Poster Session Abstracts
The 7th Annual IIIS Symposium ~ Solving the mystery of sleep ~Data Blitz by Poster Presenters

13:15 – 13:45

Poster Session

13:45 – 14:15

Poster list#

Presenter

Title

P01

Deependra Kumar

Function of the adult-born neurons in memory consolidation during sleep

P02

Zhiqiang Wang

P03

Takashi Matsui

P04

Simeng Zhang

P05

Chia-Ying Lee

P06

Kazuya Suwabe

P07

Tomoyuki Fujiyama

P08

Sakura Eri Maezono

P09

Akira Uematsu

Dissection of prefrontal cortex circuit in guiding the extinctions

P10

Noriko HottaHirashima

Longitudinal EEG/EMG measurement in mice show sleep/wakefulness maturation from infant

P11

Fuyuki Asano

Sleep/wakefulness in neuron-specific SIK3-deficient mice

6

P12

Shuntaro Izawa

Forgetting during REM sleep by hypothalamic MCH neurons

7

P13

Xuzhao Zhou

Adenosinergic mechanisms of sleep control in the nucleus accumbens

7

P14

Tomohiro Kitazono

Exploration of the intracellular molecular mechanisms for homeostatic sleep/wake regulation

8

P15

Hiroyuki Watanabe

P16

Emi Hasegawa

Search for neural circuits of narcolepsy-cataplexy9

P17

Yoan Cherasse

A neuronal subpopulation in the bed nucleus of the stria terminalis regulates NREM sleep

9

P18

Hiroyuki Mizoguchi

Roles of orexin neurons in motivated behaviors in rats

10

P19

Shinya Fujii

Acute social defeat stress increases sleep in mice

10

P20

Kanako Iwasaki

Kinase activity of sleepy mutant SIK3 visualized by newly developed FRET probe

11

P21

Chi Jung Hung

P22

Sumire Matsumoto

Cortical Neuronal Communication During Natural Slow Wave Sleep in Mice

12

P23

Chihiro Suzuki

Prevalence of self-rated sleep apnea and related factors in a Japanese working population

12

P24

JaeHoon Seol

P25

Sakthivel Srinivasan

P26

Javier Diaz

P27

Hirotoshi Hiraishi

Functional Connectivity during Moral Judgment -MEG study-

14

P28

Iyo Koyanagi

Metabolomic screening for memory consolidation during sleep

15

P29

Liqin Cao

Psycho, a potential mouse model of schizophrenia

16

Cumulative phosphorylation of SNIPPs as the molecular substrate of homeostatic
sleep need
Dopaminergic activity-dependent astrocytic glycogenolysis producing lactate in theexercising hippocampus
Sex difference in sleep architecture, thermal regulation and energy metabolism in humanLarge Scale Forward Genetic Screening Identifies Trpa1 as a Chemosensor for Predator Odorevoked Innate Fear Behaviors
Rapid stimulation of hippocampal memory circuit with acute mild exercise
Where does the Dreamless mutation take effect?
- Generation of Nalcn-FLEx and Nalcn-flox Knock-In Mice
Impaired Sleep Architecture and Learning ability in an App Knock-in Mouse Model of
Alzheimer’s Disease

to young adult

Synthesis and evaluation of 18F-labeled tetrahydroisoquinoline derivatives as novel PET
probes for imaging orexin 1 receptors

Orexin and MCH neurons double ablated mice showed severe sleep abnormality andcataplexy

Different effects of orexin receptor antagonist and GABAA agonist on physical and cognitive
functions.
Reactivation of adult-born neurons for memory consolidation during sleep
Electroencephalographic analysis of the Sleepy (sik3slp/+, sik3slp/slp) mutant mice using theenvelope characterization space

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P01
We thank all Sakaguchi lab and WPI-IIIS members, K.G.
Akers, S. Yaghisita, Y. Hayashi, K. Kobayakawa, R.
Kobayakawa, M. Hayashi, X. Hayashi, M. Oishi, G.
Changarathil, S. Narita, Y. Hirose, M. Sezaki, T.
Hosokawa, K. Shikama, S. Sugimori, Y. Mimura, N.
Hasegawa, H. Obo for technical assistance, N.
Kobayashi and M. Adachi for secretarial assistance,
This work was partially supported by grants from theWorld Premier International Research Center Initiativefrom the Japan Ministry of Education, Culture, Sports,
Science, and Technology (MEXT); JST CREST Grant
Number JPMJCR1655; Japan Society for the Promotionof Science Grants-in-Aid for Scientific Research (JSPS,
KAKENHI grant numbers 16K18359, 15F15408,
26115502, and 25116530); The Takeda Science
Foundation; Shimadzu Science Foundation; Kanae
Foundation for the Promotion of Medical Science,
Research Foundation for Opto-Science and Technology;
The Ichiro Kanehara Foundation; Kato Memorial
Bioscience Foundation; Japan Foundation for Applied
Enzymology; Senshin Medical Research Foundation;
Life Science Foundation of Japan; Uehara Memorial
Foundation; Brain Science Foundation; Kowa Life
Science Foundation; and 2016 Inamori Research Grants
Program to MS, JSPS (KAKENHI grant numbers
25000015, 18H04012) to M.K, JSPS FPD; and the
University of Tsukuba to DK, the Tokyo Biochemical
Research Foundation to SSR, NIH-NCI CCSG: P30
014195, NINDS R24 Core Grant and NEI to SK.

Function of the adult-born neurons inmemory consolidation during sleep
Deependra Kumar1, Iyo Koyanagi1, *, Alvaro CarrierRuiz2,3, *, Pablo Vergara1, *, Sakthivel Srinivasan1, *,Yuki Sugaya2,3, Masatoshi Kasuya1, Tzong-Shiue Yu4,
Kaspar Vogt1, Masafumi Muratani5, Takaaki Ohnishi6,
Sima Singh1, Catia M Teixeira7, Yoan Chérasse1,
Toshie Naoi1, Szu-Han Wang8, Pimpimon Nondhalee1,
Boran Abdel Hamid Osman1, Nahoko Kaneko9,
Kazunobu Sawamoto9,10, Steven G Kernie4, Takeshi
Sakurai1, Thomas J McHugh11, Masanobu Kano2,3,Masashi Yanagisawa1, Masanori Sakaguchi1
1International Institute for Integrative Sleep Medicine (WPI-IIIS)
and 5Department of Genome Biology, Faculty of Medicine,
University of Tsukuba, Tsukuba, Ibaraki, 2Department of
Neurophysiology, Graduate School of Medicine and 3WPI-IRCN,UTIAS, 6Graduate School of Information Science and Technology,
The University of Tokyo, Tokyo, 4Departments of Pediatrics andPathology and Cell Biology, Columbia University College ofPhysicians and Surgeons, New York, NY, USA, 7Emotional Brain
Institute, Nathan Kline Institute, New York, NY, USA, 8Centre forClinical Brain Sciences, Univ. Edinburgh, UK, 9Department ofDevelopmental and Regenerative Biology, Nagoya City
University Graduate School of Medical Sciences, Nagoya, Aichi,
10
Division of Neural Development and Regeneration, National
Institute for Physiological Sciences, Okazaki, Aichi, 11RIKEN
Center for Brain Science, Wako, Saitama, *These authors
contributed equally

Mammalian sleep contains rapid eye movement (REM)
and non-REM sleep, each employs different
mechanisms for episodic memory consolidation.
Previous reports showed that boosting slow oscillations
or inhibiting sharp wave-ripples during non-REM sleep
potentiated or interfered with memory consolidation1,2.
On the other hand, disruption of REM-related theta
rhythm impaired memory consolidation3. However, thememory circuit that is responsible for memory
consolidation during each sleep stage has not been clear.
We have shown that hippocampal adult-born neurons
are incorporated in memory circuits after learning4.
Therefore, we silence the activities of the adult-bornneurons during specific stage of sleep after learning
using optogenetic intervention, which provides
reversibility, higher time resolution and target
specificity. It revealed that the activities of the adultborn neurons are necessary for memory consolidation
during sleep.
1. Marshall et al., Nature. 2006, v444, p610
2. Girardeau et al., Nat Neurosci, 2009, v12, p1222
3. Boyce et al., Science. 2016, v352, p812
4. Arruda-Carvalho et al., J Neurosci., 2011, v31, p15113
1

P02

P03

Cumulative phosphorylation of SNIPPs as
the molecular substrate of homeostatic
sleep need

Dopaminergic activity-dependent astrocytic
glycogenolysis producing lactate in theexercising hippocampus

Zhiqiang Wang1, Jing Ma1, Chika Miyoshi1, Yuxin Li2,
Makito Sato1, Yukino Ogawa1, Tingting Lou1, Chiyu
Lee1, Xiaojie Yang1, Noriko Hotta-Hirashima1, Daniela
Klewe-Nebenius1, Aya Ikkyu1, Miyo Kakizaki1, Satomi
Kanno1, Liqin Cao1, Junmin Peng2, Yonghao Yu3,
Hiromasa Funato1, Masashi Yanagisawa1, Qinghua
Liu1,3

Takashi Matsui1, 2, 3, Ignacio Torres-Aleman3, Hideaki
Soya1, 2
1

Laboratory of Exercise Biochemistry and Neuroendocrinology,
Faculty of Health and Sport Sciences, University of Tsukuba;
2
Sport Neuroscience Division, Advanced Research Initiative for
Human High Performance (ARIHHP), University of Tsukuba;
Tsukuba 305-8574, Ibaraki, Japan;
3
Neuroendorinology Laboratory, Cajal Institute, CSIC, Avda Dr
Arce 37, 28002 Madrid, Spain.

1

International Institute for Integrative Sleep Medicine (WPI-IIIS),
University of Tsukuba
2
St. Jude Proteomics Facility, St. Jude Children's Research
Hospital
3
University of Texas Southwestern Medical Center

Hippocampal lactate produced from astrocytic glycogenis a critical energy source and/or a neuromadulator inmemory functions. Although exercise, even in mild
intensity, activates hippocampal neurons and enhances
memory functions, hippocampal glycometabolism
during exercise is less clear. Here we reveal the
dopamine (DA) D2 receptor (R)-mediated astrocytic
glycogenolysis to produce lactate in the exercising rat
hippocampus. Running exercise decreased hippocampal
glycogen and increased its lactate in a running speeddependent manner associated with DAergic activation,
independent of noradrenaline and serotonin. DA
challenge and an agonism of DAD2R, but not DAD1R,
decreased glycogen via Ca2+/cAMP/PKA signaling inprimary cultured rat astrocytes, but DA did not serve inastrocytes from mice lacking DAD2R. An antagonism
of DAD2R, but not DAD1R, inhibited glycogenolysis
and lactate production in mild exercising hippocampus.
Our findings demonstrate the DAergic activitydependent astrocytic glycogenolysis producing lactate
in the running hippocampus, providing insights into
energetics and neurotrophicity underlying exerciseenhanced memory functions.

The sleep-wake homeostasis is maintained bygeneration of a sleep need that accumulates during
waking and dissipates through sleep. In mammals, thebest known measurable index of sleep need is the slow
wave activity (SWA) or EEG delta power (1-4 Hz)
during non-rapid-eye-movement sleep (NREMS). To
investigate the molecular basis of sleep need, we
performed quantitative phosphoproteomic studies of
two opposite models of increased sleep need in mice.
Sleep deprivation induces cumulative phosphorylationof brain proteome, which dissipates during recovery
sleep. Strikingly, Sleepy (Sik3Slp/+) mutant brains,with an elevated sleep need despite chronic
hypersomnia, exhibit a hyper-phosphoproteome
mimicking sleep-deprived brains, owing to a gain-offunction mutation of protein kinase SIK3. Comparisonof two models identified 80 mostly synaptic SleepNeed-Index-PhosphoProteins(SNIPPs),
whose
phosphorylation states closely parallel changes of sleep
need. Mutant SIK3 (SLEEPY) kinase preferentiallyassociated with and phosphorylated SNIPPs. Inhibitionof SLEEPY/SIK3 activity reduced phosphorylation
state of SNIPPs and SWA of NREMS in both Sleepy andsleep-deprived mice. Our results expose an unexploredbrain phosphoproteome landscape and suggest that
SNIPPsgradually
accumulate/dissipate
phosphorylation as a function of homeostatic sleep need.
While waking encodes memories by potentiating
synapses, sleep consolidates memories and restoressynaptic homeostasis by globally downscaling
excitatory synapses. Thus, we propose that
phosphorylation and dephosphorylation of SNIPPs may
represent a major regulatory mechanism that underlies
both synaptic homeostasis and sleep-wake homeostasis.
2

P04

P05

Sex difference in sleep architecture, thermal
regulation and energy metabolism in humanLarge Scale Forward Genetic Screening
Identifies Trpa1 as a Chemosensor for
Predator Odor-evoked Innate Fear Behaviors

Simeng Zhang1, Haruka Oosumi2, Akiko Uchizawa2,
Haruka Hamada3, Insung Park1, Yoko Suzuki1, Yoshiaki
Tanaka2, Asuka Ishihara1, Katsuhiko Yajima2,4, Makoto
Satoh1, Naomi Omi2, Kumpei Tokuyama2

Chia-Ying Lee3, Yibing Wang1,2, Liqin Cao3, TomohikoMatsuo4, Kejia Wu1,2, Greg Asher3, Lijun Tang4, Tsuyoshi
Saitoh3, Jamie Russell5, Daniela Klewe-Nebenius3, Li
Wang2, Shingo Soya3, Emi Hasegawa3, Yoan Chérasse3,
Jiamin Zhou2, Yuwenbin Li2, Tao Wang5, Xiaowei Zhan5,Chika Miyoshi3, Yoko Irukayama3, Jie Cao6, Julian P.
Meeks6, Laurent Gautron7, Zhiqiang Wang3, Katsuyasu
Sakurai3, Hiromasa Funato 3,8, Takeshi Sakurai3, Masashi
Yanagisawa 3,9, Hiroshi Nagase3, Reiko Kobayakawa4, Ko
Kobayakawa4, Bruce Beutler5 & Qinghua Liu1,2,3,5,6,10

1

International Institute for Integrative Sleep Medicine, Universityof Tsukuba
2Graduate School of Comprehensive Human Sciences, Universityof Tsukuba
3
School of Health and Physical Education, University of Tsukuba
4
Faculty of Health and Nutrition, Tokyo Seiei College

1

National Institute of Biological Sciences, 102206 Beijing, China.
Department of Biochemistry, University of Texas Southwestern
Medical Center, Dallas, TX 75390, USA. 3International Institutefor Integrative Sleep Medicine (WPI-IIIS), University of Tsukuba,
Tsukuba, Ibaraki 305-8575, Japan. 4Functional Neuroscience
Lab, Kansai Medical University, Hirakata, Osaka 573-1010,
Japan. 5Center for Genetics of Host Defense, University of Texas
Southwestern Medical Center, Dallas, TX 75390, USA.
6
Department of Neuroscience, University of Texas Southwestern
Medical Center, Dallas, TX 75390, USA. 7Department of Internal
Medicine, University of Texas Southwestern Medical Center,
Dallas, TX 75390, USA. 8Department of Anatomy, Faculty of
Medicine, Toho University, Ota-Ku, Tokyo 143-8540, Japan. 9Life
Science Center, Tsukuba Advanced Research Alliance, Universityof Tsukuba, Tsukuba, Ibaraki 305-8575, Japan. 10Tsinghua
Institute of Multidisciplinary Biomedical Research, Tsinghua
University, 100084 Beijing, China.

Human sleep is generally consolidated into a single
prolonged period, and its metabolic consequence is to
impose an extended period of fasting. The energy
metabolism during sleep does not simply reflect theextension of fasting time. It has been assumed thatoxidized substrate during sleep shift from carbohydrate
to fat as sleep continues, but the time course of
respiratory quotient (RQ) indicates that carbohydrateoxidation begins to increase prior to awake. During
sleep, core body temperature decreases followed by agradual increase before wake. Gender difference in timecourse of core body temperature during sleep is known:
nadir of core body temperature appears earlier in female
than male.
The purpose of this study is to confirm whether there
is a gender difference in the time course of energy
metabolism during sleep. Energy metabolism wasmeasured by a whole room indirect calorimetry withimproved time resolution, core body temperature andskin temperature was also monitored during sleep. Sleep
was monitored using a standard PSG recording.
Sleep architecture was similar between male andfemale, and between follicular and luteal phase. Timecourse of RQ during sleep showed a gender difference,and nadir of RQ appeared earlier in female than male.
Core body temperature during the luteal phase washigher than that of the follicular phase. In the present
study heat production (energy expenditure) was similar
between follicular and luteal phase, but heat loss,
estimated from the distal-proximal gradient of skintemperature, was smaller during the luteal phase.

2

Innate fear behaviors have been believed to begenetically encoded in all organisms. But theirunderlying molecular mechanisms remain largely
unknown. Predator odor 2,4,5-trimethyl-3-thiazoline(TMT) and its potent analogue 2-methyl-2-thiazoline(2MT) elicit innate fear/defensive behaviors in naive
mice. We conduct a large-scale recessive genetics
screen based on the innate defensive responses to 2MT
in ethylnitrosourea (ENU)-mutagenized mice. We findthat Trpa1, a pungency/irritancy receptor, is related toboth 2MT and snake skin-evoked innate fear/defensive
responses. Accordingly, Trpa1-knocknout mice
(TRPA1 KO) are unable to effectively activate known
fear/stress brain centers upon 2MT exposure, despite
their apparent ability to smell and learn to fear 2MT.
Moreover, in vitro calcium influx tests indicate Trpa1
acts as a chemosensor for 2MT. Interestingly we findtrigeminal ganglia (TG) neurons are highly activated in
2MT stimulations. Unilateral lesion of TG reduced
innate freezing and TG specific AAV-Trpa1-expression
rescued freezing behavior in TRPA1 KO mice under
2MT exposure. These results suggest trigeminal
ganglion neurons contribute critically to 2MT-evoked
freezing.
3

P06

P07

Rapid stimulation of hippocampal memory
circuit with acute mild exercise

Where does the Dreamless mutation takeeffect? - Generation of Nalcn-FLEx andNalcn-flox Knock-In Mice

Kazuya Suwabe1,2, Kyeongho Byun2,3, Kazuki Hyodo1,
Zachariah M. Reagh3, Jared M. Roberts3, Akira
Matsushita4,5, Kousaku Saotome4, Genta Ochi1,
Takemune Fukuie1, Kenji Suzuki4, Yoshiyuki Sankai4,
Michael A. Yassa2,3, and Hideaki Soya1,2

Tomoyuki Fujiyama1, Seiya Mizuno2, Manabu Abe3,
Satomi Kanno1, Miyo Kakizaki1, Kanako Iwasaki1, Aya
Ikkyu1, Noriko Hotta-Hirashima1, Mana Yamada1,
Chika Miyoshi1, Makito Sato1, Takeshi Kanda1, Kenji
Sakimura3, Satoru Takahashi2, Hiromasa Funato1,4,Masashi Yanagisawa1,5

1

Laboratory of Exercise Biochemistry and Neuroendocrinology
Faculty of Health and Sport Sciences, University of Tsukuba
2Sports Neuroscience Division, Advanced Research Initiativefor Human High Performance (ARIHHP), Faculty of Healthand Sport Sciences, University of Tsukuba
3
Department of Neurobiology and Behavior, Center for the
Neurobiology of Learning and Memory, University ofCalifornia, Irvine
4Center for Cybernics Research, University of Tsukuba, Ibaraki
305-8574, Japan
5
Department of Neurology, Ibaraki Prefectural University of
Health Sciences

1

International Institute for Integrative Sleep Medicine (WPI-IIIS),
University of Tsukuba, Ibaraki, Japan
2
Laboratory Animal Resource Center, University of Tsukuba,
Ibaraki, Japan3
Department of Cellular Neurobiology, Brain Research Institute,
Niigata University, Niigata, Japan
4
Department of Anatomy, Faculty of Medicine, Toho University,
Tokyo, Japan
5
Department of Molecular Genetics, University of Texas
Southwestern Medical Center, Dallas, TX, USA

Physical exercise has beneficial effects onneurocognitive function, including hippocampusdependent episodic memory. Exercise intensity levelcan be assessed according to whether it induces a stressresponse, the most effective exercise for improving
hippocampal function remains unclear. Our prior works
using special treadmill running model in animals haveshown that stress-free mild exercise increases
hippocampal neuronal activity and promotes adultneurogenesis in the dentate gyrus (DG) of the
hippocampus, improving spatial memory performance.
However, the rapid modification from mild exercise on
hippocampal memory function and exact mechanisms
for these changes, in particular the impact on patternseparation acting in the DG and CA3 regions are yet tobe elucidated. To this end, we adopted an acute-exercise
design in human, coupled with high-resolutionfunctional MRI techniques, capable of resolving
hippocampal subfields. A single 10-minute bout of very
light intensity exercise (30%VO2peak) results in rapid
enhancement in pattern separation and an increase infunctional connectivity between hippocampal DG/CA3
and cortical regions (i.e., parahippocampal, angular, andfusiform gyri). Importantly, the magnitude of theenhanced functional connectivity predicted the extent ofmemory improvement at an individual subject level.
These results suggest that brief, very light exercise
rapidly enhances hippocampal memory function,
possibly by increasing DG/CA3-neocortical functional
connectivity.

Although sleep is a ubiquitous animal behavior, themolecular/neural basis mechanism of REM sleep(REMS) remains unknown. We performed highthroughput screening of ENU-mutagenized mice inorder to identify genes regulating sleep/wake behavior,and established the Dreamless mutant pedigree thatshows about 50% reduction in 24-h REMS time. We
identified a nucleotide change specific to Dreamlessmutant mice within the exon9 in Nalcn gene. The single
nucleotide substitution leads to a single amino acidsubstitution (N315K) of the gene product leak cation
channel, NALCN, that we termed “Dreamless”. We
adopted CRISPR/Cas system to recapitulate Dreamlessphenotype, then confirmed that the one base substitution
was responsible for REMS abnormality, suggesting thatthe identified gene is related to the regulation of daily
REMS time. To elucidate the molecular/neural basis of
REMS regulation by NALCN, we generated
genetically-modified Nalcn mutant mice bearing flox
knock-in and FLEx (flip and excision) knock-in micefor loss-of- and gain-of-function studies, respectively.
Now we are planning to analyze sleep and wakefulness
by using these two mutant mouse lines.

4

P08

P09

Impaired Sleep Architecture and Learning
ability in an App Knock-in Mouse Model of
Alzheimer’s Disease

Dissection of prefrontal cortex circuit in
guiding the extinctions

1,2

Akira Uematsu1, Mika Hamada-Uematsu2, Tomoya
Duenki1, Joshua Johansen1,3

1

Sakura Eri Maezono , Mika Kanuka , Chika
Tatsuzawa1, Miho Morita1, Mitsuaki Kashiwagi1,
Pimpimon Nondhalee1, Masanori Sakaguchi1, Takashi
Saito3, Takaomi Saido3, Yu Hayashi1

1

RIKEN Center for Brain Science
Department of Surgery and Bioengineering, Advanced ClinicalResearch Center, Institute of Medical Science, University ofTokyo
3
Department of Life Sciences, Graduate School of Arts andSciences, University of Tokyo
2

1

International Institute for Integrative Sleep Medicine, Universityof Tsukuba.
2
School of Integrative and Global Majors, University of Tsukuba
3
Laboratory for Proteolytic Neuroscience, RIKEN Center for
Brain Science

Emotional memory is crucial to guide behaviors forindividual’s survival. Organisms need to respond to
stimuli that predict dangerous or desirable outcomes.
When this contingency changes, organism canextinguish emotional memory, the process calledextinction. Extinction is essential for mental health inboth aversive and appetitive domains and malfunctionof extinction results in mental disorders. More thandecade of research indicates that the ventromedial
subregion of the prefrontal cortex (vmPFC) is importantfor extinctions of both appetitive and aversive memories.
Anatomically the vmPFC sends broad projections andeach set of projecting neurons forms distinct populationin vmPFC. However, little is known about how each
population or combination of them encodes aversive
and appetitive extinction and how they differentiate
distinct forms of extinction. To understand thesequestions, we first performed a series of optogenetic
experiments by targeting 5 projection-specificpopulations in rats. Animals were learned to associate
auditory cue with sucrose solution or foot shock andthen presented auditory cue alone repeatedly with
optogenetic manipulation. We found that each
population has distinct role on appetitive and aversive
memory extinctions. Next, to examine neuralrepresentations of each population, we recorded calcium
dynamics in vmPFC excitatory neurons with miniature
microscope and tracked same neurons during appetitive
and aversive extinctions. Our preliminary data showthat vmPFC neurons show orthogonal coding for
appetitive and aversive extinctions with the populationlevel analysis. Further results will be discussed in theposter.

Alzheimer’s disease (AD) is the major cause ofdementia. It is a neurodegenerative diseasecharacterized by aggregation of amyloid-beta (Aβ)
peptides. In addition to cognitive decline, AD patients
often suffer from sleep disorders. These symptoms not
only affect their quality of life (QOL) but also those of
the primary caregivers. Moreover, increasing studies
support that sleep deficits can contribute to theprogression of AD, consistent with the roles of sleep inmemory and brain homeostasis. Thus, it is crucial tounderstand the underlying mechanisms of sleep
disorders in AD.
Here, we unveil the sleep abnormalities exhibited by an
AD mouse model. We chose the App knock-in mouse(Saito et al., Nature Neuroscience, 2014). In this AD
mouse model, Aβ accumulates in a manner similar tohuman AD via mutation in the gene encoding amyloidbeta precursor protein (APP), and thus, various aspectsof human AD are faithfully recapitulated. In these mice,at 6 months of age, when cognitive deficits become
apparent, impairment in various aspects of the sleep
architecture were detected. These phenotypes became
more severe at 12 months of age. Consistent with thenotion that sleep is important for memory consolidation,
the AD mouse model also exhibited learning deficits.
We expect that our results provide a start point foraddressing how sleep is impaired in AD and how it canaffect cognitive functions.

5

P10

P11

Longitudinal EEG/EMG measurement inmice show sleep/wakefulness maturationfrom infant to young adultSleep/wakefulness in neuron-specific SIK3deficient mice
Fuyuki Asano1, Tomoyuki Fujiyama1, Chika Miyoshi1,
Miyo Kakizaki1, Noriko Hotta-Hirashima 1, Aya Ikkyu1,
Satomi Kanno1, Seiya Mizuno2, Fumihiro Sugiyama2,
Satoru Takahashi1,2, Hiromasa Funato1,3 and Masashi
Yanagisawa1,4,5

Noriko Hotta-Hirashima¹, Chika Miyoshi¹, Aya Ikkyu¹,
Satomi Kanno¹, Hiromasa Funato¹ ², Masashi
Yanagisawa¹
1
2

WPI-IIIS Univ Tsukuba
Toho Univ Sch Med

1

WPI-IIIS, University of Tsukuba
Laboratory Animal Science, Faculty of Medicine, University ofTsukuba3
Department of Anatomy, Toho University
4
Molecular genetics, UT Southwestern Medical Center5
Tsukuba Advanced Research Alliance, University of Tsukuba
2

Although it is postulated that sleep/wakefulness
behavior changes along with the postnatal developmentof brain, there have been very few studies on
EEG/EMG-based sleep study during infancy due totechnical difficulties in obtaining EEG/EMG signalingfrom a very small pups during breast-feeding. Here, wehave established EEG/EMG electrode implantation
surgery for pre-weaned mice (postnatal days P12) and arecording system optimized for breast-fed pups. We
obtained 24h EEG/EMG signaling from the same mouseonce a week from P15 to P57, and at P70. We observed
a drastic change in sleep/wake pattern between P15 andP22. At P15, sleep/wake stages were very unstable andrarely continued for longer than 10 seconds so that itwas very difficult to stage reliably. At P17, the durationof each sleep/wake stage became more stable and longer,
which enabled us to determine sleep stage. The
circadian change in sleep/wake behavior was getting
obvious towards P36. We also observed the upward shiftof the theta wave peak during REM sleep toward P29.
Thus, the maturation of sleep/wakefulness progresses
during infancy and after weaning with a different timecourse depending on sleep parameters.

Sleep is a fundamental behavior conserved fromvertebrates to invertebrates. SIK3, a member of AMPactivated protein kinase (AMPK) family, was identified
as a novel sleep-regulating molecule. Alanine
substitution at serine551 in SIK3 resulted in longer
NREM sleep time with increased NREM sleep deltadensity, which indicates an enhanced sleep need in
Sik3(S551A) mice. Conversely, flies and round worms
expressing a hypomorphic SIK3 orthologue reduced
sleep-like behavior. Therefore, the long sleep phenotypeof Sik3(S551A) mice may be due to a gain-of-function
effect of the mutant SIK3 protein. However, the effectof the endogenous SIK3 protein on sleep/wakefulness
regulation in mice is still unknown because almost all of
Sik3-deficient mice died on the day of their birth and a
few survived mice suffered from severe growth
retardation and malnutrition.
Here, we established Sik3-flox mice, in which exon 3 ofSik3 gene is flanked by lox P sites. Cre recombinase
excises the exon 3, which results in frameshift andcreation of premature stop codon. Nestin-Cre;
Sik3(flox/flox) mice are healthy and fertile. They didn’t
have detectable SIK3 protein in their brain. We
examined sleep/wakefulness in neuron-specific SIK3deficient mice. They showed a decreased sleep need.

6

P12

P13

Forgetting during REM sleep by
hypothalamic MCH neurons

Adenosinergic mechanisms of sleep control in
the nucleus accumbens

Shuntaro Izawa1, Ryo Inoue1, Srikanta Chowdhury1,
Yasutaka Mukai1, Daisuke Ono1, Akihiro Yamanaka1,2

Xuzhao Zhou, Yoan Cherasse, Yo Oishi and Michael
Lazarus1

International Institute for Integrative Sleep Medicine (IIIS),
University of Tsukuba

Research Institute of Environmental Medicine- Nagoya
University, Neuroscience Research, Nagoya, Japan.
2Start of Core Research for Evolutional Science and Technology
CREST, Japan Science and Technology Agency JST, Tokyo, Japan.

We have revealed a prominent role of adenosine A2A
receptor (A2AR)-expressing neurons in the nucleus
accumbens (NAc) in sleep/wake regulation andproposed a novel brain circuit for sleep control bymotivated behavior (Oishi Y., et al., Nat. Commun.,
8:article 734, 2017). This brain circuit may explain thetendency to fall asleep in the absence of motivating
stimuli, i.e., when bored. We hypothesized that theability of the NAc to induce sleep is mediated by theclassic somnogen adenosine. Adenosine can be formedby various processes in all types of cells but the neurodynamics of adenosine (release, receptor activation,etc.) in controlling sleep are still unclear. In this study,
we ablated glial fibrillary acidic protein (GFAP)positive cells in the NAc of mice by virus-mediatedexpression of diphtheria toxin (DT) receptors andintraperitoneal administration of DT. After analyzing
electroencephalogram and electromyogram recordingsof the mice, we found a remarkable increase in slowwave sleep (SWS) one week after DT treatment that was,surprisingly, accompanied by an increased number of
GFAP cells in the NAc. In-vivo microdialysis one week
after DT treatment revealed a significant increase of
extracellular adenosine in the NAc. Moreover, we foundthat chemogenetic activation of NAc GFAP cells
strongly increased the total amount of SWS. Our current
results may suggest that adenosine from NAc glial cells
plays an important role for sleep control.

Sleep is assumed to not only consolidate memory but
also impair memory. However, the neural mechanisms
behind it has been unclear. Here we found that
hypothalamic melanin concentrating hormone (MCH)
producing neurons impair memory via the hippocampus
during rapid eye movement sleep (REMS).
Using retrograde tracing beads injection into the
hippocampus, we found that MCH neurons have dense
projections from the hypothalamus to the hippocampus.
MCH neurons are known to regulate REMS, and we
identified that MCH neurons were active during REMS
by using fiber photometry calcium imaging.
To reveal the role of MCH neurons in hippocampusdependent memory, MCH neurons manipulated mice
were subjected hippocampus-dependent memory tests.
The pharmacogenetic activation of MCH neurons withhM3Dq impaired memory ability, whereas theinhibition with hM4Di improved it. MCH neuronsablation also induced memory improvement, suggesting
MCH neuronal activity induces memory impairment.
Optogenetic stimulation of MCH neurons reduced
memory score with the stimulation during memory
retention, but not encoding nor retrieval period. The
slice patch clamp recording in the hippocampus showedthat the light stimulation of MCH nerve terminals
hyperpolarized and decreased firing frequency ofhippocampal pyramidal neurons. In the behavior tests,
memory ability was impaired by the optogenetic
stimulation of MCH nerve terminals in the hippocampus.
During memory retention period, MCH neurons
inhibitionineach
sleep-wakefulness
states(Wakefulness, REMS, and Non-REMS) were
performed with optogenetic inhibition triggered by theautomatic sleep staging system. The inhibition during
REMS improved memory ability compared with
Wakefulness or Non-REMS, indicating MCH neurons
induce forgetting during REMS.

7

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P15

Exploration of the intracellular molecularmechanisms for homeostatic sleep/wakeregulation

Synthesis and evaluation of 18F-labeled
tetrahydroisoquinoline derivatives as novel
PET probes for imaging orexin 1 receptors

Tomohiro Kitazono1, Taeko Matsuoka1, Aya Ikkyu1,
Zhiqiang Wang1, Jing Ma1, Hiromasa Funato1,2,Masashi Yanagisawa1,3

Hiroyuki Watanabea, *, Kengo Fukuia, Yoichi Shimizua,b,Hideo Sajia, Masahiro Onoa,*a

Department of Patho-Functional Bioanalysis, Graduate Schoolof Pharmaceutical Sciences, Kyoto University, 46-29 Yoshida
Shimoadachi-cho, Sakyo-ku, Kyoto 606-8501, Japan.
b
Department of Diagnostic Imaging and Nuclear Medicine,
Graduate School of Medicine, Kyoto University, 54 Shogoin
Kawahara-cho, Sakyo-ku, Kyoto 606-8507, Japan.

1

WPI-IIIS, Univ of Tsukuba,
2
Dept of Anat, Sch of Med, Toho Univ,
3
HHMI, Univ of Texas Southwestern Medical Center at Dallas

Sleep is a ubiquitously conserved behavior in animals
with central nervous system, but intracellularmechanisms regulating sleep/wake homeostasis remain
unknown. Our previous study suggested that SIK3
protein kinase plays a central role in sleep/wakeregulation (Funato et al., 2016). Here, we further
analyzed the upstream and downstream regulatory
mechanisms involving SIK3. 1) Upstream analysis:
Ser551 is one of the phosphorylation sites of SIK3, andwe recently found that phosphorylation of this site wasimportant in sleep/wake regulation (Honda et al., 2018).
Therefore, we investigated the regulatory mechanismsof S551 phosphorylation in SIK3, and found that PKA
could phosphorylate this site, and phosphorylation of
PKA could be also regulated by SIK3. 2) Downstream
analysis: Although we previously reported that asplicing mutant of Sik3 gene, Sleepy mutant brains
exhibited a markedly altered phosphoproteomic status
and mimicked sleep-deprived brains (Wang et al., 2018),
the downstream signaling pathway of SIK3 in sleep
regulation is still unclear. We recently found that Sleepy
and null mutant mice of Sik3 showed opposite
phenotypeinsleep,
thus,
weexamined
phosphoproteomic status of Sik3 null mutant brain. We
found ~200 phosphorylation sites showing significant
phosphorylation state change in both mutant brains, and,surprisingly, phosphorylation of almost all sites
increased in both. In addition, we examined brain
interactome of SLEEPY2, which was a novel regulatorof sleep homeostasis found in our genetic screening, andfound that SLEEPY2 was highly associated withmultiple cell adhesion molecules. These findings will
enable us to provide new insights on the homeostatic
sleep regulation.

Introduction: Orexin 1 receptor (OX1R) is thought tobe involved in various body functions, including arousal
maintenance and emotional control, but the full detailsof its function remain unknown. OX1R imaging withpositron emission tomography (PET) would be useful inelucidating the orexin system including OX1R.
However, no PET probes targeting OX1R have beenreported. Therefore, we designed and synthesized
tetrahydroisoquinoline (THIQ) derivatives as novel
PET probes targeting OX1R, and evaluated their utility.
Methods: We succesfully synthesized THIQ-1 and
THIQ-2. To quantify their affinities for OX1R and
OX2R, we carried out in vitro competitive binding
assays using OX1R or OX2R expression cells and[125I]Orexin A. After radiosynthesis of [18F]THIQ-1 and[18F]THIQ-2, we investigated the cellular binding of[18F]THIQ-1 and [18F]THIQ-2 to cell expressing OX1R
or OX2R. Finally, to evaluate the brain uptake of[18F]THIQ-1 and [18F]THIQ-2, we performed abiodistribution study in normal mice.
Results: In an in vitro competitive binding assay, THIQ1 and THIQ-2 showed significantly higher binding to
OX1R (IC50 = 30 and 31 nM, respectively) than OX2R
(IC50 = 160 and 332 nM, respectively). These features
were also observed in the cell binding assay using[18F]THIQ-1 and [18F]THIQ-2, demonstrating their
OX1R-specific binding property in vitro. In thebiodistribution study using normal mice, [18F]THIQ-1
and [18F]THIQ-2 showed brain uptake at 2 min postinjection, but the uptake levels were not sufficient for invivo imaging with PET.
Conclusion: [18F]THIQ-1 and [18F]THIQ-2 have thepotential to become useful imaging probes for PET
targeting the OX1R.

8

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P17

Search for neural circuits of narcolepsycataplexy

A neuronal subpopulation in the bed nucleusof the stria terminalis regulates NREM sleep

Emi Hasegawa1, Takashi Maejima4, Takayuki Yoshida5,
Mitsuhiro Yoshioka5, Masashi Yanagisawa1,2,3,
Michihiro Mieda4, Takeshi Sakurai1,2,3

Yoan Cherasse, Olusolape Ladenika, Gabrielle Boix,
Yoshihiro Urade, Takeshi Sakurai
International Institute for Integrative Sleep Medicine (WPI-IIIS),
University of Tsukuba, Tsukuba, Japan

1

International Institute for Integrative Sleep Medicine;2
Faculty of Medicine
3Life Science Center for Tsukuba Advanced Research Alliance(TARA), University of Tsukuba, Japan
4
Department of Molecular Neuroscience and Integrative
Physiology, Graduate School of Medical Sciences, Kanazawa
University, Japan
5
Department of Neuropharmacology, Hokkaido University
Graduate School of Medicine, Japan.

Zinc is an essential mineral that plays an important rolein the body by acting as a cofactor for more than 300
enzymes and 1000 transcription factors. While zinc is
naturally present in food, inappropriate or insufficientfeeding behavior put more than 25% of the world
population at risk of zinc deficiency. The purpose of ourstudy was to examine the effect of zinc on sleep.
We examined the sleep-promoting activity of zinc bymonitoring
locomotor
activityandelectroencephalogram after oral administration to mice.
Zinc-containing yeast extract dose dependently
increased the total amount of non-rapid eye movement
sleep.
C-fos mRNA expression has been monitored 1 hour
after zinc administration by in situ hybridization. We
could observe a specific activation of neurons in the bed
nucleus of the stria terminalis (BNST). The specificactivation of these c-fos positive neurons bychemogenetics (DREADD-hM3Dq) or optogenetics(ChR2) led to a similar increase of NREM sleep, while
their inhibition (DREADD-hM4Di) resulted in an
inhibition of the sleep promoting effect of zinc.
Preliminary mapping experiments confirmed ananatomical connection between zinc-sensitive neurons
in the BNST and another population of neurons in the
medial septum (MS). The exact role of this BNST-MS
pathway in the regulation of NREM sleep is now underinvestigation.

Mice lacking orexin peptides, orexin neurons, or orexin
receptors recapitulate human narcolepsy phenotypes,
further highlighting a critical role for orexin signaling in
the maintenance of wakefulness. However, although thelack of orexin signaling causes the sleep disordernarcolepsy, the precise neural mechanisms by which
orexin neurons prevent narcolepsy remain unclear. In aprevious study, we found that targeted restoration oforexin receptor expression in the dorsal raphe (DR) andin the locus coeruleus (LC) of mice lacking both oforexin receptors inhibited cataplexy and pathologicalfragmentation of wakefulness (i.e., sleepiness),
respectively. These results suggested that DR
serotonergic and LC noradrenergic neurons playdifferential roles in orexin neuron-dependent regulationof sleep/wakefulness. As a next step, we used
optogenetic and chemogenetic approaches todemonstrate that DR serotonin neurons suppress
cataplexy by reducing the activity of thebasolateral/lateral amygdala that plays an important rolein emotional processing, as consistent with the fact thatstrong emotion often triggers cataplexy. Our results
suggest that the orexin neuron–DR serotonin neuron–
amygdala pathway is a critical circuit for preventing
cataplexy. Furthermore, we identified a neuronal
pathway that induces cataplexy when activated by
optogenetic manipulation. We will discuss the role ofthis pathway in emotional processing as well as in
REM-related muscle atonia.

9

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P19

Roles of orexin neurons in motivatedbehaviors in rats

Acute social defeat stress increases sleep inmice

Hiroyuki Mizoguchi1, Ayumu Inutsuka2, Kentaro
Katahira3, Kiyofumi Yamada4, Akihiro Yamanaka5

Shinya Fujii1, Mahesh K. Kaushik1, Xuzhao Zhou1,
Mustafa Korkutata1, and Michael Lazarus1

1

1Res. Ctr. Next-Generation Drug Dev., Res. Inst. Environmental
Med., Nagoya Univ. , Nagoya, Japan.
2
Dept. Physiol., Jichi Med. Univ., Shimotsuke, Japan.
3
Dept Psychol, Grad Sch Inform, Nagoya Univ, Nagoya, Japan4
Dep. Neuropsychopharmacol. Hosp. Pharm., Nagoya Univ.
Grad. Sch. Med. ,Nagoya, Japan.
5
Dept. Neuroscience II, Res. Inst. Environmental Med., Nagoya
Univ., Nagoya, Japan.

International Institute for Integrative Sleep Medicine (WPI-IIIS),
University of Tsukuba, Tsukuba, Japan

Social stress is a major risk factor for psychiatricdisorders. Social defeat stress (SDS) is often induced inanimal models to study mood and anxiety disorders.
Stress leads to insomnia, presumably by making it
difficult to fall asleep and affecting sleep quality. The
effect of SDS on sleep/wake behavior is not wellunderstood. We established a mouse model of acute
SDS based on strong aggressive mouse behavior toward
unfamiliar intruders. In response to social defeat, slowwave sleep (SWS) strongly increased for 9 h, whereas
rapid eye movement sleep increased 6 h after the stressevent for following 3 h. Slow-wave activity was only
enhanced for a short period after social defeat anddissipated long before the SWS returned to baseline.
Moreover, social defeat evoked a strong corticosterone
response that may indicate a high stress level in the
intruder mice after social defeat. These findings suggestthat SDS increases sleep in mice and thus, insomnia
does not appear to be a consequence of social stress inmice. Our social defeat model may be useful forstudying the mechanisms and function of sleep in
response to social stress.

Orexin neurons regulate physiological functions,
including not only energy homeostasis and wakefulness,but also motivated behaviors. These neurons seem toplay important roles in linking metabolic need tomotivated behaviors via the dopaminergic system.
Recently, we developed a rat model that expresses the
Cre recombinase specifically in orexin neurons.
Moreover, we established the gambling test forassessing reward motivation and decision-making underconditions of uncertainty, as well as a touch-screen
system for assessing reward sensitivity and craving.
Here, we examined the roles of orexin neurons inreward motivation and decision-making when orexin
neurons are manipulated using pharmacogenetics
approaches. In the gambling test for rodents, cellspecific excitatory manipulation of orexin neurons ofrats using the DREADD technology resulted in risky
arm-choice.
Positive, but not negative, reward
prediction error may contribute to reward-based risky
choice when orexin neurons are activated.
The
motivational values of a large reward was increased
when orexin neurons were activated by the DREADD
technology. In the probability reversal learning test,
cell-specific excitatory manipulation of orexin neurons
resulted in low performance in rats. These results
suggest that activated orexin neurons affectmotivational processes, and may alter strategy inreward-based choice behavior, potentially leading to an
action that fails to yield rewards and detect changes inreinforcement contingencies.

10

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P21

Kinase activity of sleepy mutant SIK3
visualized by newly developed FRET probe

Orexin and MCH neurons double ablated
mice showed severe sleep abnormality andcataplexy

Kanako Iwasaki1, Takeshi Kanda1, Michiyuki Matsuda2,
Hiromasa Funato1,3, and Masashi Yanagisawa1

Chi Jung Hung1,2,3, Daisuke Ono1,2,3 and Akihiro
Yamanaka1,2,3

1International Institute for Integrative Sleep Medicine, Universityof Tsukuba
2
Laboratory of Bioimaging and Cell Signaling, Graduate schoolof biostudies, Kyoto university
3
Department of Anatomy, Toho University school of medicine

1

Department of Neuroscience II, Research Institute ofEnvironmental Medicine, Nagoya University, Nagoya 464-8601,
Japan
2
Department of Neural Regulation, Nagoya University Graduate
School of Medicine, Nagoya 466-8550, Japan
3
CREST, JST, Honcho Kawaguchi, Saitama 332-0012, Japan

Salt inducible kinase (SIK) family, one of the AMPactivated protein kinase (AMPK) related kinases, has 3paralogs in vertebrate animals. Among SIK family,
SIK3 is expressed in many organs including liver,skeleton, and brain, and known to be essential for
glucose/lipid metabolism and skeletal development.
Although the function of SIK3 in the brain had been
virtually unstudied, forward-genetics analysis in
randomly mutagenized mice disclosed the role of SIK3
in regulating sleep amount. A chemically induced
nucleotide substitution at Sik3 splice donor site caused
exon 13-skipping, resulted in prolonged non-rapid eye
movement (NREM) sleep. The exon 13-encoded regionof SIK3 encompasses protein kinase A (PKA)
recognition site, but how the PKA recognition site affectthe kinase activity is unknown.
Here, we newly developed Förster resonance energy
transfer (FRET) probe that enable us to monitor spatiotemporal change of SIK3 kinase activity in living cells.
We transfected the probe with mutant SIK3 such as PKA
recognition site-deficient SIK3 to cultured cells andexamined how the kinase activity is modulated by PKA
signaling. In addition, we compared the basal kinase
activity between wild type and PKA recognition sitedeficient SIK3 in each cells with normalization ofamount of SIK3.

The lateral hypothalamus (LH) is implicated in theregulation of instinctive behaviours, including sleep.
Orexin-producing neurons (orexin neurons) andmelanin-concentrating hormone (MCH)-producing
neurons (MCH neurons) are exclusively distributed in
the LH and project to whole brain to regulate
sleep/wakefulness. Ablation of orexin neurons
decreased wakefulness in the dark period and resulted
in narcolepsy, whereas ablation of MCH neurons
increased wakefulness. These indicated that orexin and
MCH neurons play a different role in the regulation of
sleep/wakefulness. Nevertheless, it is unclear how theinteractions between orexin and MCH neurons affectsleep/wakefulness. To understand the functional
interaction between orexin and MCH neurons and its
regulation of sleep/wakefulness, transgenetic mice inwhich both orexin and MCH neurons are ablated by the
Tet-off system were used. These orexin and MCH(double) ablation mice increased wakefulness anddecreased non-rapid eye movement (NREM) sleep. Thiswas similar to MCH neurons ablated mice. However, in
addition to these, total time in REM sleep also
diminished in double ablated mice. Total time andduration of cataplexy were exacerbated in double
ablated mice compare with orexin neurons ablated mice.
These results suggest that MCH neurons have a
suppressive role in cataplexy. Interestingly, these mice
showed new state, which was not classified withconventional criteria of sleep/wakefulness or cataplexy.
Here we defined as delta/theta (DT) sleep. DT sleep wasonly observed in the double ablated mice during
wakefulness as a sudden arresting behaviour with highEEG density of δ and θ wave with low EMG power.
EEG spectrum analysis revealed that DT sleep was
similar to NREM sleep just before transition to REM
sleep. Taken together, MCH neurons might be involvedin the regulation of NREM to REM sleep transition.

11

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P23

Cortical Neuronal Communication During
Natural Slow Wave Sleep in Mice

Prevalence of self-rated sleep apnea andrelated factors in a Japanese working
population

Sumire Matsumoto1,2, Kaoru Ohyama2,3, Javier Díaz2,
Robert Greene2,4, Kaspar Vogt2

Chihiro Suzuki1,2, Emi Morita2,3, Asuka Ishihara2,4,
Sumire Matsumoto2,4, Yu Ikeda5, Mami Ishitsuka6,
Daisuke Hori7, Shotaro Doki7, Yuichi Oi7, Shinichiro
Sasahara7, Ichiyo Matsuzaki2,7, Masashi Yanagisawa2,
Makoto Satoh2

1

Ph.D. Program in Human Biology, School of Integrative andGlobal Majors, University of Tsukuba, Tsukuba, Japan
2
International Institute of Integrative Sleep Medicine (WPI-IIIS),
University of Tsukuba, Tsukuba, Japan
3
Japan Society for the Promotion of Science (JSPS) Research
Fellow
4
Department of Psychiatry&Neuroscience, Peter O’Donnell
Brain Institute, UT Southwestern Medical Center, Dallas, TX,
USA

1

Master’s Program in Medical Sciences, University of Tsukuba,
Japan
2
International Institute for Integrative Sleep Medicine (WPI-IIIS),
Japan
3
Forestry and Forest Products Research Institute, Japan
4
Ph.D. Program in Human Biology, University of Tsukuba, Japan
5
Graduate School of Comprehensive Human Sciences, Universityof Tsukuba, Japan
6
Department of nursing, Teikyo University, Japan
7
Faculty of Medicine, University of Tsukuba, JapanThe amplitude and power of slow wave activity (SWA)
during slow wave sleep (SWS), recorded fromelectroencephalograms (EEG) is a key indicator of sleep
homeostasis and therefore it likely has a central role insleep. SWA are reflected the synchronized activity ofcortical neurons. It reflects their high participation inalternating shifts between a hyperpolarized (DOWN)
and a depolarized (UP) membrane potential.
Synchronization of this activity can be observed overlarge areas of the cortex (Destexhe et al., 1999;
Vyazovskiy, et al., 2009). These phenomena raise thequestion of how those neurons achieve the wide spreadsynchronization after we fall asleep. It was reported that
the SWA is a traveling wave (Massimini, et al., 2004),
suggesting that SWA synchronization propagates
through cortical network interactions. However, theprecise mechanism of how the reactivity is modulated isstill unclear. Here we established local field potential(LFP) and unit recordings coupled with corticalstimulation using optogenetics to investigate corticalcommunication during natural waking and sleep.
Channelrhodopsin 2 expression and laser stimulation
were performed unilaterally to somatosensory cortex,
and the signals were recorded from primary motor
cortex of both hemispheres. We found that the lightevoked potential in both ipsilateral and contralateralside to the stimulation grow stronger during SWS. Moreunit activities were also recorded as a response to thestimulation during SWS. We also analyzed thestimulation during ON or OFF phase of the slow wave
activity separately, and the larger response wereobserved in both phase. This data suggest that in SWS
the cortical neurons are more responsive to the input
both during ON and OFF phase.

Purpose: Sleep disordered breathing (SDB) is one of therisk factors for cardiovascular diseases, therefore its
early detection and treatment are important. However,there are few studies investigating the self-rated
prevalence of SDB. The aim of this study is toinvestigate the percentage of individuals who are selfaware of and treated for SDB among Japanese workers,
as well as factors associated with SDB.
Methods: A survey was conducted on self-administrated
questionnaire. A total of 762 participants (men 338 andwomen 424; mean age±SD: 43.0±11.3 years) were
included in the analysis. The odds ratios were calculated
by logistic regression analysis, adjusted for age, BMI(continues variables), nightcap, smoking and exercise
habits.
Results: The number of participants who answered “noapnea”, “apnea ≥ once a week”, and “unknown” were142 (42.0%), 53 (15.7%), and 143 (42.3%) in men, and257 (60.6%), 6 (1.4%), and 161 (38.0%) in women,
respectively. Among potential patients (n=59), 12
participants (20.4%, 10 men and 2 women) had or are
currently being treated of apnea. After excluding“unknown,”, the proportion of self-rated apnea wassignificantly different between men and women(p<0.001). The adjusted odds ratio of BMI for self-rated
apnea is 1.44 (95%CI: 1.22-1.69) in men.
Discussion: Only 20.4% of participants who were awareof apnea were in treatment, suggesting that many areunaware of the importance of treatment for SDB. It is
confirmed that high BMI is a risk factor for apnea inmen. We will continue to analyze in detail, including
data from health check-ups.
12

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P25

Different effects of orexin receptor antagonist
and GABAA agonist on physical andcognitive functions.

Reactivation of adult-born neurons formemory consolidation during sleep

1

1

2

Sakthivel Srinivasan1,
Sakaguchi1

2

Jaehoon Seol , Yuya Fujii , Insung Park , Yoko Suzuki ,Fusae Kawana2, Katsuhiko Yajima1, Shoji Fukusumi2,
Tomohiro Okura1, Makoto Satoh2, Kumpei Tokuyama2,
Toshio Kokubo2, Masashi Yanagisawa2

ToshieNaoi1,

Masanori

1

International Institute for Integrative Sleep Medicine (WPI-IIIS),
University of Tsukuba, Tsukuba, Ibaraki

The mechanisms how episodic memory is consolidated
during sleep is unknown. We found that the
hippocampal adult-born neurons (ABNs) play crucial
roles in memory consolidation during sleep1. It issuggested that memory replay contribute to memory
consolidation during sleep2. However, it is not clear
whether memory reactivation during sleep is necessary
for memory consolidation. To clarify this point, we
examine the effect of silencing the reactivation of ABN
during sleep. For this purpose, we created tripletransgenic animal to specifically target ABNs, which areactivated during learning. This study will provide a
causal link between reactivation of memory engramwith its consolidation during sleep.
1. Kumar&Srinivasan et al, in prep.
2. Skaggs&McNaughton, Science, v271, p1870

1

Physical Education, Health and Sport Sciences, University ofTsukuba.
2
International Institute for Integrative Sleep Medicine, Universityof Tsukuba.

Purpose: To evaluated side-effect profile of suvorexant,compared with GABAA receptor agonists, in humancognitive and motor function during the middle of thenight upon forced awakening.
Methods: The present study was a randomized, doubleblind controlled, 3-way crossover study. The
participants were 30 healthy males, and sleep wasrecorded polysomnographically. 15 minutes before
bedtime, participants took a pill: suvorexant 20 mg,
brotizolam 0.25 mg, or placebo. A series of physical andcognitive functions tests were performed 3.5 h before
bedtime (pre-test). 90 minutes after taking a pill,
participants were forced-awoken. Participants repeatedthe same physical and cognitive functions tests (posttest). After woke up in the morning, participants
repeated the physical and cognitive functions tests(follow-test). Physical function task is adopted bodysway, agility and dynamic balance test, choice stepping
reaction time and Purdue pegboard test. Cognitive
function task used Stroop color-word test.
Results: Body sway showed significant interaction
effects (interaction P < 0.001) of time effect (P = 0.002)
and main effect (P < 0.001). The post-hoc test showedthat the rectangular area significantly increased in the
post-test under brotizolam but returned to control value.
Agility and dynamic balance and Stroop task showedsignificant interaction and time effect; deteriorated
score in post-test under brotizolam and suvorexantcompared to placebo.
Conclusion: Suvorexant didn’t show significant
adverse effects in static balance ability, while
brotizolam reduced static balance ability evaluated after
forced awakening. However, when doing intense
movements or difficult cognitive task at that time, there
may be adverse effects at midnight.

We thank all Sakaguchi lab and WPI-IIIS members, D.
Kumar, P. Vergara, T. Hosokawa, I. Koyanagi, Y.
Mimura, K. Tanaka, T. McHugh for technical assistance,
M. Adachi for secretarial assistance, This work waspartially supported by grants from the World PremierInternational Research Center Initiative from the Japan
Ministry of Education, Culture, Sports, Science, andTechnology (MEXT); JST CREST Grant Number
JPMJCR1655; Japan Society for the Promotion of
Science Grants-in-Aid for Scientific Research (JSPS,
KAKENHI grant numbers 16K18359, 15F15408,
26115502, and 25116530); The Takeda Science
Foundation; Shimadzu Science Foundation; Kanae
Foundation for the Promotion of Medical Science,
Research Foundation for Opto-Science and Technology;
The Ichiro Kanehara Foundation; Kato Memorial
Bioscience Foundation; Japan Foundation for Applied
Enzymology; Senshin Medical Research Foundation;
Life Science Foundation of Japan; Uehara Memorial
Foundation; Brain Science Foundation; Kowa Life
Science Foundation; and 2016 Inamori Research Grants
Program to MS, The Takeda Science Foundation (TSF);
The Tokyo Biochemical Research Foundation (TBRF)
to SS.

13

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P27

Electroencephalographic analysis of the
Sleepy (sik3slp/+, sik3slp/slp) mutant mice using
the envelope characterization space

Functional Connectivity during Moral
Judgment -MEG studyH. Hiraishi1, 2, T. Ikeda2, D. N. Saito2, C. Hasegawa2, S.
Kitagawa2, T. Takahashi3, M. Kikuchi2, Y. Ouchi1

J. DIAZ , K. E. VOGT , H. FUNATO , J.-C.
LETELIER (2), M. YANAGISAWA (1)
(1)

(1)

(1)

1

Department of Biofunctional Imaging, Hamamatsu University
School of Medicine
2
Research Center for Child Mental development, Kanazawa
University
3
Health Administration Center, Fukui University(1)

WPI-IIIS, Univ. of Tsukuba, Tsukuba, Japan(2)
Univ. of Chile, Santiago, Chile

We recently reported mutant mice with significant
sleep/wake abnormalities, obtained through a largescale
EEG/EMG screening of randomly mutagenenizedcohort (1). One of these mutants, named Sleepy, bearing
a splicing mutation on the sik3 gene (with exon 13
skipping), exhibits increased daily amounts of nonREM sleep with a constitutively elevated sleep need andEEG delta power during non-REM sleep. The molecularpathway for the regulation of sleep in which sik3 takes
part is yet to be determined. The EEG envelopecharacterization space (analysis based on the signal
envelope properties) can provide valuable information
about the underlying dynamics of the neuronalensembles contributing to EEG (2). This analysis
reveals that delta waves during non-REM sleep
markedly differ from the behavior of rhythmic waves,
showing instead clear Gaussian properties. Considering
these results, delta waves can be modeled as emerging
from the superposition of EEG transients, i.e., phasic
activity. In the present study, unique EEG features ofSleepy mutants were explored under the light of theenvelope characterization space. In this parameter space,
the abnormal properties of the mutant’s EEG patterns
are evident at a glance, either by visual inspection or
automated analyses. We found that the high-amplitude
non-REM delta waves characteristic of the Sleepy
mutant are associated with an increased phasic activity
in delta, theta and sigma bands. Remarkably, thisaugmented phasic activity is more pronounced in thetaand sigma bands. These results are compatible with the
hypothesis of a superposition of transient activity withbroadband spectral consequences, originating from acolored spectrum as previously proposed (2), but
implies different amplitudes and different temporal
properties characterizing the EEG transients in the
Sleepy mutant as compared with wildtype mice. Taken
together, these results help to further characterize thesleep abnormalities induced by the sik3 mutation.

Our study aimed to clarify changes in neuronalconnection by examining the functional connectivity(FC) during moral judgment.
Method: Eleven male university students participatedin this study. When pictures with three-frame cartoon
about morality (good, bad and neutral) were presented,
participants were asked to evaluate the pictures based on
their morality by pushing a button and their brain
activities were measured by a 160-channel Magnetoencepharography (MEG, Yokogawa). MEG-160
software was used for analyzing MEG data withsampling rate 1000Hz, and preprocessed data (classified
into three time-window sets) were analyzed with asoftware Brainstorm.
Results: With the intensity threshold set over 0.8, thenumber of FC (nFC) for bad moral judgment (BMJ) was
found more than that for good moral judgment (GMJ).
The nFC was gradually increasing in both BMJ and
GMJ with passage of time. No interhemispheric (global)
connectivity was found in both judgment conditions in
all three time-window sets. In the third time-window set,
however, nFC for BMJ was 39 intra-hemispheric (local)
connectivity (left=25) and that for GMJ was 29(left=15).
Discussion: There was no interhemisphericconnectivity in both judgments in the three timewindow sets. Since lowering the threshold to 0.6
showed presence of some interhemispheric connectivity
in the occipital and frontal pairs, the interhemisphericinformation processing might be existent but weaker
than the intra-hemispheric one in moral judgment. The
time-dependent greater increase in nFC in the left
temporo-parietal area in BMJ than in GMJ suggests
dynamic changes in neuronal connections occurring inestablishment of “Theory of Mind”.

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Science Grants-in-Aid for Scientific Research (JSPS,
KAKENHI grant numbers 16K18359, 15F15408,
26115502, and 25116530); The Takeda Science
Foundation; Shimadzu Science Foundation; Kanae
Foundation for the Promotion of Medical Science,
Research Foundation for Opto-Science and Technology;
The Ichiro Kanehara Foundation; Kato Memorial
Bioscience Foundation; Japan Foundation for Applied
Enzymology; Senshin Medical Research Foundation;
Life Science Foundation of Japan; Uehara Memorial
Foundation; Brain Science Foundation; Kowa Life
Science Foundation; and 2016 Inamori Research Grants
Program to MS

Metabolomic screening for memory
consolidation during sleep
Iyo Koyanagi1, Kazuhiro Sonomura2, Taro Tezuka3,
Takaaki Sato2, Takaaki Ohnishi4, Masanori Sakaguchi1
1International Institute for Integrative Sleep Medicine (WPI-IIIS),
University of Tsukuba, Japan
2
Research Institute, Shimadzu co ltd, Japan
3
Faculty of Library, Information and Media Science, Universityof Tsukuba
4
ICT University of Tokyo

Sleep plays crucial roles in memory consolidation,
however, its molecular mechanism is still elusive. It was
shown that hippocampal theta oscillation is necessary
for memory consolidation during REM sleep1. The
hippocampal dentate gyrus (DG) shows pronounced
theta oscillation during REM sleep2 and is sufficient tohold memory trace3. Indeed, we have demonstrated that
DG neurons are necessary for memory consolidation
during sleep4. Although genetic screening was
conducted in the DG for memory consolidation5, there
should be other molecular entities involved in memory
consolidation. For example, a purine metabolite,
adenosine, plays critical roles in sleep6. Therefore, we
examined around 300 metabolites in the DG uponlearning and sleep. We found that several purine
metabolites are upregulated upon both sleep andlearning. For those metabolites, we examined theirfunction in memory and sleep. This study will show thefunctional significance of metabolites in memory
consolidation during sleep, and shed new light onmolecular mechanisms how memory is consolidated
during sleep.
1. Boyce et al., Science v352, p812, 2016.
2. Montgomery et al., J Neuroscience vA56, p48, 2008.
3. Liu et al. Nature v484, p381, 2012.
4. Kumar, Koyanagi et al., in prep.
5. Sullivan et al., Journal of Neurochemistry v101
p1085, 2007.
6. Porkka-Heiskanen et al., Science, v276, p1265, 1997.
We thank all Sakaguchi lab and WPI-IIIS members, K.
Iio, N. Kutsumura. H. Nagase, T. Naoi, D. Kumar, P
Vergara, K. Shikama, S. Sugimori, N. Hasegawa, H.
Obo for technical assistance, N. Kobayashi and M.
Adachi for secretarial assistance, This work waspartially supported by grants from the World PremierInternational Research Center Initiative from the Japan
Ministry of Education, Culture, Sports, Science, andTechnology (MEXT); JST CREST Grant Number
JPMJCR1655; Japan Society for the Promotion of15

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Psycho, a potential mouse model ofschizophrenia
Liqin Cao1, Daniela Klewe-Nebenius1, Greg Asher1,
Makito Sato1, Chika Miyoshi1, Hiromasa Funato1,
Masashi Yanagisawa1, Qinghua Liu1, 2, 3
1International Institute of Integrative Sleep Medicine (WPI-IIIS),
University of Tsukuba, Japan
2
University of Texas Southwestern Medical Center, USA
3
National Institute of Biological Sciences, China

Schizophrenia is a severe psychiatric disordercharacterized by disturbances of thoughts, perceptions,emotions and cognition. The disease affects around 1%of the general population worldwide and is enormously
costly to society. Twin and adoption studies have shownthat schizophrenia is highly heritable, with disease risk
resulting from the interplay of diverse genetic variants
as well as environmental factors. However, themolecular and neural basis of schizophrenia remain
largely unclear. Schizophrenia is often associated withemotional response deficit to fear stimuli. We developed
a forward genetics screen to isolate ethylnitrosourea(ENU)-mutagenized mice with abnormal fear responses.
We identified a dominant mutant pedigree, named
Psycho, that exhibited "schizophrenia-like" symptoms,
including hyperactivity, risk-taking behaviors, reduced
motivation, and impaired cognition. We hypothesizethat Psychom/+ mice may serve as an excellent model tounderstand the molecular and neural basis of
schizophrenia.

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