Pretreatment with Rhodiola Rosea Extract Reduces Cognitive Impairment Induced by Intracerebroventricular Streptozotocin in Rats: Implication of Anti-oxidative and Neuroprotective Effects

Objective To investigate the pretreatment effects of Rhodiola rosea （R. rosea） extract on cognitive dysfunction, oxidative stress in hippocampus and hippocampal neuron injury in a rat model of Alzheimer＇s disease （AD）. Methods Male Sprague-Dawley rats were pretreated with R. rosea extract at doses of 1.5, 3.0, and 6.0 g/kg for 3 weeks, followed by bilateral intracerebroventricular injection with streptozotocin （1.5 mg/kg） on days 1 and 3. Behavioral alterations were monitored after 2 weeks from the lesion using Morris water maze task. Three weeks after the lesion, the rats were sacrificed for measuring the malondialdehyde （MDA）, glutathione reductase （GR） and reduced glutathione （GSH） levels in hippocampus and histopathology of hippocampal neurons. Results The MDA level was significantly increased while the GR and GSH levels were significantly decreased with striking impairments in spatial learning and memory and severe damage to hippocampal neurons in the model rat induced by intracerebroventricular injection of streptozotocin. These abnormalities were significantly improved by pretreatment with R. rosea extract （3.0 g/kg）. Conclusion R. rosea extract can protect rats against cognitive deficits, neuronal injury and oxidative stress induced by intracerebroventricular injection of streptozotocin, and may be used as a potential agent in treatment of neurodegenerative diseases such as AD.     

γ-secretase inhibitor DAPT prevents neuronal death and memory impairment in sepsis associated encephalopathy in septic rats

Background Brain dysfunction is a frequent complication of sepsis,usually defined as sepsis-associated encephalopathy （SAE）.Although the Notch signaling pathway has been proven to be involved in both ischemia and neuronal proliferation,its role in SAE is still unknown.Here,the effect of the Notch signaling pathway involved γ-secretase inhibitor DAPT on SAE in septic rats was investigated in a cecal ligation and puncture （CLP） model.Methods Fifty-nine Sprague-Dawley rats were randomly divided into four groups,with the septic group receiving the CLP operation.Twenty-four hours after CLP or sham treatment,rats were sacrificed and their hippocampus was harvested for Western blot analysis.TNF-αexpression was determined using an enzyme-linked immunosorbent assay （ELISA） kit.Neuronal apoptosis was assessed by TUNEL staining,and neuronal cell death was detected by H＆E staining.Finally,a novel object recognition experiment was used to evaluate memory impairment.Results Our data showed that sepsis can increase the expression of hippocampal Notch receptor intracellular domain （NICD） and poly （adenosine diphosphate [ADP]-ribose） polymerase-1 （PARP-1）,as well as the inflammatory response,neuronal apoptosis,neuronal death,and memory dysfunction in rats.The γ-secretase inhibitor N-[N-（3,5-difluorophenacetyl）-1-alanyl]-S-phenylglycine t-butyl ester （DAPT） can significantly decrease the level of NICD and PARP-1,reduce hippocampal neuronal apoptosis and death,attenuate TNF-α release and rescue cognitive impairment caused by CLP.Conclusion The neuroprotective effect of DAPT on neuronal death and memory impairment in septic rats,which could be a new therapeutic approach for treating SAE in the future.     

Effects of chronic multiple stress on learning and memory and the expression of Fyn, BDNF, TrkB in the hippocampus of rats

Background The effect of chronic stress on cognitive functions has been one of the hot topics in neuroscience. But there has been much controversy over its mechanism. The aim of this study was to investigate the effects of chronic multiple stress on spatial learning and memory as well as the expression of Fyn, BDNF and TrkB in the hippocampus of rats. Methods Adult rats were randomly divided into control and chronic multiple stressed groups. Rats in the multiple stressed group were irregularly and alternatively exposed to situations of vertical revolution, sleep expropriation and restraint lasting for 6 weeks, 6 hours per day with night illumination for 6 weeks. Before and after the period of chronic multiple stresses, the performance of spatial learning and memory of all rats was measured using the Morris Water Maze （MWM）. The expression of Fyn, BDNF and TrkB proteins in the hippocampus was assayed by Western blotting and immunohistochemical methods. The levels of Fyn and TrkB mRNAs in the hippocampus of rats were detected by RT-PCR technique. Results The escape latency in the control group and the stressed group were 15.63 and 8.27 seconds respectively. The performance of spatial learning and memory of rats was increased in chronic multiple stressed group （P〈0.05）. The levels of Fyn, BDNF and TrkB proteins in the stressed group were higher than those of the control group （P〈0.05）. The results of immunoreactivity showed that Fyn was present in the CA3 region of the hippocampus and BDNF positive particles were distributed in the nuclei of CA1 and CA3 pyramidal cells as well as DG granular cells. Quantitative analysis indicated that level of Fyn mRNA was also upregulated in the hippocampus of the stressed group （P〈0.05）. Conclusions Chronic multiple stress can enhance spatial learning and memory function of rats. The expression of Fyn, BDNF and TrkB proteins and the level of Fyn mRNA are increased in the stessed rat hippocampus. These suggest that Fyn and BDNF/TrkB signal transduction pathways may participate in the process of the enhanced learning and memory durina chronic multiple stress.     

Icariside II，a Broad-spectrum Anti-cancer Agent， Reverses Beta-amyloid-induced Cognitive Impairment through Reducing Inflammation and Apoptosis in Rats

Background：Beta-amyloid （Aβ） deposition,associated neuronal apoptosis and neuroinflammation are considered as important factors leading to cognitive deficits in Alzheimer’s disease（ AD）. Icariside Ⅱ（ ICS Ⅱ）,an active fl avonoid compound derived from Epimedium brevicornum Maxim,has been extensively used to treat erectile dysfunction,osteoporosis and dementia. Recently,ICS Ⅱ attracts great interest due to its broad-spectrum anti-cancer property. ICS Ⅱ shows an antiinflammatory potential both in cancer treatment and cerebral ischemia-reperfusion. It is not yet clear whether the anti-inflammatory effect of ICS Ⅱ could delay progression of AD. Therefore,the current study aimed to investigate the effects of ICS Ⅱ on the behavioral deficits,Aβ levels,neuroinfl ammatory responses and apoptosis in Aβ25-35 treated rats. Methods：Rats subjected to bilateral hippocampal injection of Aβ25-35 or normal saline were administered with ICS Ⅱ 20 mg·kg-1 or vehicle once a day for consecutive 15 days. Learning and memory function was evaluated using Morris water maze. The neuronal morphology in hippocampus was examined by HE staining and Nissl staining,respectively. Neuronal apoptosis was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling （TUNEL） assay. The activation of microglia and astrocytes were analyzed by immunohistochemistry. The expression of interleukin-1β（ IL-1β）,cyclooxygenase-2（COX-2）,tumor necrosis factor-α（ TNF-α） and inducible nitric oxide synthase（ iNOS） were detected by quantitative real-time polymerase chain reaction（ qRT-PCR） and Western blot. The expression of caspase-3,Bax and Bcl-2 and the content of beta-amyloid were measured by Western blot. Results：Rats treated with Aβ25-35 displayed cognitive impairment,neuronal damage,along with the increase of Aβ,infl ammation and apoptosis in the hippocampus. However,treatment with ICS Ⅱ 20 mg·kg-1 could improve the cognitive defi cits,ameliorate neuronal death,and reduce the levels of Aβ in the hippocampus. Furthermore,ICS Ⅱ could suppress microglial and astrocytic activation,inhibit expression of IL-1b,TNF-a,COX-2,and iNOS mRNA and protein,and attenuate the Aβ induced Bax/Bcl-2 ratio elevation and caspase-3 activation. Conclusions：These fi ndingssuggest that ICS Ⅱ couldreverse Aβ-induced cognitive deficits,possibly via the inhibition of neuroinfl ammation and apoptosis,which suggested a potential protectiveeffect of ICS Ⅱ on AD.     

Ginsenoside Rb1 Attenuates Isoflurane/surgery-induced Cognitive Dysfunction via Inhibiting Neuroinflammation and Oxidative Stress

Objective Anesthetic isoflurane plus surgery has been reported to induce cognitive impairment. The underlying mechanism and targeted intervention remain largely to be determined. Ginsenoside Rb1 was reported to be neuroprotective. We therefore set out to determine whether ginsenoside Rb1 can attenuate isoflurane/surgery-induced cognitive dysfunction via inhibiting neuroinflammation and oxidative stress. Methods Five-months-old C57BL/6J female mice were treated with 1.4% isoflurane plus abdominal surgery for two hours. Sixty mg/kg ginsenoside Rb1 were given intraperitoneally from 7 days before surgery. Cognition of the mice were assessed by Barnes Maze. Levels of postsynaptic density-95 and synaptophysin in mice hippocampus were measured by Western blot. Levels of reactive oxygen species, tumor necrosis factor-α and interleukin-6 in mice hippocampus were measured by ELISA. Results Here we show for the first time that the ginsenoside Rb1 treatment attenuated the isoflurane/surgery-induced cognitive impairment. Moreover, ginsenoside Rb1 attenuated the isoflurane/surgery-induced synapse dysfunction. Finally, ginsenoside Rb1 mitigated the isoflurane/surgery-induced elevation levels of reactive oxygen species, tumor necrosis factor-α and interleukin-6 in the mice hippocampus. Conclusion These results suggest that ginsenoside Rb1 may attenuate the isoflurane/surgery-induced cognitive impairment by inhibiting neuroinflammation and oxidative stress pending future studies.     

Effect of insulin on the cognizing function and expression of hippocampal Aβ1-40 of rat with Alzheimer disease

Background A model of simulated Alzheimer＇s disease （AD） induced by aggregated amyloid protein （Aβ1-40） was built in Wister rats to observe the behavioral and pathological changes of Aβ1-40 and the effect of hypodermic insulin injected on the function of study and memory and the expression of Aβ1-40 from the CA1 area of the hippocampus. Methods Experimental groups were as follows： contrast, simulated AD model, contrast of Nacl, and insulin treated. The simulated AD model was built by microinjection of aggregated Aβ1-40 at the CA1 area of the hippocampus, and was hypodermically injected with 0.9% NaCI （1 ml/kg） and insulin （0.1 U/kg） separately the next day. Two weeks after the modeling, the four groups were tested with water maze about the study and memory function of rats. Three weeks after the injection, the expression of Aβ1-40 at the CA1 area of the hippocampus was examined by pathological tests （HE, Congo red） and immunohistochemical methods. Results The study and memory abilities of rats were ameliorated significantly by the place navigation test and the spatial probe test after the application of insulin. Insulin could decrease the expression of Aβ1-40 at the CA1 area of the hippocampus to reduce the pathological damage of Aβ1-40 to the hippocampal area of rats. Conclusions The injection of aggregated Aβ1-40 to the hippocampal area could simulate the behavioral and pathological features of AD such as the difficulty of study and memory and the damage to neurons. Insulin is effective to improve the function of study and memory and amend the pathological damage of simulated AD model rats. The results give a experimental proof of insulin in the clinical treatment of AD.     

Bone Marrow-Derived Mesenchymal Stem Cells Incubated with Tanshinone IIA Aattenuates Learning and Memory Impairment Induced by β-amyloid and its Underlying Mechanisms in Rats

Objective：The present study was designed to investigate the neuroprotective effect of mesenchymal stem cells （MSC） which incubated with tanshinone IIA on β-amyloid （Aβ）-induced learning and memory impairment in rats,and further explore the underlying potential mechanisms. Methods：60 healthy male SD rats were randomly divided into 4 groups：sham-operated group,model group,MSC group,MSC+tanshinone group IIA（ n=15）.The rats of model group were injected into the hippocampal with aggregation of 5 μL Aβ25-35（ 2 μg·μL-1） to establish the model of learning and memory impairment. 1 week after surgery,the MSC were injected into the hippocampus,while the rats of model group were injected with volume-matched DMEM,instead. 2 weeks after surgery, Morris water maze （MWM） test was applied to evaluate spatial learning and memory ability of rats,then the rats were sacrifi ced. The survival status of hippocampal neurons was detected by HE and Tunel staining;the formation of hippocampal senile plaques was detected by thiofl avin S staining;the protein expression of Aβ1-42,p-Tau and AMPK in mouse hippocampus were detected by Western blot,respectively;the expression of AMPK mRNA was detected by PCR and the activity of SOD,T-AOC,GSH-PX and content of MDA in plasma by the kits.Results：Compared with sham-operated group, the mean escape latency was markedly increased and the time percentage in target quadrant showed notable decrease in model group;large number of the formation of senile plaque in the hippocampus was detected;the number of surviving neurons in hippocampus was signifi cantly decreased and the apoptosis increased;the protein expression of Aβ1-42 and p-Tau were increased;the mRNA expression and protein levels of AMPK was decreased;the activity of SOD,T-AOC,GSH-PX were decreased and content of MDA increased in plasma.However,compared with model group,the escape latency of MSC group and MSC+Tanshinone group rats was shorter,time percentage in the target quadrant and target quadrant frequency were markedly increased;moreover,the formation of senile plaque in the hippocampus was decreased;the number of surviving neurons in hippocampus was signifi cantly increased;the protein expression of Aβ1-42 and p-Tau were decreased;the mRNA expression and protein levels of AMPK was increased;the activity of SOD,T-AOC,GSH-PX were increased and content of MDA decreased in plasma.And MSC+tanshinone group was more significant than the
MSC group. Conclusion：Under the experimental conditions,MSC incubated with tanshinone ⅡA signifi cantly ameliorates spatial learning and memory impairment and reduce Aβ aggregation and tau protein phosphorylation levels induced by Aβ25-35 in rats,and its potential mechanism may be related
toanti-oxidative stress.     

Eiectroacupuncture Prevents Cognitive Deficits Caused by Topiramate （TPM） in Rats

Objective Recent studies have demonstrated that acupuncture is feasible to treat cognitive impairments. The objective of this study was to present behavioral evidence that electro-acupuncture （EA） could improve the learning and memory of TPM administrated rats. Methods The pattern of TPM-induced cognitive deficits in rats was made by administration of TPM intragastrically for 3 weeks. Of them the rats which showed damage in learning and memory （n=45） were randomly allocated to 3 groups： Impaired group （n=15）, EA group （n=15） and placebo-EA group （n=15）. Moreover, normal group （n=15） was set as control groups. EA stimulation was provided at acupoints located in either the midline of the back or of the head： Gv-20 （Baihui）, Gv-14 （Dazhui）. Morris water maze test was employed to assess spatial discriminative ability per group respectively and to analyze the curative effects of EA. Results Compared to the normal group, obvious cognitive deficits were found in the impaired and placebo-EA groups, and the statistic analysis showed that there were significant differences between normal and impaired groups in ANOVA. In the EA group, shortened mean escape latency was detected compared with the impaired effect on cognitive group during the same trial days; search strategy changed from random pattern adopted by impaired and placebo-EA rats to tendency or linear pattern popular in the normal group. Conclusion The present results suggested that EA exerted a protective impairment caused by TPM in rats, and EA has a specificity of cure. EA as a potential clinic method in treating TPM-induced cognitive impairment should be developed and investigated in the future.     

Preventive Effects of Rhodosin and Melatonin from Damage Induced by β-Amyloid1-40 in Senile Rats

Objective: To study the protective and therapeutic effects of Rhodosin and Melatonin on Alzheimers disease(AD) rats. Methods: D-galactose was intraperitoneally injected in rats for 6 weeks and β-Amyloid1-40(Aβ1-40) was injected into bilateral hippocampus to make AD models. Rhodosin and Melatonin were intraperitoneally injected in rats for 4 weeks to determine the protective and therapeutic effects on rats with AD. Y-maze test, and passive avoidance task were used to determine the ability of learning and memory. The content of lipofuscin in the central cortex, the viscous coefficient of mitochondrial membrane, the activity of superoxide dismutase and the content of malondialdehyde in both sides of hippocampus were determined. And the apoptosis of hippocampus neurons was determined with transmission electron microscopy(TEM). Results: Melatonin as an antioxidant significantly improved learning and memory deficits in the rats with AD and reduced the increase in SOD, MDA, the viscous coefficient and lipofuscin to their normal levels, and it also showed the protective effects of apoptosis. Rhodosin showed the similar effects. Conclusion: Rhodosin and Melatonin had preventive and therapeutic effects on rats with AD probably by affecting the free radical levels in rats.     

Potential association of lead exposure during early development of mice with alteration of hippocampus nitric oxide levels and learning memory

Objective Chronic lead （Pb） exposure during development is known to produce learning deficits. Nitric oxide participates in the synaptic mechanisms involved in certain forms of learning and memory. This study was designed to clarify whether Pb-induced impairment in learning and memory was associated with the changes of nitric oxide levels in mice brains. Methods Sixty Balb/c mice aged l0 days were chosen. A model of lead exposure was established by drinking 0.025%, 0.05% 0.075% lead acetate, respectively for 8 weeks. The controls were orally given distilled water. The ability to learn and memorize was examined by open field test, T-water maze test. In parallel with the behavioral data, NO level of hippocampus tissue was detected by biochemical assay. Results Compared with control groups, （1） the weight of 0.075% group was significantly reduced （P〈0.05）; （2） The number of times in mice attaining the required standards in T-water maze test was lower in 0.075% group （P〈0.01）. No significant difference was found between experimental and control groups in open field test （P〉0.05）; （3） NO level of mouse hippocampus tissue was decreased in 0.075% group （P〈0.01）. Conclusions The findings suggest that decreased hippocampus NO level may contribute to the Pb-induced deficits in learning and memory processes.     

Preventive Effect of Genetic Knockdown and Pharmacological Blockade of CysLT1R on Lipopolysaccharide（LPS）-induced Memory Deficit and Neurotoxicity in vivo

The aim of the current study was to fi nd out the role of CysLT1R on lipopolysaccharide（LPS）-induced cognitive deficit and neurotoxicity. shRNA-mediated knockdown or pharmacological blockade（ by pranlukast） of CysLT1R were performed in ICR mice for 21 days prior to systemic infusion of LPS. From day 22,LPS was administered for 7 days and then a set of behavioral,histopathological and biochemical tests were employed to test memory neuroinflammation and apoptotic responses in the mouse hippocampus. LPS （only）-treated mice showed poor performance in both Morris water maze（MWM） and Y-maze tests. However,shRNA-mediated knockdown or pranlukast-treated blockade of CysLT1R improved performance of the mice in these tests. To find out the possible underlying mechanisms,we assessed several parameters such as microglial activation（ by immunohistochemistry）,level of CysLT1R （by WB and qRT-PCR） and the inflammatory/apoptotic pathways （by ELISA or TUNEL or WB） in the mouse hippocampus. LPS-induced memory impairment was accompanied by activation of microglia,higher levels of CysLT1R,IL-1β,TNF-α and nuclear NF-κB p65. LPS also caused apoptosis in the hippocampus as detected by TUNEL staining,further supplemented by detection of increased Caspase-3 and a reduced Bcl-2/Bax ratio. All of these adverse changes in the mouse hippocampus were inhibited by pretreatment with CysLT1R-shRNA and pranlukast. Through this study we suggest that CysLT1R shares a strong correlation with LPS-associated memory defi cit,neuroinflammation and apoptosis and CysLT1R could be a novel target for preventive measures to intervene the progression of Alzheimer’s disease（ AD）-like phenotypes.     

Therapeutic benefit of Yangxue Qingnao Granule (养血清脑颗粒) on cognitive impairment induced by chronic cerebral hypoperfusion in rats

Objective:To observe the therapeutic effect of Yangxue Qingnao Granule（养血清脑颗粒, YXQNG） on cognitive impairment induced by chronic cerebral hypoperfusion and to investigate its impact on oxidative stress,apoptosis,and the cholinergic system.Methods:Adult male Wistar rats were subjected to chronic cerebral hypoperfusion by permanent occlusion of bilateral common carotid arteries（2-VO）.Thirty rats were randomly assigned to one of the five treatment groups in a 1:1:1:1:1 ratio:sham operation plus normal saline treatment,2-VO plus normal saline treatment,2-VO plus YXQNG at a dose of 2 g·kg（-1）·d^-1 or 4 g·kg（-1）·d^-1, or 2-VO plus rivastigmine 2 mgkg^-1·d^-1.The Morris water maze test was used to assess the spatial memory retrieval.Apoptosis,total antioxide capacity（T-AOC）,acetylcholine esterase（AchE） and choline acetyl transferase（ChAT） activities in the hippocampus and the cortex were investigated.Results:In the chronic cerebral hypoperfusion model,the 2-VO plus saline treatment resulted in impaired special learning as shown by the significantly prolonged escape latency and shorter swim time in the first quadrant as compared to the sham operation.The impairment was associated with apoptosis and significant decreases in T-AOC,AchE and ChAT activities in the hippocampus and the cortex.Treatment with YXQNG at either 2 g·kg（-1）·d^-1 or 4 g·kg（-1）·d^-1 dose,or rivastigmine resulted in significantly shorter escape latencies and longer swim time in the first quadrant.YXQNG at both doses,but not rivastigmine,had significant reduction in apoptosis,and significant increases in T-AOC and ChAT activity in both the hippocampus and the cortex.Unlike rivastigmine,neither dose of YXQNG showed significant reduction in AchE activity.Conclusions:YXQNG ameliorated cognitive impairment induced by chronic cerebral hypoperfusion.The protective effect may be mediated through its regulation of apoptosis and activities of T-AOC and ChAT in the hippocampus and cortex of the rats in the chronic cerebral hypoperfusion model,a mechanism that is different from rivastigmine.     

Altered angiotensin-converting enzyme and its effects on the brain in a rat model of Alzheimer disease

Background Alzheimer disease （AD） is a neurodegenerative disease related to aging. At present, its pathological mechanisms remain unclear. Family members of the renin-angiotensin system （RAS） play a role in neuronal plasticity, as well as formation of learning and memory. In this study, we explore the effects of altered angiotensin-converting enzyme （ACE）, and investigate the possible mechanisms of perindopril, an ACE inhibitor, on brain structure and function in a rat model of AD, as well as the role that ACE plays in AD. Methods Sixty Sprague-Dawley rats were selected and randomly divided into 3 groups： control, AD, and perindopril. Each group consisted of 20 rats, with 10 rats for determining pathology, and the remaining 10 rats for quantifying ACE activity. The rat AD model was established by stereotactically injecting amyloid beta protein （A-beta） 1-42 into the right hippocampus. Learning and memory functions were tested using the Y-type electric maze. The number and morphology of abnormal neurons were determined by haematoxylin and eosin staining. Amyloid deposition was measured by Congo red staining. Finally, ACE activity was estimated by spectrophotometry. Results Compared with the control group, the number of times needed to escape electrical stimuli increased （23.70±3.13, P 〈0.001）, the number of normal neurons in the CA1 region was reduced （density of 96.5±32.6/mm, P 〈0.001）, amyloid deposition was obvious, and ACE activity increased （（34.4±6.6） nmol.g-1.min-1, P 〈0.001） in the AD group. In the perindopril group, the number of times needed to escape electrical stimuli decreased （18.50±3.66, P 〈0.001）, the number of abnormal neurons increased （density of CA1 neurons was 180.8±28.5/mm, P 〈0.001）, amyloid deposition was reduced, and ACE activity was down-regulated （（26.2±6.2） nmol.g-1.min-1, P 〈0.001）. Conclusions ACE activity increased in the brains of AD rats. Perindopril improved learning and memory in AD rats, which correlated with de     

DHA Depletion in Rat Brain Is Associated With Impairment on Spatial Learning and Memory

Objective To examine the effect of docosahexaenoic acid （DHA） deficiency in brain on spatial learning and memory in rats. Methods Sprague Dawley rats were fed with an n-3 fatty acid deficient diet for two generations to induce DHA depletion in brain, DHA in seven brain regions was analyzed using the gas-liquid chromatography. Morris water maze （MWM） was employed as an assessing index of spatial learning and memory in the n-3 fatty acid deficient adult rats of second generation. Results Feeding an n-3 deficient diet for two generations depleted DHA differently by 39%-63% in the seven brain regions including cerebellum, medulla, hypothalamus, striatum, hippocampus, cortex and midbrain, The MWM test showed that the n-3 deficient rats took a longer time and swam a longer distance to find the escape platform than the n-3 Adq group. Condusion The spatial learning and memory in adult rats are partially impaired by brain DHA depletion.     

Effects of Chronic Administration of Melatonin on Spatial Learning Ability and Long-term Potentiation in Lead-exposed and Control Rats

Objective To explore the changes in spatial learning performance and long-term potentiation （LTP） which is recognized as a component of the cellular basis of learning and memory in normal and lead-exposed rats after administration of melatonin （MT） for two months. Methods Experiment was performed in adult male Wistar rats （12 controls, 12 exposed to melatonin treatment, 10 exposed to lead and 10 exposed to lead and melatonin treatment）. The lead-exposed rats received 0.2% lead acetate solution from their birth day while the control rats drank tap water. Melatonin （3 mg/kg） or vehicle was administered to the control and lead-exposed rats from the time of their weaning by gastric gavage each day for 60 days, depending on their groups. At the age of 81-90 days, all the animals were subjected to Morris water maze test and then used for extracellular recording of LTP in the dentate gyrus （DG） area of the hippocampus in vivo. Results Low dose of melatonin given from weaning for two months impaired LTP in the DG area of hippocampus and induced learning and memory deficit in the control rats. When melatonin was administered over a prolonged period to the lead-exposed rats, it exacerbated LTP impairment, learning and memory deficit induced by lead. Conclusion Melatonin is not suitable for normal and lead-exposed children.     

Effects of Ning Shen Ling Granule (宁神灵冲剂) and dehydroepiandrosterone on cognitive function in mice undergoing chronic mild stress

Objective： To investigate the changes of spontaneous and cognitive behavior, and cholinergic M receptors in the brain of mice subjected to chronic mild stress （CMS）, and to determine the effect of Ning Shen Ling Granule （宁神灵冲剂, NSL） and dehydroepiandrosterone （DHEA） on them. Methods：CMS model mice were established by applying stress every day for 3 consecutive weeks with 7 kinds of unforeseeable stress sources, and they were medicated for 1 week beginning at the 3rd week of modeling. The changes in behavior were determined by Morris Water Maze and spontaneous movement test, and M-receptor binding activity in cerebral cortex, hippocampus and hypothalamus were measured by radioactive ligand assay with 3H-QNB. Results： （1） The spontaneous movement in CMS model mice was significantly reduced, with the latency for searching platform in Morris Water Maze obviously prolonged （P〈0.01）, and these abnormal changes in behavior were improved in those treated with NSL and DHEA. （2） The binding ability of M-receptor in cerebral cortex and hippocampus of CMS mice was significantly decreased as compared with those in the control group （P〈0.05）, but could be restored to the normal level after intervention with NSL or DHEA. Conclusion： The decline of spontaneous movement and spatial learning and memory ability could be induced in animals by chronic mild stress, and that may be related to the low activity of central cholinergic M-receptors. Both NSL and DHEA could effectively alleviate the above-mentioned changes.     

Increased Expression and Altered Subcellular Distribution of Cathepsin B in Microglia Induces Cognitive Impairment through Oxidative Stress and Inflammatory Response in Mice

Objective：During normal aging，innate immunity progresses to a chronic state. However，how oxidative stress and chronic neuroinflammation arise during aging remain unclear. In this study，we intended to demonstrate the novel function of Cathepsin B （CatB） in the age related oxidative stress and chronic neuroinfl ammation. Methods：①2 month-old and 20 month-old wild-type and CatB-/- mice were used for learning and memory tests by step through passive avoidance tests. The cumulative potentiation of the fi eld excitatory postsynaptic potential（ fEPSP） slope and dendritic spine density were also examined. ②Infl ammation were examined by immunoblotting of interleukin-1β（ IL-1β），tumor necrosis factor-α（ TNF-α） and inducible nitric oxide synthase（ iNOS），and oxidative stress were examined by checking the level of 8-oxo-7，8-dihydro-2’-deoxyguanosine （8-oxo-dG） and 4-hydroxynonenal（ 4-HNE） in the hippocampal lysate of 2 month-old and 20 month-old wild-type and CatB-/- mice. ③Intra-lateral ventricle transplantation of CatB over-expressed microglia in middleaged mice，learning and memory were examined by Y-maze and novel objective tests. Results：①The retention latencies of both aged groups were significantly longer than those in the acquisition trial. The retention latency of four consecutive trials was signifi cantly longer in aged CatB-/- mice than in aged wild-type mice. Furthermore，the mean relative fEPSP slope measured at 30 min after stimulationwith 25，50 and 100 Hz in the aged CatB-/- mice was significantly larger than that in the aged wildtype mice. The mean dendritic spine density of CA1 neurons in the aged CatB-/- mice was signifi cantly
larger than that in the aged wild-type mice. ②The mean levels of CatB，iNOS，IL-1β and TNF-α in the hippocampus were signifi cantly higher in aged wild-type mice than in younger animals. Furthermore， their mean levels in the hippocampus of aged CatB-/- mice were significantly lower than those in aged wild-type mice. The mean amounts of these oxidative markers were significantly larger in the hippocampus of aged wild-type mice than in younger animals. The mean relative amount of 8-oxodG and 4-HNE was significantly lower in aged CatB-/- mice than in aged wild-type mice. ③Double immunohistochemical staining was conducted to identify the possible cellular origin of oxidative stress. In the hippocampus of aged wild-type mice，the immunoreactivities of both 8-oxo-dG and 4-HNE were found exclusively in microglia with activated morphology，but not in astrocytes or neurons. ④Intralateral ventricle transplantation of CatB over-expressed microglia signifi cantly impaired the learning and memory in the middle-aged mice. Conclusion：CatB plays a critical role in the mitochondriaderived ROS generation and infl ammatory response，resulting in impairment of learning and memory during normal aging.     

Effects of unpredictable chronic stress on behavior and brain- derived neurotrophic factor expression in CA3 subfield and dentate gyrus of the hippocampus in different aged rats

Background Brain-derived neurotrophic factor （BDNF） is a stress-responsive intercellular messenger modifying hypothalamic-pituitary-adrenal （HPA） axis activity. The interaction between stress and age in BDNF expression is currently not fully understood. This study was conducted to observe unpredictable stress effect on behavior and BDNF expression in CA3 subfield （CA3） and dentate gyrus of hippocampus in different aged rats. Methods Forty-eight Wistar rats of two different ages （2 months and 15 months） were randomly assigned to six groups： two control groups and four stress groups. The rats in the stress group received three weeks of unpredictable mild stress. The depression state and the stress level of the animals were determined by sucrose preference test and observation of exploratory behavior in an open field （OF） test. The expressions of BDNF in CA3 and dentate gyrus of the hippocampus were measured using immunohistochemistry. Results Age and stress had different effects on the behavior of different aged animals （age： F=6.173, P〈0.05, stress： F=6.056, P 〈0.05）. Stress was the main factor affecting sucrose preference （F=123.608, P 〈0.05）. Decreased sucrose preference and suppressed behavior emerged directly following stress, lasting to at least the eighth day after stress in young animals （P 〈0.05）. The older stress rats showed a lower sucrose preference than young stress rats （P 〈0.05）. Older control rats behaved differently from the younger control animals in the OF test, spending more time in the central square （P 〈0.05）, exhibiting fewer vertical movements （P 〈0.05） and less grooming （P 〈0.05）. Following exposure to stress, older-aged rats showed no obvious changes in vertical movement and grooming. This indicates that aged rats were in an unexcited state before the stress period, and responded less to stressful stimuli than younger rats. There was significantly lower BDNF expression in the CA3 and dentate gyrus regions of the hippocampus following stress in both age groups （P〈0.05）, a reduction that was still present at the eighth day after stress （P〈0.05）. Stress and age were the main factors affecting the expression of BDNF （F=9.408, P 〈0.05; F=106.303, P 〈0.05）. The aged stress group showed lower BDNF expression compared to the young stressed group at every testing time point. Conclusion Stress has age-dependent effects on behavioral responses and hippocampal BDNF expression in rats.