Behavioral stress causes mitochondrial dysfunction via ABAD up-regulation and aggravates plaque pathology in the brain of a mouse model of Alzheimer disease

Basic and clinical studies have reported that behavioral stress worsens the pathology of Alzheimer disease (AD), but the underlying mechanism has not been clearly understood. In this study, we determined the mechanism by which behavioral stress affects the pathogenesis of AD using Tg-APPswe/PS1dE9 mice, a murine model of AD. Tg-APPswe/PS1dE9 mice that were restrained for 2h daily for 16 consecutive days (2-h/16-day stress) from 6.5months of age had significantly increased Aβ(1-42) levels and plaque deposition in the brain. The 2-h/16-day stress increased oxidative stress and induced mitochondrial dysfunction in the brain. Treatment with glucocorticoid (corticosterone) and Aβ in SH-SY5Y cells increased the expression of 17β-hydroxysteroid dehydrogenase (ABAD), mitochondrial dysfunction, and levels of ROS, whereas blockade of ABAD expression by siRNA-ABAD in SH-SY5Y cells suppressed glucocorticoid-enhanced mitochondrial dysfunction and ROS accumulation. The 2-h/16-day stress up-regulated ABAD expression in mitochondria in the brain of Tg-APPswe/PS1dE9 mice. Moreover, all visible Aβ plaques were costained with anti-ABAD in the brains of Tg-APPswe/PS1dE9 mice. Together, these results suggest that behavioral stress aggravates plaque pathology and mitochondrial dysfunction via up-regulation of ABAD in the brain of a mouse model of AD.     

Metabolism of subtoxic level of selenite by double-perfused small intestine in rats

Intestinal metabolism of the subtoxic level of selenite in rats was investigated using a double-perfusion system, which is an in situ, in vitro preparation in which the intestinal lumen and its vasculature are perfused simultaneously. The toxicity of sodium selenite was determined by inhibition of 3-O-methyl glucose (3MG) absorption and by histological examination. Levels of 1.2 mM selenite were required to significantly (p<0.05) reduce 3MG intestinal absorption (58±11%, mean±SD). Cation-exchange chromatography was used to determine the chemical forms of Se from selenite after using luminal concentrations of 1–200 μM in vascular perfusates. The chemical forms were selenite, selenodiglutathione (GS-Se-SG), mixed selenoglutathione plus cysteine (GS-Se-CYS), selenodicysteine (CYS-Se-CYS), protein-bound Se, and unidentified selenocompounds. Selenite was the predominant selenocompound found in vascular perfusate, but protein-bound Se was the predominant metabolite from selenite present in the vascular effuents. There was a corresponding increase of all metabolites with increased levels of selenite with time of absorption, but not with increased concentration of luminal selenite.     

Serotoninergic modulation of GABAergic synaptic transmission in developing rat CA3 pyramidal neurons

Serotoninergic modulation of GABAergic mIPSCs was investigated in immature (postnatal 12–16-days old) rat CA3 pyramidal neurons using a conventional whole-cell patch clamp technique. Serotonin or 5-hydroxytryptamine (5-HT) (10 μmol/L) transiently and explosively increased mIPSC frequency with a small increase in the current amplitude. However, 5-HT did not affect the GABA-induced postsynaptic currents, indicating that 5-HT acts presynaptically to facilitate the probability of spontaneous GABA release. The 5-HT action on GABAergic mIPSC frequency was completely blocked by 100 nmol/L MDL72222, a selective 5-HT₃ receptor antagonist, and mimicked by mCPBG, a selective 5-HT₃ receptor agonist. The 5-HT action on GABAergic mIPSC frequency was completely occluded either in the presence of 200 μmol/L Cd²⁺ or in the Na⁺-free external solution, suggesting that the 5-HT₃ receptor-mediated facilitation of mIPSC frequency requires a Ca²⁺influx passing through voltage-dependent Ca²⁺channels from the extracellular space, and that presynaptic 5-HT₃ receptors are less permeable to Ca²⁺. The 5-HT action on mIPSC frequency in the absence or presence of extracellular Na⁺ gradually increased with postnatal development. Such a developmental change in the 5-HT₃ receptor-mediated facilitation of GABAergic transmission would play important roles in the regulation of excitability as well as development in CA3 pyramidal neurons.     

GAzer: gene set analyzer

Gene Set Analyzer (GAzer) is a web-based integrated gene set analysis tool covering previously reported parametric and non-parametric models. Based on a simulation test for the reported algorithms, we classified and implemented three main statistical methods consisting of the z-statistic, gene permutation and sample permutation for ten gene set categories including Gene Ontology (GO) for human, mouse, rat and yeast. This tool identifies significantly altered gene sets scored by z-statistics and P-values from the z-test or permutation test and provides q-values and Bonferroni P-values to correct multiple hypothesis testing. GAzer allows users to observe changes in expression of each gene in a gene set or to see the significance of the gene sets containing a gene(s) of interest, thus allowing interactive data analysis both at the gene and gene set level. Moreover, GAzer offers extensive annotation for each gene. AVAILABILITY: The GAzer gene set analyzer is freely available at http://integromics.kobic.re.kr/GAzer/. SUPPLEMENTARY INFORMATION: This can be found on the web page (http://integromics.kobic.re.kr/GAzer/supplement.jsp).     

Cardiomyocyte specific deficiency of serine palmitoyltransferase subunit 2 reduces ceramide but leads to cardiac dysfunction

The role of serine palmitoyltransferase (SPT) and de novo ceramide biosynthesis in cardiac ceramide and sphingomyelin metabolism is unclear. To determine whether the de novo synthetic pathways, rather than ceramide uptake from circulating lipoproteins, is important for heart ceramide levels, we created cardiomyocyte-specific deficiency of Sptlc2, a subunit of SPT. Heart-specific Sptlc2-deficient (hSptlc2 KO) mice had a >35% reduction in ceramide, which was limited to C18:0 and very long chain ceramides. Sphingomyelinase expression, and levels of sphingomyelin and diacylglycerol were unchanged. But surprisingly phospholipids and acyl CoAs contained increased saturated long chain fatty acids. hSptlc2 KO mice had decreased fractional shortening and thinning of the cardiac wall. While the genes regulating glucose and fatty acid metabolism were not changed, expression of cardiac failure markers and the genes involved in the formation of extracellular matrices were up-regulated in hSptlc2 KO hearts. In addition, ER-stress markers were up-regulated leading to increased apoptosis. These results suggest that Sptlc2-mediated de novo ceramide synthesis is an essential source of C18:0 and very long chain, but not of shorter chain, ceramides in the heart. Changes in heart lipids other than ceramide levels lead to cardiac toxicity.     

Zinc modulation of glycine receptors in acutely isolated rat CA3 neurons

Glycine and GABA are the primary inhibitory neurotransmitters in the spinal cord and brain stem, with glycine exerting its physiological roles by activating strychnine-sensitive ionotropic receptors. Glycine receptors are also expressed in the brain, including the cortex and hippocampus, but their physiological roles and pharmacological properties are largely unknown. Here, we report the pharmacological properties of functional glycine receptors in acutely isolated rat CA3 neurons using conventional whole-cell patch clamp techniques. Both glycine and taurine, which are endogenous agonists of glycine receptors, elicited Cl(-) currents in a concentration-dependent manner. The glycine-induced current (I(Gly)) was inhibited by strychnine, picrotoxin or cyclothiazide in a concentration-dependent manner. At lower concentrations (0.01-1 microM), ICS-205,930 potentiated I(Gly), but at higher concentrations (>10 microM) it inhibited I(Gly). These pharmacological properties strongly suggest that CA3 neurons express functional strychnine-sensitive glycine receptors containing alpha2 subunits. Furthermore, at lower concentrations (1-30 microM), Zn(2+) potentiated I(Gly), but at higher concentrations (>100 microM) it inhibited I(Gly). Considering that Zn(2+) is synaptically co-released with glutamate from mossy fiber terminals that make excitatory synapses onto CA3 neurons, these results suggest that endogenous Zn(2+) modulation of these glycine receptors may have an important role in the excitability of CA3 neurons.     

Analysis of somaclonal variation through tissue culture and chromosomal localization of rDNA sites by fluorescent in situ hybridization in wild Allium tuberosum and a regenerated variant

The effects of basal media and growth regulators on callus initiation and shoot regeneration have been investigated in wild Allium tuberosum (2n = 4x = 32). Callus initiation was greatest from flower bud explants cultured on MS medium supplemented with 2,4-D and BA at 1 mg l⁻¹ each. Maximum number of shoots was obtained from callus grown on MS medium supplemented with NAA and BA at 0.2 and 2 mg l⁻¹, respectively. The chromosome analysis of regenerants derived from callus revealed variation in ploidy, such as 2n = 28, 29, 30, 31, 33 as well as normal tetraploid. During the culture period for two generations, one aneuploid regenerant with 2n = 30 (named At30) showed better viability and growth than tetraploid plants and other aneuploid variants. In a karyotypic analysis of At30, the chromosomal positions of 5S and 18S-5.8S-26S rDNA were physically mapped by fluorescent in situ hybridization and compared to chromosomes of wild type A. tuberosum. Both wild type A. tuberosum and At30 exhibited two sets of 5S rDNA sites, one on the proximal position of the short arm of chromosome 3, and the other on the intercalary region on the long arm of chromosome 6. There was one 18S-5.8S-26S rDNA site in the secondary constriction including flanking short chromosomal segments of satellite and terminal regions on the short arm of chromosome 8 in wild type A. tuberosum. However, At30 showed only three labelled chromosome 8 indicating that this was one of the lost chromosomes of At30. This revised version was published online in June 2006 with corrections to the Cover Date.     

Comparative study on high fat diet-induced 4-hydroxy-2E-nonenal adducts in the hippocampal CA1 region of C57BL/6N and C3H/HeN mice

In the present study, we investigated the influences of a high fat diet (HD) fed for 12 weeks, on lipid peroxidation and antioxidant enzyme using 4-hydroxy-2E-nonenal (HNE)-modified proteins (HNE-mp) and Cu,Zn-superoxide dismutase (SOD1) in the hippocampal CA1 region (CA1) in C57BL/6N and C3H/HeN mice. Body weights and body weight gains were significantly higher in HD fed C57BL/6N mice than in low fat diet (LD) fed C57BL/6N and LD or HD fed C3H/HeN mice. In the HD fed C57BL/6N and C3H/HeN mice, HNE-mp immunoreactivity and protein levels were much higher than in the LD fed C57BL/6N or C3H/HeN mice. In particular, HNE-mp immunoreactivity and protein levels in HD fed C57BL/6N mice was higher than that in the HD fed C3H/HeN mice. SOD1 immunoreaction was detected in the non-pyramidal cells of C57BL/6N mice, while in the C3H/HeN mice SOD1 immunoreaction was observed in CA1 pyramidal cells. The SOD1 immunoreactivity in the LD fed C57BL/6N and C3H/HeN mice was slightly, but not significantly decreased compared to that in the HD fed C57BL/6N and C3H/HeN mice, respectively. In addition, ionized calcium-binding adapter molecule 1 (Iba-1) immunoreactive microglia in the HD fed C57BL/6N showed hypertrophy of cytoplasm, which is the characteristics of activated microglia. These results suggest that HD fed C57BL/6N mice are more susceptible to lipid peroxidation in the CA1 than in LD fed C57BL/6N and LD or HD fed C3H/HeN mice without any differences of SOD1 expression. In Koo Hwang and Il Yong Kim have contributed equally to this article.     

Anti-Oxidative Effects of Rooibos Tea (Aspalathus linearis) on Immobilization-Induced Oxidative Stress in Rat Brain

Exposure to chronic psychological stress may be related to increased reactive oxygen species (ROS) or free radicals, and thus, long-term exposure to high levels of oxidative stress may cause the accumulation of oxidative damage and eventually lead to many neurodegenerative diseases. Compared with other organs, the brain appears especially susceptible to excessive oxidative stress due to its high demand for oxygen. In the case of excessive ROS production, endogenous defense mechanisms against ROS may not be sufficient to suppress ROS-associated oxidative damage. Dietary antioxidants have been shown to protect neurons against a variety of experimental neurodegenerative conditions. In particular, Rooibos tea might be a good source of antioxidants due to its larger proportion of polyphenolic compounds. An optimal animal model for stress should show the features of a stress response and should be able to mimic natural stress progression. However, most animal models of stress, such as cold-restraint, electric foot shock, and burn shock, usually involve physical abuse in addition to the psychological aspects of stress. Animals subjected to chronic restraint or immobilization are widely believed to be a convenient and reliable model to mimic psychological stress. Therefore, in the present study, we propose that immobilization-induced oxidative stress was significantly attenuated by treatment with Rooibos tea. This conclusion is demonstrated by Rooibos tea’s ability to (i) reverse the increase in stress-related metabolites (5-HIAA and FFA), (ii) prevent lipid peroxidation (LPO), (iii) restore stress-induced protein degradation (PD), (iv) regulate glutathione metabolism (GSH and GSH/GSSG ratio), and (v) modulate changes in the activities of antioxidant enzymes (SOD and CAT).