Effects Of Neuron Autophagy Induced By Arsenic And Fluoride On Spatial Learning And Memory In Offspring Rats

Objective:To investigate the effects of arsenic and/or fluoride exposure on spatial learning and memory in early life to adult,autophagy lysosome and synaptic protein expression of neurons,and explore the mechanism of autophagy in the decline of learning memory between arsenic,fluorine alone or in a combined role.Methods:Forty female SD rats were randomly divided into 9 groups.Na As O₂（0,35,70 mg/L）and Na F（0,50,100 mg/L）were designed according to 3×3 factorial design,each group with4-5.These rats were exposed to the toxin by drinking water freely from 1 week before mating to 90 days after birth.At each timepoint（PND 21,42,90）,eight rats were selected,half male and half female.Urine was reserved for measuring the contents of arsenic and fluorine.After completing Morris Water Maze and Y Maze,animals were sacrificed and taken the bone of the right leg and brain tissue to measure fluoride content and arsenic content,respectively.The bilateral hippocampus of the remaining six pups was taken for the determination of protein expression,including autophagy-lysosome（Beclin1,LC3B,p62,Lamp2）,synaptophysin（SYN）and axon antiporter regulatory protein 1（JIP1）.Results:1.Arsenic and（or）fluoride exposure could induce significant body weight loss（P<0.05）.Except for As₇₀F₁₀₀group,the coefficient of the other exposure groups at PND21 was significantly lower than that of Con group（P<0.05）.While at PND42 and PND90,the coefficient of brain in exposed groups was increased significantly（P<0.05）.2.Arsenic and fluoride contents:The contents of arsenic in urine and brain tissue increased with the increase of arsenic dose（P<0.05）,the contents of fluoride in urine and bone increased with the increase of fluoride dose（P<0.05）.Fluoride antagonized the excretion of total arsenic in urine.3.Changes of spatial learning and memory ability:In Morris Water Maze,the escape latency of Con group was lower than that of the other groups,the escape latency in As₃₅F₅₀group was significantly higher than that in the other groups（P<0.05）.In the first 2 days and the 4th day,the number of crossing platform in each group was significantly lower than that in the other groups（P<0.05）.In Y Maze,both the present of time and the present of entry times of New Arm in exposed groups decreased to different degrees.4.Arsenic and/or fluoride exposure could induce pathological changes,such as the sparse and disordered neurons,swelling of cells and structural changes of nuclei.The ultrastructural damage of different organelles was also observed by electron microscopy. Synaptic morphology and formation of autophagic vesicles were abnormal.5.Arsenic and/or fluorine exposure increased the expression of Beclin1 and LC3B to activate autophagy.The autophagy activation of As₃₅F₅₀ group and As₃₅F₁₀₀ group was not as high as the other exposed groups.P62 increased in the arsenic or fluoride exposed groups and As₃₅F₅₀ group.Indeed,As₇₀ group was significantly higher than Con group （P<0.05）.The expression of LAMP2 increased in As₇₀ group,F₅₀ group,F₁₀₀ group and As₃₅F₅₀ group.The degradation of autophagy lysosome was inhibited.P62 in As₇₀F₅₀ group and As₇₀F₁₀₀ group and LAMP2 of As₇₀F₁₀₀ group were significantly lower than those in As₇₀ group（P<0.05）,suggesting high arsenic combined fluorine alleviated autophagy flux block.Arsenic and/or fluorine exposure reduced SYN expression （P<0.05）,disrupting synaptic structure and function.JIP1 in As₇₀F₅₀ group,As₃₅F₁₀₀ group,As₇₀F₁₀₀ group was significantly reduced（P<0.05）.Arsenic and fluoride exposure could regulate the reverse transport of axon autophagy,increase neuron synapse damage and finally induce neurotoxicity.6.There was an interaction between arsenic and fluoride exposure-induced changes of autophagic lysosome-related protein expression in hippocampus,showing antagonism.The antagonism of the high arsenic groups was stronger than that of the low arsenic groups.Conclusion:1.Exposure to arsenic and/or fluoride can impair offspring’s growth and development,spatial learning and memory,and the combined toxicity is stronger than the single toxicity.2.Exposure to arsenic and/or fluoride early in life could damage neuronal cells,internal organelles,and synaptic morphology.3.Exposure to arsenic and/or fluoride could inhibit reverse transport of autophagosomes on axons and induce autophagy defects,which damaged synaptic morphology and functions and finally induce neurotoxicity.There was an interaction between arsenic and fluorine,and the type of joint action is antagonists.