本文選題：突觸帶　切入點：發(fā)育　出處：《南方醫(yī)科大學》2017年碩士論文

【摘要】：目的了解耳蝸突觸帶發(fā)育的時間特點和空間分布,明確C57BL/6j小鼠突觸帶穩(wěn)定的時間,探討長期中等強度噪聲暴露耳蝸損傷的病理機制。方法(1)全耳蝸基底膜鋪片、免疫熒光染色后,通過共聚焦顯微鏡觀察、記錄不同發(fā)育階段耳蝸毛細胞突觸前膜上突觸帶的發(fā)育與分布;(2)應用免疫熒光、HE染色、耳蝸電鏡掃描評估耳蝸除帶狀突觸之外的結構成分的變化;(3)中等強度白噪聲,70dBSPL,8h/天,持續(xù)暴露3月,建造模擬環(huán)境噪聲慢性暴露的動物模型;(4)采用ABR檢測評估聽覺閾值和誘發(fā)電位Ⅰ波幅度;(5)應用小鼠耳蝸音頻定位圖,建立全耳蝸突觸帶與聽覺頻譜之間的對應關系。結果(1)C57小鼠出生后Od,在內外毛細胞的胞質即可觀察到CtBP2的信號表達;出生后第6天,CtBP2的信號數目在內外毛細胞的胞質和基底側膜上的表達均達到高峰,突觸帶信號的增長在外毛細胞上則表現得尤為明顯;出生后第12-60d,內外毛細胞上的CtBP2信號數目急劇減少并逐漸趨于穩(wěn)定,且在毛細胞上的分布逐漸向基底側膜聚集。功能學上,ABR可檢測聽力出現于出生后13.2天;(2)內毛細胞上突觸帶富集于耳蝸中間區(qū)域(距頂轉50%-70%),可達18.6個/細胞,明顯高于突觸帶在耳蝸頂轉10%-20%(7.4個/細胞)和底轉90%-100%(9.6個/細胞)區(qū)域的分布。外毛細胞沿耳蝸基底膜螺旋分布未見明顯差異,1-3個/細胞;(3)中等強度噪聲暴露3個月,在聽覺頻譜范圍內,各頻段均有不同程度的聽力減退,且以8-16kHz頻段閾值及ABRⅠ波幅度下降明顯;(4)借助耳蝸音頻定位,突觸帶計數顯示,沿耳蝸螺旋突觸帶數目廣泛減少,且在距耳蝸頂轉50-70%區(qū)域突觸帶減少明顯高于其他區(qū)域。(5)與對照組相比,毛細胞計數、靜纖毛排列、螺旋神經元計數未見明顯差異。結論(1)幼年C57小鼠,毛細胞與蝸神經纖維的突觸聯(lián)系出生前即已存在。出生后12d前,突觸帶經歷數目和空間分布上的劇烈變化,12d后逐漸趨于完善和功能上的成熟;出生13.2d后,小鼠開始出現聽力。因此,突觸帶成熟的時間和小鼠聽覺發(fā)生時間的吻合,提示功能成熟的突觸帶是聽覺發(fā)生的前提之一;(2)60日齡的C57小鼠,突觸帶沿耳蝸的分布趨于穩(wěn)定;(3)長期中等強度噪聲暴露可致在多個頻率上不同程度的聽閾提高,且暴露對聽覺敏感區(qū)(8-16kHz)影響最為顯著,其潛在的病理機制可能是原發(fā)性的內毛細胞與聽覺傳入纖維連接的大量減少所致。
[Abstract]:Objective to understand the distribution characteristics of time and space with the development of cochlear synapses, clear synaptic C57BL/6j mice with stable time, to investigate the pathologic mechanism of long-term moderate intensity noise exposure in cochlear injury. Methods (1) the basilar membrane preparations, immunofluorescence staining, confocal microscopy through recording, development and distribution of different synaptic ribbon developmental stages of cochlear hair cells in the presynaptic membrane; (2) by immunofluorescence, HE staining, scanning electron microscopy evaluation of cochlear structure changes of cochlear ingredients except outside the ribbon synapse; (3) moderate intensity white noise, 70dBSPL, 8h/ days, continuous exposure in March, the animal model of chronic exposure to environmental noise building simulation; (4) using ABR detection and evaluation of auditory threshold and evoked potential amplitude of 1; (5) using the mouse cochlear audio location map, establish the whole cochlear and auditory synapse with spectral correspondence between results. (1) C57 mice Od , to CtBP2 signal was observed on the inner and outer hair cells in the cytoplasm can be expressed; sixth days after birth, the number of signals in the CtBP2 expression of outer hair cell cytoplasm and the basolateral membrane of synaptic growth reached the peak, band signals in outer hair cells is particularly evident; 12-60d after birth. The number of CtBP2 signal on the inner and outer hair cells decreased sharply and gradually stabilized, and the distribution in the hair cells gradually to the basolateral membrane aggregation. Functionally, ABR can detect the hearing in 13.2 days after birth; (2) the inner hair cells on the synaptic ribbon enriched in the region in the cochlea (between from the top to 50%-70%), up to 18.6 per cell, was significantly higher than that of 10%-20% in the apical synaptic ribbon (7.4 / cell) and bottom 90%-100% (9.6 / cell) region. The distribution of the outer hair cells along the basilar membrane of the cochlea spiral distribution showed no significant difference, 1-3 / cells; (3) the medium intensity noise exposure 3 April, in the auditory frequency range, the band had different degrees of hearing loss, and to the 8-16kHz band and ABR 1 threshold amplitude decreased significantly; (4) with the aid of cochlear audio localization, synaptic counting display, along the spiral belt and reduce the number of synapses widely, with reduced significantly higher than other regions from the apical turn 50-70% synapses. (5) compared with the control group, hair cell counting, stereocilia arranged, spiral neurons count showed no significant difference. Conclusion (1) in young C57 mice, synaptic connections between hair cells and cochlear nerve fibers before birth has saved in. After the birth of 12D, with changing the number and synaptic space the distribution of 12D tends to improve gradually and functionally mature; born after 13.2d, the mice started listening. Therefore, take time to mature mice and synaptic auditory time with synaptic functional maturation with auditory occurrence One of the premises; (2) the 60 day old C57 mice, with synaptic distribution along the cochlea tends to be stable; (3) long-term medium intensity noise exposure can cause at multiple frequencies with different degrees of exposure on the auditory threshold was increased, and the sensitive area (8-16kHz) has the most significant impact, the underlying pathogenesis may be primary inner hair cells and auditory afferent fibers connecting the large decrease.

【學位授予單位】：南方醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R764.43
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本文編號：1657541