results suggest that information flow among the brain is fundamentally different between wakefulness and sleep.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-395 月経随伴症状の強さの神経基盤 Severity of premenstrual symptoms is related to the morphological characteristics of the cerebral gray matter as measured by magnetic resonance imaging 堂西倫弘(1)、寺田正樹(2)、金桶吉起(1) 1. 和歌山県立医科大学、2. 医療法人昭陽会和歌山南放射線科クリニック Tomohiro Donishi(1), Masaki Terada(2), Yoshiki Kaneoke(1) 1. Wakayama Medical University, Wakayama, Japan, 2. Wakayama-Minami Radiology Clinic, Wakayama, Japan Keyword: MENSTRUAL CYCLE, MRI, PMS Up to 90% of women in reproductive age are suffered from various symptoms especially before menstruation (premenstrual symptoms). Although these symptoms must be related to the circulating hormonal level changing with menstrual cycle, the severity could not be explained by the magnitude of hormonal levels. Thus, individual difference of the sensitivity to the hormonal change due to genetic influence is hypothesized, but the cause is largely unknown. In this study, we used structural MRI images to explore brain regions associated with severity of menstrual (particularly CNS) symptoms, measured by Menstrual Distress Questionnaire (MDQ), from 57 young healthy females (20.1±1.2 years old) with a stable menstrual cycle (25-33 days). On 1-7 (M1) and 20-24 (M2) days of menstruation, T1- and T2- weighted (T1w and T2w) images were acquired by 3T MRI. These images were normalized and calibrated using the signals outside the brain tissue. Then, the ratio images (T1w/T2w) were calculated and segmented into gray matter. Brain regions including the subcortical nuclei and the cerebellum were divided into 388 regions by the atlas (AICHA and AAL). The mean signal intensity for each region was calculated and standardized by the mean and S.D. of the signal intensities among all regions for each subject at each acquisition. Because the signal intensity for each region did not change with acquisition day (M1 and M2), the mean image was used to check the relationship with MDQ score. We found various brain regions were associated with MDQ scores: Especially, a significant negative correlation was revealed in the bilateral superior temporal gyri, while a significant positive correlation in the right dorsolateral prefrontal cortex (by the generalized linear model, FDR corrected p < 0.005). The results suggest that the morphological difference of the gray matter is related to the magnitude of premenstrual symptoms independent of circulating hormonal level change, because the signal intensity did not change with menstrual days (M1 and M2). Recent researches suggest that the gray matter signal intensity of T1w/T2w ratio image is related to the synaptic density instead of myelination. Further study using the ratio image may help investigating new treatment for the patients with severe symptoms.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-396 新規高感度オプシンを用いた冬眠様状態の誘導 Induction of mouse hibernation-like state with newly developed, highly sensitive opsin 高橋徹(1,2)、平野有沙(1,2)、櫻井武(1,2) 1. 筑波大学、2. 筑波大学国際統合睡眠医科学研究機構 (WPI-IIIS) Tohru M Takahashi(1,2), Arisa Hirano(1,2), Takeshi Sakurai(1,2) 1. University of Tsukuba, 2. WPI-IIIS Keyword: optogenetics, body temperature, preoptic area, hibernation A variety of useful optogenetic tools are currently available, but none have been reported to be effective for continuous stimulation of neurons for prolonged time periods (e.g., 24 hours). We developed an opsin (here referred to as HPN1) and applied it to induce a hibernation-like hypometabolic state (Q neurons-induced hypometabolic state: QIH) in mice. Optogenetic excitation of Q neurons using HPN1 successfully induced deep hypothermia and sustained it for a long time by delivering light at a quite low intensity - a level that cannot properly excite commonly-used opsins such as ChR2 and SSFO. The HPN1-mediated QIH recapitulated dynamics of some physiological changes observed in the natural hibernation. The optogenetics established in this study could allow for appropriate optogenetic manipulation in experiments such as circadian rhythms and thermoregulation where adverse effects of light exposure and heat are of concern, and enable us to identify the neural mechanisms underlying long-term behaviors such as hibernation.

2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-397 埋め込み型加速度計を用いた多層性エンリッチ環境における自発運動の行動解析 Behavioral analysis of voluntary physical activity using mobile accelerometer in multistory enriched environment 柳田信也(1)、久保田夏子(2) 1. 東京理科大学、2. 東京都立大学 Shinya Yanagita(1), Natsuko Kubota(2) 1. Tokyo University of Science, 2. Tokyo Metropolitan University Keyword: Enrich Environment, Exercise, Monoamine, Voluntary Running Numerous studies have shown that enriched environments (EE) could be effective for experimental rodents to improve some brain functions related to stress response and anxiolytic effect, and speculating that playfulness in EE might influence it. On the other hand, it is well known that increasing levels of physical activity could have beneficial effects as well as EE. Taken together with this evidence, the question arises: Which is effective for the improvement of brain function between playing or physical activity? To answer the question, we made a multistory enriched environment (Multi-EE) that can increase physical activity in rats. However, it is difficult to measure physical activity by existing analysis methods, such as infrared ray systems, and video tracking. Recently, a mobile accelerometer has been developed to be able to analyze the amount of physical activity without limitation. In this study, we performed the behavioral analysis of voluntary physical activity using the mobile accelerometer in our original enriched environments. We originally made Multi-EE, which are consisted of three stories. The male Wistar rats housed the Multi-EE or normal EE for 4weeks in group housing conditions (3 rats per cage). The rats housed in Multi-EE allow access to the three stories freely by ladders. Daily physical activity was recorded using a mobile accelerometer and compared Multi-EE and Normal-EE. Following 4weeks, brain monoamine levels, which are involved with increasing physical activity-induced- psychological effects, were measured by High-Performance Liquid Chromatography (HPLC) in several brain regions. Muscle and fat volume were also measured. In this study, we have been successful to analyze voluntary physical activity in both normal-EE and Multi-EE. The Multi-EE significantly changed physical activity compared to normal-EE. The voluntary physical activity in Multi-EE significantly increased the volume of soleus muscle compared to normal-EE, indicating that Multi-EE might be effective to increase the physical load. Furthermore, the Multi-EE housing was able to change the brain monoamine levels, such as serotonin and dopamine. The changing levels of these monoamines are known to have some beneficial effects on brain health. Therefore, the results of the present study suggest that increasing levels of physical activity by Multi-EE could influence some brain and physical functions.		2022年7月1日　11:00～12:00　沖縄コンベンションセンター展示棟　ポスター会場1 2P-398 体温制御機構における恒常性の強化学習 Homeostatic reinforcement learning on body temperature regulation 矢田真奈(1)、石川怜奈(1)、本田直樹(2,1,3) 1. 京都大学、2. 広島大学、3. 自然科学研究機構生命創成探究センター Mana Y Fujiwara(1), Sana Ishikawa(1), Naoki Honda(2,1,3) 1. Kyoto University, 2. Hiroshima University, 3. Exploratory Research Center on Life and Living Systems Keyword: HOMEOSTASIS, REINFORCEMENT LEARNING, DECISION MAKING, CLASSICAL CONDITIONING Animals maintain the homeostasis of their internal state in a dynamic, ever-changing environment. Homeostatic mechanisms work to prevent impact on the internal state prior to the expected changes. A paradigmatic example is the acquisition of tolerance to ethanol-induced hypothermia. This is a kind of conditioned drug tolerance; when repeated administration of ethanol, in the presence of a distinctive cue, results in tolerance, whereas the tolerance does not occur without the cue. However, the computational mechanism of how the brain learns the association between the cue stimulus and the following deviation of internal state and how the compensatory response to the cue occurs appropriately, remains to be elucidated. Here, we conducted simulations to elucidate the mechanisms that animals experience when responding to the associations between deviations of internal state and cues. We define the reward based on the needs of the body’s internal state and built a model based on the hypothesis that the system learns to respond appropriately. So, the compensation of body temperature in a reinforcement learning framework occurs. In this model, the difference from the setpoint of body temperature was set as a negative reward, and the degree of the compensatory response was set to be learned through experience. In this way, we explain how animals learn to take rational responses and behaviors that are suitable for maintaining their internal state anticipatorily, and which part is impaired when irrational responses and further behavioral choices occur. Finally, we will then discuss how reinforcement learning based on internal states and rewards is implemented in animals.