| Objective:As a common psychiatric disease,the core symptom of depression is depressed mood and a loss of interest and/or pleasure,and the global incidence is about 16%,which causes a serious social and economic burden.Traditional antidepressant drugs are targeted in the monoamine energy system,such as tricyclic antidepressants,but these drugs have disadvantages such as slow effect and inefficient.The first clinical study in 2000 showed that administered intravenously at the sub-anesthetic dose can play a fast and effective antidepressant effect,and then there has been an endless stream of studies on the mechanism of ketamine antidepressant,including the hypothesis of disinhibition hypothesis,inhibition of extra-synaptic NMDARs,the hypothesis of ketamine metabolites,the hypothesis of inhibition of NMDAR-dependent burst firing activity of lateral habenula(LHb)neurons,Recent studies have shown that calcium/calmodulin-dependent protein kinase Ⅱbeta subunits(CaMKⅡ beta)play a key role in the formation of depressive core symptoms,and the knockout of calcium/calmodulin-dependent protein kinase Ⅱalpha subunits(CaMKⅡ alpha)in the hippocampus of mice can lead to a series of psychiatric symptoms.Therefore,we speculate that CaMKⅡaplha and CaMKⅡbeta in the hippocampus may be involved in the antidepressant effects of ketamine.This research studied the changes and effects of CaMKⅡaplha and CaMKⅡbeta in the process of ketamine antidepressant by establishing lipopolysaccharide-induced depressive mice model,and discussed the possible related mechanisms.Methods:Eighty male adult C57BL/6 mices,20～30 g,randomly divided into 4 groups(N=20):Control group(CON),Control+Ketamine group(CON+KET),Acute stress group(LPS),Acute stress+Ketamine group(LPS+KET).CON Group normal feeding,LPS group and LPS+KET group on the day of the model intraperitoneal injection of 1 mg/kg LPS,LPS+KET group intraperitoneal injection of 10 mg/kg ketamine,after intraperitoneal injection of LPS 20 hours,CON+KET group at the same time point intraperitoneal injection of 10 mg/kg ketamine.After 4 hours,the Open field test,Novelty suppressed feeding test,Forced swimming test to detect their athletic ability,anxiety-like and depressive-like behavior.After behavioral detection,the hippocampus of mice was taken,the CaMKⅡalpha,CaMKⅡbeta in hippocampus was detected by Western blotting,and the Phosphorylated N-methyl D-aspartate receptor subtype 2B(p-GluN2B),N-methyl D-aspartate receptor subtype 2B(GluN2B),Death-related protein kinase-1(DAPK1),Drive protein 2 family members(KIF17),Postsynaptic density protein(Postsynaptic density 95,PSD95),Synapsinl,α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor 1(GluA1),α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor 2(GluA2)was measured.The colocation ratio of Synapsin 1/CaMKⅡalpha and Synapsin 1/CaMKⅡbeta were detected by immunofluorescence,the combination of KIF17,CaMKⅡ and GluN2B were detected by Immune Co-precipitation,and the change of miniExcitatory post-synaptic current of pyramidal neurons in hippocampus CA1 region and long term potentiation in SC-CA1 were detected by Electrophysiology.Results:In the OFT,there was no statistically difference in the total movement distance of the four groups(P>0.05).Compared with CON group,the time of the LPS group exploration in the central region was obviously shortened(P<0.05),the number of the enter central region decreased significantly(P<0.05),compared with the LPS group,the time of the LPS+KET group in the central region increased significantly(P<0.05),the number of the enter central region increased significantly(P<0.05).In FST,compared with CON group,the immobility time of LPS group was obviously prolonged(P<0.05),the swimming time was shortened obviously(P<0.05),compared with the LPS Group,the immobility time of LPS+KET group was shortened significantly(P<0.05),the time of swimming was obviously prolonged(P<0.05),and the climbing time was not statistically significant of the four groups(P>0.05).In the NSFT,compared with the CON group,the latent latency of LPS group was significantly prolonged(P<0.05),compared with that in LPS group,the latency of LPS+KET group was significantly shortened(P<0.05),and the total food intake of 15 minutes in each group was not statistically significant(P>0.05).Compared with CON group,the expression content of CaMKⅡalpha in the hippocampus cell membrane and synaptoneurosomes of LPS group increased significantly(P<0.05)in Western blot,compared with that in LPS Group,the CaMKⅡalpha expression content of hippocampus cell membrane and synaptoneurosomes in LPS+KET group decreased significantly(P<0.05).There was no statistically significant difference in the expression of CaMKⅡalpha in hippocampus total protein(P>0.05).Compared with the CON group,the expression of CaMKⅡbeta in the hippocampus cell membrane and synaptoneurosomes of LPS group decreased significantly(P<0.05),and the expression of CaMKⅡbeta in the hippocampus membrane and synaptoneurosomes of LPS+KET group was significantly higher than that in LPS group(P<0.05).There was no statistically significant difference in the expression of CaMKⅡbeta in hippocampus total protein(P>0.05).Compared with the CON group,the KIF17 expression content in total protein and the expression content of p-GluN2B、GluN2B、DAPK1 in the synaptoneurosomes increased significantly compared with LPS group(P<0.05).Compared with LPS group,the KIF17 expression content in total protein and the expression content of p-GluN2B、GluN2B、DAPK1 in the synaptoneurosomes decreased significantly compared with LPS group(P<0.05).Compared with the CON group,the expression of PSD95,Synapsin1 and GluA1 in the hippocampus synaptoneurosomes of LPS group decreased significantly(P<0.05),and the expression of PSD95,Synapsin1 and GluAl in hippocampal synaptoneurosomes of LPS+KET group increased significantly compared with that in LPS group(P<0.05).Compared with CON group,the colocation ratio of hippocampal primary neurons Synapsin 1/CaMKⅡalpha in LPS Group increased markedly(P<0.05),and the synapsin 1/CaMKⅡalpha of hippocampal neurons in LPS+KET group decreased significantly compared with that in LPS group(P<0.05).Compared with CON group,the colocation ratio of hippocampal primary neurons Synapsin 1/CaMKⅡbeta in LPS group decreased markedly(P<0.05),and the synapsin 1/CaMKⅡbeta colocation ratio of hippocampal primary neurons in LPS+KET group increased significantly compared with that in LPS group(P<0.05).In the experiment of immune co-precipitation,CaMKⅡ was combined with GluN2B specificity,and KIF17 was not combined with GluN2B.Compared with the CON group,the miniEPSC amplitude of the pyramidal neurons in the hippocampus CA1 region of LPS group decreased(P<0.05)and the frequency significantly reduced(P<0.01),compared with the LPS group,the miniEPSC amplitude of the pyramidal neurons in the CA1 region of the LPS+KET group increased significantly(P<0.05)and the frequency was significantly increased(P<0.01).Compared with the CON group,LTP in SC-CA1 area of LPS group was induced significant obstacles than that in LPS group(P<0.01),and the LTP evoked in SC-CA1 area of LPS+KET group regression normal(P<0.01).Conclusion:Acute LPS stress can cause depression and anxiety-like behavior in mice,leading to an imbalance between CaMKⅡalpha and beta subunits,increasing the membrane of GluN2B,decreasing the membrane of GluA1,causing excessive influx of calcium ions to produce neurotoxicity,and at the same time destroying the brain regional neurotransmitter delivery balance and synaptic plasticity.Ketamine can reverse these changes and exerts an antidepressant effect. |
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