BACKGROUD: Ethanol can influence neural development and the ability of learning and memory, but its mechanism of the neural toxicity is not clear till now. Endogenous nitric oxide (NO) as a gaseous messenger is proved...BACKGROUD: Ethanol can influence neural development and the ability of learning and memory, but its mechanism of the neural toxicity is not clear till now. Endogenous nitric oxide (NO) as a gaseous messenger is proved to play an important role in the formation of synaptic plasticity, transference of neuronal information and the neural development, but excessive nitro oxide can result in neurotoxicity. OBJECTIVE: To observe the effects of acute alcoholism on the learning and memory ability and the content of neuronal nitric oxide synthase (nNOS) in brain tissue of rats. DESIGN: A randomized controlled animal experiment. SETTING: Department of Physiology, Xinxiang Medical College. MATERIALS: Eighteen male clean-degree SD rats of 18-22 weeks were raised adaptively for 2 days, and then randomly divided into control group (n = 8) and experimental group (n = 10). The nNOS immunohistochemical reagent was provided by Beijing Zhongshan Golden Bridge Biotechnology Co.,Ltd. Y-maze was produced by Suixi Zhenghua Apparatus Plant. METHODS: The experiment was carried out in the laboratory of the Department of Physiology, Xinxiang Medical College from June to October in 2005. ① Rats in the experimental group were intraperitoneally injected with ethanol (2.5 g/kg) which was dissolved in normal saline (20%). The loss of righting reflex and ataxia within 5 minutes indicated the successful model. Whereas rats in the control group were given saline of the same volume. ② Examinations of learning and memory ability: The Y-maze tests for learning and memory ability were performed at 6 hours after the models establishment. The rats were put into the Y-maze separately. The test was performed in a quiet and dark room. There was a lamp at the end of each of three pathways in Y-maze and the base of maze had electric net. All the lamps of the three pathways were turned on for 3 minutes and then turned off. One lamp was turned on randomly, and the other two delayed automatically. In 5 seconds after alternation, pulsating electric current presented in the base of unsafe area to stimulate rat’s feet to run to the safe area. The lighting lasted for 15 seconds as one test. Running from unsafe area to safe area at one time in 10 seconds was justified as successful. Such test was repeated for 10 times for each rat and the successful frequency was recorded. The qualified standard of maze test was that the rat arrived in the safe area 9 times during 10 experiments. The number of trainings for the qualified standard was used to represent the result of spatial learning. ③ Determination of the content of nNOS in brain tissue: After the Y-maze test, the rats were anaesthetized, and blood was let from the incision on right auricle, transcardially perfused via the left ventricle with about 200 mL saline, then fixed by perfusion of 40 g/L paraformaldehyde. Hippocampal CA1 region, corpus striatum and cerebellum were taken to prepare serial freezing coronal sections. The nNOS contents in the brain regions were determined with the immunohistochemical methods to reflect the changes of nitric oxide in brain tissue. MAIN OUTCOME MEASURES: The changes of learning and memory ability and the changes of the nNOS contents in the brain tissue of rats with acute alcoholism were observed. RESULTS: One rat in the experimental group was excluded due to its slow reaction to electric stimulation in the Y-maze test, and the other 17 rats were involved in the analysis of results. ① The training times to reach qualifying standards of Y-maze in the experimental group was more than that in the control group [(34.33 ±13.04), (27.50±8.79) times, P < 0.05]. ② Forms and numbers of nNOS positive neurons in brain tissue: It could be observed under light microscope that in the hippocampal CA1 region, there were fewer nNOS positive neurons, which were lightly stained, and the processes were not clear enough; But the numbers of the positive neurons which were deeply stained as buffy were obviously increased in the experimental group, the cell body and cytoplasm of process were evenly stained, but the nucleus was not stained. The nNOS positive neurons in corpus striatum had similar forms and size in the experimental group and control group. The form of the nNOS positive neurons in cerebellum were similar between the two groups. The numbers of nNOS positive neurons in hippocampal CA1 region and corpus striatum in the experimental group [(18.22±7.47), (11.38±5.00) cells/high power field] were obviously higher than those in the control group [(10.15±4.24), (6.15±3.69) cells/high power field. The number of nNOS positive neurons in cerebellum had no significant difference between the two groups [(49.56±18.84), (44.43±15.42) cells/high power field, P > 0.05]. CONCLUSION: Acute alcoholism may impair learning and memory ability, and nitric oxide may be involved in mediating the neurotoxic role of ethanol.展开更多
BACKGROUND: Central adrenergic nerve and 5-serotonergic nerve can influence central cholinergic nerve on learning and memory and make easy for study; however, ginsenoside of stem and leaf (GSL) can improve functions o...BACKGROUND: Central adrenergic nerve and 5-serotonergic nerve can influence central cholinergic nerve on learning and memory and make easy for study; however, ginsenoside of stem and leaf (GSL) can improve functions of central adrenergic nerve; moreover, 5-serotonergic nerve and the combination with choline can produce synergistic effect and enhance learning and memory ability so as to improve learning and memory disorder of patients with Alzheimer disease (AD). OBJECTIVE: To observe the effects of GSL combining with choline on learning and memory of AD model rats.DESIGN: Randomized grouping design and controlled animal study.SETTING: Department of Pharmacology, Taishan Medical College. MATERIALS: The experiment was carried out in the Pharmacological Department of Medical College of Jilin University from October 1996 to January 1997. Forty healthy male Wistar rats of clean grade were randomly divided into 5 groups, including sham-injury group, model group, GSL group, choline group and combination group, with 8 rats in each group. Main medications: GSL with the volume more than 92.8% was provided by Department of Chemistry, Norman Bethune Medical College of Jilin University. Panaxatriol, the main component, was detected with thin layer scanning technique and regarded as the index of GSL quality [(55±1)%, CV = 2%, n = 5]. Choline was provided by the Third Shanghai Laboratory Factory. METHODS: 150 nmol quinolinic acid was used to damage bilateral Meynert basal nuclei of adult rats so as to establish AD models. Rats in GSL, choline and combination groups were intragastric administrated with 400 mg/kg GSL, 200 mg/kg choline (20 mL/kg), and both respectively last for 17 days starting from two days before operation. Rats in sham-injury group and model group were perfused with the same volume of distilled water once in each morning for the same days. ① Passive avoidance step-down test: Five minutes later, rats jumped up safe platform when they were shocked with 36 V alternating current. If rats jumped down from the platform and the feet touched railings, the response was wrong. Numbers of wrong response were recorded within 3 minutes, and then the test was redone after 24 hours. ② Morris water-maze spatial localization task: Swimming from jumping-off to platform directly was regarded as right response. Additionally, 4 successively right responses were regarded as the standard. Each rat was trained 10 times a day with 120 s per time for 3 successive days. The interval was 30 s. Three days later, numbers of right response were recorded. The training times were increased to 30 for unlearned rats. ③ Measurement of activity of choilne acetylase in cerebral cortex: Rats were sacrificed at 17 days after operation to obtain cerebral cortex to measure activity of choilne acetylase with radiochemistry technique. ④ Synergistic effect: It was expressed as Q value: Q value = factual incorporative effect/anticipant incorporative effect; Q ≥ 1 was regarded as synergistic effect. Anticipant incorporative effect = (EA+EB-EA·EB), EA and EB were single timing effect, respectively in GSL group and choline group. E (step-down test and Morris water maze test) = (x in model group - factual value in medicine groups)/x in model group; E (activity of choilne acetylase) = (factual value in medicine groups -x in model group)/x in model group. MAIN OUTCOME MEASURES: ① Passive avoidance step-down test and Morris water-maze spatial localization task in the study of learning and memory; ② activity of choilne acetylase. RESULTS: All 40 rats were involved in the final analysis. ① Passive avoidance response: At learning phase on first day and retesting phase on the next day, numbers of wrong responses within 3 minutes were more in model group than sham operation group, and there was significant difference [(5.88±1.46), (2.25±0.87) times; (2.63±1.06), (0.50±0.53) times; P < 0.01]; numbers of wrong responses within 3 minutes were less in combination group than model group, and there was significant difference [learning phase: (1.12±0.83), (5.88±1.46) times; retesting phase: (0.38±0.74), (2.63±1.06) times, P < 0.01]; moreover, effect was stronger than that in GSL group and choline group. The Q value was 1.07 and 1.59, respectively and it showed synergistic effect. ② Spatial localization task: Training times were more in model group than sham operation group, and there was significant difference [(2.9±2.5), (12.6±3.5) times; P < 0.01]. Training times were less in combination group than model group, and there was significant difference [(11.8±2.4), (27.9±2.5) times, P < 0.01]; moreover, effect was stronger than that in GSL group and choline group. The Q value was 1.07 and it showed synergistic effect. ③ Activity of choilne acetylase: Activity was lower in model group than sham operation group, and there was significant difference [(30.56±8.33), (61.11±8.33) nkat/g; P < 0.01]. Activity was higher in combination group than model group and there was significant difference [(50.00±8.33), (30.56±8.33) nkat/g, P < 0.01]; moreover, effect was stronger than that in GSL group and choline group. The Q value was 1.5 and it showed synergistic effect. CONCLUSION: GSL in combination with choline can synergically improve the disorder of learning and memory of AD model rats. Its mechanism may be involved in enhancing the function of central cholinergic system.展开更多
BACKGROUND: Generally speaking, anesthesia is often used in gravid body and it has been already proved that many kind of medicine can result in malformation. OBJECTIVE: To explore embryonic skeleton development and ne...BACKGROUND: Generally speaking, anesthesia is often used in gravid body and it has been already proved that many kind of medicine can result in malformation. OBJECTIVE: To explore embryonic skeleton development and neonatal learning and memory of rats anesthetized with pentobarbital sodium in gravid rats. DESIGN: A randomized control trial. SETTING: Laboratory Animal Center of Xuzhou Medical College. MATERIALS: A total of 80 adult female SD rats, of clean grade and weighing 220-240 g, were selected in this study. The main reagents were detailed as follows: pentobarbital sodium (Shanghai Xingzhi Chemical Plant, batch number: 921019); MG-2 maze test apparatus (Zhangjiagang Biomedical Instrument Factory); somatotype microscope (Beijing Taike Instrument Co., Ltd.). METHODS: ① A total of 160 SD rats of half males and females were selected in this study. All rats were copulated. The day that the plug was checked out in the vagina next day was looked as the first day of pregnancy. Gravid rats were divided randomly into four groups, including early anesthesia group, second anesthesia group, late anesthesia group and control group with 20 in each group. Rats in the early anesthesia group were injected with 25 mg/kg soluble pentobarbitone on the 7th day of pregnancy for once; rats in the second anesthesia group were anesthetized with 25 mg/kg soluble pentobarbitone on the 7th and the 14th days of pregnancy for once; rats in the late anesthesia group were anesthetized with 25 mg/kg soluble pentobarbitone on the 14th day of pregnancy for once; rats in the control group did not treat with anything. The time of anesthetizing was controlled in 3 to 4 hours and ether was absorbed while the time was not enough. ② Half of each group was sacrificed on day 20th of pregnancy and the fetus was taken out to be stained with alizarin red S. After stained, the fetal skeleton was examined. The learning and memorizing of one-month rats that were given birth by the rest gravid rats were tested through electric mare method. Determine their study ability according to their correct rate of 90% or above of arrival at the safe area in 20 s. After they finally learned to arrive at the safe area correctly, test them once more in 24 hours and record the correct rate of 15 times. MAIN OUTCOME MEASURES: The rate of malformation in fetus and ability of learning and memory in one-month rats. RESULTS: A total of 80 female rats were anesthetized in this experiment. Totally 490 immature rats were tested with maze testing machine and 196 fetuses were stained with alizarin red S to observe the development of their skeleton. However, one of the 80 female rats was led to death because of overdose. ① Malformation experiment: Learning ability of second anesthesia group was evidently different from the control group while the other two groups were not in the electric mare method. The fetal skeleton malformation rate of three experimental groups was 87.0%, 60.9% and 17.9%, respectively, while it was 5.6% in the control group. ② Electric mare method: Times of rats which arrived at the safe regions were respectively 49.0±31.0, 68.0±35.0, 47.0±31.0 and 44.0±21.0 in early anesthesia group, second anesthesia group, late anesthesia group and control group; and then, there was significant difference between the second anesthesia group and the control group (P < 0.05). Exact rates of memory of rats were respectively (64.36±14.35)%, (62.15±18.33)%, (54.19±12.28)% and (68.24±15.91)% in early anesthesia group, second anesthesia group, late anesthesia group and control group; and then, there were no significant differences as compared with the control group (P > 0.05). CONCLUSION: The influence of anesthesia with pentobarbital sodium is obvious in fetal skeleton development and learning and memory ability.展开更多
BACKGROUND: Transplantation of fetal cell suspension or blocks of fetal tissue can ameliorate the nerve function after the injury or disease in the central nervous system, and it has been used to treat neurodegenerati...BACKGROUND: Transplantation of fetal cell suspension or blocks of fetal tissue can ameliorate the nerve function after the injury or disease in the central nervous system, and it has been used to treat neurodegenerative disorders induced by Parkinson disease. OBJECTIVE: To observe the effects of the transplantation of neuron-like cells derived from bone marrow stromal cells (rMSCs) into the brain in restoring the dysfunctions of muscle strength and balance as well as learning and memory in rat models of cerebral infarction. DESIGN: A randomized controlled experiment. SETTING: Department of Pathophysiology, Zhongshan Medical College of Sun Yat-sen University. MATERIALS: Twenty-four male SD rats (3-4 weeks of age, weighing 200-220 g) were used (Certification number:2001A027). METHODS: The experiments were carried out in Zhongshan Medical College of Sun Yat-sen University between December 2003 and December 2004. ① Twenty-four male SD rats randomized into three groups with 8 rats in each: experimental group, control group and sham-operated group. Rats in the experiment al group and control group were induced into models of middle cerebral artery occlusion. After in vitro cultured, purified and identified with digestion, the Fischer344 rMSCs were induced to differentiate by tanshinone ⅡA, which was locally injected into the striate cortex (18 area) of rats in the experimental group, and the rats in the control group were injected by L-DMEM basic culture media (without serum) of the same volume to the corresponding brain area. In the sham-operated group, only muscle and vessel of neck were separated. ② At 2 and 8 weeks after the transplantation, the rats were given the screen test, prehensile-traction test, balance beam test and Morris water-maze test. ③ The survival and distribution of the induced cells in corresponding brain area were observed with Nissl stained with toluidine blue and hematoxylin and eosin (HE) staining in the groups. MAIN OUTCOME MEASURES: ① Results of the behavioral tests (time of the Morris water-maze test screen test, prehensile-traction test, balance beam test); ② Survival and distribution of the induced cells. RESULTS: All the 24 rats were involved in the analysis of results. ① Two weeks after transplantation, rats with neuron-like cells grafts in the experimental group had significant improvement on their general muscle strength than those in the control group [screen test: (9.4±1.7), (4.7±1.0) s, P < 0.01]; forelimb muscle strength [prehensile-traction test: (7.6±1.4), (5.2±1.2) s, P < 0.01], ability to keep balance [balance beam test: (7.9±0.74), (6.1±0.91) s, P < 0.01] and abilities of learning and memory [latency to find the platform: (35.8±5.9), (117.5±11.6) s, P < 0.01; distance: (623.1±43.4), (1 902.3±98.6) cm, P < 0.01] as compared with those in the control group. The functional performances in the experimental group at 8 weeks were better than those at two weeks, which were still obviously different from those in the sham-operated group (P < 0.05). ② The HE and Nissl stained brain tissue section showed that there was nerve cell proliferation at the infarcted cortex in the experiment group, the density was higher than that in the control group, plenty of aggregative or scattered cells could be observed at the site where needle was inserted for transplantation, the cells migrated directively towards the area around them, the cerebral vascular walls were wrapped by plenty of cells; In the control group, most of the cortices were destroyed, karyopyknosis and necrosis of neurons were observed, normal nervous tissue structure disappeared induced by edema, only some nerve fibers and glial cells remained. CONCLUSION: The rMSCs transplantation can obviously enhance the motor function and the abilities of learning and memory in rat models of cerebral infarction.展开更多
BACKGROUND: Calcitonin gene-related peptide (CGRP) and nerve growth actor (NGF) cam improve spatial learning and memory abilities of rats with cerebral ischemia/reperfusion; however, the effect of combination of them ...BACKGROUND: Calcitonin gene-related peptide (CGRP) and nerve growth actor (NGF) cam improve spatial learning and memory abilities of rats with cerebral ischemia/reperfusion; however, the effect of combination of them on relieving learning and memory injury following cerebral ischemia/reperfusion should be further studied. OBJECTIVE: To study the effects of exogenous CGRP and NGF on learning and memory abilities of rats with focal cerebral ischemia/reperfusion. DESIGN: Randomized controlled animal study. SETTING: Department of Neurosurgery, the Second Hospital of Xiamen; Department of Neurosurgery, the Second Affiliated Hospital of China Medical University; Department of Neurobiology, Basic Medical College of China Medical University. MATERIALS: A total of 30 healthy male SD rats, aged 8 weeks, of clean grade, weighing 250-300 g, were provided by Experimental Animal Department of China Medical University. All rats were randomly divided into sham-operation group, ischemia/reperfusion group and treatment group with 10 in each group. The main reagents were detailed as the follows: 100 g/L chloral hydrate, 0.5 mL CGRP (2 mg/L, Sigma Company, USA), and NGF (1× 106 U/L, 0.5 mL, Siweite Company, Dalian). METHODS: The experiment was carried out in the Department of Neurobiology, Basic Medical College of China Medical University from February to July 2005. Rat models of middle cerebral artery occlusion were established by method of occlusion, 2 hours after that rats were anesthetized and the thread was slightly drawn out for 10 mm under direct staring to perform reperfusion. Rats in the ischemia/reperfusion group received intraperitoneal injection of 1 mL saline via the abdomen at two hours later, while rats in the treatment group at 2 hours later received intraperitoneal injection of 2 mg/L CGRP (0.5 mL) and 1×106 U/L NGF (0.5 mL) once a day for 10 successive days. First administration was accomplished within 15 minutes after ischemia/reperfusion. Rats in the sham-operation group were separated of the vessels without occlusion or administration. The neural function was evaluated with Zea Longa 5-grade scale. Animals with the score of one, two and three points received Morris water-maze test to measure searching time on platform (omitting platform-escaping latency). Meanwhile, leaning and memory abilities of animals were reflected through testing times of passing through platform per minute. MAIN OUTCOME MEASURES: Experimental results of omitting platform-escaping latency and spatial probe. RESULTS: Three and two rats in the ischemia/reperfusion group and treatment group respectively were not in accordance with the criteria in the process, and the rest were involved in the final analysis. ① Comparisons of platform-escaping latency during Morris water-maze test in all the three groups: Ten days after modeling, the platform-escaping latency in the ischemia/reperfusion group was obviously longer than that in sham-operation group (P < 0.01), and was significantly shorter than that in the treatment group (P < 0.01). ② Comparisons of times of passing through platform in all the three groups: Times of passing through platform were remarkably less in the ischemia/reperfusion group than those in the sham-operation group [(1.79±0.39), (4.30±0.73) times/minute, P < 0.01], and those were markedly more in the treatment group than the ischemia/reperfusion group [(3.16±1.03), (1.79±0.39) times/minute, P < 0.01]. CONCLUSION: CGRP and NGF are capable of ameliorating the abilities of spatial learning and memory in MCAO rats, which indicates that CGRP and NGF can protect ischemic neurons.展开更多
Rhesus monkey neural stem cells are capable of differentiating into neurons and glial cells.Therefore,neural stem cell transplantation can be used to promote functional recovery of the nervous system.Rhesus monkey neu...Rhesus monkey neural stem cells are capable of differentiating into neurons and glial cells.Therefore,neural stem cell transplantation can be used to promote functional recovery of the nervous system.Rhesus monkey neural stem cells(1×105 cells/μL) were injected into bilateral hippocampi of rats with hippocampal lesions.Confocal laser scanning microscopy demonstrated that green fluorescent protein-labeled transplanted cells survived and grew well.Transplanted cells were detected at the lesion site,but also in the nerve fiber-rich region of the cerebral cortex and corpus callosum.Some transplanted cells differentiated into neurons and glial cells clustering along the ventricular wall,and integrated into the recipient brain.Behavioral tests revealed that spatial learning and memory ability improved,indicating that rhesus monkey neural stem cells noticeably improve spatial learning and memory abilities in rats with hippocampal lesions.展开更多
Objective:To observe the effect of the early intervention with moxibustion at "Shènshū(肾俞BL23)" on learning and memory of Alzheimer’s disease(AD) rats modeled by ovariectomy and D-Galactose(D-Gal)in...Objective:To observe the effect of the early intervention with moxibustion at "Shènshū(肾俞BL23)" on learning and memory of Alzheimer’s disease(AD) rats modeled by ovariectomy and D-Galactose(D-Gal)injection and the effect on Ca2+/CaMKII/CREB signaling pathway so as to provide the evidence for the acupoint selection in the early intervention with acupuncture and moxibustion for AD.Methods:Sixty female SD rats at the age of 3 months were selected and randomly divided into a normal group,a sham-operation group,a model group,a "BL23" group,a "Wèishū"(胃俞BL21) group,and a non-acupoint group,10 rats in each group.Except in the normal group and the sham-operation group,bilateral ovaries were removed in the other 4 groups.3 days after the operation,D-Gal was injected intraperitoneally at a dose of 150 mg/kg/d for 90 days to establish AD rat model.In the sham-operation group,only the peripheral fat of ovarian was removed and 0.9% normal saline was injected intraperitoneally of the same dose for 90 days.In the "BL23" group,the "BL21" group and the non-acupoints group,since the 2 nd day after ovariectomy,moxibustion was applied at 9:00 a.m.every day in the rats.The rats were fixed on a fixed table,the mild moxibustion with self-prepared moxa stick was applied to bilateral "BL23""BL21" and the non-acupoint of each rat,the ignited end of moxa stick was 2 cm to 3 cm far from skin surface.The infrared thermometer was adopted to measure the temperature of the epidermis of each acupoint from time to time so as to maintain the local temperature to be(41 ± 0.5)℃.Mild moxibustion lasted for 10 min each time,consecutively for 5 days a week and at an interval of 2 days.The total consecutive intervention duration was 12 weeks.In the normal group and the shamoperation group,no moxibustion intervention was performed,but the rats were restricted in a same way during the experiment.At the end of modeling and intervention,water maze test was adopted to detect the learning and memory abilities of rats in each group.The western blotting was used to detect the expressions of p-CREB,p-CaMKII,CaMKII and CREB in hippocampus of rats in each group.Results:(1)Navigation test:since Day 72,compared with the sham-operation group,the escape latencies in all of the consecutive 5 days were longer in the model group(all P <0.05).Since Day 73,compared with the normal group,the escape latency in all of the consecutive 4 days was shorter in the "BL23"group(all P <0.05).(2) Times across platform:compared with the normal group,the number of times consecutively crossing the platform was lower in the model group(P <0.05).After moxibustion,compared with the model group,the number of times across platform was higher in the "BL23"group,indicating a statistical significance(P <0.05).(3)Moving track of water maze spatial probe:the "tendency"strategy was displayed in the normal group and the sham operation group.The "marginal" strategy,which is commonly seen in the AD rats,was displayed in the model group.The "random" strategy was displayed in the "BL23" group.The "marginal" strategy was presented in the "BL21" group and the non-acupoint group.(4) Expressions of hippocampal P-CAMKII,CAMKII,CREB and P-CREB in the rats:compared with the normal group,the phosphorylation degrees of CAMKII and CREB were significantly reduced(both P<0.05) in the model group.In 12 weeks of moxibustion,compared with the model group,the levels of P-CAMKII and P-CREB were increased in the "BL23" group and the "BL21" group(all P <0.05) and the levels in the "BL23" group were higher than that of the "BL21" group and the non-acupoint group(all P<0.05).Conclusion:Moxibustion at "BL23" achieves a better effect in improving the learning and memory of AD rats modeled by ovariectomy and D-Gal injection as compared with moxibustion at "BL21" and the non-acupoint.The effect mechanism is potentially related to the effective activation of Ca2+/CaMKII/CREB signaling pathway.展开更多
基金the Natural Sci-ence Foundation of HenanProvince, No. 984021100 agrant from Key Subject Fund ofXinxiang Medical College
文摘BACKGROUD: Ethanol can influence neural development and the ability of learning and memory, but its mechanism of the neural toxicity is not clear till now. Endogenous nitric oxide (NO) as a gaseous messenger is proved to play an important role in the formation of synaptic plasticity, transference of neuronal information and the neural development, but excessive nitro oxide can result in neurotoxicity. OBJECTIVE: To observe the effects of acute alcoholism on the learning and memory ability and the content of neuronal nitric oxide synthase (nNOS) in brain tissue of rats. DESIGN: A randomized controlled animal experiment. SETTING: Department of Physiology, Xinxiang Medical College. MATERIALS: Eighteen male clean-degree SD rats of 18-22 weeks were raised adaptively for 2 days, and then randomly divided into control group (n = 8) and experimental group (n = 10). The nNOS immunohistochemical reagent was provided by Beijing Zhongshan Golden Bridge Biotechnology Co.,Ltd. Y-maze was produced by Suixi Zhenghua Apparatus Plant. METHODS: The experiment was carried out in the laboratory of the Department of Physiology, Xinxiang Medical College from June to October in 2005. ① Rats in the experimental group were intraperitoneally injected with ethanol (2.5 g/kg) which was dissolved in normal saline (20%). The loss of righting reflex and ataxia within 5 minutes indicated the successful model. Whereas rats in the control group were given saline of the same volume. ② Examinations of learning and memory ability: The Y-maze tests for learning and memory ability were performed at 6 hours after the models establishment. The rats were put into the Y-maze separately. The test was performed in a quiet and dark room. There was a lamp at the end of each of three pathways in Y-maze and the base of maze had electric net. All the lamps of the three pathways were turned on for 3 minutes and then turned off. One lamp was turned on randomly, and the other two delayed automatically. In 5 seconds after alternation, pulsating electric current presented in the base of unsafe area to stimulate rat’s feet to run to the safe area. The lighting lasted for 15 seconds as one test. Running from unsafe area to safe area at one time in 10 seconds was justified as successful. Such test was repeated for 10 times for each rat and the successful frequency was recorded. The qualified standard of maze test was that the rat arrived in the safe area 9 times during 10 experiments. The number of trainings for the qualified standard was used to represent the result of spatial learning. ③ Determination of the content of nNOS in brain tissue: After the Y-maze test, the rats were anaesthetized, and blood was let from the incision on right auricle, transcardially perfused via the left ventricle with about 200 mL saline, then fixed by perfusion of 40 g/L paraformaldehyde. Hippocampal CA1 region, corpus striatum and cerebellum were taken to prepare serial freezing coronal sections. The nNOS contents in the brain regions were determined with the immunohistochemical methods to reflect the changes of nitric oxide in brain tissue. MAIN OUTCOME MEASURES: The changes of learning and memory ability and the changes of the nNOS contents in the brain tissue of rats with acute alcoholism were observed. RESULTS: One rat in the experimental group was excluded due to its slow reaction to electric stimulation in the Y-maze test, and the other 17 rats were involved in the analysis of results. ① The training times to reach qualifying standards of Y-maze in the experimental group was more than that in the control group [(34.33 ±13.04), (27.50±8.79) times, P < 0.05]. ② Forms and numbers of nNOS positive neurons in brain tissue: It could be observed under light microscope that in the hippocampal CA1 region, there were fewer nNOS positive neurons, which were lightly stained, and the processes were not clear enough; But the numbers of the positive neurons which were deeply stained as buffy were obviously increased in the experimental group, the cell body and cytoplasm of process were evenly stained, but the nucleus was not stained. The nNOS positive neurons in corpus striatum had similar forms and size in the experimental group and control group. The form of the nNOS positive neurons in cerebellum were similar between the two groups. The numbers of nNOS positive neurons in hippocampal CA1 region and corpus striatum in the experimental group [(18.22±7.47), (11.38±5.00) cells/high power field] were obviously higher than those in the control group [(10.15±4.24), (6.15±3.69) cells/high power field. The number of nNOS positive neurons in cerebellum had no significant difference between the two groups [(49.56±18.84), (44.43±15.42) cells/high power field, P > 0.05]. CONCLUSION: Acute alcoholism may impair learning and memory ability, and nitric oxide may be involved in mediating the neurotoxic role of ethanol.
文摘BACKGROUND: Central adrenergic nerve and 5-serotonergic nerve can influence central cholinergic nerve on learning and memory and make easy for study; however, ginsenoside of stem and leaf (GSL) can improve functions of central adrenergic nerve; moreover, 5-serotonergic nerve and the combination with choline can produce synergistic effect and enhance learning and memory ability so as to improve learning and memory disorder of patients with Alzheimer disease (AD). OBJECTIVE: To observe the effects of GSL combining with choline on learning and memory of AD model rats.DESIGN: Randomized grouping design and controlled animal study.SETTING: Department of Pharmacology, Taishan Medical College. MATERIALS: The experiment was carried out in the Pharmacological Department of Medical College of Jilin University from October 1996 to January 1997. Forty healthy male Wistar rats of clean grade were randomly divided into 5 groups, including sham-injury group, model group, GSL group, choline group and combination group, with 8 rats in each group. Main medications: GSL with the volume more than 92.8% was provided by Department of Chemistry, Norman Bethune Medical College of Jilin University. Panaxatriol, the main component, was detected with thin layer scanning technique and regarded as the index of GSL quality [(55±1)%, CV = 2%, n = 5]. Choline was provided by the Third Shanghai Laboratory Factory. METHODS: 150 nmol quinolinic acid was used to damage bilateral Meynert basal nuclei of adult rats so as to establish AD models. Rats in GSL, choline and combination groups were intragastric administrated with 400 mg/kg GSL, 200 mg/kg choline (20 mL/kg), and both respectively last for 17 days starting from two days before operation. Rats in sham-injury group and model group were perfused with the same volume of distilled water once in each morning for the same days. ① Passive avoidance step-down test: Five minutes later, rats jumped up safe platform when they were shocked with 36 V alternating current. If rats jumped down from the platform and the feet touched railings, the response was wrong. Numbers of wrong response were recorded within 3 minutes, and then the test was redone after 24 hours. ② Morris water-maze spatial localization task: Swimming from jumping-off to platform directly was regarded as right response. Additionally, 4 successively right responses were regarded as the standard. Each rat was trained 10 times a day with 120 s per time for 3 successive days. The interval was 30 s. Three days later, numbers of right response were recorded. The training times were increased to 30 for unlearned rats. ③ Measurement of activity of choilne acetylase in cerebral cortex: Rats were sacrificed at 17 days after operation to obtain cerebral cortex to measure activity of choilne acetylase with radiochemistry technique. ④ Synergistic effect: It was expressed as Q value: Q value = factual incorporative effect/anticipant incorporative effect; Q ≥ 1 was regarded as synergistic effect. Anticipant incorporative effect = (EA+EB-EA·EB), EA and EB were single timing effect, respectively in GSL group and choline group. E (step-down test and Morris water maze test) = (x in model group - factual value in medicine groups)/x in model group; E (activity of choilne acetylase) = (factual value in medicine groups -x in model group)/x in model group. MAIN OUTCOME MEASURES: ① Passive avoidance step-down test and Morris water-maze spatial localization task in the study of learning and memory; ② activity of choilne acetylase. RESULTS: All 40 rats were involved in the final analysis. ① Passive avoidance response: At learning phase on first day and retesting phase on the next day, numbers of wrong responses within 3 minutes were more in model group than sham operation group, and there was significant difference [(5.88±1.46), (2.25±0.87) times; (2.63±1.06), (0.50±0.53) times; P < 0.01]; numbers of wrong responses within 3 minutes were less in combination group than model group, and there was significant difference [learning phase: (1.12±0.83), (5.88±1.46) times; retesting phase: (0.38±0.74), (2.63±1.06) times, P < 0.01]; moreover, effect was stronger than that in GSL group and choline group. The Q value was 1.07 and 1.59, respectively and it showed synergistic effect. ② Spatial localization task: Training times were more in model group than sham operation group, and there was significant difference [(2.9±2.5), (12.6±3.5) times; P < 0.01]. Training times were less in combination group than model group, and there was significant difference [(11.8±2.4), (27.9±2.5) times, P < 0.01]; moreover, effect was stronger than that in GSL group and choline group. The Q value was 1.07 and it showed synergistic effect. ③ Activity of choilne acetylase: Activity was lower in model group than sham operation group, and there was significant difference [(30.56±8.33), (61.11±8.33) nkat/g; P < 0.01]. Activity was higher in combination group than model group and there was significant difference [(50.00±8.33), (30.56±8.33) nkat/g, P < 0.01]; moreover, effect was stronger than that in GSL group and choline group. The Q value was 1.5 and it showed synergistic effect. CONCLUSION: GSL in combination with choline can synergically improve the disorder of learning and memory of AD model rats. Its mechanism may be involved in enhancing the function of central cholinergic system.
文摘BACKGROUND: Generally speaking, anesthesia is often used in gravid body and it has been already proved that many kind of medicine can result in malformation. OBJECTIVE: To explore embryonic skeleton development and neonatal learning and memory of rats anesthetized with pentobarbital sodium in gravid rats. DESIGN: A randomized control trial. SETTING: Laboratory Animal Center of Xuzhou Medical College. MATERIALS: A total of 80 adult female SD rats, of clean grade and weighing 220-240 g, were selected in this study. The main reagents were detailed as follows: pentobarbital sodium (Shanghai Xingzhi Chemical Plant, batch number: 921019); MG-2 maze test apparatus (Zhangjiagang Biomedical Instrument Factory); somatotype microscope (Beijing Taike Instrument Co., Ltd.). METHODS: ① A total of 160 SD rats of half males and females were selected in this study. All rats were copulated. The day that the plug was checked out in the vagina next day was looked as the first day of pregnancy. Gravid rats were divided randomly into four groups, including early anesthesia group, second anesthesia group, late anesthesia group and control group with 20 in each group. Rats in the early anesthesia group were injected with 25 mg/kg soluble pentobarbitone on the 7th day of pregnancy for once; rats in the second anesthesia group were anesthetized with 25 mg/kg soluble pentobarbitone on the 7th and the 14th days of pregnancy for once; rats in the late anesthesia group were anesthetized with 25 mg/kg soluble pentobarbitone on the 14th day of pregnancy for once; rats in the control group did not treat with anything. The time of anesthetizing was controlled in 3 to 4 hours and ether was absorbed while the time was not enough. ② Half of each group was sacrificed on day 20th of pregnancy and the fetus was taken out to be stained with alizarin red S. After stained, the fetal skeleton was examined. The learning and memorizing of one-month rats that were given birth by the rest gravid rats were tested through electric mare method. Determine their study ability according to their correct rate of 90% or above of arrival at the safe area in 20 s. After they finally learned to arrive at the safe area correctly, test them once more in 24 hours and record the correct rate of 15 times. MAIN OUTCOME MEASURES: The rate of malformation in fetus and ability of learning and memory in one-month rats. RESULTS: A total of 80 female rats were anesthetized in this experiment. Totally 490 immature rats were tested with maze testing machine and 196 fetuses were stained with alizarin red S to observe the development of their skeleton. However, one of the 80 female rats was led to death because of overdose. ① Malformation experiment: Learning ability of second anesthesia group was evidently different from the control group while the other two groups were not in the electric mare method. The fetal skeleton malformation rate of three experimental groups was 87.0%, 60.9% and 17.9%, respectively, while it was 5.6% in the control group. ② Electric mare method: Times of rats which arrived at the safe regions were respectively 49.0±31.0, 68.0±35.0, 47.0±31.0 and 44.0±21.0 in early anesthesia group, second anesthesia group, late anesthesia group and control group; and then, there was significant difference between the second anesthesia group and the control group (P < 0.05). Exact rates of memory of rats were respectively (64.36±14.35)%, (62.15±18.33)%, (54.19±12.28)% and (68.24±15.91)% in early anesthesia group, second anesthesia group, late anesthesia group and control group; and then, there were no significant differences as compared with the control group (P > 0.05). CONCLUSION: The influence of anesthesia with pentobarbital sodium is obvious in fetal skeleton development and learning and memory ability.
基金the National Natural Science Foundation of China, No. 03030307 the Great Special Fund of Guangdong Province, No. 2004A30201002
文摘BACKGROUND: Transplantation of fetal cell suspension or blocks of fetal tissue can ameliorate the nerve function after the injury or disease in the central nervous system, and it has been used to treat neurodegenerative disorders induced by Parkinson disease. OBJECTIVE: To observe the effects of the transplantation of neuron-like cells derived from bone marrow stromal cells (rMSCs) into the brain in restoring the dysfunctions of muscle strength and balance as well as learning and memory in rat models of cerebral infarction. DESIGN: A randomized controlled experiment. SETTING: Department of Pathophysiology, Zhongshan Medical College of Sun Yat-sen University. MATERIALS: Twenty-four male SD rats (3-4 weeks of age, weighing 200-220 g) were used (Certification number:2001A027). METHODS: The experiments were carried out in Zhongshan Medical College of Sun Yat-sen University between December 2003 and December 2004. ① Twenty-four male SD rats randomized into three groups with 8 rats in each: experimental group, control group and sham-operated group. Rats in the experiment al group and control group were induced into models of middle cerebral artery occlusion. After in vitro cultured, purified and identified with digestion, the Fischer344 rMSCs were induced to differentiate by tanshinone ⅡA, which was locally injected into the striate cortex (18 area) of rats in the experimental group, and the rats in the control group were injected by L-DMEM basic culture media (without serum) of the same volume to the corresponding brain area. In the sham-operated group, only muscle and vessel of neck were separated. ② At 2 and 8 weeks after the transplantation, the rats were given the screen test, prehensile-traction test, balance beam test and Morris water-maze test. ③ The survival and distribution of the induced cells in corresponding brain area were observed with Nissl stained with toluidine blue and hematoxylin and eosin (HE) staining in the groups. MAIN OUTCOME MEASURES: ① Results of the behavioral tests (time of the Morris water-maze test screen test, prehensile-traction test, balance beam test); ② Survival and distribution of the induced cells. RESULTS: All the 24 rats were involved in the analysis of results. ① Two weeks after transplantation, rats with neuron-like cells grafts in the experimental group had significant improvement on their general muscle strength than those in the control group [screen test: (9.4±1.7), (4.7±1.0) s, P < 0.01]; forelimb muscle strength [prehensile-traction test: (7.6±1.4), (5.2±1.2) s, P < 0.01], ability to keep balance [balance beam test: (7.9±0.74), (6.1±0.91) s, P < 0.01] and abilities of learning and memory [latency to find the platform: (35.8±5.9), (117.5±11.6) s, P < 0.01; distance: (623.1±43.4), (1 902.3±98.6) cm, P < 0.01] as compared with those in the control group. The functional performances in the experimental group at 8 weeks were better than those at two weeks, which were still obviously different from those in the sham-operated group (P < 0.05). ② The HE and Nissl stained brain tissue section showed that there was nerve cell proliferation at the infarcted cortex in the experiment group, the density was higher than that in the control group, plenty of aggregative or scattered cells could be observed at the site where needle was inserted for transplantation, the cells migrated directively towards the area around them, the cerebral vascular walls were wrapped by plenty of cells; In the control group, most of the cortices were destroyed, karyopyknosis and necrosis of neurons were observed, normal nervous tissue structure disappeared induced by edema, only some nerve fibers and glial cells remained. CONCLUSION: The rMSCs transplantation can obviously enhance the motor function and the abilities of learning and memory in rat models of cerebral infarction.
文摘BACKGROUND: Calcitonin gene-related peptide (CGRP) and nerve growth actor (NGF) cam improve spatial learning and memory abilities of rats with cerebral ischemia/reperfusion; however, the effect of combination of them on relieving learning and memory injury following cerebral ischemia/reperfusion should be further studied. OBJECTIVE: To study the effects of exogenous CGRP and NGF on learning and memory abilities of rats with focal cerebral ischemia/reperfusion. DESIGN: Randomized controlled animal study. SETTING: Department of Neurosurgery, the Second Hospital of Xiamen; Department of Neurosurgery, the Second Affiliated Hospital of China Medical University; Department of Neurobiology, Basic Medical College of China Medical University. MATERIALS: A total of 30 healthy male SD rats, aged 8 weeks, of clean grade, weighing 250-300 g, were provided by Experimental Animal Department of China Medical University. All rats were randomly divided into sham-operation group, ischemia/reperfusion group and treatment group with 10 in each group. The main reagents were detailed as the follows: 100 g/L chloral hydrate, 0.5 mL CGRP (2 mg/L, Sigma Company, USA), and NGF (1× 106 U/L, 0.5 mL, Siweite Company, Dalian). METHODS: The experiment was carried out in the Department of Neurobiology, Basic Medical College of China Medical University from February to July 2005. Rat models of middle cerebral artery occlusion were established by method of occlusion, 2 hours after that rats were anesthetized and the thread was slightly drawn out for 10 mm under direct staring to perform reperfusion. Rats in the ischemia/reperfusion group received intraperitoneal injection of 1 mL saline via the abdomen at two hours later, while rats in the treatment group at 2 hours later received intraperitoneal injection of 2 mg/L CGRP (0.5 mL) and 1×106 U/L NGF (0.5 mL) once a day for 10 successive days. First administration was accomplished within 15 minutes after ischemia/reperfusion. Rats in the sham-operation group were separated of the vessels without occlusion or administration. The neural function was evaluated with Zea Longa 5-grade scale. Animals with the score of one, two and three points received Morris water-maze test to measure searching time on platform (omitting platform-escaping latency). Meanwhile, leaning and memory abilities of animals were reflected through testing times of passing through platform per minute. MAIN OUTCOME MEASURES: Experimental results of omitting platform-escaping latency and spatial probe. RESULTS: Three and two rats in the ischemia/reperfusion group and treatment group respectively were not in accordance with the criteria in the process, and the rest were involved in the final analysis. ① Comparisons of platform-escaping latency during Morris water-maze test in all the three groups: Ten days after modeling, the platform-escaping latency in the ischemia/reperfusion group was obviously longer than that in sham-operation group (P < 0.01), and was significantly shorter than that in the treatment group (P < 0.01). ② Comparisons of times of passing through platform in all the three groups: Times of passing through platform were remarkably less in the ischemia/reperfusion group than those in the sham-operation group [(1.79±0.39), (4.30±0.73) times/minute, P < 0.01], and those were markedly more in the treatment group than the ischemia/reperfusion group [(3.16±1.03), (1.79±0.39) times/minute, P < 0.01]. CONCLUSION: CGRP and NGF are capable of ameliorating the abilities of spatial learning and memory in MCAO rats, which indicates that CGRP and NGF can protect ischemic neurons.
基金supported by the National Natural Science Foundation of China,No.31571109,81460261the Chinese-Finnish Joint Project Fund,No.813111172+2 种基金a grant from the Yunnan Key Program of Science and Technology of China,No.2014FC005the Key Science and Technology Research Project Fund of Hainan Province of China,No.ZDYF2016156the National Clinical Key Subject Construction Project Fund of China
文摘Rhesus monkey neural stem cells are capable of differentiating into neurons and glial cells.Therefore,neural stem cell transplantation can be used to promote functional recovery of the nervous system.Rhesus monkey neural stem cells(1×105 cells/μL) were injected into bilateral hippocampi of rats with hippocampal lesions.Confocal laser scanning microscopy demonstrated that green fluorescent protein-labeled transplanted cells survived and grew well.Transplanted cells were detected at the lesion site,but also in the nerve fiber-rich region of the cerebral cortex and corpus callosum.Some transplanted cells differentiated into neurons and glial cells clustering along the ventricular wall,and integrated into the recipient brain.Behavioral tests revealed that spatial learning and memory ability improved,indicating that rhesus monkey neural stem cells noticeably improve spatial learning and memory abilities in rats with hippocampal lesions.
基金Supported by National Natural Science Foundation of China Project:No.81473786No.81873380。
文摘Objective:To observe the effect of the early intervention with moxibustion at "Shènshū(肾俞BL23)" on learning and memory of Alzheimer’s disease(AD) rats modeled by ovariectomy and D-Galactose(D-Gal)injection and the effect on Ca2+/CaMKII/CREB signaling pathway so as to provide the evidence for the acupoint selection in the early intervention with acupuncture and moxibustion for AD.Methods:Sixty female SD rats at the age of 3 months were selected and randomly divided into a normal group,a sham-operation group,a model group,a "BL23" group,a "Wèishū"(胃俞BL21) group,and a non-acupoint group,10 rats in each group.Except in the normal group and the sham-operation group,bilateral ovaries were removed in the other 4 groups.3 days after the operation,D-Gal was injected intraperitoneally at a dose of 150 mg/kg/d for 90 days to establish AD rat model.In the sham-operation group,only the peripheral fat of ovarian was removed and 0.9% normal saline was injected intraperitoneally of the same dose for 90 days.In the "BL23" group,the "BL21" group and the non-acupoints group,since the 2 nd day after ovariectomy,moxibustion was applied at 9:00 a.m.every day in the rats.The rats were fixed on a fixed table,the mild moxibustion with self-prepared moxa stick was applied to bilateral "BL23""BL21" and the non-acupoint of each rat,the ignited end of moxa stick was 2 cm to 3 cm far from skin surface.The infrared thermometer was adopted to measure the temperature of the epidermis of each acupoint from time to time so as to maintain the local temperature to be(41 ± 0.5)℃.Mild moxibustion lasted for 10 min each time,consecutively for 5 days a week and at an interval of 2 days.The total consecutive intervention duration was 12 weeks.In the normal group and the shamoperation group,no moxibustion intervention was performed,but the rats were restricted in a same way during the experiment.At the end of modeling and intervention,water maze test was adopted to detect the learning and memory abilities of rats in each group.The western blotting was used to detect the expressions of p-CREB,p-CaMKII,CaMKII and CREB in hippocampus of rats in each group.Results:(1)Navigation test:since Day 72,compared with the sham-operation group,the escape latencies in all of the consecutive 5 days were longer in the model group(all P <0.05).Since Day 73,compared with the normal group,the escape latency in all of the consecutive 4 days was shorter in the "BL23"group(all P <0.05).(2) Times across platform:compared with the normal group,the number of times consecutively crossing the platform was lower in the model group(P <0.05).After moxibustion,compared with the model group,the number of times across platform was higher in the "BL23"group,indicating a statistical significance(P <0.05).(3)Moving track of water maze spatial probe:the "tendency"strategy was displayed in the normal group and the sham operation group.The "marginal" strategy,which is commonly seen in the AD rats,was displayed in the model group.The "random" strategy was displayed in the "BL23" group.The "marginal" strategy was presented in the "BL21" group and the non-acupoint group.(4) Expressions of hippocampal P-CAMKII,CAMKII,CREB and P-CREB in the rats:compared with the normal group,the phosphorylation degrees of CAMKII and CREB were significantly reduced(both P<0.05) in the model group.In 12 weeks of moxibustion,compared with the model group,the levels of P-CAMKII and P-CREB were increased in the "BL23" group and the "BL21" group(all P <0.05) and the levels in the "BL23" group were higher than that of the "BL21" group and the non-acupoint group(all P<0.05).Conclusion:Moxibustion at "BL23" achieves a better effect in improving the learning and memory of AD rats modeled by ovariectomy and D-Gal injection as compared with moxibustion at "BL21" and the non-acupoint.The effect mechanism is potentially related to the effective activation of Ca2+/CaMKII/CREB signaling pathway.
