The Exploration On The Neuroprotective Effect And Related Mechanism Of PAC1R Allosteric Modulator SPAM1 In Three Mouse Models

Objective: Pituitary adenylate cyclase activating polypeptide(PACAP)can activate Pituitary adenylate cyclase activating polypeptide receptor 1(PAC1R)in vivo through pituitary adenylate cyclase activating polypeptide receptor 1(PAC1R).Plays antioxidant,anti-apoptotic and neuroprotective effects.However,in previous studies of our group,we have successfully screened a small molecular positive allosteric modulator 1(SPAM1),which can act on the allosteric regulatory site of PAC1 R and initiate the differentiation of PAC1 R.structure regulation,thereby exerting antioxidant,anti-apoptotic and neuroprotective effects.This study mainly explored the role of SPAM1 in three different mouse models(Scop-induced mouse memory impairment model,MPTP-induced PD mouse model,and D-gal-induced progeria mouse model).Methods: This study 1)In the Scop-induced mouse memory impairment model,each experimental group was intraperitoneally injected for 14 days {the drug injections in each experimental group were as follows: NOR group(0.9Saline + 0.9Saline),Scop group(0.9% Saline + 3mg/kg/day Scop),SPAM1 group(100μmol/kg/day SPAM1 + 0.9%Saline),SPAM1+Scop group(100μmol/kg/day SPAM1 + 3mg/kg/day Scop),VIP-TAT+Scop group(100nmol/kg/day VIP-TAT+ 3mg/kg/day Scop)}.After modeling,memory impairment was assessed by dark avoidance experiments,and apoptosis and oxidation levels in brain tissue were assessed by detecting Caspase3,MDA and SOD.2)In the MPP+-induced PD cell model,the effect of SPAM1 on apoptosis was assessed by detecting the content of Caspase3.In the MPTP-induced PD mouse model,each experimental group was injected intraperitoneally,only the drug was injected without MPTP in the first 14 days,and both the drug and MPTP were injected in the next 7 days {the drug injections in each experimental group were as follows: NOR group(0.9 %Saline + 0.9%Saline),MPTP group(0.9%Saline + 25mg/kg/day MPTP),L-SPAM1 group(10μmol/kg/day SPAM1 +25mg/kg/day MPTP),H-SPAM1 group(100μmol/kg/day SPAM1 + 25mg/kg/day MPTP),DOX group(100μmol/kg/day DOX + 25mg/kg/day MPTP),PACAP38 group(100nmol/kg/day PACAP38 + 25mg/kg/day MPTP)}.After modeling,behavioral indexes were detected by pole climbing test,hanging test and open field test,and the contents of DA,DOPAC and HVA in striatum,and the contents of SOD,MDA and T-AOC in brain tissue were determined.3)In the D-gal-induced mouse model of premature aging,the drugs in each experimental group were injected subcutaneously on the back of the neck,while D-gal was injected intraperitoneally,and the two were administered simultaneously for a period of 42 days {the drug injections in each experimental group were as follows: NOR group(0.9%Saline + 0.9Saline),D-gal group(120mg/kg/day D-gal + 0.9%Saline),L-SPAM1 group(120mg/kg/day D-gal +10μmol/kg/day SPAM1),H-SPAM1(120mg/kg/day D-gal + 100μmol/kg/day SPAM1)and VIPTAT(120mg/kg/day D-gal + 100nmol/kg/day VIP-TAT)}.After modeling,the positive signal levels of PACAP and PAC1 R near the hippocampal dentate gyrus were detected by immunohistochemistry of brain tissue sections.Results: 1)In the Scop-induced memory impairment model,the administration of SPAM1 could significantly prolong the delay time in the dark avoidance experiment,up-regulate the level of SOD,and down-regulate the levels of Caspase3 and MDA,suggesting that SPAM1 has the ability to resist memory impairment,anti-apoptosis and MDA in vivo.Antioxidant effect.2)In the MPP+-induced PD cell model,the treatment of SPAM1 can significantly down-regulate the level of Caspase3,and the effect of H-SPAM1 group is better than that of L-SPAM1 group.In the MPTPinduced PD mouse model,SPAM1 administration can improve various behavioral indicators in pole climbing,hanging and open field experiments,and significantly up-regulate the levels of DA,DOPAC and HVA in the striatum,and down-regulate Caspase3 in the brain tissue.and MDA levels,and up-regulated the level of SOD in brain tissue,suggesting that SPAM1 administration has the effects of improving mouse behavioral ability,alleviating PD symptoms,anti-oxidation and antiapoptosis in PD mouse model.3)In the D-gal-induced premature aging mouse model,the administration of SPAM1 can significantly increase the positive signal levels of PACAP and PAC1 R near the hippocampal dentate gyrus.Conclusion: SPAM1 can effectively resist Scop-induced memory impairment in mice,alleviate MPTP-induced PD symptoms,and improve the behavioral,oxidative and apoptotic indexes of mice,suggesting that SPAM1 has significant antioxidant,anti-apoptotic and neuroprotective effects.At the same time,SPAM1 can also up-regulate the positive signal levels of PACAP and PAC1 R in the hippocampal dentate gyrus in D-gal-induced premature aging mice,suggesting that SPAM1 promotes the expression of PACAP and PAC1 R through allosteric regulation,thereby exerting antioxidant,anti-apoptotic and neuroprotective effect.