發(fā)布時間：2018-07-25 21:21

【摘要】：女性生殖周期雌孕激素呈現(xiàn)周期性變化，尤其是妊娠過程和分娩后激素的較大變化使得女性出現(xiàn)各種各樣情緒障礙及各種臨床癥狀。有研究認(rèn)為,婦女的以上各類精神疾病均有女性生殖周期雌、孕激素等卵巢甾體激素的波動所引發(fā)。這些臨床癥狀的出現(xiàn)與孕激素對GABAA調(diào)節(jié)系統(tǒng)的作用有關(guān)。但體內(nèi)神經(jīng)甾體激素及其中樞神經(jīng)系統(tǒng)的調(diào)控變化與此類抑郁癥相關(guān)性的認(rèn)識尚未統(tǒng)一，現(xiàn)有的研究尚不足以揭示此類疾病發(fā)生的實質(zhì)。對激素與相關(guān)情緒變化相關(guān)關(guān)系的研究，將對揭示激素與情緒變化的機理提供新的科學(xué)依據(jù)。本實驗以此為出發(fā)點,研究嚙齒類動物鼠在不同生殖周期各項指標(biāo)的變化,分析各組它們血清孕激素含量與不同腦區(qū)突觸外GABAA受體δ亞基的表達(dá)的相關(guān)性。 第一部分：雌性小鼠不同生殖周期大腦皮層和海馬突觸外GABAA受體的表達(dá)研究 目的探究雌性小鼠不同動情周期動物行為學(xué)的變化，不同生殖周期皮層及海馬突觸外GABAA受體的表達(dá)的變化。 方法以6周齡有完整動情周期雌性C57BL/6J小鼠實驗對象,采用陰道涂片法,每天分8:00、16:00和22:00三個時間段進(jìn)行動情周期的鑒別,以精確判斷小鼠動情周期,篩選出具有完整生殖周期的小鼠，確定其每個周期的持續(xù)時間。并將其分為兩組,即動情期組與動情間期組;Elisa檢測每組小鼠血清雌孕激素含量;高架十字迷宮實驗觀察每組小鼠的行為學(xué)變化;免疫組織化學(xué)法觀察每組小鼠海馬突觸外GABAA受體δ亞基的表達(dá);分析各組小鼠血清孕激素含量與突觸外GABAA受體δ亞基的表達(dá)的相關(guān)性。 結(jié)果Elisa結(jié)果顯示,動情間期組雌鼠血清孕酮含量（12.47±1.89ng/mL）高于動情期組（6.02±2.20ng/mL）,而雌激素水平動情間期（157.00±21.30pg/mL）低于動情期組（185.48±35.42pg/mL）;本研究室使用的小鼠高架十字迷宮實驗顯示，動情期組與動情間期組小鼠在高架十字迷宮開臂、閉臂以及中央平臺的活動總的活動時間無明顯差異，但是在開臂中小鼠活動的次數(shù)和時間，動情間期組高于動情期組,差異具有統(tǒng)計學(xué)意義(P＜0.05）;免疫組化結(jié)果顯示,海馬腦區(qū)GABAA受體δ亞基的陽性表達(dá)動情間期組（28.93×10~(-3)±4.92×10~(-3)）高于動情期組（18.89×10~(-3)±3.74×10~(-3)）較強,差異具有統(tǒng)計學(xué)意義(P＜0.05），，大腦皮層GABAA受體δ亞基陽性表達(dá)較強,但動情間期組（39.84×10~(-3)±2.97×10~(-3)）與動情期組（37.31×10~(-3)±1.13×10~(-3)）無明顯差異（P0.05）。動情間期與動情期海馬δ亞基陽性結(jié)果IOD值與體內(nèi)孕酮濃度的變化呈顯著性相關(guān),相關(guān)系數(shù)為r=0.9924。結(jié)論1.C57BL/6J小鼠平均動情周期為5.3天,小鼠血液雌激素濃度動情期組高于動情間期組，血液孕酮濃度動情間期組高于動情期組，雌、孕酮在小鼠體內(nèi)隨卵巢變化呈周期性的變化。2.在小鼠生理狀況下,不同生殖周期雌孕激素水平會發(fā)生波動,進(jìn)而影響GABAA受體δ亞基在不同生殖周期特定腦區(qū)的表達(dá)。海馬腦區(qū)突觸外GABAA受體δ亞基的表達(dá)在動情期低,而動情間期高,且突觸外δ亞基的表達(dá)隨孕酮濃度的增高而加強,兩者呈線性相關(guān)。3.大腦皮層突觸外GABAA受體δ亞基的表達(dá)在動情間期與動情期無明顯差異。4.行為學(xué)實驗顯示：動情間期小鼠在高架十字迷宮上進(jìn)入開臂的時間和次數(shù)大于動情期小鼠，動情間期小鼠對環(huán)境變化的適應(yīng)性增高。 第二部分：大鼠大腦海馬齒狀回、杏仁核突觸外GABAA受體表達(dá)與生殖周期雌孕激素變化相關(guān)性研究 目的明確生殖周期中卵巢甾體激素波動是否引發(fā)特定腦區(qū)神經(jīng)元突觸外GABAA受體表達(dá)變化及相關(guān)性。方法陰道涂片法篩選出具有完整生殖周期的大鼠，并將其分為兩組，動情期組與動情間期組；Elisa檢測每組大鼠血清雌孕激素含量；免疫組化法觀察每組大鼠海馬齒狀回、杏仁核突觸外GABAA受體δ亞基的表達(dá)；分析各組大鼠血清孕激素含量與突觸外GABAA受體δ亞基的表達(dá)的相關(guān)性。結(jié)果血清雌激素含量動情期組(42.51±2.42)高于動情間期組(23.17±1.45)，而孕酮含量動情期組(7.41±2.86)則低于動情間期組(10.31±1.74)，組間差異具有統(tǒng)計學(xué)意義(P0.05)。免疫組化結(jié)果顯示，海馬齒狀回GABAA受體δ亞基的陽性表達(dá)動情間期組(105.47±4.73)高于動情期組(79.23±2.89)，差異具有統(tǒng)計學(xué)意義(P0.05),杏仁核GABAA受體δ亞基的陽性表達(dá)動情間期(93.51±3.54)組高于動情期組(58.92±1.47)，差異具有統(tǒng)計學(xué)意義(P0.05)；海馬齒狀回、杏仁核突觸外δ亞基陽性結(jié)果IOD值隨著動情間期與動情期孕酮濃度值的增加而增加,兩變量呈線性相關(guān),相關(guān)系數(shù)分別為R=0.9866、R1=0.9876。結(jié)論動情間期高水平的孕激素可致大腦海馬齒狀回、杏仁核突觸外GABAA受體高表達(dá)。
[Abstract]:The female reproductive cycle has periodic changes of estrogen and progesterone, especially in the pregnancy process and the large changes in the hormone after childbirth, which make all kinds of emotional disorders and various clinical symptoms in women. Some clinical symptoms are associated with the effect of progestin on the GABAA regulation system. However, the regulation of neurosteroid and its central nervous system in the body has not been unified with this kind of depression. The present study is not enough to reveal the essence of this kind of disease. This study will provide a new scientific basis for revealing the mechanism of hormonal and emotional changes. In this experiment, the changes in the various reproductive cycle indices of rodents and the correlation between the serum progesterone content and the expression of the extracellular GABAA receptor delta subunit in different brain regions were analyzed.
Part one: the expression of GABAA receptor in the cerebral cortex and hippocampus of female mice in different reproductive cycles
Objective to explore the behavioral changes of female mice in different estrous cycles, and the expression of GABAA receptor in the cortex and hippocampus of different reproductive cycles.
Methods the estrous cycle was identified by vaginal smear method with 6 weeks old estrous cycle female C57BL/6J mice. The estrous cycle was identified by 8:00,16:00 and 22:00 every day. In order to accurately determine the estrous cycle of mice, the mice with complete reproductive cycle were screened and the duration of each cycle was determined, and it was divided into two groups. The levels of estrus and estrus were observed in the estrus group and the estrus interval group; Elisa was used to detect the serum estrogen and progesterone content in each group of mice; the elevated cross maze test was used to observe the behavioral changes in each group of mice; the expression of the GABAA receptor delta subunit in the hippocampus of the hippocampus in each group of mice was observed by immunohistochemistry, and the content of progestin and the extracellular GABAA receptor delta subunit in the blood of each group were analyzed. The correlation of expression.
Results the results of Elisa showed that the content of progesterone in the serum of estrous group (12.47 + 1.89ng/mL) was higher than that of the estrus group (6.02 + 2.20ng/mL), while the estrus level (157 + 21.30pg/mL) was lower than that of the estrus group (185.48 + 35.42pg/mL). The mice elevated cross maze test in the study room showed that the estrus group was smaller than the estrus interval group. There was no significant difference in the total activity time between the open arm of the elevated cross maze, the closed arm and the central platform, but the number and time of the activity of mice in the open arm were higher than that in the estrus group. The difference was statistically significant (P < 0.05). The immunohistochemical results showed that the positive expression of the GABAA receptor delta subunit in the hippocampus was between the estrus and the estrus. The period group (28.93 x 10~ (-3) + 4.92 x 10~ (-3)) was higher than that of the estrus group (18.89 x 10~ (-3) + 3.74 x 10~ (-3)), and the difference was statistically significant (P < 0.05), and the positive expression of the GABAA receptor delta subunit in the cerebral cortex was stronger, but there was no significant difference between the estrus interval group (39.84 x 10~ (-3) + 2.97 * 2.97). The IOD value of hippocampal delta subunit positive in estrus and estrus was significantly correlated with the changes in the concentration of progesterone in the body. The correlation coefficient was r=0.9924. conclusion that the average estrous cycle of 1.C57BL/6J mice was 5.3 days, the estrus concentration in the mice was higher than that in the estrus interval group, and the concentration of the estrone concentration in the blood was higher than that in the estrus group, the female and the pregnant women. The variation of ketone in the mice changes periodically with the changes of the ovary.2.. In the physiological state of the mice, the levels of estrogen and progesterone fluctuate in the different reproductive cycle, and then affect the expression of the GABAA receptor delta subunit in the specific brain regions of the reproductive cycle. The expression of the extracellular GABAA receptor delta subunit in the hippocampus is low in the estrous stage and high in the interval of the oestrus. The expression of the extracellular delta subunit was enhanced with the increase of progesterone concentration, and there was a linear correlation between the expression of the extracellular GABAA receptor delta subunit in the.3. cerebral cortex and no significant difference between the estrus and the estrus. The.4. behavior test showed that the time and number of time and times of entering the arm of the estrus cross maze in the elevated cross maze of the estrus mice were greater than that of the estrus mice and the estrus. The adaptability of interphase mice to environmental changes was increased.
The second part: the correlation between the expression of GABAA receptor in the amygdaloid nucleus and the reproductive cycle, estrogen and progesterone in the rat dentate gyrus.
Objective to determine whether the ovarian steroid hormone fluctuations in the reproductive cycle lead to the change and correlation of the expression of extracellular GABAA receptor in specific brain neurons. Methods rats with complete reproductive cycle were screened by vaginal smear method, and were divided into two groups, estrus group and estrus interval group, and Elisa in each group of rats serum estradiol and progesterone content; Immunohistochemical method was used to observe the expression of GABAA receptor delta subunit in the dentate gyrus and the amygdala in the amygdala. The correlation between the serum progesterone content and the expression of the GABAA receptor delta subunit in the rats was analyzed. Results the serum estrogen content in the estrous group was (42.51 + 2.42) higher than that of the estrus interval (23.17 + 1.45), while the progesterone content was estrous. The period group (7.41 + 2.86) was lower than that of the estrus interval (10.31 + 1.74), and the difference between the groups was statistically significant (P0.05). The immunohistochemical results showed that the positive expression of the GABAA receptor delta subunit in the dentate gyrus of the hippocampus was (105.47 + 4.73) higher than that of the estrous phase group (79.23 + 2.89), and the difference was statistically significant (P0.05), and the GABAA receptor delta subunit of the amygdala was in the amygdala. The positive expression of the estrus interval (93.51 + 3.54) was higher than that of the estrus group (58.92 + 1.47), the difference was statistically significant (P0.05), the hippocampal dentate gyrus and the IOD value of the extracellular delta subunit of the amygdala increased with the increase of the concentration of progesterone in estrus and estrus, and the two variables were linear correlation, and the correlation coefficients were R=0.9866, R1=0.9876. Conclusion high estrus hormone can induce high expression of extra synaptic GABAA receptor in amygdaloid nucleus of dentate gyrus.
【學(xué)位授予單位】：寧夏醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2012
【分類號】：R749.4

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