【摘要】：背景長(zhǎng)期過(guò)量飲酒對(duì)人體造成多器官、復(fù)合性嚴(yán)重危害,如心肌病變、造血功能抑制、消化系統(tǒng)潰瘍和腫瘤,更為嚴(yán)重的對(duì)神經(jīng)系統(tǒng)和生殖系統(tǒng)造成不可逆的損傷。孕齡婦女?dāng)z入酒精除對(duì)自身產(chǎn)生損害外還會(huì)對(duì)腹中胎兒產(chǎn)生嚴(yán)重、不可挽回的損傷。研究發(fā)現(xiàn)孕期攝入酒精降低子代低氧條件下攝取氧的能力；大鼠孕期攝入酒精降低新生大鼠延髓離體腦片呼吸節(jié)律性放電。這樣的實(shí)驗(yàn)結(jié)果發(fā)現(xiàn)孕期酒精暴露抑制由各級(jí)呼吸中樞的共同參與調(diào)節(jié)的動(dòng)物整體水平上的呼吸功能,產(chǎn)生基本呼吸節(jié)律性放電的延髓呼吸中樞也受到了抑制�；竟�(jié)律性呼吸起源于延髓呼吸中樞,前包欽格復(fù)合體(pre-Botzinger complex, pre-BotC)位于延髓腹外側(cè)區(qū),是延髓基本節(jié)律性呼吸產(chǎn)生的關(guān)鍵部位。多巴胺是哺乳動(dòng)物體內(nèi)廣泛存在的兒茶酚胺類神經(jīng)遞質(zhì),調(diào)節(jié)著多種生理活動(dòng),在中樞神經(jīng)系統(tǒng)內(nèi)多巴胺通過(guò)多巴胺受體參與情緒、運(yùn)動(dòng)、攝食、饑餓、內(nèi)分泌等生理活動(dòng)。現(xiàn)已發(fā)現(xiàn)延髓神經(jīng)元細(xì)胞膜上存在多巴胺D2受體,但該受體是否參與孕期攝入酒精對(duì)新生子代延髓呼吸中樞的抑制作用的研究尚未見(jiàn)報(bào)道,因此我們?cè)O(shè)計(jì)本實(shí)驗(yàn),利用神經(jīng)電生理和分子生物學(xué)的實(shí)驗(yàn)方法研究多巴胺受體2是否參與該抑制作用及其可能機(jī)制。目的研究孕期酒精暴露對(duì)新生大鼠延髓呼吸中樞節(jié)律性基本呼吸放電(rhythmic respiratory discharge activities, RRDA)的作用和多巴胺D2受體在其中的作用。方法對(duì)成年健康Sprague-Dawley大鼠進(jìn)行孕期酒精暴露,自合籠前30d至實(shí)驗(yàn)時(shí)飼喂大鼠自主飲用且只飲用8%酒精水溶液；對(duì)照大鼠常規(guī)飼養(yǎng)。本實(shí)驗(yàn)使兩組用2d新生大鼠,雌雄不拘。1.RRDA記錄：制作2d新生大鼠延髓離體腦片,使用吸附電極在腦片腹側(cè)面吸附舌下神經(jīng)根,舌下神經(jīng)根傳出的電活動(dòng)即為反映了延髓呼吸功能的節(jié)律性基本呼吸放電。在2組新生大鼠腦片灌流液中加入多巴胺受體2特異性激動(dòng)劑Quinpirole和特異性拮抗劑Raclopride,觀察Quinpirole和IRaclopride對(duì)兩組RRDA的作用差異,以明確孕期酒精暴露是否改變多巴胺2受體對(duì)新生大鼠延髓離體腦片節(jié)律性基本呼吸放電作用效果。2.將2d新生小鼠延髓離體腦片pre-BotC區(qū)切下,采用Western Blot檢驗(yàn)多巴胺D2受體在pre-BotC區(qū)神經(jīng)元上的表達(dá),研究孕期酒精暴露對(duì)受體表達(dá)的作用。3.將2d新生小鼠延髓離體腦片pre-BotC區(qū)切下,采用qRT-PCR-技術(shù)檢測(cè)多巴胺D2受體mRNA在延髓呼吸中樞pre-BotC區(qū)神經(jīng)元上的表達(dá),研究孕期酒精暴露對(duì)受體mRNA表達(dá)的作用。結(jié)果1.對(duì)照組和酒精暴露組新生大鼠延髓離體腦片呼吸節(jié)律性放電在60min內(nèi)各時(shí)間點(diǎn)之間無(wú)統(tǒng)計(jì)學(xué)改變,實(shí)驗(yàn)?zāi)Ｐ统闪�。多巴胺D2受體激動(dòng)劑Quinpirole抑制對(duì)照組和酒精暴露組腦片呼吸節(jié)律性放電,對(duì)對(duì)照組的抑制效果強(qiáng)于實(shí)驗(yàn)組；多巴胺D2受體拮抗劑Raclopride對(duì)兩組新生大鼠延髓離體腦片呼吸節(jié)律性放電都有興奮作用,酒精暴露組放電變化程度低于對(duì)照組。2. Western Blot實(shí)驗(yàn)發(fā)現(xiàn),孕期酒精暴露降低新生大鼠延髓呼吸中樞pre-BotC區(qū)神經(jīng)元多巴胺D2受體表達(dá)水平。3. qRT-PCR實(shí)驗(yàn)顯示,與對(duì)照組相比,孕期酒精暴露降低新生大鼠延髓呼吸中樞pre-BotC區(qū)神經(jīng)元多巴胺D2受體mRNA表達(dá)水平。結(jié)論1.孕期酒精暴露抑制新生大鼠延髓腦片節(jié)律性基本呼吸放電,降低多巴胺D2受體對(duì)腦片節(jié)律性基本呼吸放電的調(diào)節(jié)作用。2.孕期酒精暴露下調(diào)延髓前包欽格復(fù)合體神經(jīng)元多巴胺D2受體水平和受體mRNA表達(dá)水平,這可能是多巴胺D2受體參與孕期酒精暴露抑制子代延髓腦片律性基本呼吸放電機(jī)制之一。
[Abstract]:Background: Long-term excessive drinking can cause multiple organs and compound serious harm to the human body, such as cardiomyopathy, hemopoietic function inhibition, digestive system ulceration and tumor, and more serious irreversible damage to the nervous system and the reproductive system. The intake of alcohol in pregnant women may also cause serious and irreversible damage to the fetus in the abdomen, in addition to its own damage. The study found that the intake of alcohol during pregnancy decreased the ability of oxygen to be taken under the low-oxygen condition of the offspring, and the intake of alcohol during pregnancy decreased the respiratory rhythm of the medulla oblongata of the neonatal rats. Such experimental results showed that during pregnancy, alcohol exposure inhibited the respiratory function at the whole level of the animal regulated by the co-participation of the respiratory center at all levels, and the medulla oblongata respiratory center, which produced the basic respiratory rhythm discharge, was also inhibited. The basic rhythmic breathing originates from the medulla oblongata respiratory center, and the pre-Botzinger complex (pre-BopoC) is located in the lateral medulla of the medulla oblongata, which is the key part of the basic rhythm of the medulla oblongata. Dopamine is a widely available catecholamine neurotransmitter in a mammal, which regulates a variety of physiological activities in which dopamine is involved in physiological activities such as mood, exercise, food consumption, hunger, and endocrine. It has been found that there is a dopamine D2 receptor on the cell membrane of the medulla oblongata, but the study of whether the receptor is involved in the inhibition of the uptake of alcohol during pregnancy on the respiratory center of the medulla oblongata is not reported, so we design this experiment, Whether the dopamine receptor 2 is involved in the inhibition and its possible mechanism is studied by means of the experimental methods of neuroelectrophysiology and molecular biology. Objective To study the effects of alcohol exposure during pregnancy on the basic respiratory discharge (RRDA) and the role of dopamine D2 receptor in neonatal rats. Methods The adult healthy Sprague-Dawley rats were exposed to alcohol during pregnancy. In this experiment, two groups of newborn rats were divided into two groups. The first. RRDA records: to make 2d neonatal rat's medulla oblongata, and to use the adsorption electrode to adsorb the hypoglossal nerve roots on the ventral side of the brain, and the electrical activity from the root of the hypoglossal nerve is the basic respiratory discharge that reflects the respiratory function of the medulla oblongata. The effect of Quinpirole and IRaclopride on the two groups of RRDA was observed by the addition of a dopamine receptor 2-specific agonist, Quinpirole, and a specific antagonist, Racopride, to the brain slices of the 2-group neonatal rats. in order to determine whether the exposure of the alcohol to the dopamine-2 receptor changes the effect of the dopamine-2 receptor on the basic respiratory discharge of the medulla oblongata in the newborn rats. The expression of the dopamine D2 receptor in the pre-BopoC region was examined by Western Blot, and the effect of alcohol exposure on the expression of the receptor was studied. The expression of dopamine D2 receptor mRNA in the pre-BopoC region of the medulla oblongata was detected by the qRT-PCR-technique, and the effect of alcohol exposure on the expression of the receptor mRNA was studied. Results 1. In the control group and the alcohol exposure group, the respiratory rhythm of the medulla oblongata was not statistically changed between the time points within 60min, and the experimental model was established. in that control group, the dopamine D2 receptor agonist, Quinn pirole, inhibited the respiratory rhythm of the brain slices of the control group and the alcohol exposure group, and the inhibition effect on the control group was stronger than that of the experimental group; the dopamine D2 receptor antagonist Racopride has an excitation effect on the respiratory rhythm of the medulla oblongata of the two groups of newborn rats, The degree of discharge of the alcohol exposure group was lower than that of the control group. Western Blot experiment found that alcohol exposure during pregnancy decreased the level of dopamine D2 receptor expression in the pre-BopoC region of the neonatal rat's medulla oblongata. The qRT-PCR showed that the level of the expression of dopamine D2 receptor mRNA in the pre-BopoC region of the medulla oblongata of the neonatal rats was reduced by alcohol exposure during pregnancy as compared to the control group. Conclusion 1. The effect of the dopamine D2 receptor on the basic respiratory discharge of the brain slices was reduced by the basic respiratory discharge in the brain of the neonatal rat by alcohol exposure during pregnancy. The level of the dopamine D2 receptor and the expression of the receptor mRNA in the neurons of the submedulla oblongata complex are reduced by alcohol exposure during pregnancy, which may be one of the basic respiratory discharge mechanisms of the dopamine D2 receptor involved in the inhibition of alcohol exposure during pregnancy.
【學(xué)位授予單位】：新鄉(xiāng)醫(yī)學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R714

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