The Mechanism Of Cholinergic Neural Circuit From Basal Forebrain To Hippocampus Regulated By Chemical Genetic Technique Improving Early Cognitive Function In AD Mice Based On MRS

Objective:To detect the cholinergic metabolism in the basal forebrain and hippocampus of Alzheimer’s disease model mice at different months,and further to elucidate the mechanism of the basal forebrain-hippocampus cholinergic neural circuit regulated by the chemical genetics technique improving cognitive function in AD mice.Methods:(1)According to the results of PCR,APP/PS1 double transgenic male mice were selected as AD group,and wild-type male mice were used as control group,each group with 10 mice.Morris water maze and new object recognition experiment were performed to detect the learning-memory ability of the two groups at 2,4,6 months;Using Magnetic resonance spectroscopy to analyse the levels of Choline,N-acetylaspartate and myo-Inositol in the basal forebrain and hippocampus of the two groups.(2)Immunofluorescence and Nissl staining were respectively used to detect Aβ plaques deposition and neuron loss in the basal forebrain and hippocampus of 6-month-old APP/PS1 double transgenic mice.(3)According to the random number table method,another 20 four-month-old APP/PS1 double transgenic male mice were randomly assigned to the clozapine-N-oxide(CNO)group and the saline(SA)group;The cholinergic neurons in the basal forebrain of the two groups were transfected with h M3 Dq by AAV virus;The CNO group was intraperitoneally injected with CNO solution to activate the basal forebrain-hippocampus cholinergic neural circuit and the SA group was intraperitoneally injected with an equal dose of saline as a control group.(4)Before and after chemical genetic intervention,the new object recognition experiment and Morris water maze test were used to detect the learning-memory ability of the two groups,and the material metabolism of the basal forebrain and hippocampus were analyzed by magnetic resonance spectroscopy.(5)The basal forebrain-hippocampus cholinergic neural circuit was traced and the expression of h M3 Dq receptor on basal forebrain cholinergic neurons was observed through laser confocal imaging.Results:(1)Learning-memory ability of APP/PS1 mice at different months: Compared with the wild-type mice of the same months,the learning-memory ability of APP/PS1 double transgenic mice was not impaired at 2 month(P>0.05),while the learning-memory ability of APP/PS1 double transgenic mice was impaired at 4 month and 6 month(P<0.05 or P<0.01).(2)Material metabolism of APP/PS1 mice at different months: Compared with wild-type mice of the same months,the content of Cho in the hippocampus of 4-month-old APP/PS1 mice was increased(P<0.05),while the content of Cho in the basal forebrain and the content of NAA in the hippocampus of 6-month-old APP/PS1 mice were decreased(P<0.01).(3)Pathological manifestations of APP/PS1 mice: Aβ plaques deposition occurred in the cortex,hippocampus and basal forebrain of 6-month-old APP/PS1 mice;The neurons in the basal forebrain were lost of 6-month-old APP/PS1 mice.(4)The effects of chemical genetics technique on the learning-memory and the material metabolism of APP/PS1 mice: Before intervention of chemical genetics technique,there was no difference in learning-memory ability between the CNO group and the SA group(P>0.05);There was no difference in the material metabolism in the basal forebrain and the hippocampus of the two groups(P>0.05).After intervention,the learning-memory ability of the CNO group was improved compared with the SA group(P<0.05);The content of the Cho in the basal forebrain and the content of NAA in the hippocampus in the CNO group were increased compared with the SA group(P<0.05).(5)Basal forebrain-hippocampus cholinergic neural circuit tracing and receptor expression verification: There was single synaptic connection between the basal forebrain cholinergic neurons and hippocampus;AAV virus-transfected h M3 Dq receptor was successfully expressed on the cholinergic neurons in the basal forebrain.Conclusion:The abnormalities of Choline and N-acetylaspartate are one of the important pathological manifestations of Alzheimer’s disease in the early stage;The basal forebrain-hippocampus cholinergic neural circuit regulated by the chemical genetics technique could promote synthesis of Choline in the basal forebrain and the N-acetylaspartate in the hippocampus improving the learning-memory ability of Alzheimer’s disease.