铅暴露对快速衰老小鼠学习记忆能力的影响

Effects of lead exposure on learning and memory ability of senescence accelerated mice

[目的]探讨铅(Pb)暴露对快速衰老小鼠(SAMP8)学习记忆能力的影响及其机制。[方法]雄性SAMP8小鼠及其野生型SAMR1小鼠各20只,各随机分为2组,即SAMR1组、SAMR1+Pb组和SAMP8组、SAMP8+Pb组。SAMR1+Pb组和SAMP8+Pb组小鼠每天饲以0.05 g/L醋酸铅饮用水,SAMR1组和SAMP8组小鼠每天饲以0.05 g/L醋酸钠饮用水,共饮水9周。应用Morris水迷宫实验检测小鼠学习记忆能力;采用免疫荧光法测定实验小鼠侧脑室下区(SVZ)淀粉样前体蛋白(APP)、神经元母细胞和星形胶质样细胞特异性标志物[双皮质素(DCX)、神经胶质纤维酸性蛋白(GFAP)]的表达情况,分析神经元母细胞和星形胶质细胞的细胞比率变化;应用实时荧光定量PCR检测实验小鼠SVZ中DCX、GFAP和脑源性神经营养因子(BDNF)的m RNA表达情况。[结果]小鼠Morris水迷宫实验第3天结果显示:与SAMR1组[(10.94±3.98)s]相比,SAMP8组[(29.24±6.15)s]、SAMR1+Pb组[(34.15±6.78)s]和SAMP8+Pb组[(50.86±8.56)s]的逃避潜伏期延长,差异有统计学意义(P<0.05);且与SAMP8组和SAMR1+Pb组小鼠比较,SAMP8+Pb组小鼠的逃避潜伏期更长(P<0.05)。SAMR1组、SAMP8组、SAMR1+Pb组和SAMP8+Pb组小鼠的平均穿越平台次数分别为(6.75±1.28)、(3.25±1.28)、(3.88±1.46)、(1.25±0.46)次;SAMP8组、SAMR1+Pb组和SAMP8+Pb组小鼠穿越次数较SAMR1组均减少(P<0.05);SAMP8+Pb组小鼠穿越次数也明显低于SAMP8组和SAMR1+Pb组(P<0.05)。免疫荧光法测定结果显示:与SAMR1组小鼠比较,SAMR1+Pb组、SAMP8组和SAMP8+Pb组小鼠SVZ区APP荧光强度分别上升了79%、53%和216%;与SAMR1+Pb组和SAMP8组比较,SAMP8+Pb组小鼠APP荧光强度分别上升了76%和107%,差异均具有统计学意义(P<0.05)。与SAMR1组小鼠比较,SAMP8组和SAMP8+Pb组小鼠SVZ中GFAP(+)/DAPI(+)和DCX(+)/DAPI(+)细胞比率均降低,SAMP8+Pb组小鼠SVZ的GFAP(+)/DAPI(+)和DCX(+)/DAPI(+)细胞比率分别为SAMP8组小鼠的22%和59%。实时荧光定量PCR分析结果显示:与SAMR1组小鼠相比,SAMR1+Pb组、SAMP8组和SAMP8+Pb组小鼠SVZ中BDNF和DCX m RNA表达明显下降(P<0.05);SAMP8+Pb组小鼠BDNF m RNA表达较SAMR1+Pb组和SAMP8组小鼠分别下降了85%和83%,DCX mR NA表达则分别下降了81%和60%,差异有统计学意义(P<0.05);SAMP8组和SAMP8+Pb组小鼠的GFAP m RNA表达较SAMR1组明显下降,并且SAMP8+Pb组小鼠GFAP m RNA表达较SAMR1+Pb组和SAMP8组分别下降了89%和69%,差异有统计学意义(P<0.05)。[结论]铅暴露可导致快速衰老小鼠SVZ中APP表达增加和BDNF m RNA表达下降,同时加剧SVZ中星形胶质样细胞丢失,逃避潜伏期延长,穿台次数下降,提示铅暴露加剧快速衰老小鼠学习记忆能力的下降。

[ Objective ] To investigate the effects and related mechanism of lead （Pb） exposure on learning and memory ability of senescence accelerated mice （SAMP8） [ Methods ] Twenty male SAMP8 mice and 20 male SAMR1 mice were randomly divided into four groups： SAMR1, SAMRI＋Pb exposure, SAMP8, and SAMP8＋Pb exposure groups. Mice in the SAMRI＋Pb exposure group and the SAMP8＋Pb exposure group were treated with 0.05 g/L lead acetate via drinking water for nine weeks, and mice in the SAMR1 group and the SAMP8 group were given 0.05 g/L sodium acetate via drinking water for nine weeks. Morris water maze experiment was applied to test learning and memory ability. Protein expressions of amyloid precursor protein （APP）, doublecortin （DCX）, and glial fibrillary acidic protein （GFAP） in subventricular zone （SVZ） were observed by immunofluorescence, and the cell counts of neuronal progenitors and astrocytes were analyzed. The mRNA expressions of DCX, GFAP, and brain-derived neurophic factor （BDNF） in SVZ were detected by quantitative real-time PCR. [ Results ] The results of Morris water maze test on day 3 showed that, compared with the SAMR1 group [（10.94±3.98）s], the escape latencies of the SAMRI＋Pb exposure group [（34.15±6.78）s], the SAMP8 group [（29.24±6.15）s], and the SAMP8＋Pb exposure group [（50.86±8.56）s] were increased （P〈0.05）; compared with the SAMP8 group and the SAMRI＋Pb exposure group, the escape latency of the SAMP8＋Pb exposure group was extented （P 〈 0.05）; the number of platform crossings were 6.75± 1.28 in the SAMR1 group, 3.88 ±1.46 in the SAMRI＋Pb exposure group, 3.25 ±1.28 in the SAMP8 group, and 1.25± 0.46 in the SAMP8＋Pb exposure group; the numbers in the SAMRI＋Pb exposure group, SAMP8 group, and SAMP8＋Pb exposure group were lower than those in the SAMR1 group （P 〈 0.05）; compared with the SAMP8 group and the SAMRI＋Pb exposure group, the number in the SAMP8＋Pb exposure group was also decreased （P〈 0.05）. The results of immunoftuorescence technology showed that, compared with SAMR1 mice, the fluorescence intensity of APP increased by 79%, 53%, and 216% in the SAMRI＋Pb exposure group, SAMP8 group, and SAMP8＋Pb exposure group, respectively; compared with the SAMRI＋Pb exposure group and the SAMP8 group, the fluorescence intensity of APP in the SAMP8＋Pb exposure group increased by 76% and 107%, respectively （P 〈 0.05）. Compared with the SAMR1 group, the cell ratios of GFAP（＋）/DAPI（＋） and DCX（＋）/DAPI（＋） in SVZ of the SAMP8 group and the SAMP8＋Pb exposure group were decreased; the ratios of GFAP（＋）/DAPI（＋） and DCX（＋）/DAPI （＋） in SVZ of the SAMP8＋Pb exposure group were 22% and 59% of the SAMP8 group, respectively. Compared with the SAMRI group, the expression of BDNF and DCX mRNA in the SAMRI＋Pb exposure group, SAMP8 group, and SAMP8＋Pb exposure group were significantly decreased by real-time fluorescence quantitative PCR （P〈O.05）. Compared with the SAMRI＋Pb exposure group and the SAMP8 group, the expressions of BDNF mRNA in the SAMP8＋Pb exposure group were decreased by 85% and 83%, respectively, and the expressions of DCXmRNA were decreased by 81% and 60%, respectively （P〈0.05）. The expressions of GFAP mRNA in the SAMP8 group and SAMP8＋Pb exposure group were significantly lower than that in the SAMR1 group （P〈 0.05）; the expression of GFAP mRNA in the SAMP8＋Pb exposure group was decreased by 89% and 69% compared with the SAMRI＋Pb group and the SAMP8 group, respectively （P〈0.05）. [ Conclusion ] Our findings show that lead exposure can aggravate the increasing of APP protein expression and the decreasing of BDNF mRNA expression, as well as result in less astrocytes in SVZ, prolonged escape latency, and reduced platform crossing counts, which indicate that lead exposure may accelerate the decline of learning and memory ability in senescence accelerated mice.