Protective effect of nimodipine against quinolinic acid-induced damage of rat hippocampus in vitro.

The study was undertaken to examine the effect of nimodipine, calcium channels blocker, on the morphological alterations induced by quinolinic acid (QUIN). The experiment was performed on 21-day-old organotypic rat hippocampal cultures. Nimodipine was applied to the nutrient medium simultaneously with QUIN (both at 100 microM). Ultrastructural changes were evaluated 24 h, 5 and 7 days after the exposure to tested agent. It was shown that nimodipine induced distinct cytoprotective effect, especially considering the development of late neurotoxic injury produced by QUIN. However, the protection was not complete, indicating the participation of the other factors in the pathomechanism underlying structural damage produced by QUIN.     

慢性应激抑郁模型大鼠强迫游泳后海马中HsP70的表达

目的研究慢性应激对强迫游泳大鼠海马神经元热休克蛋白70(Hsp70)表达的影响。方法将50 只大鼠随机分为实验组和对照组各25只。实验组大鼠通过21天的应激刺激制作抑郁动物模型,此期间对照组大 鼠正常饲养。此后所有大鼠逐只进行急性强迫应激刺激。采用特异性抗体的免疫组织化学方法,观察2组大鼠在 强迫游泳后2 h、6 h、18 h、24 h和48 h各时点海马神经元Hsp70的表达情况。结果慢性应激抑郁大鼠模型接受 急性强迫游泳应激后,海马CA3区和齿状回(DG)内Hsp70蛋白的表达较对照组强迫游泳后显著降低(P<0．05)。 结论慢性应激使大鼠在急性强迫游泳应激后海马CA3区和DG内Hsp70的表达降低。     

Effect of chronic stress on cholinergic transmission in rat hippocampus

We previously demonstrated that chronic stress impaired prefrontal cortex-sensitive working memory, but not reference memory. Since the hippocampal cholinergic system is also involved in these memories, we examined the effects of chronic stress on cholinergic transmission in the rat hippocampus. A microdialysis study revealed that the stress did not affect the basal acetylcholine release, but enhanced the KCl-evoked response. These results suggest that cholinergic transmission in the chronically stressed hippocampus does not contribute to working memory impairment, but it may be involved in maintenance of reference memory.     

Protective effect of nimodipine against ischemic neuronal damage in rat hippocampus without changing postischemic cerebral blood flow

The purpose of the present study was to investigate the neuroprotective action of nimodipine. Furthermore, the influence of nimodipine on postischemic local CBF (LCBF) was examined. Forebrain ischemia of the rat was performed for 10 min by bilateral carotid clamping, administration of trimethaphan, and blood withdrawal to obtain an MABP of 40 mm Hg. LCBF was measured after 10 min of postischemic recirculation by injecting [14C]iodoantipyrine in saline solution. Nimodipine (0.1, 0.3, and 1.0 mg/kg) was suspended in miglyol oil and applied orally 60 min prior to ischemia. Histological evaluation was performed 7 days after ischemia. Hippocampal neuronal damage was determined as the percentage of necrotic neurons. After preischemic application of nimodipine, neuronal damage was significantly reduced in the hippocampal CA1 subfield. Postischemic LCBF was not affected by treatment with nimodipine. These findings show that nimodipine is able to protect neurons against ischemic damage. The neuroprotective effect of nimodipine was not mediated by a postischemic cerebral vasodilation, but by a direct action on the neurons.     

Effects of chronic stress and corticosterone on sialidase activity in the rat hippocampus

Sialidases are acid exoglycosidases that catalyse the removal of sialic acid from non-reducing end of sialoglucoconjugated substrates. Synaptic plasticity depends on sialylation state of proteins and lipids mediated by sialic acid-metabolizing enzymes. Since chronic stress causes both, hippocampal atrophy and impairment of learning, it is reasonable to investigate whether sialidase is implicated in these processes. In this study, we tested effects of chronic stress (immobilization, 2 h daily, 21 days) or chronic corticosterone administration (5 mg/kg, sc, daily) on sialidase activity and sialylated NCAMs expression in rat hippocampus.The results showed that chronic stress affects hippocampus-depended spatial learning in the Barnes maze. Both, stress (p > 0.05) and corticosterone (p < 0.001), increased latencies to enter the escape tunnel of the maze in comparison to control animals. Similar but not significant differences between control and other experimental groups were observed in the numbers of errors. Chronic stress (p > 0.05) and corticosterone (p < 0.05) decreased sialidase activity in the brain homogenates and synaptosomes (p < 0.05, both). In the stressed animals, these changes were related to significantly higher expression of polysialic acid. These results indicate that changes in sialidase activity caused by stress and chronic corticosterone administration reflect disturbances of polysialylated glycoconjugates known to be related to synaptic plasticity in hippocampus.     

Protective effect of 7-nitroindazole against DSP-4 induced noradrenaline depletion in mouse hippocampus

Summary. N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) is a selective noradrenaline (NA) uptake blocker, capable of inducing a long-lasting depletion of NA in some noradrenergic axon terminals originating from the locus coeruleus in rodents. Pretreatment with 7-nitroindazole, a fairly selective inhibitor of neuronal nitric oxide synthase in vivo, partially prevented DSP-4 induced NA depletion in mouse hippocampus measured seven days after the neurotoxic insult. Administration of L-arginine, the substrate of nitric oxide synthase, altered neither the NA depletion induced by DSP-4, nor the protective effect of 7-nitroindazole. Inhibition of nitric oxide synthesis by N(G)-nitro-L-arginine methyl ester did not attenuate the NA depleting effect of DSP-4. Thus, the contribution of neuronal nitric oxide synthase inhibition to the protective effect of 7-nitroindazole needs further studies. As 7-nitroindazole did not block NA uptake, this cannot play a part in the protective effect. The possible contribution of monoamine oxidase B enzyme inhibition by 7-nitroindazole to the protective effect is also discussed. Received July 3, 2000; accepted October 16, 2000     

Overexpression of mitochondrial Hsp70/Hsp75 protects astrocytes against ischemic injury in vitro.

Mitochondrial heat shock protein 70 (mtHsp70/Hsp75/Grp75/mortalin/TRAP-1/PBP74) is an essential mitochondrial chaperone and a member of the heat shock protein 70 (HSP70) family. Although many studies have shown the protective properties of overexpression of the cytosolic inducible member of the HSP70 family, Hsp72, few studies have investigated the protective potential of Hsp75 against ischemic injury. Mitochondria are one of the primary targets of ischemic injury in astrocytes. In this study, we analyzed the effects of Hsp75 overexpression on cellular levels of reactive oxygen species (ROS), mitochondrial membrane potential, ATP levels, and viability during the ischemia-like conditions of oxygen-glucose deprivation (OGD) or glucose deprivation (GD) in primary astrocytic cultures. We show that Hsp75 overexpression decreases ROS production and preserves mitochondrial membrane potential during GD, and preserves ATP levels and cell viability during OGD. These findings indicate that Hsp75 can provide protection against ischemia-like in vitro injury and suggest that it should be further studied as a potential candidate for protection against ischemic injury.     

慢性应激抑郁模型大鼠海马GSK-3β的表达

目的 研究慢性应激对大鼠海马糖原合成酶激酶-3β(GSK-3β)表达的影响.方法 实验组大鼠通过21d的应激刺激制作抑郁动物模型,此期间对照组大鼠正常饲养.Open-field法检测行为学改变,Western blot法测定大鼠海马GSK-3β的表达.结果 慢性应激后,实验组大鼠体重、水平穿越格数、直立次数和修饰次数均显著低于对照组(P<0.01);实验组大鼠海马GSK-3β表达的整合光密度值(28413.30±2501.31)高于对照组(23149.50±3107.25),差异有统计学意义(P<0.05).结论 慢性应激使大鼠海马GSK-3β的表达上调.     

Vitamin E protects against alcohol-induced cell loss and oxidative stress in the neonatal rat hippocampus

Oxidative stress has been proposed as a possible mechanism underlying nervous system deficits associated with Fetal Alcohol Syndrome (FAS). Current research suggests that antioxidant therapy may afford some level of protection against the teratogenic effects of alcohol. This study examined the effectiveness of antioxidant treatment in alleviating biochemical, neuroanatomical, and behavioral effects of neonatal alcohol exposure. Neonatal rats were administered alcohol (5.25 g/kg) by intragastric intubation on postnatal days 7, 8, and 9. A subset of alcohol-exposed pups were co-administered a high dose of Vitamin E (2 g/kg, or 71.9 IU/g). Controls consisted of a non-treated group, a group given the administration procedure only, and a group given the administration procedure plus the Vitamin E dose. Ethanol-exposed animals showed impaired spatial navigation in the Morris water maze, a decreased number of hippocampal CA1 pyramidal cells, and higher protein carbonyl formation in the hippocampus than controls. Vitamin E treatment alleviated the increase in protein carbonyls and the reduction in CA1 pyramidal cells seen in the ethanol-exposed group. However, the treatment did not improve spatial learning in the ethanol-exposed animals. These results suggest that while oxidative stress-related neurodegeneration may be a contributing factor in FAS, the antioxidant protection against alcohol-induced oxidative stress and neuronal cell loss in the rat hippocampus does not appear to be sufficient to prevent the behavioral impairments associated with FAS. Our findings underscore the complexity of the pathogenesis of behavioral deficits in FAS and suggest that additional mechanisms beyond oxidative damage of hippocampal neurons also contribute to the disorder.     

Cytoskeletal alterations in rat hippocampus following chronic unpredictable mild stress and re-exposure to acute and chronic unpredictable mild stress

The intrinsic dynamic instability of the cytoskeletal microtubular system is essential for neuronal development and organization. The modulation of microtubule dynamics depends on the phosphorylation of neuronal microtubule-associated proteins (MAPs). Chronic unpredicted mild stress (CUMS) affects hippocampal structure and function in the rat. The aim of the present work was to investigate the possible alteration of cytoskeleton in the hippocampus of rats exposed to CUMS and re-exposed to CUMS to mimic depression and the recurrence of depression of human. We investigated the effects of CUMS, fluoxetine and re-exposure to CUMS on α-tubulin isoforms associated with microtubule dynamics, MAP-2 and phospho-MAP-2 in the hippocampus of rats. Our results showed that rats submitted to CUMS once showed a significant reduction in locomotion and sucrose preference which indicate a state of anhedonia. These behavioral alterations were accompanied by specific alterations in hippocampal α-tubulin isoforms and phospho-MAP-2 expression, indicating less microtubule dynamics and the possible mechanism. Treatment of fluoxetine could reverse CUMS-induced impairment. Moreover, there were more dramatically changes in behaviors, α-tubulin isoforms and phospho-MAP-2 of rats re-exposed to CUMS compared to the rats exposed to CUMS once. These findings provide evidence that rats exposed to CUMS and re-exposed to CUMS showed impairment of microtubule dynamics accompanied with the decreased level of phospho-MAP-2, providing insight into the role of cytoskeleton in the depression and recurrent of depression.     

Neuroprotective role of Bacopa monniera extract against aluminium-induced oxidative stress in the hippocampus of rat brain

Bacopa monniera is a nerve tonic used extensively in traditional Indian medicinal system "Ayurveda". Reports regarding its various antioxidative, adaptogenic and memory enhancing roles have already appeared in the last few decades. In the present study, aluminium chloride (AlCl(3)) was used to generate neurotoxicity. We have investigated the neuroprotective effect of Bacopa extract against aluminium-induced changes in peroxidative products, such as thio-barbituric acid-reactive substance (TBA-RS) and protein carbonyl contents and superoxide dismutase (SOD) activity. Effect on lipofuscin (age pigments) accumulation and ultrastructural changes were also studied. Bacopa effects were compared with those of l-deprenyl. Co-administration of Bacopa extract during aluminium treatment significantly prevented the aluminium-induced decrease in SOD activity as well as the increased oxidative damage to lipids and proteins. Protective effect was also observed at microscopic level. Fluorescence and electron microscopic studies revealed considerable inhibition of intraneuronal lipofuscin accumulation and necrotic alteration in the CA1 region of the hippocampus. Observations showed that Bacopa's neuroprotective effects were comparable to those of l-deprenyl at both biochemical and microscopic levels.     

Transient elevation of synaptosomal mitoenergetic proteins and Hsp70 early in a rat model of chronic cerebrovascular hypoperfusion

Chronic cerebral hypoperfusion (CCH) might account for the cognitive deficits associated with vascular cognitive impairment, but the mechanisms of hypoperfusion insulting to the cognition remain obscure. In the present study, Wistar rats underwent permanent occlusion of bilateral common carotid arteries to induce CCH. 2D-DIGE combined with MALDI-TOF MS was applied to determine the proteins that were differentially expressed in synaptosomes of prefrontal cortex and hippocampus. ATPsynβ, NDUFS1, UQCRC1 and Hsp70 were elevated both in synaptosomes of cortex and hippocampus at week 2 after operation, but subsided to baseline at week 4 except ATPsynβ which was still upregulated in synaptosomes of hippocampus at week 4. IDH3A and PDC-E2 were increased, respectively, in synaptosomes of prefrontal cortex and hippocampus at week 2, and showed no difference when compared to control at week 4. Malate dehydrogenase showed no difference in synaptosomes of prefrontal cortex and hippocampus at week 2, but showed an elevation in synaptosomes of prefrontal cortex at week 4. Our results imply that metabolic reserve and anti-oxidative stress might transiently exist in the early stage of CCH, which probably help cognitive save.     

槲皮素对氧化应激大鼠星形胶质细胞的保护作用

目的 研究槲皮素对氧化应激损伤后大鼠星形胶质细胞的保护作用. 方法 采用终浓度为2 mmol/L的H2O2作用于体外原代培养的大鼠胶质细胞6 h,以诱导氧化应激.实验分为正常对照组、H2O2组、槲皮素+H2O2组.不同浓度(0、50、100、200μmol/L)槲皮素预处理24h后,应用速率法和LIVE/DEAD检测试剂盒分别检测氧化应激胶质细胞的乳酸脱氢酶(LDH)释放率以及细胞存活率的变化. 结果 终浓度为2 mmol/L的H2O2作用6 h即可造成细胞损伤,LDH释放率由对照组的(3.89±1.89)%增至(90.27±2.68)%,较对照组明显增多,细胞存活率由对照组的(99.25±0.08)%降至(59.73%±9.92)%,较对照组明显降低,差异均有统计学意义(P<0.05).槲皮素预处理后细胞LDH释放率降低,50、100、200 μmol/L浓度的槲皮素组LDH释放率分别减少到(48.19±13.98)%、(27.81±9.33)%和(18.13±8.28)%,与H2O2组比较差异有统计学意义(P<0.05);槲皮素预处理同时能提高细胞存活率,50、100、200 μmol/L浓度的槲皮素组细胞存活率分别提高至(86.80±3.62)%、(88.32±5.77)%和(91.18±3.03)%,与H2O2组比较差异均有统计学意义(P<0.05). 结论 槲皮素预处理对氧化应激大鼠胶质细胞有一定的保护作用.     

Hsp27 and Hsp70 administered in combination have a potent protective effect against FALS-associated SOD1-mutant-induced cell death in mammalian neuronal cells

Amyotrophic lateral sclerosis (ALS) is an adult-onset degenerative disorder characterised by the death of motor neurons in the cortex, brainstem, and spinal cord; resulting in progressive muscle weakness, atrophy, and death from respiratory paralysis, usually within 3-5 years of symptom onset. Approximately 10% of ALS cases are familial (FALS). Mutations in superoxide dismutase-1 (SOD1) cause approximately 20% of FALS cases and there is overwhelming evidence that a toxic gain of function is the cause of the disease. We have previously shown that FALS-associated SOD1 disease mutants enhanced neuronal death in response to a wide range of stimuli tested whereas wt-SOD1 protected against all insults. We demonstrate for the first time that over-expression of either heat shock protein Hsp27 or Hsp70 has a protective effect against SOD1 disease associated mutant-induced cell death. However, over-expression of Hsp27 and Hsp70 together has a greater potent anti-apoptotic effect, than when expressed singly, against the damaging effects of mutant SOD1. Our results indicate that FALS-associated SOD1 disease mutants possess enhanced death-inducing properties and lead to increased apoptosis which can be prevented by either the use of specific caspase inhibitors or Hsp27 and/or Hsp70 over-expression. This potent protective effect of Hsp27 and Hsp70 against the FALS-associated SOD1 disease mutants may be of potential therapeutic importance.     

Effects of chronic social isolation on wistar rat behavior and brain plasticity markers

Chronic stress is a contributing risk factor in the development of psychiatric illnesses, including depressive disorders. The mechanisms of their psychopathology are multifaceted and include, besides others, alterations in the brain plasticity. Previously, we investigated the effects of chronic social stress in the limbic brain structures of Wistar rats (hippocampus, HIPPO, and prefrontal cortex, PFC) and found multiple characteristics that resembled alterations described in some clinical studies of depression. We extended our investigations and followed the behavior of stressed animals by the open field test (OFT) and forced swimming test (FST), and the expression and polysialylation of synaptic plasticity markers, neural cell adhesion molecule (NCAM) and L1, in the HIPPO and PFC. We also determined the adrenal gland mass and plasma corticosterone (CORT) as a terminal part of the hypothalamic-pituitary-adrenal axis activity. Our data indicated that stressed animals avoided the central zone in the OFT and displayed decreased swimming, but prolonged immobility in the FST. The animals exhibited marked hypertrophy of the adrenal gland cortex, in spite of decreased serum CORT. Simultaneously, the stressed animals exhibited an increase in NCAM mRNA expression in the HIPPO, but not in the PFC. The synaptosomal NCAM of the HIPPO was markedly polysialylated, while cortical PSA-NCAM was significantly decreased. The results showed that chronic social isolation of Wistar rats causes both anxiety-like and depression-like behavior. These alterations are parallel with molecular changes in the limbic brain, including diminished NCAM sialylation in the PFC. Together with our previous results, the current observations suggest that a chronic social isolation model may potentially be used to study molecular mechanisms that underlie depressive symptomatology.     

Halothane anaesthesia reverses the neuroprotective effect of ketamine against ibotenic acid toxicity in the rat hippocampus

The neurodegenerative effect of ibotenic acid injected into the rat hippocampus was unaffected by the anaesthetics halothane or pentobarbital, apart from a trend to an increased toxicity at higher doses of pentobarbital (60-72 mg/kg). Its toxicity was substantially blocked only by high anaesthetic doses of the indirect acting N-methyl-D-aspartic acid antagonist, ketamine (150-180 mg/kg, i.p.). This is in contrast to its previous reported ability to protect at low concentrations in vitro. On the other hand, the protective effect of ketamine was modified under halothane or pentobarbital anaesthesia. Thus, under halothane anaesthesia, ketamine at 60 mg/kg, i.p. caused a large increase in ibotenic acid-induced neuronal death.     

Lithium attenuates the proconvulsant effect of adolescent social isolation stress via involvement of the nitrergic system

In this study, we tested whether acute administration of lithium mitigates the deleterious effect of adolescent social isolation stress (SIS) on seizure susceptibility. In comparison with socially conditioned (SC) mice, isolated conditioned (IC) mice exhibited an increase in seizure susceptibility to pentylenetetrazole. Acute administration of lithium (10 mg/kg) reversed the proconvulsant effect of SIS in IC mice, but this effect was not observed in SC mice. Coadministration of subthreshold doses of lithium (3 mg/kg) with nitric oxide synthase (NOS) inhibitors reversed the effect of SIS on seizure susceptibility and decreased hippocampal nitrite levels in IC animals. In addition, a subthreshold dose of a nitric oxide precursor reduced the protective effect of lithium on seizure susceptibility and increased nitrite levels in the hippocampus of IC mice. These results suggest that lithium exerts a protective influence against the proconvulsant effect of adolescent SIS via a nitrergic system that includes activation of neuronal NOS in the hippocampus.     

Region-specific response of the hippocampus to chronic unpredictable stress

The objective of the present study was to determine whether chronic unpredictable stress (CUS) would induce hippocampal neuroplasticity in a region-specific manner. Recent evidence suggests that the hippocampus has two functionally distinct subsections. The dorsal (septal) portion appears to be primarily associated with spatial navigation, while the ventral (temporal) region has been linked to affect-related functions, such as anxiety. Chronic stress has previously been shown to negatively affect the hippocampus by decreasing survival of progenitor cells, although it has also been shown to increase adaptive responses, such as increased expression of neuropeptide Y (NPY) and ΔFosB. Whether such events occur in a region-specific manner has not been investigated. We hypothesized that CUS would selectively impact cell survival, NPY, and ΔFosB expression in the more affect-related ventral subregion. Individually housed Long-Evans rats (n = 31) were divided into two groups: stressed and control. Stressed animals were exposed daily to an unpredictable schedule of ethologically relevant stressors, such as predator odors, forced swim, and open field exposure. All rats were injected with bromodeoxyuridine (BrdU) daily during the first 5 days of CUS in order to label dividing progenitor cells. Unbiased stereology was used to quantify BrdU+, NPY+, and ΔFosB+ cells in dorsal and ventral hippocampal subregions. In support of our hypothesis, we found that CUS selectively decreased cell survival in the ventral subregion. However, both NPY and ΔFosB were significantly increased only in the dorsal hippocampus. These results suggest that stress-induced adaptive neuroplasticity occurs primarily in the dorsal subregion, which may coincide with behavioral aspects of the stress response, such as avoidance or amelioration of the stressor.