| Objective In this study,a rat model of aluminum exposure from lactation to pre-adulthood was established to simulate aluminum exposure during infant growth and development before adulthood,and to detect the relative expression levels of mi R-204-5p,target gene Eph B2 and synaptic plasticity downstream signal pathway NMDAR-ERK-CREB-Arc in rat hippocampus,so as to explore the effects of aluminum exposure on learning and memory ability of rats during growth and development.And the expression of mi R-204-5p,target gene Eph B2 and downstream related signal pathways.Methods In this study,Wistar rats(200±20g)were selected as the research objects.The rats were habituated to the environment for a week,they were mated according to male and female(2:1),and the rats were considered to be pregnant when the vaginal plug was observed the next day.Randomly divide the pregnant mother rats into groups,namely the control group and the Al-exposed model group(low dose group,middle dose group and high dose group).The mother rats in Al began to drink 2.0g/L,4.0g/L and 8.0g/L Al Cl3 distilled water respectively.Before weaning,rats were exposed to aluminum through the mother’s milk,and after weaning,they drank Al Cl30.0g/L,2.0g/L,4.0g/L and 8.0g/L distilled aqueous solution to ingest aluminum,and the time limit for exposure was from.The first day of life for rats lasted 90 days.The weight of rats in each group and hippocampus weight were observed;the spatial memory function of rats was examined through Morris experience;the ultrastructure of hippocampal neurons was observed by transmission electron microscope,the expression of mi R-204-5p m RNA,Eph B2,NMDAR2 B,ERK1/2,CREB and Arc m RNA in hippocampus was detected by reverse transcription-polymerase chain reaction(Real time RT-PCR),the protein content of Eph B2,NMDAR2 B,ERK1/2 and Arc in hippocampus was detected by Western Blot method,and the expression of CREB protein was detected by immunohistochemical method.Results 1.Changes in body weight of rats: The body weight of rats in each aluminum exposure group increased with the extension of feeding time.At the 0th week,the body weight of rats in the low dose aluminum group was lower than that in the control group,but there was no significant difference(P > 0.05),but the body weight in the middle and high dose aluminum groups was lower than that in the control group(P < 0.05).At the 4th week,the body weight of rats exposed to aluminum was lower than that of the control group(P < 0.05).At the 8th week,the body weight of the aluminum exposed group was lower than that of the control group(P < 0.05),and the body weight of the high dose aluminum exposed group was lower than that of the low and middle dose aluminum exposed groups(P < 0.05).At the 12 th week,the body weight of aluminum exposed group was lower than that of control group(P < 0.05),and the body weight of middle and high dose aluminum exposed group was lower than that of low dose aluminum exposed group(P < 0.05).The body weight of high dose aluminum exposed group was significantly lower than that of middle dose aluminum exposed group(P < 0.05).2.Morris water maze test showed that in the location navigation test,the escape latency of the middle and high dose aluminum groups was remarkably longer than that of the control group(P < 0.05),and the escape latency of the middle dose group was longer than that of the low dose group(P < 0.05),and the escape latency of the high dose group was longer than that of the middle dose group.The results of swimming distance showed that the swimming distance of the aluminum exposed group was larger than that of the control group(P < 0.05),and the total swimming distance of the high dose aluminum exposed group was significantly larger than that of the low and middle dose aluminum exposed groups(P < 0.05).In the spatial exploration experiment,the number of crossing the platform in the first quadrant in the aluminum exposure group was lower than that in the control group(P < 0.01),and the crossing times in the high dose aluminum exposure group was significantly lower than that in the low dose group(P < 0.01).The stay time in the target quadrant in each aluminum exposure group was shorter than that in the control group(P < 0.05,P < 0.01),and the stay time in the first quadrant in the middle and high dose aluminum exposure groups was significantly less than that in the low dose group(P < 0.05,P < 0.01).3.The results showed that the hippocampal organ coefficient of rats in each aluminum exposure group increased with the increase of aluminum concentration,and the hippocampal organ coefficient in the low dose aluminum exposure group was higher than that in the control group(P < 0.05).There was no significant difference in hippocampal organ coefficient between the middle dose aluminum exposure group and the control group(P > 0.05).The organ coefficient of hippocampus in the high dose aluminum exposure group was significantly higher than that in the control group(P < 0.01).4.The results of determination of hippocampal Al showed that the hippocampal Al concentration increased with the increase of Al concentration.Among them,It showed no remarkable difference in hippocampal Al concentration between low dose Al-exposed group and control group(P >0.05),but the Al concentration in middle and high dose Alexposed groups was remarkably higher than control group(P < 0.01).5.In the control group,the structure of hippocampal neurons in CA1 region was regular,the edge of cell membrane was clear,the structure was intact,the nucleus was regular round,and the chromatin was evenly distributed.However,with the increase of aluminum exposure concentration,the morphology of hippocampal neurons in the aluminum exposure model group changed significantly,including cell structure deformation,nuclear structure incomplete and invagination,nuclear membrane shrinkage,nuclear pyknosis and chromatin pyknosis to a certain extent.6.The results of Real-time RT-PCR experiment showed that the relative expression level of mi R-204-5P in the low dose aluminum group was higher than that in the control group,and the relative expression level of mi R-204-5P in the hippocampus of rats in the middle and high dose aluminum groups was remarkably higher than that in the control group(P<0.05).The relative expression level of Eph B2 m RNA in the low dose aluminum exposure group was lower than that in the control group(P<0.01),and the relative expression level of Eph B2 m RNA in the hippocampus of rats in the middle and high dose aluminum exposure groups was lower than that in the control and low dose aluminum exposure groups(P<0.05).The relative expression level of NMDAR2 B m RNA in the hippocampus of rats exposed to low,middle and high dose aluminum was lower than that of the control group(P < 0.01).Compared with the control group,the relative expression level of ERK1 m RNA in the hippocampus of the low dose aluminum group was lower than that of the control group(P<0.01),the relative expression level of ERK2 m RNA in the hippocampus of the low dose aluminum group was remarkably lower than that of the control group(P<0.05),and the relative expression level of ERK1/2 m RNA in the hippocampus of the middle and high dose aluminum groups was remarkably decreased(P<0.01).The relative expression level of CREB m RNA in hippocampus of rats exposed to aluminum in low and middle dose groups was lower than that in control group(P<0.01),and the relative expression level of CREB m RNA in hippocampus in high dose aluminum exposure group was lower than that in other dose groups(P < 0.05,P < 0.01).There was no significant difference in the relative expression of Arc m RNA between the low dose aluminum exposure group and the control group(P>0.05),but the relative expression level of Arc m RNA in the middle dose aluminum exposure group was lower than that in the control group(P<0.01),and the relative expression of Arc m RNA in the high dose aluminum exposure group was lower than that in other dose groups(P < 0.05,P < 0.01).7.Protein expression results: in the results of Western blot detection,there was no significant difference in the relative expression of Eph B2 protein in the hippocampus of the low dose aluminum group compared with the control group(P>0.05),while the relative expression of Eph B2 protein in the middle and high dose aluminum exposure groups was significantly lower than that in the control and low dose aluminum groups(P < 0.05).There was no significant difference in the relative expression of ERK1/2 protein between the low dose aluminum group and the control group(P>0.05),but the relative expression of ERK1/2 protein in the middle and high dose aluminum groups was significantly lower than that in the control and low dose groups(P < 0.05).There was no significant difference in the relative expression of NMDAR2 B protein between the low dose aluminum exposure group and the control group(P>0.05),but the relative expression of NMDAR2 B protein in the middle and high dose aluminum exposure groups was remarkably lower than that in the control and low dose aluminum exposure groups(P<0.05).There was no significant difference in the relative expression of ERK1/2 protein between the low dose aluminum exposure group and the control group(P>0.05),but the relative expression of ERK1/2 protein in the middle and high dose aluminum exposure groups was remarkably lower than that in the control and low dose aluminum exposure groups(P<0.05).There was no significant difference in the relative expression of Arc protein between the low dose aluminum exposure group and the control group(P>0.05),but the relative expression of Arc protein in the middle and high dose aluminum exposure groups was remarkably lower than that in the control and low dose aluminum exposure groups(P<0.05).There was no significant difference in the expression of CREB protein between the low dose aluminum group and the control group(P>0.05),but the expression of CREB protein in the hippocampus of the middle dose aluminum group was significantly decreased(P < 0.05).The expression of CREB protein in hippocampus of rats exposed to aluminum in high dose group was lower than that in control and low dose aluminum groups(P < 0.05).Conclusions 1.Aluminum exposure can accumulate in the hippocampus and increase the aluminum content,damage the morphology of nerve cells in the CA1 region of the hippocampus,and lead to impairment of learning and memory in rats.2.Aluminum exposure can promote the expression of mi R-204-5P m RNA in rat hippocampus,and inhibit the expression of Eph B2,NMDAR2 B,ERK1/2,CREB and Arc m RNA and protein.3.One of the mechanisms of aluminum exposure leading to impaired learning and memory in rats may be due to the regulation of mi R-204-5P m RNA expression to inhibit the expression of the target gene Eph B2 and its downstream related signaling pathway NMDAR-ERK-CREB-Arc. |
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