Alteration of the sialylation pattern and memory deficits by injection of Aβ(25–35) into the hippocampus of rats

Sialic acid in glycoconjugates participates in important cellular functions associated with normal development, growth, and communication. Therefore we evaluated the sialylation pattern and memory deficits caused by the injection of Aβ_(25–35) into the hippocampus (Hp) of rats. The eight-arm maze spatial-learning and memory test indicated that the injection of Aβ_(25–35) into subfield CA1 of the Hp impaired both learning and memory. The sialylation pattern was examined using sialic acid-specific lectins. Our results showed that Maackia amurensis agglutinin (MAA, specific for Neu5Acα2,3Gal) showed reactivity in the CA1 and dentate gyrus (DG) subfields of the Hp mainly in the group injected with vehicle, whereas Macrobrachium rosenbergii lectin (MRL, specific for Neu5,9,7Ac) and Sambucus nigra agglutinin (SNA, specific for Neu5Acα2,6Gal-GalNAc) had increased reactivity in the CA1 and DG subfields of the Hp in the Aβ_(25–35)-injected group. The staining pattern of the antibody specific for polysialic acid (a linear homopolymer of α-2,8-linked sialic acid) increased in the CA1 and DG subfields of the Hp of the Aβ_(25–35) group compared to the control group. Our results suggest that injection of Aβ_(25–35) causes impairment in spatial memory and alters the sialylation pattern in response to compensatory reorganization and-or sprouting of dendrites and axons of the surviving neurons.     

Neuroprotective effect of alpha-asarone on spatial memory and nitric oxide levels in rats injected with amyloid-β(25–35)

The chemical α-asarone is an important active substance of the Acori graminei rhizome (AGR). It has pharmacological effects that include antihyperlipidemic, antiinflammatory, and antioxidant activity. Our aim was to study the effects α-asarone on nitric oxide (NO) levels in the hippocampus and temporal cortex of the rat after injection of the fraction 25–35 from amyloid-β (Aβ_(25–35)). In addition we examined the working spatial memory in an eight-arm radial maze. Our results showed a significant increase of nitrites in the hippocampus and temporal cortex of Aβ_(25–35)-treated rats. Other evidence of neuronal damage was the expression of a glial-fibrillar-acid protein and a silver staining. There were impairments in the spatial memory evaluated in the eight-arm radial maze. We wanted to determine whether α-asarone improves the memory correlated with NO overproduction and neuronal damage caused by the injection of Aβ_(25–35) into rats. Then animals received a 16-day treatment of α-asarone before the Aβ_(25–35) injection. Our results show a significant decrease of nitrite levels in the hippocampus and temporal cortex, without astrocytosis and silver-staining cells, which correlates with memory improvement in the α-asarone-treated group. Our results suggest that α-asarone may protect neurons against Aβ_(25–35)–caused neurotoxicity by inhibiting the effects of NO overproduction in the hippocampus and temporal cortex.     

Effects of memantine on soluble Αβ25-35-induced changes in peptidergic and glial cells in Alzheimer's disease model rat brain regions

Soluble forms of amyloid-β (Aβ) have been considered responsible for cognitive dysfunction prior to senile plaque formation in Alzheimer's disease (AD). As its mechanism is not well understood, we examined the effects of repeated i.c.v. infusion of soluble Αβ_25-35 on peptidergic system and glial cells in the pathogenesis of AD. The present study aims to investigate the protective effects of memantine on Aβ_25-35-induced changes in peptidergic and glial systems. Infusion of Αβ_25-35 decreased the level of immunoreactive somatostatin (SS) and substance P (SP) in the hippocampus prior to neuronal loss or caspase activation, which is correlated with the loss of spine density and activation of inducible nitric-oxide synthase (iNOS). Biochemical experiment with peptide-degrading enzymes, prolyl oligopeptidase (POP) and endopeptidase 24.15 (EP 24.15) activities demonstrated a concomitant increase with the activation of glial marker proteins, glial fibrillary acidic protein (GFAP) and CD11b in the Aβ-treated hippocampus. Double immunostaining experiments of EP 24.15 and GFAP/CD11b antibodies clearly demonstrated the co-localization of neuro peptidases with astrocytes and microglia. Treatment with memantine, a non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist significantly attenuated Aβ_25-35-induced changes of neuropeptides, their metabolizing enzymes, glial marker proteins, and activation of iNOS. Taken together, the data implies that memantine exerts its protective effects by modulating the neuropeptide system as a consequence of suppressing the glial cells and oxidative stress in AD model rat brain regions.     

Amyloid-β25–35 induces impairment of cognitive function and long-term potentiation through phosphorylation of collapsin response mediator protein 2

Alzheimer's disease (AD) is characterized by amyloid-β (Aβ) protein and tau deposition in the brain. Numerous studies have reported a central role of Aβ in the development of AD, but the pathogenesis is not well understood. Collapsin response mediator protein 2 (CRMP2), an intracellular protein mediating a repulsive axon guidance molecule, Semaphorin3A, is also accumulated in neurofibrillary tangles in AD brains. To gain insight into the role of CRMP2 phosphorylation in AD pathogenesis, we investigated the effects of Aβ neurotoxicity in CRMP2 phosphorylation-deficient knock-in (crmp2^ki/ki) mice, in which the serine residue at 522 was replaced with alanine. Intracerebroventricular (i.c.v.) injection of Aβ_25–35 peptide, a neurotoxic fragment of Aβ protein, to wild-type (wt) mice increased hippocampal phosphorylation of CRMP2. Behavioral assessment revealed that i.c.v. injection of Aβ_25–35 peptide caused impairment of novel object recognition in wt mice, while the same peptide did not in crmp2^ki/ki mice. In electrophysiological recording, wt and crmp2^ki/ki mice have similar input–output basal synaptic transmission and paired-pulse ratios. However, long-term potentiation was impaired in hippocampal slices of Aβ_25–35 peptide-treated wt but not those of crmp2^ki/ki. Our findings indicate that CRMP2 phosphorylation is required for Aβ-induced impairment of cognitive memory and synaptic plasticity.     

Involvement of cysteinyl leukotriene receptor 1 in Aβ1–42-induced neurotoxicity in vitro and in vivo

Accumulation of amyloid-β (Aβ) is thought to be associated with the progressive neuronal death observed in Alzheimer's disease, but the mechanisms underlying neurotoxicity triggered by Aβ remain elusive. In the current study, we investigated the roles of cysteinyl leukotriene receptor 1 (CysLT₁R) in Aβ_1–42-induced neurotoxicity in vitro or in vivo. In vitro exposure of mouse primary neurons to Aβ_1–42 caused a gradual increases in CysLT₁R expression. In vivo bilateral intrahippocampal injection of Aβ_1–42 also elicited time-dependent increases of CysLT₁R expression in the hippocampus and cortex of mice. The CysLT₁R antagonist pranlukast not only reversed Aβ_1–42-induced upregulation of CysLT₁R, but also suppressed Aβ_1–42-triggered neurotoxicity evidenced by enhanced nuclear factor-kappa B p65, activated caspase-3, decreased B-cell lymphoma-2 and cell viability and impaired memory. Furthermore, chronic treatment with pranlukast produced similar beneficial effects on memory behavior and hippocampal long-term potentiation to memantine or donepezil in intrahippocampal Aβ_1–42-injected mice. Our data indicate that CysLT₁R is involved in Aβ_1–42-induced neurotoxicity, and that blockade of CysLT₁R, such as application of CysLT₁R antagonist, could be a novel and promising strategy for the treatment of Alzheimer's disease.     

Effects of Hericium erinaceus on amyloid β(25-35) peptide-induced learning and memory deficits in mice

The mushroom Hericium erinaceus has been used as a food and herbal medicine since ancient times in East Asia. It has been reported that H. erinaceus promotes nerve growth factor secretion in vitro and in vivo. Nerve growth factor is involved in maintaining and organizing cholinergic neurons in the central nervous system. These findings suggest that H. erinaceus may be appropriate for the prevention or treatment of dementia. In the present study, we examined the effects of H. erinaceus on amyloid β(25-35) peptide-induced learning and memory deficits in mice. Mice were administered 10 μg of amyloid β(25-35) peptide intracerebroventricularly on days 7 and 14, and fed a diet containing H. erinaceus over a 23-d experimental period. Memory and learning function was examined using behavioral pharmacological methods including the Y-maze test and the novel-object recognition test. The results revealed that H. erinaceus prevented impairments of spatial short-term and visual recognition memory induced by amyloid β(25-35) peptide. This finding indicates that H. erinaceus may be useful in the prevention of cognitive dysfunction.     

The anti-amnesic effects of luteolin against amyloid β25–35 peptide-induced toxicity in mice involve the protection of neurovascular unit

Luteolin (3′,4′,5,7-tetrahydroxyflavone) is an important member of the flavonoid family. It exhibits strongly anti-inflammatory, antioxidant and phytoestrogen-like activities. In the present study, we examined the anti-amnesic and protective effects of luteolin against Aβ_25-35-induced toxicity in mice. Mice were given an i.c.v. injection of aggregated Aβ_25-35 peptide. The learning and memory impairments, ultrastructural changes of cerebral cortex, cerebrovascular dysfunction and neuronal changes were detected after oral administration of luteolin continuously for 8 days. Our results demonstrate that oral administration of luteolin for 8 days for those Aβ_25-35-induced amnesic mice conferred robust neurovascular protection in Aβ_25-35-induced amnesia, involving the improvement of the spatial learning and memory capabilities, the modulation of microvascular function, the increase of regional cerebral blood flow values, the clearance of reactive oxygen species, the improvement of cholinergic neuronal system, and the increase of brain-derived neurotrophic factor level and its receptor tyrosine kinase B expression in cerebral cortex.     

Agmatine protects against β-amyloid25-35-induced memory impairments in the rat

Amyloid beta fragment 25-35 (Aβ_25-35) is the neurotoxic domain of the full-length Aβ_1-42 and causes memory impairments in rodents. Recent research suggests that agmatine, decarboxylated arginine, has a neuroprotective role. This study investigated the effects of a single bilateral i.c.v. infusion of aggregated Aβ_25-35 (30 nmol) in a battery of behavioural tests conducted during the period 4–6 (Experiment 1) and 4–14 (Experiment 2) weeks post-Aβ_25-35 infusion, and evaluated the protective effect of agmatine (40 mg/kg) administered i.p. 30 min prior to Aβ_25-35 infusion and once daily for a further nine consecutive days. In Experiment 1, Aβ_25-35 rats with saline treatment were not impaired in the elevated plus maze and open field and mildly impaired in the reference memory version of the water maze task, but performed poorly in the working memory version of the water maze task and the object recognition memory task, relative to the control rats that received the i.c.v. infusion of Aβ_35-25 (inactive peptide) and saline treatment. By contrast, Aβ_25-35 rats with agmatine treatment did not show performance impairments in the working memory version of the water maze task and the object recognition memory task. In Experiment 2, Aβ_25-35 rats with saline treatment were significantly impaired in the standard radial arm maze task, but only displayed no or very mild impairments in the delayed non-match to position and reference memory versions of the radial arm maze task, T-maze, object recognition memory task, both the reference and working memory versions of the water maze task, elevated plus maze and open field. By contrast, Aβ_25-35 rats with agmatine treatment were not impaired in the standard radial arm maze and performed even better than the controls in the reference memory version of the task. These results demonstrate that agmatine is able to protect against Aβ_25-35-induced memory deficits.     

Role of α2/δ subunit in the development of morphine-induced rewarding effect and behavioral sensitization

Previous data demonstrate that L-type voltage-gated calcium channel (VGCC) blockers, which bind to α₁ subunits of VGCC to suppress Ca²⁺ entry into cells, inhibit the development of psychological dependence on drugs of abuse, suggesting the upregulation of L-type VGCC in the development of psychological dependence. However, there are few available data on changes of the auxiliary subunit α₂/δ modifying L-type VGCC under such conditions. We therefore investigated here the role of α₂/δ subunits of VGCCs in the brain of mouse after repeated treatment with morphine. The treatment with morphine increased α₂/δ subunit expression in the frontal cortex and the limbic forebrain of mice showing rewarding effect and sensitization to hyperlocomotion by morphine. The morphine-induced behavioral sensitization and place preference were also suppressed by gabapentin, which binds to an exofacial epitope of the α₂/δ auxiliary subunits of VGCCs. These findings indicate that the upregulation of α₂/δ subunit as well as α₁ subunits of VGCC in the frontal cortex and the limbic forebrain plays a critical role in development of morphine-induced rewarding effect and behavioral sensitization following neuronal plasticity.     

Association of cerebrospinal fluid Aβ42 with A2M gene in cognitively normal subjects

Low cerebrospinal fluid (CSF) Aβ₄₂ levels correlate with increased brain Aβ deposition in Alzheimer's disease (AD), which suggests a disruption in the degradation and clearance of Aβ from the brain. In addition, APOE ε4 carriers have lower CSF Aβ₄₂ levels than non-carriers. The hypothesis of this investigation was that CSF Aβ₄₂ levels would correlate with regulatory region variation in genes that are biologically associated with degradation or clearance of Aβ from the brain. CSF Aβ₄₂ levels were tested for associations with Aβ degradation and clearance genes and APOE ε4. Twenty-four SNPs located within the 5′ and 3′ regions of 12 genes were analyzed. The study sample consisted of 99 AD patients and 168 cognitively normal control subjects. CSF Aβ₄₂ levels were associated with APOE ε4 status in controls but not in AD patients; A2M regulatory region SNPs were also associated with CSF Aβ₄₂ levels in controls but not in AD patients, even after adjusting for APOE ε4. These results suggest that genetic variation within the A2M gene influences CSF Aβ₄₂ levels.     

Anti-inflammatory effect of α,β-amyrin, a triterpene from Protium heptaphyllum, on cerulein-induced acute pancreatitis in mice

Objective  

To evaluate the anti-inflammatory effect of α,β-amyrin, a pentacyclic triterpenoid from Protium heptaphyllum, on cerulein-induced acute pancreatitis in mice.     

The protective effect of trihexyphenidyl on the beta-amyloid peptide 25–35-induced cytotoxicity in PC12 cells

In the development and progression of Alzheimer’s disease (AD), β-amyloid peptide (Aβ) that induced cytotoxicity containing apoptosis and excess production of reactive oxygen species (ROS) is considered as a causal role. The aim of present study is to investigate the protective effect of Trihexyphenidyl (THY) on Aβ_25–35-induced cytotoxicity in cultured rat pheochromocytoma (PC12) cells. In this report, the cell survival was measured by MTT assay, the enzyme activity of superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX), the contents of lipid peroxidation products malondialdehyde (MDA) and ROS in the cells were determined. Acridine orange (AO) was used to observe the morphological characteristic of apoptotic cells. Mitochondrial membrane potential in PC12 cells were monitored by fluorospectrophotometer combining with Rh123. As a cell permeable fluorescent probe, Fura-2/AM was employed to detect intracellular [Ca²⁺]. The results showed that after incubation with Aβ_25–35 (10 μM) for 24 h, there were decreased changes in cell viability, SOD, and GSH-PX activity as well as mitochondrial membrane potential, in contrast, the levels of [Ca²⁺]_i, ROS, and MDA were increased, THY significantly attenuated all the changes induced by Aβ_25–35, indicating that THY exhibited protective effect against Aβ_25–35-induced cytotoxicity, which may represent the cellular mechanisms of the action.     

The effect of APOE genotype on brain levels of oxysterols in young and old human APOE ε2, ε3 and ε4 knock-in mice

Despite apolipoprotein E's important role in cholesterol transport and metabolism in the brain as well as its influence on Alzheimer's disease, the impact of the human APOE genotype on cholesterol metabolism in brain has not been fully examined. This study was carried out to investigate APOE genotype effects on oxysterols measured. In this study the measurement of cholesterol and several oxysterols in the brains of human APOE ε2, ε3 and ε4 knock-in mice at 8 weeks and 1 year of age using gas chromatography mass spectrometry (GC–MS) demonstrated no APOE genotype or age effect on total brain cholesterol and the oxysterol 24-hydroxycholesterol. The level of 27-hydroxycholesterol was elevated in 1 year old animals for all APOE genotypes. Interestingly, lathosterol an indicator of cholesterol synthesis was significantly reduced in the 1 year old animals for all APOE genotypes. APOE ε4 expressing mice exhibited statistically lower levels of lathosterol compared to APOE ε2 in both the young and old mice. Oxidized cholesterol metabolites were significantly lower in APOE ε2 mice compared to other genotypes at 8 weeks old. Although minimal differences were observed between APOE E3 and E4 knock-in (KI) mice, these findings indicate that there are some clear APOE genotype specific effects on brain cholesterol synthesis and associated metabolic pathways, particularly in APOE ε2 KI mice.     

Protective effect of pyrroloquinoline quinone against Aβ-induced neurotoxicity in human neuroblastoma SH-SY5Y cells

The neurotoxicity of aggregated β-amyloid (Aβ) has been implicated as a critical cause in the pathogenesis of Alzheimer's disease (AD). It can cause neurotoxicity in AD by evoking a cascade of oxidative damage-dependent apoptosis to neurons. In the present study, we for the first time investigated the protective effect of pyrroloquinoline quinone (PQQ), an anionic, water soluble compound that acts as a redox cofactor of bacterial dehydrogenases, on Aβ-induced SH-SY5Y cytotoxicity. Aβ_25–35 significantly reduced cell viability, increased the number of apoptotic-like cells, and increased ROS production. All of these phenotypes induced by Aβ_25–35 were markedly reversed by PQQ. PQQ pretreatment recovered cells from Aβ_25–35-induced cell death, prevented Aβ_25–35-induced apoptosis, and decreased ROS production. PQQ strikingly decreased Bax/Bcl-2 ratio, and suppressed the cleavage of caspase-3. These results indicated that PQQ could protect SH-SY5Y cells against β-amyloid induced neurotoxicity.     

The effect of β-glucan on formation and functionality of neutrophil extracellular traps in carp (Cyprinus carpio L.)

The formation of neutrophil extracellular traps (NETs) has been characterised as a novel antimicrobial host defence strategy of neutrophils besides phagocytosis and degranulation, which may lead to entrapment and subsequent immobilisation and/or killing of bacterial pathogens. Here we studied the effect of the feed additive β-glucan, namely MacroGard®, on the formation and functionality of NETs in carp. Therefore, common carp (Cyprinus carpio) head kidney and kidney cells were isolated and treated with or without β-glucan over time. The formation of NETs was analysed by immunofluorescence microscopy and revealed a distinct increase of NET-formation with β-glucan. Furthermore the subsequent entrapment of Aeromonas hydrophila, an important fish pathogen, was increased after stimulating the cells with β-glucan. However, β-glucan did not lead to a stimulation of antimicrobial activity of neutrophils against A. hydrophila. In conclusion, the data underline the fact that the feed additive β-glucan is able to modulate carp neutrophil functions.     

Protective effect of (−)clausenamide against Aβ-induced neurotoxicity in differentiated PC12 cells

The neurotoxicity of aggregated β-amyloid (Aβ) has been implicated as a critical cause in the pathogenesis of Alzheimer's disease (AD). In the present study, we investigated the effect of (−)clausenamide ((−)Clau), an aqueous extract of leaves of Clausena lassium (lour) skeel, on the neurotoxicity of Aβ_25–35. The viability of differentiated PC12 cells was determined by MTT assay. Apoptosis was detected by flow cytometry. DCFH-DA was used for assessment of intracellular ROS generation, JC-1 and Rhodamine 123 for measurement of mitochondrial transmembrane potential (MMP). The intracellular calcium was determined with Fluo-3. The phosphorylation of p38 MAPK and the expression of Bcl-2, Bax, P53, Caspase 3 were examined by Western blot. The results showed that (−)Clau significantly elevated cell viability. Furthermore, (−)Clau arrested the apoptotic cascade by reversing overload of calcium, preventing ROS generation, moderated the dissipation of MMP and the misbalance of Bcl-2 and Bax, inhibiting the activation of p38 MAPK and the expression of P53 and cleaved Caspase 3. Our results suggested that (−)Clau may be a therapeutic agent for AD.     

Aβ25-35致PC12细胞凋亡和线粒体跨膜电位损伤关系研究

目的:探讨不同浓度Aβ25-35致PC12细胞凋亡和线粒体跨膜电位相关性。方法:体外培养大鼠嗜铬细胞瘤PC12细胞,Aβ25-35160、80、40、20、10、5、2.5μmol.L-17个浓度刺激PC-12细胞,CCK-8法观察细胞活力,Annexin-V/FITC双染后流式细胞仪检测细胞凋亡率,荧光显微镜观察hochest33342/PI双染后细胞形态,罗丹明染色后荧光显微镜检测细胞线粒体跨膜电位变化。结果:同正常组比较,Aβ25-35刺激PC-12细胞后,细胞活力逐渐下降,并呈时间和剂量依赖性,20μmol.L-1以上浓度Aβ25-35刺激12 h后PC-12细胞的细胞活力呈明显下降(P<0.05),凋亡率明显增加(P<0.05)。PC-12细胞有核浓缩和凋亡小体的产生,甚至出现死亡现象,线粒体跨膜电位下降。20μmol.L-1以下各浓度组对PC12细胞活力、细胞凋亡率、线粒体跨膜电位无明显影响(P>0.05)。结论:Aβ25-35刺激PC-12后致细胞凋亡与线粒体跨膜电位下降呈正相关,20μmol.L-1刺激12 h即可致理想的PC12细胞凋亡模型。     

Effects of β-amyloid (25–35) on learning in the common snail

The effects of neurotoxic β-amyloid fragment (25–35) on the formation of behavioral sensitization and a conditioned defensive reflex to food were studied. Administration of β-amyloid (25–35) to common snails before the start of training led to a significant reduction in sensitization of the defensive reaction, weakening of the formation of the conditioned defensive reflex to food, and impairment of memory. These impairments to behavioral plasticity may be mediated by changes in synaptic plasticity previously observed in the presence of β-amyloid. __________ Translated from Zhurnal Vysshei Nervnoi Deyatel’nosti imeni I. P. Pavlova, Vol. 57, No. 2, pp. 229–236, March–April, 2007.