| Objective:1.To investigate the possible caspase pathway of exposure of long-term aluminum lead to neuronal apoptosis in rats.2.To explore the relation between the neuronal apoptosis in rats induced by aluminum and synaptic plasticity injury.Methods:1.Fourty male healthy SD rats were randomly divided into four groups: control group(0mg/kg.bw), low dose group(10mg/kg.bw), medium dose group(30mg/kg.bw) and high dose group(90mg/kg.bw). The rats were daily gavaged for 1 and 3 months in order to build a model of aluminum lactate exposure.2.MWM was employed to detect the ability of spatial learning and memory in rats, and Open-field test was designed to observe the ability of exploratory behavior and cognitive abilities in rats.3.The hippocampal LTP in rats was measured with electrophysiological grapher.4.The hippocampal neuronal apoptosis in rats was detected with Flow cytometry.5.The relative expression of gene which includes caspase-3, caspase-8,caspase- 9 was measured by Real-time PCR.6.The relative expression of protein which includes caspase-3, caspase-8, caspase-9,NR1,NR2 A,CYCS was measured by ELISA.Results:1. Neurobehavioral test: During the hidden platform test, escape latency of each treated rats which exposed to aluminum for 1 and 3months decreased obviously with the extension of training time(P<0.05).The escape latency of rats which exposed to aluminum for 3 months increased with increasing exposure dose(P<0.05).According to the results of LSD assay in rats exposed to aluminum for three months, the escape latency of mediumgroup, and high group increased compared with control group(P<0.05); the escape latency of high group increased compared with low group(P<0.05). According to repeat measure analyze assay, the escape latency of each group in the hidden platform test was affected by training time and aluminum dose, but not found interaction. According to the result of the test which exposed for 1 months during probe trains, low group, medium group, and high group decreased the time spent in the target quadrant compare with control group(P<0.05); low group, medium group, and high group decreased the number of crossing the platform(P<0.05). According to the result of the test which exposed for 3 months, we found that low group, medium group, and high group decreased the time spent in the target quadrant compare with control group(P<0.05); medium group, and high group decreased the number of crossing the platform(P<0.05). high group decreased the time spent in the target quadrant and the number of crossing the platform compare with low group(P<0.05). According to the result of the test which exposure for 1 months during the open-field test, high group increased central lattice residence time compare with control group and low group(P<0.05). According to the result of the test which exposed for 3 months, medium group, and high group increased central lattice residence time compare with control group(P<0.05); high group increased central lattice residence time compare with low group(P<0.05).2.electrophysiological grapher: According to the result of the test which exposed for 1 months, the average of f EPSPs was decreased at different time point in all exposed groups compared with control group(P<0.05); the average of f EPSPs was decreased at 30 min,40min,50 min,60min in medium group, and high group compared with low group(P<0.05). According to the result of the test which exposed for 3 months, the average of f EPSPs was decreased at different time point in all exposed groups compared with control group(P<0.05); the average of f EPSPs was decreased at different time point in medium group, and high group compared with low group(P<0.05); the average of f EPSPs was decreased at 30 min,40min,50 min,60min in high group compared with medium group(P<0.05).3. The result of flow cytometer: The rate of apoptosis of the rats which exposed for 1,3months was significantly increased with increasing aluminum dose. Compared with control group, the rate of apoptosis which exposed for 1 month was significantly increasedin medium dose group and the dose group(P<0.05); Compared with low group, the rate of apoptosis which exposed for 1 month was significantly increased in high dose group(P<0.05). Compared with control group, the rate of apoptosis which exposed for 3 month was significantly increased in medium dose group and high dose group(P<0.05); Compared with low group, the rate of apoptosis which exposed for 3 month was significantly increased in medium dose group and high dose group(P<0.05).4. The result of real-time PCR: We found it about the first month result that the gene relative expression of caspase-3 in medium dose group and high dose group was significantly increased compared with control group(P < 0.05); the gene relative expression of caspase-8 in high dose group was significantly increased compared with control group(P<0.05); the gene relative expression of caspase-9 which have a certain trend was not significantly increased compared with control group(P>0.05). The third month result showed that the gene relative expression of caspase-3 in medium dose group and high dose group was significantly increased compared with control group(P<0.05); the gene relative expression of caspase-3 in medium dose group and high dose group was significantly increased compared with low group(P<0.05); the gene relative expression of caspase-9 in medium dose group and high dose group was significantly increased compared with control group(P<0.05); the gene relative expression of caspase-9 in medium dose group and high dose group was significantly increased compared with low group(P<0.05); the gene relative expression of caspase-8 which have a certain trend was not significantly increased compared with control group(P<0.05).5. The result of Elisa: We found that caspase-3,caspase-8, caspase-9,NMDAR1, NMDAR2 A,and cytochrome C had a certain dose dependent in the result of first month. The protein relative expression of NMDAR2 A in medium dose group and high dose group was significantly increased compared with control group and the low dose group(P<0.05). The protein relative expression of cytochrome C in high dose group was significantly increased compared with control group(P<0.05). The result of the third month displayed that the protein relative expression of caspase-3 in low dose group, medium dose group and high dose group was significantly increased compared with control group(P<0.05); the protein relative expression of caspase-9 in low dose group, medium dose group and high dose group was significantly increased compared with control group(P<0.05); theprotein relative expression of NMDAR2 A in low dose group, medium dose group and high dose group was significantly increased compared with control group(P<0.05); the protein relative expression of cytochrome C NMDAR1 in medium dose group and high dose group was significantly increased compared with control group(P<0.05); the protein relative expression of NMDAR2 A,cytochrome C in high dose group was significantly increased compared with low group(P<0.05).6. The anaylsis of relation: The correlation between the rate of hippocampal neuron apoptosis and the f EPSP is-0.692(P<0.05) in the result of first month.; The correlation between the rate of hippocampal neuron apoptosis and the f EPSP is-0.755(P<0.05) in the result of third month.Conclusions:1. Aluminum lactate exposure may induced the impairment of neurobehavioral function in SD rats, and the damage would more serious with the increase of the exposure dose and time.2. Aluminum lactate exposure may inhibited the LTP of the hippocampus in rats, and the inhibition was interaction with the exposure dose and time.3. The rate of apoptosis increased with the increase of the aluminum lactate exposure dose and time.4. Aluminum exposure may increased the gene and protein relative expression, and induced neuronal apoptosis eventually. The possible mechanism of neuronal apoptosis was carried out by both the death receptor pathway and the mitochondrial pathway, but the function of the mitochondrial pathway was more important.5.There was a relation between neuron apoptosis and the damage of hippocampal synaptic plasticity induced by Aluminum, and the correlation would enhanced with the increase of time. |
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