硫化汞纳米颗粒对秀丽隐杆线虫不同神经递质类型神经元的影响

Effect of mercuric sulfide nanoparticles on different neurotransmitter-type neurons in Caenorhabditis elegans

目的 探究硫化汞纳米颗粒(HgS-NP)暴露对秀丽隐杆线虫(简称线虫)不同神经递质类型神经元的影响。方法 采用野生型N2线虫以及绿色荧光蛋白特异性标记转运蛋白或生物合成酶的γ-氨基丁酸(GABA)能、谷氨酸(Glu)能、多巴胺(DA)能、乙酰胆碱(ACh)能和5-羟色胺(5-HT)能神经元的转基因线虫(EG1285,DA1240,BZ555,LX929和GR1366),通过固体暴露的方式,将同步化后的L4期线虫经不同浓度HgS-NP处理。(1) 5种转基因线虫暴露于HgS-NP 0(溶剂对照组),250,500和1000 mg·L^(-1)72 h,采用荧光显微镜观察转基因线虫不同神经递质类型神经元绿色荧光的表达。(2)野生型N2线虫暴露于HgS-NP 0(溶剂对照组)和1000 mg·L^(-1)72 h,采用实时荧光定量PCR(RT-qPCR)检测33种GABA能和Glu能神经递质系统相关基因mRNA表达水平。结果 (1)与同基因型溶剂对照组线虫相比,暴露于不同浓度HgS-NP后,EG1285线虫出现GABA能神经元胞体缩小、树突断裂和神经元缺失,神经元相对荧光强度显著降低(P<0.05);暴露于HgS-NP 1000 mg·L^(-1)后,DA1240线虫头部Glu能神经元缺失,相对荧光强度显著降低(P<0.01);然而,暴露于不同浓度HgS-NP后,BZ555、LX929和GR1366线虫的DA能、ACh能和5-HT能神经元形态及相对荧光强度均无明显改变。(2)与溶剂对照组相比,野生型N2线虫暴露于HgS-NP 1000 mg·L^(-1)后,仅编码非N-甲基-D-天冬氨酸类Glu受体的glr-7和glr-8的mRNA表达水平升高(P<0.05),其余31种GABA能和Glu能神经递质系统相关基因mRNA表达水平均无明显改变。结论 较高剂量的HgS-NP暴露72 h可损伤线虫的GABA能和Glu能神经元,而对DA能、ACh能和5-HT能神经元无明显影响。

OBJECTIVE To investigate the impact of mercuric sulfide nanoparticles(HgS-NP)on various neurotransmitter-type neurons in Caenorhabditis elegans(C.elegans).METHODS Wild-type N2 C.elegans,as well as the transgenic C.elegans expressing green fluorescent protein specifically in gamma-amino-butyric acid(GABA)-ergic,glutamatergic,dopaminergic,cholinergic,and serotoninergic neurons(strains EG1285,DA1240,BZ555,LX929,and GR1366),were synchronized at the L4 stage and treated with different concentrations of HgS-NP via solid exposure.①Five transgenic strains of C.elegans were exposed to HgS-NP 0(control group),250,500,and 1000 mg·L^(-1) for 72 h.Fluorescence microscopy was used to observe the expression of green fluorescence in different neurotransmitter-type neurons of the corresponding transgenic C.elegans.②Wild-type N2 C.elegans were exposed to HgS-NP 0(control group)and 1000 mg·L^(-1) for 72 h.Real-time quantitative PCR(RT-qPCR)was performed to detect the mRNA expression levels of 33 genes related to the GABAergic and glutamatergic neurotransmitter systems.RESULTS①Compared with the control group of the same genotype,the EG1285 C.elegans exhibited soma shrinkage,dendrite fragmentation,and neuronal loss,with a significant decrease in the relative fluorescence intensity of GABAergic neurons(P<0.05)after exposure to different concentra-tions of HgS-NP.After exposure to HgS-NP 1000 mg·L^(-1),the DA1240 C.elegans showed a loss of glutamatergic neurons in the head region and a significant decrease in the relative fluorescence intensity(P<0.01).However,there were no significant changes in the morphology or relative fluorescence intensity of dopaminergic,cholinergic,and serotoninergic neurons in BZ555,LX929,and GR1366 C.elegans after exposure to HgS-NP,respectively.②Compared with the control group,wild-type N2 C.elegans exposed to HgS-NP 1000 mg·L^(-1) showed increased mRNA expression levels of glr-7 and glr-8,which encoded non-N-methyl-D-aspartate glutamate receptors(P<0.05),while the expression levels of the remaining 31 genes related to the GABAergic and glutamatergic neurotransmitter systems showed no significant changes.CONCLUSION Exposure to a high dose of HgS-NP for 72 h may potentially damage the GABAergic and glutamatergic neurons in C.elegans,but have no significant effect on dopaminergic,cholinergic or serotoninergic neurons.