Neurodevelopmental Toxic Effects Of A Novel Nanomaterial ZIF-67 On Zebrafish(Danio Rerio) Embryos

Zeolite-imidazolate framework materials（ZIFs）are a subfamily of metal-organic frameworks（MOFs）.Tetrahedral coordination is formed mainly by transition metals such as metal ions,zinc,cobalt and imidazolester frames by self-assembly.Among them,ZIF-67 is synthesized from transition metal ions Co and 2-methylimidazole,and its pores are neatly arranged,with large specific surface area,heat resistance and topological structure,strong chemical stability than other MOFs materials,and are widely used in catalytic synthesis,adsorption of harmful substances,biological imaging,protection and storage of biological macromolecules,drug delivery and other fields.ZIFs materials can affect the system and function of organisms,but there are few reports on neurodevelopmental toxicity,so ZIF-67 was used in this study to explore the neurodevelopmental effects of ZIFs exposure on zebrafish embryos.In order to study the toxic effect and mechanism of ZIF-67 on the neurodevelopmental process of zebrafish embryos,zebrafish embryos were exposed to different concentrations of ZIF-67,and the growth and development parameters of zebrafish embryos were recorded during exposure.Results showed that ZIF-67 exposure caused increased mortality,decreased heart rate,decreased hatchability,increased malformation rate,and significantly inhibited body length of zebrafish embryos.Then,the effects of ZIF-67 on the early motor behavior of zebrafish embryos were evaluated by assessing the frequency of voluntary movement and mechanical touch avoidance behavior.Results showed that ZIF-67 exposure led to a decrease in the frequency of voluntary movements of zebrafish embryos and a decrease in the number of mechanical touch evasions.Then,high-throughput transcriptome sequencing was performed on zebrafish embryos,and the results showed that 1673 differentially expressed genes（DGEs）were significantly up-regulated and 756 DEGs were significantly down-regulated in zebrafish embryos after ZIF-67 exposure.GO enrichment analysis showed that DEGs were significantly enriched in immune system processes and immune responses in biological processes.The cellular components are mainly enriched in the extracellular matrix;under the molecular function category,it was mainly enriched in intrapeptide chain enzyme activity.After KEGG Pathway analysis,the neurodevelopmental toxicity of ZIF-67 on zebrafish embryos may be induced by affecting glutathione metabolism,apoptosis,and NOD-like receptor signaling pathways.Then,the transcriptional expression of the neurodevelopment-related genes gap43,gfap,elavl₃ andα1-tublin in zebrafish embryos exposed to ZIF-67 was analyzed by quantitative Real-Time q PCR（RT-q PCR）.Results showed that the m RNA expression of gap43,gfap,elavl3 andα1-tublin was reduced significantly.Secondly,the m RNA expression of neurotransmitter receptor genes of zebrafish embryo was measured,and the results showed that with the increase of exposure concentration,the m RNA expression of DA receptor drd2a and drd2b decreased significantly.The m RNA expression of 5-HT receptor 5-htr1ab and GABA receptor gabra was reduced significantly after ZIF-67exposure.The m RNA expression ofα1-andα3-nicotinic cholinergic receptors chrna1and chrna3 was reduced significantly,while the m RNA expression ofγnicotinic cholinergic receptor chrng was increased.The m RNA expression of acetylcholinesterase gene ache was reduced significantly and the activity of acetylcholinesterase in zebrafish embryos was also decreased after ZIF-67 exposure.It was deduced that ZIF-67 exposure induces neurotoxic effects by affecting the growth,the development of nerves and interfering with neurotransmitter pathways which lead to the disordered early motor behavior.