永井 拓

Neurogenesis in the hippocampus occurs throughout life in a wide range of species and could be associated with hippocampus-dependent learning and memory. Stress is well established to seriously perturb physiological/psychological homeostasis and affect hippocampal function. In the present study, to investigate the effect of chronic restraint stress in early life on hippocampal neurogenesis and hippocampus-dependent memory, 3-week-old mice were subjected to restraint stress 6 days a week for 4 weeks. The chronic restraint stress significantly decreased the hippocampal volume by 6.3% and impaired hippocampal neurogenesis as indicated by the reduced number of Ki67-, 5-bromo-2'-deoxyuridine- and doublecortin-positive cells in the dentate gyrus. The chronic restraint stress severely impaired hippocampus-dependent contextual fear memory without affecting hippocampus-independent fear memory. The expression level of brain-specific transcription factor neuronal PAS domain protein 4 (Npas4) mRNA in the hippocampus was down-regulated by the restraint stress or by acute corticosterone treatment. Npas4 immunoreactivity was detected in progenitors, immature and mature neurons of the dentate gyrus in control and stressed mice. Our findings suggest that the chronic restraint stress decreases hippocampal neurogenesis, leading to an impairment of hippocampus-dependent fear memory in mice. Corticosterone-induced down-regulation of Npas4 expression may play a role in stress-induced impairment of hippocampal function.

The high-mobility group A protein 1a (HMGA1a) is a well-documented DNA-binding protein acting as an architectural transcription regulator. Recently, HMGA1a protein has been identified as a hypoxia-inducible RNA-binding trans-acting factor for aberrant splicing of presenilin-2 (PS2) pre-mRNA observed in the brains of sporadic Alzheimer's disease. Interestingly, this aberrant splicing of PS2 was also observed in the brains of bipolar disorder and schizophrenia. Many downstream genes under the control of HMGA1a could be associated with schizophrenia. On the other hand, many gene transcripts are aberrantly spliced in schizophrenia. Therefore, we examined the expression at the mRNA and protein levels of this DNA- and RNA-binding factor HMGA1a in the lymphoblastoid cell lines obtained from 16 schizophrenia patients with age-matched controls. We observed markedly higher HMGA1a mRNA and the increased HMGA1a protein in the nuclear fractions of schizophrenia patients. In contrast, there were no significant differences in the expression levels of HMGA1b, which is an alternatively spliced isoform of HMGA1a. The present study is the first to report a significant upregulation of HMGA1a in schizophrenia, suggesting its potential roles in both transcription and splicing of target genes linked with schizophrenia. (c) 2010 Elsevier Ltd. All rights reserved.

Cognitive deficits are a core feature of patients with methamphetamine (METH) abuse. It has been reported that repeated METH treatment impairs long-term recognition memory in the novel object recognition test (NORT) in mice. Recent studies indicate that silibinin, a flavonoid derived from the herb milk thistle, has potent neuroprotective effects in cell cultures and several animal models of neurological diseases. However, its effect on the cognitive deficit induced by METH remains unclear. In the present study, we attempt to clarify the effect of silibinin on impairments of recognition memory caused by METH in mice. Mice were co-administered silibinin with METH for 7 days and then cognitive function was assessed by NORT after 7-day withdrawal. Tissue levels of dopamine and serotonin as well as their metabolites in the prefrontal cortex and hippocampus were measured 1 day after NORT. Silibinin dose-dependently ameliorated the impairment of recognition memory caused by METH treatment in mice. Silibinin significantly attenuated the decreases in the dopamine content of the prefrontal cortex and serotonin content of the hippocampus caused by METH treatment. We also found a correlation between the recognition values and dopamine and serotonin contents of the prefrontal cortex and hippocampus. The effect of silibinin on cognitive impairment may be associated with an amelioration of decreases in dopamine and serotonin levels in the prefrontal cortex and hippocampus, respectively. These results suggest that silibinin may be useful as a pharmacological tool to investigate the mechanisms of METH-induced cognitive impairments. (C) 2009 Elsevier B.V. All rights reserved.

Dystrobrevin binding protein-1 gene (DTNBP1), which encodes dysbindin protein, has been identified as a schizophrenia susceptibility gene. Dysbindin has been shown to contribute to the regulation of exocytosis and formation of synaptic vesicles. Although hypofrontality in schizophrenia underlies its pathophysiology, the molecular function of dysbindin in synaptic neurotransmission remains unclear. In the present study, we investigated depolarization-evoked dopamine (DA) and serotonin (5-HT) release in the prefrontal cortex (PFC) of sandy (sdy) mice, which have a deletion mutation in the gene encoding DTNBP1. In vivo microdialysis analysis revealed that extracellular DA levels in the PFC of wild-type mice were increased by 60 mM KCl stimulation, and the KCl-evoked DA release was significantly decreased in sdy mice compared with wild-type mice. Extracellular 5-HT levels in the PFC of wild-type mice were also increased by 60 mM KCl stimulation. The KCl-evoked 5-HT release did not differ between wild-type and sdy mice. There was no difference in basal levels of DA and 5-HT before the stimulation between two groups. Behavioral sensitization after repeated methamphetamine (METH) treatment was significantly reduced in sdy mice compared with wild-type mice whereas no difference was observed in METH-induced hyperlocomotion between two groups. These results suggest that dysbindin may have a role in the regulation of depolarization-evoked DA release in the PFC and in the development of behavioral sensitization induced by repeated METH treatment. (C) 2010 Elsevier Ireland Ltd. All rights reserved.

Gene-environment interaction may play a role in the etiology of schizophrenia. Transgenic mice expressing dominant-negative DISC1 (DN-DISC1 mice) show some histological and behavioral endophenotypes relevant to schizophrenia. Viral infection during neurodevelopment provides a major environmental risk for schizophrenia. Neonatal injection of polyriboinosinic-polyribocytidylic acid (polyI:C), which mimics innate immune responses elicited by viral infection, leads to schizophrenia-like behavioral alteration in mice after puberty. To study how gene-environmental interaction during neurodevelopment results in phenotypic changes in adulthood, we treated DN-DISC1 mice or wild-type littermates with injection of polyI:C during the neonatal stage, according to the published method, respectively, and the behavioral and histological phenotypes were examined in adulthood. We demonstrated that neonatal polyI:C treatment in DN-DISC1 mice resulted in the deficits of short-term, object recognition, and hippocampus-dependent fear memories after puberty, although polyI:C treatment by itself had smaller influences on wild-type mice. Furthermore, polyI:C-treated DN-DISC1 mice exhibited signs of impairment of social recognition and interaction, and augmented susceptibility to MK-801-induced hyperactivity as compared with vehicle-treated wild-type mice. Of most importance, additive effects of polyI:C and DN-DISC1 were observed by a marked decrease in parvalbumin-positive interneurons in the medial prefrontal cortex. These results suggest that combined effect of neonatal polyI:C treatment and DN-DISC1 affects some behavioral and histological phenotypes in adulthood.

Intracellular amyloid-beta peptide (A beta) has been implicated in neuronal death associated with Alzheimer's disease. Although A beta is predominantly secreted into the extracellular space, mechanisms of A beta transport at the level of the neuronal cell membrane remain to be fully elucidated. We demonstrate that receptor for advanced glycation end products (RAGE) contributes to transport of A beta from the cell surface to the intracellular space. Mouse cortical neurons exposed to extracellular human A beta subsequently showed detectable peptide intracellularly in the cytosol and mitochondria by confocal microscope and immunogold electron microscopy. Pretreatment of cultured neurons from wild-type mice with neutralizing antibody to RAGE, and neurons from RAGE knockout mice displayed decreased uptake of A beta and protection from A beta-mediated mitochondrial dysfunction. A beta activated p38 MAPK, but not SAPK/JNK, and then stimulated intracellular uptake of A beta-RAGE complex. Similar intraneuronal co-localization of A beta and RAGE was observed in the hippocampus of transgenic mice overexpressing mutant amyloid precursor protein. These findings indicate that RAGE contributes to mechanisms involved in the translocation of A beta from the extracellular to the intracellular space, thereby enhancing A beta cytotoxicity.

In Alzheimer's disease (AD), the deposition of amyloid peptides is invariably associated with oxidative stress and inflammatory responses. Silibinin (silybin), a flavonoid derived from the herb milk thistle, has potent anti-inflammatory and antioxidant activities. However, it remains unclear whether silibinin improves amyloid beta (A beta) peptide-induced neurotoxicity. In this study, we examined the effect of silibinin on the fear-conditioning memory deficits, inflammatory response, and oxidative stress induced by the intracerebroventricular injection of A beta peptide(25-35) (A beta(25-35)) in mice. Mice were treated with silibinin (2, 20, and 200 mg/kg p.o., once a day for 8 days) from the day of the A beta(25-35) injection (day 0). Memory function was evaluated in cued and contextual fear-conditioning tests (day 6). Nitrotyrosine levels in the hippocampus and amygdala were examined (day 8). The mRNA expression of inducible nitric-oxide synthase (iNOS) and tumor necrosis factor-alpha (TNF-alpha) in the hippocampus and amygdala was measured 2 h after the A beta(25-35) injection. We found that silibinin significantly attenuated memory deficits caused by A beta(25-35) in the cued and contextual fear-conditioning test. Silibinin significantly inhibited the increase in nitrotyrosine levels in the hippocampus and amygdala induced by A beta(25-35). Nitrotyrosine levels in these regions were negatively correlated with memory performance. Moreover, real-time RT-PCR revealed that silibinin inhibited the overexpression of iNOS and TNF-alpha mRNA in the hippocampus and amygdala induced by A beta(25-35). These findings suggest that silibinin (i) attenuates memory impairment through amelioration of oxidative stress and inflammatory response induced by A beta(25-35) and (ii) may be a potential candidate for an AD medication.

Background and purpose: Accumulated evidence suggests that oxidative stress is involved in amyloid beta (A beta)-induced cognitive dysfunction. Silibinin (silybin), a flavonoid derived from the herb milk thistle (Silybum marianum), has been shown to have antioxidative properties; however, it remains unclear whether silibinin improves A beta-induced neurotoxicity. In the present study, we examined the effect of silibinin on the memory impairment and accumulation of oxidative stress induced by A beta(25-35) in mice. Experimental approach: Aggregated A beta(25-35) (3 nmol) was intracerebroventricularly administered to mice. Treatment with silibinin (2, 20 and 200 mg center dot kg(-1), once a day, p.o.) was started immediately after the injection of A beta(25-35). Locomotor activity was evaluated 6 days after the A beta(25-35) treatment, and cognitive function was evaluated in a Y-maze and novel object recognition tests 6-11 days after the A beta(25-35) treatment. The levels of lipid peroxidation (malondialdehyde) and antioxidant (glutathione) in the hippocampus were measured 7 days after the A beta(25-35) injection. Key results: Silibinin prevented the memory impairment induced by A beta(25-35) in the Y-maze and novel object recognition tests. Repeated treatment with silibinin attenuated the A beta(25-35)-induced accumulation of malondialdehyde and depletion of glutathione in the hippocampus. Conclusions and implications: Silibinin prevents memory impairment and oxidative damage induced by A beta(25-35) and may be a potential therapeutic agent for Alzheimer's disease.

Social isolation (SI) rearing in rodents causes a variety of behavioral changes, including hyperlocomotion, anxiety, impulsivity, aggression, and learning and memory deficits. These behavioral abnormalities in rodents may be related to the symptoms in patients with neuro psychiatric disorders, such as attention-deficit hyperactivity disorder, obsessive-compulsive disorder, autism, schizophrenia and depression. In this study, we examined the effect of long-term SI rearing after weaning on emotional behaviors and cognitive function in mice. Furthermore, the effects of methylphenidate (MPH), clozapine (CLZ)and fluoxetine (FIX) on SI-induced behavioral changes were examined to measure the predictive validity of SI-reared mice as an animal model for these neuropsychiatric disorders. MPH improved SI-induced anxiety-like behavior in the elevated-plus maze test, but had no effect on aggressive behavior. In contrast, CLZ ameliorated aggressive behavior, but not anxiety-like behavior in SI-reared mice. Repeated FIX treatment prevented SI-induced aggressive behavior and social interaction deficits. These findings suggest that SI-induced behavioral abnormality is a psychobehavioral complex relevant to various clinical symptoms observed in neuro psychiatric disorders and that SI-reared mice are a useful animal model to study the pathophysiology/pathogenesis of these diseases. (c) 2009 Elsevier B.V. All rights reserved.

It has been reported that viral infection in the first and second trimesters of pregnancy in humans increases the risk of subsequently developing schizophrenia. To develop a mouse model of immune activation during the early postnatal period, neonatal ICR mice were repeatedly injected with polyriboinosinic-polyribocytidilic acid (polyI:C; an inducer of strong innate immune responses) for 5 days (postnatal day 2-6) which may correspond, in terms of brain development, to the early second trimester in human. Cognitive and emotional behavior as well as the extracellular level of glutamate in the hippocampus were analyzed at the age of 10-12 weeks old. PolyI:C-treated mice showed anxiety-like behavior, impairment of object recognition memory and social behavior, and sensorimotor gating deficits, as compared to the saline-treated control group. Depolarization-evoked glutamate release in the hippocampus was impaired in polyI:C-treated mice compared to saline-treated control mice. Furthermore, to investigate the effect of neonatal immune activation on the expression levels of schizophrenia-related genes, we analyzed mRNA levels in the hippocampus 2 and 24h after polyI:C treatment. No significant differences or only transient and marginal changes were observed between polyI:C-treated and saline-treated control mice in the expression levels of schizophrenia-related genes in the hippocampus.

Early-life stress during the postnatal period could precipitate long-lasting alterations in the functional properties underlying emotional expression in humans,but how the psychological stress of cross-fostering affects emotional behavior during adulthood in mice remains primarily unknown. The purpose of the present study was to examine the long-term effects of cross-fostering on the emotional behavior and cognitive functions of ICR offspring in adulthood. Cross-fostering was performed from postnatal day 7 for 3 weeks. Mice were divided into three groups: (1) biological group: pups born from ICR dams fostered by their original mothers; (2) in-foster group: pups born from ICR dams but adopted by other ICR dams and (3) cross-foster group: ICR pups adopted by C57 dams. ICR mice were subjected to behavioral experiments at the age of 8 weeks. Emotional behaviors in the cross-fostered mice were significantly altered in the open-field, elevated plus maze and forced swimming tests, as well as social interaction tests. However, the cross-fostered mice showed normal memory function in the Y-maze and novel object recognition tests. The contents of serotonin metabolisms were decreased in the prefrontal cortex and hippocampus indicated the deficit of serotoninergic neuronal function by cross-fostering. These findings suggested that the early-life stress of cross-fostering induced long-lasting emotional abnormalities, which might be possibly related to alterations of serotonin metabolisms. (C) 2008 Elsevier B.V. All rights reserved.

Cognitive deficits, including memory impairment, are regarded as a core feature of schizophrenia. Aripiprazole, an atypical antipsychotic drug, has been shown to improve disruption of prepulse inhibition and social interaction in an animal model of schizophrenia induced by phencyclidine (PCP); however, the effects of aripiprazole on recognition memory remain to be investigated. In this study, we examined the effect of aripiprazole on cognitive impairment in mice treated with PCP repeatedly. Mice were repeatedly administered PCP at a dose of 10mg/kg for 14days, and their cognitive function was assessed using a novel-object recognition task. We investigated the therapeutic effects of aripiprazole (0.01-1.0mg/kg) and haloperidol (0.3 and 1.0mg/kg) on cognitive impairment in mice treated with PCP repeatedly. Single (1.0mg/kg) and repeated (0.03 and 0.1mg/kg, for 7days) treatment with aripiprazole ameliorated PCP-induced impairment of recognition memory, although single treatment significantly decreased the total exploration time during the training session. In contrast, both single and repeated treatment with haloperidol (0.3 and 1.0mg/kg) failed to attenuate PCP-induced cognitive impairment. The ameliorating effect of aripiprazole on recognition memory in PCP-treated mice was blocked by co-treatment with a dopamine D(1) receptor antagonist, SCH23390, and a serotonin 5-HT(1A) receptor antagonist, WAY100635; however, co-treatment with a D(2) receptor antagonist raclopride had no effect on the ameliorating effect of aripiprazole. These results suggest that the ameliorative effect of aripiprazole on PCP-induced memory impairment is associated with dopamine D(1) and serotonin 5-HT(1A) receptors.

In this study, we investigated the effects of GABA(A) and GABA(B) receptor agonists on the methamphetamine-induced impairment of recognition memory in mice. Repeated treatment with methamphetamine at a dose of 1 mg/kg for 7 days induced an impairment of recognition memory. Baclofen, a GABA(B) receptor agonist, ameliorated the repeated methamphetamine-induced cognitive impairment, although gaboxadol, a GABA(A) receptor agonist, had no significant effect. GABA(B) receptors may constitute a putative new target in treating cognitive deficits in patients suffering from schizophrenia, as well as methamphetamine psychosis. (C) 2008 Elsevier B.V. All rights reserved.

We have previously demonstrated that pallidotegmental GABAergic neurons play a crucial role in prepulse inhibition (PPI) of the startle reflex in mice through the activation of GABA(B) receptors in pedunculopontine tegmental neurons. In this study, we investigated whether PPI disruption induced by methamphetamine ( METH) or MK-801 is associated with the dysfunction of pallidotegmental neurons. Furthermore, we examined the effects of baclofen, a GABA(B) receptor agonist, on METH- and MK-801-induced PPI impairment. Acute treatment with METH ( 3 mg/kg, subcutaneouly (s.c.)) and MK-801 (>0.3 mg/kg, s.c.) significantly disrupted PPI, accompanied by the suppression of c-Fos expression in lateral globus pallidus induced by PPI. Furthermore, acute treatment with METH and MK-801 stimulated c-Fos expression in the caudal pontine reticular nucleus (PnC) in mice subjected to the PPT test, although PPI alone had no effect on c-Fos expression. Repeated treatment with 1 mg/kg METH for 7 days, which did not affect PPI acutely, showed similar effects on PPI and c-Fos expression to acute treatment with METH ( 3 mg/kg). Baclofen dose-dependently ameliorated PPI impairment induced by acute treatment with METH ( 3 mg/kg) and MK-801 ( 1 mg/kg), and decreased METH- and MK-801-stimulated c-Fos expression in PnC to the basal level. These results suggest that dysfunction of pallidotegmental neurons is involved in PPI disruption caused by METH and MK-801 in mice. GABA(B) receptor may constitute a putative target in treating neuropsychiatric disorders with sensorimotor gating deficits, such as schizophrenia and METH psychosis.

Tyrosine nitration of proteins at an extensive level is widely associated with the cognitive pathology induced by amyloid beta peptide (A beta). However, the precise identity and explicit consequences of protein nitration have scarcely been addressed. In this study, we examined the detectable nitration of proteins in the hippocampus of mice with cognitive impairment (day 5) induced by the i.c.v. injection of A beta(25-35) (day 0). The intensity of the nitration of proteins was inversely associated with the level of recognition memory in mice. The detectable tyrosine nitrations were revealed in proteins with a single size of approximately 70 kDa. The specific nitrated proteins at this size were identified using the liquid chromatography/mass spectrometry/mass spectrometry analysis and immunodetection methods. Intense nitration of the neurofilament light chain (NFL) was observed. The increased nitration of NFL was associated with its serine hyperphosphorylation and weak interaction with the nuclear distribution element-like, a protein essential for the stable assembly of neurofilaments. No changes in cell numbers in the hippocampus were found (day 5) in mice that received A beta(25-35) injections. These findings suggested that extensive nitration of NFL is associated with the A beta-induced impairment of recognition memory in mice.

Experiences during brain development may influence the pathogenesis of developmental disorders. Thus, social isolation (SI) rearing after weaning is a useful animal model for studying the pathological mechanisms of such psychiatric diseases. In this study, we examined the effect of SI on neurogenesis in the hippocampal dentate gyrus (DG) relating to memory and emotion-related behaviors. When newly divided cells were labeled with 5-bromo-2'-deoxyuridine (BrdU) before SI, the number of BrdU-positive cells and the rate of differentiation into neurons were significantly decreased after 4-week SI compared with those in group-housed mice. Repeated treatment of fluoxetine prevented the SI-induced impairment of survival of newly divided cells and ameliorated spatial memory impairment and part of aggression in SI mice. Furthermore, we investigated the changes in gene expression in the DG of SI mice by using DNA microarray and real-time PCR. We finally found that SI reduced the expression of development-related genes Nurr1 and Npas4. These findings suggest that communication in juvenile is important in the survival and differentiation of newly divided cells, which may be associated with memory and aggression, and raise the possibility that the reduced expression of Nurr1 and/or Npas4 may contribute to the impairment of neurogenesis and memory and aggression induced by SI.

We have previously demonstrated that repeated, but not acute, methamphetamine (METH) treatment increases tissue plasminogen activator (tPA) activity in the brain, which is associated with the development of behavioral sensitization to METH. In this study, we investigated whether the tPA-plasmin system is involved in the development of sensitization in METH-induced dopamine release in the nucleus accumbens (NAc). There was no difference in acute METH-induced increase in extracellular dopamine levels in the NAc between wild-type and tPA-deficient (tPA-/-) mice. Repeated METH treatment resulted in a significant enhancement of METH- induced dopamine release in wild-type mice, but not tPA-/- mice. Microinjection of exogenous tPA or plasmin into the NAc of wild-type mice significantly potentiated acute METH- induced dopamine release. Degradation of laminin was evident in brain tissues incubated with tPA plus plasminogen or plasmin in vitro although tPA or plasminogen alone had no effect. Immunohistochemical analysis revealed that microinjection of plasmin into the NAc reduced laminin immunoreactivity without neuronal damage. Our findings suggest that the tPA-plasmin system participates in the development of behavioral sensitization induced by repeated METH treatment, by regulating the processes underlying the sensitization of METH-induced dopamine release in the NAc, in which degradation of laminin by plasmin may play a role.