Author Correction: Behavioral abnormalities with disruption of brain structure in mice overexpressing VGF.

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GPNMB Induces BiP Expression by Enhancing Splicing of BiP Pre-mRNA during the Endoplasmic Reticulum Stress Response.

Glycoprotein nonmetastatic melanoma protein B (GPNMB) has a neuroprotective effect against neuronal cell death caused by the accumulation of abnormal mutated proteins. It is known that the accumulation of pathological proteins induces endoplasmic-reticulum (ER) stress leading to cell damage. The aim of this study was to determine the role of GPNMB in the ER stress response. GPNMB was greatly up-regulated by thapsigargin-induced ER stress. Under the ER stress conditions, GPNMB relocated to the nucleus and specifically up-regulated expression of BiP at the mRNA level by promoting the BiP pre-mRNA splicing, not through the pathways initiated by the three major transducers of the unfolded protein response: IRE1, PERK, and ATF6. Furthermore, we found that the protein level of BiP and the infarction were increased and attenuated, respectively, in Gpnmb-transgenic mice after occlusion of the middle cerebral artery, in comparison with wild-type mice. Thus, our findings indicate that GPNMB enhances the BiP expression by promoting the splicing (thereby preventing cell death caused by ER stress) and could be a therapeutic target in ER stress-related disorders.

Behavioral abnormalities with disruption of brain structure in mice overexpressing VGF.

VGF nerve growth factor inducible (VGF) is a neuropeptide induced by nerve growth factor and brain-derived neurotrophic factor. This peptide is involved in synaptic plasticity, neurogenesis, and neurite growth in the brain. Patients with depression and bipolar disorder have lower-than-normal levels of VGF, whereas patients with schizophrenia and other cohorts of patients with depression have higher-than-normal levels. VGF knockout mice display behavioral abnormalities such as higher depressive behavior and memory dysfunction. However, it is unclear whether upregulation of VGF affects brain function. In the present study, we generated mice that overexpress VGF and investigated several behavioral phenotypes and the brain structure. These adult VGF-overexpressing mice showed (a) hyperactivity, working memory impairment, a higher depressive state, and lower sociality compared with wild-type mice; (b) lower brain weight without a change in body weight; (c) increased lateral ventricle volume compared with wild-type mice; and (d) striatal morphological defects. These results suggest that VGF may modulate a variety of behaviors and brain development. This transgenic mouse line may provide a useful model for research on mental illnesses.

Distribution and function of hyaluronan binding protein involved in hyaluronan depolymerization (HYBID, KIAA1199) in the mouse central nervous system.

HYBID (HYaluronan Binding Protein Involved in hyaluronan [HA] Depolymerization, KIAA1199) is one of the HA binding proteins that is involved in the depolymerization of HA. HYBID mRNA is highly expressed in the brain, however, the role of HYBID in the brain remains unclear. In this study, we bred Hybid knock-out (KO) mice and evaluated the function of Hybid in the central nervous system. Hybid mRNA was expressed in the brain, especially in the hippocampus and cerebellum, in wild-type mice. Hybid KO mice demonstrated decreased mnemonic ability in novel object recognition and Morris water maze tests. The average molecular mass of hippocampal HA increased in KO mice, accompanied by a significant increase in the total HA amount. Hybid KO mice did not differ in behavior from wild-type mice in the open field test, evaluation of acoustic startle responses, or drug-induced seizure test. In real-time PCR, Hyal1 and Hyal2 mRNA levels, which code hyaluronidases 1 and 2, respectively, did not differ between the Hybid KO and wild-type mouse brain. These results indicate that Hybid plays a key role in memory function in the brain.

Cilostazol ameliorates collagenase-induced cerebral hemorrhage by protecting the blood-brain barrier.

Intracranial hemorrhage remains a devastating disease. Among antiplatelet drugs, cilostazol, a phosphodiesterase 3 inhibitor, was recently reported to prevent secondary hemorrhagic stroke in patients in a clinical trial. The aim of this study was to evaluate whether pre-treatment with cilostazol could decrease the intracranial hemorrhage volume and examine the protective mechanisms of cilostazol. We evaluated the pre-treatment effects of the antiplatelet drug cilostazol on the collagenase-induced intracranial hemorrhage volume and neurological outcomes in mice. To estimate the mechanism of collagenase injury, we evaluated various vascular components in vitro, including endothelial cells, vascular smooth muscle cells, pericytes, and a blood-brain barrier model. Cilostazol pre-treatment reduced the intracranial hemorrhage volume with sufficient inhibition of platelet aggregation, and motor function was improved by cilostazol treatment. Blood-brain barrier permeability was increased by collagenase-induced intracranial hemorrhage, and cilostazol attenuated blood-brain barrier leakage. Terminal deoxynucleotidyl transferase dUTP nick-end labeling and western blot analysis showed that cilostazol prevented pericyte cell death by inducing cyclic adenosine monophosphate-responsive element-binding protein phosphorylation. Cilostazol also prevented endothelial cell death and protected collagen type 4, laminin, and vascular endothelial- and N-cadherins from collagenase injury. In conclusion, cilostazol reduced collagenase-induced intracranial hemorrhage volume by protecting the blood-brain barrier.

Memantine, an NMDA receptor antagonist, improves working memory deficits in DGKß knockout mice.

Diacylglycerol kinase (DGK) ß is a type 1 isozyme of the DGK family. We previously reported that DGKß was deeply involved in neurite spine formation, and DGKß knockout (KO) mice exhibited behavioral abnormalities concerning spine formation, such as cognitive, emotional, and attentional impairment. Moreover, some of these abnormalities were ameliorated by the administration of a mood stabilizer. However, there is no data about how memory-improving drugs used in the treatment of Alzheimer's disease affect DGKß KO mice. In the present study, we evaluated the effect of an anti-Alzheimer's drug, memantine on the working memory deficit observed in DGKß KO mice. In the Y-maze test, the administration of memantine significantly improved working memory of DGKß KO mice. We also found that the expression levels of the NR2A and NR2B N-methyl-d-aspartate (NMDA) receptor subunits were increased in the prefrontal cortex, but decreased in the hippocampus of DGKß KO mice. These altered expression levels of NR2 subunits might be related to the effect of an NMDA receptor antagonist, memantine. Taken together, these findings may support the hypothesis that DGKß has a pivotal role in cognitive function.

Crocetin protects ultraviolet A-induced oxidative stress and cell death in skin in vitro and in vivo.

Crocetin, the aglycone of crocin, is a carotenoid found in fruits of gardenia (Gardeina jasminoides Ellis) and saffron (Crocus sativus L.). We investigated the protective effects of crocetin against ultraviolet-A (UV-A)-induced skin damage and explored the underlying mechanism. Human skin-derived fibroblasts cells (NB1-RGB) were damaged by exposure to UV-A irradiation (10J/cm(2)). Crocetin protected these cells against cell death and reduced the production of reactive oxygen species induced by UV-A irradiation. Crocetin treatment also suppressed induction of caspase-3 activation by UV-A irradiation. The effects of crocetin against oxidative stress were also examined by imaging of Keap1-dependent oxidative stress detector (OKD) mice. UV-A irradiation upregulated oxidative stress in the OKD mice skin, while crocetin administration (100mg/kg, p.o.) ameliorated this oxidative stress. Crocetin administration also decreased lipid peroxidation in the skin. These findings suggest that crocetin its observed protective effects against UV-A induced skin damage by reducing reactive oxygen species production and cell apoptosis.

Modulation of endoplasmic reticulum stress in Parkinson's disease.

Parkinson's disease is a neurodegenerative disease characterized by the loss of dopaminergic neurons in the substantia nigra in the midbrain. However, the etiology of the reduction in dopaminergic neurons remains unclear. Recently, it has been suggested that oxidative stress, endoplasmic reticulum (ER) stress, and mitochondrial dysfunction are involved in neuronal cell death in the pathology of Parkinson's disease. Furthermore, it has been suggested that some existing anti- Parkinson's disease drugs have protective effects against cell death. Among these, zonisamide exerts neuroprotective effects partly by modulating ER stress. Elucidating the involvement of ER stress in in vitro and in vivo Parkinson's disease models and investigating the mechanisms against ER stress will contribute to the search for new therapeutic agents for Parkinson's disease.

Japanese Huperzia serrata extract and the constituent, huperzine A, ameliorate the scopolamine-induced cognitive impairment in mice.

Huperzia serrata has been used as a Chinese folk medicine for many years. It contains huperzine A, which has a protective effect against memory deficits in animal models; however, it is unclear if H. serrata extract exerts any effects in Alzheimer's disease (AD) models. We used H. serrata collected in Japan and determined its huperzine A content using HPLC. We determined its inhibitory effects on acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) activity. H. serrata extract (30 mg/kg/day) and donepezil (10 mg/kg/day) were orally administrated for 7 days. After repeated administration, we performed the Y-maze and passive avoidance tests. H. serrata extract contained 0.5% huperzine A; H. serrata extract inhibited AChE, but not BuChE. H. serrata extract ameliorated cognitive function in mice. These results indicate that Japanese H. serrata extract ameliorates cognitive function deficits by inhibiting AChE. Therefore, H. serrata extract may be valuable for the prevention or treatment of dementia in AD.

Glucagon-like peptide-1 protects the murine hippocampus against stressors via Akt and ERK1/2 signaling.

Alzheimer's disease (AD) is a common neurodegenerative disease characterized by cognitive dysfunction and neuronal cell death in the hippocampus and cerebral cortex. Glucagon-like peptide-1 (GLP-1) is an insulinotropic peptides. GLP-1-associated medicines are widely used as treatments for type 2 diabetes. In addition, they have been shown to ameliorate pathology in AD mouse models. Here, we investigated the effects of GLP-1 on different stressors in murine hippocampal HT22 cells. GLP-1 (7-36) prevented H2O2-, l-glutamate-, tunicamycin-, thapsigargin-, and amyloid ß1-42-induced neuronal cell death in a concentration-dependent manner. GLP-1 (7-36) treatment for 1 h significantly increased phosphorylated Akt and extracellular signal-regulated kinase 1 and 2 (ERK1/2) when compared with vehicle-treatment. These results suggest that GLP-1 (7-36) is protective against these stressors via activation of survival signaling molecules, such as Akt and ERK1/2 in HT22 cells. In conclusion, GLP-1 and activators of the GLP-1 receptor might be useful targets for the treatment of AD.

The effects of valproate and olanzapine on the abnormal behavior of diacylglycerol kinase ß knockout mice.

BACKGROUND: Diacylglycerol kinase (DGK) is an enzyme that converts diacylglycerol to phosphatidic acid. Previously, we reported that DGKß knockout (KO) mice showed mania-like behaviors such as hyperactivity, reduced anxiety, and cognitive impairment. Furthermore, lithium ameliorated the hyperactivity and reduced anxiety of DGKß KO mice. In this study, we investigated the effects of the clinically active antimanic drugs valproate and olanzapine on the abnormal behaviors of DGKß KO mice. METHODS: Valproate (100mg/kg/day) and olanzapine (1mg/kg/day) were administered intraperitoneally. Following drugs treatments, behavioral tests were performed to investigate locomotor activity, anxiety levels, and cognitive function of the mice. RESULTS: A single treatment of valproate and olanzapine did not ameliorate the hyperactivity or abnormal anxiety level of DGKß KO mice. Chronic treatment with valproate and olanzapine significantly decreased locomotor activity and abnormal anxiety levels of DGKß KO mice. Additionally, valproate also ameliorated cognitive function of DGKß KO mice. CONCLUSION: These results suggest that the abnormal behaviors of DGKß KO mice is responsive to antimanic drugs, and that DGKß KO mice are useful as an animal model of mania.

Zonisamide suppresses endoplasmic reticulum stress-induced neuronal cell damage in vitro and in vivo.

Zonisamide has been reported to have protective effects on epilepsy and Parkinson׳s disease and to work via various mechanisms of action, such as inhibition of monoamine oxidase-B and enhancement of tyrosine hydroxylase. Recently, it has been suggested that zonisamide itself shows neuroprotective actions. Therefore, in the present study we investigated the neuroprotective effects of zonisamide against endoplasmic reticulum (ER) stress. We used human neuroblastoma (SH-SY5Y) cells and investigated the protective effects of zonisamide against tunicamycin- and thapsigargin-induced neuronal cell death. In addition, we investigated the effect of zonisamide against 1-methyl-4-phenylpyridinium (MPP⁺)-induced cell death and the mechanism of protection against ER stress. In vivo, we investigated the effect of zonisamide (20 mg/kg, p.o.) in the 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced mouse model of Parkinson׳s disease. Zonisamide not only suppressed MPP⁺-induced cell death, but also inhibited ER stress-induced cell death and suppressed the expression of ER stress-related factors such as C/EBO homologous protein (CHOP) in vivo. Furthermore, zonisamide inhibited the activation of caspase-3 in vitro. These results suggest that zonisamide affected ER stress via caspase-3. We think that ER stress, particularly the mechanism via caspase-3, is involved in part of the neuroprotective effect of zonisamide against the experimental models of Parkinson׳s disease.

The extracellular fragment of GPNMB (Glycoprotein nonmelanosoma protein B, osteoactivin) improves memory and increases hippocampal GluA1 levels in mice.

Glycoprotein nonmelanoma protein B (GPNMB, alias osteoactivin), a type I transmembrane glycoprotein, is cleaved by extracellular proteases, resulting in release of an extracellular fragment (ECF). GPNMB is widely expressed by neurons within the CNS, including the hippocampus; however, its function in the brain remains unknown. Here, we investigated the role of GPNMB in memory and learning by using transgenic (Tg) mice over-expressing GPNMB (Tg mice on a BDF-1 background) and ECF-treated mice. In the hippocampus of both wild-type and Tg mice, GPNMB was highly expressed in neurons and astrocytes. Tg mice exhibited memory improvements in two types of learning tasks but were impaired in a passive-avoidance test. In Tg mice, the hippocampus displayed increased levels of the α-amino-3-hydroxy-5-methylisoxazole-4-propionate receptor subunit GluA1. Intracerebroventricular administration of ECF (50 ng) to Institute of Cancer Research (ICR) mice also improved memory in a passive-avoidance test and increased hippocampal GluA1 levels 24 h after treatment. In Tg mice and ECF (0.25 µg/mL)-treated hippocampal slices, long-term potentiation was promoted. These findings suggest that GPNMB may be a novel target for research on higher order brain functions.

The roles of diacylglycerol kinases in the central nervous system: review of genetic studies in mice.

Diacylglycerol kinase (DGK) is an enzyme that converts diacylglycerol to phosphatidic acid. To date, 10 isoforms of DGKs (α, ß, γ, δ, ε, ζ, η, θ, ι, and κ) have been identified in mammals, and these DGKs show characteristic expression patterns and roles. The expression levels of DGKs are comparatively higher in the central nervous system than in other organs and may play several important roles in regulating higher brain functions. Currently, many studies have been performed to reveal the roles of DGKs by knocking down or overexpression of DGKs in vitro. Additionally, knockout or overexpression mice of several DGKs have been generated, and phenotypes of these mice have been studied. In this review, we discuss the roles of DGKs in the central nervous system based on recent findings in genetic models.

Imipramine protects mouse hippocampus against tunicamycin-induced cell death.

Endoplasmic reticulum (ER) stress is implicated in various diseases. Recently, some reports have suggested that the sigma-1 receptor may play a role in ER stress, and many antidepressants have a high affinity for the sigma-1 receptor. In the present study, we focused on imipramine, a widely used antidepressant, and investigated whether it might protect against the neuronal cell death induced by tunicamycin, an ER stress inducer. In mouse cultured hippocampal HT22 cells, imipramine inhibited cell death and caspase-3 activation induced by tunicamycin, although it did not alter the elevated expressions of 78 kDa glucose-regulated protein (GRP78) and C/EBP-homologous protein (CHOP). Interestingly, in such cells application of imipramine normalized the expression of the sigma-1 receptor, which was decreased by treatment with tunicamycin alone. Additionally, NE-100, a selective sigma-1 receptor antagonist, abolished the protective effect of imipramine against such tunicamycin-induced cell death. Imipramine inhibited the reduction of mitochondrial membrane potential induced by tunicamycin, and NE-100 blocked this modulating effect of imipramine. Furthermore, in anesthetized mice intracerebroventricular administration of tunicamycin decreased the number of neuronal cells in the hippocampus, particularly in the CA1 and dentate gyrus (DG) areas, and 7 days' imipramine treatment (10mg/kg/day; i.p.) significantly suppressed these reductions in CA1 and DG. These findings suggest that imipramine protects against ER stress-induced hippocampal neuronal cell death both in vitro and in vivo. Such protection may be partly due to the sigma-1 receptor.

Diacylglycerol kinase ß knockout mice exhibit attention-deficit behavior and an abnormal response on methylphenidate-induced hyperactivity.

BACKGROUND: Diacylglycerol kinase (DGK) is an enzyme that phosphorylates diacylglycerol to produce phosphatidic acid. DGKß is one of the subtypes of the DGK family and regulates many intracellular signaling pathways in the central nervous system. Previously, we demonstrated that DGKß knockout (KO) mice showed various dysfunctions of higher brain function, such as cognitive impairment (with lower spine density), hyperactivity, reduced anxiety, and careless behavior. In the present study, we conducted further tests on DGKß KO mice in order to investigate the function of DGKß in the central nervous system, especially in the pathophysiology of attention deficit hyperactivity disorder (ADHD). METHODOLOGY/PRINCIPAL FINDINGS: DGKß KO mice showed attention-deficit behavior in the object-based attention test and it was ameliorated by methylphenidate (MPH, 30 mg/kg, i.p.). In the open field test, DGKß KO mice displayed a decreased response to the locomotor stimulating effects of MPH (30 mg/kg, i.p.), but showed a similar response to an N-methyl-d-aspartate (NMDA) receptor antagonist, MK-801 (0.3 mg/kg, i.p.), when compared to WT mice. Examination of the phosphorylation of extracellular signal-regulated kinase (ERK), which is involved in regulation of locomotor activity, indicated that ERK1/2 activation induced by MPH treatment was defective in the striatum of DGKß KO mice. CONCLUSIONS/SIGNIFICANCE: These findings suggest that DGKß KO mice showed attention-deficit and hyperactive phenotype, similar to ADHD. Furthermore, the hyporesponsiveness of DGKß KO mice to MPH was due to dysregulation of ERK phosphorylation, and that DGKß has a pivotal involvement in ERK regulation in the striatum.

Luteolin shows an antidepressant-like effect via suppressing endoplasmic reticulum stress.

Depression is a significant public health problem and some reports indicate an association between depression and endoplasmic reticulum stress. Luteolin is a flavonoid contained in many plants and with a variety of known pharmacological properties such as anti-inflammatory, anti-anxiety, and memory-improving effects, suggesting that luteolin penetrates into the brain. In the present study, we investigated the effects of luteolin on endoplasmic reticulum stress-induced neuronal cell death. Luteolin significantly suppressed tunicamycin-induced cell death at 1 to 10 µM in human neuroblastoma cells. Luteolin increased in the expression of the 78 kDa glucose-regulated protein and 94 kDa glucose-regulated protein and decreased in the cleavage activation of caspase-3. Additionally, to investigate whether chronic luteolin treatment has an antidepression effect, we performed some behavioral tests. Chronic luteolin treatment showed antidepressant-like effects in behavioral tests and, luteolin attenuated the expression of endoplasmic reticulum stress-related proteins in the hippocampus of corticosterone-treated depression model mice. These findings indicate that luteolin has antidepressant-like effects, partly due to the suppression of endoplasmic reticulum stress.

Diacylglycerol kinase ß knockout mice exhibit lithium-sensitive behavioral abnormalities.

BACKGROUND: Diacylglycerol kinase (DGK) is an enzyme that phosphorylates diacylglycerol (DG) to produce phosphatidic acid (PA). DGKß is widely distributed in the central nervous system, such as the olfactory bulb, cerebral cortex, striatum, and hippocampus. Recent studies reported that the splice variant at the COOH-terminal of DGKß was related to bipolar disorder, but its detailed mechanism is still unknown. METHODOLOGY/PRINCIPAL FINDINGS: In the present study, we performed behavioral tests using DGKß knockout (KO) mice to investigate the effects of DGKß deficits on psychomotor behavior. DGKß KO mice exhibited some behavioral abnormalities, such as hyperactivity, reduced anxiety, and reduced depression. Additionally, hyperactivity and reduced anxiety were attenuated by the administration of the mood stabilizer, lithium, but not haloperidol, diazepam, or imipramine. Moreover, DGKß KO mice showed impairment in Akt-glycogen synthesis kinase (GSK) 3ß signaling and cortical spine formation. CONCLUSIONS/SIGNIFICANCE: These findings suggest that DGKß KO mice exhibit lithium-sensitive behavioral abnormalities that are, at least in part, due to the impairment of Akt-GSK3ß signaling and cortical spine formation.