The Distribution Of Serotonin In The Brain Of SOD1 G93A Transgenic Mice And Its Correlation With The Mechanism Of Amyotrophic Lateral Sclerosis

Objective:Amyotrophic Lateral Sclerosis is a chronic progressive neurodegenerative disease characterized by a series of changes in upper and lower motor neurons.The pathogenesis of ALS is currently unknown,no specific treatment.Serotonin(5-HT)participates in the pathogenesis of ALS,but its alteration features and potential effects haven’t completely clarified.Tryptophan hydroxylase 2(TPH2)is a rate-limiting enzyme for 5-HT synthesis.Therefore,we systematically observed and analyzed the distribution characteristics and expression changes of 5-HT and TPH2 in the brain of SOD1 G93 A transgenic mice.The expression of Mammalian achaete-scute homologue mash1,an upstream regulation gene necessary for the development of serotonin neurons,in the brain of SOD1 G93 A transgenic mice was also observed.Methods:The G93 A SOD1 C57BL/6J transgenic mouse model was established,and the transgenic mice were detected and screened by PCR.Wild type WT mice and G93 A SOD1 transgenic mice were divided into pre-onset stage(60-70 days),onset stage(90-100 days)and progression stage(120-130 days).The wild-type WT mice and G93 A SOD1 transgenic mice were stained with fluorescence immunohistochemistry in different brain sections,TPH2 and 5-HT were labeled with TPH2 antibody and Serotonin antibody,respectively.mash1 antibody was used to label cells expressing mash1.The distribution characteristics of 5-HT,TPH2 and mash1 were observed and analyzed under the same field of vision and magnification under a fluorescence microscope,the positive expression distribution of wild-type WT mice and G93 A SOD1 transgenic mice in different anatomic areas at different stages of disease was statistically analyzed,and the correlation between the change of TPH2 expression and the change of mash1 positive expression was analyzed.Results:1.Animal immunofluorescence assay showed that 5-HT and TPH2 had the same expression sites and the same morphological characteristics,and positive expression of TPH2 could be seen in different functional areas of the brain.Distributing in glomerular layer of the olfactory bulb(GI)、accumbens nucleus(Acb)、cingulum(cg)、fimbria of hippocampus(fi)、mediodorsal thalamic nucleus(MD)、habenular nucleus(Hb)、 ventromedial hypothalamic nucleus(VMH)、 lateral hypothalamic area(LH).The expression of LH was the most abundant in the brain,followed by VMH and MD.TPH2 was distributed in the nucleus cluster in GI、Hb、MD、VMH,axonal distribution in LH,Acb,filamentous distribution in fi,curved track distribution in cg,and cell-like distribution in brainstem.2.Compared with the normal control group,the distribution of TPH2 expression in SOD1 G93 A mice was significantly decreased in various functional expression sites of the brain at various stages of disease.The expression distribution of GI,Acb,cg,fi,MD,Hb and VMH in WT mice at the onset stage was the highest,and lowest in the progression stage,while the expression distribution of TPH2 in LH was gradually increased with the increase of age of mice.The expression distribution of TPH2 in fi,MD,Hb,VMH and LH was the highest in the SOD1 G93 A group during the onset stage,and the lowest in the progression stage,cg gradually decreases with the progression of the disease,while Acb and GI are the lowest in the onset stage and the highest in the pre-onset stage.3.The protein expression of mash1 was mainly in the subependymal zone(SVZ).Compared with WT mice,the positive expression of mash1 in SVZ was significantly decreased in SOD1 G93 A mice,consistented with changes in positive TPH2 expression in the brain.Correlation analysis showed that in WT mice and SOD1 G93 A mice,the change of mash1 positive expression in SVZ was positively correlated with the change of TPH2 expression in fi,Hb,MD,VMH and LH at different stages.Conclusions:This study revealed the distribution and expression changes of 5-TH,TPH2 and mash1 in the brain anatomical area,and TPH2 was significantly reduced in several brain regions of SOD1 G93 A mice.Compared with the normal control group,the positive expression of mash1 in SOD1 G93 A mice was significantly reduced.The expression of TPH2 in the mouse brain was significantly correlated with the expression of mash1 in the SVZ region.5-TH participates in the pathogenic of ALS-like mice,it mainly regulates the expression of the upstream regulatory gene mash1,which is necessary for the development of serotonin neurons,in SVZ.