Expression Of Annexin A7 In Brain Tissue Of Epileptic Mouse And Its Effects On Neuronal Excitability

Part one Expression of Annexin A7 in brain tissue of epileptic mouseObjectives:Epilepsy is a common chronic disease in the nervous system. With the existing drug treatment, there are still 30% of people with epilepsy bearing the pain of this disease. Therefore, further study of the pathogenesis in epilepsy is very important. In recent years, abnormal calcium homeostasis in neurons is playing an important role in epilepsy. And Annexin A7 can regulate intracellular calcium homeostasis. We aimed to established pilocarpine model of epilepsy in mouse, and then initially verificate Annexin A7 expression in the brain tissue of epileptic mouse using Western blot and immunofluorescence techniques.Methods:Thirty healthy adult male C57BL/6 mice were randomly divided into experimental group and control group. Epilepsy mouse model was established in the experimental group and behavioural characteristics of mice were observed in two groups. Expression of Annexin A7 was detected respectively using western blot and immunofluorescence labeling techniques.Results:1. In cortical tissue of epileptic mouse, expression of Annexin A7 is significantly higher than in the control group (P<0.05).2. In hippocampus tissue of epileptic mouse, expression of Annexin A7 is significantly higher than in the control group (P<0.05).3. In the cortex and hippocampus of epileptic mouse, Annexin A7 and neurons labeled molecules-NeuN exist coexpression; Annexin A7 and neuronal dendrites labeled molecules-MAP2 exist near expression; Annexin A7 and astrocytes labeled molecules-GFAP does not exist coexpression or near expression.Conclusion:1. Expressions of Annexin A7 in the hippocampus and temporal cortex of experimental group were markedly higher than that of control group.2. In the brain tissue of epilepsy mouse model, Annexin A7 was expressed in the neurons rather than astrocytes.Part two Effects of Annexin A7 on hippocampal neuronal excitability in magnesium-free brain slicesObjectives:Studies have shown that abnormal discharge of neurons in epileptic activity comes from the imbalance of excitatory and inhibitory neurotransmitter release. Higher excitatory synaptic transmission or lower inhibitory synaptic transmission in in neurons are likely to make it in an over-excited state. Therefore, we aim to detect the impact of Annexin A7 on hippocampal neuronal excitability.Methods:Lentiviruses were stereotactic injected into mouse hippocampus, then EGFP fluorescence was collected with confocal fluorescence microscope to verify the success of lentivirus injection into hippocampus and transfected into neurons; Western blot technique was used to verify whether lentiviruses had successfully interferred with protein expression of Annexin A7; then whole-cell patch-clamp technique was used to perform electrophysiological test on hippocampal CA1 neurons in magnesium-free mice brain slices.Results:1. Efficiency detection of lentiviral transfection:EGFP green fluorescence was detected in the hippocampus region of the slices under confocal fluorescence microscope; In addition, Western blot showed expression of Annexin A7 protein in LV-ANXA7 group was significantly lower than in the LV-empty group.2. The electrophysiological results showed that the frequency of action potential in hippocampal CA1 neurons in LV-ANXA7 group was significantly less than that in LV-empty group (P<0.05).3. The amplitude and inner-event interval of mlPSC in LV-ANXA7 group and LV-empty group had no significant differences.4. Inner-event interval of mEPSC in LV-ANXA7 group was significantly longer than in LV-empty group, and no differences were found in mEPSC amplitude between the two groups.Conclusion:1. In magnesium-free model of brain slices, Annexin A7 increased the overall excitability of hippocampal CA1 neurons.2. In magnesium-free model of brain slices, Annexin A7 increased excitatory synaptic transmission of hippocampal CA1 neurons, while did nothing to the inhibitory synaptic transmission. And the increase of excitatory synaptic transmission may due to alteration of presynaptic components.3. Annexin A7 may be involved in the development and progression of epilepsy.