謝湘林，劉蕾，謝宇，方圣博，杜冰，睢大筼，劉宏雁*(.吉林大學(xué)藥學(xué)院藥理教研室,長春300;.吉林大學(xué)口腔醫(yī)院修復(fù)科,長春300)

研究報(bào)告

5年生種植人參對小鼠學(xué)習(xí)記憶的影響

謝湘林1，劉蕾1，謝宇2，方圣博1，杜冰1，睢大筼1，劉宏雁1*
(1.吉林大學(xué)藥學(xué)院藥理教研室,長春130021;2.吉林大學(xué)口腔醫(yī)院修復(fù)科,長春130021)

目的觀察5年生種植人參原粉、水提物和醇提物長期應(yīng)用對小鼠學(xué)習(xí)記憶的影響,為人參的食用提供實(shí)驗(yàn)依據(jù).方法正常成年昆明小鼠140只,雌雄各半,體重18～22g,隨機(jī)分為10組,即:對照組,5年生種植人參原粉小、中、大劑量(0.25、0.5、1.0g生藥/kg)組,5年生種植人參水提物小、中、大劑量(0.25、0.5、1.0g生藥/kg)組,5年生種植人參醇提物小、中、大劑量(0.25、0.5、1.0g生藥/ kg)組,每組14只,連續(xù)灌胃給藥3個(gè)月后進(jìn)行東莨菪堿致學(xué)習(xí)記憶獲得障礙實(shí)驗(yàn).結(jié)果與模型組比較,人參醇提物大劑量小鼠第1天的到達(dá)平臺潛伏期、游程有縮短的趨勢,但無統(tǒng)計(jì)學(xué)意義,第4天的游程明顯縮短(P〈0.05),其他指標(biāo)無明顯變化.人參原粉小、中、大劑量組,人參水提物小、中、大劑量組,人參醇提物小、中劑量組第1天至第5天到達(dá)平臺的潛伏期、游程、朝向角、游泳速度無明顯變化,第6天小鼠在1.5min內(nèi)穿越平臺的次數(shù)、穿越有效區(qū)的次數(shù)、平臺停留距離、平臺區(qū)的逗留時(shí)間、有效區(qū)停留距離、有效區(qū)停留時(shí)間、平臺停留距離/總路程比、在平臺區(qū)的逗留時(shí)間/總時(shí)間、有效區(qū)停留距離/總路程、有效區(qū)停留時(shí)間/總時(shí)間均無明顯變化.結(jié)論長期食用5年生種植人參原粉、水提物及醇提物對小鼠東莨菪堿致學(xué)習(xí)記憶獲得障礙無明顯的改善作用.

五年生種植人參;食用;學(xué)習(xí)記憶;小鼠

人參為珍貴的中藥材,其含有皂苷類、糖類、有機(jī)酸等多種成分,其中主要有效成分為人參皂苷和人參多糖[1].人參作用廣泛,大量的研究結(jié)果顯示人參對中樞神經(jīng)系統(tǒng)、循環(huán)系統(tǒng)、內(nèi)分泌等多種系統(tǒng)均有顯著的藥理作用[2～4].

大量的研究結(jié)果顯示,人參有效成分及某些單體對學(xué)習(xí)記憶功能有明顯的改善作用[5～7],但人參原粉及粗提物長期應(yīng)用對學(xué)習(xí)記憶的影響未見相關(guān)報(bào)道,本研究預(yù)觀察長期應(yīng)用人參原粉、水提物、醇提物對小鼠學(xué)習(xí)記憶影響,為人參的食用提供實(shí)驗(yàn)依據(jù).

1　材料與方法

1.1實(shí)驗(yàn)動(dòng)物昆明小鼠140只,雌雄各半,體重18-22g.由吉林大學(xué)動(dòng)物研究中心提供,批號:SCXK-(吉)2007-2003.動(dòng)物于專用實(shí)驗(yàn)室內(nèi)分籠飼養(yǎng),自由攝食和飲水.

1.2實(shí)驗(yàn)藥品及儀器實(shí)驗(yàn)用生曬參(5年生)產(chǎn)于吉林省撫松縣.人參原粉系200目的細(xì)粉;人參水提物系兩次水煎煮提取,過濾,濃縮,烘干得人參干粉末,提取率為40.09%,其中人參總皂苷含量約為0.99%,人參多糖含量為7.54%;人參醇提物系70%乙醇加熱回流提取人參粗粉,過濾,濃縮,烘干得人參浸膏,提取率為30.8%,其中人參總皂苷含量約為2.33%,人參多糖含量為0.76%.上述三種實(shí)驗(yàn)藥品均由長春中醫(yī)藥大學(xué)研發(fā)中心生物工程實(shí)驗(yàn)室提供,批號分別為:20101216、20110407及20110331;氫溴酸東莨菪堿注射液,國藥控股沈陽有限公司長春分公司,批號:090801.Morris水迷宮裝置為成都泰盟科技有限公司產(chǎn)品,自制跳臺裝置.

1.3實(shí)驗(yàn)方法將140只小鼠隨機(jī)分為對照組,5年生種植人參原粉小、中、大劑量(0.25、0.5、1.0g生藥/kg)組,5年生種植人參水提物小、中、大劑量(0.25、0.5、1.0g生藥/kg)組,5年生種植人參醇提物小、中、大劑量(0.25、0.5、1.0g生藥/kg)組.各組大鼠每天灌胃給藥1次,連續(xù)3個(gè)月后,進(jìn)行東莨菪堿致小鼠學(xué)習(xí)記憶獲得障礙實(shí)驗(yàn),實(shí)驗(yàn)期間繼續(xù)給藥.

水迷宮實(shí)驗(yàn)連續(xù)進(jìn)行6天,前5天,在1、2、3、4象限四個(gè)不同的入水點(diǎn),測定小鼠到達(dá)平臺潛伏期、游程、朝向角及平均速度,第6天撤掉平臺,測定小鼠在1.5min經(jīng)過平臺次數(shù)、經(jīng)過有效區(qū)次數(shù)、平臺停留距離、平臺停留時(shí)間、有效區(qū)停留距離、有效區(qū)停留時(shí)間、平臺停留距離/總路程、平臺停留時(shí)間/總時(shí)間、有效區(qū)停留距離/總路程、有效區(qū)停留時(shí)間/總時(shí)間.每天給予東莨菪堿2mg/kg,20分鐘后進(jìn)行水迷宮實(shí)驗(yàn).

2　結(jié)果

與模型組比較,人參醇提物大劑量小鼠第1天的到達(dá)平臺潛伏期、游程有縮短的趨勢,但無統(tǒng)計(jì)學(xué)意義,第4天的游程明顯縮短(P<0.05),其他指標(biāo)無明顯變化.與模型組比,人參原粉小、中、大劑量組,人參水提物小、中、大劑量組,人參醇提物小、中第1天至第5天到達(dá)平臺的潛伏期、游程、朝向角、游泳速度無明顯變化,第6天小鼠在2min內(nèi)穿越平臺的次數(shù)、穿越有效區(qū)的次數(shù)、平臺停留距離、平臺區(qū)的逗留時(shí)間、有效區(qū)停留距離、有效區(qū)停留時(shí)間、平臺停留距離/總路程比、在平臺區(qū)的逗留時(shí)間/總時(shí)間、有效區(qū)停留距離/總路程、有效區(qū)停留時(shí)間/總時(shí)間均無明顯變化,結(jié)果見表1～5.

表1　人參對成年小鼠水迷宮潛伏期的影響(s)

表1　人參對成年小鼠水迷宮潛伏期的影響(s)

*與模型組比較P<0.05,**P<0.01

組別例數(shù)1第1天第2天第3天第4天第5天對照組1141.97±32.33**14.35±10.64**9.82±4.40**8.72±3.69**8.34±3.35**模型組1193.72±22.2271.49±36.1553.69±34.3066.35±34.0351.74±31.17原粉小8102.58±19.8876.49±14.6961.10±42.3563.73±25.0847.82±23.45原粉中1096.32±27.4274.80±38.7472.53±39.6054.08±35.7536.20±37.50原粉大884.54±32.7248.06±30.5242.70±23.6343.90±27.1535.53±37.64水提物小881.30±26.0855.38±25.6667.84±34.8264.69±33.3150.65±40.04水提物中1085.39±26.0769.82±30.8164.92±36.7356.02±33.2652.61±40.00水提物大1085.83±21.1365.69±36.9261.60±40.4838.58±31.0462.56±39.15醇提物小893.07±25.0497.92±32.1673.52±33.9885.08±37.0565.35±37.42醇提物中1189.79±30.7672.83±31.1774.38±41.3573.32±32.8957.24±45.60醇提物大1074.27±29.7858.96±37.4944.35±32.5747.54±43.2556.30±35.99

表2　人參對成年小鼠水迷宮游程的影響(cm)

表2　人參對成年小鼠水迷宮游程的影響(cm)

*與模型組比較P<0.05,**P<0.01

組別例數(shù)第1天第2天第3天第4天第5天對照組11773.33±489.12**288.55±186.65**222.20±49.70**181.41±72.00**180.43±68.43**模型組111801.48±604.48 1366.33±734.34 1046.84±679.00 1344.05±807.24 1042.79±642.90原粉小81808.00±397.91 1543.09±306.41 1218.74±845.95 1326.19±454.90 1032.11±623.32原粉中101637.34±431.531398.70±67.581399.63±697.12 1029.09±615.50697.73±673.23原粉大81479.57±669.56923.71±552.98874.49±450.07859.64±559.61688.32±723.90水提物小81554.86±512.65 1191.17±503.21 1459.60±702.79 1429.82±768.82 1113.02±814.06水提物中101434.27±288.14 1095.38±386.35 1142.64±660.79965.52±593.93942.96±43.56水提物大101548.22±347.82 1279.27±584.12 1183.03±659.86846.03±573.181250.21±684.29醇提物小81495.90±459.61 1562.06±495.89 1350.38±568.56 1531.31±593.45 1213.74±616.34醇提物中111677.89±606.20 1402.02±542.63 1345.87±722.28 1420.73±590.97 1113.96±788.66醇提物大101360.50±6050.201021.48±508.27883.73±527.58701.12±506.45*1079.87±643.01

表3　人參對成年小鼠水迷宮朝向角的影響(°)

表3　人參對成年小鼠水迷宮朝向角的影響(°)

*與模型組比較P<0.05,**P<0.01

組別例數(shù)第1天第2天第3天第4天第5天對照組1145.13±22.3953.40±32.6238.54±28.1643.82±24.0140.83±21.24模型組1147.71±19.5042.69±16.5157.00±23.2151.59±21.7155.80±28.14原粉小849.87±33.1447.33±24.0947.58±24.2154.64±20.0337.68±13.01原粉中1044.68±26.6555.05±21.6541.59±17.7149.24±24.5541.70±15.94原粉大850.53±22.4057.09±26.4355.9±917.9759.46±19.8145.60±23.82水提物小841.98±19.7841.29±21.6750.11±17.8748.63±25.6444.52±22.75水提物中1049.51±18.3244.89±24.7843.7±320.5852.89±28.3349.17±28.84水提物大1056.99±14.6243.86±19.6152.22±22.3943.17±23.6155.04±25.87醇提物小861.93±14.5947.42±17.5962.54±22.0349.74±25.8045.52±20.68醇提物中1133.39±17.5153.47±17.0848.46±21.4852.201±9.8538.95±25.26醇提物大1046.71±21.7553.74±29.4149.33±20.9346.41±22.9851.08±25.50

表4　人參對成年小鼠水迷宮平均速度的影響(cm/s)

表4　人參對成年小鼠水迷宮平均速度的影響(cm/s)

*與模型組比較P<0.05,**P<0.01

組別例數(shù)第1天第2天第3天第4天第5天對照組1121.0±56.8621.32±6.3326.72±13.4120.72±3.8022.76±7.75模型組1119.6±84.8620.64±4.7121.94±4.1620.98±4.2721.96±5.36原粉小817.67±2.1421.15±3.2320.54±3.1622.50±6.0922.02±2.64原粉中1017.10±1.8018.89±2.6219.73±2.7320.15±4.9222.89±4.90原粉大817.70±3.7618.88±2.1320.45±2.5720.78±2.3922.50±3.98水提物小819.92±3.3122.20±2.5121.55±1.6122.66±2.8722.63±11.75水提物中1017.58±3.4018.45±5.3719.19±4.0318.23±3.8120.46±6.82水提物大1018.97±2.8220.40±2.6619.82±2.6825.85±13.6221.40±3.98醇提物小816.22±2.6217.35±3.6618.84±2.1019.23±3.9920.15±3.42醇提物中1118.24±1.7119.09±3.4318.69±3.2923.35±6.9522.13±4.29醇提物大1018.94±6.4219.54±4.2521.18±3.1718.80±6.4120.84±4.37

表5　人參對成年小鼠第6天水迷宮實(shí)驗(yàn)的影響

表5　人參對成年小鼠第6天水迷宮實(shí)驗(yàn)的影響

*與模型組比較P<0.05,**P<0.01

組別n經(jīng)過平臺次數(shù)經(jīng)過有效區(qū)次數(shù)平臺停留距離(cm)平臺停留時(shí)間(s)有效區(qū)停留距離(cm)正常對照119.90±3.69*19.90±4.22*88.91±36.804.37±1.87774.5460±205.8002模型組115.45±4.7113.72±6.0554.85±54.102.80±2.99520.9671±384.2406原粉小83.12±3.449.37±6.8831.83±35.861.45±1.68331.1489±263.3315原粉中106.40±4.9716.40±8.3962.34±52.602.69±2.69520.6510±333.0155原粉大83.50±3.4610.12±5.1931.31±28.741.60±1.62326.6338±225.3414水提物小84.00±3.0213.62±8.5334.57±30.771.46±1.44416.0658±278.1003水提物中105.10±4.3312.90±7.7544.32±41.462.17±2.11447.0368±323.1549水提物大105.50±4.4514.70±9.3453.91±46.162.32±1.91495.3344±332.1806醇提物小81.75±2.869.00±6.1817.38±34.211.08±2.30293.7998±244.2904醇提物中115.90±5.4612.81±8.8858.90±57.822.94±2.88457.8216±363.5375醇提物大105.50±3.1015.60±4.5051.01±29.072.15±1.60538.4719±201.3847 Groupn有效區(qū)停留時(shí)間(s)有效區(qū)停留時(shí)間/總時(shí)間X100%正常對照1140.30±9.664.18±2.06*3.64±1.5536.89±14.54**33.66±12.95**模型組1125.91±22.241.95±2.082.33±2.5018.30±13.4917.37±13.18原粉小814.36±12.281.22±1.351.21±1.3912.75±10.7012.01±10.38原粉中1022.65±16.672.42±2.352.24±2.2320.16±15.0318.89±14.37原粉大817.57±16.851.48±1.581.33±1.3414.86±13.2314.65±14.02水提物小817.38±11.981.26±1.161.22±1.2015.31±10.9214.50±10.51水提物中1021.43±15.501.95±1.781.80±1.7619.27±13.9717.8713.39水提物大1022.03±14.552.06±1.671.93±1.5919.27±13.0718.40±12.76醇提物小814.58±13.780.81±1.570.90±1.9212.64±11.3312.53±11.51醇提物中1121.72±16.222.26±2.452.45±2.3917.57±14.3316.28±13.05醇提物大1024.29±13.402.09±1.671.79±1.3321.56±12.8420.26±12.24平臺停留距離/總路程X100%平臺停留時(shí)間/總時(shí)間X100%有效區(qū)停留距離/總路程X100%

3　討論

東莨菪堿為M受體阻斷劑,通過與M受體特異性結(jié)合,阻斷乙酰膽堿作用,破壞了腦內(nèi)的信息傳遞通路,使動(dòng)物發(fā)生學(xué)習(xí)記憶獲得障礙.水迷宮實(shí)驗(yàn)是考察動(dòng)物定位航行能力及空間探索能力.

本研究結(jié)果顯示長期食用人參原粉、水提物及醇提物,對小鼠膽堿系統(tǒng)損傷致獲得性記憶障礙(定位航行能力及空間探索能力),無明顯的改善.

人參有效成分、某些單體對學(xué)習(xí)記憶損傷有明顯的改善作用[5～7],而本研究結(jié)果顯示人參原粉、水提物及醇提物長期應(yīng)用對東莨菪堿致學(xué)習(xí)記憶損傷無明顯的改善作用,其原因是人參各種成分相互作用的結(jié)果,給藥時(shí)間的影響,還是其他機(jī)制?人參原粉、水提物及醇提物長期應(yīng)用對其他原因?qū)е碌膶W(xué)習(xí)記憶損傷的如何影響?這些都有待于進(jìn)一步研究.

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The effect of Ginseng-5year on learning and memory in mice

XIE Xiang-lin LIU Lei FANG Sheng-Bo,DU Bing,SUI Da-yun,LIU Hong-Yan*
(Department of Pharmacology,School of Pharmacy,Jilin University,Changchun 130021,China)

Aim To observe the long-term effect of Ginseng-5year raw powder、aqueous extract and ethanol extract on learning and memory in mice to provide experimental basis for edible Ginseng.Methods 140 Kun Ming mice(70 male and 70 female,18-22g)were divide to 10 groups,normal group, Ginseng raw powder little,middle and large dose group(0.25,0.5 and 1.0g crude drug/kg),Ginseng aqueous extract little,middle and large dose group(0.25,0.5 and 1.0g crude drug/kg),Ginseng ethanol extract little,middle and large dose group(0.25,0.5 and 1.0g crude drug/kg),14 mice a group,do the learning and memory abilities experiment of acquisition impairment model mice induced by scopolamine after intragastric administration for 3 months.Result Compared with normal group,incubation period and run-length of Ginseng ethanol extract large dose group had a decreasing trend on the 1st day,but there was no statistical significance,and the run-length shortened on the 4th day(P〈0.05),at the same time other indicators have no obvious change.There was no obvious change in incubation period,run-length,orientation Angle and swimming speed betweenGinseng raw powder little,middle,large dose group,Ginseng aqueous extract little, middle,large dose group,Ginseng ethanol extract little and middle group on the 1st day to 5th day.There was no obvious change in the times of crossing the platform,times of crossing the effective area,distance in the platform,distance in the effective area,standing time in the platform, standing time in the effective area,the ratio of residence time in the platform area/total time,the ratio of residence time in the effective area/total time,the ratio of distance in the effective area/ total distance,the ratio of distance in the platform/total distance in 1.5min on the 6th day. Conclusion 5-year-Ginseng raw powder、aqueous extract and ethanol extract have not obvious effect on learning and memory abilities of acquisition impairment model mice induced by scopolamine for edible Ginseng.

5-year-Ginseng,edible,learning and memory,mice學(xué)習(xí)與記憶功能為腦的高級功能,是人類最具特色的生理特征之一,有良好的學(xué)習(xí)記憶功能,人類才能與外界多變的環(huán)境相適應(yīng),才能參加社會(huì)實(shí)踐活動(dòng),才能認(rèn)識和改造客觀世界.

吉林省重大科技攻關(guān)項(xiàng)目,項(xiàng)目編號:10ZDGG013.

謝湘林,男,高級實(shí)驗(yàn)師,主要從事中樞藥理研究.

劉宏雁,女,教授,主要從事中樞藥理學(xué)研究.E-mail:hongyl@jlu.edu.cn.