Effects Of Chronic Exposure To 5 Typical Nanomaterials On Locomotory Behaviors And Related Neurons In Caenorhabditis Elegans

Nanomaterials have unique properties,and are extensively used in fields of biomedicine,telecommunication,environmental protection,energy,national security in recent years.Among nanomaterials,graphene(G)and graphene oxide(GO)are most common kinds;silver nanoparticles(Ag-NPs)and zinc oxide nanoparticles(ZnO-NPs)are recently applied in new materials and medical technology.In the process of production,utilization and disposal,fragments of nanomaterials will occur in the air,water,soil and organisms.Nanomaterials will probably threaten the ecological safety,and its bio-toxicity and the potential risks to human health receive much concern.Biological effects including reproductive toxicity and developmental toxicity of some nanomaterials have been revealed in previous studies.However,their impacts on environment and health are still not well understood,especially about risks of chronic neurotoxicity.In this study,Caenorhabditis elegans was selected as a model organism,due to the advantages of a short life cycle and a simple nervous system.Five kinds of commonly used nanomaterials,graphene,graphene oxide,graphene quantum dots(GQDs),Ag-NPs and ZnO-NPs were studied in detail.After phy-chemical properties of these nanomaterials were characterized,the lethal concentrations were measured for 6 d exposure of suspension liquids in C.elegans.The effects of chronic exposure to these nanomaterials on locomotor behaviors of nematodes and their neurotoxicity were fully investigated under sub-lethal exposure conditions.The toxic effects of these nanomaterials on dopaminergic,glutamate and GABAergic neurons were compared using fluorescence-labeled transgenic nematodes.Main results include that::(1)G and GO are in the form of flakes,with a thickness of 0.8-1.2 nm.GQDs,Ag-NPs and ZnO-NPs are in the form of granules with diameters of 30-50 nm.After chronically exposed to less than 100mg/L G and GQDs for 6 d,C.elegans showed no lethal effects;while 50-100mg/L GO resulted in significant lethality,the median lethal concentration(LC50)was 51.22 mg/L.However,Ag-NPs and ZnO-NPs had strong lethal effects on nematodes;LC50 was 2.723 mg/L and 1.852 mg/L,respectively.(2)After exposure,graphene and GO nanomaterials had extensive accumulated and distributed in the body of nematodes.For example,G and GO nanosheets were primarily distributed in pharynx and intestine;while GQDs were widely distributed in various organs of nematodes in dose-dependent manners.Chronic exposure to five nanomaterials caused inhibitory effects on the growth of the nematodes,such as shorter body than the controls.(3)Chronic exposure to five nanomaterials caused inhibitory effects on the movement behavior of the nematodes in concentration dependent manners.Nanomaterials resulted in significant declines in the frequency of head thrashing,body bending and pharyngeal pumping,which indicated the damage of locomotor behaviors.Furthermore,we found that crawling tracks were irregularly curved,crawling distances were shortened,amplitude of bending wave became smaller and the crawling angles changed slower in the exposed nematodes in comparison with the controls.These results indicated that nematodes presented characteristics of movement impairment including bradykinesia,hypokinesia and unbalance and so on.(4)Using transgenic nematodes,results showed that high concentrations(50-100mg/L)of three kinds of graphene-based nanomaterials could inhibit the protein expression of dat-1?GFP and eat-4::GFP,but unc-47::GFP expression did not change.Morphology results showed deformation of the cell body,dendritic fracture and other damages in dopaminergic and glutamate neurons.Exposure of Ag-NPs and ZnO-NPs resulted in a significant decrease in the fluorescence intensity of dat-1::GFP and unc-47::GFP,but no changes in the expression of eat-4? GFP.The results indicated that these metal nanomaterials were susceptible to dopaminergic neurons and GABAergic neurons.These results indicate that:(1)Five kinds of commonly used nanomaterials possess chronic toxicity to Caenorhabditis elegans.To comparison with LC50 and growth inhibition effects,toxicity of two metal nanoparticles is stronger than that of graphene-based nanomaterials.The order of toxicity is:ZnO-NPs>Ag-NPs>GO>G>GQDs.(2)Chronic exposure to sub-lethal concentrations of these nanomaterials causes neurotoxicity with different degrees.Chronic neurotoxicity includes behavioral deficits and neural damages in C.elegans.Behavioral toxicity shows impairment character such as bradykinesia,hypokinesia and unbalance and so on.In addition,metal and graphene-based nanomaterials have different susceptibility to dopaminergic neurons,GABAergic neurons and glutamate neurons,and detailed mechanisms need to be further studiedIn the present study,we have fully revealed the health risk of commonly used nanomaterials,behavioral deficits and neurotoxicity induced.In recent year,nanomaterials have been increasing used in daily life.These findings provide important references for risk evaluation,safety management and standard establishment of nanomaterials.This study also builds the foundation for the further study on the toxicity mechanism and preventive measures of nanomaterials.