工頻電磁環(huán)境對(duì)果蠅發(fā)育和節(jié)律行為的影響研究

鄒鑫平,王曉麗,端正花,張紫燕,孫永彥,3*

工頻電磁環(huán)境對(duì)果蠅發(fā)育和節(jié)律行為的影響研究

鄒鑫平1,王曉麗1,端正花1,張紫燕2,3,孫永彥1,3*

(1.天津理工大學(xué)環(huán)境科學(xué)與安全工程學(xué)院,天津 300384；2.中國(guó)科學(xué)院城市環(huán)境研究所,福建 廈門 361021；3.中國(guó)科學(xué)院上海營(yíng)養(yǎng)與健康研究所,上海 200031)

比較了PF-EMF暴露下W1118野生型和突變型果蠅發(fā)育、睡眠和運(yùn)動(dòng)節(jié)律行為的差異.連續(xù)暴露處理3d后去除親本果蠅,記錄子代發(fā)育過(guò)程并監(jiān)測(cè)運(yùn)動(dòng)和睡眠節(jié)律行為.結(jié)果表明,PF-EMF暴露后W1118型成蛹量、成蠅量分別顯著提高56.2%和57.5%,型果蠅成蛹量無(wú)顯著差異,成蠅量顯著增加85.2%,子代雌雄比例未顯著變化.從行為數(shù)據(jù)看,兩株系果蠅暴露后運(yùn)動(dòng)和睡眠節(jié)律相位未見顯著差異,W1118型表現(xiàn)為24h平均運(yùn)動(dòng)次數(shù)增加37.3%,24h睡眠片段次數(shù)減少10.6%,型24h平均運(yùn)動(dòng)次數(shù)顯著增加50.0%,24h睡眠片段次數(shù)減少51.5%,總睡眠時(shí)長(zhǎng)顯著降低45.7%.和W1118型相比,型果蠅發(fā)育、運(yùn)動(dòng)、睡眠情況改變更顯著,提示型果蠅電磁暴露更為敏感.總之PF-EMF環(huán)境對(duì)昆蟲發(fā)育和節(jié)律行為均可帶來(lái)影響,造成后代數(shù)量上升、顯著活躍果蠅運(yùn)動(dòng)行為并引起睡眠障礙.本研究為進(jìn)一步了解電磁暴露對(duì)昆蟲類生物的影響,合理評(píng)估PF-EMF的生態(tài)效應(yīng)提供依據(jù).

工頻電磁場(chǎng)；黑腹果蠅；節(jié)律行為；發(fā)育

工頻電磁場(chǎng)(Power frequency electromagnetic field,PF-EMF)指電力傳輸線路、設(shè)施產(chǎn)生的頻率在50Hz的電磁場(chǎng).隨著我國(guó)用電量激增,大規(guī)模戶外輸變電工程產(chǎn)生的人造電磁場(chǎng)改變了電磁環(huán)境背景[1],對(duì)動(dòng)植物乃至生態(tài)系統(tǒng)產(chǎn)生影響.例如一些農(nóng)業(yè)活動(dòng)中關(guān)注的昆蟲類動(dòng)物如蜜蜂、蚜蟲等,已有報(bào)道顯示其對(duì)電磁環(huán)境敏感,一些電磁技術(shù)在害蟲防治方面得到了應(yīng)用[2].英國(guó)研究人員的一項(xiàng)Meta分析顯示,WiFi、移動(dòng)通信網(wǎng)絡(luò)和電線構(gòu)成的全球通訊系統(tǒng)所產(chǎn)生的電磁輻射可能會(huì)擾亂鳥類和昆蟲的方向感和運(yùn)動(dòng),并影響其代謝活動(dòng)[3].同時(shí)也有部分研究者認(rèn)為PF-EMF暴露不會(huì)對(duì)昆蟲的生活史產(chǎn)生顯著影響[4].鑒于電氣化時(shí)代背景下PF-EMF環(huán)境農(nóng)業(yè)生態(tài)效應(yīng)評(píng)估仍不充分[5],為應(yīng)對(duì)該形勢(shì),本研究擬利用模式生物黑腹果蠅(,以下簡(jiǎn)稱“果蠅”)來(lái)分析PF-EMF對(duì)昆蟲類生物發(fā)育和節(jié)律行為的影響.已有報(bào)道顯示昆蟲作為數(shù)量龐大的生物類群對(duì)各種電磁環(huán)境變化和脅迫敏感.例如將粘蟲放置于亥姆赫茲線圈產(chǎn)生的強(qiáng)磁場(chǎng)中時(shí),粘蟲群體共同定向行為消失[6].關(guān)于電磁場(chǎng)對(duì)果蠅的影響迄今研究并不廣泛深入.我們前期探索發(fā)現(xiàn)PF-EMF聯(lián)合高溫脅迫環(huán)境下果蠅體內(nèi)熱激蛋白表達(dá)水平發(fā)生響應(yīng),PF-EMF暴露可提高果蠅對(duì)溫度的耐受能力,緩解高溫環(huán)境對(duì)果蠅的損傷效應(yīng)[7].對(duì)于電氣化背景下輸變電工程帶來(lái)的PF-EMF電磁場(chǎng)的昆蟲生物學(xué)效應(yīng)目前來(lái)講還未有明確定論.本文以果蠅為研究對(duì)象來(lái)探究PF-EMF的生物效應(yīng)具有創(chuàng)新性.一是果蠅具備昆蟲類生物的明顯特征,具有繁殖快、后代數(shù)目多、發(fā)育過(guò)程性狀明顯等優(yōu)點(diǎn);二是果蠅的節(jié)律行為具有代表性,其運(yùn)動(dòng)和睡眠節(jié)律等行為指標(biāo)可在實(shí)驗(yàn)室條件下進(jìn)行檢測(cè),并且參與果蠅睡眠調(diào)控的生物鐘基因十分保守,與哺乳動(dòng)物有同源相似性,可作為開展節(jié)律行為研究的常用模型[8].本實(shí)驗(yàn)利用野生型W1118和突變體模型果蠅來(lái)研究PF-EMF暴露對(duì)昆蟲發(fā)育、睡眠和運(yùn)動(dòng)節(jié)律行為的影響,基因又名red eye 基因,編碼煙堿乙酰膽堿受體α4(nAChRα4)復(fù)合物的四通道跨膜通道蛋白.已知當(dāng)突變后,四通道跨膜通道蛋白功能異常造成果蠅睡眠減少、睡眠碎片化[9].本實(shí)驗(yàn)可為明確PF-EMF的昆蟲生物學(xué)效應(yīng),明確和制定合理的PF-EMF暴露生態(tài)健康風(fēng)險(xiǎn)防控規(guī)范提供理論支持.

1 材料與方法

1.1 果蠅培養(yǎng)

野生型黑腹果蠅() W1118品系和突變型黑腹果蠅12414R3品系,購(gòu)自中國(guó)科學(xué)院上海生命科學(xué)研究院生化細(xì)胞所果蠅資源與技術(shù)平臺(tái).以玉米粉-蔗糖-酵母為基本培養(yǎng)基,培養(yǎng)于光照周期12L:12D(06:00開燈,18:00關(guān)燈),溫度(25±1)℃,相對(duì)濕度(60±1)%的人工氣候箱中.

1.2 工頻電磁場(chǎng)暴露艙

PF-EMF實(shí)驗(yàn)暴露艙如圖1所示,主要利用交流變頻電源、低溫恒溫循環(huán)器和亥姆霍茲線圈來(lái)營(yíng)造一個(gè)穩(wěn)定可調(diào)控的極低頻電磁場(chǎng).亥姆霍茲線圈平放在支架上,通過(guò)電線與能夠穩(wěn)定調(diào)控輸出電壓和頻率的交流變頻電源相連,接通電源后,可在兩線圈中間形成一個(gè)穩(wěn)定的磁場(chǎng).為了實(shí)現(xiàn)對(duì)實(shí)驗(yàn)的溫濕度、光照等環(huán)境的控制,兩個(gè)亥姆霍茲線圈放于生化培養(yǎng)箱中.其能夠?qū)崿F(xiàn)頻率40～499.9Hz;磁場(chǎng)強(qiáng)度:0.01～10mT范圍內(nèi)的模擬.此次實(shí)驗(yàn)的暴露條件為:頻率50Hz、磁場(chǎng)強(qiáng)度6mT.

A:示意圖:1.生化培養(yǎng)箱2.亥姆霍茲線圈 3. 冷凝循環(huán)器 4. 溫度記錄儀5. 變頻電源;B:實(shí)物圖

1.3 PF-EMF對(duì)果蠅子代的影響實(shí)驗(yàn)

實(shí)驗(yàn)在黑暗條件下進(jìn)行,共設(shè)置4組,分別為對(duì)照-W1118組、PF-EMF-W1118組、對(duì)照-組和PF-EMF-組.收集48h內(nèi)羽化的果蠅,接種10對(duì)果蠅于新鮮培養(yǎng)基中,每組10管,暴露組培養(yǎng)于發(fā)射磁場(chǎng)的線圈中,對(duì)照組培養(yǎng)于磁場(chǎng)強(qiáng)度為0的人工培養(yǎng)箱中,連續(xù)暴露三日后去除親本,然后將帶有卵的果蠅管繼續(xù)暴露,直至最后一只子代果蠅羽化出來(lái)停止實(shí)驗(yàn),實(shí)驗(yàn)重復(fù)三次.期間統(tǒng)計(jì)各果蠅管中的成蛹數(shù)、成蠅數(shù)前三天成蛹率以及雌雄比,繪制成蛹進(jìn)程曲線,具體統(tǒng)計(jì)方法如下:

成蛹(蠅)數(shù):統(tǒng)計(jì)每管中一個(gè)生活史周期內(nèi)所出現(xiàn)的蛹或(羽化的果蠅)的數(shù)量.

前三天成蛹率:從第一個(gè)蛹出現(xiàn)開始計(jì)為第一天,統(tǒng)計(jì)前三天成蛹數(shù)量占總成蛹量比值.

雌雄比:每管中雌果蠅與雄果蠅的數(shù)量比.

1.4 PF-EMF對(duì)果蠅節(jié)律行為的影響實(shí)驗(yàn)

本實(shí)驗(yàn)選用DAM2行為監(jiān)測(cè)器(Drosophila Activity Monitor,美國(guó)TriKinetics公司)用于記錄果蠅運(yùn)動(dòng)和睡眠節(jié)律,DAM2能夠一次性測(cè)量32只果蠅樣本的運(yùn)動(dòng)行為,每只果蠅單獨(dú)放在該系統(tǒng)配套玻璃管中,當(dāng)果蠅穿過(guò)試管中點(diǎn)時(shí)就切割一次紅外光束,信號(hào)自動(dòng)傳感計(jì)數(shù)一次,被視為果蠅運(yùn)動(dòng)一次.按照1.3所述實(shí)驗(yàn)條件和分組,收集48h內(nèi)羽化的雄蠅將其裝進(jìn)監(jiān)測(cè)管中并每五分鐘記錄一次數(shù)據(jù).果蠅裝進(jìn)玻璃檢測(cè)管中適應(yīng)至少24h后再開展實(shí)驗(yàn),實(shí)驗(yàn)持續(xù)處理3d,重復(fù)3次.統(tǒng)計(jì)分析各組果蠅的運(yùn)動(dòng)和睡眠情況,計(jì)算指標(biāo)如下:

(1)平均運(yùn)動(dòng)次數(shù):統(tǒng)計(jì)每只果蠅的總運(yùn)動(dòng)次數(shù)后計(jì)算所有果蠅的總運(yùn)動(dòng)次數(shù)的平均值,分析白天、夜晚和24h(整天)三個(gè)指標(biāo).白天總運(yùn)動(dòng)次數(shù)為從上午06:00～18:00時(shí)間段內(nèi)總運(yùn)動(dòng)次數(shù)的平均值;夜晚平均運(yùn)動(dòng)次數(shù)表示從上午18:00至第二天06:00 時(shí)段;24h總運(yùn)動(dòng)次數(shù)為從下午18:00至第二天18:00全天所有果蠅總運(yùn)動(dòng)次數(shù)的平均值.

(2)單位時(shí)間運(yùn)動(dòng)量:果蠅清醒狀態(tài)每分鐘的平均運(yùn)動(dòng)次數(shù)即為單位時(shí)間運(yùn)動(dòng)量.由白天、夜晚或24h每一時(shí)間段范圍內(nèi)的總運(yùn)動(dòng)次數(shù)除以對(duì)應(yīng)時(shí)間段的總運(yùn)動(dòng)時(shí)間.計(jì)算公式為:單位時(shí)間運(yùn)動(dòng)量=每個(gè)時(shí)間段的總運(yùn)動(dòng)次數(shù)/(每個(gè)時(shí)間段總時(shí)長(zhǎng)-每個(gè)時(shí)間段的總睡眠時(shí)間).

(3)睡眠總時(shí)間:在監(jiān)測(cè)過(guò)程中,果蠅連續(xù)5min沒(méi)有穿越監(jiān)測(cè)玻璃管中間位置,運(yùn)動(dòng)次數(shù)計(jì)為0,視為果蠅處于睡眠狀態(tài)狀態(tài)[10],統(tǒng)計(jì)果蠅白天、夜晚和24h的總睡眠時(shí)長(zhǎng).

(4)睡眠片段:果蠅在各時(shí)間段睡眠片段次數(shù).分別出統(tǒng)計(jì)果蠅在白天、夜晚和24h內(nèi)的睡眠次數(shù).

2 結(jié)果與分析

2.1 PF-EMF對(duì)果蠅子代發(fā)育數(shù)量的影響

由實(shí)驗(yàn)結(jié)果圖2A可知,PF-EMF暴露后W1118型成蛹量、成蠅量分別顯著提高56.2%和57.5%(< 0.05),型果蠅成蛹量雖無(wú)顯著差異但呈現(xiàn)增加趨勢(shì),成蠅量顯著增加85.2%(<0.01),表明PF-EMF暴露可引起果蠅后代數(shù)量增加,無(wú)論是野生型還是突變型果蠅,均對(duì)其生殖有促進(jìn)作用.細(xì)胞水平研究顯示工頻電磁場(chǎng)可促進(jìn)細(xì)胞增殖和分裂,比如多次照射PF-EMF后可對(duì)正常淋巴細(xì)胞和肝細(xì)胞的增殖產(chǎn)生刺激作用[11].除交變PF-EMF外,一些學(xué)者認(rèn)為中等強(qiáng)度的靜態(tài)磁場(chǎng)(1mT～1T)能加快昆蟲胚后發(fā)育,增加昆蟲體重[12],這些證據(jù)提示PF- EMF電磁場(chǎng)可能對(duì)昆蟲繁殖具有相同的效應(yīng).從機(jī)理層面分析,一些研究提示PF-EMF促進(jìn)繁殖可能與細(xì)胞間隙連接有關(guān).間隙連接是細(xì)胞通訊的一種方式,在代謝偶聯(lián)、神經(jīng)沖動(dòng)信息傳遞、早期胚胎發(fā)育和細(xì)胞分化過(guò)程中具有重要作用.姜槐等[13]發(fā)現(xiàn)工頻磁場(chǎng)對(duì)間隙連接具有抑制或協(xié)同抑制作用,而間隙連接異常往往引起細(xì)胞間物質(zhì)交換障礙,進(jìn)而引發(fā)細(xì)胞分裂增殖異常.PF-EMF可能通過(guò)改變果蠅細(xì)胞間隙連接通訊促進(jìn)了細(xì)胞分裂,最終導(dǎo)致子代數(shù)量的增加.由實(shí)驗(yàn)結(jié)果圖2B可知,野生型和突變型兩種果蠅在PF-EMF暴露處理后子代性別比均未發(fā)生顯著差改變(圖2B,>0.05),表明PF-EMF暴露處理不會(huì)造成果蠅后代性別偏離,只引起數(shù)量增加,提示PF-EMF可能不是影響果蠅繁殖過(guò)程中性別比例的關(guān)鍵因子.一些流行病資料也證實(shí)極低頻電磁場(chǎng)波段范圍輻射因其強(qiáng)度偏低,可能對(duì)子代出生性別比例影響很小.如Saadat等[14]發(fā)現(xiàn)51名高壓架線工經(jīng)PF-EMF暴露后也未造成其后代發(fā)生顯著性別偏離,但值得注意的是這些試驗(yàn)雖無(wú)統(tǒng)計(jì)學(xué)差異,但子代中雄性比例有下降的趨勢(shì),本研究中野生型果蠅也出現(xiàn)類似趨勢(shì).結(jié)合下文中PF-EMF暴露引發(fā)了果蠅運(yùn)動(dòng)水平提高(圖4A),分析果蠅后代數(shù)量增加的原因可能還與PF-EMF提高了運(yùn)動(dòng)次數(shù)進(jìn)而增大了果蠅交配行為發(fā)生可能性相關(guān),從而提高了繁殖成功率,但未顯著影響后代性別比例.例如已有研究發(fā)現(xiàn)PF-EMF可引發(fā)蝗蟲變得活躍,飛行增加,進(jìn)而增大其成功產(chǎn)卵概率,從而提高繁殖數(shù)量[15],本實(shí)驗(yàn)研究也得出了類似的結(jié)論,這些結(jié)果啟示我們應(yīng)當(dāng)關(guān)注PF-EMF對(duì)昆蟲生殖過(guò)程的影響,對(duì)PF-EMF環(huán)境暴露的生態(tài)閾值進(jìn)一步加以管理,比如要進(jìn)一步通過(guò)實(shí)驗(yàn)研究明確PF-EMF是否會(huì)造成昆蟲的性別比例發(fā)生變化,且這種變化是否可能通過(guò)世代累積效應(yīng)顯現(xiàn),后續(xù)應(yīng)當(dāng)討論和關(guān)注.

A:成蛹量、成蠅量;B:雌雄比

*<0.05 **<0.01,下同

2.2 PF-EMF暴露對(duì)果蠅成蛹進(jìn)程的影響

由圖3A來(lái)看,PF-EMF暴露處理后,W1118果蠅前3d成蛹率顯著降低(<0.01),進(jìn)一步結(jié)合成蛹時(shí)間曲線分析可以發(fā)現(xiàn)(圖3B),前3d的成蛹量未受PF-EMF處理影響,從第11d開始,暴露組成蛹量顯著高于對(duì)照組,PF-EMF處理導(dǎo)致后代蛹量顯著增加,因此計(jì)算出的前三天成蛹率數(shù)值暴露組顯著低于對(duì)照組.分析原因可能是PF-EMF的時(shí)間累積效應(yīng).隨著暴露時(shí)間的延長(zhǎng),W1118果蠅表現(xiàn)出的響應(yīng)更明顯.PF-EMF的時(shí)間累積效應(yīng)這一結(jié)論也有其他生物證據(jù)支持.例如范純武等[16]采用50Hz, 1.5mT工頻磁場(chǎng)環(huán)境暴露健康人外周血細(xì)胞,培養(yǎng)24～96h后發(fā)現(xiàn)試驗(yàn)組淋巴細(xì)胞轉(zhuǎn)化率隨磁場(chǎng)作用時(shí)間的延長(zhǎng)而遞增,反映50Hz工頻磁場(chǎng)能對(duì)淋巴細(xì)胞產(chǎn)生損傷性生物效應(yīng), 且這種效應(yīng)隨輻射時(shí)間的延長(zhǎng)而加劇.已知PF-EMF暴露處理后昆蟲胚后發(fā)育加快,細(xì)胞增殖和分裂能力異?；钴S,在本實(shí)驗(yàn)中,果蠅成蛹進(jìn)程的加速效應(yīng)可能隨著暴露時(shí)間推移而加劇,因而導(dǎo)致了W1118前期成蛹量不變,后期急劇增加的情況.

A:前三天成蛹率;B:成蛹進(jìn)程曲線

繼續(xù)分析果蠅成蛹過(guò)程發(fā)現(xiàn),前3d的成蛹率和對(duì)照組相比降低,但未達(dá)到顯著水平(圖3A),由圖3B可知,PF-EMF暴露組的成蛹數(shù)量整體顯著多于對(duì)照組,提示PF-EMF暴露引發(fā)了整個(gè)階段內(nèi)的蛹數(shù)量增加.PF-EMF暴露主要影響W1118果蠅成蛹中后期階段,對(duì)成蛹前期無(wú)顯著作用,而PF-EMF可導(dǎo)致果蠅整個(gè)發(fā)育進(jìn)程子代數(shù)量顯著上升,表明對(duì)于PF-EMF暴露脅迫,果蠅比W1118果蠅更敏感.已知突變型生物個(gè)體存在睡眠節(jié)律障礙,睡眠比野生型水平更低,因而體質(zhì)更加羸弱[17],這可能是突變型果蠅對(duì)外界一些環(huán)境刺激比野生型更易感的原因之一,果蠅可能比野生型更難以抵御PF-EMF脅迫.從運(yùn)動(dòng)水平分析,由于果蠅睡眠減少,比野生型更活躍,結(jié)合下文中運(yùn)動(dòng)行為監(jiān)測(cè)結(jié)果,發(fā)現(xiàn)PF-EMF能促進(jìn)果蠅運(yùn)動(dòng)更加活躍(圖4A),因而果蠅其交配概率比野生型高,產(chǎn)生的蛹數(shù)量也明顯多于野生型,這也可能是在成蛹階段早期果蠅就表現(xiàn)出高成蛹水平的原因.

2.3 PF-EMF對(duì)果蠅節(jié)律行為的影響

2.3.1 PF-EMF對(duì)果蠅運(yùn)動(dòng)行為的影響 實(shí)驗(yàn)中對(duì)果蠅運(yùn)動(dòng)行為的監(jiān)測(cè)結(jié)果顯示,PF-EMF暴露處理后無(wú)論是W1118還是果蠅其運(yùn)動(dòng)水平明顯提高(圖4A,*<0.05,**<0.01),野生型果蠅夜間運(yùn)動(dòng)次數(shù)顯著增多,24h全天運(yùn)動(dòng)次數(shù)顯著提升37.3%,白天運(yùn)動(dòng)次數(shù)雖升高但未出現(xiàn)明顯差異,表明PF-EMF可導(dǎo)致野生型果蠅夜間異常活躍,提示PF-EMF對(duì)昆蟲的運(yùn)動(dòng)節(jié)律存在潛在危害.已有報(bào)道表示,PF-EMF可以顯著增加蝗蟲翼拍頻率,蝗蟲種群表現(xiàn)更加活躍,飛行擴(kuò)散范圍顯著增加[18].蜜蜂暴露于電磁場(chǎng)后展現(xiàn)出更加積極的攻擊行為[19],這些結(jié)果與本實(shí)驗(yàn)中果蠅PF-EMF暴露后造成運(yùn)動(dòng)頻繁十分相似.認(rèn)為PF-EMF作為一種環(huán)境脅迫刺激,可能使昆蟲產(chǎn)生壓力應(yīng)激反應(yīng).本實(shí)驗(yàn)中,果蠅為應(yīng)對(duì)PF-EMF暴露環(huán)境壓力進(jìn)而表現(xiàn)活躍,具體分析分子機(jī)制可能與PF-EMF誘導(dǎo)熱激蛋白(Heat Shock Proteins,HSPs)表達(dá)相關(guān).熱激蛋白又稱作熱休克蛋白,多以“分子伴侶” (molecullar chaperones) 的形式與代謝活動(dòng)中蛋白酶相結(jié)合, 糾正蛋白錯(cuò)誤卷曲, 從而保護(hù)酶活性, 提高機(jī)體耐受能力.當(dāng)機(jī)體遭受紫外線、低氧等不良脅迫環(huán)境時(shí),機(jī)體處于應(yīng)激狀態(tài),表現(xiàn)為HSP表達(dá)升高.已有報(bào)道顯示運(yùn)動(dòng)應(yīng)激能誘導(dǎo)骨骼肌細(xì)胞中HSP合成表達(dá)上升,如發(fā)現(xiàn)低頻電短期刺激可延長(zhǎng)大鼠游泳時(shí)間,增加其運(yùn)動(dòng)能力,蛋白水平檢測(cè)發(fā)現(xiàn)大鼠此過(guò)程中增強(qiáng)了HSP70表達(dá)[20].本課題組前期的研究已經(jīng)發(fā)現(xiàn)PF-EMF暴露可誘導(dǎo)果蠅、及的等熱激蛋白基因表達(dá)水平上升[21],由此可以推出PF-EMF暴露可能引發(fā)了果蠅運(yùn)動(dòng)行為增加.

與野生型相比,突變體果蠅白天和夜晚運(yùn)動(dòng)次數(shù)均升高,說(shuō)明PF-EMF對(duì)突變體果蠅影響更顯著,這可能也是由于突變體果蠅對(duì)PF-EMF更易感.由圖4B進(jìn)一步分析果蠅活躍的具體表現(xiàn),我們發(fā)現(xiàn)暴露組野生型果蠅單位時(shí)間運(yùn)動(dòng)次數(shù)顯著提升(**<0.01),而突變型果蠅變化不顯著,提示兩株系果蠅運(yùn)動(dòng)活躍機(jī)制有差異.對(duì)于野生型果蠅而言,PF-EMF暴露后表現(xiàn)為較明顯的應(yīng)激反應(yīng),單位時(shí)間內(nèi)活動(dòng)頻繁;而對(duì)突變體果蠅來(lái)講,PF- EMF的影響則更體現(xiàn)為減少果蠅的睡眠時(shí)長(zhǎng),運(yùn)動(dòng)時(shí)長(zhǎng)隨之增加,因而表現(xiàn)為運(yùn)動(dòng)總量增加而單位運(yùn)動(dòng)量未變.分析分子機(jī)制,推測(cè)在野生型果蠅體內(nèi),PF- EMF暴露效應(yīng)主要體現(xiàn)為一種類似于熱應(yīng)激反應(yīng),由于HSP表達(dá)增多伴隨果蠅行為活躍,與上文探討一致;而在果蠅體內(nèi),已知蛋白是編碼煙堿乙酰膽堿受體α4(nAChRα4)復(fù)合物的四通道跨膜通道蛋白,當(dāng)發(fā)生突變后,乙酰膽堿跨膜轉(zhuǎn)運(yùn)受阻干擾了信號(hào)傳導(dǎo)過(guò)程,進(jìn)而發(fā)生睡眠質(zhì)量碎片化的現(xiàn)象.實(shí)驗(yàn)表明nAChRα4 突變會(huì)導(dǎo)致有機(jī)體對(duì)睡眠環(huán)境更為敏感,比如nAChRα4 突變致癲癇的患者睡眠時(shí)對(duì)溫度、光環(huán)境質(zhì)量要求提高[22].已有證據(jù)提示極低頻電磁場(chǎng)作為干擾因子,在長(zhǎng)期職業(yè)接觸后可能會(huì)導(dǎo)致工人抑郁、焦慮和睡眠質(zhì)量變差[23],本研究中也發(fā)現(xiàn)了PF-EMF暴露減少果蠅睡眠時(shí)長(zhǎng)的結(jié)果.當(dāng)果蠅經(jīng)PF-EMF處理后,此時(shí)類熱應(yīng)激反應(yīng)的表現(xiàn)不明顯,而對(duì)于突變導(dǎo)致的睡眠剝奪效應(yīng)的影響更為凸顯.

2.3.2 PF-EMF對(duì)果蠅睡眠行為的影響 由圖5A可知,W1118野生型果蠅的總睡眠時(shí)長(zhǎng)在PF-EMF暴露后未發(fā)生顯著變化,基因突變型果蠅白天、夜間和24h全天的睡眠時(shí)長(zhǎng)均顯著下降(<0.01).該結(jié)果進(jìn)一步印證了基因突變型果蠅比野生型果蠅對(duì)PF-EMF環(huán)境脅迫更加敏感.同時(shí)W1118型果蠅的睡眠時(shí)長(zhǎng)在暴露后未顯著改變,這也驗(yàn)證了上文對(duì)W1118型果蠅活躍表現(xiàn)的原因分析,主要是由于果蠅活動(dòng)頻率增加,單位時(shí)間內(nèi)運(yùn)動(dòng)次數(shù)上升,并非是擠占了睡眠時(shí)間.圖5B可以看出對(duì)照組果蠅睡眠片段化次數(shù)明顯多于對(duì)照組W1118野生型果蠅,這是由于蛋白突變?cè)斐?PF-EMF暴露后W1118的24h睡眠片段次數(shù)顯著減少10.6%,白天和夜間統(tǒng)計(jì)結(jié)果與對(duì)照組未出現(xiàn)顯著差異,而果蠅的夜間睡眠片段次數(shù)減少,24h睡眠片段次數(shù)下降51.5%(*<0.05,**<0.01),總睡眠時(shí)間也顯著減少45.7%,表明突變型果蠅暴露后形成失眠的情況比野生型更顯著,結(jié)合上文運(yùn)動(dòng)節(jié)律分析,果蠅因失眠造成運(yùn)動(dòng)時(shí)間延長(zhǎng),進(jìn)而總運(yùn)動(dòng)量增大.

2.3.3 PF-EMF對(duì)果蠅節(jié)律相位的影響 由圖6分析可知無(wú)論是W1118野生型果蠅還是基因突變型果蠅,PF-EMF并未改變其運(yùn)動(dòng)和睡眠整體節(jié)律相位.兩株系果蠅都呈現(xiàn)出明顯運(yùn)動(dòng)節(jié)律峰(圖6A)和睡眠節(jié)律峰(圖6B),與W1118相對(duì)比,果蠅受PF-EMF影響更大,運(yùn)動(dòng)量增加和睡眠減少更顯著.圖6B顯示,W1118果蠅與果蠅均在中午12:00出現(xiàn)睡眠高峰,但果蠅對(duì)照組與暴露組的高峰峰值的差距比野生型明顯,果蠅在暴露后明顯減少了高峰期睡眠時(shí)間,且隨著暴露時(shí)間延長(zhǎng),W1118果蠅與果蠅的睡眠高峰峰值在PF-EMF暴露后差異均越來(lái)越明顯,這可能是由于暴露的累積效應(yīng)所致.

A:平均運(yùn)動(dòng)次數(shù);B:單位時(shí)間運(yùn)動(dòng)量

A:睡眠總時(shí)長(zhǎng);B:睡眠片段

A:運(yùn)動(dòng)規(guī)律;B:睡眠規(guī)律

3 結(jié)論

3.1 人造輸變電工程的產(chǎn)生的工頻電磁環(huán)境對(duì)昆蟲發(fā)育和節(jié)律行為有影響.PF-EMF暴露對(duì)果蠅成蛹量和成蛹速率都有促進(jìn)作用,但不改變出生子代的性別比例,未發(fā)生性別偏移.

3.2 PF-EMF暴露對(duì)果蠅運(yùn)動(dòng)和睡眠的節(jié)律相位未造成顯著影響,但PF-EMF暴露后果蠅運(yùn)動(dòng)量明顯增大,睡眠時(shí)間顯著減少,睡眠片段增多,PF-EMF暴露引起野生型果蠅運(yùn)動(dòng)行為增加,活動(dòng)頻繁,表現(xiàn)為一種類熱應(yīng)激反應(yīng)效應(yīng).

3.3突變型果蠅作為短睡眠個(gè)體與野生型相比,體質(zhì)相對(duì)羸弱,因而對(duì)PF-EMF環(huán)境壓力的感知更敏感,效應(yīng)表現(xiàn)比野生型更顯著,突變型表現(xiàn)為睡眠片段次數(shù)減少,運(yùn)動(dòng)量增大主要由于失眠所致,PF-EMF顯著降低了突變體果蠅的睡眠水平,加劇其睡眠障礙情況.

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Effects of power frequency electromagnetic environment on development and rhythmic behavior of.

ZOU Xin-ping1, WANG Xiao-li1, DUAN Zheng-hua1, ZHANG Zi-yan2,3, SUN Yong-yan1,3*

(1.School of Environmental Science and Safety Engineering, Tianjin University of Technology, Tianjin 300384, China；2.Institute of Urban Environment, Chinese Academy of Sciences, Xiamen 361021, China；3.Shanghai Institute of Nutrition and Health, Chinese Academy of Sciences, Shanghai 200031, China)., 2023,43(11)：6149～6155

Differences in development, sleep and movement rhythmic behaviors between W1118 wild-type flies andmutants were investigated under PF-EMF exposure. After continuous exposure for 3 days, the parental flies were removed. Developmental process, movement and sleep rhythmic behaviors of offspring were monitored. The results showed that , the pupal and fly population of W1118 significantly increased respectively by 56.2% and 57.5% under PF-EMF exposure. No significant changes were observed in pupal numbers oftype flies, but the fly population significantly increased by 85.2%. No significant change of sex ratio was found under PF-EMF exposure. For W1118 flies, the average number of 24-hour movements was increased by 37.3% after PF-EMF exposure, and 10.6% decrease in the number of 24-hour sleep segments was calculated. Fortype flies, the average counts of 24-hour movement significantly increased by 50.0 % after exposure, and 51.5% decrease of 24-hour sleep segment counts was investigated. In the meanwhile, 45.7% decrease of the total sleep duration were also found. Compared with the W1118 type, themutants showed more significant changes, which suggested thatmutants were more sensitive to PF-EMF exposure. In brief, the PF-EMF environment can impact development and rhythmic behaviors of insects. Increased fly offspring numbers movement counts, and disturbed sleep were found after PF-EMF exposure. Further understandings on the entomological effects of PF-EMF exposure, and reasonable evaluations on the ecological effects of PF-EMF can be provided in this study.

PF-EMF；；behavioral rhythms；development

X171.5

A

1000-6923(2023)11-6149-07

鄒鑫平(1997-),男,江西撫州人,天津理工大學(xué)碩士,主要從事物理環(huán)境與睡眠健康相關(guān)研究.發(fā)表論文1篇.zouxinping918@163.com.

鄒鑫平,王曉麗,端正花,等.工頻電磁環(huán)境對(duì)果蠅發(fā)育和節(jié)律行為的影響研究 [J]. 中國(guó)環(huán)境科學(xué), 2023,43(11):6149-6155.

Zou X P, Wang X L, Duan Z H, et al. Effects of power frequency electromagnetic environment on development and rhythmic behavior of[J]. China Environmental Science, 2023,43(11):6149-6155.

2023-03-09

自然資源部海洋生物遺傳資源重點(diǎn)實(shí)驗(yàn)室開放課題項(xiàng)目(HY202106);國(guó)家自然科學(xué)基金資助項(xiàng)目(32201017)

* 責(zé)任作者, 講師, yongyansun@email.tjut.edu.cn