Chronic social defeat stress induces the down-regulation of the Nedd4L-GLT-1 ubiquitination pathway in the prefrontal cortex of mice.

Stressful life events contribute to the onset of major depressive disorder (MDD). We recently demonstrated abnormalities in ubiquitination in the pathophysiology of MDD. However, the underlying molecular mechanisms remain unclear. We investigated the involvement of the ubiquitination system-mediated glutamatergic dysfunction in social impairment induced by chronic social defeat stress (CSDS). Adult C57BL/6J mice were exposed to aggressor ICR male mice for 10 consecutive days. Social impairment was induced by CSDS in the social interaction test 1 days after the last stress exposure. In terms of brain microdialysis, CSDS reduced depolarization-evoked glutamate release in the prefrontal cortex (PFC), which was reversed by a glutamate transporter 1 (GLT-1) inhibitor. Interestingly, the expression of ubiquitinated, but not total GLT-1, was decreased in the PFC of mice exposed to CSDS. The expression of neural precursor cells expressing developmentally downregulated gene 4-like (Nedd4L: E3 ligase for GLT-1), and ubiquitin-conjugating enzyme E2D2 (Ube2d2: E2 ubiquitin-conjugating enzyme for Nedd4L) was also reduced in CSDS mice. Furthermore, the downregulation of the Nedd4L-GLT-1 ubiquitination pathway decreased SIT ratio, but up-regulation increased it even in non-CSDS mice. Taken together, the decrease in GLT-1 ubiquitination may reduce the release of extracellular glutamate induced by high-potassium stimulation, which may lead to social impairment, while we could not find differences in GLT-1 ubiquitination between susceptible and resistant CSDS mice. In conclusion, GLT-1 ubiquitination could play a crucial role in the pathophysiology of MDD and is an attractive target for the development of novel antidepressants.

Deficiency of kynurenine 3-monooxygenase exacerbates impairment of prepulse inhibition induced by phencyclidine.

Phencyclidine (PCP) causes mental symptoms that closely resemble schizophrenia through the inhibition of the glutamatergic system. The kynurenine (KYN) pathway (KP) generates metabolites that modulate glutamatergic systems such as kynurenic acid (KA), quinolinic acid (QA), and xanthurenic acid (XA). Kynurenine 3-monooxygenase (KMO) metabolizes KYN to 3-hydroxykynurenine (3-HK), an upstream metabolite of QA and XA. Clinical studies have reported lower KMO mRNA and higher KA levels in the postmortem brains of patients with schizophrenia and exacerbation of symptoms in schizophrenia by PCP. However, the association between KMO deficiency and PCP remains elusive. Here, we demonstrated that a non-effective dose of PCP induced impairment of prepulse inhibition (PPI) in KMO KO mice. KA levels were increased in the prefrontal cortex (PFC) and hippocampus (HIP) of KMO KO mice, but 3-HK levels were decreased. In wild-type C57BL/6 N mice, the PPI impairment induced by PCP is exacerbated by KA, while attenuated by 3-HK, QA and XA. Taken together, KMO KO mice were vulnerable to the PPI impairment induced by PCP through an increase in KA and a decrease in 3-HK, suggesting that an increase in the ratio of KA to 3-HK (QA and XA) may play an important role in the pathophysiology of schizophrenia.

Loureirin C and Xanthoceraside Prevent Abnormal Behaviors Associated with Downregulation of Brain Derived Neurotrophic Factor and AKT/mTOR/CREB Signaling in the Prefrontal Cortex Induced by Chronic Corticosterone Exposure in Mice.

Brain derived neurotrophic factor (BDNF) is one of the most abundant neurotrophic factors, and its deficits are involved in the pathogenesis of major depressive disorders (MDD). Loureirin C (Lou C) is a compound derived from red resin extracted from the stems of Chinese dragon's blood. Xanthoceraside (Xan) is a triterpenoid saponin extracted from the husks of Xanthoceras sorbifolia Bunge. These compounds have neuroprotective effects through upregulation of BDNF. The present study aimed to evaluate whether Lou C and Xan attenuate abnormal behaviors induced by chronic corticosterone (CORT) administration. CORT was administered subcutaneously to mice for 3 weeks, and Lou C and Xan, dispensed orally once a day during the last 2 weeks of CORT administration. Chronic CORT administration induced abnormal behaviors such as prolonged starting latency in the open field test, decreased social interaction time in the social interaction test and prolonged latency to eat in the novelty suppressed feeding test. Chronic CORT administration decreased the expression levels of BDNF and the phosphorylation of protein kinase B (Akt), mammalian target of rapamycin (mTOR), and the cAMP response element binding protein (CREB) in the prefrontal cortex. Lou C and Xan dose-dependently prevented the abnormal behaviors and decreased the expression levels of BDNF and in phosphorylation of AKT, mTOR, and CREB in the prefrontal cortex of CORT mice. These results suggest that Lou C and Xan could be attractive candidates for pharmacotherapy of MDD at least in part, given their propensity to increase BDNF expression and phosphorylation of AKT, mTOR, and CREB.

Loureirin C and Xanthoceraside Attenuate Depression-Like Behaviors and Expression of Interleukin-17 in the Prefrontal Cortex Induced by Chronic Unpredictable Mild Stress in Mice.

Major depressive disorder (MDD) is the most prevalent and serious psychiatric disease involving inflammation. Loureirin C and Xanthoceraside are extracts of dragon's blood and Xanthoceras sorbifolia Bunge, respectively, which have neuroprotective and anti-inflammatory properties. In this study, we examined whether Loureirin C and Xanthoceraside attenuated depression-like behaviors and inflammation induced by chronic unpredicted mild stress (CUMS) in mice. Adult C57BL/6 J mice exposed to CUMS for 4 weeks showed depression-like behaviors characterized by hyperactivity in a novel environment, decreased interaction time in the social interaction test, prolongation of eating latency in the novelty suppressed feeding test, and increased immobility in the forced swimming test. CUMS increased the expression of interleukin-17 (IL-17) in the prefrontal cortex (PFC). One week after exposure to CUMS, the mice were treated with Loureirin C (0.64 mg/kg) or Xanthoceraside (1.28 mg/kg) once a day for 3 weeks during CUMS. Loureirin C and Xanthoceraside significantly attenuated CUMS-induced behavioral impairment. Furthermore, both Loureirin C and Xanthoceraside prevented IL-17 expression induced by CUMS in the PFC. This data suggests that Loureirin C and Xanthoceraside have antidepressant-like properties that may be associated with the inhibition of IL-17 expression.

Exploring the factors underlying remyelination arrest by studying the post-transcriptional regulatory mechanisms of cystatin F gene.

Remyelination plays an important role in determining the fate of demyelinating disorders. However, it is arrested during chronic disease states. Cystatin F, a papain-like lysosomal cysteine proteinase inhibitor, is a crucial regulator of demyelination and remyelination. Using hemizygous proteolipid protein transgenic 4e (PLP4e/- ) mice, an animal model of chronic demyelination, we found that cystatin F mRNA expression was induced at 2.5 months of age and up-regulated in the early phase of demyelination, but significantly decreased in the chronic phase. We next investigated cystatin F regulatory factors as potential mechanisms of remyelination arrest in chronic demyelinating disorders. We used the CysF-STOP-tetO::Iba-mtTA mouse model, in which cystatin F gene expression is driven by the tetracycline operator. Interestingly, we found that forced cystatin F mRNA over-expression was eventually decreased. Our findings show that cystatin F expression is modulated post-transcriptionally. We next identified embryonic lethal, abnormal vision, drosophila like RNA-binding protein 1 (ELAVL-1), and miR29a as cystatin F mRNA stabilizing and destabilizing factors, respectively. These roles were confirmed in vitro in NIH3T3 cells. Using postmortem plaque samples from human multiple sclerosis patients, we also confirmed that ELAVL-1 expression was highly correlated with the previously reported expression pattern of cystatin F. These data indicate the important roles of ELAVL-1 and miR29a in regulating cystatin F expression. Furthermore, they provide new insights into potential therapeutic targets for demyelinating disorders.

Shati/Nat8l deficiency disrupts adult neurogenesis and causes attentional impairment through dopaminergic neuronal dysfunction in the dentate gyrus.

Successful completion of daily activities relies on the ability to select the relevant features of the environment for memory and recall. Disruption to these processes can lead to various disorders, such as attention-deficit hyperactivity disorder (ADHD). Dopamine is a neurotransmitter implicated in the regulation of several processes, including attention. In addition to the higher-order brain function, dopamine is implicated in the regulation of adult neurogenesis. Previously, we generated mice lacking Shati, an N-acetyltransferase-8-like protein on a C57BL/6J genetic background (Shati/Nat8l-/- ). These mice showed a series of changes in the dopamine system and ADHD-like behavioral phenotypes. Therefore, we hypothesized that deficiency of Shati/Nat8l would affect neurogenesis and attentional behavior in mice. We found aberrant morphology of neurons and impaired neurogenesis in the dentate gyrus of Shati/Nat8l-/- mice. Additionally, research has suggested that impaired neurogenesis might be because of the reduction of dopamine in the hippocampus. Galantamine (GAL) attenuated the attentional impairment observed in the object-based attention test via increasing the dopamine release in the hippocampus of Shati/Nat8l-/- mice. The α7 nicotinic acetylcholine receptor antagonist, methyllycaconitine, and dopamine D1 receptor antagonist, SCH23390, blocked the ameliorating effect of GAL on attentional impairment in Shati/Nat8l-/- mice. These results suggest that the ameliorating effect of GAL on Shati/Nat8l-/- attentional impairment is associated with activation of D1 receptors following increased dopamine release in the hippocampus via α7 nicotinic acetylcholine receptor. In summary, Shati/Nat8l is important in both morphogenesis and neurogenesis in the dentate gyrus and attention, possible via modulation of dopaminergic transmission. Cover Image for this issue: https://doi.org/10.1111/jnc.15061.

Prefrontal cortex miR-874-3p prevents lipopolysaccharide-induced depression-like behavior through inhibition of indoleamine 2,3-dioxygenase 1 expression in mice.

Indoleamine 2,3-dioxygenase 1 (IDO1) is the first rate-limiting enzyme that metabolizes tryptophan to the kynurenine pathway. Its activity is highly inducible by pro-inflammatory cytokines and correlates with the severity of major depressive disorder (MDD). MicroRNAs (miRNAs) are involved in gene regulation and the development of neuropsychiatric disorders including MDD. However, the role of miRNAs in targeting IDO1 in the pathophysiology of MDD is still unknown. In this study, we investigated the role of novel miRNAs in the regulation of IDO1 activity and its effect on lipopolysaccharide (LPS)-induced depression-like behavior in mice. LPS up-regulated miR-874-3p concomitantly with increase in IDO1 expression in the prefrontal cortex (PFC), increase in immobility in the forced swimming test as depression-like behavior and decrease in locomotor activity as sickness behavior without motor dysfunction. The miR-874-3p increased in both neuron and microglia after LPS. Its mimic significantly suppressed LPS-induced IDO1 expression in the PFC. Infusion of IDO1 inhibitor (1-methyl-l-tryptophan) and miR-874-3p into PFC prevented an increase in immobility in the forced swimming test, but did not decrease in locomotor activity induced by LPS. These results suggest that miR-874-3p may play an important role in preventing the LPS-induced depression-like behavior through inhibition of IDO1 expression. This may also serve as a novel potential target molecule for the treatment of MDD.

Heat-sterilized Bifidobacterium breve prevents depression-like behavior and interleukin-1ß expression in mice exposed to chronic social defeat stress.

Major depressive disorder (MDD) is a common and serious psychiatric disease that involves brain inflammation. Bifidobacterium breve is commonly used as a probiotic and was shown to improve colitis and allergic diseases by suppressing the inflammatory response. Heat-sterilized B. breve has beneficial effects on inflammation. We hypothesize, therefore, that this probiotic might reduce depression symptoms. We tested this is a mouse model of social defeat stress. C57BL/6J mice exposed to chronic social defeat stress (CSDS) for five consecutive days developed a mild depression-like behavior characterized by a social interaction impairment. CSDS also altered the gut microbiota composition, such as increased abundance of Bacilli, Bacteroidia, Mollicutes, and Verrucomicrobiae classes and decreased Erysipelotrichi class. The prophylactic effect of heat-sterilized B. breve as a functional food ingredient was evaluated on the depression-like behavior in mice. The supplementation started two weeks before and lasted two weeks after the last exposure to CSDS. Two weeks after CSDS, the mice showed deficits in social interaction and increased levels of inflammatory cytokines, including interleukin-1ß (IL-1ß) in the prefrontal cortex (PFC) and hippocampus (HIP). Heat-sterilized B. breve supplementation significantly prevented social interaction impairment, suppressed IL-1ß increase in the PFC and HIP, and modulated the alteration of the gut microbiota composition induced by CSDS. These findings suggest that heat-sterilized B. breve prevents depression-like behavior and IL-1ß expression induced by CSDS through modulation of the gut microbiota composition in mice. Therefore, heat-sterilized B. breve used as an ingredient of functional food might prevent MDD.

Indoleamine 2,3-dioxygenase 2 depletion suppresses tumor growth in a mouse model of Lewis lung carcinoma.

Tryptophan metabolism is important to induce immune tolerance in tumors. To date, 3 types of tryptophan-metabolizing enzymes have been identified: indoleamine 2,3-dioxygenase 1 and 2 (IDO1 and IDO2) and tryptophan 2,3-dioxygenase 2. Numerous studies have focused on IDO1 as its expression is enhanced in various cancers. Recently, IDO2 has been identified as a tryptophan-metabolizing enzyme that is involved in several immune functions and expressed in cancers such as pancreatic cancer. However, the biological role of IDO2 in the induction of immune tolerance in tumors has not yet been reported. In the present study, we examined the effects of Ido2 depletion on tumor growth in a mouse model of Lewis lung carcinoma by using Ido2-knockout mice. Ido2-knockout mice had reduced tumor volumes compared to WT mice. Furthermore, Ido2 depletion altered the tumor microenvironment, such as tryptophan accumulation and kynurenine reduction, leading to enhancement of immune cell invasion. Finally, enzyme-linked immunospot assay revealed that Ido2 depletion enhanced γ-interferon secretion in the tumor. In conclusion, Ido2 is an important immune regulator in the tumor microenvironment. Our data indicate that IDO2 is a potential target for cancer treatment and drug development.

Dysregulation of schizophrenia-related aquaporin 3 through disruption of paranode influences neuronal viability.

Myelinated axons segregate the axonal membrane into four defined regions: the node of Ranvier, paranode, juxtaparanode, and internode. The paranodal junction consists of specific component proteins, such as neurofascin155 (NF155) on the glial side, and Caspr and Contactin on the axonal side. Although paranodal junctions are thought to play crucial roles in rapid saltatory conduction and nodal assembly, the role of their interaction with neurons is not fully understood. In a previous study, conditional NF155 knockout in oligodendrocytes led to disorganization of the paranodal junctions. To examine if disruption of paranodal junctions affects neuronal gene expression, we prepared total RNA from the retina of NF155 conditional knockout, and performed expression analysis. We found that the expression level of 433 genes changed in response to paranodal junction ablation. Interestingly, expression of aquaporin 3 (AQP3) was significantly reduced in NF155 conditional knockout mice, but not in cerebroside sulfotransferase knockout (CST-KO) mice, whose paranodes are not originally formed during development. Copy number variations have an important role in the etiology of schizophrenia (SCZ). We observed rare duplications of AQP3 in SCZ patients, suggesting a correlation between abnormal AQP3 expression and SCZ. To determine if AQP3 over-expression in NF155 conditional knockout mice influences neuronal function, we performed adeno-associated virus (AAV)-mediated over-expression of AQP3 in the motor cortex of mice and found a significant increase in caspase 3-dependent neuronal apoptosis in AQP3-transduced cells. This study may provide new insights into therapeutic approaches for SCZ by regulating AQP3 expression, which is associated with paranodal disruption.

Social Behavior in Animal Models of Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder characterized by social deficits and stereotyped, repetitive patterns of behaviors, limited interests, and cognitive impairment. Especially, social deficit has been considered a core feature of ASD. Because of the limitations of the experimental approach in humans, valid animal models are essential in an effort to identify novel therapeutics for social deficits in ASD. The genetic and environmental factors are clinically relevant to the pathophysiology of ASD. Epidemiological studies demonstrate environmental interventions such as prenatal exposure to valproic acid (VPA). Prenatal exposure to VPA represents a robust model of ASD exhibiting face, construct, and predictive validity. Here, we introduce protocols of the social interaction test and the three-chamber test for evaluating social deficits in mice prenatally exposed to VPA.

Indoleamine 2,3-dioxygenase 2 deficiency associates with autism-like behavior via dopaminergic neuronal dysfunction.

Indoleamine 2,3-dioxygenase 2 (IDO2) is an enzyme of the tryptophan-kynurenine pathway that is constitutively expressed in the brain. To provide insight into the physiological role of IDO2 in the brain, behavioral and neurochemical analyses in IDO2 knockout (KO) mice were performed. IDO2 KO mice showed stereotyped behavior, restricted interest and social deficits, traits that are associated with behavioral endophenotypes of autism spectrum disorder (ASD). IDO2 was colocalized immunohistochemically with tyrosine-hydroxylase-positive cells in dopaminergic neurons. In the striatum and amygdala of IDO2 KO mice, decreased dopamine turnover was associated with increased α-synuclein level. Correspondingly, levels of downstream dopamine D1 receptor signaling molecules such as brain-derived neurotrophic factor and c-Fos positive proteins were decreased. Furthermore, decreased abundance of ramified-type microglia resulted in increased dendritic spine density in the striatum of IDO2 KO mice. Both chemogenetic activation of dopaminergic neurons and treatment with methylphenidate, a dopamine reuptake inhibitor, ameliorated the ASD-like behavior of IDO2 KO mice. Sequencing analysis of exon regions in IDO2 from 309 ASD samples identified a rare canonical splice site variant in one ASD case. These results suggest that the IDO2 gene is, at least in part, a factor closely related to the development of psychiatric disorders.

Ido2 Deficiency Exacerbates Motor Impairment and Reduces Aryl Hydrocarbon Receptor Activity through Decreased Kynurenine in a Chronic Demyelinating Mouse Model.

Demyelinating diseases including multiple sclerosis (MS) are chronic inflammatory diseases of the central nervous system. Indoleamine 2,3-dioxygenase 2 (Ido2) is a recently identified as catalytic enzyme involved in the rate-limiting step of the tryptophan-kynurenine pathway that influences susceptibility to inflammatory diseases. However, the pathological role of Ido2 in demyelination remains unclear. In this study, we investigated whether Ido2 deficiency influences the pathogenesis of proteolipid protein transgenic (Plp tg) mice, an animal model of chronic demyelination. Ido2 deficiency exacerbates impairments of motor function in the locomotor activity test, wire hanging test, and rotarod test. Ido2 deficiency caused severe demyelination associated with CD68-positive microglial activation in Plp tg mice. In the cerebellum of Plp tg mice, Ido2 deficiency significantly increased the expression of Tnfα. Ido2 deficiency reduced tryptophan metabolite kynurenine (KYN) levels and subsequent aryl hydrocarbon receptor (AhR) activity, which play an important role in anti-inflammatory response. These results suggest that Ido2 has an important role in preventing demyelination through AhR. Taken together, Ido2 could be a potential therapeutic target for demyelinating diseases.

[Diagnoses and new therapeutic strategy focused on physiological alteration of tryptophan metabolism].

The monoamine hypothesis has been common hypotheses for the pathophysiology of major depressive disorder (MDD). Since mainstream antidepressants are selective serotonin (5-HT) reuptake inhibitors, hypo-serotonergic function has been implicated in the MDD. However, one-third of patients are refractory to the treatment with antidepressants. Tryptophan (TRP) is metabolized via the kynurenine (KYN) and 5-HT pathways. Indoleamine 2,3-dioxygenase 1 (IDO1) is the first metabolizing enzyme in the TRP-KYN pathway which is inducible by pro-inflammatory cytokines, involved depression-like behavior via 5-HT depletion due to decreased level of TRP in the 5-HT pathway. Kynurenine 3-monooxygenase (KMO) is the enzyme in the metabolism of KYN to 3-hydroxykynurenine. KMO deficiency increases level of kynurenic acid (KA), a KYN metabolite by kynurenine aminotransferases (KATs) and induces depression-like behavior. Interestingly, Chronic unpredictable mild stress (CUMS) is associated with a disruption of the hypothalamus-pituitary-adrenocortical (HPA) system and increases KA level with decreased KMO expression in the prefrontal cortex. The decrease of KMO may be related to the reduction in expression of microglia, since KMO is mainly found in microglia in the nervous system. CUMS increases KA level via alternation of enzymes from KMO to KAT. KA is α7 nicotinic acetylcholine receptor (α7nAChR) antagonist. Activation of α7nAChR by nicotine or galantamine attenuates CUMS-induced depression-like behaviors. Taken together, depletion of 5-HT by induction of IDO1 and α7nAChR antagonism by KA via decreased KMO expression cause depression-like behavior, suggesting that metabolic alterations in TRP-KYN pathway are highly involved in the pathophysiology of MDD. Therefore, TRP-KYN pathway is expected to be an attractive target for the development of novel diagnosis of MDD and antidepressants.

Treatment of tubular damage in high-fat-diet-fed obese mice using sodium-glucose co-transporter inhibitors.

A long-term high-fat diet (HFD) causes obesity and changes in renal lipid metabolism and lysosomal dysfunction in mice, causing renal damage. Sodium-glucose co-transporter inhibitors, including phlorizin, exert nephroprotective effects in patients with chronic kidney disease, but the underlying mechanism remains unclear. A HFD or standard diet was fed to adult C57BL/6J male mice, and phlorizin was administered. Lamellar body components of the proximal tubular epithelial cells (PTECs) were investigated. After phlorizin administration in HFD-fed mice, sphingomyelin and ceramide in urine and tissues were assessed and label-free quantitative proteomics was performed using kidney tissue samples. Mitochondrial elongation by fusion was effective in the PTECs of HFD-fed obese mice under phlorizin administration, and many lamellar bodies were found in the apical portion of the S2 segment of the proximal tubule. Phlorizin functioned as a diuretic, releasing lamellar bodies from the apical membrane of PTECs and clearing the obstruction in nephrons. The main component of the lamellar bodies was sphingomyelin. On the first day of phlorizin administration in HFD-fed obese mice, the diuretic effect was increased, and more sphingomyelin was excreted through urine than in vehicle-treated mice. The expressions of three peroxisomal ß-oxidation proteins involved in fatty acid metabolism were downregulated after phlorizin administration in the kidneys of HFD-fed mice. Fatty acid elongation protein levels increased with phlorizin administration, indicating an increase in long-chain fatty acids. Lamellar bodies accumulated in the proximal renal tubule of the S2 segment of the HFD-fed mice, indicating that the urinary excretion of lamellar bodies has nephroprotective effects.

High salt induces cognitive impairment via the interaction of the angiotensin II-AT1 and prostaglandin E2-EP1 systems.

BACKGROUND AND PURPOSE: High salt (HS) intake has been associated with hypertension and cognitive impairment. It is well known that the angiotensin II (Ang II)-AT1 receptor and prostaglandin E2 (PGE2)-EP1 receptor systems are involved in hypertension and neurotoxicity. However, the involvement of these systems in HS-mediated hypertension and emotional and cognitive impairments remains unclear. EXPERIMENTAL APPROACH: Mice were loaded with HS solution (2% NaCl drinking water) for 12 weeks, and blood pressure was monitored. Subsequently, effects of HS intake on emotional and cognitive function and tau phosphorylation in the prefrontal cortex (PFC) and hippocampus (HIP) were investigated. The involvement of Ang II-AT1 and PGE2-EP1 systems in HS-induced hypertension and neuronal and behavioural impairments was examined by treatment with losartan, an AT1 receptor blocker (ARB), or EP1 gene knockout. KEY RESULTS: We demonstrate that hypertension and impaired social behaviour and object recognition memory following HS intake may be associated with tau hyperphosphorylation, decreased phosphorylation of Ca2+ /calmodulin-dependent protein kinase II (CaMKII), and postsynaptic density protein 95 (PSD95) expression in the PFC and HIP of mice. These changes were blocked by pharmacological treatment with losartan or EP1 receptor gene knockout. CONCLUSIONS AND IMPLICATIONS: Our findings suggest that the interaction of Ang II-AT1 receptor and PGE2-EP1 receptor systems could be novel therapeutic targets for hypertension-induced cognitive impairment.

Pharmacological blockade of dopamine D1- or D2-receptor in the prefrontal cortex induces attentional impairment in the object-based attention test through different neuronal circuits in mice.

Dopamine is a key neurotransmitter that regulates attention through dopamine D1 and D2-receptors in the prefrontal cortex (PFC). We previously developed an object-based attention test (OBAT) to evaluate attention in mice. Disruption of the dopaminergic neuronal system in the PFC induced attentional impairment in the OBAT. However, previous studies have not systematically examined which specific brain regions are associated with the blockade of PFC dopamine D1 and D2-receptors in the OBAT. In this study, we investigated the association of dopamine D1 and D2-receptors in the PFC with attention and neuronal activity in diverse brain regions. We found that both dopamine D1 and D2-receptor antagonists induced attentional impairment in the OBAT by bilateral microinjection into the PFC of mice, suggesting that both dopamine D1 and D2-receptors were associated with attention in the OBAT. Our analysis of the neuronal activity as indicated by c-Fos expression in 11 different brain regions showed that based on the antagonist types, there was selective activation of several brain regions. Overall, this study suggests that both dopamine D1 and D2-receptors play a role in attention through different neuronal circuits in the PFC of mice.

Kynurenine 3-monooxygenase deficiency induces depression-like behavior via enhanced antagonism of α7 nicotinic acetylcholine receptors by kynurenic acid.

Tryptophan (TRP) is metabolized via the kynurenine (KYN) pathway, which is related to the pathogenesis of major depressive disorder (MDD). Kynurenine 3-monooxygenase (KMO) is a pivotal enzyme in the metabolism of KYN to 3-hydroxykynurenine. In rodents, KMO deficiency induces a depression-like behavior and increases the levels of kynurenic acid (KA), a KYN metabolite formed by kynurenine aminotransferases (KATs). KA antagonizes α7 nicotinic acetylcholine receptor (α7nAChR). Here, we investigated the involvement of KA in depression-like behavior in KMO knockout (KO) mice. KYN, KA, and anthranilic acid but not TRP or 3-hydroxyanthranilic acid were elevated in the prefrontal cortex of KMO KO mice. The mRNA levels of KAT1 and α7nAChR but not KAT2-4, α4nAChR, or ß2nAChR were elevated in the prefrontal cortex of KMO KO mice. Nicotine blocked increase in locomotor activity, decrease in social interaction time, and prolonged immobility in a forced swimming test, but it did not decrease sucrose preference in the KMO KO mice. Methyllycaconitine (an α7nAChR antagonist) antagonized the effect of nicotine on decreased social interaction time and prolonged immobility in the forced swimming test, but not increased locomotor activity. Galantamine (an α7nAChR allosteric agonist) blocked the increased locomotor activity and prolonged immobility in the forced swimming test, but not the decreased social interaction time in the KMO KO mice. In conclusion, elevation of KA levels contributes to depression-like behaviors in KMO KO mice by α7nAChR antagonism. The ameliorating effects of nicotine and galantamine on depression-like behaviors in KMO KO mice are associated with the activation of α7nAChR.

Prefrontal cortex, dorsomedial striatum, and dentate gyrus are necessary in the object-based attention test in mice.

Disturbances of attention are a common behavioral feature associated with neuropsychiatric disorders with largely unknown underlying causes. We previously developed an object-based attention test (OBAT) as a simple and practical method for evaluating attention in mice. Since its establishment, the test has become a popular method for assessing attention and related underlying mechanisms in various mouse models. However, the underlying neuronal network involved in this test has yet to be studied. The purpose of this study was to identify the principal brain regions activated in the OBAT. Accordingly, C57BL/6J mice were subjected to the OBAT and thereafter prepared for immunohistochemical quantification of c-Fos, an immediate early gene that is frequently used as a marker of neuronal activity, in 13 different brain regions. The number of c-Fos-positive cells was significantly higher in the prefrontal cortex (PFC), dorsomedial striatum (DMS), and dentate gyrus (DG) in the test group as compared to the control group. The neuronal activation of these brain regions during the OBAT indicates that these brain regions are necessary for the regulation of attention in this test. This was supported by excitotoxic lesioning of these brain regions, leading to impaired attention without causing locomotor dysfunction. This study is one of the first attempts to analyze the brain regions that regulate attention in the OBAT. These findings provide an initial insight into the role of these brain regions and ideas for studying the underlying neural and molecular mechanisms.

Focal loss of the paranodal domain protein Neurofascin155 in the internal capsule impairs cortically induced muscle activity in vivo.

Paranodal axoglial junctions are essential for rapid nerve conduction and the organization of axonal domains in myelinated axons. Neurofascin155 (Nfasc155) is a glial cell adhesion molecule that is also required for the assembly of these domains. Previous studies have demonstrated that general ablation of Nfasc155 disorganizes these domains, reduces conduction velocity, and disrupts motor behaviors. Multiple sclerosis (MS), a typical disorder of demyelination in the central nervous system, is reported to have autoantibody to Nfasc. However, the impact of focal loss of Nfasc155, which may occur in MS patients, remains unclear. Here, we examined whether restricted focal loss of Nfasc155 affects the electrophysiological properties of the motor system in vivo. Adeno-associated virus type5 (AAV5) harboring EGFP-2A-Cre was injected into the glial-enriched internal capsule of floxed-Neurofascin (NfascFlox/Flox) mice to focally disrupt paranodal junctions in the cortico-fugal fibers from the motor cortex to the spinal cord. Electromyograms (EMGs) of the triceps brachii muscles in response to electrical stimulation of the motor cortex were successively examined in these awake mice. EMG analysis showed significant delay in the onset and peak latencies after AAV injection compared to control (Nfasc+/+) mice. Moreover, EMG half-widths were increased, and EMG amplitudes were gradually decreased by 13 weeks. Similar EMG changes have been reported in MS patients. These findings provide physiological evidence that motor outputs are obstructed by focal ablation of paranodal junctions in myelinated axons. Our findings may open a new path toward development of a novel biomarker for an early phase of human MS, as Nfasc155 detects microstructural changes in the paranodal junction.