Yasushi Kajii

Schema-Dependent Gene Activation and Memory Encoding in Neocortex

New hippocampal-dependent learning is in parallel consolidated with existing memories in the neocortex. When new learning occurs against the background of established prior knowledge, relevant new information can be assimilated into a schema and thereby expand the knowledge base. An animal model of this important component of memory consolidation reveals that systems memory consolidation can be very fast. In experiments with rats, we found that the hippocampal-dependent learning of new paired associates is associated with a striking up-regulation of immediate early genes in the prelimbic region of the medial prefrontal cortex, and that pharmacological interventions targeted at that area can prevent both new learning and the recall of remotely and even recently consolidated information. These findings challenge the concept of distinct fast (hippocampal) and slow (cortical) learning systems, and shed new light on the neural mechanisms of memory assimilation into schemas.

On the relationship between the two branches of the kynurenine pathway in the rat brain in vivo.

In the mammalian brain, kynurenine aminotransferase II (KAT II) and kynurenine 3‐monooxygenase (KMO), key enzymes of the kynurenine pathway (KP) of tryptophan degradation, form the neuroactive metabolites kynurenic acid (KYNA) and 3‐hydroxykynurenine (3‐HK), respectively. Although physically segregated, both enzymes use the pivotal KP metabolite l‐kynurenine as a substrate. We studied the functional consequences of this cellular compartmentalization in vivo using two specific tools, the KAT II inhibitor BFF 122 and the KMO inhibitor UPF 648. The acute effects of selective KAT II or KMO inhibition were studied using a radiotracing method in which the de novo synthesis of KYNA, and of 3‐HK and its downstream metabolite quinolinic acid (QUIN), is monitored following an intrastriatal injection of 3H‐kynurenine. In naïve rats, intrastriatal BFF 122 decreased newly formed KYNA by 66%, without influencing 3‐HK or QUIN production. Conversely, UPF 648 reduced 3‐HK synthesis (by 64%) without affecting KYNA formation. Similar, selective effects of KAT II and KMO inhibition were observed when the inhibitors were applied acutely together with the excitotoxin QUIN, which impairs local KP metabolism. Somewhat different effects of KMO (but not KAT II) inhibition were obtained in rats that had received an intrastriatal QUIN injection 7 days earlier. In these neuron‐depleted striata, UPF 648 not only decreased both 3‐HK and QUIN production (by 77% and 66%, respectively) but also moderately raised KYNA synthesis (by 27%). These results indicate a remarkable functional segregation of the two pathway branches in the brain, boding well for the development of selective KAT II or KMO inhibitors for cognitive enhancement and neuroprotection, respectively.

Attenuation by the 5-HT1A receptor agonist osemozotan of the behavioral effects of single and repeated methamphetamine in mice

This study examined the effects of the selective 5-HT1A receptor agonist osemozotan on repeated methamphetamine (METH)-induced behavioral sensitization and single METH-induced locomotor stimulant effect in mice, and then the neurochemical mechanisms using in vivo microdialysis. Repeated administration of METH for 7 days enhanced METH challenge-induced locomotor activity, and this sensitization was observed even after its withdrawal for 7-14 days. Administration of osemozotan to METH-sensitized mice inhibited the maintenance of behavioral sensitization. This effect was blocked by a low dose of WAY100635, a selective 5-HT1A receptor antagonist. A METH challenge increased the extracellular levels of dopamine (DA), 5-HT, and noradrenaline in the prefrontal cortex, but only the increase in 5-HT release was enhanced by repeated METH administration. This augmented response of 5-HT release was attenuated by osemozotan in a WAY100635-sensitive way. A single administration of osemozotan to drug naïve mice inhibited METH-induced locomotor stimulant effect and reduced METH-induced increase in prefrontal 5-HT, but not DA, release. These results suggest that prefrontal 5-HT release is involved at least partly in the effects of osemozotan on single and repeated METH-induced behavioral effects in mice, and imply that the 5-HT1A receptors may have a potential therapeutic value in the remission of schizophrenia.

A developmentally regulated and psychostimulant-inducible novel rat gene mrt1 encoding PDZ-PX proteins isolated in the neocortex

Single or repeated exposure to psychostimulants such as amphetamines and cocaine after postnatal week 3 leads to an enduring enhancement in the psychotomimetic responses elicited by a subsequent challenge of a stimulant in rodents. This behavioral sensitization phenomenon has been considered to be the neural consequences of stimulant-induced alterations in gene expression in the brain after a critical period of postnatal development. Using a differential cloning technique, RNA arbitrarily primed PCR, we have now identified from the rat neocortex a novel and developmentally regulated methamphetamine (MAP)-inducible gene mrt1 (MAP responsive transcript 1). mrt1 encodes two major types of PDZ- and PX-domains containing proteins of approximately 62 kDa in size with different carboxy termini, Mrt1a and Mrt1b. The mrt1 mRNAs for Mrt1a, mrt1a, and for Mrt1b, mrt1b, are predominantly expressed in various brain regions and the testes, respectively. Acute MAP injection upregulated mrt1b expression in the neocortex after postnatal week 3 in a D1 receptor antagonist-sensitive manner without affecting mrt1a expression. This upregulation was mimicked by another stimulant, cocaine, whereas pentobarbital and D1 antagonist failed to change the mrt1b transcript levels. Moreover, repeated daily treatment of MAP, but not MAP plus D1 antagonist, for 5 days caused an augmentation of the basal expression of mrt1b 2 and 3 weeks after the drug discontinuation. These late-developing, cocaine-crossreactive, D1 antagonist-sensitive and long-term regulations of mrt1b by MAP are similar to the pharmacological profiles of stimulant-induced behavioral sensitization, and therefore may be associated with the initiation and/or maintenance of the long-term neuronal adaptation.

Transgenic up-regulation of alpha-CaMKII in forebrain leads to increased anxiety-like behaviors and aggression.

BackgroundPrevious studies have demonstrated essential roles for alpha-calcium/calmodulin-dependent protein kinase II (alpha-CaMKII) in learning, memory and long-term potentiation (LTP). However, previous studies have also shown that alpha-CaMKII (+/-) heterozygous knockout mice display a dramatic decrease in anxiety-like and fearful behaviors, and an increase in defensive aggression. These findings indicated that alpha-CaMKII is important not only for learning and memory but also for emotional behaviors. In this study, to understand the roles of alpha-CaMKII in emotional behavior, we generated transgenic mice overexpressing alpha-CaMKII in the forebrain and analyzed their behavioral phenotypes.ResultsWe generated transgenic mice overexpressing alpha-CaMKII in the forebrain under the control of the alpha-CaMKII promoter. In contrast to alpha-CaMKII (+/-) heterozygous knockout mice, alpha-CaMKII overexpressing mice display an increase in anxiety-like behaviors in open field, elevated zero maze, light-dark transition and social interaction tests, and a decrease in locomotor activity in their home cages and novel environments; these phenotypes were the opposite to those observed in alpha-CaMKII (+/-) heterozygous knockout mice. In addition, similarly with alpha-CaMKII (+/-) heterozygous knockout mice, alpha-CaMKII overexpressing mice display an increase in aggression. However, in contrast to the increase in defensive aggression observed in alpha-CaMKII (+/-) heterozygous knockout mice, alpha-CaMKII overexpressing mice display an increase in offensive aggression.ConclusionUp-regulation of alpha-CaMKII expression in the forebrain leads to an increase in anxiety-like behaviors and offensive aggression. From the comparisons with previous findings, we suggest that the expression levels of alpha-CaMKII are associated with the state of emotion; the expression level of alpha-CaMKII positively correlates with the anxiety state and strongly affects aggressive behavior.

Targeting Kynurenine Aminotransferase II in Psychiatric Diseases: Promising Effects of an Orally Active Enzyme Inhibitor

Increased brain levels of the tryptophan metabolite kynurenic acid (KYNA) have been linked to cognitive dysfunctions in schizophrenia and other psychiatric diseases. In the rat, local inhibition of kynurenine aminotransferase II (KAT II), the enzyme responsible for the neosynthesis of readily mobilizable KYNA in the brain, leads to a prompt reduction in extracellular KYNA levels, and secondarily induces an increase in extracellular glutamate, dopamine, and acetylcholine levels in several brain areas. Using microdialysis in unanesthetized, adult rats, we now show that the novel, systemically active KAT II inhibitor BFF-816, applied orally at 30 mg/kg in all experiments, mimics the effects of local enzyme inhibition. No tolerance was seen when animals were treated daily for 5 consecutive days. Behaviorally, daily injections of BFF-816 significantly decreased escape latency in the Morris water maze, indicating improved performance in spatial and contextual memory. Thus, systemically applied BFF-816 constitutes an excellent tool for studying the neurobiology of KYNA and, in particular, for investigating the mechanisms linking KAT II inhibition to changes in glutamatergic, dopaminergic, and cholinergic function in brain physiology and pathology.

Psychotomimetic-induction of tissue plasminogen activator mRNA in corticostriatal neurons in rat brain

We have studied in the rat the effects of acute subcutaneous injection of psychotomimetics including methamphetamine (MAP), cocaine and phencyclidine (PCP) on the expression of a brain plasticity‐related molecule, tissue plasminogen activator (tPA) mRNA, using non‐radioactive in situ hybridization histochemistry. In addition to the constitutive expression of tPA mRNA in cerebellar Purkinje cells, ventricular ependymal cells and meningeal blood vessel‐associated cells, MAP (1–4 mg/kg), cocaine (30 mg/kg) and PCP (1.25–5 mg/kg) caused a transient and dose‐dependent induction of the transcript with its peak at 3 h postinjection in a group of neurons of the medial and insular prefrontal cortices, and the piriform cortex. Another indirect dopamine agonist nomifensine (20–40 mg/kg) mimicked the tPA mRNA induction in the prefrontal cortical areas. Moreover, MAP induction of tPA mRNA was markedly inhibited by pretreatment with a D1 (R(+)‐SCH23390: R(+)‐7‐chloro‐8‐hydroxy‐3‐methyl‐1‐phenyl‐2,3,4,5‐tetra‐hydro‐1H‐3‐benzazepine hydrochloride) or a D2 (haloperidol) dopamine receptor‐preferring antagonist. Intramedial striatum, but not intrathalamic, application of a fluorescent tracer, fluorogold, retrogradely labelled the cortical cells expressing tPA mRNA. The present results suggest that acute injections of the above psychotomimetic drugs may induce tPA mRNA in a group of the prefrontal cortical neurons that project to the medial striatum. This tPA mRNA expression may be due to the activation of the dopamine neurotransmission. Because it is well documented that single or repeated administration of methamphetamine, cocaine and PCP produces enduring changes in responses to these drugs in humans and experimental animals (e.g. behavioural sensitization), the psychotomimetic‐induction of tPA mRNA could be implicated in an initial step in the plastic rearrangements in the neuronal circuits underlying long‐lasting changes in behavioural expression.

Cuprizone short-term exposure: astrocytic IL-6 activation and behavioral changes relevant to psychosis.

A growing body of evidence suggests the involvement of inflammatory processes in the pathophysiology of schizophrenia. Four- to 8-week exposure to cuprizone, a copper chelator, causes robust demyelination and has been used to build a model for multiple sclerosis. In contrast, we report here the effects of 1-week cuprizone exposure in mice. This short-term cuprizone exposure elicits behavioral changes that include augmented responsiveness to methamphetamine and phencyclidine, as well as impaired working memory. The cellular effects of 1-week cuprizone exposure differ substantially from the longer-term exposure; perturbation of astrocytes and microglia is induced without any sign of demyelination. Furthermore, the proinflammatory cytokine interleukin-6 was significantly up-regulated in glial fibrillary acidic protein (GFAP)-positive cells. We propose that this cuprizone short-term exposure may offer a model to study some aspects of biology relevant to schizophrenia and related conditions.

Age Dependency of Inhibition of α7 Nicotinic Receptors and Tonically Active N -Methyl-d-aspartate Receptors by Endogenously Produced Kynurenic Acid in the Brain

In the mouse hippocampus normal levels of kynurenic acid (KYNA), a neuroactive metabolite synthesized in astrocytes primarily by kynurenine aminotransferase II (KAT II)-catalyzed transamination of l-kynurenine, maintain a degree of tonic inhibition of α7 nicotinic acetylcholine receptors (nAChRs). The present in vitro study was designed to test the hypothesis that α7 nAChR activity decreases when endogenous production of KYNA increases. Incubation (2–7 h) of rat hippocampal slices with kynurenine (200 μM) resulted in continuous de novo synthesis of KYNA. Kynurenine conversion to KYNA was significantly decreased by the KAT II inhibitor (S)-(−)-9-(4-aminopiperazine-1-yl)-8-fluoro-3-methyl-6-oxo-2,3,5,6-tetrahydro-4H-1-oxa-3a-azaphenalene-5carboxylic acid (BFF122) (100 μM) and was more effective in slices from postweaned than preweaned rats. Incubation of slices from postweaned rats with kynurenine inhibited α7 nAChRs and extrasynaptic N-methyl-d-aspartate receptors (NMDARs) on CA1 stratum radiatum interneurons. These effects were attenuated by BFF122 and mimicked by exogenously applied KYNA (200 μM). Exposure of human cerebral cortical slices to kynurenine also inhibited α7 nAChRs. The α7 nAChR sensitivity to KYNA is age-dependent, because neither endogenously produced nor exogenously applied KYNA inhibited α7 nAChRs in slices from preweaned rats. In these slices, kynurenine-derived KYNA also failed to inhibit extrasynaptic NMDARs, which could, however, be inhibited by exogenously applied KYNA. In slices from preweaned and postweaned rats, glutamatergic synaptic currents were not affected by endogenously produced KYNA, but were inhibited by exogenously applied KYNA. These results suggest that in the mature brain α7 nAChRs and extrasynaptic NMDARs are in close apposition to KYNA release sites and, thereby, readily accessible to inhibition by endogenously produced KYNA.

Increasing effects of S-methyl-L-cysteine on the extracellular D-serine concentrations in the rat medial frontal cortex.

In an in vivo dialysis experiment, the intra-medial frontal cortex infusion of a system A and Asc-1 transporter inhibitor, S-methyl-l-cysteine, caused a concentration-dependent increase in the dialysate contents of an endogenous coagonist for the N-methyl-d-aspartate (NMDA) type glutamate receptor, d-serine, in the cortical portion. These results suggest that these neutral amino acid transporters could control the extracellular d-serine signaling in the brain and be a target for the development of a novel threapy for neuropsychiatric disorders with an NMDA receptor dysfunction.