Atsushi Oyagi

Induction of BiP, an ER-resident protein, prevents the neuronal death induced by transient forebrain ischemia in gerbil.

Endoplasmic reticulum (ER) stress, which is caused by the accumulation of unfolded proteins in the ER lumen, is associated with stroke and neurodegenerative diseases such as Parkinsons and Alzheimers diseases. We evaluated the effect of a selective inducer of immunoglobulin heavy chain binding protein (BiP) (BiP inducer X; BIX) against both tunicamycin-induced cell death (in SH-SY5Y cells) and the effects of global transient forebrain ischemia (in gerbils). BIX significantly induced BiP expression both in vitro and in vivo. Pretreatment with BIX at 2 or 5 microM reduced the cell death induced by tunicamycin in SH-SY5Y cells. In gerbils subjected to forebrain ischemia, prior treatment with BIX (intracerebroventricular injection at 10 or 40 microg) protected against cell death and decreased TUNEL-positive cells in the hippocampal CA1 subfield. These findings indicate that this selective inducer of BiP could be used to prevent the neuronal damage both in vitro and in vivo.

Sirtuin 1 overexpression mice show a reference memory deficit, but not neuroprotection.

Sirtuin 1 (SIRT1) is the closest mammalian homologue of yeast silent information regulator 2 (Sir2) and has a role in lifespan modulation. Reportedly, SIRT1 is also linked to neurodegenerative diseases. However, there are limited studies that report the relation between SIRT1 and neurodegenerative diseases using in vivo transgenic (Tg) methods. In the present study, we generated neuron-specific enolase (NSE) SIRT1 Tg mice that overexpress human SIRT1 in neurons. We examined possible neuroprotective effects of SIRT1 overexpression and compared their higher brain functions with those of wild-type (WT) mice. Overexpression of SIRT1 did not have any neuroprotective effects against the neuronal damage induced by ischemia or 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). However, SIRT1 Tg mice exhibited a reference memory deficit. These findings suggest that an excessive expression of SIRT1 might induce the memory deficit in mice, but not neuroprotective effects.

Essential Role of Neuron-Enriched Diacylglycerol Kinase (DGK), DGKβ in Neurite Spine Formation, Contributing to Cognitive Function

Background Diacylglycerol (DG) kinase (DGK) phosphorylates DG to produce phosphatidic acid (PA). Of the 10 subtypes of mammalian DGKs, DGKβ is a membrane-localized subtype and abundantly expressed in the cerebral cortex, hippocampus, and caudate-putamen. However, its physiological roles in neurons and higher brain function have not been elucidated. Methodology/Principal Findings We, therefore, developed DGKβ KO mice using the Sleeping Beauty transposon system, and found that its long-term potentiation in the hippocampal CA1 region was reduced, causing impairment of cognitive functions including spatial and long-term memories in Y-maze and Morris water-maze tests. The primary cultured hippocampal neurons from KO mice had less branches and spines compared to the wild type. This morphological impairment was rescued by overexpression of DGKβ. In addition, overexpression of DGKβ in SH-SY5Y cells or primary cultured mouse hippocampal neurons resulted in branch- and spine-formation, while a splice variant form of DGKβ, which has kinase activity but loses membrane localization, did not induce branches and spines. In the cells overexpressing DGKβ but not the splice variant form, DGK product, PA, was increased and the substrate, DG, was decreased on the plasma membrane. Importantly, lower spine density and abnormality of PA and DG contents in the CA1 region of the KO mice were confirmed. Conclusions/Significance These results demonstrate that membrane-localized DGKβ regulates spine formation by regulation of lipids, contributing to the maintenance of neural networks in synaptic transmission of cognitive processes including memory.

Diacylglycerol Kinase β Knockout Mice Exhibit Lithium-Sensitive Behavioral Abnormalities

Background Diacylglycerol kinase (DGK) is an enzyme that phosphorylates diacylglycerol (DG) to produce phosphatidic acid (PA). DGKβ 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 DGKβ was related to bipolar disorder, but its detailed mechanism is still unknown. Methodology/Principal Findings In the present study, we performed behavioral tests using DGKβ knockout (KO) mice to investigate the effects of DGKβ deficits on psychomotor behavior. DGKβ 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, DGKβ KO mice showed impairment in Akt-glycogen synthesis kinase (GSK) 3β signaling and cortical spine formation. Conclusions/Significance These findings suggest that DGKβ KO mice exhibit lithium-sensitive behavioral abnormalities that are, at least in part, due to the impairment of Akt-GSK3β signaling and cortical spine formation.

Protective effects of a gastrointestinal agent containing Korean red ginseng on gastric ulcer models in mice

BackgroundKorean red ginseng (KRG) is a ginseng that has been cultivated and aged for 4-6 years or more, and goes through an extensive cleaning, steaming and drying process. KRG contains more than 30 kinds of saponin components and has been reported as having various biological properties, such as anti-fatigue action, immune restoration, and neurovegetative effect. The purpose of this study was to assess the effects of a KRG-containing drug (KRGCD) on gastric ulcer models in mice.MethodsStomach ulcers were induced by oral ingestion of hydrochloride (HCl)/ethanol or indomethacin. Treatment with KRGCD (30, 100, and 300 mg/kg, p.o.) occurred 1 hr before the ulcer induction. Effect of KRGCD on anti-oxidant activity and gastric mucosal blood flow with a laser Doppler flowmeter in mice stomach tissue was evaluated.ResultsKRGCD (100 and 300 mg/kg, p.o.) significantly decreased ethanol- and indomethacin-induced gastric ulcer compared with the vehicle-treated (control) group. KRGCD (100 and 300 mg/kg) also decreased the level of thiobarbituric acid reactive substance (TBARS) and increased gastric mucosal blood flow compared with the control group.ConclusionsThese results suggest that the gastroprotective effects of KRGCD on mice ulcer models can be attributed to its ameliorating effect on oxidative damage and improving effect of gastric mucosal blood flow.

Generation and Characterization of Conditional Heparin-Binding EGF-Like Growth Factor Knockout Mice

Recently, neurotrophic factors and cytokines have been shown to be associated in psychiatric disorders, such as schizophrenia, bipolar disorder, and depression. Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a member of the EGF family, serves as a neurotrophic molecular and plays a significant role in the brain. We generated mice in which HB-EGF activity is disrupted specifically in the ventral forebrain. These knockout mice showed (a) behavioral abnormalities similar to those described in psychiatric disorders, which were ameliorated by typical or atypical antipsychotics, (b) altered dopamine and serotonin levels in the brain, (c) decreases in spine density in neurons of the prefrontal cortex, (d) reductions in the protein levels of the NR1 subunit of the N-methyl-D-aspartate (NMDA) receptor and post-synaptic protein-95 (PSD-95), (e) decreases in the EGF receptor, and in the calcium/calmodulin-dependent protein kinase II (CaMK II) signal cascade. These results suggest the alterations affecting HB-EGF signaling could comprise a contributing factor in psychiatric disorder.

Diacylglycerol kinase β 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. DGKβ is one of the subtypes of the DGK family and regulates many intracellular signaling pathways in the central nervous system. Previously, we demonstrated that DGKβ 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 DGKβ KO mice in order to investigate the function of DGKβ in the central nervous system, especially in the pathophysiology of attention deficit hyperactivity disorder (ADHD). Methodology/Principal Findings DGKβ 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, DGKβ 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 DGKβ KO mice. Conclusions/Significance These findings suggest that DGKβ KO mice showed attention-deficit and hyperactive phenotype, similar to ADHD. Furthermore, the hyporesponsiveness of DGKβ KO mice to MPH was due to dysregulation of ERK phosphorylation, and that DGKβ has a pivotal involvement in ERK regulation in the striatum.

The protective effect and action mechanism of Vaccinium myrtillus L. on gastric ulcer in mice.

Vaccinium myrtillus L. anthocyanoside (VMA) is used as a folk medicine to treat diseases related to gastric ulcers in northern Europe. However, the effects of VMA and its detailed mechanism on gastric ulcer have not been investigated sufficiently. Therefore, the aim of the present study was to investigate the protective effects of VMA on gastric mucosal damage in a murine gastric ulcer model. First the effects of VMA on ethanol‐induced gastric ulcers in mice were investigated. Then, the levels of lipid peroxide in murine stomach homogenates were measured to investigate the antioxidative effects of VMA. In addition, the free radical scavenging activity of VMA and its main anthocyanidins were evaluated by electron spin resonance measurement. Oral administration of VMA (10, 30 and 100 mg/kg) significantly protected gastric mucosa against HCl/ethanol‐induced gastric ulcers. Furthermore, VMA inhibited lipid peroxide levels in a concentration‐dependent manner and showed high scavenging activity against the superoxide anion radical (·O2‐) and the hydroxyl radical (·OH). Anthocyanidins also showed scavenging activity against the ·O2‐, while only delphinidin showed high scavenging activity against the ·OH. These findings indicate that the protective effects of VMA on HCl/ethanol‐induced gastric mucosal injury may be partially due to the antiperoxidative effects of anthocyanidins. Copyright

Heparin-binding EGF-like growth factor is required for synaptic plasticity and memory formation

Heparin-binding epidermal growth factor-like growth factor (HB-EGF), a member of epidermal growth factor (EGF) family, is a potent mitogenic peptide for various types of cells. HB-EGF is widely expressed in central nervous system, including hippocampus and cerebral cortex, and is considered to play pivotal roles in the developing and adult nervous system. In this study, we assessed the role of HB-EGF in learning and memory by testing HB-EGF conditional knock-out mice (KO) in two different learning tasks, and evaluated the long-term potentiation (LTP) in hippocampus slices from these mice. The HB-EGF KO mice were impaired in spatial memory in the Morris water maze and in fear learning in a passive avoidance test. HB-EGF KO mice also showed an impaired LTP, and reduction in activity of Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) and phosphorylated GluR1. We also found that the levels of neurotrophic factors, such as nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), or glial cell line-derived neurotrophic factor (GDNF), were altered in several brain regions in the HB-EGF KO mice. These results confirm the importance of the HB-EGF in synaptic plasticity and memory formation.

Protective effects of SUN N8075, a novel agent with antioxidant properties, in in vitro and in vivo models of Parkinson's disease

SUN N8075 is a novel antioxidant with neuroprotective properties. This study was designed to elucidate its neuroprotective effects against 6-hydroxy dopamine (6-OHDA)-induced cell death and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced neurotoxicity (known as in vitro and in vivo models of Parkinsons disease, respectively). In the in vitro study, on human neuroblastoma SH-SY5Y cells, SUN N8075 decreased the hydrogen peroxide (H2O2)-induced production of reactive oxygen species and protected against 6-OHDA-induced cell death. In the in vivo study, SUN N8075, when injected intraperitoneally (i.p.) twice with a 5-h interval, inhibited lipid peroxidation (viz. the production of thiobarbituric acid reactive substance) in the mouse forebrain at 1 h after the second injection. Mice were injected i.p. with MPTP (10 mg/kg) four times at 1-h intervals, and brains were analyzed 7 days later. SUN N8075 at 30 mg/kg (i.p., twice) exhibited a protective effect against the MPTP-induced decrease in tyrosine hydroxylase (TH)-positive fibers in the striatum. Moreover, SUN N8075 at 10 and 30 mg/kg (i.p., twice) had a similar protective effect against the MPTP-induced decrease in TH-positive cells in the substantia nigra. Further, SUN N8075 30 mg/kg (i.p. twice) markedly suppressed the MPTP-induced accumulation of 8-hydroxy-deoxyguanosine (8-OHdG) in the striatum. These findings indicate that SUN N8075 exerts protective effects, at least in part via an anti-oxidation mechanism, in these in vitro and in vivo models of Parkinsons disease.