Mitsue Ishisaka

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.

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.

Modulation of endoplasmic reticulum stress in Parkinson's disease.

Parkinsons 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 Parkinsons disease. Furthermore, it has been suggested that some existing anti- Parkinsons 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 Parkinsons disease models and investigating the mechanisms against ER stress will contribute to the search for new therapeutic agents for Parkinsons disease.

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

Alzheimers 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 β1-42-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.

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. Repeated treatment with Huperzia serrata extract (30 mg/kg/day) for six or seven days ameliorated the memory impairment induced by scopolamine in the two behavioral tests.

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.

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.

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.

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.

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.