Effects Of Electroacupuncture On Macroscopic And Microscopic Structure,Function Of Visual Cortex In Amblyopic Rats

Objective:Clinical studies have found that electroacupuncture can improve the visual acuity of amblyopic patients,but the specific mechanism is still unclear.This experiment used the classical method of amblyopia modeling to study the changes of macroscopic and microscopic structure,function of visual cortex of amblyopic rats in electroacupuncture intervention group and non-intervention group,to explore the possible mechanism of electroacupuncture treatment of amblyopia.Method:1 To observe the effect of electroacupuncture on macro-brain functional connectivity in visual cortex of form deprivation amblyopia rats.Adult SD rats were caged and delivered about 22 days pregnant later.Eighteen newborn rats were randomly selected.Random number table method was used to divide them into three groups:normal control group,amblyopia model group,amblyopia model+electroacupuncture intervention group.Rats in amblyopia model group and amblyopia model+electroacupuncture intervention group were sutured right eyelid 13 days after birth to make monocular form deprivation（MD）amblyopia model.The sutured eyelids were cut45 days after birth.From 30 days to 180 days（totally five months）,the rats in amblyopia model+electroacupuncture intervention group were given electroacupuncture intervention.The intervention points were bilateral Hegu,Taiyang,right side Cuanzhu and middle Baihui,of which right side Cuanzhu,Taiyang,left side Hegu,Taiyang were given electroacupuncture intervention,other points were given ordinary acupuncture.Rats were intervened for 30 minutes a day,at least 6 times a week.The other two groups restraint operation for the same length of time during acupuncture intervention.Behavioral acuity test was conducted 60 days after birth in three groups.After completion of the test,blood oxygen level dependent functional magnetic resonance imaging（BOLD-f MRI）was performed to observe the functional connectivity of the brain in resting state.2 To observe the effect of electroacupuncture on the microstructure in visual cortex of form deprivation amblyopia rats and its molecular mechanism.The changes of synaptic density and the percentage of mitochondrial area per unit area of visual field in the bilateral primary visual cortex（V1M）of the three groups were detected by transmission electron microscopy.The contents of dopamine（DA）,gamma-aminobutyric acid（GABA）and its receptors（D₁,GABA_A）m RNA expression change in the contralateral visual cortex of amblyopic eyes after short-term electroacupuncture intervention were detected by high performance liquid chromatography（HPLC）and quantitative polymerase Chain Reaction（q PCR）.Results:1 Effects of electroacupuncture on behavioral acuity and brain functional connection in visual cortex of MD amblyopic rats.（1）The results of behavioral visual acuity test showed that there were significant differences among the three groups(F_（2,15）=384.68,P<0.05).Compared with the normal control group,amblyopia model group and amblyopia model+electroacupuncture interve-ntion group showed a significant decrease in amblyopia behavior acuity（P<0.05,P<0.05）,while amblyopia model+electroacupuncture intervention group showed a significant incr-ease in behavioral acuity compared with amblyopia model group（P<0.05）.（2）The results of BOLD-f MRI scan of visual cortex were as follows:The functional connectivity between V1M and retrosplenial dysgranular cortex（RSD）was significantly different among the three groups（P<0.05）.The functional connectivity of the two brain regions in amblyopia model group and amblyopia model+electroacupuncture intervention group was significantly lower than that in normal control group（P<0.05,P<0.05）,while that in amblyopia model+electroacupuncture intervention group was significantly higher than that in amblyopia model group（P<0.05）.The functional connectivity between V1M and the secondary visual cortex（V2）was significantly different among the three groups（P<0.05）.The functional connectivity of the two brain regions in amblyopia model group and amblyopia model+electroacupuncture intervention group was significantly lower than that in normal control group（P<0.05,P<0.05）.There was no significant difference between amblyopia model+electroacupuncture intervention group and amblyopia model group（P>0.05）.The functional connectivity between V1M and primary visual cortex binocular area（V1B）was significantly different among the three groups（P<0.05）.The functional connectivity of the two brain regions in amblyopia model group and amblyopia model+electroacupuncture intervention group was significantly lower than that in normal control group（P<0.05,P<0.05）.There was no significant difference between amblyopia model+electroacupuncture intervention group and amblyopia model group（P>0.05）.2 Effects of electroacupuncture on microstructure of visual cortex in MD amblyopic rats.（1）The observation of synaptic density in visual cortex by transmission electron microscopy was as follows:There was significant difference among the three groups(F_（4,16）=10.423,P<0.01).Compared with the normal control group,the synaptic density of bilateral visual cortex in amblyopia model group decreased significantly（P<0.01,P<0.05）,and that in contralateral visual cortex of amblyopic eye decreased more significantly.Compared with amblyopia model group,the synaptic density of bilateral visual cortex in amblyopia model+electroacupuncture intervention group increased signi-ficantly（P<0.01,P>0.05）,and there was no statistical difference compared with the normal control group（P>0.05,P>0.05）.Comparisons of synaptic density of bilateral visual cortex in amblyopia model group:the contralateral visual cortex of amblyopic eyes was significantly lower than that of healthy eyes（P<0.01）.The synaptic density of bilateral visual cortex in amblyopia model+electroacupuncture intervention group:there was no statistical difference between the contralateral visual cortex of amblyopic eyes and that of healthy eyes（P>0.05）.（2）The percentage of mitochondria in visual field per unit area of visual cortex was observed by transmission electron microscopy as follows:There were significant differences among the three groups(F_（4,15）=7.489,P<0.01).Compared with the normal control group,the percentage of mitochondrial total area per unit area of visual field in the bilateral visual cortex of amblyopia model group decreased significantly（P<0.01,P<0.05）,and that in the contralateral visual cortex of amblyopia eye decreased more significantly.Compared with the amblyopia model group,the percentage of mitochondrial total area in the bilateral visual cortex of amblyopia model+electroacupuncture interven-tion group increased significantly（P<0.01,P<0.05）,and compared with the normal control group:there was no significant difference between the two groups（P>0.05,P>0.05）.Comparing the percentage of mitochondrial total area of bilateral visual cortex between amblyopia model group and amblyopia model+electroacupuncture intervention group:there was no significant difference between the two groups（P>0.05,P>0.05）.（3）The content of DA and the expression of its receptor D₁ in the contralateral visual cortex of amblyopic eyes were as follows:after the establishment of amblyopic model,the content of DA and the expression of its receptor D₁ in amblyopic model group were significantly lower than those in normal control group（P<0.01,P<0.01）,while the content of DA and the expression of its receptor D₁ in amblyopic model+electroacupunct-ure intervention group were significantly higher than those in amblyopic model group（P<0.01,P>0.05）,although still lower than those in normal control group（P<0.01,P<0.01）.（4）The content of GABA and the expression of its receptor GABA_A in the contralateral visual cortex of amblyopic eyes were as follows:the content of GABA and the expression of its receptor GABA_A in amblyopic model group were significantly lower than those in normal control group（P<0.01,P<0.01）,while in the amblyopic model+electroacupuncture intervention group,the content of GABA and the expression of its receptor GABA_A were significantly higher than those in the amblyopic model group（P<0.01,P<0.05）,although still lower than those in the normal control group（P>0.05,P<0.01）.Conclusion:1 Behavioral visual acuity of amblyopia rats decreased significantly after modeling,which indicated that the model of amblyopia was successful（in line with the definition of amblyopia）.After electroacupuncture treatment,the visual acuity of amblyopic eyes has been significantly improved,which proves the therapeutic effect of electroacupuncture on amblyopia,and lays a foundation for the follow-up study on the mechanism of electroacupuncture treatment of amblyopic rats.2 The effect of amblyopia on the visual cortex of the brain is not local,but multi-directional and multi-range damage.The functional connection between V1M area and RSD area,V2 area and V1M area in amblyopic model group was significantly lower than that in normal control group,suggesting that the integrated ability of visual signals in amblyopic rats was decreased.Electroacupuncture treatment could significantly improve the functional connection between V1M area and RSD area,and enhance the processing and integration of visual signals in amblyopic rats.3 The synaptic density in the bilateral visual cortex of the amblyopia model group was significantly lower than that of the normal control group,which indicated that MD amblyopia model in the sensitive period of visual development could cause synaptic damage in the visual cortex.After electroacupuncture intervention,the synaptic density of bilateral visual cortex in amblyopia model+electroacupuncture intervention group increa-sed significantly,suggesting that electroacupuncture treatment shows a positive effect on the synaptic density of visual cortex in amblyopic rats,and electroacupuncture can significantly antagonize the inhibitory effect of amblyopia on visual system.4 The percentage of total mitochondrial area per unit area of visual field in the bilateral visual cortex of the brain decreased significantly after amblyopia modeling compared with the normal control group,suggesting that the occurrence of amblyopia may be related to the damage of brain mitochondria.Mitochondria not only provide energy for the life activities of cells,but also control the release of Ca²⁺.The concentration of Ca²⁺also regulates the connection between the functions of various brain regions.From the above results,it can be concluded that the occurrence and development of amblyopia will inevitably lead to changes in the macroscopic and microscopic structure,function of visual cortex.After electroacupuncture intervention,the percentage of total mitochondrial area in bilateral visual cortex increased significantly,indicating that electroacupuncture treatment can promote the repair of mitochondria in visual cortex of amblyopic rats,thereby regulating the concentration of Ca²⁺and increasing energy supply,and strengthening functional connectivity between brain regions.5 The content of DA,GABA and their receptors in the visual cortex of the brain decreased after amblyopia modeling,and increased after electroacupuncture intervention,It suggests that the occurrence and development of amblyopia is related to the content of DA and GABA in the visual cortex.Electroacupuncture treatment of amblyopia plays a role by interfering with the content of these two neurotransmitters.