| Objectives: Previous studies have revealed that spatial memory deficit and cognition dysfunction were impaired by central insulin resistance,while improved via treatment with hypoglycemia drugs.The underlying mechanisms mainly involve inflammation,oxidative stress,synaptic transmission,and neuronal apoptosis.Unfortunately,there are only a few studies on cerebral metabolism,namely astrocyte-neuron metabolic cooperation,and how cerebral metabolism plays a role in memory deficit and cognitive dysfunction is yet unclear.In this study,we evalutated the changes in spatial memory and cerebral metabolism in C57BL/6J mice with central insulin resistance induced by high-fat diet,and then determined therapeutic effects of various hypoglycemic drugs to evaluate the thrapeutic effects.We also investigated the mechanisms of central insulin resistance possibly causing spatial memory deficit and cognitive impairment in order to provide a potential theoretical basis for treatment of diabetes-related cognitive dysfunction and new idears for delaying diabetes-related nerval aging.Methods: Male C57BL/6J mice with weight of 16-18 g were randomly divided into five groups: control(Ctrl)group,high-fat(HF)group,alogliptin(ALO)group,exenatide(EXE)group,and pioglitazone(PIO)group.Ctrl group was fed on normal diet and the others were fed on high-fat diet for 12 weeks.Insulin resistance were evaluated by the oral glucose tolerance test(OGTT)after 12 weeks.From the 13 th week,ALO group,EXE group,and PIO group were administrated with alogliptin(25 mg/kg/day,i.g.,q.d.),exenatide(30 ug/kg/day,s.c.,b.i.d.),and pioglitazone(10 mg/kg/day,i.g.,q.d.),respectively,while Ctrl group and HF group were given saline(10 ml/kg,i.g.,q.d.)for 4 weeks.Body weight was measured weekly.Food intake was recorded and fasting blood glucose was measured using tail vein-blood every two weeks.The OGTT was performed every four weeks.Spatial memory was accessed using the Morris Water Maze test after 16 weeks.Three to five days after completion of behavioral tests mice were sacrificed,and their blood and brains were rapidly collected.The expression of astrocyte-neuron metabolic cooperation-related transporters and enzymes,aging-related factors,and synaptic plasticity-related factors were evaluated by Western blot.The energy metabolic substrates in hippocampus,cortex,and blood samples were detected by kit.Cellular morphology and cell number of hippocampal and cortex tissue were determined by Hematoxylin eosin (HE)staining.Nissl bodies in hippocampus and cortex was detected by Nissl staining and cell apoptosis in hippocampal and cortex tissue was detected by Td T-mediated d UTP Nick-End Labeling(TUNEL)kit.All data were collected and analyzed using SPSS21.0 and Graph Pad7.0.Statistical analysis was performed using the one-way analysis of variance(ANOVA)test,except that two-way ANOVA test were used for Morris Water Maze test.Measurement data were presented as mean±SD and P <0.05 was considered statistically significance.Results: 1.Energy intake and zoometric parameters in animal models After 16 weeks of high-fat diet,increased caloric intake,body weight,body mass index(BMI),body fat(Lee's index),and abdominal perimeter were observed in the HF group as compared with the Ctrl group,while a decrease in the above parameter was noted in the ALO group,EXE group,and PIO group as compared with the HF group.2.Fasting plasma glucose,lipid parameters and insulin resistance in animal models After 16 weeks of high-fat diet,increased fasting plasma glucose,lipid parameters,and level of homeostasis model assessment of insulin resistance(HOMA-IR),while decreased level of homeostasis model assessment insulin secrete index(HOMA-IS)was detected in the HF group as compared with the Ctrl group.Insulin signaling-related factors,phosphorylated insulin receptor substrate 1/insulin receptor substrate 1(p IRS1/IRS1),phosphoinositide 3-kinase(PI3K),and phosphorylated protein kinase B/protein kinase B(p Akt/Akt)were downregulated in hippocampus and cortex of the HF group.However,fasting plasma glucose,lipid parameters,HOMA-IR,HOMA-IS,and insulin signaling-related factors were reversed after 4-week treatment with alogliptin,exenatide,and pioglitazone.3.Spatial memory ability in animal models Spatial memory ability was evaluated with Morris Water Maze test at the 16 th week.Comparative analysis indicated a significant increased latency in the HF group.Frequency of crossing target quadrant,frequency of crossing platform,time spent in the target quadrant,and distance travelled in the target quadrant was decreased in the HF group as compared with the Ctrl group.Moreover,latencies were decreased,while frequency of crossing target quadrant and frequency of crossing platform were increased in the ALO group,EXE group,and PIO group.Time spent in target quadrant was increased only in the PIO group as compared with the HF group.4.Neuronal morphology,structure,quantity,apoptosis and senescence in hippocampus and cortex Destroyed morphology and neuronal loss,Nissl bodies dissolution and accelerated apoptosis in the HF group was detected by HE staining,Nissl's staining and TUNEL kit,respectively.There was no significant difference in silencing information regulator 2(SIR2)-related enzyme 1(SIRT1),a biomarker of aging,in hippocampus and cortex between the Ctrl group and the HF group.Morphology,quantity,structure,and apoptosis of neuronal cells in hippocampus and cortex were alleviated after 4-week treatment with alogliptin,exenatide,and pioglitazone.In addition,exenatide and pioglitazone upregulated the expression of SIRT1,but alogliptin had no effects.5.Neuronal synaptic plasticity in hippocampus and cortex Synaptic plasticity-related factors were downregulated in hippocampus and cortex of the HF group as compared with the Ctrl group.These factors are brain-derived neurotrophic factor(BDNF),activity-regulated cytoskeleton–associated protein(Arc),phosphorylated c AMP-response element binding protein/c AMP-response element binding protein(p CREB/CREB),postsynaptic density protein 95(PSD95),and synapsin-1.Except that pioglitazone had no effects on Arc,all of the above factors were upregulated in hippocampus and cortex after 4-week treatment with alogliptin,exenatide,and pioglitazone.6.Astrocyte-neuron metabolic cooperation in hippocampus and cortex The results showed the decreased expression of glucose transporter(GLUT)1,3,monocarboxylate transporter(MCT)1,2,4,excitatory amino-acid transporter 2(EAAT2),and lactate dehydrogenase(LDH)1 in both hippocampus and cortex in the HF group as compared with the Ctrl group.In the HF group,the expression of LDH5 was decreased in hippocampus,while not changed in the cortex.All of the above factors were respectively upregulated in different degree via 4-week treatment with alogliptin,exenatide,and pioglitazone.In addition,metabolin including lactate,pyruvate,and glycogen were further evaluated by kit and periodic acid-schiff(PAS)staining.Firstly,in hippocampus,there was higher level of lactate and lower level of glycogen in the HF group as compared with the Ctrl group.Lower levels of lactate and higher levels of glycogen in ALO group,EXE group,and PIO group were detected as compared with the HF group.There was no statistical difference in pyruvate among five groups.Secondly,in cortex,comparative analysis showed no difference in lactate among five groups.Pyruvate level was increased and glycogen level was decreased in the HF group as compare with the Ctrl group.ALO group reversed the levels of pyruvate and glycogen.The other two therapeutic groups had no effects on pyruvate,but upregulated glycogen as compared with the HF group.Thirdly,in serum,HF group had lower levels of lactate and pyruvate as compared with the Ctrl group.EXE group had higher level of lactate as compared with the HF group,while no effects on level of pyruvate.Furthermore,ALO group and PIO group had no effects on the levels of lactate and pyruvate as compared with the HF group.Conclusions: Our findings indicated that spatial memory was impaired by inhibition of astrocyte-neuron metabolic cooperation in hippocampus and cortex of C57BL/6J mice with central insulin resistance induced by high-fat diet.In addition,alogliptin,exenatide,and pioglitazone effectively improved spatial memory in C57BL/6J mice with central insulin resistance by increasing astrocyte-neuron lactate shuttle(ANLS)and glucose intake,reducing neuronal structure and function damages,and delaying aging in brain tissue. |
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