| BackgroundAlzheimer’s disease（AD）is a neurodegenerative disorder,and its main clinical feature is progressive memory deficits and cognitive impairments.There are more than30 million AD patients worldwide in 2010 and an expected 106 million by 2050.For families,individuals and health care systems,AD is a heavy burden.So far,it has been proven that drug treatment is limited,and more and more research is focused on improving and delaying the onset and progression of AD by non-pharmacological interventions.Studies on the pathogenesis of AD have never stopped.In some previous research,the deposition of amyloid precursor protein（Aβ）to form plaques and neuronal tangles is considered to be the main disease reason of AD,but in recent years,studies have found that AD brain energy metabolism disorders are closely related to cognitive impairment,and are earlier than Aβdeposition and neuronal tangles.Therefore,intervention in the treatment of AD brain energy metabolism disorders may be a new therapeutic direction.The normal adult brain take up approximately 2%of the body weight and consumes approximately 20%of glucose in the body（about 120 g of glucose per day）.Glucose is the primary source of energy in the energy metabolism of the brain,but glucose does not pass through the cell membrane freely.It needs to enter the cell with the help of glucose transporters（GLUTs）to participate in energy metabolism in the brain.Similarly,glucose aerobic metabolism is the main source of energy in the nervous system,and major energy is used to maintain excitatory synaptic transmission and interactions between neurons,astrocytes,and cells.These cells express different subtypes of glucose transporters（GLUTs）that promote glucose-mediated sodium-independent transmembrane transport.GLUT1 and GLUT3 are expressed in the nervous system,GLUT1（encoded by the SLC2A1 gene）mainly express in endothelial cells and astrocytes;whereas GLUT3（encoded by the SLC2A3 gene）mainly express in neurons.In general,GLUT1 and GLUT3 are essential for energy metabolism in the brainA series of observational studies confirmed that AD is closely related to unhealthy lifestyles,including lack of physical activity.Therefore,we propose a hypothesis:regular exercise can enhance the expression of GLUT1 and GLUT3 in the brain,thereby improving the energy metabolism disorder in the brain,and further delaying the cognitive impairment of AD,especially learning and memory skills.PurposesIn this experiment,APP/PS1 double transgenic mice were used to express various pathological features of AD.The standard for regular exercise in mice was forced swimming for 4 weeks（2 h/day,6 days/week）in a pool.The learning and memory ability was detected by Morris water maze was used to detect the ability of cognition,and the effects of regular exercise on the expression of GLUT1 and GLUT3 in hippocampus and cortex were further examined.The experiment explored the effects of regular exercise on the cognitive function of AD by improving the energy metabolism state in the brain of mice,aiming to provide new treatment methods for delaying the occurrence and development of AD.Methods（1）Identification of AD offspring and swimming trainingAD offspring were used to extract DNA,the DNA of AD offspring was amplified by PCR,and then identified by agarose gel electrophoresis.The band which was 350bp was AD positive mice.Subsequently,AD mice and same-month-old wild-type（WT）mice were grouped:I WT untrained group（WT-NT）;II WT training group（WT-T）;III AD untrained group（AD-NT）;IV AD training group（AD-T）.The training teams were trained one month by swimming training（2 hour/per day,6 days/per week）.（2）Morris water mazeHidden platform experiment:When a mouse swims in the water,its instinct urges it to find a place to rest.This is a complex memory process,including the collection and processing of spatial information and visual information to find a hidden platform.Base on this fundamental,the hidden platform experiment of the mouse can also be called the positioning navigation experiment which detect the learning and memory ability of the mouse.The water temperature was kept at about 25°C.In order to create an appropriate environment,the pool is covered with curtains.In the computer software（Smart 3.0）,the circular pool is divided into four quadrants（I,II,III,IV）,the platform is placed in the second quadrant,and the mice were placed from the midpoint of each quadrant,and the automatic imaging analysis system tracked and recorded mice swimming path.Statistical analysis was performed on the time that the mouse found and climbed the platform（Latency to target）and distance（Total Distance）.The test was continued for 6 days.Space exploration experiment:After the end of the navigation experiment,the platform was removed,and the mice were placed in the water from entry point of quadrant which away from the platform quadrant.The number of mice crossing the platform area and path in the 60s were recorded and analyzed to compare the spatial positioning ability.（3）Transmission electron microscopy was used to detect the change of mitochondrial morphological structure and synaptic number.Mitochondria are closely related to the state of energy metabolism in the brain,the energy metabolism in the brain is imbalanced when mitochondrial dysfunction occurs,and the density of synapses is important to maintain normal brain function.The change of mitochondrial morphological structure and synaptic density was detected by transmission electron microscopy to observe the effects of regular exercise on energy metabolism and normal brain function in the brain.（4）Detection of GLUT1 and GLUT3 expression levels in the hippocampus and cortex of the brain.In order to detect the effect of regular exercise on the expression of GLUTs in the brain of AD model mice,the expression of GLUT1 and GLUT3 in different brain subareas of mice was detected by Western blot.To further detect the influence of regular exercise on GLUTs in different regions of the brain,the IF method was used.The expression levels of GLUTs in hippocampal CA1 and CA3 and cortex of different groups of mice was examined.Results（1）After DNA was extracted from AD offspring,PCR amplification and agarose detection were performed,and 350bp was an AD positive mouse.（2）The learning and memory ability of the AD group was significantly lower than that of the WT group（**p<0.01）.After regular exercise intervention,the learning and memory ability(##p<0.01)and spatial exploration ability of the AD-T group(##p<0.01)was significantly improved relative to the AD-NT group of mice.（3）The result of electron microscopy showed that when AD group compared with the WT group,the mitochondrial cristae was broken and the mitochondria outer membrane edge was blurred and the number of synapses was significantly decreased（***p<0.001）.The number of synapses in the AD-T group was higher than that in the AD-NT group(##p<0.01),and the mitochondria cristae was more dense and intact,and the mitochondrial outer membrane was clearer.（4）Western blot results showed that the expression level of GLUT1 and GLUT3 in the brain of AD group were significantly lower than that in WT group（***p<0.001）,whether in the cerebral cortex or hippocampus.And regular exercise significantly enhanced the expression levels of GLUT1(##p<0.01)and GLUT3（#p<0.05）in the brain endothelium and hippocampus of AD mice.（5）IF results showed that the expression levels of GLUT1 and GLUT3 in the brain of AD group were significantly lower than those in WT group（***p<0.001）,whether in the cerebral cortex or hippocampus.Compared with the AD-NT group,the expression levels of GLUT1（#p<0.05）and GLUT3(##p<0.01)in the brain endothelium and hippocampal CA1 and CA3 regions were significantly increased.ConclusionThe experimental results show that regular exercise can increase the expression of GLUT1 and GLUT3 in the brain of AD mice,thereby improving the energy metabolism disorder in the brain,further enhancing the learning and memory ability of AD mice,and delaying the occurrence and development of AD. |
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