Dehydroevodiamine attenuates beta-amyloid peptide-induced amnesia in mice

Dehydroevodiamine has been reported to have anticholinesterase activity and an anti-amnesic effect. This study examined the effects of dehydroevodiamine on scopolamine- and beta-amyloid peptide-(25--35)-induced amnesia in mice, using a step-through passive avoidance test. Similarly to the cholinesterase inhibitor, physostigmine (0.03--0.3 mg/kg, i.p.), dehydroevodiamine (0.75--12.0 mg/kg, i.p.) administered 30 min before the training trial, immediately after the training trial, and 30 min before the retention test significantly improved scopolamine- and beta-amyloid peptide-(25--35)-induced amnesia. In beta-amyloid peptide-(25--35)-induced amnesia, the rank order of anti-amnesic potency in these three administration schedules for dehydroevodiamine was different from that for physostigmine. Furthermore, dehydroevodiamine was more potent to improve beta-amyloid peptide-(25--35)-induced amnesia than scopolamine-induced amnesia when administered before the training trial. These results suggested that dehydroevodiamine may have an action other than that of an anticholinesterase and may be a novel and effective ligand for improvement of beta-amyloid type amnesia.     

Mycelial extract of Cordyceps ophioglossoides prevents neuronal cell death and ameliorates beta-amyloid peptide-induced memory deficits in rats

Ameliorating effects of methanol extracts of Basidiomycetes against in vitro and in vivo model of Alzheimer's disease were investigated. The extracts of Cordyceps ophioglossoides and Hypocrea citrina var. citrina prevented the beta-amyloid((25-35)) (Abeta((25-35)))-induced cell death in SK-N-SH neuronal cells. However, in rat model of Alzheimer's disease, 30-d intraperitoneal administration with only the extract of Cordyceps ophioglossoides significantly prevented spatial memory loss by intracranial injection of Abeta((25-35)), which was assessed in water maze task. Interestingly, the scavenging activity of the reactive oxygen species (ROS) generated in Abeta((25-35))-treated cells was also found in the extract of Cordyceps ophioglossoides, but not in the extract of Hypocrea citrina var. citrina. These results suggest that the extract of Cordyceps ophioglossoides may protect the Abeta-induced neuronal cell death and memory loss through free radical scavenging activity. These results further suggest that Cordyceps ophioglossoides mycelium may be valuable for the protection from Alzheimer's dementia.     

Quercetin attenuates oxygen-glucose deprivation- and excitotoxin-induced neurotoxicity in primary cortical cell cultures

The possible role of quercetin, a naturally occurring plant flavonoid, in protecting against oxygen-glucose deprivation (OGD)-, excitotoxins-, and free radical-induced neuronal injury in mouse cortical cell cultures was investigated. Pre- and co-treatment with quercetin (100 microM) inhibited 50 min OGD-, 20 microM N-methyl-D-aspartate (NMDA)-, and 50 microM kainate-induced neurotoxicity by 36, 22, and 61%, respectively. Quercetin significantly ameliorated free radical-induced neuronal injury caused by buthionine sulfoximine, sodium nitroprusside, ZnCl(2), and FeCl(2). These results suggest that quercetin may contribute a neuroprotective action against ischemic neural injury, partially via antioxidant actions.     

阿米替林对谷氨酸损伤中枢神经细胞的保护作用

目的：研究阿米替林（Ami)对谷氨酸损伤培养大鼠皮层神经细胞的保护作用。方法：分离培养15－18d胎龄的大鼠神经细胞，加入谷氨酸（Glu)，造成神经细胞的损伤，测定乳酸脱氢酶，一氧化氮，丙二醛及超氧化物歧化酶的含量，观察谷氨酸对神经细胞的损伤作用及阿米替林的保护作用。结果：加入谷氨酸后，神经细胞出现明显损伤性变化，死亡率升高，培养上清液中乳酸脱氢酶（LDH），一氧化氮（NO）含量增加，细胞匀浆中丙二醛（MDA）生成增加，超氧化物歧化酶（SOD）含量明显减少，Ami10^-8-10^-6mol/L^-1能不同程度地减轻上述损伤性变化。结论：Ami对谷氨酸所致的神经细胞损伤具有保护作用。其作用机制可能与抗脂质过氧化作用有关。     

Cyanide-induced neurotoxicity: calcium mediation of morphological changes in neuronal cells

Calcium channel blockade decreases the elevation of brain calcium as well as the tremors produced by cyanide in mice. To determine if cyanide-induced morphological changes could also be inhibited by calcium channel blockade, the effect of diltiazem was studied in cultured rat pheochromocytoma (PC12) cells, a neuronal model. Incubation with KCN (1 to 10 mM for 1 to 2 hr) caused depletion of secretory granules, alignment of remaining granules along the plasma membrane, and mitochondrial swelling. All these effects were inhibited by pretreatment with 0.01 mM diltiazem. Scanning electron microscopy revealed that cyanide (1 to 10 mM for 1 to 2 hr) produced loss of microvilli and bleb formation in PC12 cells. These changes were partially inhibited by preincubation with 0.01 mM diltiazem. Incubation of cells with 10 mM cyanide increased release of lactic dehydrogenase (LDH) into the culture media at 60 and 120 min. A decrease in cell viability, as determined by trypan blue dye exclusion, paralleled the release of LDH. At 120 min of cyanide incubation, 24% of the cells excluded dye. Both the release of LDH and decreased cell viability were attenuated by pretreatment with diltiazem. The results indicate that the influx of extracellular calcium is an important factor mediating cyanide-induced morphologic changes in neuronal cells.     

Role of hydroxyl radical formation in neurotoxicity as revealed by in vivo free radical trapping

Reactive oxygen species have been implicated in dopaminergic toxicity caused by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and iron. Although MPTP produces a parkinsonian syndrome after its conversion to 1-methyl-4-phenylpyridine (MPP(+)) by type B monoamine oxidase (MAO-B) in the brain, the etiology of this disease remains obscure. MPP(+) is a highly potent dopaminbergic-releasing agents and dopamine (DA) autoxidation catalyzed by iron and oxidative stress may be involved in the pathogenesis of Parkinson's disease. Neuromelanine synthesis from DA produce highly reactive free radicals. Although the controversy possible neurotoxin and/or neuroprotective roles of nitric oxide (NO) was discussed, NO contributes to oxidative injury to brain neurons in vivo. An environmental estrogen-like chemical also related to MPP(+)-induced *OH generation. This review describes actual mechanism of the free radicals formation by dialysis studies of in vivo free radical trapping in the pathogenesis of neurodegenerative disorders, including in the Parkinson's disease, Alzheimer disease and traumatic brain injuries.     

Pre-treatment and post-treatment with alpha-tocopherol attenuates hippocampal neuronal damage in experimental cerebral hypoperfusion

Alpha-tocopherol, a potent antioxidant, has been widely investigated as a dietary supplement with which to reduce the risk of atherosclerosis, and has recently been considered as a potential supplement to moderate oxidative neuronal damage in Alzheimer's disease patients. Since alpha-tocopherol appears beneficial in vascular and neurodegenerative disorders, we set out to identify its neuroprotective action in a rat model of chronic cerebral hypoperfusion-induced brain injury. The bilateral common carotid arteries of male Wistar rats were permanently occluded (2VO). Sham-operated animals served as controls. Half of the animals were pre- or post-treated repeatedly with alpha-tocopherol (5x100 mg/kg daily, i.p.), the other half receiving only soybean oil, the alpha-tocopherol vehicle. One week after the onset of 2VO, the spatial learning capacity of the animals was assessed in the Morris water maze. After testing, hippocampal slices were stained with cresyl violet in order to examine the pyramidal cell layer integrity. The density of microtubule-associated protein-2 (MAP-2)-positive dendrites and the OX-42-labeled microglial activation level were determined immunocytochemically. Finally, alpha-tocopherol was determined in the peripheral tissues, blood and brain. Alpha-tocopherol moderated the 2VO-induced learning impairment. The various forms of alpha-tocopherol treatment, and particularly the post-treatment, prevented the 2VO-induced pyramidal cell death and the activation of microglia in the hippocampus CA1 region, and the degeneration of MAP-2-positive dendrites in the CA3 region. The alpha-tocopherol concentration was elevated in the peripheral tissues and the blood, but not in the brain. The data indicate that alpha-tocopherol, particularly when administered as post-treatment, is neuroprotective in chronic cerebral hypoperfusion.     

Idebenone protects hippocampal neurons against amyloid β-peptide-induced neurotoxicity in rat primary cultures

The application of amyloid β-peptide (Aβ) 1–40 (10 μM) caused neurodegeneration of hippocampal neuronal cells, as indicated by the release of lactate dehydrogenase (LDH) into the culture medium. Treatment with idebenone (10–1000 nM), a potent antioxidant in mitochondria, protected the hippocampal neurons against the Aβ1–40 (10 μM)-induced neurotoxicity. To determine the morphological change in neurons during the Aβ1–40-induced cytotoxicity, the cells were immunostained with anti-MAP2 antibodies. After 4-day exposure to 10 μM Aβ1–40, the number of neurons was reduced, and the surviving neurons had an apparently reduced number of neurites which were shorter than those of control neurons. When idebenone was added to the culture medium with Aβ1–40, the number of surviving neurons was significantly increased, and their neurites were as long as seen in control culture. These results suggest that reactive oxygen species mediate neurotoxicity of Aβ1–40, and idebenone protects neurons against the Aβ1–40-induced neurotoxicity. Received: 5 June 1998 / Accepted: 25 August 1998     

藻酸双酯钠对培养神经细胞谷氨酸兴奋毒性的保护作用

目的 :观察藻酸双酯钠 (PSS)对培养神经细胞谷氨酸兴奋毒性的保护作用。方法 :体外培养大鼠胚胎皮层神经细胞 ,加入谷氨酸观察谷氨酸对神经细胞的兴奋毒性及PSS的保护作用。结果 :PSS 5 0 ,1 0 0 ,1 5 0mg·L-1能显著减少细胞死亡 ,降低乳酸脱氢酶(LDH)漏出量 ,一氧化氮 (NO)含量及丙二醛 (MDA)生成 ,提高超氧歧化酶 (SOD)活性。结论 :PSS对培养神经细胞谷氨酸兴奋毒性具有显著的保护作用 ,其机制可能与PSS抗脂质过氧化作用有关。     

神经生长因子对培养的脑皮质神经元谷氨酸毒性的影响

在原代培养的大鼠胎鼠脑皮质神经元，观察神经生长因子(NGF)对谷氨酸损伤的影响. 谷氨酸(0.2-0.8 mmol·L^-1) 促进神经元死亡及乳酸脱氢酶(LDH)释放，增加丙二醛(MDA)含量. NGF 3-100 μg·L^-1浓度依赖地抑制谷氨酸引起的神经元死亡及LDH和MDA的增加. NGF 30 μg·L^-1提高超氧化物歧化酶和谷胱甘肽过氧化酶活力. NGF浓度依赖地增加谷氨酸引起的抗氧化酶水平降低. 结果提示NGF通过抑制脂质过氧化物生成及提高抗氧化酶活性来保护大脑皮质细胞抗谷氨酸毒性.     

Neuroprotective effect of mithramycin against endoplasmic reticulum stress-induced neurotoxicity in organotypic hippocampal slice cultures

Endoplasmic reticulum (ER) stress has been implicated in the pathogenesis of various neurodegenerative diseases. Although the underlying mechanisms of these diseases have been suggested by many studies, therapeutic drugs have yet to be found. In this study, experiments were performed to examine the effect of mithramycin (MTM), a clinically approved guanosine-cytosine (GC)-rich DNA sequence-binding antitumor antibiotic, on ER stress-induced neurotoxicity in organotypic hippocampal slice cultures (OHCs). Time-dependent induction of the ER chaperones, glucose-regulated protein (GRP) 78 and GRP94, was observed after treatment with tunicamycin (TM) (80?μg/mL). Western blot analysis showed that treatment of OHCs with TM increased the expression of CHOP and the cleaved forms of caspase-12. Simultaneous application of MTM suppressed TM-induced cell death in all areas of OHCs with a concomitant decrease in the level of CHOP. In contrast, MTM had no effect on excitotoxic cell death induced by ibotenic acid, a potent N-methyl-d-aspartate (NMDA) agonist in OHCs. Moreover, RNA interference to CHOP or simultaneous treatment with MTM attenuated TM-induced cell death in primary cultured hippocampal neurons. These results suggest that CHOP plays a critical role in the mechanisms underlying ER-stress-induced neurotoxicity in the hippocampus, and that MTM could be a protective agent against ER stress-induced hippocampal neuronal death through attenuation of ER stress-associated signal proteins.     

Effect of nerve growth factor on glutamate-induced neurotoxicity in cerebral cortical neuronal cultures

在原代培养的大鼠胎鼠脑皮质神经元，观察神经生长因子（ＮＧＦ）对谷氨酸损伤的影响．谷氨酸（０．２－０．８ｍｍｏｌ·Ｌ－１）促进神经元死亡及乳酸脱氢酶（ＬＤＨ）释放，增加丙二醛（ＭＤＡ）含量．ＮＧＦ３－１００μｇ·Ｌ－１浓度依赖地抑制谷氨酸引起的神经元死亡及ＬＤＨ和ＭＤＡ的增加．ＮＧＦ３０μｇ·Ｌ－１提高超氧化物歧化酶和谷胱甘肽过氧化酶活力．ＮＧＦ浓度依赖地增加谷氨酸引起的抗氧化酶水平降低．结果提示ＮＧＦ通过抑制脂质过氧化物生成及提高抗氧化酶活性来保护大脑皮质细胞抗谷氨酸毒性．     

Blockade of N-methyl-D-aspartate receptors prevents cyanide-induced neuronal injury in primary hippocampal cultures.

Cyanide-induced alterations of cytosolic calcium levels and cytotoxicity were examined in primary cultures of rat hippocampus. Cytosolic free Ca2+ ([Ca2+]i) levels were measured in hippocampal neurons using the fluorescent probe, fura 2. A concentration-dependent rise in [Ca2+]i occurred rapidly following exposure of cells to 0.5-10 mM NaCN. In normal medium (1.3 mM Ca2+), 2 mM NaCN produced an increase in [Ca2+]i (172 +/- 27% of control), 45 sec following exposure. Ca2+ elevation produced by NaCN was blocked by removal of Ca2+ from the external medium or by pretreatment with the N-methyl-D-aspartate (NMDA) receptor antagonist, 2-amino-5-phosphonovalerate (APV). The cytotoxicity of cyanide, assessed by measuring the efflux of lactate dehydrogenase, was blocked by APV. These results indicate that in hippocampal neurons, cytosolic Ca2+ accumulation induced by cyanide originates from the extracellular compartment and the NMDA receptor ionophore is a significant route for Ca2+ entry. It is proposed that excitotoxic mechanisms may contribute to altered neuronal homeostasis and injury associated with cyanide.     

Huperzine A attenuates cognitive dysfunction and neuronal degeneration caused by beta-amyloid protein-(1-40) in rat

Huperzine A, a promising therapeutic agent for Alzheimer's disease, was examined for its potential to antagonize the deleterious neurochemical, structural, and cognitive effects of infusing beta-amyloid protein-(1-40) into the cerebral ventricles of rats. Daily intraperitoneal administration of huperzine A for 12 consecutive days produced significant reversals of the beta-amyloid-induced deficit in learning a water maze task. This treatment also reduced the loss of choline acetyltransferase activity in cerebral cortex, and the neuronal degeneration induced by beta-amyloid protein-(1-40). In addition, huperzine A partly reversed the down-regulation of anti-apoptotic Bcl-2 and the up-regulation of pro-apoptotic Bax and P53 proteins and reduced the apoptosis that normally followed beta-amyloid injection. The present findings confirm that huperzine A can alleviate the cognitive dysfunction induced by intracerebroventricular infusion of beta-amyloid protein-(1-40) in rats. The beneficial effects are not confined to the cholinergic system, but also include favorable changes in the expression of apoptosis-related proteins and in the extent of apoptosis in widespread regions of the brain.     

Cholesterol reduction attenuates 5-HT1A receptor-mediated signaling in human primary neuronal cultures

Lipid rafts regulate functions of various G protein-coupled receptors and signaling proteins. We show that human primary neuronal cultures contain high levels of 5-HT(1A) receptors. Stimulation with the 5-HT(1A/7) receptor agonist, 8-OH-DPAT, reduced P-T(185)/Y(187)-ERK2. This reduction could be blocked by the 5-HT(1A) receptor antagonist, WAY100635. Pretreatment with the cholesterol sequestering agent, methyl-beta-cyclodextrin, before adding 8-OH-DPAT, significantly counteracted the inhibitory influence of 8-OH-DPAT on P-T(185)/Y(187)-ERK2 and P-S(133)-CREB. These data indicate that reduction of cholesterol levels significantly influence signaling via 5-HT(1A) receptors in intact neurons.     

Effects of nicotine on hydroxyl free radical formation in vitro and on MPTP-induced neurotoxicity in vivo

Parkinson’s disease (PD) is one of the most frequent disorders of the basal ganglia. From epidemiological studies there is a controversial discussion on the question whether tobacco smoking is correlated with a decreased incidence of PD. The present study aimed to elucidate the role of nicotine and its potential neuroprotective effects in a rodent model of PD. These effects may be related to an altered hydroxyl radical formation; this possibility was studied in vitro. Nicotine and α-phenyl-N-tert-butyl nitrone (PBN) were examined in a cell-free in vitro Fenton system (Fe³⁺/EDTA + H₂O₂) for their radical scavenging properties using the salicylate trapping method. Salicylic acid (0.5 mM) was incubated in the presence and absence of nicotine or PBN and the main products of the reaction of hydroxyl radicals with salicylic acid, namely 2,3- and 2,5-dihydroxybenzoic acid, were immediately determined using HPLC in combination with electrochemical detection. Nicotine and PBN were both able to significantly reduce hydroxyl radical levels at concentrations of 1, 2.5 and 5 mM. Interestingly, at 5 mM nicotine was able to reduce hydroxyl radical levels significantly more than the radical scavenger PBN (5 mM). To investigate the in vivo effects of nicotine, male C57BL/6 mice were used in the MPTP mouse model of PD. Nicotine (0.1 or 0.4 mg/kg s.c.) was administered twice daily for a period of 14 days. On day 8 a single injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP, 30 mg/kg s.c.) was given as well as an enhanced protocol of nicotine treatment (0.1 or 0.4 mg/kg s.c., 30 min before MPTP and 30, 90, 210, 330, 450, 570 min after MPTP) for a total of seven injections of nicotine. High dosage nicotine treatment significantly increased the MPTP-induced loss of body weight and resulted in a significantly decreased striatal dopamine content and an increased dopamine turnover in comparison with the MPTP-treated controls at day 15. However, the lower dosage of nicotine did not significantly alleviate the MPTP-induced effects, although some parameters showed a slight tendency in this direction. These results demonstrate that in vitro nicotine has radical scavenging properties which might suggest neuroprotective effects. In vivo experiments with nicotine, however, showed that a low dosage of nicotine did not alleviate the MPTP-induced dopamine depletion, but a large dosage even enhanced it. Received: 20 March 1998 / Accepted: 23 June 1998     

Activation of GABA(A) receptors by taurine and muscimol blocks the neurotoxicity of beta-amyloid in rat hippocampal and cortical neurons

The β-amyloid peptide (Aβ) is centrally related to the pathogenesis of Alzheimer's disease (AD) and is potently neurotoxic to central nervous system neurons. The neurotoxicity of Aβ has been partially related to the over activation of glutamatergic transmission and excitotoxicity. Taurine is a naturally occurring β-amino acid present in the mammalian brain. Due to its safety and tolerability, taurine has been clinically used in humans in the treatment of a number of non-neurological disorders. Here, we show that micromolar doses of taurine block the neurotoxicity of Aβ to rat hippocampal and cortical neurons in culture. Moreover, taurine also rescues central neurons from the excitotoxicity induced by high concentrations of extracellular glutamate. Neuroprotection by taurine is abrogated by picrotoxin, a GABA_A receptor antagonist. GABA and muscimol, an agonist of the GABA_A receptor, also block neuronal death induced by Aβ in rat hippocampal and cortical neurons. These results suggest that activation of GABA_A receptors protects neurons against Aβ toxicity in AD-affected regions of the mammalian brain and that taurine should be investigated as a novel therapeutic tool in the treatment of AD and of other neurological disorders in which excitotoxicity plays a relevant role.     

Mechanisms of manganese-induced neurotoxicity in primary neuronal cultures: the role of manganese speciation and cell type

Manganese (Mn) is an essential trace element required for the proper functioning of a variety of physiological processes. However, chronic exposures to Mn can cause neurotoxicity in humans, especially when it occurs during critical stages of the central nervous system development. The mechanisms mediating this phenomenon as well as the contribution of Mn speciation and the sensitivity of different types of neuronal cells in such toxicity are poorly understood. This study was aimed to investigate the mechanisms mediating the toxic effects of MnCl(2), Mn(II) citrate, Mn(III) citrate, and Mn(III) pyrophosphate in primary cultures of neocortical (CTX) and cerebellar granular (CGC) neurons. Cell viability, mitochondrial function, and glutathione levels were evaluated after Mn exposure. CGC were significantly more susceptible to Mn-induced toxicity when compared with CTX. Moreover, undifferentiated CGC were more vulnerable to Mn toxicity than mature neurons. Mitochondrial dysfunction was observed after the exposure to all the tested Mn species. Ascorbate protected CGC against Mn-induced neurotoxicity, and this event seemed to be related to the dual role of ascorbate in neurons, acting as antioxidant and metabolic energetic supplier. CTX were protected from Mn-induced toxicity by ascorbate only when coincubated with lactate. These findings reinforce and extend the notion of the hazardous effects of Mn toward neuronal cells. In addition, the present results indicate that Mn-induced neurotoxicity is influenced by brain cell types, their origins, and developmental stages as well as by the chemical speciation of Mn, thus providing important information about Mn-induced developmental neurotoxicity and its risk assessment.     

Geniposide from Gardenia jasminoides attenuates neuronal cell death in oxygen and glucose deprivation-exposed rat hippocampal slice culture

Geniposide from Gardenia jasminoides protected neuronal cells from damage in oxygen and glucose deprivation-exposed hippocampal slice culture. Geniposide showed a greater protective effect on the granule cell layer than on the pyramidal cell layer including CA 1 and CA 3. On the basis of the experimental results, geniposide may be a therapeutic agent for ischemia in patients.