The Changes Of Acrylamide On Motor Function And Neural Mechanisms Of Toxicity Of Rats

ObjectiveAcrylamide (acrylamide,ACR) is a widely used industrial chemical with strong toxicity.It is widely used in wastewater treatment,oil,paper,resin processing,textiles,leather and other fields. In recent years,people begin to focus on high-temperature cooking food on human health.ACRis known to produce central-peripheral distal axonopathy,which is charactrized by ataxia and distal skeletal muscle weakness.The major pathological hallmarks are sistal swellings and secondary degeneration both in experimental animals and human companioning excessive accumulation of neurofilaments(NFs)in the distal swollen axon.However,the exact mechanism of action of its neurotoxicity has not been completely clear until now.Based on the pathological alterations,we built ACR-intoxicated rats models and investigated the relative levels of motor function of rats and changes in learning and memory,then measured the changes of calcium (Ca2+),calpain activity and phosphorylation of the central and peripheral nervous tissue and serum.At last,we detected pathological changes of the brain,spinal cord and sciatic nerve,which will offer the cues and evidences for exploring the mechanism of ACR neurotoxicity and searching biomarkers for early clinical diagnosis.Methods 1 Male Wistar rats,weighing 180-220g were randomly divided into 3 groups(n=14 rats per group).The rats in group 1served as control,and received normal saline.The animals in group 2 and 3 were given ACR dissolved in physioligical saline(20,40mg/kg ip 3days/week) for 8 weeks.2 Neurobehavioral index determination:Observe general changes of rats Motor function indexes was determined per week.Special tests included observations of spontaneous locomotion in an open field,measures of swim and grip time. Learing and memory ability was tested by Morris Water maze (MWM).3 The spinal cord tissue were Separated and digested with trypsin into cells.Then the cells were loaded with Fura-2/AM,which were determined with Fluorescence spectrophotometer.The decapitated bloods were sampled,and the serum were separated routinely.The calcium, of serum were investigated with calcium kit.4 The activity of Calpain of brain, spinal cord and sciatic nerve were determined by using calpain substrateâ…¡with fluorescence spectrophotometer to derive fluorescence value.5 The morphology of the brain,spinal cord and sciatic nerve was observed by light microscopy and electron microscope,and we determine whether there is morphological changes.6 We use immunohistochemistry (SP) to measure the changes of NF non-phosphorylated and phosphorylation of spinal cord and brain.Results1 General observation and Neurobehavioral index;The weight of rats treated with acrylamide was decreased (P<0.05).Gait score of rats at High-dose group was increased. Compared with the control group,Swim time of rats at low-dose and high-dose group was significantly Shorten.Grip time in grip test at low-dose and high-dose group was significantly Shorten,too.Between ACR and the control groups there were significant differences in MWM (P<0.05),the latency at high-dose group was extended and the distance was increased.2 Cell calcium of spinal cord and serum:Cell calcium of spinal cord of rats treated with acrylamide was creased (P<0.05).Cell calcium of serum was shorten (P<0.05).3 Results of calpain activity show that,compared with control group,brain activity calpain was reduced (P<0.05),while calpain activity of spinal cord and the sciatic nerve was increased (P<0.05).4 Morphologic detection1) Compared to control,HE staining did not reveal overt neuronal loss in the cerebral cortex and the spinal cord after ACR administration;We did not see the changes of cells,but see concentrated dense chromatin in both groups;sciatic nerve were found myelin swelling, axonal swelling or disappear.2) Electron microscopy:Spinal cord and sciatic nerve were found fibrosis, osteoporosis,deformation,shrinkage,fracture and disintegration.wo saw that NF were increased and gathered in Sciatic nerve myelin.5 Immunohistochemistry confirmed that the neurons of non-phosphorylated in Spinal anterior horn was stained compared control,and phosphorylated was light staining.The neurons of phosphorylated in brain were stained which did not have axons.Conclusion1 Acrylamide can damage motor function and learning-memory capability of rats.2 Acrylamide can cause calpain activity changes in brain,spinal cord and sciatic nerve.3 Acrylamide can cause cell calcium content changes of spinal cord of rats.4 Acrylamide can cause morphological changes of brain and spinal cord,rat sciatic nerve morphological changes.5 ACR can cause phosphorylation of changes of brain and spinal cord, and this has inevitable connection within the cell calcium and calpain activity.