Research On The Improvement Effect And Mechanism Of L-stephenidine On Methamphetamine-induced Learning And Memory Impairment

Objective: Drug addiction is a chronic,recurrent brain disease that is characterized by a continuous and compulsive intake of morphine,heroin,methamphetamine(METH),ketamine and other dependent drugs for non-medical purposes,which seriously endangers the health and safety of individuals,families and society.METH is a kind of amphetamine-type central stimulant,which is seriously harmful to the health of human,and brings new problems and challenges to narcotics control in China.Chronic use of METH has serious effects on cognitive functions,including decision making and learning and memory function mainly through the dopamine(DA)system.Many studies have suggested that DA neurotransmission is closely related to learning and memory ability.The inhibition of DRD1(D1 dopamine receptor,DRD1)can cause the decline of learning and memory,while the activation of DRD2(D2 dopamine receptor,DRD2)can also cause the decline of learning and memory.l-sepholidine(l-SPD),a tetrahydroprotoberberine alkaloid,exerts dual actions on brain DA receptors,eliciting partial DRD1 agonisttic activity and acting as DRD2 antagonistic activity.The unique pharmacological properties of l-SPD make it possible to improve the methamphetamine-induced spatial learning and memory impairment.Therefore,we investigated the effects of l-SPD on METH-induced memory impairment through the morris water maze and novel object recognition test experiments.Meanwhile,we investigated the expression of dendritic spines in theprefrontal cortex(PFC)through the Golgi silver staining,and investigated the expression of learning and memory related proteins in PFC though the Western blot.Through the above experiments,we explored the mechanism of the improvement effect of l-SPD on METH-induced learning and memory impairment,and provided a possible basis for clinical treatment of METH-induced learning and memory impairment by l-SPD.Methods: 1.Establishing a model of METH-induced memory impairment.A mice model of learning and memory impairment induced by METH was established by injected with METH 10mg/kg(i.p.)for 7 consecutive days.2.Animal behavior experiment.The effect of l-SPD on METH-induced spatial learning and memory impairment of mice was examined using the Morris water maze test.Then,the memory ability of mice was tested by the Novel Object Recognition Test.Through the above experiments,we explored whether l-SPD could improve METH-induced learning and memory impairment.3.Western blot experiment.We detected the changes in the expression of the learning and memory-related signaling pathway proteins in the prefrontal cortex(PFC),such as,DRD1,DRD2,dopamine transporter(DAT),protein kinase A(PKA),phosphorylated protein kinase A(p-PKA),cAMP-response element binding protein(CREB),phosphorylation of phosphorylated cAMP-response element binding protein(p-CREB)and brain derived neurotrophic factor(BDNF),to explore the mechanism of 1-SPD to improve METH-induced learning and memory impairment.4.Golgi silver staining experiment.The expression of dendritic spines in theprefrontal cortex was examined by the Golgi silver staining.We studied the mechanism of l-SPD to improve the METH-induced dendritic spine injury from the morphology.Results: 1.Compared with the control group,the escape latency in learning phase was significantly prolonged in the model group on the second/third/forth day(P<0.05,n=12),indicating that the spatial learning ability was impaired in the model group.Compared with the model group,the escape latency of l-SPD-treated groups in learning phase showed no significant difference,which indicated that l-SPD could not improve METH-induced learning impairment.Compared with the control,the number of crossing the platform in the model group was significantly reduced(P<0.05,n=12),indicating that the memory ability of the model group was impaired.Compared with the model group,the numbers of crossing the platform of the medium and high doses of l-SPD-treated groups were significantly increased(P<0.05,n=12),indicating that the medium and high doses of l-SPD treated groups could improve METH-induced memory impairment.Compared with the model group,the preference index of the model group for the new thing was significantly reduced in novel object recognition test(P<0.05,n=12),indicating that the memory ability of the model group was impaired.Compared with the model group,the preference index of l-SPD-treated groups for the new thing was significantly increased(P<0.05,n=12),indicating that l-SPD could improve METH-induced memory impairment.2.Compared with the model group,the protein expressions of DRD1,DAT,p-PKA,p-CREB,BDNF were significantly increased in the l-SPD treated group,indicatingthat l-SPD might improve METH-induced memory impairment by activating the DA system and the downstream PKA signaling pathway.3.Golgi silver staining experiments showed that the number of dendritic spines in the model group was significantly reduced compared with the control group(P<0.05,n=4),indicating that METH injured the dendritic spines on neurons.Compared with the model group,the number of dendritic spines of the l-SPD-treated groups in PFC neurons was increased(P<0.05,n=4).Conclusions: 1.Behavioral experiments showed that l-SPD could improve METH-induced memory impairment.2.Western blot analysis showed that l-SPD might improve METH-induced memory impairment by DRD1-PKA-BDNF pathway.3.Golgi silver staining experiments showed that l-SPD improved the damage of dendritic spine in PFC induced by METH.