Publications by authors named "Manabu Abe"

229 Publications

Studies on the application of the Paternò-Büchi reaction to the synthesis of novel fluorinated scaffolds.

Chemistry 2021 Sep 15. Epub 2021 Sep 15.

Hiroshima University, Department of Chemistry, JAPAN.

In the context of new scaffolds obtained by photochemical reactions, Paternò-Büchi reactions between heteroaromatic, trifluoromethylphenyl ketone and electron rich alkenes to give oxetanes are described. A comprehensive study has then been carried out on the reaction of aromatic ketones with fluorinated alkenes. Depending on the substitution pattern at the oxetane ring, a metathesis reaction is described as a minor side process to give mono fluorinated alkenes. Overall, this last reaction corresponds to a photo-Wittig reactions and yield amid isosteres. In order to explain the uncommon regioselectivity of the Paternò-Büchi reaction with these alkenes, electrostatic-potential derived charges (ESP) have been determined. In a second computational study, the relative stabilities of the typical 1,4-diradical intermediates of the PaternòBüchi reaction have been determined. The results well explain the regioselectivity. Further transformations of the oxetanes or previous functionalization of the fluoroalkenes open perspectives for oxetanes as core structures for biologically active compounds.
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http://dx.doi.org/10.1002/chem.202102621DOI Listing
September 2021

SOMO-HOMO Conversion in Triplet Cyclopentane-1,3-diyl Diradicals.

ACS Omega 2021 Sep 24;6(35):22773-22779. Epub 2021 Aug 24.

Department of Chemistry, Graduate School of Science, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima, Hiroshima 739-8526, Japan.

According to the Aufbau principle, singly occupied molecular orbitals (SOMOs) are energetically higher lying than a highest doubly occupied molecular orbital (HOMO) in the electronically ground state of radicals. However, in the last decade, SOMO-HOMO-converted species have been reported in a limited group of radicals, such as distonic anion radicals and nitroxides. In this study, SOMO-HOMO conversion was observed in triplet 2,2-difluorocyclopentane-1,3-diyl diradicals , , and 2-fluorocyclopentante-1,3-diyl diradical , which contain the anthracyl unit at the remote position. The high HOMO energy in the anthracyl moiety and the low-lying SOMO-1 due to the fluoro-substituent effect are the key to the SOMO-HOMO conversion phenomenon. Furthermore, the cation radical generated through the one-electron oxidation of was found to be a SOMO-HOMO-converted monoradical.
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http://dx.doi.org/10.1021/acsomega.1c03125DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8427779PMC
September 2021

Direct Detection of Singlet Cyclopentane-1,3-diyl Diradicals By Infrared and Ultraviolet-Visible Spectroscopy at Cryogenic Temperature and Their Photoreactivity.

J Org Chem 2021 Sep 11;86(17):12046-12053. Epub 2021 Aug 11.

Department of Chemistry, Graduate School of Science, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima, Hiroshima 739-8526, Japan.

Photolysis of a 7,7-difluoro-1,4-diphenyl-2,3-diazabicyclo[2.2.1]hept-2-ene derivative () using a 365 nm light-emitting diode in an Ar matrix at 4 K resulted in the formation of a planar singlet 2,2-difluoro-1,3-diphenylcyclopentane-1,3-diyl diradical derivative, S--pl (λ = 520 nm). A singlet cyclopentane-1,3-diyl diradical system (S--pl) was directly detected by steady-state infrared (IR) spectroscopy. Due to the photolability of S--pl, initial photolysis of also yielded the ring-closed product ret- and migration products - and/or -, which were observed using IR spectra. Monitoring of prolonged photolysis using IR and ultraviolet-visible (UV-vis) spectra demonstrated the formation of the allylic cation (λ = 470 nm). On the other hand, photolysis of a 7,7-dimethoxy-1,4-diphenyl-2,3-diazabicyclo[2.2.1]hept-2-ene derivative () yielded a puckered conformer (instead of planar) of the corresponding diradical S--puc, which was detected by IR and UV-vis spectroscopy in an Ar matrix at 4 K. This spectroscopic characterization opens a new strategy to obtain more detailed information about the structure and reactivity of singlet cyclopentane-1,3-diyl diradicals.
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http://dx.doi.org/10.1021/acs.joc.1c01410DOI Listing
September 2021

Fis1 ablation in the male germline disrupts mitochondrial morphology and mitophagy, and arrests spermatid maturation.

Development 2021 Aug 12;148(16). Epub 2021 Aug 12.

Division of Biology and Biological Engineering, California Institute of Technology, Pasadena CA 91125, USA.

Male germline development involves choreographed changes to mitochondrial number, morphology and organization. Mitochondrial reorganization during spermatogenesis was recently shown to require mitochondrial fusion and fission. Mitophagy, the autophagic degradation of mitochondria, is another mechanism for controlling mitochondrial number and physiology, but its role during spermatogenesis is largely unknown. During post-meiotic spermatid development, restructuring of the mitochondrial network results in packing of mitochondria into a tight array in the sperm midpiece to fuel motility. Here, we show that disruption of mouse Fis1 in the male germline results in early spermatid arrest that is associated with increased mitochondrial content. Mutant spermatids coalesce into multinucleated giant cells that accumulate mitochondria of aberrant ultrastructure and numerous mitophagic and autophagic intermediates, suggesting a defect in mitophagy. We conclude that Fis1 regulates mitochondrial morphology and turnover to promote spermatid maturation.
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http://dx.doi.org/10.1242/dev.199686DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8380467PMC
August 2021

Photolysis of 3-Azido-3-phenyl-3H-isobenzofuran-1-one at Ambient and Cryogenic Temperatures.

Photochem Photobiol 2021 Aug 4. Epub 2021 Aug 4.

Department of Chemistry, University of Cincinnati, Cincinnati, OH, USA.

Although alkyl azides are known to typically form imines under direct irradiation, the product formation mechanism remains ambiguous as some alkyl azides also yield the corresponding triplet alkylnitrenes at cryogenic temperatures. The photoreactivity of 3-azido-3-phenyl-3H-isobenzofuran-1-one (1) was investigated in solution and in cryogenic matrices. Irradiation (λ = 254 nm) of azide 1 in acetonitrile yielded a mixture of imines 2 and 3. Monitoring of the reaction progress using UV-Vis absorption spectroscopy revealed an isosbestic point at 210 nm, indicating that the reaction proceeded cleanly. Similar results were observed for the photoreactivity of azide 1 in a frozen 2-methyltetrahydrofuran (mTHF) matrix. Irradiation of azide 1 in an argon matrix at 6 K resulted in the disappearance of its IR bands with the concurrent appearance of IR bands corresponding to imines 2 and 3. Thus, it was theorized that azide 1 forms imines 2 and 3 via a concerted mechanism from its singlet excited state or through singlet alkylnitrene 1N, which does not intersystem cross to its triplet configuration. This proposal was supported by CASPT2 calculations on a model system, which suggested that the energy gap between the singlet and triplet configurations of alkylnitrene 1N is 33 kcal/mol, thus making intersystem crossing inefficient.
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http://dx.doi.org/10.1111/php.13500DOI Listing
August 2021

Protonation-Enhanced Reactivity of Triplet State in Dearomative Photocycloaddition of Quinolines to Olefins.

Org Lett 2021 Aug 29;23(16):6257-6261. Epub 2021 Jul 29.

Department of Chemistry, Faculty of Science, Gakushuin University, 1-5-1 Mejiro, Toshima-ku, Tokyo 171-8588, Japan.

The intermolecular dearomative cycloaddition of acidified bicyclic azaarenes with olefins was recently reported. We report here the crucial role of the acid in the dearomative photocycloaddition of quinolines to olefins. Experimental and theoretical results show that the key role of the protonation of quinolines is not to promote the energy transfer but to enhance the reactivity of the triplet state of quinolines toward olefins.
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http://dx.doi.org/10.1021/acs.orglett.1c02026DOI Listing
August 2021

Optical manipulation of local cerebral blood flow in the deep brain of freely moving mice.

Cell Rep 2021 Jul;36(4):109427

Department of Neuropsychiatry, Keio University School of Medicine, Tokyo 160-8582, Japan. Electronic address:

An artificial tool for manipulating local cerebral blood flow (CBF) is necessary for understanding how CBF controls brain function. Here, we generate vascular optogenetic tools whereby smooth muscle cells and endothelial cells express optical actuators in the brain. The illumination of channelrhodopsin-2 (ChR2)-expressing mice induces a local reduction in CBF. Photoactivated adenylyl cyclase (PAC) is an optical protein that increases intracellular cyclic adenosine monophosphate (cAMP), and the illumination of PAC-expressing mice induces a local increase in CBF. We target the ventral striatum, determine the temporal kinetics of CBF change, and optimize the illumination intensity to confine the effects to the ventral striatum. We demonstrate the utility of this vascular optogenetic manipulation in freely and adaptively behaving mice and validate the task- and actuator-dependent behavioral readouts. The development of vascular optogenetic animal models will help accelerate research linking vasculature, circuits, and behavior to health and disease.
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http://dx.doi.org/10.1016/j.celrep.2021.109427DOI Listing
July 2021

Structural Investigation of Photochemical Intermediates in Solution by Cold UV Spectroscopy in the Gas Phase: Photosubstitution of Dicyanobenzenes by Allylsilanes.

J Phys Chem A 2021 Jul 9;125(28):6238-6245. Epub 2021 Jul 9.

Department of Chemistry, Graduate School of Advanced Science and Engineering, Hiroshima University, Higashi-Hiroshima 739-8526, Japan.

Electrospray ion sources with an in-line quartz cell were constructed to produce photochemical intermediates in solution. These ion sources can detect photochemical intermediates having lifetimes longer than a few seconds. Intermediates formed by photosubstitution of 1,4-dicyanobenzene (DCB) by allyltrimethylsilane (AMS) in acetonitrile using a Xe lamp were injected into the mass spectrometer. The cationic intermediate (CHN·H) was observed at / = 171, but no anionic intermediate was found, although CHN was expected based on prior studies. Theoretical studies suggested that CHN was simultaneously converted to neutral CHN and cationic CHN·H species, which can be stable intermediates in the photosubstitution reaction. The UV photodissociation (UVPD) spectrum of CHN·H under cold (∼10 K) gas-phase conditions determined the conformation of the CHN unit of the CHN·H cation. This report demonstrates that cold gas-phase UV spectroscopy is a prospectively powerful tool for investigation of the electronic and geometric structures of photochemical intermediates produced in solution.
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http://dx.doi.org/10.1021/acs.jpca.1c04807DOI Listing
July 2021

Ddx20, DEAD box helicase 20, is essential for the differentiation of oligodendrocyte and maintenance of myelin gene expression.

Glia 2021 Nov 7;69(11):2559-2574. Epub 2021 Jul 7.

Division of Neurobiology and Anatomy, Graduate School of Medical and Dental Sciences, Niigata University, Niigata, Japan.

Oligodendrocytes form myelin sheaths that surround axons, contributing to saltatory conduction and proper central nervous system (CNS) function. Oligodendrocyte progenitor cells (OPCs) are generated during the embryonic stage and differentiate into myelinating oligodendrocytes postnatally. Ddx20 is a multifunctional, DEAD-box helicase involved in multiple cellular processes, including transcription, splicing, microRNA biogenesis, and translation. Although defects in each of these processes result in abnormal oligodendrocyte differentiation and myelination, the involvement of Ddx20 in oligodendrocyte terminal differentiation remains unknown. To address this question, we used Mbp-Cre mice to generate Ddx20 conditional knockout (cKO) mice to allow for the deletion of Ddx20 from mature oligodendrocytes. Mbp-Cre;Ddx20 cKO mice demonstrated small body sizes, behavioral abnormalities, muscle weakness, and short lifespans, with mortality by the age of 2 months old. Histological analyses demonstrated significant reductions in the number of mature oligodendrocytes and drastic reductions in the expression levels of myelin-associated mRNAs, such as Mbp and Plp at postnatal day 42. The number of OPCs did not change. A thin myelin layer was observed for large-diameter axons in Ddx20 cKO mice, based on electron microscopic analysis. A bromodeoxyuridine (BrdU) labeling experiment demonstrated that terminal differentiation was perturbed from ages 2 weeks to 7 weeks in the CNS of Mbp-Cre;Ddx20 cKO mice. The activation of mitogen-activated protein (MAP) kinase, which promotes myelination, was downregulated in the Ddx20 cKO mice based on immunohistochemical detection. These results indicate that Ddx20 is an essential factor for terminal differentiation of oligodendrocytes and maintenance of myelin gene expression.
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http://dx.doi.org/10.1002/glia.24058DOI Listing
November 2021

Impact of the macrocyclic structure and dynamic solvent effect on the reactivity of a localised singlet diradicaloid with π-single bonding character.

Chem Sci 2020 Nov 10;12(2):613-625. Epub 2020 Nov 10.

Department of Chemistry, Graduate School of Science, Hiroshima University 1-3-1 Kagamiyama, Higashi-Hiroshima Hiroshima 739-8526 Japan

Localised singlet diradicals are key intermediates in bond homolysis processes. Generally, these highly reactive species undergo radical-radical coupling reaction immediately after their generation. Therefore, their short-lived character hampers experimental investigations of their nature. In this study, we implemented the new concept of "stretch effect" to access a kinetically stabilised singlet diradicaloid. To this end, a macrocyclic structure was computationally designed to enable the experimental examination of a singlet diradicaloid with π-single bonding character. The kinetically stabilised diradicaloid exhibited a low carbon-carbon coupling reaction rate of 6.4 × 10 s (155.9 μs), approximately 11 and 1000 times slower than those of the first generation of macrocyclic system (7.0 × 10 s, 14.2 μs) and the parent system lacking the macrocycle (5 × 10 s, 200 ns) at 293 K in benzene, respectively. In addition, a significant dynamic solvent effect was observed for the first time in intramolecular radical-radical coupling reactions in viscous solvents such as glycerin triacetate. This theoretical and experimental study demonstrates that the stretch effect and solvent viscosity play important roles in retarding the σ-bond formation process, thus enabling a thorough examination of the nature of the singlet diradicaloid and paving the way toward a deeper understanding of reactive intermediates.
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http://dx.doi.org/10.1039/d0sc05311bDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8179019PMC
November 2020

The Climate Response to Emissions Reductions Due to COVID-19: Initial Results From CovidMIP.

Geophys Res Lett 2021 Apr;48(8):e2020GL091883

Irish Centre for High-End Computing (ICHEC) I Trinity Technology & Enterprise Campus Dublin 2 Ireland.

Many nations responded to the corona virus disease-2019 (COVID-19) pandemic by restricting travel and other activities during 2020, resulting in temporarily reduced emissions of CO, other greenhouse gases and ozone and aerosol precursors. We present the initial results from a coordinated Intercomparison, CovidMIP, of Earth system model simulations which assess the impact on climate of these emissions reductions. 12 models performed multiple initial-condition ensembles to produce over 300 simulations spanning both initial condition and model structural uncertainty. We find model consensus on reduced aerosol amounts (particularly over southern and eastern Asia) and associated increases in surface shortwave radiation levels. However, any impact on near-surface temperature or rainfall during 2020-2024 is extremely small and is not detectable in this initial analysis. Regional analyses on a finer scale, and closer attention to extremes (especially linked to changes in atmospheric composition and air quality) are required to test the impact of COVID-19-related emission reductions on near-term climate.
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http://dx.doi.org/10.1029/2020GL091883DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8206678PMC
April 2021

SOMO-HOMO Conversion in Triplet Carbenes.

Org Lett 2021 Jul 28;23(13):4955-4959. Epub 2021 May 28.

Department of Chemistry, Graduate School of Science, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima 739-8526, Hiroshima, Japan.

In this study, the SOMO-HOMO conversion has been shown for the first time in triplet carbenes embedded in cycloparaphenylene units. The high-lying HOMO originating from the curved π-conjugated system and the low-lying SOMO-1 originating due to the small carbene angle are the key to endowing this interesting electronic configuration. Furthermore, simple planar triplet carbenes such as fluorenylidene were found to possess SOMO-HOMO energy-converted electronic configurations.
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http://dx.doi.org/10.1021/acs.orglett.1c01137DOI Listing
July 2021

1,3-Diradicals Embedded in Curved Paraphenylene Units: Singlet versus Triplet State and In-Plane Aromaticity.

J Am Chem Soc 2021 May 26;143(19):7426-7439. Epub 2021 Apr 26.

Department of Chemistry, Graduate School of Science, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima, Hiroshima 739-8526, Japan.

Curved π-conjugated molecules and open-shell structures have attracted much attention from the perspective of fundamental chemistry, as well as materials science. In this study, the chemistry of 1,3-diradicals (s) embedded in curved cycloparaphenylene (s) structures, -(+3)s ( = 0-5), was investigated to understand the effects of the curvature and system size on the spin-spin interactions and singlet versus triplet state, as well as their unique characteristics such as in-plane aromaticity. A triplet ground state was predicted for the larger 1,3-diradicals, such as the seven- and eight-paraphenylene-unit-containing diradicals - ( = 4) and - ( = 5), by quantum chemical calculations. The smaller-sized diradicals -(+3)s ( = 0-3) were found to possess singlet ground states. Thus, the ground-state spin multiplicity is controlled by the size of the paraphenylene cycle. The size effect on the ground-state spin multiplicity was confirmed by the experimental generation of - in the photochemical denitrogenation of its azo-containing precursor (-). Intriguingly, a unique type of in-plane aromaticity emerged in the smaller-sized singlet states such as S-- ( = 1), as proven by nucleus-independent chemical shift calculations (NICS) and an analysis of the anisotropy of the induced current density (ACID), which demonstrate that homoconjugation between the 1,3-diradical moiety arises because of the curved and distorted bonding system.
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http://dx.doi.org/10.1021/jacs.1c01329DOI Listing
May 2021

CAPS2 Deficiency Impairs the Release of the Social Peptide Oxytocin, as Well as Oxytocin-Associated Social Behavior.

J Neurosci 2021 May 12;41(20):4524-4535. Epub 2021 Apr 12.

Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Chiba 278-8510, Japan

Ca-dependent activator protein for secretion 2 (CAPS2) regulates dense-core vesicle (DCV) exocytosis to facilitate peptidergic and catecholaminergic transmitter release. CAPS2 deficiency in mice has mild neuronal effects but markedly impairs social behavior. Rare alterations also occur in autism spectrum disorder, although whether CAPS2-mediated release influences social behavior remains unclear. Here, we demonstrate that CAPS2 is associated with DCV exocytosis-mediated release of the social interaction modulatory peptide oxytocin (OXT). CAPS2 is expressed in hypothalamic OXT neurons and localizes to OXT nerve projection and OXT release sites, such as the pituitary. KO mice exhibited reduced plasma albeit increased hypothalamic and pituitary OXT levels, indicating insufficient release. OXT neuron-specific conditional KO supported CAPS2 function in pituitary OXT release, also affording impaired social interaction and recognition behavior that could be ameliorated by exogenous OXT administered intranasally. Thus, CAPS2 appears critical for OXT release, thereby being associated with social behavior. The role of the neuropeptide oxytocin in enhancing social interaction and social bonding behavior has attracted considerable public and neuroscientific attention. A central issue in oxytocin biology concerns how oxytocin release is regulated. Our study provides an important insight into the understanding of oxytocin-dependent social behavior from the perspective of the CAPS2-regulated release mechanism.
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http://dx.doi.org/10.1523/JNEUROSCI.3240-20.2021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8152601PMC
May 2021

Activity-induced secretion of semaphorin 3A mediates learning.

Eur J Neurosci 2021 05 5;53(10):3279-3293. Epub 2021 Apr 5.

Department of Physiology, Yokohama City University Graduate School of Medicine, Yokohama, Japan.

The semaphorin family is a well-characterized family of secreted or membrane-bound proteins that are involved in activity-independent neurodevelopmental processes, such as axon guidance, cell migration, and immune functions. Although semaphorins have recently been demonstrated to regulate activity-dependent synaptic scaling, their roles in Hebbian synaptic plasticity as well as learning and memory remain poorly understood. Here, using a rodent model, we found that an inhibitory avoidance task, a hippocampus-dependent contextual learning paradigm, increased secretion of semaphorin 3A in the hippocampus. Furthermore, the secreted semaphorin 3A in the hippocampus mediated contextual memory formation likely by driving AMPA receptors into hippocampal synapses via the neuropilin1-plexin A4-semaphorin receptor complex. This signaling process involves alteration of the phosphorylation status of collapsin response mediator protein 2, which has been characterized as a downstream molecule in semaphorin signaling. These findings implicate semaphorin family as a regulator of Hebbian synaptic plasticity and learning.
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http://dx.doi.org/10.1111/ejn.15210DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8252788PMC
May 2021

Light-triggered elimination of CO and absorption of O (artificial breathing reaction) in photolysis of 2-(4-nitrophenyl)-1H-indole derivatives.

Photochem Photobiol Sci 2021 Mar 26;20(3):421-434. Epub 2021 Feb 26.

Department of Chemistry, Graduate School of Science, Hiroshima University, Higashihiroshima, Hiroshima, 739-8526, Japan.

A new chromophore, 2-(4-nitrophenyl)-1H-indole (NPI), was synthesized as a potential photolabile protecting group. Caged benzoic acids featuring the NPI chromophore were synthesized as model compounds. Benzoic acid was released in moderate yields (~ 40-60%) upon photolysis of the caged benzoic acids without any additional chemical reagents. Interestingly, an aldehyde, 1-(5-(1-formyl-1H-indol-2-yl)-2-nitrophenyl)ethyl benzoate, was isolated in ≈ 20% together with benzoic acid (≈ 40%) in photolysis of a caged benzoic acid, 2-(2-(3-(1-(benzoyloxy)ethyl)-4-nitrophenyl)-1H-indol-1-yl)acetic acid. The functional group, CHCOOH, at the indole nitrogen was transformed into the aldehyde group, CHO, under photolysis conditions in air. The similar photochemical transformation was observed in the photolysis of 2-(2-(4-nitrophenyl)-1H-indol-1-yl)acetic acid, in which the benzoate group is not attached at the nitrophenyl ring. Products analysis, transient absorption spectroscopy, and computational study suggested that intramolecular electron transfer is key for the elimination of CO and absorption of O for the formation of the aldehyde. The artificial breathing-type reaction can apply to transition metal-free oxidation of amino acids under mild conditions.
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http://dx.doi.org/10.1007/s43630-021-00031-zDOI Listing
March 2021

Ezh1 regulates expression of Cpg15/Neuritin in mouse cortical neurons.

Drug Discov Ther 2021 May 8;15(2):55-65. Epub 2021 Mar 8.

Neuroscience 2, Laboratory for Drug Discovery and Disease Research, Shionogi & Co. Ltd., Toyonaka, Osaka, Japan.

Immature neurons undergo morphological and physiological maturation in order to establish neuronal networks. During neuronal maturation, a large number of genes change their transcriptional levels, and these changes may be mediated by chromatin modifiers. In this study, we found that the level of Ezh1, a component of Polycomb repressive complex 2 (PRC2), increases during neuronal maturation in mouse neocortical culture. In addition, conditional knockout of Ezh1 in post-mitotic excitatory neurons leads to downregulation of a set of genes related to neuronal maturation. Moreover, the locus encoding Cpg15/Neuritin (Nrn1), which is regulated by neuronal activity and implicated in stabilization and maturation of excitatory synapses, is a direct target of Ezh1 in cortical neurons. Together, these results suggest that elevated expression of Ezh1 contributes to maturation of cortical neurons.
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http://dx.doi.org/10.5582/ddt.2021.01017DOI Listing
May 2021

Discovery of 4-alkoxy-2-aryl-6,7-dimethoxyquinolines as a new class of topoisomerase I inhibitors endowed with potent in vitro anticancer activity.

Eur J Med Chem 2021 Apr 9;215:113261. Epub 2021 Feb 9.

Department of Chemistry, Graduate School of Science, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima, Hiroshima, 739-8526, Japan. Electronic address:

In our attempt to develop potential anticancer agents targeting Topoisomerase I (TOP1), two novel series of 4-alkoxy-2-arylquinolines 14a-p and 19a-c were designed and synthesized based on structure activity relationships of the reported TOP1 inhibitors and structural features required for stabilization of TOP1-DNA cleavage complexes (TOP1ccs). The in vitro anticancer activity of these two series of compounds was evaluated at one dose level using NCI-60 cancer cell lines panel. Compounds 14e-h and 14m-p, with p-substituted phenyl at C2 and propyl linker at C4, were the most potent and were selected for assay at five doses level in which they exhibited potent anticancer activity at sub-micromolar level against diverse cancer cell lines. Compound 14m was the most potent with full panel GI MG-MID 1.26 μM and the most sensitive cancers were colon cancer, leukemia and melanoma with GI MG-MID 0.875, 0.904 and 0.926 μM, respectively. Melanoma (LOX IMVI) was the most sensitive cell line to all tested compounds displaying GI from 0.116 to 0.227 μM, TGI from 0.275 to 0.592 μM and LC at sub-micromolar concentration against almost of the tested compounds. Compounds 14e-h and 14m-p were assayed using TOP1-mediated DNA cleavage assay to evaluate their ability to stabilize TOP1ccs resulting in cancer cell death. The morpholino analogs 14h and 14p exhibited moderate TOP1 inhibitory activity compared to 1 μM camptothecin suggesting their use as lead compounds that can be optimized for the development of more potent anticancer agents with potential TOP1 inhibitory activity. Finally, Swiss ADME online web tool predicted that compounds 14h and 14p possessed good oral bioavailability and druglikeness characteristics.
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http://dx.doi.org/10.1016/j.ejmech.2021.113261DOI Listing
April 2021

A comparative analysis of kainate receptor GluK2 and GluK5 knockout mice in a pure genetic background.

Behav Brain Res 2021 05 22;405:113194. Epub 2021 Feb 22.

Division of Oral Biochemistry, Faculty of Dentistry & Graduate School of Medical and Dental Sciences, Niigata University, Niigata 951-8514, Japan. Electronic address:

Kainate receptors (KARs) are members of the glutamate receptor family that regulate synaptic function in the brain. Although they are known to be associated with psychiatric disorders, how they are involved in these disorders remains unclear. KARs are tetrameric channels assembled from a combination of GluK1-5 subunits. Among these, GluK2 and GluK5 subunits are the major heteromeric subunits in the brain. To determine the functional similarities and differences between GluK2 and GluK5 subunits, we generated GluK2 KO and GluK5 KO mice on a C57BL/6N background, a well-characterized inbred strain, and compared their behavioral phenotypes. We found that GluK2 KO and GluK5 KO mice exhibited the same phenotypes in many tests, such as reduced locomotor activity, impaired motor function, and enhanced depressive-like behavior. No change was observed in motor learning, anxiety-like behavior, or sociability. Additionally, we identified subunit-specific phenotypes, such as reduced motivation toward their environment in GluK2 KO mice and an enhancement in the contextual memory in GluK5 KO mice. These results revealed that GluK2 and GluK5 subunits not only function in a coordinated manner but also have a subunit-specific role in regulating behavior. To summarize, we demonstrated subunit-specific and common behavioral effects of GluK2 and GluK5 subunits for the first time. Moreover, to the best of our knowledge, this is the first evidence of the involvement of the GluK5 subunit in the expression of depressive-like behavior and contextual memory, which strongly indicates its role in psychiatric disorders.
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http://dx.doi.org/10.1016/j.bbr.2021.113194DOI Listing
May 2021

Simple generation of various α-monofluoroalkyl radicals by organic photoredox catalysis: modular synthesis of β-monofluoroketones.

Chem Commun (Camb) 2021 Mar;57(21):2609-2612

School of Materials and Chemical Technology, Tokyo Institute of Technology, R1-27, 4259 Nagatsuta-cho, Midori-ku, Yokohama 226-8503, Japan and Laboratory for Chemistry and Life Science, Institute of Innovative Research, Tokyo Institute of Technology, Japan.

A metal-free and operationally simple strategy for the generation of various α-monofluoroalkyl radicals has been developed. A combination of 1,4-bis(diarylamino)naphthalene photocatalyst and sulfoximine-based fluoroalkylating reagents is the key to success. The protocol can be applied to modular synthesis of β-monofluoroketones through radical monofluoroalkylation of alkenyl acetates.
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http://dx.doi.org/10.1039/d0cc08060hDOI Listing
March 2021

Dysregulation of sphingolipid metabolic enzymes leads to high levels of sphingosine-1-phosphate and ceramide in human hepatocellular carcinoma.

Hepatol Res 2021 May 9;51(5):614-626. Epub 2021 Mar 9.

Division of Digestive and General Surgery, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Niigata, Japan.

Aim: Sphingosine-1-phosphate (S1P) and ceramide are bioactive sphingolipids known to be important in regulating numerous processes involved in cancer progression. The aim of this study was to determine the absolute levels of sphingolipids in hepatocellular carcinoma (HCC) utilizing data obtained from surgical specimens. In addition, we explored the clinical significance of S1P in patients with HCC and the biological role of S1P in HCC cells.

Methods: Tumors and normal liver tissues were collected from 20 patients with HCC, and sphingolipids were measured by mass spectrometry. The Cancer Genome Atlas (TCGA) cohort was utilized to evaluate gene expression of enzymes related to sphingolipid metabolism. Immunohistochemistry of phospho-sphingosine kinase 1 (SphK1), an S1P-producing enzyme, was performed for 61 surgical specimens. CRISPR/Cas9-mediated SphK1 knockout cells were used to examine HCC cell biology.

Results: S1P levels were substantially higher in HCC tissue compared with normal liver tissue. Levels of other sphingolipids upstream of S1P in the metabolic cascade, such as sphingomyelin, monohexosylceramide and ceramide, were also considerably higher in HCC tissue. Enzymes involved in generating S1P and its precursor, ceramide, were found in higher levels in HCC compared with normal liver tissue. Immunohistochemical analysis found that phospho-SphK1 expression was associated with tumor size. Finally, in vitro assays indicated that S1P is involved in the aggressiveness of HCC cells.

Conclusions: Sphingolipid levels, including S1P and ceramide, were elevated in HCC compared with surrounding normal liver tissue. Our findings suggest S1P plays an important role in HCC tumor progression, and further examination is warranted.
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http://dx.doi.org/10.1111/hepr.13625DOI Listing
May 2021

Identification and Therapeutic Targeting of GPR20, Selectively Expressed in Gastrointestinal Stromal Tumors, with DS-6157a, a First-in-Class Antibody-Drug Conjugate.

Cancer Discov 2021 Jun 12;11(6):1508-1523. Epub 2021 Feb 12.

Daiichi Sankyo, Co., Ltd., Tokyo, Japan.

Currently, the only approved treatments for gastrointestinal stromal tumor (GIST) are tyrosine kinase inhibitors (TKI), which eventually lead to the development of secondary resistance mutations in KIT or PDGFRA and disease progression. Herein, we identified G protein-coupled receptor 20 (GPR20) as a novel non-tyrosine kinase target in GIST, developed new GPR20 IHC, and assessed GPR20 expression in cell lines, patient-derived xenografts, and clinical samples from two institutes (United States and Japan). We studied GPR20 expression stratified by treatment line, KIT expression, GIST molecular subtype, and primary tumor location. We produced DS-6157a, an anti-GPR20 antibody-drug conjugate with a novel tetrapeptide-based linker and DNA topoisomerase I inhibitor exatecan derivative (DXd). DS-6157a exhibited GPR20 expression-dependent antitumor activity in GIST xenograft models including a GIST model resistant to imatinib, sunitinib, and regorafenib. Preclinical pharmacokinetics and safety profile of DS-6157a support its clinical development as a potential novel GIST therapy in patients who are refractory or have resistance or intolerance to approved TKIs. SIGNIFICANCE: GPR20 is selectively expressed in GIST across all treatment lines, regardless of / genotypes. We generated DS-6157a, a DXd-based antibody-drug conjugate that exhibited antitumor activity in GIST models by a different mode of action than currently approved TKIs, showing favorable pharmacokinetics and safety profiles..
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http://dx.doi.org/10.1158/2159-8290.CD-20-1434DOI Listing
June 2021

Induction of Mutant Allele in Neurons in Late Infancy Increases Sleep Need.

J Neurosci 2021 03 8;41(12):2733-2746. Epub 2021 Feb 8.

International Institute for Integrative Sleep Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan

Sleep is regulated in a homeostatic manner. Sleep deprivation increases sleep need, which is compensated mainly by increased EEG δ power during non-rapid eye movement sleep (NREMS) and, to a lesser extent, by increased sleep amount. Although genetic factors determine the constitutive level of sleep need and sleep amount in mice and humans, the molecular entity behind sleep need remains unknown. Recently, we found that a gain-of-function () mutation in the () gene, which produces the mutant SIK3(SLP) protein, leads to an increase in NREMS EEG δ power and sleep amount. Since mice express SIK3(SLP) in various types of cells in the brain as well as multiple peripheral tissues from the embryonic stage, the cell type and developmental stage responsible for the sleep phenotype in mice remain to be elucidated. Here, we generated two mouse lines, and mice, which enable inducible Cre-mediated, conditional expression of SIK3(SLP) in neurons on tamoxifen administration. Administration of tamoxifen to mice during late infancy resulted in higher recombination efficiency than administration during adolescence. SIK3(SLP) expression after late infancy increased NREMS and NREMS δ power in male mice. The expression of SIK3(SLP) after adolescence led to a higher NREMS δ power without a significant change in NREMS amounts. Thus, neuron-specific expression of SIK3(SLP) after late infancy is sufficient to increase sleep. The propensity to accumulate sleep need during wakefulness and to dissipate it during sleep underlies the homeostatic regulation of sleep. However, little is known about the developmental stage and cell types involved in determining the homeostatic regulation of sleep. Here, we show that allele induction in mature neurons in late infancy is sufficient to increase non-rapid eye movement sleep amount and non-rapid eye movement sleep δ power. SIK3 signaling in neurons constitutes an intracellular mechanism to increase sleep.
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http://dx.doi.org/10.1523/JNEUROSCI.1004-20.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8018738PMC
March 2021

Activin a Receptor Type 2A Mutation Affects the Tumor Biology of Microsatellite Instability-High Gastric Cancer.

J Gastrointest Surg 2021 Jan 8. Epub 2021 Jan 8.

Division of Digestive and General Surgery, Niigata University Graduate School of Medical and Dental Sciences, 1-757 Asahimachi-dori, Chuo-ku, Niigata, Niigata, 951-8510, Japan.

Background: Activin A receptor type 2A (ACVR2A) is one of the most frequently mutated genes in microsatellite instability-high (MSI-H) gastric cancer. However, the clinical relevance of the ACVR2A mutation in MSI-H gastric cancer patients remains unclear. The aims of this study were to explore the effect of ACVR2A mutation on the tumor behavior and to identify the clinicopathological characteristics of gastric cancer patients with ACVR2A mutations.

Methods: An in vitro study was performed to investigate the biological role of ACVR2A via CRISPR/Cas9-mediated ACVR2A knockout MKN74 human gastric cancer cells. One hundred twenty-four patients with gastric cancer were retrospectively analyzed, and relations between MSI status, ACVR2A mutations, and clinicopathological factors were evaluated.

Results: ACVR2A knockout cells showed less aggressive tumor biology than mock-transfected cells, displaying reduced proliferation, migration, and invasion (P < 0.05). MSI mutations were found in 10% (13/124) of gastric cancer patients, and ACVR2A mutations were found in 8.1% (10/124) of patients. All ACVR2A mutations were accompanied by MSI. The 5-year overall survival rates of ACVR2A wild-type patients and ACVR2A-mutated patients were 57% and 90%, respectively (P = 0.048). Multivariate analysis revealed that older age (P = 0.015), distant metastasis (P < 0.001), and ACVR2A wild-type status (P = 0.040) were independent prognostic factors for overall survival.

Conclusions: Our study demonstrated that gastric cancer patients with ACVR2A mutation have a significantly better prognosis than those without. Dysfunction of ACVR2A in MKN74 human gastric cancer cells caused less aggressive tumor biology, indicating the importance of ACVR2A in the progression of MSI-H tumors.
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http://dx.doi.org/10.1007/s11605-020-04889-9DOI Listing
January 2021

Global knockdown of glutamate decarboxylase 67 elicits emotional abnormality in mice.

Mol Brain 2021 01 7;14(1). Epub 2021 Jan 7.

Department of Genetic and Behavioral Neuroscience, Gunma University Graduate School of Medicine, Maebashi, 371-8511, Japan.

Reduced expression of glutamate decarboxylase 67 (GAD67), encoded by the Gad1 gene, is a consistent finding in postmortem brains of patients with several psychiatric disorders, including schizophrenia, bipolar disorder and major depressive disorder. The dysfunction of GAD67 in the brain is implicated in the pathophysiology of these psychiatric disorders; however, the neurobiological consequences of GAD67 dysfunction in mature brains are not fully understood because the homozygous Gad1 knockout is lethal in newborn mice. We hypothesized that the tetracycline-controlled gene expression/suppression system could be applied to develop global GAD67 knockdown mice that would survive into adulthood. In addition, GAD67 knockdown mice would provide new insights into the neurobiological impact of GAD67 dysfunction. Here, we developed Gad1 biallelic knock-in mice using Gad1 and Gad1 knock-in mice, and compared them with Gad1 mice. The expression level of GAD67 protein in brains of Gad1 mice treated with doxycycline (Dox) was decreased by approximately 90%. The GABA content was also decreased in the brains of Dox-treated Gad1 mice. In the open-field test, Dox-treated Gad1 mice exhibited hyper-locomotor activity and decreased duration spent in the center region. In addition, acoustic startle responses were impaired in Dox-treated Gad1 mice. These results suggest that global reduction in GAD67 elicits emotional abnormalities in mice. These GAD67 knockdown mice will be useful for elucidating the neurobiological mechanisms of emotional abnormalities, such as anxiety symptoms associated with psychiatric disorders.
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http://dx.doi.org/10.1186/s13041-020-00713-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7789591PMC
January 2021

Heterogeneity of microglial proton channel in different brain regions and its relationship with aging.

J Neurochem 2021 05 20;157(3):624-641. Epub 2021 Jan 20.

Integrative Physiology, Graduate School of Medicine & Frontier Biosciences, Osaka University, Suita, Japan.

The properties of microglia largely differ depending on aging as well as on brain regions. However, there are few studies that investigated the functional importance of such heterogeneous properties of microglia at the molecular level. Voltage-gated proton channel, Hv1/VSOP, could be one of the candidates which confers functional heterogeneity among microglia since it regulates brain oxidative stress in age-dependent manner. In this study, we found that Hv1/VSOP shows brain region-dependent heterogeneity of gene expression with the highest level in the striatum. We studied the importance of Hv1/VSOP in two different brain regions, the cerebral cortex and striatum, and examined their relationship with aging (using mice of different ages). In the cortex, we observed the age-dependent impact of Hv1/VSOP on oxidative stress, microglial morphology, and gene expression profile. On the other hand, we found that the age-dependent significance of Hv1/VSOP was less obvious in the striatum than the cortex. Finally, we performed a battery of behavioral experiments on Hv1/VSOP-deficient mice both at young and aged stages to examine the effect of aging on Hv1/VSOP function. Hv1/VSOP-deficient mice specifically showed a marked difference in behavior in light/dark transition test only at aged stages, indicating that anxiety state is altered in aged Hv1/VSOP mice. This study suggests that a combination of brain region heterogeneity and animal aging underscores the functional importance of Hv1/VSOP in microglia.
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http://dx.doi.org/10.1111/jnc.15292DOI Listing
May 2021

p62/SQSTM1-droplet serves as a platform for autophagosome formation and anti-oxidative stress response.

Nat Commun 2021 01 4;12(1):16. Epub 2021 Jan 4.

Department of Physiology, Juntendo University Graduate School of Medicine, Bunkyo-ku, Tokyo, 113-8421, Japan.

Autophagy contributes to the selective degradation of liquid droplets, including the P-Granule, Ape1-complex and p62/SQSTM1-body, although the molecular mechanisms and physiological relevance of selective degradation remain unclear. In this report, we describe the properties of endogenous p62-bodies, the effect of autophagosome biogenesis on these bodies, and the in vivo significance of their turnover. p62-bodies are low-liquidity gels containing ubiquitin and core autophagy-related proteins. Multiple autophagosomes form on the p62-gels, and the interaction of autophagosome-localizing Atg8-proteins with p62 directs autophagosome formation toward the p62-gel. Keap1 also reversibly translocates to the p62-gels in a p62-binding dependent fashion to activate the transcription factor Nrf2. Mice deficient for Atg8-interaction-dependent selective autophagy show that impaired turnover of p62-gels leads to Nrf2 hyperactivation in vivo. These results indicate that p62-gels are not simple substrates for autophagy but serve as platforms for both autophagosome formation and anti-oxidative stress.
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http://dx.doi.org/10.1038/s41467-020-20185-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7782522PMC
January 2021

Generation of Thyroid Tissues From Embryonic Stem Cells Blastocyst Complementation .

Front Endocrinol (Lausanne) 2020 14;11:609697. Epub 2020 Dec 14.

Department of Medical Oncology, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Japan.

The generation of mature, functional, thyroid follicular cells from pluripotent stem cells would potentially provide a therapeutic benefit for patients with hypothyroidism, but differentiation remains difficult. We earlier reported the generation of lung organs blastocyst complementation in fibroblast growth factor 10 (), compound, heterozygous mutant ( Ex1/Ex3) mice. Fgf10 also plays an essential role in thyroid development and branching morphogenesis, but any role thereof in thyroid organogenesis remains unclear. Here, we report that the thyroids of Ex1/Ex3 mice exhibit severe hypoplasia, and we generate thyroid tissues from mouse embryonic stem cells (ESCs) in Ex1/Ex3 mice blastocyst complementation. The tissues were morphologically normal and physiologically functional. The thyroid follicular cells of Ex1/Ex3 chimeric mice were derived largely from GFP-positive mouse ESCs although the recipient cells were mixed. Thyroid generation blastocyst complementation will aid functional thyroid regeneration.
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http://dx.doi.org/10.3389/fendo.2020.609697DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7767966PMC
May 2021

A Discrete Glycinergic Neuronal Population in the Ventromedial Medulla That Induces Muscle Atonia during REM Sleep and Cataplexy in Mice.

J Neurosci 2021 02 28;41(7):1582-1596. Epub 2020 Dec 28.

Faculty of Medicine, University of Tsukuba, Tsukuba, Ibaraki 305-8575, Japan

During rapid eye movement (REM) sleep, anti-gravity muscle tone and bodily movements are mostly absent, because somatic motoneurons are inhibited by descending inhibitory pathways. Recent studies showed that glycine/GABA neurons in the ventromedial medulla (VMM; Gly neurons) play an important role in generating muscle atonia during REM sleep (REM-atonia). However, how these REM-atonia-inducing neurons interconnect with other neuronal populations has been unknown. In the present study, we first identified a specific subpopulation of Gly neurons that play an important role in induction of REM-atonia by virus vector-mediated tracing in male mice in which glycinergic neurons expressed Cre recombinase. We found these neurons receive direct synaptic input from neurons in several brain stem regions, including glutamatergic neurons in the sublaterodorsal tegmental nucleus (SLD; Glu neurons). Silencing this circuit by specifically expressing tetanus toxin light chain (TeTNLC) resulted in REM sleep without atonia. This manipulation also caused a marked decrease in time spent in cataplexy-like episodes (CLEs) when applied to narcoleptic mice. We also showed that Gly neurons play an important role in maintenance of sleep. This present study identified a population of glycinergic neurons in the VMM that are commonly involved in REM-atonia and cataplexy. We identified a population of glycinergic neurons in the ventral medulla that plays an important role in inducing muscle atonia during rapid eye movement (REM) sleep. It sends axonal projections almost exclusively to motoneurons in the spinal cord and brain stem except to those that innervate extraocular muscles, while other glycinergic neurons in the same region also send projections to other regions including monoaminergic nuclei. Furthermore, these neurons receive direct inputs from several brainstem regions including glutamatergic neurons in the sublaterodorsal tegmental nucleus (SLD). Genetic silencing of this pathway resulted in REM sleep without atonia and a decrease of cataplexy when applied to narcoleptic mice. This work identified a neural population involved in generating muscle atonia during REM sleep and cataplexy.
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http://dx.doi.org/10.1523/JNEUROSCI.0688-20.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7896014PMC
February 2021

Specific Neuroligin3-αNeurexin1 signaling regulates GABAergic synaptic function in mouse hippocampus.

Elife 2020 12 23;9. Epub 2020 Dec 23.

Brudnick Neuropsychiatric Research Institute, Department of Neurobiology, University of Massachusetts Medical School, Worcester, United States.

Synapse formation and regulation require signaling interactions between pre- and postsynaptic proteins, notably cell adhesion molecules (CAMs). It has been proposed that the functions of neuroligins (Nlgns), postsynaptic CAMs, rely on the formation of trans-synaptic complexes with neurexins (Nrxns), presynaptic CAMs. Nlgn3 is a unique Nlgn isoform that localizes at both excitatory and inhibitory synapses. However, Nlgn3 function mediated via Nrxn interactions is unknown. Here we demonstrate that Nlgn3 localizes at postsynaptic sites apposing vesicular glutamate transporter 3-expressing (VGT3+) inhibitory terminals and regulates VGT3+ inhibitory interneuron-mediated synaptic transmission in mouse organotypic slice cultures. Gene expression analysis of interneurons revealed that the αNrxn1+AS4 splice isoform is highly expressed in VGT3+ interneurons as compared with other interneurons. Most importantly, postsynaptic Nlgn3 requires presynaptic αNrxn1+AS4 expressed in VGT3+ interneurons to regulate inhibitory synaptic transmission. Our results indicate that specific Nlgn-Nrxn signaling generates distinct functional properties at synapses.
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http://dx.doi.org/10.7554/eLife.59545DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7758064PMC
December 2020
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