Papers by Ken-ichiro Kubo
Biochemical and Biophysical Research Communications, 2010
Disrupted-in-Schizophrenia 1 (DISC1) is a promising genetic risk factor for major mental disorder... more Disrupted-in-Schizophrenia 1 (DISC1) is a promising genetic risk factor for major mental disorders. Many groups repeatedly reported a role for DISC1 in brain development in various strains of mice and rats by using RNA interference (RNAi) approach. Nonetheless, due to the complexity of its molecular disposition, such as many splice variants and a spontaneous deletion in a coding exon of the DISC1 gene in some mouse strains, there have been debates on the interpretation on these published data. Thus, in this study, we address this question by DISC1 knockdown via short-hairpin RNAs (shRNAs) against several distinct target sequences with more than one delivery methodologies into several mouse strains, including C57BL/6, ICR, and 129X1/SvJ. Here, we show that DISC1 knockdown by in utero electroporation of shRNA against exons 2, 6, and 10 consistently results in neuronal migration defects in the developing cerebral cortex, which are successfully rescued by coexpression of full-length DISC1. Furthermore, lentivirus-mediated shRNA also led to migration defects, which is consistent with two other methodologies already published, such as plasmidmediated and retrovirus-mediated ones. The previous study by Song's group also reported that, in the adult hippocampus, the phenotype elicited by DISC1 knockdown with shRNA targeting exon 2 was consistently seen in both C57BL/6 and 129S6 mice. Taken together, we propose that some of DISC1 isoforms that are feasible to be knocked down by shRNAs to exon 2, 6, and 10 of the DISC1 gene play a key role for neuronal migration commonly in various mouse strains and rats.
The analysis of the role of Disc1 in the hippocampal layer formation in vivo
Neuroscience Research, 2011
s / Neuroscience Research 71S (2011) e108–e415 e231 Research fund: KAKENHI(21500330), Keio Univer... more s / Neuroscience Research 71S (2011) e108–e415 e231 Research fund: KAKENHI(21500330), Keio University Special Grant-in-Aid for Innovative Collaborative Research Projects. doi:10.1016/j.neures.2011.07.1005 P3-e11 Intramolecular regulation of Dab1 protein function Satoshi Kikkawa , Yoshimi Takahashi, Tomohiro Namikawa, Toshio Terashima Div of Dev Neurobiol, Kobe Univ Grad Sch of Med, Kobe, Japan In mammalians, the Reelin signal pathway plays important roles to coordinate neuronal migration during the brain development. When Reelin binds to its receptors, ApoER2 and VLDLR, the intracellular adaptor protein Disabled-1 (Dab1) is recruited to the NPXY domain of receptors and tyrosinephosphorylated by Src family kinases (SFKs). The downstream pathways seem to vary depending on brain areas, neuronal types, and/or developmental stages. We have previously identified two zebrafish orthologs of Dab1, Dab1a and Dab1b. These two isoforms showed spatially different expression patterns. When zebrafish Dab1s are overexpressed in CHO cells, the cells transfected with Dab1a or b alone showed a normal fusiform shape while the cells transfected with Dab1a/b and the Reelin receptors showed an abnormally round shape. Even when tyrosine phosphorylation by SFKs was blocked with the selective inhibitor PP2, the cells transfected with Dab1a/b and receptors showed the same round shape. We next transfected the CHO cells with various truncated-forms of Dab1a/b. The C-terminal polypeptides which lack all four putative tyrosine phosphorylation sites and the PTB domain failed to induce morphological change even when transfected with the receptors. In contrast, the N-terminal polypeptides containing the PTB domain induced morphological change in the expressed cells without co-expression of the receptors. Interestingly, when the N-terminal polypeptides were co-expressed with their complementary C-terminal polypeptides, the transfected cells showed the normal shape. These results suggest an intramolecular regulation mechanism between the Nand C-terminal domains of the Dab1 protein. doi:10.1016/j.neures.2011.07.1006 P3-e12 Expressions of tricellulin, claudin-19 and junctional adhesion molecule-C in myelinating mouse Schwann cells Shin Kikuchi 1 , Takafumi Ninomiya 1, Takashi Kojima 2, Haruyuki Tatsumi 1 1 Dept. of Anatomy, Sapporo Medical Univ. Sch. of Med., Sapporo, Japan 2 Dept. of Pathology, Sapporo Medical Univ. Sch. of Med., Sapporo, Japan Autotypic tight junctions, tight junctions connecting between same Schwann cell membranes, in mouse myelin are observed in noncompact myelin such as paranodes, Schmidt–Lanterman incisures and mesaxons. In noncompact myelin, some tight junctional (TJ) proteins are detected and thought to contribute to the integrity of myelin function. However, the role of TJ proteins in noncompact myelin is not clear. To determine in part the role of TJ proteins in noncompact myelin, we examined expressions of TJ proteins during development in mouse sciatic nerves and cultured dorsal root ganglion cells. Analyzed TJ proteins were Claudin-19 (Cldn19), junctional adhesion molecule C (JAM-C) and tricellulin (TRIC). Myelin protein zero (MPZ) was used as a marker of compact myelin. Sciatic nerves of the C57/BL mice at postnatal days (P)1, P3, P5, P7, P14, P21 and P35, and cultured dorsal root ganglion cells in 2 and 4 weeks of culture were carried out to analyze the TJ proteins in myelination by immunocytochemical staining or Western blot. In sciatic nerves, MPZ was detected at P3. Cldn19 and JAM-C were observed at P5 in paranode and TRIC was at P21. In the incisures, the expression of TRIC was observed earlier than Cldn19 or JAM-C. The expression or concentration patterns in noncompact myelin differed among the researched TJ proteins. In cultured Schwann cells, TRIC, Cldn19, JAM-C and MPZ were detected by immunostaining or Western blot in 4 weeks culture, but no they were detected in 2 weeks culture. TRIC was expressed only in mesaxon, not paranodes and the incisures. On the other hands, Cldn19 and JAM-C immunoreactivities existed at both paranodes and incisures like in vivo. TRIC was not concentrated in paranodal loops and Schmidt–Lanterman incisures under the culture condition. These results suggest that Cldn19 and JAMC play a structural maintaining role in myelination and TRIC has not only structural role but also functional one for maintaining the myelin sheath. doi:10.1016/j.neures.2011.07.1007 P3-e13 Functional balance between a microtubuleassociated protein RP1 and ciliary kinase Mak regulates ciliary length and survival of retinal photoreceptor cells Taro Chaya 1,2,3 , Yoshihiro Omori 1,2,4, Kimiko Katoh 1, Takahisa Furukawa 1,2 1 Dept Dev Biol, Osaka Biosci Inst, Suita, Osaka, Japan 2 JST, CREST, Japan 3 Dept Mol Dev, Grad Sch Med, Kyoto Univ, Kyoto, Japan 4 JST, PRESTO, Japan Cilia are evolutionally conserved microtubule-based organelles that extend from basal bodies and form on the apical surface of cells. In…
The Journal of Neuroscience, 2014
The hippocampus plays important roles in brain functions. Despite the importance of hippocampal f... more The hippocampus plays important roles in brain functions. Despite the importance of hippocampal functions, recent analyses of neuronal migration have mainly been performed on the cerebral neocortex, and the cellular mechanisms responsible for the formation of the hippocampus are not yet completely understood. Moreover, why a prolonged time is required for hippocampal neurons to complete their migration has been unexplainable for several decades. We analyzed the migratory profile of neurons in the developing mouse hippocampal CA1 region and found that the hippocampal pyramidal neurons generated near the ventricle became postmitotic multipolar cells and accumulated in the multipolar cell accumulation zone (MAZ) in the late stage of development. The hippocampal neurons passed through the pyramidal layer by a unique mode of migration. Their leading processes were highly branched and made contact with many radial fibers. Time-lapse imaging revealed that the migrating cells changed their ...
[Neuronal distribution in the cerebral cortex and its significance in schizophrenia]
Brain and nerve = Shinkei kenkyū no shinpo, 2013
Abnormal cortical architectures have been identified as one of the microscopic neuropathological ... more Abnormal cortical architectures have been identified as one of the microscopic neuropathological findings of schizophrenia. Among such findings, the altered distribution of neurons is suggestive of deficits during brain development and is consistent with the neurodevelopmental hypothesis of schizophrenia. However, the relationship between altered neuronal distribution and emergence of psychosis is unclear. In this review, we discuss the neurodevelopmental hypothesis of schizophrenia and describe the normal architecture of the neocortex and the recent findings regarding neocortical development. Thereafter, we summarize the abnormal cortical architectures reported for the brains of patients with schizophrenia. Furthermore, on the basis of the recent findings regarding cortical development, we suggest a mechanism for the formation of abnormal cortical architectures in schizophrenia and speculate how it could relate to some of the symptoms of schizophrenia.
Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling
SummaryIn mammalian cerebral neocortex, different regions have different cytoarchitecture, neuron... more SummaryIn mammalian cerebral neocortex, different regions have different cytoarchitecture, neuronal birthdates and functions. In most regions, neuronal migratory profiles have been speculated similar to each other based on observations using thymidine analogues. Few reports investigated regional migratory differences from mitosis at the ventricular surface. Here, in mice, we applied FlashTag technology, in which dyes are injected intraventricularly, to describe migratory profiles. We revealed a mediolateral regional difference in migratory profiles of neurons that is dependent on the developmental stages, e.g., neurons labeled at E12.5-15.5 reached their destination earlier dorsomedially than dorsolaterally even where there were underlying ventricular surfaces, reflecting sojourning below the subplate. This difference was hardly recapitulated by thymidine analogues, which visualize neurogenic gradient, suggesting biological significance different from neurogenic gradient. These obse...
Human neocortical development as a basis to understand mechanisms underlying neurodevelopmental disabilities in extremely preterm infants
Journal of Obstetrics and Gynaecology Research
Psychiatry and Clinical Neurosciences
Both excitatory and inhibitory neurons transiently form clusters at the outermost region of the developing mammalian cerebral neocortex
Journal of Comparative Neurology
Proceedings of the National Academy of Sciences
Reelin is an essential glycoprotein for the establishment of the highly organized six-layered str... more Reelin is an essential glycoprotein for the establishment of the highly organized six-layered structure of neurons of the mammalian neocortex. Although the role of Reelin in the control of neuronal migration has been extensively studied at the molecular level, the mechanisms underlying Reelin-dependent neuronal layer organization are not yet fully understood. In this study, we directly showed that Reelin promotes adhesion among dissociated neocortical neurons in culture. The Reelin-mediated neuronal aggregation occurs in an N-cadherin–dependent manner, both in vivo and in vitro. Unexpectedly, however, in a rotation culture of dissociated neocortical cells that gradually reaggregated over time, we found that it was the neural progenitor cells [radial glial cells (RGCs)], rather than the neurons, that tended to form clusters in the presence of Reelin. Mathematical modeling suggested that this clustering of RGCs could be recapitulated if the Reelin-dependent promotion of neuronal adhes...
The Journal of Toxicological Sciences
The aryl hydrocarbon receptor (AhR) avidly binds dioxin, a ubiquitous environmental contaminant. ... more The aryl hydrocarbon receptor (AhR) avidly binds dioxin, a ubiquitous environmental contaminant. Disruption of downstream AhR signaling has been reported to alter neuronal development, and rodent offspring exposed to dioxin during gestation and lactation showed abnormalities in learning and memory, emotion, and social behavior. However, the mechanism behind the disrupted AhR signaling and developmental neurotoxicity induced by xenobiotic ligands remains elusive. Therefore, we studied how excessive AhR activation affects neuronal migration in the hippocampal CA1 region of the developing mouse brain. We transfected constitutively active (CA)-AhR, AhR, or control vector plasmids into neurons via in utero electroporation on gestational day 14 and analyzed neuronal positioning in the hippocampal CA1 region of offspring on postnatal day 14. CA-AhR transfection affected neuronal positioning, whereas no change was observed in AhR-transfected or control hippocampus. These results suggest that constitutively activated AhR signaling disrupts neuronal migration during hippocampal development. Further studies are needed to investigate whether such developmental disruption in the hippocampus leads to the abnormal cognition and behavior of rodent offspring upon maternal exposure to AhR xenobiotic ligands.
PLOS ONE
The basic helix-loop-helix (bHLH) transcription factors exert multiple functions in mammalian cer... more The basic helix-loop-helix (bHLH) transcription factors exert multiple functions in mammalian cerebral cortex development. The aryl hydrocarbon receptor (AhR), a member of the bHLH-Per-Arnt-Sim subfamily, is a ligand-activated transcription factor reported to regulate nervous system development in both invertebrates and vertebrates, but the functions that AhR signaling pathway may have for mammalian cerebral cortex development remains elusive. Although the endogenous ligand involved in brain developmental process has not been identified, the environmental pollutant dioxin potently binds AhR and induces abnormalities in higher brain function of laboratory animals. Thus, we studied how activation of AhR signaling influences cortical development in mice. To this end, we produced mice expressing either constitutively active-AhR (CA-AhR), which has the capacity for ligandindependent activation of downstream genes, or AhR, which requires its ligands for activation. In brief, CA-AhR-expressing plasmid and AhR-expressing plasmid were each transfected into neural stems cells in the developing cerebrum by in utero electroporation on embryonic day 14.5. On postnatal day 14, mice transfected in utero with CA-AhR, but not those transfected with AhR, exhibited drastically reduced dendritic arborization of layer II/III pyramidal neurons and impaired neuronal positioning in the developing somatosensory cortex. The effects of CA-AhR were observed for dendrite development but not for the commissural fiber projection, suggesting a preferential influence on dendrites. The present results indicate that over-activation of AhR perturbs neuronal migration and morphological development in mammalian cortex, supporting previous observations of impaired dendritic structure, cortical dysgenesis, and behavioral abnormalities following perinatal dioxin exposure.
JCI insight, Jan 18, 2017
Many extremely preterm infants (born before 28 gestational weeks [GWs]) develop cognitive impairm... more Many extremely preterm infants (born before 28 gestational weeks [GWs]) develop cognitive impairment in later life, although the underlying pathogenesis is not yet completely understood. Our examinations of the developing human neocortex confirmed that neuronal migration continues beyond 23 GWs, the gestational week at which extremely preterm infants have live births. We observed larger numbers of ectopic neurons in the white matter of the neocortex in human extremely preterm infants with brain injury and hypothesized that altered neuronal migration may be associated with cognitive impairment in later life. To confirm whether preterm brain injury affects neuronal migration, we produced brain damage in mouse embryos by occluding the maternal uterine arteries. The mice showed delayed neuronal migration, ectopic neurons in the white matter, altered neuronal alignment, and abnormal corticocortical axonal wiring. Similar to human extremely preterm infants with brain injury, the surviving...
Frontiers in cellular neuroscience, 2016
Proper neuronal migration and laminar formation during corticogenesis is essential for normal bra... more Proper neuronal migration and laminar formation during corticogenesis is essential for normal brain function. Disruption of these developmental processes is thought to be involved in the pathogenesis of some neuropsychiatric conditions. Especially, Reelin, a glycoprotein mainly secreted by the Cajal-Retzius cells and a subpopulation of GABAergic interneurons, has been shown to play a critical role, both during embryonic and postnatal periods. Indeed, animal studies have clearly revealed that Reelin is an essential molecule for proper migration of cortical neurons and finally regulates the cell positioning in the cortex during embryonic and early postnatal stages; by contrast, Reelin signaling is closely involved in synaptic function in adulthood. In humans, genetic studies have shown that the reelin gene (RELN) is associated with a number of psychiatric diseases, including Schizophrenia (SZ), bipolar disorder (BP) and autistic spectrum disorder. Indeed, Reln haploinsufficiency has b...
Molecular Neuropsychiatry, 2016
If citing, it is advised that you check and use the publisher's definitive version for pagination... more If citing, it is advised that you check and use the publisher's definitive version for pagination, volume/issue, and date of publication details. And where the final published version is provided on the Research Portal, if citing you are again advised to check the publisher's website for any subsequent corrections.
Frontiers in Endocrinology, 2016
Bisphenol A (BPA) has been known to have endocrine-disrupting activity to induce reproductive and... more Bisphenol A (BPA) has been known to have endocrine-disrupting activity to induce reproductive and behavioral abnormalities in offspring of laboratory animal species. However, morphological basis of this abnormality during brain development is largely unknown. Cerebral cortex plays a crucial role in higher brain function, and its precisely laminated structure is formed by neuronal migration. In the present study, transfecting a plasmid (pCAG-mCherry) by in utero electroporation (IUE), we visualized developing neurons and investigated the possible effects of in utero BPA exposure on neuronal migration. Pregnant mice were exposed to BPA by osmotic pump at estimated daily doses of 0, 40 (BPA-40), or 400 (BPA-400) μg/kg from embryonic day 14.5 (E14.5) to E18.5. IUE was performed at E14.5 and neuronal migration was analyzed at E18.5. Compared with the control group, neuronal migration in the cortical plate was significantly decreased in the BPA-40 group; however, there was no significant difference in the BPA-400 group. Among several neuronal migration-related genes and cortical layer-specific genes, TrkB in the BPA-400 group was found significantly upregulated. In conclusion, in utero exposure to low BPA dose was found to disrupt neuronal migration in the cerebral cortex in a dose-specific manner.
Resilience in schizophrenia: A comparative study between a remote island and an urban area in Japan
Schizophrenia research, Jan 21, 2016
The resilience levels between patients with schizophrenia residing in a rural island and a metrop... more The resilience levels between patients with schizophrenia residing in a rural island and a metropolitan area in Tokyo, Japan, was compared and the factors associated with resilience were explored. The Resilience Scale (RS) and EuroQol were assessed, together with biological markers and multiple demographic variables. No significant difference was found in the RS scores between the two groups (40 subjects each). However, longer duration of illness and higher EuroQol score were significantly associated with a greater RS score, which indicates that potentially successful adaptation and subjective perspectives appear more pertinent than the degree of urbanicity in determining resilience levels.
Climbing mode of migration: A novel migration mode of pyramidal neurons in the developing hippocampus
International journal of developmental neuroscience : the official journal of the International Society for Developmental Neuroscience, 2015
Developmental origin of abnormal dendritic growth in the mouse brain induced by in utero disruption of aryl hydrocarbon receptor signaling
Neurotoxicology and Teratology, 2015
Increased prevalence of mental disorders cannot be solely attributed to genetic factors and is co... more Increased prevalence of mental disorders cannot be solely attributed to genetic factors and is considered at least partly attributable to chemical exposure. Among various environmental chemicals, in utero and lactational dioxin exposure has been extensively studied and is known to induce higher brain function abnormalities in both humans and laboratory animals. However, how the perinatal dioxin exposure affects neuromorphological alterations has remained largely unknown. Therefore, in this study, we initially studied whether and how the over-expression of aryl hydrocarbon receptor (AhR), a dioxin receptor, would affect the dendritic growth in the hippocampus of the developing brain. Transfecting a constitutively active AhR plasmid into the hippocampus via in utero electroporation on gestational day (GD) 14 induced abnormal dendritic branch growth. Further, we observed that 14-day-old mice born to dams administered with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; doses: 0, 0.6, or 3.0μg/kg) on GD 12.5 exhibited disrupted dendritic branch growth in both the hippocampus and amygdala. Finally, we observed that 16-month-old mice born to dams exposed to perinatal TCDD as described above exhibited significantly reduced spine densities. These results indicated that abnormal micromorphology observed in the developing brain may persist until adulthood and may induce abnormal higher brain function later in life.
Proceedings of the National Academy of Sciences, 2015
The preoptic area (POa) of the rostral diencephalon supplies the neocortex and the amygdala with ... more The preoptic area (POa) of the rostral diencephalon supplies the neocortex and the amygdala with GABAergic neurons in the developing mouse brain. However, the molecular mechanisms that determine the pathway and destinations of POa-derived neurons have not yet been identified. Here we show that Chicken ovalbumin upstream promoter transcription factor II (COUP-TFII)–induced expression of Neuropilin-2 (Nrp2) and its down-regulation control the destination of POa-derived GABAergic neurons. Initially, a majority of the POa-derived migrating neurons express COUP-TFII and form a caudal migratory stream toward the caudal subpallium. When a subpopulation of cells steers toward the neocortex, they exhibit decreased expression of COUP-TFII and Nrp2. The present findings show that suppression of COUP-TFII/Nrp2 changed the destination of the cells into the neocortex, whereas overexpression of COUP-TFII/Nrp2 caused cells to end up in the medial part of the amygdala. Taken together, these results ...
Frontiers in neuroscience, 2015
A fine structure of the hippocampus is required for proper functions, and disruption of this form... more A fine structure of the hippocampus is required for proper functions, and disruption of this formation by neuronal migration defects during development may play a role in some psychiatric illnesses. During hippocampal development in rodents, pyramidal neurons in the Ammon's horn are mostly generated in the ventricular zone (VZ), spent as multipolar cells just above the VZ, and then migrate radially toward the pial surface, ultimately settling into the hippocampal plate. Although this process is similar to that of neocortical projection neurons, these are not identical. In addition to numerous histological studies, the development of novel techniques gives a clear picture of the cellular dynamics of hippocampal neurons, as well as neocortical neurons. In this article, we provide an overview of the cellular mechanisms of rodent hippocampal neuronal migration including those of dentate granule cells, especially focusing on the differences of migration modes between hippocampal neur...
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Papers by Ken-ichiro Kubo