本文選題：睡眠剝奪 + 學(xué)習(xí)�。� 參考：《天津醫(yī)科大學(xué)》2007年碩士論文

【摘要】： 目的 睡眠是腦的高級功能之一，是生命賴以維持的基本條件。睡眠剝奪是由于各種原因?qū)е碌乃卟蛔愕臓顟B(tài)，睡眠剝奪能夠?qū)C(jī)體產(chǎn)生廣泛的影響。本研究利用小平臺水環(huán)境法制備大鼠睡眠剝奪動物模型，探討不同持續(xù)時間睡眠剝奪對大鼠學(xué)習(xí)能力的影響并初步分析其可能機(jī)制。 方法 1實(shí)驗(yàn)動物及分組 選用體重240～260g的健康成年SD大鼠，雌雄各半。隨機(jī)分為正常對照組、大平臺對照組、睡眠剝奪2d、4d、6d組。 2睡眠剝奪模型制備 制作30cm×30cm×30cm睡眠剝奪箱，內(nèi)置一直徑6.3cm的小平臺，高8cm，箱中注水，水面低于平臺面約1.0cm。利用此法制備大鼠睡眠剝奪2d、4d、6d組模型。大平臺對照組平臺直徑為18cm，大鼠置于大平臺環(huán)境6天，其余條件同睡眠剝奪組。正常對照組籠養(yǎng)。 3一般狀態(tài)觀察 觀察睡眠剝奪后大鼠精神狀態(tài)、皮毛、飲水飲食、反應(yīng)能力等改變。 4學(xué)習(xí)能力測試 大鼠學(xué)習(xí)能力測試在三等分“Y”型迷宮中進(jìn)行，環(huán)境為安靜暗室。Y迷宮每臂頂端設(shè)1個信號燈，以此提示“安全區(qū)”，通電后安全區(qū)燈亮，該區(qū)內(nèi)無電流刺激，其余區(qū)域被通以36V交流電。實(shí)驗(yàn)時，將大鼠放入迷宮，先適應(yīng)5分鐘，然后開始實(shí)驗(yàn)。規(guī)定大鼠受電擊后從起步區(qū)直接逃至亮臂安全區(qū)為“正確反應(yīng)”。實(shí)驗(yàn)中每次通電刺激間隔時間為30秒，大鼠連續(xù)10次反應(yīng)正確時，即為達(dá)標(biāo)，記錄大鼠達(dá)標(biāo)時所需的總訓(xùn)練次數(shù)，以此代表學(xué)習(xí)能力。 5海馬組織超微結(jié)構(gòu)透射電鏡檢查 取SD-6d大鼠海馬組織制作電鏡標(biāo)本，觀察超微結(jié)構(gòu)改變。 6海馬組織神經(jīng)遞質(zhì)測定 將大鼠迅速斷頭，取雙側(cè)海馬。高效液相色譜法檢測五羥色胺、去甲腎上腺素、谷氨酸、γ-氨基丁酸含量；堿性羥胺比色法檢測乙酰膽堿含量。 7統(tǒng)計學(xué)分析方法 各組數(shù)據(jù)以(?)±s表示。利用單因素方差分析進(jìn)行假設(shè)檢驗(yàn)。組間差異利用Dunnett-t檢驗(yàn)，檢驗(yàn)水準(zhǔn)α=0.05。 結(jié)果 1大鼠睡眠剝奪后一般狀態(tài)的改變 SD組動物在實(shí)驗(yàn)早期表現(xiàn)出一定的興奮性行為，，如活動增多，對外界聲、光等刺激敏感。而在實(shí)驗(yàn)中后期(SD-4d以后)，SD動物行為由興奮逐漸轉(zhuǎn)為抑制，并表現(xiàn)出情緒上的“激惹”現(xiàn)象。 2睡眠剝奪對大鼠學(xué)習(xí)能力的影響 與CC組比較，SD-4d組和SD-6d組大鼠學(xué)習(xí)能力下降，達(dá)標(biāo)所需訓(xùn)練次數(shù)增多(p＜0.01)；SD2d組學(xué)習(xí)能力提高，達(dá)標(biāo)所需訓(xùn)練次數(shù)減少(p＜0.05)。 3睡眠剝奪大鼠海馬超微結(jié)構(gòu)改變 海馬神經(jīng)元細(xì)胞水腫，暗細(xì)胞變，向凋亡或類凋亡方向過渡；部分神經(jīng)元出現(xiàn)核染色質(zhì)溶解，胞質(zhì)內(nèi)粗面內(nèi)質(zhì)網(wǎng)和核糖體明顯減少或缺失，線粒體減少，結(jié)構(gòu)模糊；部分突觸水腫，突觸小泡減少；血腦屏障結(jié)構(gòu)明顯異常，內(nèi)皮細(xì)胞暗調(diào)型或細(xì)胞變性甚至出現(xiàn)胞質(zhì)斷裂等嚴(yán)重?fù)p傷的表現(xiàn)，星形膠質(zhì)細(xì)胞足突嚴(yán)重水腫；在神經(jīng)元內(nèi)及血腦屏障結(jié)構(gòu)等處均可見到髓磷體明顯增多。 4睡眠剝奪對海馬神經(jīng)遞質(zhì)含量的影響 乙酰膽堿：SD-4d組、SD-6d組大鼠海馬Ach含量低于CC組，具有顯著性差異(p＜0.05，p＜0.01)。 去甲腎上腺素：SD-4d組、SD-6d組大鼠海馬NE含量低于CC組(p＜0.05)；TC組及SD-2d組NE含量高于CC組(p＜0.05)。 五羥色胺：高效液相法測得SD-2d組、SD-4d組大鼠海馬5-HT含量明顯高于CC組和TC組(p＜0.01，p＜0.05)；SD-6d組大鼠海馬5-HT含量明顯低于CC組和TC組(p＜0.01)；TC組5-HT含量與CC組相比無顯著差異。 谷氨酸、γ-氨基丁酸及二者比值：SD-2d組大鼠海馬GLU含量高于CC組(p＜0.01)，SD-6d組大鼠海馬GLU含量低于CC組(p＜0.05)，與TC組比較，SD-2d組和SD-4d組GLU含量增加(p＜0.01，p＜0.01)，SD-6d組GLU含量減少(p＜0.05)。SD各組大鼠海馬GABA含量均高于CC組(p＜0.05，p＜0.01)；與CC組比較，SD-4d組和SD-6d組GLU／GABA值顯著降低(p＜0.01)；SD-2d和TC組GLU／GABA值無顯著變化。 結(jié)論 1．較長時間(4天以上)睡眠剝奪會造成大鼠學(xué)習(xí)能力降低，表現(xiàn)為Y型迷宮訓(xùn)練達(dá)標(biāo)所需訓(xùn)練次數(shù)增多。 2．睡眠剝奪引起大鼠海馬結(jié)構(gòu)損傷，表現(xiàn)為神經(jīng)元細(xì)胞水腫，向凋亡或類凋亡方向過渡；突觸水腫，突觸小泡減少；血腦屏障結(jié)構(gòu)明顯異常；髓磷體明顯增多。 3．睡眠剝奪可以造成大鼠海馬多種神經(jīng)遞質(zhì)(乙酰膽堿、去甲腎上腺素、五羥色胺、谷氨酸、γ-氨基丁酸及GLU／GABA比值)分布異常。 4．睡眠剝奪引起的海馬結(jié)構(gòu)損傷及神經(jīng)遞質(zhì)含量異常是SD損害動物學(xué)習(xí)能力的可能機(jī)制。
[Abstract]:objective
Sleep is one of the advanced functions of the brain. It is the basic condition for the maintenance of life. Sleep deprivation is a state of sleep deprivation caused by various reasons. Sleep deprivation can exert a wide influence on the body. This study uses a small platform water environment to prepare rat sleep deprivation animal models and discuss the effect of sleep deprivation on different duration of sleep. The influence of learning ability of rats was analyzed and its possible mechanism was preliminarily analyzed.
Method
1 experimental animals and groups
Healthy adult SD rats weighing 240 to 260g were randomly divided into normal control group, large platform control group, sleep deprived 2D group, 4D group and 6D group.
The preparation of 2 sleep deprivation model
The 30cm x 30cm x 30cm sleep deprivation box was built with a small platform with a constant diameter of 6.3cm, high 8cm, water injection in the box, and the water surface was lower than the platform of about 1.0cm. to make rats sleep deprived of 2D, 4D, 6D group model. The platform diameter of the large platform control group was 18cm, the rats were placed in the large platform environment for 6 days, and the rest were kept in the same condition as the sleep deprivation group. Normal control group cage.
3 general state observation
The changes of mental state, fur, drinking water and reactivity after sleep deprivation were observed.
4 learning ability test
The rats' learning ability test was carried out in the three equal "Y" maze labyrinth, the environment for the quiet dark room.Y maze at the top of each arm set up 1 signal lights, which prompted the "safety zone", the safety zone light after the electricity, the area without current stimulation, the rest of the area was switched to 36V alternating current. Experiment, the rats in the maze, first adapt to 5 minutes, and then start the experiment It is required that the rats receive a correct response from the start area to the bright arm safety area after the electric shock. The interval of the stimulation is 30 seconds each time in the experiment. When the rats are correct for 10 consecutive times, the rats are up to the standard to record the total training times required when the rats reach the standard, which represents the learning ability.
Ultrastructural transmission electron microscopic examination of 5 hippocampal tissue
The hippocampal tissues of SD-6d rats were made to observe the ultrastructural changes by electron microscopy.
Determination of neurotransmitter in 6 hippocampal tissue
The rats were quickly broken and the hippocampus was taken. The content of five hydroxytryptamine, norepinephrine, glutamic acid and gamma aminobutyric acid were detected by high performance liquid chromatography, and the content of acetylcholine was detected by alkaline hydroxylamine colorimetric assay.
7 statistical analysis method
The data of each group were expressed as (+) s. Hypothesis test was conducted by one-way ANOVA. Dunnett-t test was used to test the level of alpha =0.05..
Result
Changes in the general state of 1 rats after sleep deprivation
In the early experiment, the animals of group SD showed certain excitatory behavior, such as increased activity, external sound, light and other irritation. In the later period of the experiment (after SD-4d), the behavior of SD animal was gradually turned from excitement to inhibition and showed emotional "irritability".
The effect of 2 sleep deprivation on the learning ability of rats
Compared with the CC group, the learning ability of rats in group SD-4d and group SD-6d was decreased, and the number of training times needed to meet the standard was increased (P < 0.01); the learning ability of the SD2d group was improved and the number of training required to meet the standard was reduced (P < 0.05).
The ultrastructural changes of hippocampus in 3 sleep deprived rats
Hippocampal neurons were edema, dark cell change, transition to apoptosis or apoptosis, nucleus chromatin dissolved in some neurons, apparent decrease or loss of rough endoplasmic reticulum and ribosome in the cytoplasm, mitochondria decreased, structure blurred, partial synaptic edema, synaptic vesicles decreased; blood brain barrier structure was obviously abnormal, endothelial cell tone type The appearance of severe injury, such as cell degeneration or even cytoplasmic breakage, is a serious edema of the astrocyte podocyte, and a significant increase in myelin body can be seen in the neurons and the structure of the blood brain barrier.
The effect of 4 sleep deprivation on the content of hippocampal neurotransmitters
Acetylcholine: in group SD-4d, the content of Ach in hippocampus of SD-6d group was lower than that of CC group, with significant difference (P < 0.05, P < 0.01).
In norepinephrine group SD-4d, the content of NE in hippocampus of SD-6d group was lower than that of CC group (P < 0.05); NE content in TC group and SD-2d group was higher than that in CC group (P < 0.05).
Five serotonin: SD-2d group was measured by high performance liquid chromatography. The content of 5-HT in hippocampus of rats in group SD-4d was significantly higher than that in group CC and TC group (P < 0.01, P < 0.05), and the content of 5-HT in hippocampus of SD-6d group was significantly lower than that of CC group and TC group (P < 0.01).
The ratio of glutamic acid, gamma aminobutyric acid and two groups was higher than that of group CC (P < 0.01), and the content of GLU in hippocampus of group SD-6d was lower than that of group CC (P < 0.05). Compared with group TC (P < 0.05), the content of GLU in SD-2d and SD-4d group was increased (0.01, 0.01), and the content of hippocampus in hippocampus was higher than that of 0. group (0.05). 05, P < 0.01); compared with group CC, the GLU / GABA values of group SD-4d and group SD-6d were significantly lower (P < 0.01); SD-2d / TC group had no significant change in GLU / GABA value.
conclusion
1. sleep deprivation in a longer period of time (more than 4 days) will reduce the learning ability of rats. The result shows that the number of training required for Y maze training is increased.
2. sleep deprivation caused hippocampal damage in rats, characterized by edema of neuron cells, transition to apoptosis or apoptotic direction, synaptic edema, decrease of synaptic vesicles, obvious abnormal structure of blood brain barrier, and significantly increased myelin body.
3. sleep deprivation can cause a variety of neurotransmitters in the hippocampus of rats (acetylcholine, norepinephrine, five serotonin, glutamic acid, gamma aminobutyric acid and GLU / GABA ratio) distribution.
4. sleep deprivation induced hippocampal structural damage and abnormal neurotransmitter content are the possible mechanisms of SD damaging the learning ability of animals.

【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2007
【分類號】：R363

【引證文獻(xiàn)】

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