Taku Nagai

The present study examined the kinetics of glutathione (GSH) concentration during maturation and after fertilization in pig oocytes and its relevance to the ability of pig oocytes to form a male pronucleus after in vitro fertilization. The GSH concentration was significantly higher in pig oocytes matured in Waymouth medium than in pig oocytes matured in either modified (m) TCM-199 or mTLP media. The addition of 0.04-0.57 mM cysteine (CySH) to mTLP significantly increased both the GSH concentrations in oocytes matured in vitro and the rate of male pronucleus formation as compared to those in oocytes cultured in mTLP alone. When pig oocytes were cultured 12, 24, or 36 h in mTLP plus 0.14 mM CySH, their GSH concentrations were significantly higher than in uncultured oocytes. After fertilization, the GSH concentration in pig oocytes declined significantly. GSH concentrations in oocytes matured in vivo did not differ from those in oocytes matured in mTLP plus 0.14 or 0.57 mM CySH. The results indicate that 1) the composition of maturation medium affects the GSH concentration in pig oocytes; 2) the addition of CySH to maturation medium permits GSH synthesis by the pig oocytes; 3) GSH levels in pig oocytes change during maturation and after fertilization; and 4) GSH synthesis during oocyte maturation is an important factor for promoting their ability to form a male pronucleus after fertilization.

A new gene, termed klotho, is associated with the suppression of several aging phenotypes. Because high expression of klotho gene was detected in the brain, it would be plausible that klotho gene is involved in the regulation of brain aging. We investigated the changes in mnemonic function accompanying aging in klotho mutant mice. Cognitive function measured by novel-object recognition and conditioned-fear tests in klotho mutant mice was normal at the age of 6 wk, but markedly impaired at the age of 7 wk. Lipid (malondialdehyde) and DNA (8-hydroxy-2'-deoxyguanosine) peroxide levels in the hippocampus of klotho mutant mice increased at the age of 5 wk, 2 wk before the development of cognition deficits. Pro-death Bax increased, whereas anti-death Bcl-2 and Bcl-XL decreased, and apoptotic TUNEL-positive cells were detected in the hippocampus of klotho mutant mice at the age of 7 wk. A potent antioxidant, a-tocopherol, prevented cognition impairment and lipid peroxide accumulation and decreased the number of apoptotic cells in klotho mutant mice. These results suggest that oxidative stress has a crucial role in the aging-associated cognition impairment in klotho mutant mice. Klotho protein may be involved in the regulation of antioxidative defense.

Intracellular amyloid-beta peptide (Abeta) has been implicated in neuronal death associated with Alzheimer's disease. Although Abeta is predominantly secreted into the extracellular space, mechanisms of Abeta transport at the level of the neuronal cell membrane remain to be fully elucidated. We demonstrate that receptor for advanced glycation end products (RAGE) contributes to transport of Abeta from the cell surface to the intracellular space. Mouse cortical neurons exposed to extracellular human Abeta subsequently showed detectable peptide intracellularly in the cytosol and mitochondria by confocal microscope and immunogold electron microscopy. Pretreatment of cultured neurons from wild-type mice with neutralizing antibody to RAGE, and neurons from RAGE knockout mice displayed decreased uptake of Abeta and protection from Abeta-mediated mitochondrial dysfunction. Abeta activated p38 MAPK, but not SAPK/JNK, and then stimulated intracellular uptake of Abeta-RAGE complex. Similar intraneuronal co-localization of Abeta and RAGE was observed in the hippocampus of transgenic mice overexpressing mutant amyloid precursor protein. These findings indicate that RAGE contributes to mechanisms involved in the translocation of Abeta from the extracellular to the intracellular space, thereby enhancing Abeta cytotoxicity.

Experiences during brain development may influence the pathogenesis of developmental disorders. Thus, social isolation (SI) rearing after weaning is a useful animal model for studying the pathological mechanisms of such psychiatric diseases. In this study, we examined the effect of SI on neurogenesis in the hippocampal dentate gyrus (DG) relating to memory and emotion-related behaviors. When newly divided cells were labeled with 5-bromo-2'-deoxyuridine (BrdU) before SI, the number of BrdU-positive cells and the rate of differentiation into neurons were significantly decreased after 4-week SI compared with those in group-housed mice. Repeated treatment of fluoxetine prevented the SI-induced impairment of survival of newly divided cells and ameliorated spatial memory impairment and part of aggression in SI mice. Furthermore, we investigated the changes in gene expression in the DG of SI mice by using DNA microarray and real-time PCR. We finally found that SI reduced the expression of development-related genes Nurr1 and Npas4. These findings suggest that communication in juvenile is important in the survival and differentiation of newly divided cells, which may be associated with memory and aggression, and raise the possibility that the reduced expression of Nurr1 and/or Npas4 may contribute to the impairment of neurogenesis and memory and aggression induced by SI.