The Effectof High-intensity Interval Trainingonthe Mitochondrial Quality Control Systemin Mice Hippocampusand Analysisofthe Role Of Lactate

Objective:High-intensity interval training(HIIT)has been proved to promote Brain-derived neurotrophic factor(BDNF)expression and effectively improve cognition.However,the underlying neurobiological mechanism has not been fully elucidated.One of the potential mechanisms is the enhanced energetic efficiency of mitochondria production since exercise can improve mitochondrial adaptation and antioxidants.Recently,lactate,known as the classical metabolite of HIIT,is proved to regulate mitochondrial-related genes and proteins expression in L6 and SY5Y cells.Therefore,we speculate that the increased brain lactate,produced by intensive exercise,is responsible for the superior effect of HIIT on synaptic plasticity through regulating the mitochondrial quality control system.In vivo and in vitro studies were used for exploring the mitochondrial-related mechanism of synaptic plasticity improvement and revealing the role of lactate.Methods:?in vivo study:64 male mice were 7 weeks of age at the start of the experiments.After one week acclimatization period,they were divided randomly into two groups(n=32 each group):the control group(Ctl group)and the high-intensity interval training group(HIIT group).Mice in the HIIT group completed 6-week HIIT after 5-day adaptive training.Blood lactate was detected by Lactate-Scout immediately at the end of 2^nd,4^th,and 6^thweek after completing the 10 bouts of training.10 of all mice in each group were collected from tail tip blood.At the end of the 6-week training and after the anesthetization,20 mice(n=10 each group)were sacrificed immediately.Hippocampal lactate levels were measured using the microplate reader.At the end of the 6-week training,24 hours after the last training and after the anesthetization,30 mice were sacrificed(n=15 each group).24 mice hippocampus(n=12 each group)were carefully dissected out for detecting physiological and biochemical indexes.The other 6 mice brain(n=3 each group)were dissected out for morphological observation.The distinction between two groups of dendritic branches and spine density in CA1 pyramidal neurons was observed by Golgi staining for understanding the effect of HIIT on mice hippocampal synaptic plasticity.The monocarboxylic acid transporter(MCT1 and MCT4),l-lactate receptor GPR81,mitochondrial fusion(OPA1,MFN1,and MFN2),mitochondrial fission(DRP1and FIS1),mitophagy(PINK1,PARKIN,P62,and LC3),and mitochondrial biogenesis(PGC-1?,NRF-1,NRF-2,TFAM,and mt DNA)were detected by Western blot and RT-PCR.The five complex genes of mitochondrial OXPHOS(NDUFS8?SDHb?Uqcrc1?COX5b?Atp5a1)were detected by RT-PCR.The ATP levels were detected by automatic microplate reader.The remaining mice(n=7 per group)were asked for completing the 5-day Morris water maze task.?in vitro study:Primary hippocampal cells were obtained from 1-day-old neonatal C56BL/6J mice with the neurobasal medium.On the 7^thday,hippocampal cells were treated with l-lactate or GPR81 agonist 3Cl-5OH-BA.The experiment was divided into the Ctl group,l-lac group and 3Cl-5OH-BA group.The above indicators in vivo were detected.Results:?in vivo study:(1)The effect of HIIT on the lactate level and GPR81expression in mice:The blood lactate levels in the HIIT group can reach 6-10 m M immediately after training(p<0.01),which are significantly increased compared with the Ctl group.Meanwhile,the hippocampal lactate levels in the HIIT group were also significantly higher than those in the Ctl group(p<0.05).Moreover,HIIT promoted m RNA(p<0.01,p<0.05)and protein expression(p<0.01,p<0.01)of MCT1 and MCT4 in the mice hippocampus.Besides,HIIT significantly increased the GPR81protein and m RNA expression in mice hippocampus(p<0.01,p<0.05).(2)The effect of HIIT on the synaptic plasticity and the ability of learning and memory:HIIT significantly promoted BDNF protein expression(p<0.01).Meanwhile,the spine density(p<0.01)and the times across the platform(p<0.05)were also increased in the HIIT group.(3)The effect of HIIT on the mitochondrial function in mice hippocampus:HIIT significantly promoted COX5b gene expression(p<0.01).Meanwhile,we found that HIIT notably elevated hippocampal ATP levels(p<0.05).(4)The effect of HIIT on the mitochondrial quality control system in mice hippocampus:HIIT promoted the fusion protein expression of OPA1,MFN1,and MFN2(p<0.05,p<0.01,p<0.01),but significantly inhibited the fission proteins expression of DRP1 and FIS1(p<0.05,p<0.01).On the mitophagy,there was no significant difference between the two groups in protein expression of PINK1,PARKIN,and LC3,although the protein expression of P62 was decreased in the HIIT group compared to the Ctl group(p<0.01).On the mitochondrial biogenesis,HIIT promoted hippocampal PGC-1?and NRF2 protein expression(p<0.05,p<0.01),accompanied by the mitochondrial copy number increased significantly(p<0.01).?in vitro study:(1)The effect of l-lactate and GPR81 on the mitochondrial function in mice hippocampal cells:The ATP levels were increased as early as 15m M l-lactate treatment for 3 h in mice hippocampal cells(p<0.05).Then we found the gene levels of Uqcrc1 and Atp5a1 significantly increased after lactate treatment(p<0.01,p<0.05).The ATP levels were also increased as early as 3Cl-5OH-BA treatment for 3 h in mice hippocampal cells(p<0.01).However,there is no difference between the two groups in the OXPHOS related genes expression in mice hippocampal cells with3Cl-5OH-BA treatment.(2)The effect of l-lactate and GPR81 on BDNF expression in mice hippocampal cells:Both l-lactate(p<0.05)and GPR81 treatment(p<0.05)can boost the BDNF protein expression in the mice hippocampal cells.(3)The effect of l-lactate and GPR81 on the mitochondrial quality control system related proteins in mice hippocampal cells:L-lactate treatment significantly upregulated the expression of mitochondrial fusion protein MFN1 and MFN2(p<0.05,p<0.01).Meanwhile,the expression of mitochondrial fission protein DRP1 and FIS1 was inhibited after l-lactate treatment(p<0.05,p<0.05).As for 3Cl-5OH-BA treatment,it promoted the fusion protein expression of OPA1,MFN1,and MFN2(p<0.01,p<0.01,p<0.05).But it had no effect on the fission proteins expression of DRP1 and FIS1.On the mitophagy,l-lactate treatment had no effect on the mitophagy related proteins expression of PINK1,PARKIN,and P62,except for the significantly downregulated protein expression of LC3(p<0.05).And 3Cl-5OH-BA treatment had no effect these four mitophagy-related proteins.On the mitochondrial biogenesis,the l-lactate treatment improved the protein expression of PGC-1?,NRF2,and TFAM(p<0.05,p<0.01,p<0.05)and the mitochondria DNA copy number(p<0.05).And so well as the 3Cl-5OH-BA treatment.Conclusion:?HIIT can elevate the blood lactate and increase the expression of lactate transporter to promoting the brain lactate level and lactate receptor GPR81expression.?HIIT promoted mitochondrial fusion,inhibited fission,and improved mitochondrial biogenesis.This can contribute to the mitochondrial function improvement and thus synaptic plasticity.?Lactate mediated the effect of HIIT on the mitochondrial quality control system.